Quercetin exerts anti-melanoma activities and inhibits STAT3 signaling
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现代消化及介入诊疗 2020年第25卷第7期ModernDigestion&Intervention2020牞Vol.25牞No.7981 道检查,及时的结肠镜检查有助于早期发现结肠转移,提高患者的生存率。
参考文献[1] 李忠武,王燕,薛卫成,等.Galectin 1与galectin 3在黑色素细胞病变中的表达及预后意义[J].中华病理学杂志,2013,42(12):801 805.[2] 庄步强,张蓬波,任泽强.以全身多发肿块为首发症状的恶性黑色素瘤1例报告[J].中国肿瘤外科杂志,2014,6(2):128 129.[3] 赵滨,阴亚坤.皮肤黑色素瘤中PITX1表达及其与临床病理特征和预后的关系[J].中国中西医结合皮肤性病学杂志,2020,19(1):20 24.[4] 谢炜星,晋大祥.以骨转移灶症状为首发表现的恶性黑色素瘤1例报告[J].中国矫形外科杂志,2020,28(1):88 90.[5] MillsAM,Policarpio NicholasML,AgaimyA,etal.SclerosingMicrocysticAdenocarcinomaoftheHeadandNeckMucosa:ANeoplasmCloselyResemblingMicrocysticAdnexalCarcinoma[J].HeadNeckPathol,2016,10(4):501 508.[6] 石衡,任帅帅,范建平,等.男性飞行员黑色素瘤发生风险的Meta分析[J].西北国防医学杂志,2019,40(8):502 506.[7] 夏翠锋,李云峰,李强,等.原发性降结肠恶性黑色素瘤1例[J].世界华人消化杂志,2009,17(36):3765 3767.[8] VigliarE,MarinoG,MatanoE,etal.Signet ring cellcarcinomaofstomachmetastatictothebladder:acasereportwithcytologicalandhistologicalcorrelationandliteraturereview[J].IntJSurgPathol,2013,21(1):72 75.[9] 沈超,叶颖江,颜艺超,等.胃肠道恶性黑色素瘤二例并文献复习[J].中华临床医师杂志(电子版),2014,8(24):4356 4631.[10]SamoS,SheridM,HuseinH,etal.Metastaticmalignantmelanomatothecolon:acasereportandreviewoftheliterature[J].JGastrointestCancer,2014,45(2):221 224.[11]ParkJS,NgKS,SawRPM,etal.MetastaticMelanomatotheColon,Rectum,andAnus:A50 YearExperience[J].AnnSurgOncol,2018,25(8):2178 2183.[12]TessierDJ,McConnellEJ,Young FadokT,etal.Melanomametastatictothecolon:caseseriesandreviewoftheliteraturewithoutcomeanalysis[J].DisColonRectum,2003,46(4):441 447.[13]HarrisZ,DonovanMG,BrancoGM,etal.QuercetinasanEmergingAnti MelanomaAgent:AFour FocusAreaTherapeuticDevelopmentStrategy[J].FrontNutr,2016,3:48.[14]SchützB,KoppensteinerA,Sch rghoferD,etal.Generationofmetastaticmelanomaspecificantibodiesbyaffinitypurification[J].SciRep,2016,6:37253.[15]CaputyGG,DonohueJH,GoellnerJR,etal.Metastaticmelanomaofthegastrointestinaltract.Resultsofsurgicalmanagement.ArchSurg,1991,126(11):1353 1358.[16]SerinG,Doˇganav爧argilB,Cali爧kanC,etal.Colonicmalignantmelanoma,primaryormetastatic?Casereport[J].TurkJGastroenterol,2010,21(1):45 49.[17]秦文华.33例恶性黑色素瘤患者临床病理特点和BRAF基因突变分析[J].肿瘤基础与临床,2019,32(6):527 529[18]BenderGN,MaglinteDD,McLarneyJH,etal.Malignantmelanoma:patternsofmetastasistothesmallbowel,reliabilityofimagingstudies,andclinicalrelevance[J].AmJGastroenterol,2001,96(8):2392 2400.[19]CSCO黑色素瘤专家委员会.中国黑色素瘤诊治指南(2011版)[J].临床肿瘤学杂志,2012,17(2):159 171.[20]施春雨,张磊超,侯睿智,等.原发性乙状结肠恶性黑色素瘤1例报告[J].中国普通外科杂志,2015,24(10):1497 1500.[21]王锡山.肛管直肠恶性黑色素瘤诊治指南解读[J].中华结直肠疾病电子杂志,2015,4(2):15 17.[22]战雪林,张瑜娟,闵卫平.黑色素瘤治疗新进展[J].南昌大学学报(医学版),2018,58(5):90 96.(收稿日期:2020 05 04)(本文编辑:青海涛)·病例报告·自身免疫性胰腺炎的诊治:2例报告及文献复习龚帅,任泽强,张蓬波,吴耐,张冲,张易,张秀忠 【关键词】 自身免疫性胰腺炎;诊治;报告中图分类号:R571 文献标志码:A DOI:10.3969/j.issn.1672-2159.2020.07.037作者单位:221002徐州医科大学附属医院普外科通信作者:张秀忠,E mail:13776780677@163.com 自身免疫性性胰腺炎(autoimmunepancreatitis,AIP)是一种以自身免疫介导的特殊类型的慢性胰腺炎,有其特定的病理学、组织学以及临床特征[1-2]。
HIGHLIGHTS OF PRESCRIBING INFORMATIONThese highlights do not include all the information needed to use YASMIN safely and effectively. See full prescribing information for YASMIN.YASMIN (drospirenone/ethinyl estradiol) tablets, for oral useInitial U.S. Approval: 2001WARNING: CIGARETTE SMOKING AND SERIOUSCARDIOVASCULAR EVENTSSee full prescribing information for complete boxed warning• Women over 35 years old who smoke should not use Yasmin. (4) • Cigarette smoking increases the risk of serious cardiovascular events from combination oral contraceptive (COC) use. (4)----------------------------RECENT MAJOR CHANGES-------------------------Warnings and Precautions, Hyperkalemia (5.2) 5/2015----------------------------INDICATIONS AND USAGE---------------------------Yasmin is an estrogen/progestin COC indicated for use by women to prevent pregnancy. (1)---------------------DOSAGE AND ADMINISTRATION---------------------• Take one tablet daily by mouth at the same time every day. (2.1)• Tablets must be taken in the order directed on the blister pack. (2.1)---------------------DOSAGE FORMS AND STRENGTHS----------------------Yasmin consists of 28 film-coated, biconvex tablets in the following order (3): • 21 yellow tablets, each containing 3 mg drospirenone (DRSP) and0.03 mg ethinyl estradiol (EE)• 7 inert white tablets-------------------------------CONTRAINDICATIONS-----------------------------• Renal impairment (4)• Adrenal insufficiency (4)• A high risk of arterial or venous thrombotic diseases (4)• Undiagnosed abnormal uterine bleeding (4)• Breast cancer or other estrogen-or progestin-sensitive cancer (4)• Liver tumors or liver disease (4)• Pregnancy (4)-----------------------WARNINGS AND PRECAUTIONS-----------------------• Vascular risks: Stop Yasmin if a thrombotic event occurs. Stop at least 4 weeks before and through 2 weeks after major surgery. Start no earlierthan 4 weeks after delivery, in women who are not breastfeeding (5.1).COCs containing DRSP may be associated with a higher risk of venous thromboembolism (VTE) than COCs containing levonorgestrel or some other progestins. Before initiating Yasmin in a new COC user or awoman who is switching from a contraceptive that does not containDRSP, consider the risks and benefits of a DRSP-containing COC inlight of her risk of a VTE (5.1).•Hyperkalemia: DRSP has anti-mineralocorticoid activity. Do not use in patients predisposed to hyperkalemia. Check serum potassiumconcentration during the first treatment cycle in women on long-termtreatment with medications that may increase serum potassiumconcentration. (5.2, 7.1, 7.2)•Liver disease: Discontinue Yasmin if jaundice occurs. (5.4)•High blood pressure: Do not prescribe Yasmin for women with uncontrolled hypertension or hypertension with vascular disease. (5.5) •Carbohydrate and lipid metabolic effects: Monitor prediabetic and diabetic women taking Yasmin. Consider an alternate contraceptivemethod for women with uncontrolled dyslipidemia. (5.7) •Headache: Evaluate significant change in headaches and discontinue Yasmin if indicated. (5.8)•Uterine bleeding: Evaluate irregular bleeding or amenorrhea. (5.9)------------------------------ADVERSE REACTIONS------------------------------The most frequent adverse reactions (≥2%) are premenstrual syndrome (13.2%), headache /migraine (10.7%), breast pain/tenderness/discomfort (8.3%), nausea/vomiting (4.5%), abdominal pain/tenderness/discomfort(2.3%), mood changes (2.3%). (6.1)To report SUSPECTED ADVERSE REACTIONS, contact Bayer HealthCare Pharmaceuticals Inc. at 1-888-842-2937 or FDA at 1-800FDA-1088 or /medwatch------------------------------DRUG