Soluble HLA-G circulates in maternal blood during pregnancy

系统医学 2023 年 12 月第 8 卷第 24期血清降钙素原检测、微生物培养的临床应用价值廖龙波荔浦市人民医院检验科，广西荔浦546699[摘要]目的探究分析血清降钙素原以及微生物培养在细菌感染中的价值。

方法回顾性选取2022年1—12月荔浦市人民医院检验科500例疑似细菌感染患者的临床资料，均实施血清降钙素原检测、微生物培养（细菌培养或/和血培养），以微生物培养结果为金标准，分析血清降钙素原检测结果及效能。

结果微生物培养结果是金标准，有菌生长173例，无菌生长327例。

血清降钙素原检测显示，阳性296例，阴性204例，阳性率59.20%、阴性率40.80%，敏感度是82.08%、特异度是52.91%、准确度是63.00%、误诊率是47.09%、漏诊率是17.92%、阳性预测值是47.97%、阴性预测值是84.80%，Kappa值为0.841。

结论血清降钙素原具有检测速度快、敏感度高等优点，可及时识别细菌感染，若要明确病原菌种类，建议进一步进行微生物培养，辅助临床尽早确诊与治疗。

[关键词]血清降钙素原；微生物培养；有菌生长；无菌生长；敏感度；特异度[中图分类号]R446.1 [文献标识码] A [文章编号]2096-1782（2023）12(b)-0049-04 Clinical Application Value of Serum Procalcitonin Detection and Microbial CultureLIAO LongboDepartment of Laboratory, Lipu People's Hospital, Lipu, Guangxi Zhuang Autonomous Region, 546699 China [Abstract] Objective To explore the value of serum procalcitonin and microbial culture in bacterial infection. Methods the clinical data of 500 patients with suspected bacterial infections in the Laboratory Department of Lipu City People's Hospital from January to December 2022 were retrospective selected, all of whom were implemented se⁃rum calcitoninogen detection, microbial culture (bacterial culture or/and blood culture), and the results of the micro⁃bial culture results were used as the gold standard to analyze the results and efficacy of serum calcitoninogen detec⁃tion. Results The results of microbial culture were the gold standard, as shown below: 173 cases had bacterial growth and 327 cases had aseptic growth. Serum procalcitonin test showed 296 positive cases, 204 negative cases, positive rate of 59.20%, negative rate of 40.80%. Sensitivity was 82.08%, specificity was 52.91%, accuracy was 63.00%, mis⁃diagnosis rate was 47.09%, missed diagnosis rate was 17.92%, positive predictive value was 47.97%, negative predic⁃tive value was 84.80%, and Kappa value was 0.841. Conclusion Serum procalcitonin has the advantages of fast detec⁃tion speed and high sensitivity, which can timely identify bacterial infections. If the type of pathogen needs to be iden⁃tified, it is recommended to further conduct microbial culture to assist clinical diagnosis and treatment as early as pos⁃sible.[Key words] Serum procalcitonin; Microbial culture; Bacterial growth; Aseptic growth; Sensitivity; Specificity血流感染是导致医院危重症患者预后不佳的主要原因，临床多在怀疑患者发生血流感染后，以血培养为首选诊断方案，将其作为血流感染诊断金标准[1-2]。

英语单词表细胞 Cellhistology [his’tɔlǝdƷi] 组织学embryology [e mbri’ɔlǝdƷi]胚胎学light microscope[laitmai krɔskoup]光学显微镜electron microscope ['maɪkrə,skop] 电子显微镜hematoxylin [hem☯t ksilin]苏木精eosin [ i:☯sin] 伊红basophil [beis fil] 嗜碱性acidophil [asid fil] 嗜酸性neutrophil [ nju tr☯fil] 中性histochemistry [ histou kemistri] 组织化学immuno histochemistry免疫组织化学cell coat [k☯ut] 细胞衣cell membrane [ membrein] 细胞膜cytoplasm [ sait pl✌zm] 细胞质organelle [ g☯nel] 细胞器mitochondria [mait☯u k ndri☯] 线粒体Golgi complex [k mpleks] 高尔基复合体smooth endoplasmic reticulum滑面内质网[smu: end☯u’pl✌smik ri’tikjulem] rough endoplasmic reticulum [r✈f]粗面内质网ribosome [raib s☯um]核蛋白体lysosome [lais s☯um] 溶酶体microtubule[maikr☯’tu:bjul]微管microfilament [maikr☯’fil☯m☯nt] 微丝centrosome [’sentr s☯um] 中心粒inclusion [in’klu:✞☯n] 包含物lipid droplet [’lipid ’dr plit] 脂滴nucleus [’nju:kli☯s] 细胞核nuclear envelope [’envil☯up] 核膜chromatin [kr☯um☯tin] 染色质euchromatin [ju:kr☯um☯tin] 常染色质heterochromatin [heter☯’kr☯um☯tin] 异染色质chromosome [’kr☯um☯soum] 染色体nucleous [nju:’kli:☯l☯s] 核仁上皮组织Epithelial Tissue epithelium [ epi i:li☜m] 上皮free surface[fri: s☜:fis] 游离面basement membrane 基底膜microvillus [mаikr☐✞vil☜♦] 微绒毛cilium [ sili☜m] 纤毛mesothelium [mes☜i:li☯m] 间皮endothelium [endou i:li☜m] 内皮simple squamous epithelium单层扁平上皮simple cuboidal epithelium 单层立方上皮simple columnar epithelium单层柱状上皮pseudostratified ciliated columnar epithelium[sju:d☜str✌tifaid]假复层纤毛柱状上皮transitional epithelium [tr✌n si☞n☯l] 变移上皮intercellular substance 细胞间质stratified squamous epithelium复层扁平上皮junctional complex 连接复合体plasma membrane infolding 质膜内褶protein-secreting cel l 蛋白质分泌细胞steroid-secreting cell类固醇分泌细胞glandular epithelium 腺上皮serous cell [ si☜r☜] 浆液细胞mucous cell [ mju:k☜s] 黏液细胞endocrine gland [end krain] 内分泌腺exocrine gland 外分泌腺结缔组织Connective Tissuemesenchyme 间充质loose connective tissue 疏松结缔组织elastic fiber[i'læstik] 弹性纤维collagenous fiber 胶原纤维collagenous fibril['faibril] 胶原原纤维reticular fiber 网状纤维ground substance[graund's☜bst☜ns]基质fibroblast['faibr☜blæst] 成纤维细胞macrophage['ma:kr☜feid☜] 巨噬细胞plasma cell['plæzm☜] 浆细胞mast cell [mæst] 肥大细胞fat cell[fat] 脂肪细胞dense connective tissue[dens]致密结缔组织adipose tissue['ædipous] 脂肪组织reticular tissue[ri'tikjul☜] 网状组织血液与血细胞发生Blood and Hematopoisisplasma ['p1æzm☜] 血浆serum ['si☜r☜rn] 血清erythrocyte [ i'ri☜rousait ] 红细胞hemoglobin （Hb）[hi:mou'gloubin] 血红蛋白reticulocyte [ ri'tikjul☜sait ] 网织红细胞leucocyte ['lju:k☜sait] 白细胞neutrophilic granulocyle, neutrophil [ nju:tr☜'filik'grænjul☜sait] [ nju:tr☜'fil ] 中性粒细胞eosinophilic granulocyte, eosinophil [i:ousinou'filik] 嗜酸性粒细胞basophilic granulocyte, basophil [beis☜'filik] 嗜碱性粒细胞monocyte ['m☜nousait ] 单核细胞lymphocyte [ 'limf☜sait ] 淋巴细胞hemopoietic stem cell [ hi:'m☜p☜i'etik stem] 造血干细胞blood platelet ['pleitlet ] 血小板软骨和骨Cartilage and Bonecartilage tissue 软骨组织hyaline cartilage 透明软骨fibrous cartilage 纤维软骨elastic cartilage 弹性软骨cartilage lacuna 软骨陷窝cartilage capsule 软骨囊perichondrium 软骨膜osseous tissue 骨组织bone matrix 骨基质bone lamella 骨板osteocyte 骨细胞bone lacuna 骨陷窝bone canaliculi 骨小管osteoprogenic cell 骨祖细胞osteoblast 成骨细胞osteoclast 破骨细胞osteoid 类骨质circumferential lamella 环骨板osteon 骨单位interstitial lamella 间骨板intramembranous ossification 膜内成骨endochondral ossification 软骨内成骨肌组织Muscle Tissuemuscle tissue 肌组织skeletal muscle 骨骼肌cardiac muscle 心肌smooth muscle 平滑肌sarcolemma 肌膜myofibril 肌原纤维sarcomere 肌节transverse tubule 横小管sarcoplasmic reticulum 肌浆网triad三联体intercalated disk 闰盘densebody 密体神经组织Nerve Tissuenervous tissue 神经组织neuron 神经元dendrite 树突axon 轴突axon hillock 轴丘axoplasm 轴浆Nissl body 尼氏体synapse 突触presynaptic element 突触前成分synaptic cleft 突触间隙postsynaptic element 突触后成分synapse vesicle 突触小泡neuroglia cell 神经胶质细胞astrocyte 星形胶质细胞oligodendrocyte 少突胶质细胞microglia 小胶质细胞Schwann cell 施万细胞satellite cell 卫星细胞nerve fiber 神经纤维Ranvier node 郎飞氏结neurofibril 神经原纤维myelin sheath 髓鞘neurolemma 神经膜眼和耳retina 视网膜rod cell 视杆细胞cone cell 视锥细胞ganglion cell 节细胞spiral organ 螺旋器循环系统circulatory system tunica intima 内膜tunica media 中膜tunica adventitia 外膜internal elastic membrane 内弹性膜external elastic membrane 外弹性膜elastic artery 弹性动脉muscular artery 肌性动脉capillary 毛细血管sinusoid 血窦endocardium 心内膜myocardium 心肌膜epicardium 心外膜皮肤skinepidermis 表皮dermis 真皮hair 毛hair shaft 毛干hair root 毛根hair follicle 毛囊hair bulb 毛球hair papilla 毛乳头sweat gland 汗腺sebaceous gland 皮脂腺免疫系统Immunesystemlymphoid tissue [lim f id tisju:]淋巴组织thymocyte [ aim sait] 胸腺细胞lymph node [ limf noud] 淋巴结trabecula [tr☜bekjul☜] 小梁paracortical zone 副皮质区lymphoid sinus [lim f id sain☜s] 淋巴窦medullary cord [me d✈l☜ri k :d] 髓索spleen [spli:n] 脾脏white pulp [hwait p✈lp] 白髓red pulp [red p✈lp] 红髓splenic sinusoid [ spli:nik sain☜s id] 脾窦内分泌系统Endocrine systemhormone [ h :moun] 激素target cell [ t :git si:l] 靶细胞follicle [ f likl] 滤泡pituicyte [pi tju（:）isait] 垂体细胞parafollicular cell滤泡旁细胞zona glomerulosa[gl m☜ru l s☜]球状带zona fasciculata [zoun f✌sikju leit] 束状带zona reticularis [zoun ritikju lris] 网状带chromaffin cell [krou m✌fin si:l] 嗜铬细胞chromophobe cell [ kroumfoub si:l]嫌色细胞消化管digestive tract digestive tract 消化管mucosa [mju: kous☜] 粘膜lamina propria [ læmin☜ proupri☜]固有层muscularis mucosa粘膜肌层submucosa [s✈bmju kous☜]粘膜下层muscularis [m✈s kju:l☜ris] 肌层adventitia [ædven ti☞i☜] 外膜gastric gland [gæstrik glænd] 胃腺parietal cell [p☜raiitl sel] 壁细胞chief cell [t☞i:f sel] 主细胞mucosa neck cell颈粘液细胞intestinal villus [in testinl vil☜s] 小肠绒毛goblet cell [g :blit sel ] 杯状细胞small intestinal gland小肠腺paneth cell [pænis sel] 潘氏细胞消化腺Digestive Gland pancreas islet [ pænkri☜s ailit] 胰岛hepatic lobule [hi pætik l bju:l] 肝小叶hepatic plate [ hi pætik pleit] 肝板hepatic cord [ hi pætik k :rd] 肝索hepatocyte [hi pæt sait] 肝细胞hepatic sinusoid [hi pætik sains id] 肝血窦Kupffer cell 枯否细胞perisinusoidal space 窦周隙bile canaliculi [bail kæn☜likjuli] 胆小管portal area [p :tl ☪☯ri☜] 门管区呼吸系统Respiratory Systembronchial tree 支气管树pulmonary lobule 肺小叶bronchiole 细支气管terminal bronchiole 终末细支气管respiratory bronchiole 呼吸性细支气管alveolar duct肺泡管alveolar sac 肺泡囊pulmonary alveoli肺泡type I alveolar cell I型肺泡细胞type II alveolar cell II型肺泡细胞pulmonary surfactant肺泡表面活性物质alveolar septum肺泡隔alveolar pore 肺泡孔pulmonary macrophage肺巨噬细胞dust cell 尘细胞blood-air barrier 气血屏障泌尿系统Urinary Systemrenal pyramid 肾锥体medullary ray 髓放线cortical labyrinth 皮质迷路nephron 肾单位medullary loop 髓袢renal corpuscle 肾小体glomerulus 血管球renal capsule 肾小囊podocyte 足细胞slit membrane 裂孔膜filtration barrier 滤过屏障renal tubule 肾小管proximal tubule 近端小管distal tubule 远端小管thin segment 细段collecting tubule 集合小管juxtaglomerular complex 球旁复合体macula densa 致密斑ureter 输尿管urinary bladder 膀胱男性生殖系统Male Reproductive system seminiferous tubule 生精小管spermatogenic epithelium 生精上皮spermatogonium 精原细胞primary spermatocyte 初级精母细胞secondary spermatocyte 次级精母细胞spermatid 精子细胞spermatozoon 精子blood-testis barrier 血-睾屏障spermatogenesis 精子发生spermiogenesis 精子形成女性生殖系统Female Reproductive system primordial follicle 原始卵泡primary follicle 初级卵泡secondary follicle 次级卵泡mature follicle 成熟卵泡corpus luteum 黄体atresic follicle 闭锁卵泡endometrium 子宫内膜proliferation phase 增生期secretory phase 分泌期menstrual phase 月经期胚胎学Embryology fertilization [f☜:tilaize☞☜n] 受精fertilized ovum [ouv☜m] 受精卵cleavage [kli:vid✞] 卵裂morula [m☜rula] 桑椹胚blastocyst[blæst sist] 胚泡（囊胚）trophoblast[trof blæst] 滋养层inner cell mass 内细胞群implantation [impla:n’tei☞☜n] 植入embryonic disc [embri nik] 胚盘endoderm [end d☜:m] 内胚层ectoderm [ekt☜d☜:m] 外胚层mesoderm [mes☜d☜:m] 中胚层primitive streak [praimitiv strik] 原条notochord [nout☜k :d] 脊索neural tube [nju☜r l] 神经管placenta [pl☜’sent☜] 胎盘。

4Bile acids,obesity,and the metabolic syndromeHuijuan Ma,MD,Adult Endocrinologist a ,Mary Elizabeth Patti,MD,Investigator andAdult Endocrinologist b ,*aDepartment of Endocrinology and Metabolism,Hebei General Hospital,Shijiazhuang,Hebei 050051,China b Research Division,Joslin Diabetes Center,and Harvard Medical School,Boston,MA 02215,USAKeywords:Bile acidsFXRTGR5ObesityInsulin resistance a b s t r a c tBile acids are increasingly recognized as key regulators of systemic metabolism.While bile acids have long been known to play important and direct roles in nutrient absorption,bile acids also serve as signalling molecules.Bile acid interactions with the nuclear hormone receptor farnesoid X receptor (FXR)and themembrane receptor G-protein-coupled bile acid receptor 5(TGR5)can regulate incretin hormone and fibroblast growth factor 19(FGF19)secretion,cholesterol metabolism,and systemic energyexpenditure.Bile acid levels and distribution are altered in type 2diabetes and increased following bariatric procedures,in parallelwith reduced body weight and improved insulin sensitivity andglycaemic control.Thus,modulation of bile acid levels andsignalling,using bile acid binding resins,TGR5agonists,and FXRagonists,may serve as a potent therapeutic approach for thetreatment of obesity,type 2diabetes,and other components of themetabolic syndrome in humans.©2014Elsevier Ltd.All rights reserved.IntroductionBile is a mixture of bile acids (BAs),cholesterol,phosphatidylcholine,and bilirubin.Of these,BAs are essential constituents and play critical roles in regulation of metabolism in both humans and animal *Corresponding author.Joslin Diabetes Center,1Joslin Place,Boston,MA 02215,USA.Tel.:þ16173091966;fax:þ16173092593.E-mail address:Mary.Elizabeth.Patti@ (M.E.Patti).Contents lists available at ScienceDirectBest Practice &Research ClinicalGastroenterology/10.1016/j.bpg.2014.07.0041521-6918/©2014Elsevier Ltd.All rights reserved.Best Practice &Research Clinical Gastroenterology 28(2014)573e 583models.Bile acids have long been recognized to aid in the absorption of fat and fat-soluble vitamins and modulate cholesterol levels.However,recent data indicate that bile acids also play an important role in glucose and lipid homeostasis by activating both the nuclear receptor LXR and the cell surface receptor G protein-coupled bile acid receptor 5(TGR5)[1e 3].Moreover,modulation of plasma bile acid levels and the total bile acid pool can affect glycaemic control,body weight,and insulin sensitivity[4e 6].In this review,we will focus on the relation between bile acids and regulation of systemic meta-bolism and the potential for bile acids as a therapeutic approach for obesity,insulin resistance,type 2diabetes (T2D),and other components of the metabolic syndrome.Bile acid synthesis and regulationBile acid synthesisBAs are amphipathic molecules with a steroid backbone which are synthesized from cholesterol in hepatocytes.It is estimated that about half of the 800mg of cholesterol synthesized daily is used for bile acid synthesis,totalling about 200e 600mg daily in humans [7].Bile acids are synthesized from cholesterol through two dominant pathways:the classic pathway and the alternative pathway (Fig.1).In the classic (or neutral)pathway,CYP7A1catalyses the initial and rate-limiting step converting cholesterol into 7a -hydroxycholesterol,with CYP8B1subsequently regulating synthesis of 12a -hydroxysterols including cholic acid (CA).In the alternative (or acidic)pathway,CYP27A1first hydroxylates the cholesterol side chain,converting cholesterol into 27-hydroxycholesterol,which is then 7a -hydroxylated by CYP7B1prior to CYP8B1action.In humans,the classical pathway produces the primary BA cholic acid (CA)and chenodeoxycholic acid (CDCA)in Fig.1.Bile acid synthesis pathway.Cholesterol is converted to two primary bile acids in human liver,CA and CDCA.Key regulatory enzymes in these pathways include CYP7A1,CYP8B1,CYP27A1,and CYP7B1.CYP7A1initiates the classic (neutral)biosynthetic pathway,while CYP27A1initiates the alternative (acidic)pathway in liver and macrophages.CA and CDCA can be conjugated with glycine (G)and taurine (T).In the intestine,conjugated CA and CDCA are deconjugated and then dehydroxylated at the 7a -position to the secondary bile acids DCA and LCA,respectively.H.Ma,M.E.Patti /Best Practice &Research Clinical Gastroenterology 28(2014)573e 583574H.Ma,M.E.Patti/Best Practice&Research Clinical Gastroenterology28(2014)573e583575 roughly equal amounts,whereas the alternative pathway produces mainly CDCA[8].Most bile acids are conjugated with either glycine or taurine,with a3:1predominance of glycine over taurine[3,5,9].Synthesized BA are stored in the gallbladder and secreted into the duodenum in response to feeding,contributing to digestion of lipids and lipid-soluble vitamins.The primary BA CA and CDCA can be dehydroxylated at the7a position by gut microbiota to produce the secondary BAs,predominantly deoxycholic acid(DCA)and lithocholic acid(LCA)[10,11].Thus,bile acid levels and relative composition can be modulated by gut microbiota populations[12].In the terminal ileum,BAs are efficiently absorbed by both active transport and passive diffusion, transported back to the liver via the portal vein,taken up at the sinusoidal membrane of hepatocytes, and secreted into bile again.Each BA molecule may complete4e12cycles of this enterohepatic cir-culation per day[1,13].This process is highly efficient,as only about5%of bile acids are lost in feces [3,14].This process is similar in mice,although different bile acid species dominate.CDCA is efficiently converted into muricholic acid(MCA),and BAs are conjugated to taurine.Given that different BA have different structures,hydrophobicity,and affinities for membrane and nuclear receptors,interindividual differences in BA pool composition resulting from differential regulation of the complex BA synthesis pathway may have functional consequences for systemic metabolism.Regulation of Bile Acid SynthesisBA are potent regulators of their own synthesis,serving to limit excessive accumulation of BA in the circulation via multiple redundant pathways.Feeding bile acids to rats strongly reduces CYP7A1 enzyme activity and bile acid synthesis[8,15].This self-regulation of BA synthesis involves activation of the nuclear receptor FXR(farnesoid X receptor,official gene name NR1H4)[16].FXR knockout mice have increased BA synthesis and Cyp7a1expression,verifying the central role for FXR in mediating bile acid inhibition of Cyp7a1[17].FXR has both direct effects on Cyp7a1expression as well as indirect effects,mediated by induction of small heterodimer partner(SHP)which in turn inhibits trans-activation of CYP7A1and CYP8B1by the transcription factors Hepatocyte Nuclear Factor4a(HNF4a) and liver-related homolog-1(LRH-1)at the bile acid response element.FXR also increases BA conju-gation and upregulates expression of several genes which promote bile acid efflux from hepatocytes into the bile.An additional feedback repression mechanism involves FXR regulation of several indi-vidualfibroblast growth factors(FGFs).For example,bile acid activation of FXR results in secretion of FGF-19from hepatocytes and enterocytes.In turn,FGF-19can bind tofibroblast growth factor receptor 4(FGFR4)receptors on hepatocytes,leading to suppression of CYP7A1expression and bile acid syn-thesis via a SHP-independent,but c-Jun N-terminal kinase(JNK)dependent mechanism[18].Consis-tent with this mechanism,FGFR4knockout mice have increased expression of CYP7A1,in parallel with increased fecal bile acids and bile acid pool size[19].Other nuclear receptors,such as the pregnane X receptor(PXR)[20]and vitamin D receptor(VDR)[21],can also regulate BA synthesis by suppressing CYP7A1.Moreover,activation of the nuclear receptor ROR a can modulate12a-hydroxylase(CYP8B1) expression[22,23].BA differ markedly in their potency to activate FXR.The hydrophobic bile acid CDCA is the most potent BA ligand of FXR,followed by lithocholic acid(LCA),deoxycholic acid(DCA),and CA;by contrast, the hydrophilic bile acids ursodeoxycholic acid(UDCA)and muricholic acid(MCA)do not activate FXR[24].BA are also regulated in response to other elements of systemic metabolism.Early studies showed that the bile acid pool size is increased in insulin-deficient diabetic rats,with a3-fold increase in the cholic acid pool[25].Conversely,insulin treatment reduces bile acid pool size,inhibits CYP7A1and CYP8B1activity,and alters bile acid composition[26].CYP7A1can also be regulated by steroid hor-mones,activated protein kinase C,and proinflammatory cytokines[8].Bile acids and regulation of systemic metabolismFXR and TGR5signalling mechanisms appear to dominate for BA effects on regulation of glucose, lipid,and energy metabolism[1,27e30].As noted above,BA are natural ligands for FXR a,a nuclearreceptor highly expressed in liver,intestine,kidney and adrenal glands [17,27].BAs can also activate TGR5,a membrane-bound G protein-coupled BA receptor.TGR5is expressed in many organs and tissues,with high expression in macrophages/monocytes,placenta,gallbladder,liver and intestine[31,32].BA stimulation of energy expenditure in brown adipose tissue (BAT)and skeletal muscle ap-pears to be mediated via TGR5[2].Bile acids and lipid metabolismBAs exert an important regulatory role in cholesterol and triglyceride (TG)metabolism.Increased bile acid synthesis increases utilization of cholesterol as substrate.Bile acid synthesis rates are correlated with serum triglyceride levels in hyperlipidemic patients [33].Bile acid sequestrants or other interruptions in enterohepatic circulation also increase both bile acid and VLDL triglyceride synthesis.Conversely,CDCA-mediated increases in the BA pool size leads to inhibition of BA synthesis and reduced serum triglycerides in hyperlipidemic patients [34].These effects of BA to modulate TG metabolism are likely mediated via several distinct mechanisms,predominantly BA activation of FXR [35,36].FXR alters the transcription of several genes involved in fatty acid and triglyceride synthesis and lipoprotein metabolism.In mice,administration of FXR ago-nists [GW4064,6a-ethyl-chenodeoxycholic acid]reduces plasma triglyceride and cholesterol levels[37e 41]via repression of the lipogenic genes sterol-regulatory-element-binding protein-1c (SREBP1c)and fatty acid synthase (FAS)in liver [36].FXR also induces expression of peroxisome proliferator activated receptor (PPAR)a ,a nuclear receptor that promotes lipid oxidation [42],and of pyruvate dehydrogenase kinase,isoenzyme 4(PDK4),leading to inhibition of pyruvate dehydrogenase and increased fatty acid oxidation [43].Additional FXR target genes include the apolipoproteins A-V,C-III,apoE,syndecan-1,and the VLDL receptor [1,44].Conversely,FXR-null mice have higher serum TG levels and increased synthesis of apolipoprotein (apo)B-containing lipoproteins [17].Thus,bile acids play central roles in lipid metabolism and in the control of TG levels,in part via FXR and downstream transcriptional targets.Additional effects of BA on lipid metabolism may be independent of FXR.For example,the bile acid tauroursodeoxycholic acid (TUDCA)also acts as a chaperone,modulating endoplasmic reticulum stress[45].TUDCA reduces adipogenesis in human adipocyte stem cells [46].Similarly,in another study,UDCA (but not TUDCA)profoundly inhibits adipogenesis,in parallel with activation of extracellular regulated protein kinases 1and 2(ERK 1/2)[47].Bile acids and glucose metabolismBile acids are also implicated in regulation of glucose metabolism [48e 50].Increasing hepatic bile acid synthesis can inhibit gluconeogenesis and stimulate glycolysis.Effects on glycogen metabolism appear complex [51].Some studies have shown bile acids stimulate glycogen phosphorylase (GP)and glycogen breakdown to glucose-1-P [52],while other data indicate bile acids also activate glycogen synthesis (GS)[53].Additional effects of BA on glucose metabolism and insulin action may also be mediated via reductions in endoplasmic reticulum (ER)stress,a key mediator of insulin resistance [45].In addition to direct effects,many of the bene ficial effects of BA on glucose metabolism are mediated via FXR.FXR is an important regulator of glucose metabolism,as demonstrated by reduced plasma glucose and reduced hepatic glycogen levels in FXR-null mice [49,50,54].Conversely,activation of FXR is associated with increased phosphoenolpyruvate carboxykinase (PEPCK)and glucose-6-phosphatase expression and glucose output from primary rat hepatocytes.However,in vivo pharmacologic stimulation of FXR in two mouse models of obesity and T2D (db/db or KKA(y)mice)causes inhibition of gluconeogenesis,hypoglycemia,and increased insulin sensitivity [49,50].Thus,FXR may have a dominant effect to inhibit gluconeogenesis in diabetes,perhaps via inhibition of PEPCK by SHP-dependent inhibition of HNF4a and FoxO1[55].Activation of FXR can also stimulate the insulin/Akt pathway,promoting glycogen synthesis in liver [55],GLUT2activation in pancreatic b -cells [56],and improving insulin resistance in obese ob/ob mice [57].In this context,inhibition of gluconeogenesis,improved insulin action,and stimulation of glycogen syn-thesis may synergize to improve plasma glucose,insulin secretion,insulin sensitivity,and glucoseH.Ma,M.E.Patti /Best Practice &Research Clinical Gastroenterology 28(2014)573e 583576H.Ma,M.E.Patti/Best Practice&Research Clinical Gastroenterology28(2014)573e583577 tolerance.In parallel,however,GW4064increases susceptibility to high fat diet-induced obesity and diabetes[50,58].These complex data highlight the importance of the specific context in which FXR is activated.Beneficial effects of BA on glucose metabolism may also be mediated via TGR5.TGR5is expressed in many organs and tissues,with highest expression in macrophages/monocytes,placenta,gallbladder, liver and intestine[32].Activation of TGR5in enterocytes can stimulate the secretion of the incretin hormone glucagon-like peptide GLP1,promoting glucose-dependent insulin secretion[59,60].More-over,BA stimulation of cell-surface TGR5on neurons may also modulate GLP-1secretion[61].In pe-ripheral tissues,TGR5activation may increase activation of the type2deiodinase,resulting in increased active thyroid hormone,mitochondrial oxidative capacity,and energy expenditure[2].A recent human genetic study demonstrates that a single nucleotide polymorphism(SNP)at the TGR5 locus(rs3731859)is associated with BMI,waist circumference,intramyocellular lipids,and fasting GLP-1levels[62].Furthermore,TGR5-null mice have a25%reduction in bile acid pool size,and female Tgr5 null mice show increased weight gain and fat accumulation when fed a high fat diet[63].Further support for the regulatory role of TGR5in glucose homeostasis comes from thefinding that TGR5 agonists decrease blood glucose in animals[64].Bile acids in humansIn healthy individuals,BA levelsfluctuate with cycles of fasting and refeeding.BA robustly increase in response to an oral glucose load[65].Such responses are increased in insulin sensitive individuals and are blunted in individuals with prediabetes[55],suggesting dysregulation of bile acid pools and/or intestinal regulation in insulin resistance.The majority of studies have analysed bile acid levels and species distribution in the fasting state; many of these demonstrate alterations in insulin resistance and diabetes.For example,one study demonstrated1.6-fold increases in deoxycholic acid(DCA)in T2D[66].Similarly,Haeusler et al. demonstrated that12a e hydroxylated species(sum of CA,DCA,and their conjugates)are significantly higher in patients with T2D[67].Furthermore,ratios of12-hydroxylated/non e12-hydroxylated BAs are associated with key features of IR,including higher insulin,glucose,and triglyceride(TG)levels and lower HDL cholesterol[67].Brufau and colleagues showed that individuals with T2D had higher cholic acid(CA)synthesis rates and enlarged DCA pool size[68].Our group recently reported that concen-trations of total taurine-conjugated BA were higher in T2D and intermediate in individuals with impaired glucose tolerance[69].Interestingly,total taurine-BA were positively associated with fasting and post-load glucose levels,fasting insulin,and HOMA-IR.However,insulin-mediated glycaemic improvement in T2D patients did not change fasting serum total BA,or BA composition,suggesting dysregulation of BA levels is not directly linked to glycaemic burden but possible due to other aspects of insulin resistance or the metabolic syndrome.Bile acids are increased following bariatric surgeryBariatric surgery is increasingly recognized as a robust method to not only reduce body weight,but also to reduce glycaemia and medication requirement(so called diabetes‘remission’)[70].Bariatric surgical procedures include Roux-en-Y gastric bypass(RYGB),vertical sleeve gastrectomy(VSG), laparoscopic adjustable gastric banding(LAGB)and biliopancreatic diversion(BPD).In this context,it is interesting that several studies have demonstrated that bile acids are markedly increased following bariatric surgery[5,6,71,72].Interestingly,total bile acids in post-bypass patients are correlated with improvement in several key metabolic parameters;bile acids are inversely correlated with post-prandial glucose,triglycerides,and positively correlated with adiponectin and peak GLP1levels following a mixed meal test.These intriguing data suggest increased serum bile acid levels could contribute to improvements in insulin sensitivity,incretin secretion,and postprandial glycaemia in response to bariatric surgery[6].Despite these intriguing data,it remains uncertain whether increased BA are absolutely essential for metabolic improvements following bariatric surgery,particularly during the early postoperative period.For example,longitudinal studies in humans demonstrate that increases in BA are not detecteduntil 1year postoperatively [73],despite improved glucose levels and reduced hepatic glucose output within one week of surgery [74,75].By contrast,increases in both fasting and postprandial BA are also observed as early as 14days following VSG in rodents [76,77].In addition,both RYGB and VSG can alter the distribution of bile acid species [8][78].By contrast,circulating BA do not change signi ficantly after LAGB [4,73,79],and could contribute to reduced ef ficacy of this procedure for long-term resolution of T2D as compared with RYGB or VSG.Importantly,increased BA action,via FXR-mediated effects on both gut microbiome and transcriptional pathways,appears to be required to achieve the metabolic effects of surgery,at least in rodent models [77].Intriguingly,direct modulation of small intestinal anatomy can alter bile acid levels and composi-tion.Mid-to-distal small intestinal resection,with preservation of the terminal ileum,increases bile acid levels [80].Similarly,interposition of the ileum into more proximal segments of gut also increases bile acid levels [81].Nonsurgical approaches may also be effective;for example,endoluminal sleeves,which allow luminal contents to bypass the duodenal mucosa,also increase bile acids in rodents and improve glucose metabolism [82].While there are many unanswered questions,these data suggest that modulation of segmental intestinal anatomy,and/or changes in intestinal flora which result from these modi fications,may improve systemic metabolism via BA-dependent mechanisms.Modulating bile acids as a potential therapeutic approach for obesity and T2DThe close links between plasma levels of bile acids and a host of key metabolic parameters raise the possibility that modulation of BA could be used as a therapeutic approach for the treatment of metabolic diseases.Dietary supplementation with cholic acid (CA)increases energy expenditure,reducing weight gain during high-fat feeding [83].While the precise mechanism remains uncertain,BA increase expression and activity of the type 2iodothyronine deiodinase (D2)via a TGR5-cAMP-mediated pathway,thus increasing active thyroid hormone (T3)availability and energy expenditure in tissues critical for thermogenesis,such as BAT [10].Similarly,CDCA has been shown to increase energy expenditure by induction of UCP1and activation of thermogenesis in BAT in mice [84].Bile acid sequestrants,which increase the CA pool size (while reducing CDCA and DCA pools),also reduce glucose,hemoglobin A1c,and cholesterol levels in patients with type 2diabetes [85e 87].Thus,therapeutic strategies which increase circulating BA levels or modulate relative distribution of active bile acid species could be an effective approach to improving systemic metabolism.Bile acid-binding resinsBile acid binding resins (BABR)(cholestyramine,colestipol,colestimide,and colesevelam)are positively charged nondigestible resins that bind to bile acids in the intestine to form an insoluble complex that is excreted in the feces.BABR have been successfully employed for the treatment of hypercholesterolaemia for many years.Recently,the second-generation BABR colestimide and cole-sevelam were found to improve glycaemic control in patients with T2D [86,88e 91].Several possible mechanisms have been proposed for these effects.BABR not only increase plasma BA but also change the composition of the BA pool [92].These changes in BA may promote increased energy expenditure,as observed in rodents [93].In addition,BABRs stimulate the expression of proglucagon and release of GLP1,thus improving incretin-mediated insulin secretion and reducing plasma glucose [94].This effect of colesevelam was achieved through regulation of FXR/FGF19and TGR5/GLP-1signalling pathways[89].Colestimide also decreases postprandial plasma glucose and increases GLP-1secretion in patients with T2D [85].Despite these intriguing possibilities,there is not a clear relationship between BA metabolism and improvements in glycaemia,and changes in FGF19or energy expenditure are not consistently observed in human studies [68,95].Nevertheless,BA sequestrants regulate glucose ho-meostasis,potentially at least in part by modulating FXR-and TGR5-mediated pathways.FXR agonistsSince FXR modulates many of the metabolic effects of bile acids,activation of FXR could be another approach to treat metabolic disease [96].Treatment with the FXR ligand GW4064signi ficantlyH.Ma,M.E.Patti /Best Practice &Research Clinical Gastroenterology 28(2014)573e 583578H.Ma,M.E.Patti/Best Practice&Research Clinical Gastroenterology28(2014)573e583579 decreases plasma glucose,triglycerides,and cholesterol in both wild-type and diabetic db/db mice [50].6-ECDCA,another FXR agonist,can decrease glucose,cholesterol,free fatty acid,and triglyceride levels in Zucker fa/fa rats[54]by enhancing glucose disposal,reducing body weight,in parallel with reduced expression of PEPCK and glucose-6-phosphates(G6Pase)in ob/ob mice.Interestingly,these effects of FXR agonists are achieved despite decreased bile acid levels,largely through direct activation of hepatic FXR-SHP and intestinal FXR-FGF15/19pathways[96].These data again highlight the importance of FXR pathways in mediating metabolic effects of BA.TGR5agonistsThe cell surface receptor TGR5represents another novel pharmacological target for the treat-ment of the metabolic syndrome and related disorders[97,98].TGR5can be activated by either synthetic ligands[10,80],or by BA in a dose-dependent manner,with potency of activation depending on specific BA species:lithocholic acid>deoxycholic acid>chenodeoxycholic acid >cholic acid.In rodents,synthetic TGR5agonists decrease plasma glucose and insulin levels and protect against weight gain induced by a high-fat diet[64].The expression and activity of the type2 deiodinase and energy expenditure can be increased with incubation of skeletal muscle with a synthetic TGR5agonist[10].Therapeutic modulation of BA via manipulation of the intestinal tract and its residentflora As noted above,bariatric surgery is associated with increased plasma BA levels and alterated composition of BA species.In turn,increased BA may contribute to enhanced FXR and TGR5acti-vation,leading to reduced hepatic glucose production,increased GLP1secretion,and increased systemic energy expenditure.In support of this concept,recent data implicate FXR activity as a critical mediator of the beneficial effects of bariatric surgery in rodents[85].Given the important role of the gut microbiome in modulating bile acid pool size,composition,and enterohepatic recirculation,it is intriguing to consider whether modification of the gut microbiome could pro-mote BA-mediated beneficial changes in metabolism.This could potentially be achieved by altered dietary fatty acid composition[99]or with probiotics which modulate gutflora[100,101].For example,one recent study demonstrated that probiotics could increase BA deconjugation,increase fecal BA excretion,and increase hepatic BA synthesis in an FGF-dependent mechanism[101].These intriguing data provide hope that BA action on the gut microbiome and systemic metabolism could be harnessed to yield beneficial effects similar to those observed after bariatric surgery,but in the absence of invasive surgery.This line of investigation will be an important question for future studies.SummaryTaken together,data from both humans and preclinical animal models demonstrate that BA are important signalling molecules which contribute to regulation of whole-body glucose and lipid metabolism and body weight.Such effects of BA are largely mediated by the nuclear receptor FXR and the G protein-coupled receptor TGR5.Future research employing proteomic,metabolomic,and lip-idomic approaches are likely to help in identifying bile acid-related biomarkers which may be useful for predicting and assessing response to BA-related therapy for human obesity and metabolic syn-drome.Moreover,novel approaches to altering biliaryflow and enterohepatic recirculation,plasma BA levels,composition of BA pools,and downstream effectors of BA signalling pathways should be pur-sued,as they may be effective strategies for the management of obesity,insulin resistance,type2 diabetes,and other components of the metabolic syndrome.Conflict of interestNone.AcknowledgementsThe authors gratefully acknowledge grant support from Hebei Health Department of Scienti fic Research Fund (to HM),P30DK036836DRC (Joslin Diabetes Center),and the American Diabetes As-sociation (to MEP).References[1]Houten SM,Watanabe M,Auwerx J.Endocrine functions of bile acids.EMBO J 2006;25:1419e 25.[2]Watanabe M,Houten SM,Mataki C,Christoffolete MA,Kim BW,Sato H,et al.Bile acids induce energy expenditure bypromoting intracellular thyroid hormone activation.Nature 2006;439:484e 9.[3]Chiang JY.Bile acid metabolism and pr Physiol 2013;3:1191e 212.[4]Kohli R,Bradley D,Setchell KD,Eagon JC,Abumrad N,Klein S.Weight loss induced 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联合克拉屈滨预处理方案在难治／复发性急性髓系白血病患者异基因造血干细胞移植中的疗效观察王晴晴，夏凌辉ＯｂｓｅｒｖａｔｉｏｎｏｆｃｕｒａｔｉｖｅｅｆｆｅｃｔｏｆｃｏｍｂｉｎｅｄｔｈｅｒａｐｙｗｉｔｈｃｌａｄｒｉｂｉｎｅｉｎａＩｌｏｇｅｎｅｉｃｈｅｍａｔｏ ｐｏｉｅｔｉｃｓｔｅｍｃｅｌｌｔｒａｎｓｐｌａｎｔａｔｉｏｎｏｆｒｅｆｒａｃｔｏｒｙ／ｒｅｌａｐｓｅｄａｃｕｔｅｍｙｅｌｏｉｄｌｅｕｋｅｍｉａｐａｔｉｅｎｔｓＷＡＮＧＱｉｎｇｑｉｎｇ，ＸＩＡＬｉｎｇｈｕｉＩｎｓｔｉｔｕｔｅｏｆＨｅｍａｔｏｌｏｇｙ，ＵｎｉｏｎＨｏｓｐｉｔａｌ，ＴｏｎｇｊｉＭｅｄｉｃａｌＣｏｌｌｅｇｅｏｆＨｕａｚｈｏｎｇＵｎｉｖｅｒｓｉｔｙｏｆＳｃｉｅｎｃｅａｎｄＴｅｃｈｎｏｌｏｇｙ，ＨｕｂｅｉＷｕｈａｎ４３００２２，Ｃｈｉｎａ．【Ａｂｓｔｒａｃｔ】　Ｏｂｊｅｃｔｉｖｅ：Ｔｏｅｖａｌｕａｔｅｔｈｅｅｆｆｉｃａｃｙｏｆｃｏｍｂｉｎｅｄｔｒｅａｔｍｅｎｔｗｉｔｈｃｌａｄｒｉｂｉｎｅｉｎｔｈｅｔｒｅａｔｍｅｎｔｏｆｒｅｆｒａｃｔｏｒｙ／ｒｅｌａｐｓｅｄａｃｕｔｅｍｙｅｌｏｉｄｌｅｕｋｅｍｉａ（ＡＭＬ）ｗｉｔｈａｌｌｏｇｅｎｅｉｃｈｅｍａｔｏｐｏｉｅｔｉｃｓｔｅｍｃｅｌｌｔｒａｎｓｐｌａｎｔａｔｉｏｎ（ａｌｌｏ－ＨＳＣＴ）．Ｍｅｔｈｏｄｓ：Ｔｈｅｃｌｉｎｉｃａｌｄａｔａｏｆ１７ｐａｔｉｅｎｔｓｗｉｔｈｒｅｆｒａｃｔｏｒｙ／ｒｅｌａｐｓｅｄＡＭＬｗｈｏｕｎｄｅｒｗｅｎｔｃｏｍｂｉｎｅｄｃｌａｄｒｉｂｉｎｅｐｒｅｔｒｅａｔｍｅｎｔｐｌａｎａｎｄｒｅｃｅｉｖｅｄａｌｌｏ－ＨＳＣＴｉｎｔｈｅｌａｍｉｎａｒｆｌｏｗｗａｒｄｏｆｏｕｒｈｏｓｐｉｔａｌｆｒｏｍＡｐｒｉｌ２０１７ｔｏＯｃｔｏｂｅｒ２０１９ｗｅｒｅｃｏｌｌｅｃｔｅｄａｎｄａｎａｌｙｚｅｄ．Ｒｅｓｕｌｔｓ：Ｈｅｍａｔｏｐｏｉｅｔｉｃｒｅｃｏｎｓｔｒｕｃｔｉｏｎｗａｓｃｏｍｐｌｅｔｅｄｉｎａｌｌｔｈｅ１７ｐａｔｉｅｎｔｓ，ａｎｄｔｈｅｍｅｄｉａｎｔｉｍｅｏｆｇｒａｎｕｌｏｃｙｔｅｉｍｐｌａｎｔａｔｉｏｎｗａｓ１２ｄ（９～２０ｄ），ａｎｄｔｈｅｍｅｄｉａｎｔｉｍｅｏｆｐｌａｔｅｌｅｔｉｍｐｌａｎｔａｔｉｏｎｗａｓ１１ｄ（９～３０ｄ）．Ｄｕｒｉｎｇｔｈｅｐｒｅｔｒｅａｔｍｅｎｔ，ｏｎｅｐａｔｉｅｎｔｄｅｖｅｌｏｐｅｄｈｅｍｏｒｒｈａｇｉｃｃｙｓｔｉｔｉｓ．４ｐａｔｉｅｎｔｓｏｃｃｕｒｒｅｄⅠ－ⅡｄｅｇｒｅｅｏｆａＧＶＨＤ．Ａｔｔｈｅｅｎｄｏｆｆｏｌｌｏｗ－ｕｐ，８ｏｆｔｈｅ１１ｓｕｒｖｉｖｉｎｇｐａｔｉｅｎｔｓｈａｄｌｏｃａｌｉｚｅｄｃＧＶＨＤ．Ｆｉｖｅｐａｔｉｅｎｔｓｒｅｌａｐｓｅｄａｆｔｅｒｔｒａｎｓｐｌａｎｔａｔｉｏｎ，ｗｉｔｈａｍｅｄｉａｎｒｅｃｕｒｒｅｎｃｅｔｉｍｅｏｆ４ｍｏｎｔｈｓ（２～１９ｍｏｎｔｈｓ）．Ｔｈｅ１－ｙｅａｒｏｖｅｒａｌｌｓｕｒｖｉｖａｌｒａｔｅｗａｓ５７．２％，ａｎｄｔｈｅ１－ｙｅａｒｄｉｓｅａｓｅ－ｆｒｅｅｓｕｒｖｉｖａｌｒａｔｅｗａｓ５５．２％．Ｃｏｎｃｌｕｓｉｏｎ：Ｔｈｅｓｈｏｒｔ－ｔｅｒｍｅｆｆｅｃｔｏｆｃｌａｄｒｉｂｉｎｅｃｏｍｂｉｎｅｄｗｉｔｈｐｒｅｔｒｅａｔｍｅｎｔｉｓｂｅｔｔｅｒ，ｗｉｔｈｏｕｔｉｎｃｒｅａｓｉｎｇｔｈｅａｄｖｅｒｓｅｒｅａｃｔｉｏｎｓｒｅｌａｔｅｄｔｏｐｒｅｔｒｅａｔｍｅｎｔ．Ｉｔｃａｎｅｆｆｅｃｔｉｖｅｌｙｉｍｐｒｏｖｅｔｈｅｓｕｒｖｉｖａｌｒａｔｅｏｆｐａｔｉｅｎｔｓａｎｄｉｍｐｒｏｖｅｔｈｅｐｒｏｇｎｏｓｉｓｏｆｐａｔｉｅｎｔｓ．【Ｋｅｙｗｏｒｄｓ】ｃｌａｄｒｉｂｉｎｅ，ａｌｌｏｇｅｎｅｉｃｈｅｍａｔｏｐｏｉｅｔｉｃｓｔｅｍｃｅｌｌｔｒａｎｓｐｌａｎｔａｔｉｏｎ，ａｃｕｔｅｍｙｅｌｏｉｄｌｅｕｋｅｍｉａＭｏｄｅｒｎＯｎｃｏｌｏｇｙ２０２１，２９（０８）：１４１９－１４２２【摘要】　目的：评价联合克拉屈滨预处理方案在异基因造血干细胞移植（ａｌｌｏ－ＨＳＣＴ）治疗难治／复发性急性髓系白血病（ＡＭＬ）中的疗效。