INTERACTIONS------------------------------Drugs or herbal products that induce certain enzymes (for example, CYP3A4) may decrease the effectiveness of COCs or increase breakthrough bleeding. Counsel patients to use a back-up or alternative method of contraception when enzyme inducers are used with COCs. (7.1)-----------------------USE IN SPECIFIC POPULATIONS-----------------------Nursing mothers: Not recommended; can decrease milk production. (8.3) See 17 for PATIENT COUNSELING INFORMATION and FDA-approved patient labeling.Revised: 5/2015FULL PRESCRIBING INFORMATION: CONTENTS*WARNING: CIGARETTE SMOKING AND SERIOUS CARDIOVASCULAR EVENTS1 INDICATIONS AND USAGE2 DOSAGE AND ADMINISTRATION2.1 How to Take Yasmin2.2 How to Start Yasmin2.3 Advice in Case of Gastrointestinal Disturbances3 DOSAGE FORMS AND STRENGTHS4 CONTRAINDICATIONS5 WARNINGS AND PRECAUTIONS5.1 Thromboembolic Disorders and Other Vascular Problems5.2 Hyperkalemia5.3 Carcinoma of the Breasts and Reproductive Organs5.4 Liver Disease5.5 High Blood Pressure5.6 Gallbladder Disease5.7 Carbohydrate and Lipid Metabolic Effects5.8 Headache5.9 Bleeding Irregularities5.10 COC Use Before or During Early Pregnancy5.11 Depression5.12 Interference with Laboratory Tests5.13 Monitoring5.14 Other Conditions6 ADVERSE REACTIONS6.1 Clinical Trials Experience6.2 Postmarketing Experience7 DRUG INTERACTIONS7.1 Effects of Other Drugs on Combined Oral Contraceptives7.2 Effects of Combined Oral Contraceptives on Other Drugs7.3 Interference with Laboratory Tests8 USE IN SPECIFIC POPULATIONS8.1 Pregnancy8.3 Nursing Mothers8.4 Pediatric Use8.5 Geriatric Use8.6 Patients with Renal Impairment8.7 Patients with Hepatic Impairment8.8 Race10 OVERDOSAGE11 DESCRIPTION12 CLINICAL PHARMACOLOGY12.1 Mechanism of Action12.2 Pharmacodynamics12.3 Pharmacokinetics13 NONCLINICAL TOXICOLOGY13.1 Carcinogenesis, Mutagenesis, Impairment of Fertility14 CLINICAL STUDIES15 REFERENCES16 HOW SUPPLIED/STORAGE AND HANDLING16.1 How Supplied16.2 Storage17 PATIENT COUNSELING INFORMATION*Sections or subsections omitted from the full prescribing information are not listed__________________________________________________________________________________________________ FULL PRESCRIBING INFORMATION1 INDICATIONS AND USAGEYasmin® is indicated for use by women to prevent pregnancy.2 DOSAGE AND ADMINISTRATION2.1 How to Take YasminTake one tablet by mouth at the same time every day. The failure rate may increase when pills are missed or taken incorrectly.To achieve maximum contraceptive effectiveness, Yasmin must be taken as directed, in the order directed on the blister pack. Single missed pills should be taken as soon as remembered.2.2 How to Start YasminInstruct the patient to begin taking Yasmin either on the first day of her menstrual period (Day 1 Start) or on the first Sunday after the onset of her menstrual period (Sunday Start).Day 1 StartDuring the first cycle of Yasmin use, instruct the patient to take one yellow Yasmin daily, beginning on Day 1 of her menstrual cycle. (The first day of menstruation is Day 1.) She should take one yellow Yasmin daily for 21 consecutive days, followed by one white tablet daily on Days 22 through 28. Yasmin should be taken in the order directed on the package at the same time each day, preferably after the evening meal or at bedtime with some liquid, as needed. Yasmin can be taken without regard to meals. If Yasmin is first taken later than the first day of the menstrual cycle, Yasmin should not be considered effective as a contraceptive until after the first 7 consecutive days of product administration. Instruct the patient to use a non-hormonal contraceptive as back-up during the first 7 days. The possibility of ovulation and conception prior to initiation of medication should be considered.Sunday StartDuring the first cycle of Yasmin use, instruct the patient to take one yellow Yasmin daily, beginning on the first Sunday after the onset of her menstrual period. She should take one yellow Yasmin daily for 21 consecutive days, followed by one white tablet daily on Days 22 through 28. Yasmin should be taken in the order directed on the package at the same time each day, preferably after the evening meal or at bedtime with some liquid, as needed. Yasmin can be taken without regard to meals. Yasmin should not be considered effective as a contraceptive until after the first 7 consecutive days of product administration. Instruct the patient to use a non-hormonal contraceptive as back-up during the first 7 days. The possibility of ovulation and conception prior to initiation of medication should be considered.The patient should begin her next and all subsequent 28-day regimens of Yasmin on the same day of the week that she began her first regimen, following the same schedule. She should begin taking her yellow tablets on the next day after ingestion of the last white tablet, regardless of whether or not a menstrual period has occurred or is still in progress. Anytime a subsequent cycle of Yasmin is started later than the day following administration of the last white tablet, the patient should use another method of contraception until she has taken a yellow Yasmin daily for seven consecutive days. When switching from a different birth control pillWhen switching from another birth control pill, Yasmin should be started on the same day that a new pack of the previous oral contraceptive would have been started.When switching from a method other than a birth control pillWhen switching from a transdermal patch or vaginal ring, Yasmin should be started when the next application would have been due. When switching from an injection, Yasmin should be started when the next dose would have been due. When switching from an intrauterine contraceptive or an implant, Yasmin should be started on the day of removal. Withdrawal bleeding usually occurs within 3 days following the last yellow tablet. If spotting or breakthrough bleeding occurs while taking Yasmin, instruct the patient to continue taking Yasmin by the regimen described above. Counsel her that this type of bleeding is usually transient and without significance; however, advise her that if the bleeding is persistent or prolonged, she should consult her healthcare provider.Although the occurrence of pregnancy is low if Yasmin is taken according to directions, if withdrawal bleeding does not occur, consider the possibility of pregnancy. If the patient has not adhered to the prescribed dosing schedule (missed one or more active tablets or started taking them on a day later than she should