To locate English-language literature regarding the solubilization of proteins by arginine, several avenues can be pursued for search and acquisition:1.Academic Databaseso PubMed: PubMed, maintained by the National Library of Medicine (NLM) in the United States, is a comprehensive biomedical literature database. By entering keywords such as "arginine" and "protein solubilization" or "protein refolding," relevant English-language articles can be retrieved.o Google Scholar: Google Scholar serves as another powerful academic search engine, indexing a vast array of academic resources including scholarly articles, theses, and book chapters. Similar keyword combinations can be utilized to uncover pertinent English-languagepublications.2.Scientific Journalso Journal of Biological Chemistry: As a top-tier journal in the field of biochemistry, it frequently publishes research papers on protein structure and function, biomolecular interactions, and related topics. Access the journal's official website or search within academic databases to find articles on arginine-mediated protein solubilization.o Protein Expression and Purification: This journal specializes in the expression, purification, and characterization of proteins, and it may also contain articles related to the solubilization of proteins by arginine.3.Academic Institutions and University LibrariesMany academic institutions and university libraries possess extensive literature collections, both in print and electronic formats. Visit the library website of your institution or university and utilize the library's online catalog (e.g., OPAC) to conduct searches. Libraries often offer interlibrary loan services, enabling the borrowing of materials from other libraries if unavailable locally.4.Professional Websites and ForumsSome professional websites and forums may provide access to English-language literature or discussions on arginine-mediated protein solubilization. These platforms, maintained by industry experts or research institutions, serve as a venue for researchers to exchange ideas and share research findings.5.Optimized Search Engine QueriesWhen using search engines like Google, experiment with more specific keyword combinations or enclose phrases in quotation marks for precise searches. Additionally, leverage advanced search features to filter results by document type (e.g., academic articles), publication year, and other criteria.SummaryTo access English-language literature on the solubilization of proteins by arginine, a multifaceted approach is recommended, encompassing academic databases, scientific journals, academic institutions and university libraries, professional websites and forums, and optimized search engine queries. By leveraging these various channels, a more comprehensive and accurate collection of literature resources can be obtained.。