have), consider the possibility of pregnancy at the time of the first missed period and take appropriate diagnostic measures. If the patient has adhered to the prescribed regimen and misses two consecutive periods, rule out pregnancy. Discontinue Yasmin if pregnancy is confirmed.The risk of pregnancy increases with each active yellow tablet missed. For additional patient instructions regarding missed pills, see the “WHAT TO DO IF YOU MISS PILLS” section in the FDA-Approved Patient Labeling. If breakthrough bleeding occurs following missed tablets, it will usually be transient and of no consequence. If the patient misses one or more white tablets, she should still be protected against pregnancy provided she begins taking a new cycle of yellow tablets on the proper day.For postpartum women who do not breastfeed or after a second trimester abortion, start Yasmin no earlier than 4 weeks postpartum due to the increased risk of thromboembolism. If the patient starts Yasmin postpartum and has not yet had a period, evaluate for possible pregnancy, and instruct her to use an additional method of contraception until she has taken Yasmin for 7 consecutive days.2.3 Advice in Case of Gastrointestinal DisturbancesIn case of severe vomiting or diarrhea, absorption may not be complete and additional contraceptive measures should be taken. If vomiting occurs within 3-4 hours after tablet-taking, this can be regarded as a missed tablet.3 DOSAGE FORMS AND STRENGTHSYasmin (drospirenone/ethinyl estradiol) tablets are available in blister packs.Each blister pack contains 28 film-coated, round, bi-convex tablets in the following order:• 21 yellow tablets each containing 3 mg drospirenone (DRSP) and 0.03 mg ethinyl estradiol (EE) embossed with a “DO” in a regular hexagon on one side• 7 inert white tablets embossed with a “DP” in a regular hexagon on one side4 CONTRAINDICATIONSDo not prescribe Yasmin to women who are known to have the following:• Renal impairment• Adrenal insufficiency• A high risk of arterial or venous thrombotic diseases. Examples include women who are known to: o Smoke, if over age 35 [see Boxed Warning and Warnings and Precautions (5.1)]o Have deep vein thrombosis or pulmonary embolism, now or in the past [see Warnings and Precautions (5.1)] o Have cerebrovascular disease [see Warnings and Precautions (5.1)]o Have coronary artery disease [see Warnings and Precautions (5.1)]o Have thrombogenic valvular or thrombogenic rhythm diseases of the heart (for example, subacute bacterial endocarditis with valvular disease, or atrial fibrillation) [see Warnings and Precautions (5.1)] o Have inherited or acquired hypercoagulopathies [see Warnings and Precautions (5.1)]o Have uncontrolled hypertension [see Warnings and Precautions (5.5)]o Have diabetes mellitus with vascular disease [see Warnings and Precautions (5.7)]o Have headaches with focal neurological symptoms or have migraine headaches with or without aura if over age 35 [see Warnings and Precautions (5.8)]• Undiagnosed abnormal uterine bleeding [see Warnings and Precautions (5.9)]• Breast cancer or other estrogen-or progestin-sensitive cancer, now or in the past [see Warnings and Precautions(5.3)]• Liver tumor (benign or malignant) or liver disease [see Warnings and Precautions (5.4) and Use in Specific Populations (8.7)]• Pregnancy, because there is no reason to use COCs during pregnancy [see Warnings and Precautions (5.10) and Use in Specific Populations (8.1)]5 WARNINGS AND PRECAUTIONS5.1 Thromboembolic Disorders and Other Vascular ProblemsStop Yasmin if an arterial or venous thrombotic (VTE) event occurs.Based on presently available information on Yasmin, DRSP-containing COCs may be associated with a higher risk of venous thromboembolism (VTE) than COCs containing the progestin levonorgestrel or some other progestins. Epidemiologic studies that compared the risk of VTE reported that the risk ranged from no increase to a three-fold increase. Before initiating use of Yasmin in a new COC user or a woman who is switching from a contraceptive that does not contain DRSP, consider the risks and benefits of a DRSP-containing COC in light of her risk of a VTE. Known risk factors for VTE include smoking, obesity, and family history of VTE, in addition to other factors that contraindicate use of COCs [see Contraindications (4)].A number of studies have compared the risk of VTE for users of Yasmin to the risk for users of other COCs, including COCs containing levonorgestrel. Those that were required or sponsored by regulatory agencies are summarized in Table 1.Table 1: Estimates (Hazard Ratios) of Venous Thromboembolism Risk in Current Users of Yasmin Compared to Users of Oral Contraceptives that Contain Other Progestins Epidemiologic Study Comparator Product Hazard Ratio (HR) (Author, Year of (all are low-dose COCs; with (95% CI) Publication) ≤0.04 mg of EE)Population Studiedi3 Ingenix All COCs available in the US HR: 0.9 (Seeger 2007)Initiators, including newusers aduring the conduct of the study b (0.5-1.6)All COCs available in Europe HR: 0.9 EURAS during the conduct of the study c (0.6-1.4) (Dinger 2007)Initiators, including new users a Levonorgestrel/EE HR: 1.0(0.6-1.8)“FDA-funded study” (2011)New users aAll users(i.e., initiation and continuing use of study combination hormonal contraception) Other COCs available during thecourse of the study dLevonorgestrel/0.03 mg EEOther COCs available during thecourse of the study dLevonorgestrel/0.03 mg EEHR: 1.8(1.3-2.4)HR: 1.6(1.1-2.2)HR: 1.7(1.4-2.1)HR: 1.5(1.2-1.8)a) “New users” -no use of combination hormonal contraception for at least the prior 6 monthsb) Includes low-dose COCs containing the following progestins: norgestimate, norethindrone, levonorgestrel,desogestrel, norgestrel, medroxyprogesterone, or ethynodiol diacetatec) Includes low-dose COCs containing the following progestins: levonorgestrel, desogestrel, dienogest, chlormadinoneacetate, gestodene, cyproterone acetate, norgestimate, or norethindroned) Includes low-dose COCs containing the following progestins: norgestimate, norethindrone, or levonorgestrelIn addition to these “regulatory studies,” other studies of various designs have been conducted. Overall, there are two prospective cohort studies (see Table 1): the US post-approval safety study Ingenix [Seeger 2007], the European post-approval safety study EURAS (European Active Surveillance Study) [Dinger 2007]. An extension of the EURAS study, the Long-Term Active Surveillance Study (LASS), did not enroll additional subjects, but continued to assess VTE risk. There are three retrospective cohort studies: one study in the US funded by the FDA (see Table 1), and two from Denmark [Lidegaard 2009, Lidegaard 2011]. There are two case-control studies: the Dutch MEGA study analysis [van Hylckama Vlieg 2009] and the German case-control study [Dinger 2010]. There are two nested case-control studies that evaluated the risk of non-fatal idiopathic VTE: the PharMetrics study [Jick 2011] and the GPRD study [Parkin 2011]. The results of all of these studies are presented in Figure 1.Figure 1: VTE Risk with Yasmin Relative to LNG-Containing COCs (adjusted risk#)Risk ratios displayed on logarithmic scale; risk ratio < 1 indicates a lower risk of VTE for DRSP, > 1 indicates an increased risk of VTE for DRSP.*Comparator “Other COCs”, including LNG-containing COCs† LASS is an extension of the EURAS study#Some adjustment factors are indicated by superscript letters: a) Current heavy smoking, b) hypertension, c) obesity, d) family history, e) age, f) BMI, g) duration of use, h) VTE history, i) period of inclusion, j) calendar year, k) education, l) length of use, m) parity, n) chronic disease, o) concomitant medication, p) smoking, q) duration of exposure, r) site(References: Ingenix [Seeger 2007]1, EURAS (European Active Surveillance Study) [Dinger 2007]2, LASS (Long-Term Active Surveillance Study) [Dinger, unpublished document on file], FDA-funded study [Sidney 2011]3, Danish [Lidegaard 2009]4, Danish reanalysis [ Lidegaard 2011]5, MEGA study [van Hylckama Vlieg 2009]6, German Case-Control study [Dinger 2010]7, PharMetrics [Jick 2011]8, GPRD study [Parkin 2011]9)Although the absolute VTE rates are increased for users of hormonal contraceptives compared to non-users, the rates during pregnancy are even greater, especially during the post-partum period (see Figure 2). The risk of VTE in women using COCs has been estimated to be 3 to 9 per 10,000 woman-years. The risk of VTE is highest during the first year of use. Data from a large, prospective cohort safety study of various COCs suggest that this increased risk, as compared to that in non-COC users, is greatest during the first 6 months of COC use. Data from this safety study indicate that the greatest risk of VTE is present after initially starting a COC or restarting (following a 4 week or greater pill-free interval) the same or a different COC.The risk of thromboembolic disease due to oral contraceptives gradually disappears after COC use is discontinued. Figure 2 shows the risk of developing a VTE for women who are not pregnant and do not use oral contraceptives, for women who use oral contraceptives, for pregnant women, and for women in the postpartum period. To put the risk of developing a VTE into perspective: If 10,000 women who are not pregnant and do not use oral contraceptives are followed for one year, between 1 and 5 of these women will develop a VTE.Figure 2 Likelihood of Developing a VTEIf feasible, stop Yasmin at least 4 weeks before and through 2 weeks after major surgery or other surgeries known to have an elevated risk of thromboembolism.Start Yasmin no earlier than 4 weeks after delivery, in women who are not breastfeeding. The risk of postpartum thromboembolism decreases after the third postpartum week, whereas the risk of ovulation increases after the third postpartum week.Use of COCs also increases the risk of arterial thromboses such as strokes and myocardial infarctions, especially in women with other risk factors for these events.COCs have been shown to increase both the relative and attributable risks of cerebrovascular events (thrombotic and hemorrhagic strokes), although, in general, the risk is greatest among older (>35 years of age), hypertensive women who also smoke. COCs also increase the risk for stroke in women with other underlying risk factors.Oral contraceptives must be used with caution in women with cardiovascular disease risk factors.Stop Yasmin if there is unexplained loss of vision, proptosis, diplopia, papilledema, or retinal vascular lesions. Evaluate for retinal vein thrombosis immediately. [See Adverse Reactions (6).]5.2 HyperkalemiaYasmin contains 3 mg of the progestin DRSP, which has anti-mineralocorticoid activity, including the potential for hyperkalemia in high-risk patients, comparable to a 25 mg dose of spironolactone. Yasmin is contraindicated in patients with conditions that predispose to hyperkalemia (that is, renal impairment, hepatic impairment, and adrenal insufficiency). Women receiving daily, long-term treatment for chronic conditions or diseases with medications that may increase serum potassium concentration should have their serum potassium concentration checked during the first treatment cycle. Medications that may increase serum potassium concentration include ACE inhibitors, angiotensin–II receptor antagonists, potassium-sparing diuretics, potassium supplementation, heparin, aldosterone antagonists, and NSAIDs. Consider monitoring serum potassium concentration in high-risk patients who take a strong CYP3A4 inhibitor long-term and concomitantly. Strong CYP3A4 inhibitors include azole antifungals (e.g. ketoconazole, itraconazole, voriconazole), HIV/HCV protease inhibitors (e.g., indinavir, boceprevir), and clarithromycin [see Clinical Pharmacology (12.3)].5.3 Carcinoma of the Breasts and Reproductive OrgansWomen who currently have or have had breast cancer should not use Yasmin because breast cancer is a hormonallysensitive tumor.There is substantial evidence that COCs do not increase the incidence of breast cancer. Although some past studies have suggested that COCs might increase the incidence of breast cancer, more recent studies have not confirmed such findings. Some studies suggest that COCs are associated with an increase in the risk of cervical cancer or intraepithelial neoplasia. However, there is controversy about the extent to which these findings may be due to differences in sexual behavior and other factors.5.4 Liver DiseaseDiscontinue Yasmin if jaundice develops. Steroid hormones may be poorly metabolized in patients with impaired liver function. Acute or chronic disturbances of liver function may necessitate the discontinuation of COC use until markers of liver function return to normal and COC causation has been excluded.Hepatic adenomas are associated with COC use. An estimate of the attributable risk is 3.3 cases/100,000 COC users. Rupture of hepatic adenomas may cause death through intra-abdominal hemorrhage.Studies have shown an increased risk of developing hepatocellular carcinoma in long-term (> 8 years) COC users. However, the attributable risk of liver cancers in COC users is less than one case per million users.Oral contraceptive-related cholestasis may occur in women with a history of pregnancy-related cholestasis. Women with a history of COC-related cholestasis may have the condition recur with subsequent COC use.5.5 High Blood PressureFor women with well-controlled hypertension, monitor blood pressure and stop Yasmin if blood pressure rises significantly. Women with uncontrolled hypertension or hypertension with vascular disease should not use COCs.An increase in blood pressure has been reported in women taking COCs, and this increase is more likely in older women and with extended duration of use. The incidence of hypertension increases with increasing concentration of progestin. 