Compare drug release profiles of water poor soluble drugs from a novel chitosan and polycarbophil interpolyelectrolytes complexation (PCC) and hydroxylpropyl - methylcellulose (HPMC) based matrix tabletsZhilei Lu*, Weiyang Chen, Eugene Olivier, Josias H., HammanDepartment of Pharmaceutical Sciences, Tshwane University of Technology, Private Bag X680, P retoria, 0001, South Africa资料个人收集整理，勿做商业用途*Corresponding author:Zhilei Lu (Dr.)Department of Pharmaceutical Sciences,Tshwane University of Technology,Private Bag X680,Pretoria, 0001,South Africa (e-mail: luzj@tut.ac.za)AbstractThe aim of this study was to compare the drug release behaviours of water poor soluble drugs from an interpolyelectrolyte complex (IPEC) of chitosan with polycarbophil (PCC) and hydroxylpropylmethylcellulose (HPMC) based matrix tablets. A novel interpoly - electrolyte complex (IPEC) of chitosan with polycarbophil (PCC) was synthesized and characterized. Water poor soluble drugsHydrochlorothiazide and Ketoprofen were used in this study as model drugs.Polymers (including PCC, HPMC K100M and HPMC K100LV) based matrix tablets drug controlled release system were prepared using direct compression method.The results illustrate PCC based-matrix tablets offer a swelling controlled release system for water poor soluble drug and drug release mechanism from this matrix drug delivery system can be improved by addition of microcrystalline cellulose (Avicel).Analysis of the in vitro release kinetic parametersof the matrix tablets, PCC based matrix tablets exhibited similar or higher drug release exponent (n) and mean dissolution time (MDT) values compared to the HPMC based matrix tablets. It demonstrated that PCC polymer can be successfully used as a matrixcontrolled release system for the water poor soluble model drugs such as hydroxylpropylmethylcellulose (HPMC). 资料个人收集整理，勿做商业用途1 IntroductionOver the last three decades years, as the expense and complications involved in marketing new drug entities have increased, with concomitant recognition of the therapeutic advantages of controlled drug delivery, greater attention has been focused on the development of novel and controlled release drug delivery systemsto provide a long-term therapeutic of drugs at the site of action following a single dose (Mandal, 2000; Jantzen and Robinson, 2002). Many formulation techniques have been used tobuild t”he barrier into the peroral dosage form to provide slow release of the maintenance dose. These techniques include the use of coatings, embedding of the drug in a wax, polymeric or plastic matrix, microencapsulation, chemical bindingto ion-exchange resins and incorporation into an osmotic pump (Collett and Moreton, 2002:293). Among different technologies used in controlled drug delivery, polymeric matrix systems are the most majority because of the simplicity of formulation, ease of manufacturing, low cost and applicability to drugs with wide range of solubility (Colombo, et al., 2000; Jamzad and Fassihi, 2006). 资料个人收集整理，勿做商业用途Drugs release profiles from polymeric matrix system can influence by different factors, but the type, amount, and physicochemical properties of the polymers used play a primary role (Jamzad and Fassihi, 2006). Hydroxylpropyl-methylcellulose (HPMC) is the most important hydrophilic carrier material used for oral drug sustained delivery systems (Colombo, 1993; Siepmann and Peppas, 2001). BecauseHPMC is water soluble polymer, it is generally recognized that drug release fromHPMC matrices follows two mechanisms, drug diffusion through the swelling gellayer and release by matrix erosion of the swollen layer (Ford et al., 1987; Raoet al., 1990; Colombo, 1993; Tahara et al., 1995; Reynolds, Gehrke et al., 1998; Siepmann et al., 1999; Siepmann and Peppas, 2001). However, diffusion, swelling and erosion are most important rate-controlling mechanisms of commercial available controlled release products (Langer and Peppas, 1983), the major advantages of swelling/erosionHPMC based matrix drug delivery system are: (i) minimum the drug burst release; (ii) the different physicochemical drugs release rate approach a constant; (iii) the possibility to predict the effect of the device design parameters (e.g. shape, size and composition of HPMC-based matrix tablets) on the resulting drug release rate, thus facilitating the development of new pharmaceutical products (Colombo, 1993;Siepmann and Peppas, 2001资).料个人收集整理，勿做商业用途Interpolyelectrolyte complexes (IPEC) are formed as precipitates by two oppositely charged polyelectrolytes in an aqueous solution, have been reported as a new class of polymer carriers, which play an important role in creating new oral drug delivery systems (Peppas and Khare, 1993; Berger et al., 2004). A variety chemical structure and stoichiometry of both components in interpolyelectrolyte complexes depends onthe pH values of the media, ionic strength, concentration, mixing ratio, and temperature (Peppas, 1986; Dumitriu and Chornet, 1998; Berger et al., 2004;Moustafine et al., 2005a). Chitosan is a positively charged (amino groups) deacetylated derivative of the natural polysaccharide, chitin (Paul and Sharma, 2000).Chitosan has already been successfully used to form complexes with natural anionic polymers such as carboxymethylcellulose, alginic acid, dextran sulfate,carboxymethyl dextran, heparin, carrageenan,pectin methacrylic acid copolymers ? (Eudragit polymers) and xanthan (Dumitriu and Chornet, 1998, Berger et al., 2004,Sankalia et al., 2007, Margulis and Moustafine, 2006)资. 料个人收集整理，勿做商业用途In this study, a novel polymer - IPEC between chitosan and polycarbophil (PCC) was synthesized, characterized and used as direct compressedexcipients in the matrix tablet. Although it have been well known that various IPEC have been used as a polymer carriers in drug controlled release system (Peppas and Khare, 1993, Garcia and Ghaly, 1996, Lorenzo-Lamoza et al., 1998, Soppirnath and Aminabhavi, 2002,Chen et al., 2004, Nam et al., 2004, Moustafine et al., 2005b), IPEC chitosan and polycarbophil was used as a polymer carriers have been investigated by Lu et al., (2006, 2007a, 2007 b, 2008a; 2008b资料个人收集整理，勿做商业用途The aim of this study was to comparein vitro water poor soluble drugs release profile of HPMC based matrices system to PCC based matrices system at same formulation.Water poor soluble model drugs Hydrochlorothiazide and ketoprofen were used in thisstudy. Two types HPMC (K100M and K100LV) and PCC polymers were used indirect compressedpolymers based matrix drug release system. The results of the hydration and erosion studies showed PCC based matrix systems have superior swelling properties. Drug release exponent (n) of each formulation PCC based matrices tablets are higher than HPMC based matrices tablets at pH 7.4 buffer solutions. It demonstrated that PCC has high potential to use in polymer based matrix drug con trolled released delivery for water poor soluble drugs资料个人收集整理，勿做商业用途2. Materials and methods2.1 MaterialsChitosan (Warren Chem Specialities, South Africa, Deacetylation Degree =91.25%),Polycarbophil (Noveon, Cleveland, USA), Hydroxylpropylmethylcellulose (MethocelK100M, K100LV Premium, Colorcon Limited, Kent, England), Ketoprofen (Changzhou Siyao Pharma. China), Hydrochlorothiazide (Huzhou Konch Pharmaceutical Co., Ltd. China), Microcrystalline cellulose (Avicel, pH101, FMC corporation NV, Brussels, Belgium), Sodium carboxymethyl starch (Explotab, Edward Mendell Co., Inc New York, USA). All other chemicals were of analytical grade and used as receive资料个人收集整理，勿做商业用途2.2 Preparation of interpolyelectrolytes complexation between chitosan and P olycarb op hil (PCC)资料个人收集整理，勿做商业用途Chitosan (30 g) was dissolved in 1000 ml of a 2% v/v acetic acid solution andpolycarbophil (30 g) was dissolved in 1000 ml of a 2% v/v acetic acid solution. Thechitosan solution was slowly added to the polycarbophil solution underhomogenisation (5200 rpm, ZKA , Germany) over a period of 20 minutes. Themixture was then mechanically stirred for a period of 1 hour at a speed of 1200 rpm(Heidolph RZR2021, Germany). The gel formed was separated by centrifuging for 5 min at 3000 rpm and then washed several times with a 2% v/v acetic acidsolution toremove any unreacted polymeric material. The gel was freeze dried for a period of 48 hours (Jouan LP3, France) and the lyophilised powder was screened through a 300prn sieve资料个人收集整理，勿做商业用途2.3Differential scanning calorimetry (DSC)DSC thernograns of the PCC were recorded with a Shinadzu DSC50 (Kyoto, Japan) instrument. The thermal behaviour was studied by sealing 2 mg samples of the material in aluminium crimp cells and heating it at a heating rate of 10o C per min under the flow of nitrogen at a flow rate of 20 ml/min. The calorimeter was calibrated with 2 mg of indium (Kyoto, Japan, melting point 156.4o C) at a heating rate of 10o C per min.资料个人收集整理，勿做商业用途2.4Fourier transforn infrared (FT-IR)Fourier transforn infrared (FT-IR) spectral data of the PCC polyner was obtained on a FTS-175C spectrophotoneter (BIO-RAD, USA) using the KBr disk nethod. 资料个人收集整理，勿做商业用途2.5Preparation of the natrix tabletsIn order to conpare the release profiles of water poor soluble drugs fron polyner based natrix tablets, nonolithic natrix type tablets containing hydrochlorothiazide or ketoprofen were prepared by conpressing a nixture of the ingredients with varying concentrations of the PCC, HPMC K100M and HPMC K100LV as indicated in Table 1. The ingredients of the different fornulationswere nanually pre-nixed by stirring in a 1000 nl glass beaker for 30 ninutes with a spatula. After the addition of 0.05 g of nagnesiun stearate (0.5% w/w), the powder nass was nixed for 10 nin. The powdernixture was conpressed using a rotating tablet press (Cadnach, India) fitted with round, shallow pun ches to p roduce matrix type tablets with a 6 mm diameter资料个人收集整理，勿做商业用途26 Weight, hard ness, thick ness and friability of tablets资料个人收集整理，勿做商业用途Weight variation was tested by weighing 20 randomly selected tablets individually, the n calculati ng the average weight and comparing the in dividual tablet weights to the average. The specification for weight variation is 10% from the average weight if the average weight < 0.08 g (USP 2006资料个人收集整理，勿做商业用途The hardnessof ten randomly selected matrix type tablets of each formulation was determined using a hardness tester (TBH 220 ERWE K A, Germany). The force (N) n eeded to break each tablet was recorded料个人收集整理，勿做商业用途The thick ness of each of 10 ran domly selected matrix type tablets were measured witha vernier calliper (accuracy = 0.02 mm). The thickness of the tablet should be within 5% variation of the average value资料个人收集整理，勿做商业用途A friability test was con ducted on the tablets using an Erweka Friabilator (TA3R,Germany). Twenty matrices were randomly selected from each formulation and any loose dust was removed with the aid of a soft brush. The selected tablets were weighed accurately and p laced in the drum of the friabilator. The drum was rotated at 25 rpm for 4 minu tes after which the matrices were removed. Any loose dust was removed from the matrices before they were weighed again. The friability maximal limit is 1% (USP 2006) was calculated using the following equation资料个人收集整理，勿做商业用途F (%) = W before (g)「W曲(g)X 100%(1)W after (g)Where F is the friability, W before is the initial weight of the matrices and W after is the weight of the matrices after test ing资料个人收集整理，勿做商业用途2.7 Swelli ng and erosi on studiesSwelling and erosion studies were carried out for all formulations matrix tablets. The matrices were weighed in dividually before they were pl aced in 900 ml p hos phate buffer (pH 7.4) at 37.0 0.寸C.± The medium was stirred with a paddle at a rotation speed of 50 rpm in a USP dissolution flask. At each time point, three tablets of each formulatio n were removed from the dissoluti on flask and gen tly wiped with a tissue toremove surface water, weighed and then placed into a plastic bowel. The matrix tablets were dried at 60°C until constant weight was achieved. The mean weights were determ ined for the three tablets at each time in terval. The data obta ined from this exp erime nt was used to calculate the swelli ng in dex and p erce ntage mass loss.料个人收集整理，勿做商业用途2.7.1Swelli ng indexThe swelli ng in dex (or degree of swelli ng) was calculated accord ing to the followi ngequati on资料个人收集整理，勿做商业用途s,=WJ—00W dWhere SI is the swelling index, W s and W d are the swollen and dry matrix weights, resp ectively, at immersio n timet in the buffer soluti on.资料个人收集整理，勿做商业用途2.7.2P erce ntage of matrix erosi onThe p erce ntage of matrix erosi on is calculated in relatio n to the in itial dry weight of the matrices, accord ing to the followi ng equation 资料个人收集整理，勿做商业用途Erow 册件"00%Where: dry weight (t) is the weight at time t.28 Assay of hydrochlorothiazide and ket oprofen in matrix tablets.料个人收集整理，勿做商业用途The drug content of the matrix type tablets was determ ined by crush ing 10 ran domly selected tablets from each formulatio n in a mortar and p estle. App roximately 80 mg po wder from the hydrochlorothiazide or ket oprofen containing matrices were weighed accurately and individually transferred into a 200 ml volumetric flasks, which were then made up to volume with p hos phate buffer soluti on (pH 7.4). This mixture was stirred for 30 minutes to allow compiete release of the drug. After filtration through a 0.45 阿filter membrane, the solution was assayed using ultraviolet (UV) spectrophotometry (Helios a Thermo , England) at a wavelength of 271 nm for hydrochlorothiazide and 261 nm for ketoprofen. The assay for drug content wasperformed in triplicate for each formulation. The percentage drug content of the tablets was calculated by mea ns of the followi ng equation:料个人收集整理，勿做商业用途DC (% w/w^W dru^x100%WmtWhere DC is the drug content, W drug is the weight of the drug and W mt is the weight of the matrix tablet .资料个人收集整理，勿做商业用途2.9 Release an alysisThe USP (2006) dissoluti on app aratus 2 (i.e. p addle) was used to determ ine the in vitro drug release from the different polymers based matrix tablets. The dissolution medium (900 ml) consisted of phosphate buffer solution (pH 7.4) at 37 0.5 o C and a ± rotation speed of 50 rpm was used. Three hydrochlorothiazide or ketoprofen matrix tablets of each formulatio n were in troduced into each of three dissoluti on vessels (i.e.?in triplicate) in a six station dissolution apparatus (TDT-08L, Electrolab , India).Samp les (5 ml) were withdraw n at sp ecially in tervals, and 5 ml of p reheated dissolution medium was replaced immediately. Sink conditions were maintained throughout the study. The samp les were filtered through a 0.45 阿membra ne, hydrochlorothiazide or ketoprofen content in the solution was determined using ultraviolet (UV) spectrop hotometry at a wavele ngth of 271 or 261 nm, res pectively.An alyses were p erformed in tripli cate资料个人收集整理，勿做商业用途2.9.1 Kin eticCon trolled release drug delivery systems may be classified accord ing to their mecha ni sms of drug release, which in cludes diffusi on-con trolled, dissoluti on con trolled, swelli ng con trolled and chemically con trolled systems (La nger et al., 1983). Drug release from sim pie swellable and erosi on systems may be described by the well-known power law expression and is defined by the following equation(Ritger and Pepp as, 1987; P illay and Fassihi, 1999资料个人收集整理，勿做商业用途Where M t is the amount of drug released at time t, M is the overall amount of drug released, K is the release con sta nt; n is the release or diffusi onal exponent; M/M is the cumulative drug concen trati on released at time t (or fractio nal drug release)料个人收集整理，勿做商业用途The release exponent (n) is used for the in terpretati on of the release mecha nism from poly meric matrix con trolled drug release systems (Peppas 1985). For the case of < 0.45 corrosFickdantdiffusi on release (Case I<an89homalous (non-Fickia n) transport, n = 0.89 toa zero-order (Case II) release kin etics, and n > 0.89to a super Case II transport (Ritger and Pepp as, 1987资料个人收集整理，勿做商业用途 The dissoluti on data were modelled by using the Po wer law equati on (Eq 7) withgraphs analysis software (Origin Scientific Graphing and Analysis software, Version 7, Origi nLab Corpo rati on, USA) using the Gaussia n-Newt on (Leve nberg-Hartely) app roach 资料个人收集整理，勿做商业用途2.9.2 Mea n dissolutio n time (MDT)MDT is a statistical moment that describes the cumulative dissolution process and provides a quantitative estimation of the drug release rate. It is defined by the following equation (Reppas and Nicolaides, 2000; Sousa^t al., 2002):资料个人收集整理，勿 做商业用途nMDTt i M t/M^ i =±Where MDT is the mean dissolution time, M t is the amount of the drug released at time t; t i is the time (min) at the midpoint between i and i-1 and M 乂 is the overall amount of the drug released.料个人收集整理，勿做商业用途cyli ndrical, i n sp ecially, n diffusi on al), 0.45 < n2.9.3 Differe nt factor f i and Similarityfactor f 2 The different factor f i is a measure of the relative error between two dissoluti on curves and the similarity factor f 2 is a measure of similarity in the p erce ntage dissoluti on betwee n two dissoluti on curves (Moore and Fla nn er, 1996). Assu ming that the p erce ntage dissolved values for two p rofiles cannot be higher tha n 100, the differe nt factor f 1 can have values from 0 (whe n no differe nee the two curves exists) to 100 (when maximum differenee exists). With the same assumption holding, the similarity factor f 2 can have values from 100 (when no differenee between the two curves exists) to 0 (when maximum differenee exists) (Pillay and Fassihi, 1999;Moore and Fla nn er, 1996; Re ppas and Nicolaides, 2000). In this study, these factors were used to confirm the relative of release p rofiles of water poor soluble model drugs from poly mers based matrix tablets of the same formulati ons. They are defi ned bythe followi ng equati ons:资料个人收集整理，勿做商业用途 f^100xn Z |Rt —Tt t 吕 n z R f2"0^「100hXG (Rt -T , I V ny 丿］Where n is the number of sample withdrawal points, R t is the percentage of the refere nee dissolved at time t, T is the p erce ntage of the test dissolved at time 资料个人 收集整理，勿做商业用途 3 Results and discussion 3.1 Prep arati on and characterisati on ofPCC The ion ic bond of the interpo "electrolyte comp lex (IP EC) betwee n chitosa n andpo lycarb op hil was con firmed by means p reviously p ublished differe ntial sea nning calorimetry (DSC) (Lu et al., 2007b) and Fourier tran sform in frared (FT-IR). Fig.1shows the FT-IR sp ectra of chitosa n, po lycarb op hil and the PCC poly mer.资料个人收集整理，勿做商业用途-1A peak that appears at 1561 cm on the IR spectrum of the PCC, which might be assigned to the carboxylate groups that formed ionic bonds with the protonated amino groups of chitosan as previously illustrated for the interaction between Eudragit E andEudragit L (Moustafine at al., 2005). This ionic bond seems to be the primary bin di ng force for the formatio n of a comp lex betwee n chitosa n and po lycarb op hil.资料个人收集整理，勿做商业用途Chitosan is a cationic polymer of natural origin with excellent gel and film forming properties. Polycarbophil can also be considered as polyanions with negatively charged carboxylate groups. Mixing chitosan and polycarbophil in acidic solution (2% acetic acid solution was used in the study), ionic bonds should form between the protonated free amino groups of chitosan and carboxylate groups of polycarbophil.According to the results obtained from DSC and FT-IR, the possible process of formatio n of interpo "electrolyte comp lexes may be described as illustrated in Fig.2资料个人收集整理，勿做商业用途3.2Physical characteristics and drug content of poly mers based matrix tablets^ 料个人收集整理，勿做商业用途As summarised in Table 2, the physical characteristics of matrix tablets showed the good thickness uniformity, as ranged from 3.40 0.04 to 4.12± 0.0±4mm, a variationof matrix tablets weight from 73.3 2.4 mg to±87.9 4.0±mg, furthermore the weight variation of all formulation tablets is very low (< 10% from the average weight) (USP 2006). Hardness of the matrix tablets shows a range from 68 ±14 to 94 ±12N.The tablets also pasted the friability test (<1%), confirm that all formulations tablets are within USP (2006) limits. Drugs content of all formulations ranged from 4.60 0.65 to 5.01 0.1±1%.资料个人收集整理，勿做商业用途3.3Swelli ng and erosi on prop erties of the poly mers based matrices tablets资料个人收集整理，勿做商业用途Investigation of matrix hydration and erosion by gravimetrical analysis is a valuable exercise to better understand the mechanism of release and the relative importance of participating parameters (Jamzad and Fassihi, 2006). Fig.3 and Fig.4 illustrate the water uptake profiles and Fig.5， Fig.6 illustrate percentage of matrix erosion of all formulation tablets, respectively. Swelling properties of the all formulation matrix tablets based on the content of PCC, HPMC K100M and HPMC K100LV in the matrices tablets. Water uptake and percentage of matrix erosion values of these matrix tablets show superior swelling characteristics either HPMC K100LV based matrix tablets, or PCC based matrix tablets. 资料个人收集整理，勿做商业用途IPEC betwee n chitosa n and po lycarb op hil is a three -dime nsional n etworks water insoluble poly (acrylic acid) polymer with free hydroxy groups. Hydroxy groups ofPCC contribute hydrophilic capacity significantly and polymer erosion characteristics depend on the reaction ratio of chitosan and polycarbophil while polymer synthesis.While the PCC based matrixes were put into the buffer solution, the electrostatic repulsion between fixed charges (hydroxy groups) uncoiled the polymers chains.The counterion diffusion inside the PCC gel creates an additional osmotic pressure difference across the gel, consequently lead to higher water uptake (Peppas and Khare, 1993; Lu, et al., 2007b). During the matrix erosion, the ionic bonds between chitosan and polycarbophil were not broken by the matrix swelling. PCC based matrix tablets (F1 and F7 formulation) have superior swelling behaviors compare to the HPMC based matrices. Swelling index values of F1 and F7 formulation matrix tablets are 1599.62±216.68 % and 1579.82 ±118.05 % at 12 hours, respectively.Furthermore, addition of microcrystalline cellulose (Avicel) can increase matrices erosion significantly. Compare the erosion behaviors of F1, F7 and F2, F8formulation (containing 20% Avicel), F1 and F7 matrix tablets erode 5.74 1.62 % and 6.59 1±.18 % on 12 hours only, cont rary F2 and F8 matrix tablets erode 55.59 1.43 and 100 % respectively. Microcrystalline cellulose (Avicel) is widely used in pharmaceutical, primarily as a binder/diluent, also has some disintegrant properties on oral tablet and capsule formulations where it is used in both wet granulation and direct-compression process (Wheatley, 2000). In this study, matrix erosion behaviours were act by microcrystalline cellulose facilitating the transport of liquid into the pore of matrix tablets. It demonstrates that PCC polymer have capacity to form swelli ng only or swelli ng-erosi on matrix drug delivery system.资料个人收集整理，勿做商业用途It also was confirmed that PCC based matrix tablets have much better swelling behaviors than HPMC based matrix tablets by comparing swelling curves in Fig.3 andFig.4. Swelling index values of F1 and F7 formulation matrix tablets are 1599.62216.68 % and 1579.82 11±8.05 % at 12 hours, contrary F3 and F9 matrix tablets are545.96 ±4.32% and 547.72 2±6.27%. HPMC K100LV based matrix tablets have excellence erosion curves in this study, F5, F6, F11 and F12 formulation matrixtablets eroded 100% on 12 hours, but F2 and F8 (PCC based tablets) formulation matrix tablets can eroded 55.59 ±1.43 and 100 % with microcrystalline cellulosefacilitating. 资料个人收集整理，勿做商业用途3.4Drug releaseIn vitro drug release was performed in pH 7.4 phosphate buffer solution for 12 hours.Results of percentage drug release versus time for hydrochlorothiazide and ketoprofen in different formulations matrices tablets are presented in Fig.7 and Fig. 8, while theMDT and drug release kinetics values were present in Table 3资.料个人收集整理，勿做商业用In this study, water poor soluble model drugs hydrochlorothiazide and ketoprofen release from polymers based matrix tablets was controlled by the polymer matrices swelling or swelling combination with erosion. Percentage of drug release, matrix swelling and erosion of F7 were summarised in Fig 9. The percentageketoprofen release curve follows the percentagematrix tablets swelling curve, it demonstrates that PCC based matrix drug delivery system is the swelling dependent drug release system for water poor soluble model drugs. Same as F7 matrix tablets, F1 matrix tablets is also a swelling only drug delivery system, in these matrix systems drugs release behaviour primarily depend on the matrix swelling characteristics. Because as the superior swelling capacity of PCC based matrix tablets, liquid environments inside of the matrix provide that the model drugs release are zero order drug release.As described in Table 3, release exponentsn)( of F1 and F7 are 0.83 0.03 a±nd 0.99 ± 0.02 during the experimental time, respectively. 资料个人收集整理，勿做商业用途Addition of microcrystalline cellulose (Avicel) influence the model drugs release profiles from PCC based matrix tablets significantly. Cumulative drug release of F2 and F8 formulation tablets is 93.7 4.13 % a±nd 99.6 4.2±5% at 12 hours, relativelyF1 and F7 formulation tablets is 73.8 1.13 % an±d 47.2 4.5±3 % only. This can be explained by drugs release mechanism were swelling and erosion instead of swelling only, consequently accelerate the drugs release. The adjustable capacity of PCC based matrix drug delivery system by addition of microcrystalline cellulose (Avicel) dem on strates the poten tial useful of PCC poly mer in drug con trolled release field 资料个人收集整理，勿做商业用途Compare to the PCC based matrix tablets, the drugs release profiles of HPMC based matrix tablets were adjusted difficultly. The relatives f1 and f2 values of difference polymers including PCC, HPMC 100M, HPMC 100LV based matrix tablets containing hydrochlorothiazide under same formulation were show in Table 4. As describedf1 and f2 values in Table 4, F3 and F4, F5 and F6 formulation tablets have similar drug release behaviours, but F1 and F2 formulation tablets illustrate different drug release behaviours. This phenomenacan be explained by the superior water uptake capacity of PCC polymer, more water containing can easier broken the physical tensility between the polymer particles. 资料个人收集整理，勿做商业用途However, HPMC 100LV polymer has excellence erosion characteristics, in this study model drugs release from HPMC 100LV based matrix tablets illustrate matrix erosion dependent properties. In generally, drug release from swelling and erosion matrix system shows zero order release pare the drug release exponentsn(), release constant (k1), and mean dissolution time (MDT) of F2 to F5, F6, they have not significantly different as described in Tablet 3, furthermore the relatives f1 and f2 values between F2 and F5, F6 in Table 4 show they are similar release profiles. It imply PCC based matrix tablets can become a swelling and erosion drug delivery system by the addition of microcrystalline cellulose (Avicel), this drug delivery system illustrate similar drug release p rofiles as HPMC 100LV based matrix tablets资料个人收集整理，勿做商业用途Although it is very complex process that the model drugs release from swelling and。

人绒毛膜促性腺激素检测溯源问题刘奉亭青岛兰信医学检验所，青岛266100摘要目的：探讨HCG检测的溯源性及实验间检验结果互相认可中存在的问题。

方法：结合国际国内HCG检测现状，对有关HCG检测的影响因素及国际标准物质方面的问题进行综述。

结果：HCG分子及其裂解产物种类繁多，抗原性相似又不完全相同，不同厂家试剂使用的抗体组合不同，对HCG类分子的识别不同。

一些试剂的标签不正确或不清楚，一些实验人员对HCG检验的分子种类不清楚，国内HCG收费项目不全，是导致有意无意错报检验项目的原因。

目前发布的HCG 国际标准物质含有杂质，并且对于无活性的α亚基和β亚基赋值错误。

这些都给HCG检测结果的实验室间互相认可带来困难。

结论：HCG检测的溯源及检验结果的实验室间互相认可尚存在许多问题。

要解决这些问题，需要新的、以摩尔浓度赋值的HCG国际标准的发布，各试剂厂商以新的国际标准物质作为抗原来制备抗体，并用新的抗体制备的HCG检测试剂溯源到新的国际标准。

另外，还要进一步普及有关HCG类分子及其裂解产物的知识宣传。

血清人绒毛膜促性腺激素（HCG）的测定对于妊娠的确认和监测，异位妊娠及其治疗的监测，产前筛查，先兆流产、滋养层疾病、非滋养层肿瘤、生殖细胞瘤、膀胱癌、睾丸癌、肺癌等的诊断和治疗效果的监测具有十分重要的意义[1-7]。

人绒毛膜促性腺激素（HCG）与促黄体生成激素（LH）、促卵泡激素（FSH）及促甲状腺激素（TSH）都是由一个α亚基和一个β亚基所组成[8,9] 。

这些激素都拥有共同的α亚基，它们的区别在β亚基[10]。

HCG的β亚基与FSH、TSH不同，但与LH的β亚基类似，LH的β亚基含有121个氨基酸，而HCG的β亚基含有145个氨基酸，在LH的β亚基的C末端延伸了24个氨基酸[11]。

早期，人们采用生物学方法进行妊娠实验，将尿液注入幼鼠等动物，通过观察动物的反应而判断是否怀孕。

1960年Wide和Gemzell建立了首个妊娠实验的免疫学方法――红细胞凝集法[12]。

DOI:10.12280/gjszjk.20200730谢奇君，李欣，赵纯，凌秀凤△【摘要】辅助生殖技术（ART）已被广泛应用于不孕症的治疗，包括人工授精（AI）和体外受精-胚胎移植（IVF-ET），及其相关衍生技术如胞浆内单精子注射（ICSI）、胚胎植入前遗传学筛查（PGS）、卵母细胞体外成熟（IVM）、胚胎辅助孵化（AH）等技术。