5.6 Gallbladder DiseaseStudies suggest a small increased relative risk of developing gallbladder disease among COC users.5.7 Carbohydrate and Lipid Metabolic EffectsCarefully monitor prediabetic and diabetic women who are taking Yasmin. COCs may decrease glucose tolerance in a dose-related fashion.Consider alternative contraception for women with uncontrolled dyslipidemia. A small proportion of women will have adverse lipid changes while on COCs.Women with hypertriglyceridemia, or a family history thereof, may be at an increased risk of pancreatitis when using COCs.5.8 HeadacheIf a woman taking Yasmin develops new headaches that are recurrent, persistent, or severe, evaluate the cause and discontinue Yasmin if indicated.An increase in frequency or severity of migraine during COC use (which may be prodromal of a cerebrovascular event) may be a reason for immediate discontinuation of the COC.5.9 Bleeding IrregularitiesUnscheduled (breakthrough or intracyclic) bleeding and spotting sometimes occur in patients on COCs, especially during the first three months of use. If bleeding persists or occurs after previously regular cycles, check for causes such as pregnancy or malignancy. If pathology and pregnancy are excluded, bleeding irregularities may resolve over time or with a change to a different COC.Data from ten contraceptive efficacy clinical trials (N=2,467) show that the percent of women who took Yasmin and experienced unscheduled bleeding decreased over time from 12% at cycle 2 to 6% (cycle 13). A total of 24 subjects out of 2,837 in the Yasmin trials (<1%) discontinued due to bleeding complaints. These are described as metrorrhagia, vaginal hemorrhage, menorrhagia, abnormal withdrawal bleeding, and menometrorrhagia.The average duration of scheduled bleeding episodes in the majority of subjects (86%-88%) was 4-7 days. Women who use Yasmin may experience absence of withdrawal bleeding, even if they are not pregnant. Based on subject diaries fromcontraceptive efficacy trials, during cycles 2–13, 1-11% of women per cycle experienced no withdrawal bleeding. Some women may encounter post-pill amenorrhea or oligomenorrhea, especially when such a condition was pre-existent.If withdrawal bleeding does not occur, consider the possibility of pregnancy. If the patient has not adhered to the prescribed dosing schedule (missed one or more active tablets or started taking them on a day later than she should have), consider the possibility of pregnancy at the time of the first missed period and take appropriate diagnostic measures. If the patient has adhered to the prescribed regimen and misses two consecutive periods, rule out pregnancy.5.10 COC Use Before or During Early PregnancyExtensive epidemiological studies have revealed no increased risk of birth defects in women who have used oral contraceptives prior to pregnancy. Studies also do not suggest a teratogenic effect when COCs are taken inadvertently during early pregnancy, particularly in so far as cardiac anomalies and limb-reduction defects are concerned.The administration of oral contraceptives to induce withdrawal bleeding should not be used as a test for pregnancy [see Use in Specific Populations (8.1)].5.11 DepressionWomen with a history of depression should be carefully observed and Yasmin discontinued if depression recurs to a serious degree.5.12 Interference with Laboratory TestsThe use of COCs may change the results of some laboratory tests, such as coagulation factors, lipids, glucose tolerance, and binding proteins. Women on thyroid hormone replacement therapy may need increased doses of thyroid hormone because serum concentrations of thyroid-binding globulin increase with use of COCs [see Drug Interactions (7.2)].DRSP causes an increase in plasma renin activity and plasma aldosterone induced by its mild anti-mineralocorticoid activity.5.13 MonitoringA woman who is taking COCs should have a yearly visit with her healthcare provider for a blood pressure check and for other indicated healthcare.5.14 Other ConditionsIn women with hereditary angioedema, exogenous estrogens may induce or exacerbate symptoms of angioedema. Chloasma may occasionally occur, especially in women with a history of chloasma gravidarum. Women with a tendency to chloasma should avoid exposure to the sun or ultraviolet radiation while taking COCs.6 ADVERSE REACTIONSThe following serious adverse reactions with the use of COCs are discussed elsewhere in the labeling:•Serious cardiovascular events and stroke [see Boxed Warning and Warnings and Precautions (5.1)]•Vascular events [see Warnings and Precautions (5.1)]•Liver disease [see Warnings and Precautions (5.4)]Adverse reactions commonly reported by COC users are:•Irregular uterine bleeding•Nausea•Breast tenderness•Headache6.1 Clinical Trials ExperienceBecause clinical trials are conducted under widely varying conditions, the adverse reaction rates observed cannot be directly compared to rates in other clinical trials and may not reflect the rates observed in practice.。
详解美国保健品的各种功效,不要乱吃!美国的保健品市场是全世界最大,也是最先进的市场,国内来美国的人越来越多,回国带物,保健品也是其中一个重要的采购方面。
也有些保健品是华人家庭的必备品,但是具体的功效你都了解吗?买保健品也需要对症下药,吃前一定了解保健品的功效是否适合你,不要乱吃。
【Canker-Rid 口腔溃疡/炎症/疼痛速效康】可迅速起效,缓解口腔溃疡、唇疱疹、热病疱疹等口腔炎症和疼痛,同时加速口腔炎症康复。
使用时先用纸巾擦干患处,用棉签点涂在患处即可,即刻见效,94个满分好评!【Nature Made 三倍强效维骨力150粒】适用于治疗和预防全身所有部位的骨关节炎,联合葡萄糖胺、软骨素和MSM三种有效成分,可有效减轻关节疼痛、肿胀,增强关节灵活性,缓解运动损伤,预防骨质疏松。
送老人,骨关节炎、关节软骨伤和骨折患者都不错。
服用方法:每日两次,每次一粒,随餐服用。
或者每日一次,每次2粒。
【Genesis Today 液体超浓缩新西兰胶原蛋白】胶原不仅和美丽有关,人在35岁后胶原分泌量大幅度减少导致关节问题比较多。
大幅度摄入胶原不仅可以营养肌肤,而且对骨关节健康大有好处,但摄入安全高浓缩是关键。
Genesis T oday的胶原采用新西兰小牛胶原,液体更易吸收。
【新章超临界萃取肉桂精华胶囊 New Chapter Cinnamonforce 120粒】新章超临界萃取肉桂精华,比较适合糖尿病患者作为平时日常辅助的保健品服用,能有效帮助改善睡眠,改善身体的血糖指标。
【Herbs Etc. Deep Sleep 深度睡眠补剂120粒】美国制造,共120胶囊,睡觉前20分钟服用一粒,能够有效促进睡眠质量,恢复精神。
液体胶囊,快速起效。