随着ART的普及和妊娠率的逐步提高，其带来的并发症及其安全性也越来越受到重视。

能够及时发现和处理并发症，可以最大限度地使不孕症患者获得安全优质的妊娠结局。

目前，ART相关并发症发生的原因仍存在争议，有部分研究认为是由于ART导致的多胎妊娠或不孕症病因，也有研究认为是由于ART本身。

故对近年来相关文献进行分析，总结ART相关并发症的最新研究进展。

【关键词】生殖技术，辅助；并发症；不育，女（雌）性；体外受精；胚胎移植Research Progress of Complications Related to Assisted Reproductive Technology XIE Qi-jun,LI Xin,ZHAO Chun,LING Xiu-feng.Reproductive Center,Women′s Hospital of Nanjing Medical University(NanjingMaternity and Child Health Care Hospital),Nanjing210004,ChinaCorresponding author:LING Xiu-feng,E-mail:************************【Abstract】Assisted reproductive technology(ART)has been widely used in the treatment of infertility,including artificial insemination(AI)and in vitro fertilization-embryo transfer(IVF-ET),and many derivativetechniques such as intracytoplasmic sperm injection(ICSI),preimplantation genetic screening(PGS),in vitromaturation(IVM),and assisted hatching(AH).With the popularity of ART and the gradual improvement ofpregnancy rate,more and more attention has been paid to the complications of ART and the safety of ART.Detection and treatment of complications in time can maximize the safety and high quality of pregnancy,and thegood outcomes for patients.At present,the causes of ART-related complications are still controversial.Somestudies believe that the increased rate of multiple pregnancy caused by ART or the factors related to infertility arethe main causes.However,other studies believe that the factors of ART itself cannot be overlooked.In this review,we analyzed the complications related to ART and the causes.【Keywords】Reproductive techniques，assisted；Complications；Infertility，female；Fertilization in vitro；Embryo transfer（ＪＩｎｔＲｅｐｒｏｄＨｅａｌｔｈ蛐ＦａｍＰｌａｎ，2021，40：204-208）·综述·基金项目：国家自然科学基金（81871210）作者单位：210004南京医科大学附属妇产医院（南京市妇幼保健院）生殖中心通信作者：凌秀凤，E-mail：************************△审校者辅助生殖技术（assisted reproductive technology，ART）是一种应用各种技术处理精子或卵子，以帮助不孕症夫妇实现生育的方法，包括人工授精（artificial insemination，AI）、体外受精-胚胎移植（invitro fertilization-embryo transfer，IVF-ET）及相关技术，如胞浆内单精子注射（intracytoplasmic sperminjection，ICSI）、胚胎植入前遗传学筛查（preimplantation genetic screening，PGS）、卵母细胞体外成熟（in vitro maturation，IVM）、胚胎辅助孵化技术（assisted hatching，AH）和卵母细胞玻璃化冷冻技术等。

Characterization of acid-soluble collagen from the skinof walleye pollock (Theragra chalcogramma )Mingyan Yan,Bafang Li *,Xue Zhao,Guoyan Ren,Yongliang Zhuang,Hu Hou,Xiukun Zhang,Li Chen,Yan FanCollege of Food Science and Technology,Ocean University of China,No.5,Yushan Road,Qingdao,Shandong Province 266003,PR ChinaReceived 10July 2007;received in revised form 14September 2007;accepted 4October 2007AbstractAcid-soluble collagen (ASC)was extracted from the skin of walleye pollock (Theragra chalcogramma )and partially characterized.It exhibited a maximum absorbance at 220nm,but little absorbance near to 280nm.Amino acid composition and SDS-PAGE suggested that the collagen might be classified as type I collagen.Moreover,FTIR investigations showed the existence of helical arrangements of collagen.The denaturation temperature (T d )and shrinkage temperature (T s )were 24.6°C and 47°C,respectively,both lower than those of mammalian collagens.However,T d of walleye pollock skin collagen was higher than that of cod skin collagen reported previously.These results indicate that walleye pollock skin is a potential source of collagen and provide the theoretical basis for further research.Ó2007Elsevier Ltd.All rights reserved.Keywords:Characterization;Collagen;Thermal stability;Walleye pollock1.IntroductionCollagen is an abundant protein in animal tissues and constitutes approximately 30%of total animal protein (Muyonga,Cole,&Duodu,2004).It is widely distributed in skin,bones,cartilage,tendons,ligaments,blood vessels,teeth,cornea and all other organs of vertebrates (Sena-ratne,Park,&Kim,2006).Collagen has a wide range of applications in leather and film industries,pharmaceutical,cosmetic and biomedical materials,and food (Kittiphattan-abawon,Benjakul,Visessanguan,Nagai,&Tanaka,2005).The highest utilization of collagen is in pharmaceutical applications including production of wound dressings,vit-reous implants and as carriers for drug delivery.Moreover,collagen is used for the production of cosmetics because it has a good moisturizing property (Swatschek,Schatton,Kellermann,Muller,&Kreuter,2002).In addition,colla-gen has been utilized to produce edible casings,which areneeded in the meat processing industries (sausages/sal-ami/snack sticks),and heat-denatured collagen,called gel-atin,is important in food manufacturing (Senaratne et al.,2006).For industrial purposes,the main sources of collagen are limited to those of land-based animals,such as bovine or porcine skin and bone.However,the outbreak of bovine spongiform encephalopathy (BSE),transmissible spongi-form encephalopathy (TSE)and the foot-and-mouth dis-ease (FMD)crisis have resulted in anxiety among users of collagen and collagen-derived products of land animal origin (Jongjareonrak,Benjakul,Visessanguan,Nagai,&Tanaka,2005).In addition,the collagen extracted from porcine sources cannot be used as a component of some foods,due to religious barriers.Therefore,alternative sources of collagen should be sought.Scientists have found that skin,bone,scale,fin and cartilage of freshwater and marine fish,scallop mantle (Shen,Kurihara,&Takahashi,2007),adductor of pearl oyster (Mizuta,Miyagi,Nish-imiya,&Yoshinaka,2002)and the muscle layer of the ascidian (Mizuta,Isobe,&Yoshinaka,2002)can be used as new sources of collagen.0308-8146/$-see front matter Ó2007Elsevier Ltd.All rights reserved.doi:10.1016/j.foodchem.2007.10.027*Corresponding author.Tel.:+8653282031662.E-mail address:mingyan012003@ (B.Li)./locate/foodchemAvailable online at Food Chemistry 107(2008)1581–1586Food ChemistryPollock is one of the commercially importantfish species in China.Approximately400–500thousand of tons pollock were processed annually,mainly in Shandong Province. Seafood processors generate more than30thousand of tonsfish skins through their processing of pollock for food service per year(data were provided by Bureau of Fisher-ies,Ministry of Agriculture,People’s Republic of China). The skin is dumped without utilization,which would cause environmental pollution and resources waste.However, 70%of the pollock skin dry matter is collagen(Zhang, Luo,Zhang,Song,&Jiang,2003).For making effective use of the dumped skin as a collagen resource,it is neces-sary to obtain fundamental information about the pollock skin collagen.Therefore,the present paper describes the isolation and physicochemical properties of the collagen from walleye pollock skin.2.Materials and methods2.1.Fish skin preparationWalleye pollock(Theragra chalcogramma)were caught from the Bering Sea by commercialfishing boat in August, 2006,stored atÀ18°C,immediately after gutting,and transported to the dock in Qingdao.After arrival at a local fish processing factory,frozenfish were thawed using run-ning water,and skins were removed and descaled manu-ally.These skins were transported to the laboratory and stored atÀ20°C until used.All other reagents used were of analytical grade.2.2.Extraction of collagenAll procedures were performed at4°C,as previously described(Nagai&Suzuki,2000)with a slight modification. The skin was extracted with0.1M NaOH to remove non-collagenous materials effectively and to exclude the effect of endogenous proteases on collagen(Sato,Yoshinaka, Sato,&Shimizu,1987),then thoroughly rinsed with distilled water until a neutral pH was reached.Minced skins were slowly stirred with0.5M acetic acid solution for48h,and the extract was centrifuged at10,000g for30min.Then the acid-soluble collagen(ASC)in the supernatant was salted out by adding NaCl to afinal concentration of0.9M.The solution was left overnight,and the resultant precipitate, collected by centrifugation at8000g for20min,was dis-solved in0.5M acetic acid,dialyzed against0.1M acetic acid for1d(1:15,v/v,changed every4h),distilled water for2d(1:15,v/v,changed every4h),and then lyophilized.2.3.UV–vis spectraA collagen sample was dissolved in0.5M acetic acid to obtain a concentration of2g lÀ1.The UV–vis adsorption spectra were recorded by a Shimadzu spectrophotometer UV-2550(Shimadzu,Tokyo,Japan)from190to400nm at a scanning rate of210nm min-1.2.4.Amino acid compositionASC samples were hydrolyzed under reduced pressure with6M HCl at110°C for24h,and the hydrolysates were analyzed on a Hitachi835-50amino acid analyzer(Hitachi, Tokyo,Japan).2.5.Sodium dodecyl sulphate polyacylamide-gel electrophoresis(SDS-PAGE)SDS-PAGE was performed by the method of Laemmli (1970),using the discontinuous tris-HCl/glycine buffer sys-tem with7.5%resolving gel and5%stacking gel.Proteins were stained with0.1%(w/v)Coomassie Brilliant Blue R-250dissolved in water,methanol and acetic acid(9:9:2, v/v/v)for20min,then destained using a solution contain-ing water,methanol and acetic acid(8:1:1,v/v/v).2.6.Fourier transform infrared spectroscopy(FTIR)FTIR spectra were obtained from discs containing 0.2mg lyophilized collagen and about10mg potassium bromide(KBr)ground together under dry conditions. The spectra were recorded using infrared spectrophotome-ter(Nicolet200SXV)from4000to500cmÀ1at a data acquisition rate of2cmÀ1per point.The resulting spectra were analyzed using Omnic6.0software(Thermo-Nicolet, Madison,Wisconsin).2.7.Determination of denaturation temperatureThe denaturation temperature was measured from the viscosity changes,using an Ubbelohde viscometer,accord-ing to the method of Zhang et al.(2007)with some modi-fication.Ten millilitres of0.03%collagen solution in0.1M acetic acid+0.2M sodium acetate buffer(pH5.0)were used for viscosity measurements.The thermal determina-tion curve was obtained by measuring solution viscosity at several temperatures from16to42°C,and the temper-ature was raised stepwise and maintained for30min.Frac-tional viscosity at the given temperature was calculated by the equation:Fractional viscosity¼ðg spðTÞÀg spð42 CÞÞ=ðg spð16 CÞÀg spð42 CÞÞ;where g sp is the specific viscosity.These fractional viscosi-ties were plotted against the temperatures and the denatur-ation temperature was taken to be the temperature where fractional viscosity was0.5.2.8.Differential scanning calorimetryDifferential scanning calorimetry(DSC)was performed on a Netzsch DSC200PC(Netzsch,Bavaria,Germany) instrumentfitted with an air cooling compressor and a liquid nitrogen cooler at ambient temperature(Cui et al.,1582M.Yan et al./Food Chemistry107(2008)1581–15862007).The temperature was effectively calibrated using indium as standard.Collagen fibre was weighed (3mg)accurately and sealed in aluminium pans (BO 6.239.2–64.502).At least triplicate samples were heated from 20to 100°C at a scanning rate of 2K min -1,with an empty sealed pan as a reference.The shrinkage temperature was measured at the top of the transition peak.3.Results and discussion 3.1.UV–vis spectraAs can be seen from the UV–vis spectra (Fig.1),the dis-tinct absorbance of the collagen was obtained near 220nm,which is contributed by n ?p *transition of C @O in the peptide bond.Generally,tyrosine and phenylalanine are sensitive chromophores and absorb UV light at 283nm and 251nm (Liu &Liu,2006),where ASC has no evident absorbance.Therefore,acid-soluble collagen from walleye pollock skin well supports the property of collagen that there is absorbance at 220–230nm,with little or no absor-bance near 280nm.Thus the protein is collagen.3.2.Amino acid compositionThe amino acid composition of the acid-soluble collagen from walleye pollock skin is shown in Table 1.Glycine was the major amino acid although it was only 19.7%,signifi-cantly lower than that of brown backed toadfish (Sena-ratne et al.,2006),but similar to that of Nile perch (Muyonga et al.,2004)and channel catfish (Liu,Li,&Guo,2007).The reason that the glycine content was lower may be related to contamination by other proteins.The collagen was found to be very low in tyrosine,histidine and isoleucine,and no cystine was detected.It also con-sisted of imino acids (proline +hydroxyproline),which were unique amino acids found in collagen.The imino acid content was 18.4%,similar to the 18.6%of grass carp col-lagen (Zhang et al.,2007),but lower than the 22%of por-cine skin collagen (Ikoma,Kobayashi,Tanaka,Walsh,&Mann,2003).Hydroxyproline is derived from proline by post-translational hydroxylation mediated by prolylhy-droxylase (Li,Liu,Gao,&Chen,2004).The degree of hydroxylation of proline residues in collagen from walleye pollock skin was 37.5%,similar to the 39%of ocellate puf-fer fish (Nagai,Araki,&Suzuki,2002),but lower than the 43%of channel catfish (Liu et al.,2007)and 48%of cuttle-fish (Nagai,Yamashita,Taniguchi,Kanamori,&Suzuki,2001).The distribution patterns of amino acid composi-tion,similar to that of acid-soluble collagen from the skin of channel catfish (Liu et al.,2007),indicate that ASC might be classified as type I collagen.3.3.SDS-PAGEThe acid-soluble collagen from walleye pollock skin was examined by SDS-PAGE,using a 7.5%resolving gel (Fig.2).This collagen consisted of a chains (a 1and a 2chain),which showed two distinct species varying in their mobility,and their dimer (b chain),and small amounts of c components were also found.The existence of at least two different subunits shows that a major collagen from walleye pollock skin might be type I collagen.There was no clear difference between the electrophoretic patterns,with or without b -mercaptoethanol,suggesting absence of disulphide bonds.This is consistent with the observation that the collagen was almost devoid of sulphur-containing amino acids (Muyonga et al.,2004).From the electropho-retic patterns of collagen,we could not determine whether the collagen contained an a 3chain or not.If present,the a 3chain could not be separated under the conditionemployedFig.1.UV–vis spectra of acid-soluble collagen from walleye pollock skin.Table 1Amino acid composition of acid-soluble collagen from walleye pollock skin Amino acid Amino acid residues/1000total amino acid residues Hydroxyproline 69Aspartic acid 54Threonine 23Serine66Glutamic acid 116Glycine 197Alanine 104Cystine 0Valine21Methionine 24Isoleucine 13Leucine 27Tyrosine4Phenylalanine 21Lysine 35Histidine 13Arginine 98Proline115M.Yan et al./Food Chemistry 107(2008)1581–15861583because a 3(I)migrates electrophoretically to the same posi-tion as a 1(I)(Kimura,1992).3.4.Fourier transform infrared spectroscopyFig.3shows the FTIR spectra of the acid-soluble colla-gen from walleye pollock skin,similar to that exhibited by other collagens (Muyonga et al.,2004;Liu et al.,2007).The amide A band is associated with the N–H stretching frequency.A free N–H stretching vibration occurs in the range of 3400–3440cm À1,and when the NH group of apeptide is involved in a hydrogen bond,the position is shifted to lower frequency,usually near 3300cm À1(Li et al.,2004).The amide A band of walleye pollock skin col-lagen was found at 3328cm À1,which shows that there were NH groups involved in hydrogen bonds.The amide B band of collagen was found at 3080cm À1,which is related to asymmetrical stretch of CH 2(Muyonga et al.,2004).The amide I band position was observed at 1648cm À1,which is the absorption band of C @O stretching.It is asso-ciated with the secondary structure of the protein.The absorption between the 1236cm À1(amide III)and 1452cm À1bands demonstrated the existence of helical structure (Liu et al.,2007).Therefore,the FTIR investiga-tions show the existence of helical arrangements of walleye pollock skin collagen.3.5.Thermal stabilityThermal stability of collagen is usually described by the denaturation temperature (T d )in solution and the shrink-age temperature (T s )of fibre.The temperature at which the triple helix structure of collagen in solution is disinte-grated into random coils is taken as T d (Hao &Li,1999).Fig.4shows the thermal denaturation curve of acid-soluble collagen from walleye pollock skin.T d of collagen was 24.6°C,lower by about 12°C than that of collagen from porcine skin (Nagai et al.,1999).The shrinkage tempera-ture refers to the temperature at which fibre shrinks to one third of its length (Fathima,Madhan,Rao,Nair,&Ramasami,2004).In the shrinkage process,a phase transi-tion involving the conversion of a crystalline triple helical collagen structure to an amorphous random coil form (Fat-hima,Balaraman,Rao,&Nair,2003)occurs.T s of colla-gen from walleye pollock skin was 47°C (Fig.5),lower by about 15°C than that of type I collagen from bovine skin (Cui et al.,2007).These results proved that thehelicesFig.2.SDS-PAGE patterns of acid-soluble collagen from walleye pollock skin on 7.5%ne 1:ASC (the sample solution without b -mercaptoethanol);lane 2:ASC (the sample solution with b -mercap-toethanol);lane 3:proteinmarkers.Fig.3.Fourier transform infrared spectra of acid-soluble collagen from walleye pollock skin.1584M.Yan et al./Food Chemistry 107(2008)1581–1586of collagen from walleye pollock skin were less stable than those of mammalian collagens.We also found that the DSC thermogram was broader,which reveals smaller cooperatives among the participating subunits(Usha& Ramasami,2004).There were still two small peaks within the temperature range20–35°C.The former could be observed in the DSC thermogram of collagen from calf and sea cucumber(Stichopus japonicus)(Cui et al.,2007). The latter is a pre-denaturational transition,which has been recorded in previous studies,but its nature has not been ultimately identified(Komsa-Penkova,Koynova, Kostov,&Tenchov,1996).Some studies ascribe it to shortened or nicked collagen fragments(Condell,Sakai, Mercado,&Larenas,1988)or protein oxidation(Komsa-Penkova et al.,1996).The difference between T s and T d of walleye pollock skin collagen was about22°C,which is obviously consistent with the conclusion that the differ-ence between T s and T d of marine collagen is not influenced by species,about(20–25)°C(Hao&Li,1999).The thermal stability is influenced by the imino acid con-tent.The higher the imino acid content,the more stable are the helices,because the molecular structure of collagen is maintained mainly by restrictions on changes in the sec-ondary structure of the polypeptide chain,imposed by the pyrrolidine rings of proline and hydroxyproline,and also partially maintained by the hydrogen bond ability through the hydroxy group of hydroxyproline(Zhang et al.,2007).Therefore,porcine and bovine skin collagens, having higher imino acid contents(220and215residues per1000residues,respectively)(Ikoma et al.,2003;Cui et al.,2007)than walleye pollock skin collagen(184resi-dues per1000residues),denature at higher temperatures. It is known that the stability of collagen is also correlated with environmental and body temperature.Bigeye snapper and Brownstripe red snapper are tropicalfish,so the colla-gens are more stable(Kittiphattanabawon et al.,2005; Jongjareonrak et al.,2005).T d of walleye pollock skin col-lagen in this study was higher than that of cod skin colla-gen,determined by Rigby(1968),probably due to the environmental and body temperature.4.ConclusionASC extracted from walleye pollock skin was classified as type I collagen.It has a distinct amino acid composition and thermal stability,different from those of bovine or por-cine skin collagens.FTIR investigations show the existence of helical arrangements of collagen.Thus,walleye pollock skin is a good source of collagen.This study provides a the-oretical basis for collagen modification and utilization.AcknowledgementThis work was supported by the High Technology Re-search and Development Programme of China(No. 2006AA09Z438).Fig.5.Thermal transition curve of acid-soluble collagen from walleye pollock skin,as shown by DSC.M.Yan et al./Food Chemistry107(2008)1581–15861585ReferencesCondell,R. 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-1108-宁夏医学杂志2020年12月第42卷第12期Ningxia Med J,Dec.2020,Vol.42,No.12 Doi：10.13621/j.1001-5949.2020.12.1108•实验研究•成骨细胞源性外泌体的分离提取和鉴定蔡则成】，杨绍兵2,马荣3,张彦龙】，梁思敏3［摘要］目的分离成骨细胞来源的外泌体，并对其进行鉴定。

方法体外培养成骨细胞，用差速离心法分离、提取成骨细胞分泌的外泌体，经透射电子显微镜观察其形态及大小，采用蛋白质印迹法（WB）检测外泌体跨膜蛋白CD9和成骨细胞特异性碱性磷酸酶（AKP）的表达。