帮助您快速入睡,带到深度睡眠效果,恢复精力。
以新鲜草药为原料,不会形成依赖性。
醒来后不打瞌睡。
【Carlson Labs 卡尔森 Co-Q-10 辅酶 50mg 60片】因为 CoQ10能够显著的保护心脏,所以在许多国家被称为心脏的发动机。
姜黄素多酚类似物对酪氨酸酶抑制活性的研究涂增清;杜志云;张焜;潘文龙;汤志恺;莫容清【期刊名称】《日用化学工业》【年(卷),期】2011(41)1【摘要】酪氨酸酶抑制剂广泛应用于皮肤美白化妆品中.通过芳香醛分别与丙酮和环戊酮在酸性条件下催化缩合,合成了两个系列共10个姜黄素多酚类似物(A1-5和B1-5),并研究了其对酪氨酸酶的抑制活性.结果表明,含邻二酚羟基的姜黄素多酚类似物(A4和B4)对酪氰酸酶具有很强的抑制作用,半抑制浓度(IC50)分别为1.19,1.24 μmol/L,比曲酸(IC50为28.59μmoL/L)的活性强20多倍.抑制动力学研究表明,该类姜黄素多酚类似物对酪氨酸酶的抑制属于非竞争性抑制类型.%Tyrosinase inhibitors are widely used in skin whitening cosmetics.Two series covering ten compounds of curcumin polyphenolic analogs (A1~5 and B1~5 ) were synthesized by acidic catalytic condensation of an appropriate aromatic aldehyde with acetone and cyclopentanone respectively.Their tyrosinase inhibition activity was evaluated, and the results indicated that compound A4 and B4 that containing two hydroxy groups of ortho orientation similar to catechol exhibit very strong inhibitory effects on tyrosinase with IC50 values of 1.19,1.24 μmoL/L respectively, which are more than twenty times higher than that of Kojic Acid (IC50 = 28.59 μmoL/L).Inhibition kinetic study showed that the tyrosinase inhibition of this kind of curcumin polyphenolic analogs belong to non -competitive type.【总页数】5页(P27-31)【作者】涂增清;杜志云;张焜;潘文龙;汤志恺;莫容清【作者单位】广东工业大学,轻工化工学院,广东,广州,510006;广东工业大学,轻工化工学院,广东,广州,510006;广东工业大学,轻工化工学院,广东,广州,510006;中国广州分析测试研究中心,广东,广州,510070;广东工业大学,轻工化工学院,广东,广州,510006;广东工业大学,轻工化工学院,广东,广州,510006【正文语种】中文【中图分类】TQ658【相关文献】1.含氟姜黄素类似物的合成及其抑制酪氨酸酶活性研究 [J], 韦星船;郑成;何浏;罗颖;赵文忠;林学镁2.含氮姜黄素类似物的合成及其酪氨酸酶抑制效应 [J], 韦星船;霍梦月;郑成;段彦飞;杨前程;蔡伟平3.新型碳糖苷类熊果苷类似物的合成及其对酪氨酸酶的抑制活性 [J], 朱晨江;石焱;吕遐;唐燕辉;陈国荣4.白及须根多酚提取工艺优化及其抗氧化、酪氨酸酶抑制活性研究 [J], 陈向阳;宋武;檀小菲;王婷婷;耿玉闯;王卫东;康清俊;吴永祥5.木瓜皮多酚和黄酮提取工艺优化及酪氨酸酶与胰脂肪酶抑制活性研究 [J], 裴文清;吕泸楠;王靖宇;浦思琦;雷霜;王春丽因版权原因,仅展示原文概要,查看原文内容请购买。
2010,Vol.34,N o.4 187P rogress in Phar m aceutical Sc iences信息广角2010年第34卷 第4期 第187页(orlista,t即罗氏公司的处方药产品Xen i c al和葛兰素史克公司的非处方药产品A lli)致肝损伤的不良反应报告进行调查研究。
另外,刚刚递交了上市申请的更多减肥新药是否会因安全性问题而被淘汰,人们正拭目以待。
在美国,2009年12月,V ivus和Arena两家制药公司分别向FDA递交了其减肥药产品Qnexa(芬特明+托吡酯)和lorcaseri n的上市申请,而O rex igen 公司计划于2010年递交其减肥复方产品Contrave (安非他酮+纳曲酮)的上市申请。
目前许多分析人士看好Qnexa,认为该复方产品是新一轮减肥药中的佼佼者,称其具有令人印象深刻!的疗效,其用于某些患者群,可致患者体重下降14 7%(即13 8kg),若获准上市,预计其会有可观的市场份额。
然而,也有人担忧:Qnexa可能产生与托吡酯相关的不良反应。
药物不良反应报告抗HIV药去羟肌苷致非肝硬化性门静脉高压[关键词] 去羟肌苷;核苷逆转录酶抑制剂;非肝硬化性门静脉高压[中图分类号] R978 7;R969 3美国FDA最近发出警告,抗H I V药去羟肌苷(didanosine,包括百时美施贵宝公司的产品V idex 和V i d ex EC及其仿制药)可增加用药者发生非肝硬化性门静脉高压的风险。
去羟肌苷是一种核苷逆转录酶抑制剂(NRTI),于2000年在美国上市用于治疗H I V感染,其产品标签上现已标有黑框警告∀∀∀本品可能导致胰腺炎、乳酸酸中毒以及肝肿大并伴脂肪变性。
但FDA认为,对于某些患者,该药的疗效仍超过其风险。
非肝硬化性门静脉高压是由于肝脏的主要静脉∀∀∀门静脉中血流减缓所致,从而引发胃肠道中食管静脉严重扩张,直至破裂和严重出血。
美国赛布瑞
【产品名称】
通用名称:美国赛布瑞
英文名称:Joint Health Original
【成分】
本品专利NEM®成分:II型胶原蛋白、葡糖氨基葡聚糖、透明质酸、钙等成分。
【适用症】
适用于类风湿关节炎,类风湿关节炎及其引发的关节疼痛、肿胀、晨僵、活动受限等
【用法用量】
饭前30分钟服用,每次1~2粒,温水送服
【不良反应】
纯天然生物复合胶囊,无激素化学添加成分,无不良症状
【特殊事项】
孕妇或哺乳期儿童需酌情食用,对鸡蛋过敏的不建议服用
【药物相互作用】
本品为纯天然提取物,可与其他药物同时服用
【有效期】
3年
【生产企业】
美国原瓶原装进口,美国REDD REMEDIES
【储存方法】
置于阴凉干燥处密封保存,避免阳光直射,并远离儿童
本品不含糖分、钠、酵母、麦粉、谷物等成分,可以每天使用。
melatonin翻译中文melatonin翻译中文:褪黑激素用法:褪黑激素是一种在治疗失眠、时差反应以及其他与生物钟有关的问题方面有用的药物。
它通常在晚上服用,并且应该在安静和黑暗的环境中服用。
用药量和治疗时间应根据个体需要和建议而定。
双语例句:1. Melatonin is commonly used to help regulate sleep patterns and treat insomnia.褪黑激素通常用于帮助调节睡眠模式和治疗失眠。
2. Taking melatonin in a quiet and dark environmentbefore bed can help improve sleep quality.在安静和黑暗的环境中睡前服用褪黑激素可以帮助改善睡眠质量。
3. Melatonin supplements are available over the counterin most drug stores and health food stores.多数药店和保健食品店都可以买到非处方褪黑激素补充剂。
4. Melatonin production is regulated by the body's natural circadian rhythms.褪黑激素的产生受到生物钟的自然节律的调节。
5. Some studies suggest that melatonin may help reduce symptoms of seasonal affective disorder.一些研究表明褪黑激素可能有助于减轻季节性情感障碍的症状。
6. Melatonin has been shown to have antioxidant properties and may help protect against cellular damage.褪黑激素被证明具有抗氧化性质,并可能有助于预防细胞损伤。
甘草中α-葡萄糖苷酶抑制剂的筛选(英文)
许有瑞;倪京满;孟庆刚;张承忠;高燕;王锐
【期刊名称】《中国药学:英文版》
【年(卷),期】2006(15)1
【摘要】目的从甘草中筛选α-葡萄糖苷酶抑制剂。
方法从甘草中分别提取纯化甘草酸、甘草次酸、甘草黄酮、生物碱和多糖,并进行活性比较。
然后以体外α-葡萄糖苷酶抑制试验为指导,从甘草黄酮中提取分离活性成分,并对其抑制动力学特性进
行研究。
结果甘草黄酮和甘草次酸具有较强的α-葡萄糖苷酶抑制活性,从甘草黄酮中分离鉴定了一活性成分—甘草酚,其表现为快速的非剂量依赖性的非竞争性抑制
类型,且IC50=0.26μg·mL-1,而甘草次酸却表现为快速的剂量依赖性的不可逆抑制类型,IC50=102.4μg·mL-1。
结论甘草酚可作为一种有效的α-葡萄糖苷酶抑制剂。
【总页数】4页(P24-27)
【关键词】α-葡萄糖苷酶抑制剂;甘草;甘草酚;甘草次酸
【作者】许有瑞;倪京满;孟庆刚;张承忠;高燕;王锐
【作者单位】兰州大学药学院;金川公司医院药剂科;兰州大学生命科学院
【正文语种】中文
【中图分类】R284.2;R285.5
【相关文献】
1.甘草酸提取废液α-葡萄糖苷酶抑制剂的筛选与鉴定 [J], 余颖;樊金玲;程源斌;朱
文学
2.用固定化酶筛选模型从天然产物中筛选α-葡萄糖苷酶抑制剂 [J], 卢大胜;吕敬慈;雍克岚;陈旭;冯偲慜;顾慧娟
3.一种用微孔板筛选alpha-葡萄糖苷酶抑制剂的方法(英文) [J], 李婷;张小东;宋聿文;刘建文
因版权原因,仅展示原文概要,查看原文内容请购买。
口服伊曲康唑治疗脂溢性皮炎的一项开放性非对照试验Kose O.;Erbil H.;Gur A.R.;崔荣【期刊名称】《世界核心医学期刊文摘:皮肤病学分册》【年(卷),期】2005(000)008【摘要】Background: Seborrhoeic dermatitis is an inflammatory cutaneous disorder in which the colonization of the affected area by Malassezia has been proved to play a key role. Objective: To perform a noncomparative open clinical study with oral itraconazole capsule (200 mg/day× 7 days) and consecutive usage 200 mg/day for the first 2 days of the following 2 months in patients with seborrhoeic dermatitis. Methods: Twenty- nine patients were enrolled to determine the efficacy and safety of oral itraconazole. The patients were evaluated according to itching, burning, erythema, desquamation and seborrhoea, each scored on a 0- 4 scaleon days15 (T15), 30 (T30), 60 (T 60) and 90 (T90). Itraconazole capsule 100 mg was given twice a day for 1 week and then, after a 3- week interval, patients used itraconazole capsule 200 mg/day for the first 2 days of the following 2 months. The clinical response was graded as markedly effective, effective, moderate or ineffective. Results: A clinical improvement (evaluated as markedly effective or effective) was observed in 23 patients (83% ) at T15, 21 (76% ) at T30, 20 (72% ) at T60 and 17 (61% ) at T90. At baseline, the mean ± .SD total clinical scores were 10.44 ± 2.45, 1.98 ± 0.5, 2.97 ± .1.12, 3.15 ± 1.74 and 3.30 ± 1.90 at T0, T15, T30, T60 and T90,respectively. Compared with baseline values, itraconazole capsule significantly reduced the mean ± SD total score as well as individ ual erythema and desquamation (Wilcoxon’ s signed test- twotailed)(P < 0.0001). No drug- related systemic adverse event was observed during the study. Conclusions: Seborrhoeic dermatitis shows marked reduction in inflammation when treated with itraconazole. The anti- inflammatory activity of oral itraconazole and efficacy on Malessezia suggests that itraconazole capsule will be first oral treatment option in future in severe seborrhoeic dermatitis.【总页数】2页(P27-28)【作者】Kose O.;Erbil H.;Gur A.R.;崔荣【作者单位】Queen Mary School of Medicine Cutaneous Research Centre 2 Newark Street Whitechapel London E1 2AT United Kingdom【正文语种】中文【中图分类】R758.732【相关文献】1.口服伊曲康唑治疗及预防中重度脂溢性皮炎复发作用的临床疗效分析 [J], 李宁;冯登超2.结直肠癌试验解读之瑞戈非尼单药治疗用于既往治疗过的转移性结直肠癌(CORRECT):一项国际、多中心、随机、安慰剂对照、Ⅲ期试验 [J], 戴广海;3.结直肠癌试验解读之瑞戈非尼单药治疗用于既往治疗过的转移性结直肠癌(CORRECT):一项国际、多中心、随机、安慰剂对照、Ⅲ期试验 [J], 戴广海4.氯吡格雷、依贝沙坦预防房颤患者血管事件的试验(ACTIVE W)中氯吡格雷联合阿司匹林与单用口服抗凝药治疗房颤的比较:一项随机对照试验 [J], 吴晓燕5.中重度脂溢性皮炎复发治疗和预防中伊曲康唑口服的临床价值分析 [J], 姚小卫因版权原因,仅展示原文概要,查看原文内容请购买。