结果成骨细胞源性外泌体为直径30-100nm的圆形或椭圆形结构,高表达CD9和AKP蛋白。

结论差速离心法提取的成骨细胞源性外泌体，具有外泌体的一般特性，并高表达成骨细胞特异性蛋白AKP。

［关键词］结核分枝杆菌；外泌体;成骨细胞［中图分类号］Q2-33［文献标识码］AIsolation，extraction and identification of exosomes derived from osteoblasts CAI Zecheng，Y ANG Shaobing2，MA Rong3，ZHANG Yanlong1，LIANG Simin3. 1.Ningxia Medical University，Yinchun750004，China Department of Cardiology，General Hospital of Ningxia Medical University，Y inchun750004，C hina；3.Department of Orthopae­dics，General Hospital of Ningxia Medical University，Y inchun750004，C hinaCorresponding author:L IANG Simin,Email：148376463@[Abstract]Objective The exosomes from osteoblasts were isolated and identified.Methods Osteoblasts were cultured in vitro.The exosomes secreted from osteoblasts were separated and extracted by differential centrifugation.The morphology and size of exo­somes were observed by transmission electron microscopy.The expression of CD9and alkaline phosphatase in osteoblasts were detected by Western blotting.Results The exosomes of osteoblasts were round or oval structures with a diameter of30〜100nm,which highly ex­pressed CD9and AKP protein.Conclusion The osteoblast derived from exosomes have the general characteristics of exosomes and high­ly express the osteoblast specific protein AKP.[Key words]Mycobacterium tuberculosis；E xosomes；Osteoblasts脊柱结核是由结核分枝杆菌引起的慢性感染性疾病,以进行性骨质破坏为主要病理特征［1］，破骨细胞在脊柱结核骨质破坏的发生发展中起着极其重要的作用。

益生菌对阿尔茨海默病作用的研究进展发布时间：2021-12-14T06:08:15.523Z 来源：《中国结合医学杂志》2021年12期作者：宋鑫萍1,2，李盛钰2，金清1[导读] 阿尔茨海默病已成为威胁全球老年人生命健康的主要疾病之一，患者数量逐年攀升，其护理的经济成本高，给全球经济造成重大挑战。

近年来研究显示，益生菌在适量使用时作为有益于宿主健康的微生物，在防治阿尔茨海默病方面具有积极影响，其作用机制可能通过调节肠道菌群，影响神经免疫系统，调控神经活性物质以及代谢产物，通过肠-脑轴影响该病发生和发展。

宋鑫萍1,2，李盛钰2，金清11.延边大学农学院，吉林延吉 1330022.吉林省农业科学院农产品加工研究所，吉林长春 130033摘要：阿尔茨海默病已成为威胁全球老年人生命健康的主要疾病之一，患者数量逐年攀升，其护理的经济成本高，给全球经济造成重大挑战。

近年来研究显示，益生菌在适量使用时作为有益于宿主健康的微生物，在防治阿尔茨海默病方面具有积极影响，其作用机制可能通过调节肠道菌群，影响神经免疫系统，调控神经活性物质以及代谢产物，通过肠-脑轴影响该病发生和发展。

本文综述了近几年来国内外益生菌对阿尔茨海默病的作用进展，以及其预防和治疗阿尔茨海默病的潜在作用机制。

关键词：益生菌；阿尔茨海默病；肠道菌群；机制Recent Progress in Research on Probiotics Effect on Alzheimer’s DiseaseSONG Xinping1,2，LI Shengyu2，JI Qing1*(1.College of Agricultural, Yanbian University, Yanji 133002，China)(2.Institute of Agro-food Technology, Jilin Academy of Agricultural Sciences, Chanchun 130033, China)Abstract：Alzheimer’s disease has become one of the major diseases threatening the life and health of the global elderly. The number of patients is increasing year by year, and the economic cost of nursing is high, which poses a major challenge to the global economy. In recent years, studies have shown that probiotics, as microorganisms beneficial to the health of the host, have a positive impact on the prevention and treatment of Alzheimer’s disease. Its mechanism may be through regulating intestinal flora, affecting the nervous immune system, regulating the neuroactive substances and metabolites, and affecting the occurrence and development of the disease through thegut- brain axis. This paper reviews the progress of probiotics on Alzheimer’s disease at home and abroad in recent years, as well as its potential mechanism of prevention and treatment.Key words：probiotics; Alzheimer’s disease; gut microbiota; mechanism阿尔茨海默病（Alzheimer’s disease, AD），系中枢神经系统退行性疾病，属于老年期痴呆常见类型，临床特征主要包括：记忆力减退、认知功能障碍、行为改变、焦虑和抑郁等。

英文单词：组织学与胚胎学（白皮）Histology 组织学 [hɪˈstɒlədʒi]Embryology 胚胎学 [embriˈɒlədʒi]tissue 组织 [ˈtɪʃu:]Extracellular matrix 细胞外基质 [ˌekstrəˈseljuləˈmeɪtrɪks] Light microscope 光学显微镜 [lait ˈmaikrəskəup] Electron microscope 电子显微镜 [iˈlektrɔn ˈmaikrəskəup] Paraffin sectioning 石蜡切片 [ˈpærəfɪn ˈsekʃən]Hematoxylin cosin staining 苏木精-伊红染剂[hi:məˈtɔksilin ˈi:əusin ˈsteini]Histochemistry 组织化学Immunohistochemistry 免疫组织化学[ɪmjʊnəʊhɪstəʊ'kemistri] Cell culture 细胞培养 [sel ˈkʌltʃə] Tissue engineering 组织工程Epithelium 上皮 [ˌepɪ'θi:lɪəm] Endothelium 内皮 [ˌendəʊ'θi:lɪəm] Mesothelium 间皮 [ˌmezə'θi:lɪəm]Exocrine gland 外分泌腺 [ˈeksəukrain ɡlænd]Endocrine gland 内分泌腺 [ˈendəukrain ɡlænd]Acinus 腺泡['æsɪnəs]Serous cell 浆液细胞 [ˈsiərəs sel]Mucous cell 粘液细胞 [ˈmju:kəs sel]Serous demilune 浆液半月 [ˈsiərəs ˈdemilu:n]Micro villus 微绒毛 [maɪkrəʊ'vɪləs]Cilium 纤毛 ['sɪlɪəm]Desmosome 桥粒 ['desməsəm]Junctional complex 连接复合体Basement membrane 基膜 [ˈbeismənt ˈmemˌbreɪn]Basal lamina 基板 [ˈbeisəl ˈlæminə]Reticular lamina 网板 [riˈtikjuləˈlæminə]loose connective tissue 疏松结缔组织 [kəˈnektɪv ˈtisju:]Connective tissue proper 固有结缔组织 [kəˈnektɪv ˈtisju: ˈprɔpə] Mesenchyme 间充质 ['mezənkaɪm]Fibroblast 成纤维细胞 ['faɪbrəblæst]Fibrocyte 纤维细胞 ['faɪbroʊsaɪt]Plasma cell 巨噬细胞 [ˈplæzmə sel]Macrophage 浆细胞[ˈmækrəfeɪdʒ]mast cell 肥大细胞[mɑ:st sel]fat cell 脂肪细胞 [fat sel]mesenchymal cell 间充质细胞 [mes'eŋkɪməl][sel] Collagenous fiber 胶原纤维 [kɒ'lɑ:dʒenəs]['faɪbə] Elastic fiber 弹性纤维 [iˈlæstik ˈfaibə]Reticular fiber 网状纤维 [rɪ'tɪkjʊlə]['faɪbə] Groung substance 基质 [ɡraund ˈsʌbstəns] Adipose tissue 脂肪组织 [ˈædɪpəʊs][ˈtɪʃu:] Reticular tissue 网状组织 [rɪ'tɪkjʊlə][ˈtɪʃu:] plasma 血浆 [ˈplæzmə] Serum 血清 [ˈsɪərəm]wright staining 瑞氏染色 [rait][steɪnɪŋ] erythrocyte ,red blood cell 红细胞 [ɪˈrɪθrəsaɪt] Hemoglobin 血红蛋白 [ˌhi:məʊ'gləʊbɪn] erythrocyte membrane skeleton 红细胞膜骨架[ɪˈrɪθrəsaɪt] ['membreɪn][ˈskelɪtn]Hemolysis 溶血 [hɪ'mɒlɪsɪs] reticulocyte 网织红细胞 [rɪ'tɪkjʊləsaɪt] leukocyte,white blood cell 白细胞 ['lu:kəˌsaɪt] neutrophilic granulocyte ,neutrophil 中性粒细胞[nju:trə'fɪlɪk] ['grænjʊləsaɪt]， ['nju:trəfɪl]azurophilic granule 嗜天青颗粒[æʒʊərə'fɪlɪk][ˈgrænju:l]specific granule 特殊颗粒 [spəˈsɪfɪk][ˈgrænju:l] basophilic granulocyte 嗜碱性颗粒[bæsə'fɪlɪk]['grænjʊləsaɪt]eosinophilic granulocyte,eosinophil 嗜酸性颗粒[ˌi:əˌsɪnə'fɪlɪk] ['grænjʊləsaɪt]，[ˌi:ə'sɪnəfɪl]monocyte 单核细胞 ['mɒnəsaɪt] lymphocyte 淋巴细胞 [ˈlɪmfəsaɪt] blood platelet 血小板 [blʌd] [ˈpleɪtlət] bone marrow 骨髓 [bəʊn] [ˈmæro]hemopoietic stem cell 造血干细胞[ˌhi:məpɔɪ'i:tɪk] [stem] [sel]multipotential stem cell 多能干细胞[mʌltɪpəʊ'tenʃl] [stem] [sel]Cartilage tissue 软骨组织 [ˈkɑ:tlɪdʒˈtisju:]Chondrocyte 软骨细胞 [kʌdrɒsɪt]Cartilage lacuna 软骨陷窝 [ˈkɑ:tlɪdʒ ləˈkju:nə]Isogenous group 同源细胞群 [aiˈsɔdʒinəs ɡru:pCartilage capsule 软骨囊 [ˈkɑ:tlɪdʒˈkæpsju:l]Hyaline cartilage 透明软骨 [ˈhaiəli:n ˈkɑ:tlɪdʒFibrous cartilage 纤维软骨 [ˈfaɪbrəs]Elastic cartilage 弹性软骨[ɪˈlæstɪk]Chondroblast 成软骨细胞 ['kɒndrəʊblɑ:st]Osseous tissue 骨组织 [ˈɔsi:əs ˈtisju:]Bone matrix 骨基质 [bəun ˈmeɪtrɪks]Osteoid 类骨质['ɒstɪɔɪd]Bone lamella 骨板[bəun ləˈmelə]Osteoprogenitor cell 骨祖细胞Osteoblast 成骨细胞 ['ɒstɪəblæst]Matrix vesicle 基质小炮 [ˈvɛsɪkəl]Osteocyte 骨细胞['ɒstɪəsaɪt]Bone lacuna 骨陷窝 [bəun ləˈkju:nə]Bone canaliculus 骨小管 [bəun ˌkænəˈlikjuləs] Osteoclast 破骨细胞Perforating canal 穿通管Circumferential lamella 环骨板 [səˌkʌmfəˈrenʃəl ləˈmelə] Haversian system 哈弗斯系统 [həˈvə:ʃən ˈsistəm] Osteon 骨单位 ['ɒstɪɒn]Skeletal muscle 骨骼肌 [ˈskelitl ˈmʌsl]Cardiac muscle 心肌 [ˈkɑ:diæk]Smooth muscle 平滑肌Myofibril 肌原纤维 [ˌmaɪə'faɪbrəl]Sarcomere 肌节 ['sɑ:kəmɪə]Sarcoplasm 肌浆 ['sɑ:kəʊˌplæzəm] Sarcoplasmic reticulum 肌浆网 ['sɑ:kəʊˌplæzəm][rɪ'tɪkjʊləm] Intercalated disk 闰盘Transverse tubule 横小管 [ˈtrænzvɜ:s]['tju:bju:l] Longitudinal tubule 纵小管 [ˌlɒŋgɪˈtju:dɪnl]['tju:bju:l] Terminal cisternae 终池 [si'stə:ni:]Triad 三联体 [ˈtraɪæd]Thick filament 粗肌丝 [θik ˈfɪləmənt]Thin filament 细肌丝 [θin ˈfɪləmənt] nervous tissue 神经组织 [ˈnə:vəs ˈtisju:] neuron 神经元 [ˈnʊərˌɔn, ˈnjʊər-] Neuroglial cell 神经胶质细胞 [n'jʊərəʊɡlɪəl ‘ sel] Nissl body 尼氏体 [ˈbɒdi] Neurotransmitter 神经胶质 [ˈnjʊərəʊtrænzmɪtə(r)] Neurofibril 神经原纤维 [ˌnjʊərə'faɪbrɪl] Dendrite 树突 [ˈdendraɪt]Axon 轴突 [ˈæksɒn]Axolemma 轴膜['æksəʊlemə]Axoplasm 轴浆[æk'sɒplæzəm] Pseudounipolar neuron 假单极神经元[sju:dəʊnaɪ'pəʊlə][ˈnʊərˌɔn, ˈnjʊər-]Synapse 突触 ['saɪnæps]Presynaptic element 突触前成分 [prisiˈnæptik ˈelimənt] Synaptic cleft 突触间隙 [sɪˈnæptɪk kleft] Postsynaptic element 突触后成分 [pəustsiˈnæptik ˈelimənt] Postsynaptic membrane 突触后膜 [pəustsiˈnæptik ˈmemˌbreɪn] presynaptic membrane 突触前膜 [prisiˈnæptik ˈmemˌbreɪn] Synaptic knob 突触小体 [sɪˈnæptɪk nɔb]Astrocyte 星形胶质细胞['æstrəsaɪt] Oligodendrocyte 少突胶质细胞 ['ɒlɪgəʊ'dendrəsaɪt] Ependymal cell 室管膜细胞 [e'pendɪməl ‘sel] Schwann cell 施万细胞 [ʃwɔn ‘sel]Myelin sheath 髓鞘 [ˈmaiəli(:)n ʃi:θ] Myelinated nerve fiber 有髓神经纤维 [ˈmaiəlineitid nə:v ˈfaɪbə] Ranvier node 郎飞结 [‘ræviə‘nəʊd] Internode 中间体 ['ɪntənəʊd]Tactile corpuscle 触觉小体 [ˈtæktəl ˈkɔ:pəsəl] Lamellar corpuscle 环层小体 [lə'melə][ˈkɔ:pʌsl] Neuromuscular junction 神经肌连接[ˌnjʊərəʊ'mʌskjʊlə 'dʒʌŋkʃn]epineurium 神经外膜 [ˌepɪ'njʊərɪəm] perineurium 神经束膜 [ˌperə'nju:rɪəm] endoneurium 神经内膜 [endəʊ'nju:rɪəm]motor end plate 运动终板 [ˈməutə end pleit] tunica intima 内膜 [ˈtju:nikəˈintimə] Tunica media 中膜 [ˈtju:nikəˈmi:djə] Tunica adventitia 外膜 [ˈtju:nikəˌædvenˈtiʃjə] Endocardium 心内膜 [endəʊ'kɑ:dɪəm] Myocardium 心肌膜 [maɪə'kɑ:dɪəm] Epicardium 心外膜 [ˌepɪ'kɑ:dɪəm]arteriole 微动脉 [ɑ:ˈtɪəriəʊl]Venule 微静脉 ['venju:l]Capillary 毛细血管 [kəˈpɪləri]elastic membrane 弹性膜 [iˈlæstik ˈmemˌbreɪn] Pericyte 周细胞 [peri:'saɪt]continuous capillary 连续毛细血管[kənˈtinjuəs ˈkæpəˌleri:]Fenestrated capillary 有孔毛细血管 [fiˈnestreitid ˈkæpəˌleri:] Sinusoid capillary 血窦 ['saɪnəsɔɪd ˈkæpəˌleri:] Purkinje fiber 浦肯野纤维[pu ken ye~（就是音译） 'faɪbə]Microcirculation 微循环 [maɪkrəʊsɜ:kjʊ'leɪʃn]skin 皮肤[skɪn]epidermis 表皮 [,epɪ'dɜːmɪs]keratinocyte 角质形成细胞 [kə'rætinəsait]stratum basale 基底层 [ˈstrɑ:təm] [beɪseɪl] stratum spinosum 棘层 [ˈstrɑ:təm][spaɪ'nəʊsʌm] stratum granulosum 颗粒层 [ˈstrɑ:təm]stratum lucidum 透明层 [ˈstrɑ:təm] ['lu:si:dəm] stratum corneum 角质层 [ˈstrɑ:təm]['kɔ:niəm] melanocyte 黑素细胞 ['melənəsaɪt]langerhans cell 朗格汉斯细胞 [sel]dermis 真皮 ['dɜːmɪs]hair 毛 [heə]sebaceous gland 皮脂腺 [sɪ'beɪʃəs][glænd]sweat gland 汗腺[swet][glænd] recirculation of lymphocyte 淋巴细胞再循环 [ri:'s ɜ:kjʊ'leɪʃən] [ˈlɪmfəsaɪt]mononuclear phagocytic system 单核吞噬细胞系统[mɒnəʊn'ju:klɪər][fægə'sɪtɪk][ˈsɪstəm]dendritic cell 树突状细胞 [ˌden'drɪtɪk][sel] Diffuse lymphoid tissue 弥散淋巴组织[dɪˈfju:s]['lɪmfɔɪd][ˈtɪʃu:]Lymphoid nodule 淋巴小结 [ˈnɒdju:l]Germinal center 生发中心 [ˈdʒə:minl]['sentə] Thymic lobule 胸腺小叶 ['θaɪmɪk] ['lɒbju:l] Thymocyte 胸腺细胞 ['θaɪməsaɪt]Thymic corpuscle 胸腺小体 [ˈθaimik ˈkɔ:pəsəl] Blood-thymus barrier 血胸屏障[ˈθaɪməs][ˈbæriə(r)] Supercial cortex 浅层皮质 [ˌsu:pəˈfɪʃl][ˈkɔ:teks] Paracortex zone 副皮质区 [pærə'kɔ:teks]Cortical sinus 皮质淋巴窦 ['kɔ:tɪkl]Medullary cord 髓索 ['medələrɪ]Medullary sinus 髓窦 ['medələrɪ]White pulp 白髓 [pʌlp]Red pulp 红髓 [pʌlp]Periarterial lymphatic sheath 动脉周围淋巴鞘[pɪə'rɪətɪərɪəl][lɪm'fætɪk][ʃi:θ]Marginal zone 边缘区Splenic cord 脾索 ['splenɪk][kɔ:d]Splenic sinus 脾血窦 ['splenɪk] [ˈsaɪnəs] endocrine system 内分泌系统 [ˈend əukrain ˈsistəm]hormone 激素 ['hɔ:məʊn]paracrine 旁分泌 [pəræk'raɪn]thyroid follicle 甲状旁腺滤泡 [ˈθaɪˌrɔɪd ˈfɔlɪkəl] parafollicular cell 滤泡旁细胞[pærə'fɒlɪkjʊlə]zona glomerulosa 球状带['zoʊnə][ɡlɒmrjʊ'loʊzə]zona fasciculate 束状带 ['zoʊnə][fə'sɪkjʊˌleɪt] zona reticularis 网状带 ['zoʊnə]chromaffin cell 嗜铬细胞 [ˈkroməfɪn sɛl]pars distalis 远侧部 [pɑ:z] [dɪs'təlɪs] acidophil 嗜酸性细胞['æsɪdoʊˌfɪl]basophil 嗜碱性细胞[bæsə'fɪl] chromophobe cell 嫌色细胞 [ˈkroməˌfob sɛl]herring body 赫令体 [ˈhɛrɪŋ]gonadotroph 促性腺激素细胞 [ɡənədət'rɒf]pituicyte 垂体细胞 [pɪ'tju:ɪˌsaɪt] somatotroph 生长激素细胞 ['soʊmətətroʊf] hypophyseal portal system 垂体门脉系统 [haɪ'pɒfəsi:l]['pɔ:tl] Digestive system 消化系统 [daɪˈdʒestɪv ˈsistəm] Mucosa 粘膜 [mju:'kəʊsə] Submucosa 粘膜下层 [sʌbmju:'kəʊsə] Muscularis 肌层 ['mʌskjʊlærɪs] Adventitia 外膜 [ˌædvən'tɪʃɪə]Plica 皱襞 ['plaɪkə]Serosa 浆膜 [sɪ'rəʊsə]Gastric area 胃小凹 [ˈgæstrɪk ˈɛəriə] Fundic gland 胃底腺 [ˈfʌndik ɡlænd] Parietal cell 壁细胞 [pəˈraiətəl sel] Oxyntic cell 泌酸细胞 ['ɒksɪntɪk sel]Chief cell 主细胞 [tʃi:f sel] Intracellular secretory canaliculus 细胞内分泌小管[ˌɪntrəˈseljələ siˈkri:təri ˌkænəˈlikjuləs]Intestinal villus 肠绒毛 [ɪnˈtestənəl ˈviləs] Absorptive cell 吸收细胞 [əbˈsɔ:ptiv sel] Paneth cell 潘氏细胞Duodenal gland 十二指肠腺 [ˌdju(:)əuˈdi:nl ɡlænd] Central lacteal 中央乳糜管 [ˈsentrəl ˈlæktiəl] digestive gland 消化腺 [daɪˈdʒestɪv ɡlænd] intercalated duct 闰管 [ɪntɜ:kə'leɪtɪd dʌkt] centroacinar cells 泡心细胞pancreas islet 胰岛 [ˈpæŋkri:əs ˈailit] hepatic lobule 肝小叶 [hɪˈpætɪk ˈlɔbju:l] central vein 中央静脉 [ˈsentrəl vein] Hepatocyte 肝细胞 ['hepətəsaɪt]hepatic plate 肝板 [hɪˈpætɪk pleit]Kupffer cell 肝巨噬细胞perisinusoidal space 窦周隙bile canaliculi 胆小管 [baɪl kænə'likjulai] portal area 门管区 [ˈpɔ:təl ˈɛəriə] respiratory system 呼吸系统 [ˈrespərəˌtɔ:ri]['sɪstəm] trachea 气管 [trə'ki:ə]brush cell 刷细胞 [brʌʃ][sel]ciliated cell 纤毛细胞 ['sɪlɪeɪtɪd][sel] bronchus 支气管 [ˈbrɒŋkəs]lung 肺 [lʌŋ]respiratory bronchiole [ˈrespərəˌtɔ:ri] alveolar duct 肺泡管[ælˈvi:ələ(r)][dʌkt]alveolar sac 肺泡囊[ælˈvi:ələ(r)][sæk] pulmonary alveolus 肺泡 [ˈpʌlmənəri][ælˈvi:ələs] alveolar septum 肺泡隔[ælˈvi:ələ(r)][ˈseptəm] pulmonary macrophage 肺巨噬细胞 [ˈpʌlmənəri][ˈmækrəfeɪdʒ] blood-air barrier 气-血屏障 [blʌd] [eə(r)] [ˈbæriə(r)] Nephron 肾单位 ['nefrɒn]Medullary ray 髓放线 [meˈdʌləri rei] Uriniferous tubule 泌尿小管['jʊərə'nɪfərəs] ['tju:bju:l]Renal corpuscle 肾小体 [ˈri:nəl ˈkɔ:pəsəl] Glomerulus 血管球 [gləʊ'meərjʊləs]Renal capsule 肾小囊 [ˈri:nəl ˈkæpsju:l]Renal tubule 肾小管 [ˈri:nəl ˈtju:bju:l] Podocyte 足细胞 [pɒdə'saɪt]Proximal tubule 近端小管 [ˈprɔksiməl ˈtju:bju:l] Distal tubule 远端小管 [ˈdistəl ˈtju:bju:l]Brush border 刷状缘 [ˈdistəl ˈtju:bju:l] Macula densa 致密斑 ['mækjʊlə]Filtration barrier 滤过屏障[filˈtreiʃən ˈbæriə]Renin 肾素Juxtaglomerular complex 球旁复合体[ˌdʒʌkstəˌɡlɔˈmeruləˈkɔmpleks]seminiferous tubule 生精小管 [ˌsemə'nɪfərəs] ['tju:bju:l] Spermatozoa 精子细胞 [ˌspɜ:mətəˈzəʊə]spermatogonium 精原细胞 [ˌspɜ:mətə'gəʊnɪəm] Spermatocyte 精母细胞 [spə'mætəsaɪt] spermatogenesis 精子发生 [spɜ:mətəʊ'dʒenɪsɪs] spermatogenic cell 生精细胞 [ˌspə:mətəˈdʒenik sel] Acrosome 顶体 ['ækrəˌsəʊm] Sperminogenesis 精子形成[ˌspɜ:mɪəʊ'dʒenəsɪs] Epididymis 附睾Sustentacular cell 支持细胞[ˌsʌstenˈtækjulə sel] Prostate 前列腺 [ˈprɒsteɪt]Blood-testis barrier 血-睾屏障 [ˈtestɪs]Testicular interstitial cell 睾丸间质细胞Androgen binding protein 雄激素结合蛋白[ˈændrədʒən ˈbaɪndɪŋˈprəuti:n]Female reproductive system 女性生殖系统['fi:meɪl][ˌri:prəˈdʌktɪv]['sɪstəm]Vary 卵巢 ['veərɪ]Follicle 卵泡 [ˈfɒlɪkl]Primordial follicle 原始卵泡 [praɪˈmɔ:di:əl ˈfɔlɪkəl] Primary follicle 初级卵泡 [ˈpraiməri ˈfɔlɪkəl] Follicular theca 卵泡膜 [fəˈlikjuləˈθi:kə]Secondary follicle 次级卵泡 [ˈsekəndəri ˈfɔlɪkəl] Mature follicle 成熟卵泡 [məˈtjuəˈfɔlɪkəl] Oogonia 卵原细胞Secondary oocyte 次级卵母细胞 [ˈsekəndəri ˈəuəsait] Ovulation 排卵 [ˌɒvjʊ'leɪʃn]Ovum 卵细胞 [ˈəʊvəm]Zona pellucida 透明带 [ˈzəʊnəpəˈlu:sɪdə, pelˈju:-] Corona radiata 放射冠 [kə'rəʊnə]Corpus luteum 黄体 ['kɔ:pəs][ˈlu:ti:əm] Granulosa lutein cell 颗粒黄体细胞[grænjʊ'ləʊsə]['lu:tɪɪn][sel]Theca lutein cell 膜黄体细胞 [ˈθi:kəˈlu:tiin sel] Uterus 子宫 ['ju:tərəs]Uterine gland 子宫腺 [ˈju:tərain ɡlænd] Mammary gland 乳腺 [ˈmæməri ɡlænd]Germ cell 生殖细胞 [dʒə:m sel]Gamete 配子 [ˈgæmi:t]Capacitation 获能 [kəpæsɪ'teɪʃən] Fertilization 受精 [ˌfɜ:təlaɪ'zeɪʃn] Acrosome reaction 顶体反应['ækrəˌsəʊm riˈækʃn] Zone reaction 透明带反应 [zəʊn riˈækʃn]Male pronucleus 雄原核 [meil prəˈnju:kliəs]Female pronucleus 雌原核 [ˈfi:meil prəˈnju:kliəs] Fertilized ovum/zygote 受精卵/合子[ˈfɜ:təlaɪzd ˈəʊvəm/ˈzaɪgəʊt]Cleavage 卵裂 [ˈkli:vɪdʒ]Blastomere 卵裂球 [blɑ:stə'mɪə]Morula 桑椹胚 ['mɔ:rʊlə]Blastocyst 胚泡['blæstəsɪst] blastocoele 胚泡腔 [blɑ:stəʊ'kəʊl] Trophoblast 滋养层 ['trɒfəblæst]Inner cell mass 内细胞群 [ˈinəsel mæs] Implantation/imbed 植入/着床 [ˌɪmplɑ:n'teɪʃn]/[imˈbed] Syncytiotrophoblast 合体滋养层 [sɪnsɪti:əʊt'rɒfəʊblæst] Cytotrophoblast 细胞滋养层 [saɪtəʊ'trɒfəblæst] Decidua 蜕膜 [dɪ'sɪdjʊə]Decidual cell 蜕膜细胞 [dɪsɪd'jʊəl]Decidua basalis 基蜕膜 [dɪ'sɪdjʊə 'beɪsəlɪs ] Decidua capularis 包蜕膜 [dɪ'sɪdjʊə？]Decidua parietalis 壁蜕膜 [dɪ'sɪdjʊə？] Embryonic disc 胚盘 [ˌembri:ˈɔnɪk disk] Epiblast 上胚层 ['epɪblæst]Hypoblast 下胚层 ['haɪpəblæst]body stalk 体蒂 ['bɒdɪ] [stɔːk]primitive streak/node/pit 原条 ['prɪmɪtɪv][stri:k]/[nəʊd]/[pɪt] intraembryonic mesoderm 胚内中胚层 [ɪntrə,embrɪ'ɒnɪk] ['mesədɜ:m]mesoderm 中胚层 ['mesədɜ:m]ectoderm 外胚层 ['ektəʊˌdɜ:m]endoderm 内胚层['endəʊdɜ:m]notochord 脊索 ['nəʊtəˌkɔ:d]neural plate/groove/fold/tube 神经板/沟/褶/管[ˈnjʊərəl][pleɪt][fəʊld][tju:b]neural crest 神经嵴[ˈnjʊərəl] [krest]mesenchyme 间充质['mezənkaɪm]paraxial mesoderm 轴旁中胚层[pæ'ræksɪəl]['mesədɜ:m] intermediate mesoderm 间质中胚层 [ˌɪntəˈmi:diət] ['mesədɜ:m]lateral mesoderm 侧中胚层 [ˈlætərəl]['mesədɜ:m]parietal mesoderm 体壁中胚层 [pə'raɪɪtl]['mesədɜ:m]visceral mesoderm 脏壁中胚层[ˈvɪsərəl]['mesədɜ:m]afterbirth 衣胞 [ˈɑ:ftəbɜ:θ]fetal membrane 胎膜['fi:tl][ˈmembreɪn]chorion 绒毛膜 ['kɔ:rɪɒn]amnion 羊膜['æmnɪən]amniotic fluid 羊水[ˌæmnɪ'əʊtɪk][ˈflu:ɪd]yolk sac 卵黄囊 [jəʊk][sæk]umbilical cord 脐带[ʌm'bɪlɪkəl][kɔ:d] placenta 胎盘[pləˈsentə]placental septum 胎盘隔[pləˈsentl][ˈseptəm] placental membrane /placental barrier 胎盘膜/胎盘屏障[pləˈsentl][ˈmembreɪn]/[pləˈsentl][ˈbæriə(r)]twins 双胎 [twɪnz]multiplets 多胎 ['mʌltɪpləts]conjoined twins 联体双胎 [kən'dʒɔɪnd] [twɪnz] Frontonasal process 额鼻 [f'rʌntəʊnəsl][ˈprəʊses] Heart process 心突 [ˈprəʊses]Branchial arch/groove 鳃弓/沟 [b'rɑ:nkɪəl] [gru:v] Pharyngeal pouch 咽囊 [fəˈrɪndʒiəl] [paʊtʃ] Branchial membrane/apparatus 鳃膜/器[b'rɑ:nkɪəl][ˈmembreɪn]/ [ˌæpəˈreɪtəs]Maxillary/mandibular process 上/下颌突[mæk'sɪlərɪ]/[mæn'dɪbjʊlə] Stomodeum 口凹/原始口腔 [stəmɒ'di:əm]Nasal placode/pit 鼻扳/窝 [ˈneɪzl] ['plækəʊd]/[pɪt] Median palatine process 正中腭突 [ˈpælətaɪn]Lateral palatine process 外侧腭突 [ˈlætərəl][ˈpælətaɪn] Dental lamina 牙扳 [ˈdentl] ['læmənə]Tooth bud 牙蕾 [tu:θ bʌd]Enamel organ 造釉器 [iˈnæməl ˈɔ:ɡən]Ameloblast 成釉质细胞[æmə'lɒblæst]Dental papilla 牙乳头 [ˈdentl][pə'pɪlə]Limb bud 上/下肢牙 [lim‘bʌd]Cleft lip 唇裂 [kleft lip]Cleft palate 腭裂 [ˈpælət]Oblique facial 面斜裂 [əˈbli:k]primitive digestive duct 原始消化管 [ˈprɪmətɪv daɪˈdʒestɪv dʌkt] foregut 前肠 ['fɔ:gʌt] midgut 中肠 ['mɪdgʌt] hindgut 后肠 ['haɪndgʌt] midgut loop 中肠袢 [ˈmidˌgʌt lu:p] caecal bud 盲肠突/盲肠芽 ['si:kə bʌd] umbilical coelom 脐腔 [ˌʌmbiˈlaikəl ˈsi:ləm] cloaca 泄殖腔 [kləʊ'eɪkə] urorectal septum 尿直肠隔[ˈseptəm] urogenital sinus 尿生殖窦 [juərəuˈdʒenitl ˈsaɪnəs] urogenital membrance 尿生殖膜 [juərəuˈdʒenitlˈmembreɪn] anal menbrance 肛膜 [ˈeɪnlˈmembreɪn]hepatic diverticulum 肝憩室 [hɪˈpætɪk ˌdaivə:ˈtikjuləm] ventral pancreatic bud 腹胰芽 [ˈventrəl ˌpæŋkriˈætik bʌd] dorsal pancreatic bud 背胰芽 [ˈdɔ:səl ˌpæŋkriˈætik bʌd] thyroglossal cyst 甲状舌管囊肿 [θaɪ'roʊɡlɒsl]Meckel's diverticulum 梅克尔憩室 [ˌdaɪvɜ:'tɪkjʊləm] umbilical fistula 脐瘘；脐粪瘘 [ˌʌmbiˈlaikəl ˈfistjulə] congenital umbilical hernia 先天性脐疝[kənˈdʒenɪtl ˌʌmbiˈlaikəl ˈhə:njə]laryngotracheal groove 喉气管沟 [ˌləriŋɡəuˌtrəˈkiəl ɡru:v] laryngotracheal diverticulum 喉气管憩室[ˌləriŋɡəuˌtrəˈkiəl ˌdaivə:ˈtikjuləm]lung bud 肺芽 [lʌŋ bʌd] tracheoesophageal fistula 气管食管瘘[treikiəui:ˌsɔfəˈdʒi:əl ˈfistjulə]hyaline membrane disease 透明膜病[ˈhaiəli:n ˈmemˌbreɪn diˈzi:z]nephrotome 生肾节 ['nefrəˌtəʊm]urogenital ridge 尿生殖嵴 [juərəuˈdʒenitl ridʒ] mesonephric ridge 中肾嵴 [mi:sə'nefrɪk ridʒ] genital ridge 生殖腺嵴 [ˈdʒenitl ridʒ] pronephros 前肾 [prəʊ'nefrɒs] mesonephros 中肾 [ˌmesəʊ'nefrəs] metanephros 后肾 [ˌmetə'nefrɒs] mesonephric duct/Wolffian duct 中肾管 [mi:sə'nefrɪk]ureteric bud 输尿管芽 [bʌd]metanephrogenic tissue 生后肾组织 [metənɪfrəd'ʒenɪk]primordial germ cell 原始生殖细胞[praɪˈmɔ:di:əl dʒə:m sel]paramesonephric duct 中肾旁管Blood island 血岛 [blʌd ˈailənd] Primitive cardiovascular system 原始心血管系统[ˈprɪmətɪv ˌkɑ:diəʊˈvæskjələ(r) 'sɪstəm]Vitelline artery 卵黄动脉 [viˈtelin ˈɑ:təri] Umbilical artery 脐动脉 [ˌʌmbiˈlaikəl ˈɑ:təri] Aortic artery 弓动脉 [eɪ'ɔ:tɪk ˈɑ:təri] Anterior cardinal vein 前主静脉[ænˈtɪəri:əˈkɑ:dinl vein] Posterior cardinal vein 后主静脉 [pɔˈstɪəri:əˈkɑ:dinl vein] Common cardinal vein 总主静脉 [ˈkɔmən ˈkɑ:dinl vein] Vitelline vein 卵黄静脉 [viˈtelin vein] Umbilical vein 脐静脉 [ˌʌmbiˈlaikəl vein] Pericardial coelom 围心腔 [ˌperiˈkɑ:diəl ˈsi:ləm] Cardiogenic plate 生心扳 [ˌkɑ:diəuˈdʒenik pleit] Cardiogenic tube 心管Myoepicardial mantle 心肌外套层 [ˌmaɪə'kɑ:dɪəl] [ˈmæntl] Cardiac jelly 心胶质 [ˈkɑ:di:ˌæk ˈdʒeli:] Bulbus cordis 心球 ['bʌlbʌs 'kɔ:dɪs]Sinus venosus 静脉窦 [vi:ˈnəʊsəs]Truncus arteriosus 动脉干 [ˈtrʌŋkəs]Bulboventricular loop 球室袢 [bʌlbʌvent'rɪkjʊlə lu:p] Atrioventricular canal 房室管[ˌeitriəuvenˈtrikjulə kəˈnæl]Endocardial cushion 心内膜垫 [ˌendəʊ'kɑ:dɪəl ˈkuʃənz] Foramen ovale 卵圆孔 [əʊˈvæli:, -ˈveɪli:, -ˈvɑ:-] Truncal ridge 动脉干嵴 ['trʌŋkl rɪdʒ]Bulbar ridge 球嵴 [ˈbʌlbə ridʒ]Aorico-pulmonary septum 主动脉肺动脉隔[eɪɔ:tɪkəʊ'pʌlmənərɪˈseptəm]Atrial septal defect 房间隔缺损 [ˈeitriəl ˈseptl diˈfekt] Ventricular septal defect 室间隔缺[venˈtrikjuləˈseptl diˈfekt] Tetralogy of Fallot 法络四联症[te'trælədʒɪ] [fæˈləʊ]。

不同脑小血管病负荷评分与伴无症状腔隙的脑小血管病患者认知功能的关系杜晓光，魏荣，刘琦慧，于力群，周丽摘要：目的　探讨不同脑小血管病（CSVD）负荷评分与伴无症状腔隙的CSVD患者认知功能的关系。

方法　纳入2021年7月—2023年10月就诊于潍坊市人民医院神经内科的128例伴无症状腔隙的CSVD患者，运用蒙特利尔量表（MoCA）、CSVD总负荷评分和改良负荷评分统计受试者的认知功能和CSVD负荷，分为认知障碍组（MoCA<26分）和无认知障碍组（MoCA≥26分），比较两组患者的人口社会学信息、血管病危险因素及CSVD负荷评分的差异。

采用线性回归分析MoCA评分与两种CSVD负荷评分的关系，采用趋势检验分析伴无症状腔隙的CSVD 患者认知障碍的发病趋势。

结果　研究共纳入伴无症状性腔隙的CSVD患者128例，其中认知障碍组68例（53.1%），无认知障碍组60例（46.9%），两组患者人口社会学信息及血管病危险因素差异无统计学意义（P>0.05）。

两组患者的CSVD总负荷评分和改良负荷评分比较均存在统计学差异（P<0.05）。

Spearman秩相关分析显示，CSVD 总负荷评分和改良负荷评分均与MoCA评分呈负相关（P<0.001）。

线性趋势χ2检验分析显示，伴无症状腔隙的CSVD患者认知障碍发病风险随CSVD改良负荷评分增加而增加（P trend<0.05），该发病风险与CSVD总负荷评分间趋势分析无统计学意义（P trend=0.069）。

结论　CSVD总负荷评分和改良负荷评分均可用于筛检伴无症状腔隙的CSVD认知障碍患者。

改良负荷评分可能在识别认知障碍高风险患者方面更具优势。

关键词：脑小血管病；认知；腔隙中图分类号：R743 文献标识码：ARelationship between cerebral small vessel disease burden scores and cognitive function in patients with cerebral small vessel disease with asymptomatic lacunes　DU Xiaoguang，WEI Rong，LIU Qihui， et al.（Department of Neurol⁃ogy， Weifang People′s Hospital， Weifang 261000， China）Abstract：Objective To investigate the relationship between cerebral small vessel disease （CSVD） burden scores and cognitive function in patients with CSVD with asymptomatic lacunes.Methods A total of 128 patients with CSVD with asymptomatic lacunes who visited the Department of Neurology of Weifang People′s Hospital from July 2021 to October 2023 were included. All the patients were scored using the Montreal Cognitive Assessment （MoCA） for cognitive function and using the total CSVD score and the modified CSVD score for CSVD burden. They were divided into cognitive impairment group （MoCA score<26）and non-cognitive impairment group （MoCA score≥26）.The demographic information，vascular disease risk factors， and the CSVD scores of the two groups were compared. A linear regression analysis was performed to as⁃sess the relationship between the MoCA score and the two CSVD scores. A trend analysis was conducted to analyze the trend of incidence of cognitive impairment in patients with CSVD with asymptomatic lacunes.Results Among the 128 patients with CSVD with asymptomatic lacunes， 68 （53.1%） were in the cognitive impairment group and 60 （46.9%） were in the non-cognitive impairment group. There were no significant differences in the demographic information and vascular disease risk factors between the two groups （P>0.05）. The total CSVD score and the modified CSVD score differed significantly be⁃tween the two groups （P<0.05）. The Spearman correlation analysis showed that the total and modified CSVD scores were sig⁃nificantly negatively correlated with the MoCA score （P<0.001）. The chi-square test for linear trend revealed that the cogni⁃tive impairment risk increased significantly with the modified CSVD score in patients with CSVD with asymptomatic lacunes （P trend<0.05）， but with no significance for the total CSVD score （P trend=0.069）.Conclusion Both the total and modified CSVD scores are useful tools to detect cognitive impairment in patients with CSVD with asymptomatic lacunes， and the modi⁃fied CSVD score may be superior in identifying patients at high risk of cognitive impairment.Key words：Cerebral small vessel disease；Cognition；Lacune脑小血管病（cerebral small vessel disease，CSVD）是血管性认知障碍的主要原因［1］。

海藻酸盐基微囊膜的物质传递性能王云红;刘袖洞;于炜婷;戴小敏;马小军【摘要】Natural polysaccharide alginate shows good biocompatibility,biodegradability,adhesi-on,as well as unique deformation stress-release characteristics,so that alginate-based microcapsules with outer polyelectrolyte complex membrane are promising carriers for cell culture and sustained/controlled release.Membrane permeability can decide the diffusion and mass transfer properties of alginate-based microcapsules,which is critical for its application in the biomedical field.We mainly discussed the mathematical models of mass transfer on microcapsule membrane,the diffusion behavior of smallmolecules,macromolecules (such as proteins) and oxygen in alginate-based microcapsules membrane,summarized the influence rules of parameters such as materials properties,pH on membrane permeability.It is hopeful to provide reference for the preparation and application of alginate-based microcapsules.%天然多糖海藻酸钠具有良好的生物相容性、生物降解性、黏附性以及独特的应力形变-释放特性,因此,海藻酸钠外覆聚电解质复合膜形成的微胶囊是很有前途的细胞培养和药物缓控释载体.膜的渗透分离特性决定了海藻酸盐基微囊的物质扩散传递行为,对其在生物医学领域的应用至关重要.本文主要综述了海藻酸盐基微囊膜的传质数学模型研究,概述了小分子物质、大分子蛋白以及氧气在海藻酸盐基微囊膜上的扩散传递行为,总结了材料物化性能、pH值等参数对微囊膜渗透性能的影响规律,对海藻酸盐基微胶囊的制备和应用具有较大的参考价值.【期刊名称】《膜科学与技术》【年(卷),期】2017(037)002【总页数】7页(P6-11,18)【关键词】海藻酸钠;微囊膜;物质传递【作者】王云红;刘袖洞;于炜婷;戴小敏;马小军【作者单位】大连大学环境与化学工程学院,大连116622;大连大学环境与化学工程学院,大连116622;中国科学院大连化学物理研究所,大连116023;大连大学环境与化学工程学院,大连116622;中国科学院大连化学物理研究所,大连116023【正文语种】中文【中图分类】TQ028.8海藻酸钠(简写为NaAlg或Alg)是存在于褐藻类海洋生物中的线性阴离子天然多糖，由β-D-甘露糖醛酸(mannuronate,M)和α-L-古罗糖醛酸(guluronate,G)通过(1-4)糖苷键连接，以poly-GG、poly-MG、poly-MM片段随机排列组成的共聚物[1-2].海藻酸钠通过与二价阳离子(如Ca+, Ba2+)间的离子移变作用发生凝胶化反应，条件温和，工艺简单；且材料具有良好生物相容性、生物降解性、价格低廉、易于加工成球形微囊等特点，使得海藻酸盐基微胶囊技术在动植物细胞培养、微生物及酶的固定化、蛋白质等大分子药物控释等领域有大量的研究报道.海藻酸钙凝胶微球是常用的一种形式，但它在应用过程中的稳定性不理想.研究者通过聚阳离子与海藻酸盐羧基的相互作用生成聚电解质复合(polyelectrolyte complex, PEC)高分子膜，以增强微胶囊的机械稳定性.络合的聚阳离子材料有聚氨基酸类(如聚赖氨酸[3]、聚鸟氨酸、聚精氨酸、聚组氨酸[4]等),聚胺类(如聚乙烯亚胺[5]、聚亚甲基胍[6]、聚N-乙烯基己内酰胺[7]、羧基-丙基-丙烯酰胺共聚物、DEAE-dextran[8]、氨基聚乙二醇[9]等),壳聚糖等.PEC膜的引入，在微囊表面形成一层更致密的物质选择透过膜，从而直接影响微胶囊的物质传递行为，而微囊膜的传质行为又是决定微胶囊应用效果的重要因素.因此，本文综述了海藻酸盐基微囊膜的传质行为及其影响因素，为海藻酸盐基微胶囊技术在医药领域的应用提供参考和依据.1 海藻酸盐基微囊膜的形成原理1.1 海藻酸盐凝胶的形成原理海藻酸钠分子结构如图1所示，以Ca+为例，通过理论分析与实验研究，认为Ca+ 引发海藻酸盐凝胶机理如下[10]：1个Ca+与海藻酸钠分子链段中2个GG片段通过4个配位键形成具有2个六元环结构的稳定螯合物，即“蛋格”结构(图2)，其中，由G 单元的5-COO-和2-OH参与配位键形成.图1 海藻酸钠分子结构Fig.1 Molecular structure of sodium alginate图2 海藻酸钙凝胶的“蛋格”结构Fig.2 The “egg-box” structure of calcium alginate gel1.2 海藻酸盐基微囊膜的形成原理海藻酸盐凝胶通常作为微胶囊囊芯，而其表面分子与聚阳离子通过静电络合作用生成PEC膜，常被称为海藻酸盐基微囊膜，其多孔网络结构赋予了物质选择透过性，即利用膜孔洞的尺寸筛分效应选择性地使得具有一定分子量和空间结构的物质扩散进出.以壳聚糖为例，壳聚糖分子链上的伯氨基与海藻酸盐分子上的羧基发生静电络合作用而形成PEC膜.壳聚糖的结构式及壳聚糖与海藻酸盐的反应机理如图3[11]、图4[12]所示.图3 壳聚糖结构式Fig.3 The structure of chitosan图4 壳聚糖与海藻酸盐反应机理Fig.4 Reaction mechanism of chitosan and alginate2 海藻酸盐基微囊膜的传质规律2.1 海藻酸盐基微囊膜的传质扩散数学模型目前用于描述微囊膜传质扩散的数学模型多建立在Fick定律基础上，但是对于某些比较复杂的微囊化细胞培养系统，小分子物质受微胶囊分布以及细胞等的影响，浓度处于不断变化的过程中，由于Fick扩散定律过于简化，所以不太适用这种复杂的过程.Qi等[13]假设溶质在微囊内外介质中均匀分布，但在膜相中非均匀分布，根据化学势平衡建立了如下数学模型：(1)式中， Cs为t时刻蛋白质在分散介质中的浓度；为蛋白质的在分散介质中的平衡浓度；为蛋白质在分散介质中的初始浓度；τ为时间常数.Song Kedong[14]以海藻酸钙-明胶混合微囊为研究对象，建立了一种非稳态下均匀球体的混合扩散数学模型.具体公式如下：(2)底物溶质从本体溶液扩散到微囊内部，则球体中的底物浓度Cr可通过以下公式来表示：(3)(4)在足够的搅拌条件下，微胶囊周围液体薄膜的阻力可忽略不计.因此，底物在微球表面的浓度等于其在溶液中的浓度，可通过下面的公式来计算：(5)(6)根据Yao[15]的模型，Dm，D1和D2之间的关系可以被定义为如下：(7)在上面的公式中， C0为溶质在溶液中的初始浓度，α是液体体积与固体球体的体积比，R是球体的半径，V是本体溶液的体积，ra是微胶囊的内部半径，而rb是微胶囊的外部半径，Dm为微胶囊的混合扩散系数，D1、D2为微分别为溶质在微胶囊膜中的扩散系数和在微胶囊液芯中的扩散系数，其中溶质在微胶囊液芯中的扩散过程可以被认为等于其在纯水中的扩散过程(扩散系数为Dw)： D2=Dw.通过借助MATLAB软件来解方程，可以求出底物在微囊膜中的扩散系数.2.2 不同物质在海藻酸盐基微囊膜上的传质行为2.2.1 小分子物质在海藻酸盐基微囊膜上的传质行为朱敏莉等[16]考察了4种小分子物质谷氨酸、L-苯丙氨酸、葡萄糖和乳糖在海藻酸钠-壳聚糖微囊膜上的传质性能.结果表明，小分子物质均能顺利通过微囊膜进入囊内，而且物质切割分子量越大，通过微囊膜的时间越长，扩散系数越小，进入微胶囊的难度越大.张宏亮等[17]以传统细胞培养的重要碳源葡萄糖与柠檬酸、氮源L-谷氨酸为小分子模型,发现它们可以自由透过微囊膜进行扩散，并且谷氨酸根或柠檬酸根会与海藻酸根竞争Ca+形成新的配合物，破坏海藻酸钙凝胶“蛋格”结构，微球结构逐渐疏松，微球溶胀.由此可见，小分子物质在海藻酸盐微囊中接近自由扩散，扩散阻力主要集中在微囊膜上.但Ca+的螯合剂会引起海藻酸盐凝胶结构解聚，在微囊化细胞培养时，应考虑培养体系中营养物质对微胶囊结构的影响.2.2.2 大分子物质在海藻酸盐基微囊膜上的传质行为用于评价微囊膜传质的大分子模型主要包括：蛋白及多糖.其中，免疫球蛋白G(IgG)可用来预测微囊膜的免疫隔离性能；牛血清白蛋白(BSA)是球形分子且分子量为66 000，常被用来评价营养物在微囊膜的通透性；中性多糖如葡聚糖、支链淀粉等由于能制备出系列分子量产物，可通过普适标定原则计算微囊膜的通透性.蛋白类分子由于更接近细胞生长所需的营养成分，因此结果可直接回馈到细胞培养过程，但由于其与微胶囊之间更易发生特异性吸附而影响扩散结果的准确性.孙学战等[18]发现，BSA浓度越大，在海藻酸钙-几丁聚糖微囊膜表面吸附量越多，释放速率越快.刘映薇等[19]发现海藻酸钠-壳聚糖微囊膜更像一个“诱捕区”而非简单的屏障，BSA先吸附于膜上，再渗透进入微胶囊内.谢红国等[20]通过研究纤维蛋白原在海藻酸钠-壳聚糖微囊膜表面吸附行为，从微囊膜表面电荷、表面粗糙度等表面性质及蛋白分子的电荷、回转半径等方面揭示了蛋白在微囊膜的吸附本质. 由此可见，蛋白在微囊膜上的扩散过程除受到膜的三维结构、孔径、孔隙率等因素影响外，蛋白质本身的性质、微囊膜的表面性质等都会通过影响蛋白的吸附过程，进而影响其在微囊膜上的传质行为.2.2.3 氧气在在海藻酸盐基微囊膜上的传质行为氧气作为一种难溶且细胞大量消耗的营养成分，在载细胞生物微囊膜的传递中倍受人们的关注，微胶囊内部缺氧是导致内部细胞团出现中心坏死的主要原因.氧气在微囊膜上的扩散行为将直接决定微囊内细胞的生长代谢行为.赵伟等[21]发现，海藻酸钠-壳聚糖聚电解质复合膜是氧传质主要的阻力部位，其溶氧扩散系数为水中的23.3%～43.3%，且随海藻酸钠特性黏度、壳聚糖分子量的增大而减小.Sharp等[22]通过如下公式对微囊化细胞培养系统的氧气传递过程进行了理论分析.(C*-Ci,m,i)≥Q=QO2x(8)式中， VL为液相体积； r0和ri分别为胶囊球体的内外半径；klα为氧气由气相进人液相的物质传递系数；ksα为由液相到固相物质传递系数； DO2,m为微胶囊膜中氧气的扩散系数； C*为平衡状态下气相氧气浓度；Ci,m,i微胶囊膜内表面氧气浓度；为单个微胶囊中细胞的氧气消耗速度；QO2为单个细胞的氧气消耗速度； x为细胞密度.对于微囊化细胞培养来说，只有当氧气由气相进入微囊膜内的传递速度OTRG大于或等于胶囊内细胞生长速度时，才能保证其正常的存活和增殖.3 影响海藻酸盐基微囊膜物质传递性能的因素3.1 材料物性参数对微囊膜传质性能的影响刘博等[23]在固定化酶的研究发现，海藻酸钠浓度越大，形成凝胶的孔径越小，扩散阻力越大，制约微囊膜内脂肪酶与微囊膜外底物的结合；而海藻酸钠的浓度过低，会导致凝胶孔径增大，酶易流失.聚阳离子通过控制成膜反应影响微囊膜的传质性能.研究发现，壳聚糖浓度越高，在浓度差的推动下，其扩散进入海藻酸钙凝胶珠内部越深，形成的微囊膜越厚；同时会增加溶液中的位点，使溶液中与海藻酸钙凝胶分子中—COO-结合的概率增大，形成的微囊膜更致密，传质阻力增大.姜恒丽等[24]、徐旻等[25]也得出与此相一致的结论.Yu等[26]研究表明，壳聚糖分子量越小，分子链段越短，空间位阻越小，越容易扩散到海藻酸钙凝胶网络结构较深区域，但短分子链不能有效拉动相邻海藻酸钠分子链形成致密交联，形成的微囊膜厚而疏松；相反，大分子量壳聚糖形成的微囊膜薄而致密.此外，壳聚糖的脱乙酰度也会影响微囊膜的传质性能[27]，即随着壳聚糖脱乙酰度的提高，微囊膜更致密，传质性能降低.3.2 pH值对微囊膜传质性能的影响海藻酸钠具有pH响应性，对海藻酸盐基微囊膜的传质性能产生影响.通常在酸性介质中，海藻酸盐基微胶囊收缩，而在中性或碱性介质中微胶囊膨胀.这一性能正好符合口服肠道生化微反应器作为基因工程药物生产与释放系统的功能要求,即微胶囊在胃内酸性环境收缩使膜致密,以保护囊内包埋物质免受胃酸及酶的破坏，而微胶囊在肠道近中性环境中膨胀而使膜疏松,以利于包埋物(如益生菌、大分子蛋白类药物等)释放.王慧萍等[28]、冯志云等[29]、唐文等[30]以及张守庆等[31]的研究均得到一致的结论.He等[32]利用精蛋白吸附和仿生硅化的方式制备出海藻酸钙/精蛋白/二氧化硅杂化微胶囊(APSi)，并研究了亚甲基蓝、VB 12、4 000和10 000的葡聚糖这4种不同切割分子量物质在该微囊膜上的扩散性能.当分子大小与微囊膜孔径尺寸相匹配(如VB 12和4 000的FITC-葡聚糖)时，APSi杂化微囊膜会表现出明显的pH 响应控释特性.即当pH>4.5时，海藻酸钙凝胶网络带负电，此时带正电的精蛋白分子由于静电作用被吸附到海藻酸钙凝胶网络上，海藻酸钙凝胶网络结构的扩散通道处于“开”状态；当pH<4.5时，海藻酸钙凝胶网络呈电中性，带正电的精蛋白分子相互排斥地分布于凝胶网络的扩散通道内，因而扩散通道处于"关"的状态.这种pH响应性能在肠靶向智能药物载体方面极具应用广阔的前景[33].3.3 凝胶浴离子强度对微囊膜传质性能的影响徐旻等[25]研究表明，交联剂Ca+的浓度会影响海藻酸盐微囊膜的渗透性能，Ca+浓度越高，海藻酸钙网络结构越紧密，微囊膜的通透性越低，但是当Ca+浓度过高时，形成的凝胶网络尺寸减小，与之成膜的聚阳离子分子进入凝胶的深度浅，且大量海藻酸根链段上的羧基与Ca+结合，使得与聚阳离子结合的羧基数量明显减少，因此形成的膜薄而松散，微囊膜的通透性反而增大.在实验中还发现，在凝胶浴中，引入反凝胶离子如Na+，能显著提高微囊膜的通透性.3.4 其他因素对微囊膜扩散性能的影响除了以上几种因素外，模型物质本身的性质会影响海藻酸盐微囊膜的扩散性能.刘映薇等[19]发现罗丹明异硫氰酸酯(RBITC)标记的胰蛋白酶、卵清白蛋白、溶菌酶等几乎都能够扩散进入微囊膜，但BSA却有部分被微囊膜截留，原因是其回转半径较大.微胶囊粒径大小会影响传质表面积和物质传递距离，因此会明显影响物质在微囊膜的扩散动力学[34].另外，通过层层自组装技术(LBL法)制备的微囊膜，其传质性能与LBL层数呈现显著的相关性.李思阳等[35]的结果表明，随着聚电解质PLL/ALG交替层数的增加，微囊膜变厚，微囊膜的通透性降低，导致药物10-羟基喜树碱(HCPT)释放速率降低，通过调节微胶囊的包覆层数可以控制药物的释放率.该结论在其他报道中也得到证实[36-39].此外，Yu等[26]发现，作为细胞载体的海藻酸钠-壳聚糖微胶囊，膜的通透性能随着细胞的增殖而降低，原因是在壳聚糖成膜及细胞增殖过程中，在静电力及囊内渗透压作用下，微囊内的海藻酸钠分子向膜内表面移动，结合细胞增殖过程产生的大分子代谢产物均挤压在微囊膜的内表面，形成新的“表观膜”，从而增加了物质的扩散阻力.4 结语和展望海藻酸盐基微胶囊因其良好的生物相容性、pH响应性、体积相变性、选择透过性等被广泛应用于细胞培养微反应器、药物控释载体、基因运载工具等方面.微囊膜传质性能是决定其应用效果的关键，尽管海藻酸盐基微囊膜传质行为的相关研究已有很多报道，但实验研究较多，理论研究相对不足.数学模型有利于更好地分析物质在微囊膜上的传质行为，但由于制备材料与工艺参数不同而导致的微囊膜结构差异、物质分子结构及电性差异、微囊内含物性质或活性差异(尤其是细胞的生长情况)以及应用环境差异等都决定了实际应用过程中微囊膜上的物质扩散传递较为复杂.同时，建立模型时为便于计算而引入的假设(如：分子在微囊内均匀分布)与实际情况(存在浓度梯度)不符，导致理论模型往往无法真正得到有效应用.因此，需要借助或建立一些在微囊应用过程中的在线检测方法(如微探针)尽可能获取物质扩散传递的动态信息，再结合计算机软件的强大数据分析功能，去获得更为接近真实情况下的物质扩散模型，为微囊化细胞的规模化培养和自动化控制或者微囊化药物控释剂型研制提供理论指导.参考文献:[1] 刘袖洞, 于炜婷, 王为, 等. 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