Quercetin exerts anti-melanoma activities and inhibits STAT3signalingHui-Hui Cao,Anfernee Kai-Wing Tse,Hiu-Yee Kwan,Hua Yu,Chi-Yan Cheng,Tao Su,Wang-Fun Fong,Zhi-Ling Yu*School of Chinese Medicine,Hong Kong Baptist University,Hong Kong,China1.IntroductionMelanoma is a highly aggressive cancer that resists mostconventional treatment.Its incidence and mortality rates haveincreased,alarmingly,world-wide in the least30years[1].Despitea wide variety of therapies were used to treat melanoma,theprognosis for melanoma remains very poor,with a5-year survivalrate of less than5%[2,3].Most conventional chemotherapy drugshave failed in melanoma treatment because of the low responserates[4]and significant toxicities[5,6].Therefore,new effectiveand safe treatments for melanoma are urgently needed.Many signaling pathways are involved in melanoma develop-ment and progression,including STAT3signaling[7].STAT3(signaltransducers and activators of transcription3)is a member of afamily of seven proteins(STATs1,2,3,4,5a,5b,and6)that transmitsignals from the plasma membrane to the nucleus,where theymodulate the transcription of genes involved in the regulation of avariety of critical functions,including cell differentiation,prolifer-ation,survival,angiogenesis,metastasis,and immune responses[8,9].STAT3plays an important role in melanoma developmentand progression.The activation of STAT3promotes melanoma cellsgrowth and survival[10,11];persistent STAT3activity promotes invivo angiogenesis and stimulates invasion and metastasis[12,13].Blockade of STAT3signaling with dominant-negative STAT3variant Stat3b can suppress the growth of the murine melanomaB16tumor in vivo[11].Inhibition of STAT3activation using smallmolecule inhibitors can induce cell apoptosis and reduce humanmelanoma growth in nude mice[14–17].Thus,STAT3is animportant target in melanoma therapy and chemotherapy target-ing STAT3should be able to assault melanoma on multiple fronts.Quercetin(3,30,40,5,7-pentahydroxyflavone,Fig.1A)is a dietaryflavonoid with no carcinogenic activity in vivo[18]and lowintrinsic toxicity[19].It is abundant in many fruits and vegetablesas well as various kinds of Chinese medicinal herbs.It has multipleBiochemical Pharmacology87(2014)424–434A R T I C L E I N F OArticle history:Received26September2013Accepted15November2013Available online23November2013Keywords:MelanomaSTAT3ApoptosisInvasionMigrationA B S T R A C TMelanoma is highly resistant to chemotherapy,and the mortality rate is increasing rapidly worldwide.STAT3signaling has been implicated in the pathogenesis of melanoma and constitutive activated STAT3has been validated can as a target for melanoma therapy.Quercetin,a noncarcinogenic dietaryflavonoidwith low toxicity,has been shown to exert anti-melanoma activity.However,the anti-melanomamechanisms of quercetin are not fully understood.In this study,we sought to test the involvement ofSTAT3signaling in the inhibitory effects of quercetin on melanoma cell growth,migration and invasion.Our results showed that exposure to quercetin resulted in inhibition of proliferation of melanoma cells,induction of cell apoptosis,and suppression of migratory and invasive properties.Mechanistic studyindicated that quercetin inhibited the activation of STAT3signaling by interfering with STAT3phosphorylation,and reducing STAT3nuclear localization.This inhibited STAT3transcription activityand down-regulated STAT3targeted genes Mcl-1,MMP-2,MMP-9and VEGF,which are involved in cellgrowth,migration and invasion.Importantly,overexpression of constitutively active STAT3partiallyrescued the growth inhibiting effects induced by quercetin.Furthermore,quercetin suppressed A375tumor growth and STAT3activities in xenografted mice model,and inhibited murine B16F10cells lungmetastasis in an animal model.Overall,these results indicate that the antitumor activity of quercetin isat least partially due to inhibition of STAT3signaling in melanoma cells.Ourfindings provided newinsight into the action of quercetin potently inhibits the STAT3signaling pathway,suggesting it has apotential role in the prevention and treatment of melanoma.ß2013Elsevier Inc.All rights reserved.*Corresponding author.E-mail address:zlyu@.hk(Z.-L.Yu).Contents lists available at ScienceDirectBiochemical Pharmacologyj o u rn a l h om e p a g e:w w w.e l s e v i e r.c o m/l o c a t e/bi o c h e m p h a r m0006-2952/$–see front matterß2013Elsevier Inc.All rights reserved./10.1016/j.bcp.2013.11.008biological functions including anti-oxidant,anti-inflammatory,cardioprotective and anti-cancer activities [20–22].In melanoma,quercetin has been found to be able to inhibit murine melanoma cell B16-BL6metastasis in an animal model,and this has been correlated with decreased matrix metalloproteinase-9through the protein-kinase C (PKC)pathway [23–25].Quercetin also induces B16-BL6apoptosis in in vitro study,which has been attributed to target mitochondria,decreases Bcl-2expression,and inhibits PKC-a expression and PKC-d translocation [25,26].Quercetin was found to inhibit the cell proliferation of various human melanoma cells such as MNT1,M10,M14,C32and A375[27,28],and induce apoptosis in human melanoma Mel-Juso and G361cells [29].Moreover,quercetin has also been reported as a potent STAT3inhibitor in human glioblastoma and gastric cancer cells [30,31].In this study,A375and A2058,two human melanoma cell lines that STAT3activated [10,32]and B16F10,a murine melanoma cell line that widely used in lung metastasis study [33,34]were employed.We showed that quercetin potently inhibited STAT3signaling pathway in melanoma cells by interfering STAT3phosphorylation and reducing STAT3nuclear localization.We found that quercetin induced melanoma cell apoptosis,reduced cell proliferation,migration and invasion through down-regulation of STAT3signaling and its targeted genes expression,and quercetin also showed the inhibitory effects on tumor growth and metastasis in in vivo study.2.Materials and methods2.1.ReagentsAntibodies against phospho-AKT (Ser473),AKT,phospho-ERK (Thr202/Tyr204),ERK,phospho-STAT3(Tyr705),STAT3,phosphor-JAK2(Y1007/1008),JAK2,phosphor-Src (Tyr 416),Src,poly (ADP-ribose)polymerase (PARP),Mcl-1and Bcl-xL were obtained from cell signaling technology (Beverly,MA,USA).Antibodies against b -actin,VEGF and SP1were purchased from Santa Cruz Biotechnol-ogy (Santa Cruz,CA,USA).Anti-FLAG antibody was supplied by Sigma–Aldrich (St.Louis,MO,USA).Goat anti-rabbit IgG,goat anti-mouse IgG and protein marker were supplied by Bio-Rad (Hercules,CA,USA).Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI)apoptosis detection kit was purchased from BD Biosciences (San Jose,CA,USA).Other chemicals were obtained from Sigma–Aldrich (St.Louis,MO,USA).Quercetin was obtained from Chromadex (USA).The stock solution of 100mM quercetin was prepared in dimethyl sulfoxide (DMSO)and stored at À208C.2.2.Cell cultureHuman melanoma A375and A2058cells,and murine melano-ma B16F10cells,were obtained from American Type Culture Collection (ATCC,USA),and were maintained at 378C and 5%CO2Fig.1.Quercetin reduced viability and induced apoptosis in melanoma cells.(A)Chemical structure of quercetin.(B)The viability of A375and A2058cells after treatment with various concentrations of quercetin or vehicle control for 24h and 48h was measured by MTT assay.(C)The growth inhibition effect of quercetin on B16F10cells was measured by violet staining assay and photographs were showed as the 24h treatment (left)and 48h treatment (right).(D)A375and A2058cells were treated with various concentrations of quercetin or vehicle control for 48h.Apoptosis was analyzed by flow cytometry by using Annexin V/PI double staining.The percentage of apoptotic cells was presented as the mean ÆSD of three independent experiments.*P <0.05.(E)RARP cleavage in A375and A2058cells after treated with various concentrations of quercetin or vehicle control for 24h were detected by using Western blot analysis (left panel)and relative expression level (right panel)was analyzed by Image J software.Data were shown as mean ÆSD from three independent experiments,*P <0.05,**P <0.01.H.-H.Cao et al./Biochemical Pharmacology 87(2014)424–434425in a humidified atmosphere in high glucose Dulbecco’s modified Eagle’s medium(DMEM,GIBCO,USA),supplemented with10% fetal bovine serum(FBS,GIBCO,USA)and1%penicillin/strepto-mycin(P/S,GIBCO,USA).2.3.Cell viability assayThe cytotoxic effect of quercetin on A375and A2058cells were determined by MTT assay.Cells(5000/well)were seeded in96-well plates,treated with either vehicle control(DMSO)or various concentrations of quercetin.After incubation for24or 48h,30m l of MTT solution(5mg/ml)was added to each well and incubated for an additional3h.The formazan crystal formed was dissolved with100m l of DMSO;absorbance was detected at570nm by a microplate spectrophotometer(BD Biosciences,USA).The cytotoxic effect of quercetin on B16F10 was determined by crystal violet staining.Cells(2.5Â104)were seeded in60mm dishes,exposed to quercetin or vehicle control (DMSO)for24or48h,then cells werefixed with10%formalin for10min,followed by staining with0.05%crystal violet solution(CV)in distilled water for30min.Finally,CV was removed,the cells were washed twice with tap water and images were photographed.2.4.Apoptosis assayApoptotic effects in A375and A2058cells were evaluated by Annexin V/PI double staining with the Apoptosis Detection Kit (BD Biosciences,San Jose,CA,USA)according to the manufac-turer’s instructions.Briefly,cells were harvested after48h treatment and1Â105cells were then incubated in100m l labeling solution(5m l of AnnexinV-FITC,5m l of PI,10m l of10Âbinding buffer and80m l of H2O)in darkness at room temperature for15min,after that,400m l of1Âbinding buffer was added to stop the staining reaction.Flow cytometric analyses were performed on a FACSCalibur TM(BD Biosciences, USA)utilizing10,000events.2.5.In vitro cell migration assayThe wound healing assay was used to evaluate the ability of cell migration.When cells were grown to80–90%confluence in6-well plates.Cell monolayers were scratched with a sterile10m l pipette tip across the center of the well to generate a clean,straight wound area,and then were washed with PBS to remove detached cells from the plates.After that,cells were incubated with vehicle control or quercetin in serum-free DMEM medium,migration of cells into the wound area was photographed at the0and16h time points by microscope(Leica,Germany).2.6.In vitro cell invasion assayCell invasion was determined by using BD BioCoat TM Ma-trigel TM invasion chamber(24well plate,8-m m pore size) according to the manufacturer’s instruction.Briefly,0.75ml of 10%FBS culture medium was placed in the lower chamber,while 1.25Â105cells in500m l of DMEM-0.1%BSA containing quercetin or vehicle control were added to the upper chamber. Chambers were assembled and incubated for24h at378C. Afterwards,matrigel and the cells remaining in the upper chamber were removed by scrubbing with a cotton swab, the cells on the lower surface of the membrane,which were considered to have invaded,werefixed with4%parafor-maldehyde and stained with crystal violet.Cells infive microscope areas(at200Âmagnification)were counted and photographed.2.7.Preparation of cytoplasmic and nuclear fractionsQuercetin-treated and vehicle-treated cells were collected and suspended in hypotonic buffer[20mM Tris–HCl(pH7.9), 1.5mM MgCl2and10mM KCl].After incubation on ice for 15min,12m l10%(v/v)NP-40was added,and the combination was vortex for10s and kept on ice again for another10min, after centrifuged at14,000rpm at48C for1min,the super-natants,which were considered to be the cytoplasmic extracts, were transferred to fresh tubes.The pellets were washed once with100m l hypotonic buffers,and resuspended in high salt buffer[20mM Tris–HCl pH7.9,25%glycerol,0.42M NaCl, 1.5mM MgCl2and0.2mM EDTA].After30min incubation on ice,the lysates were centrifuged at14,000rpm at48C for 10min;the resultant supernatants were kept as nuclear extracts.2.8.Western blot analysisAfter treatment with different dosages of quercetin or vehicle control,cells were harvested and then lysed with RIPA buffer [50mM Tris–HCl,1%NP-40,0.35%sodium-deoxycholate, 150mM NaCl,1mM EDTA(pH7.4),1mM phenylmethylsulfonyl fluoride,1mM NaF,1mM Na3VO4and10m g/ml each of aprotinin,leupetin and pepstatin A]on ice for30min.Then the lysates were centrifuged at14,000rpm at48C for20min. After centrifuging,the supernatant were transferred into new tubes and stored atÀ808C.Protein concentrations were determined by using Bio-Rad protein assay reagent.40m g proteins mixed with5Âloading dye and2-mercaptoethanol were heated at958C for5min and then separated on8%,10%or 12%SDS-PAGEs along with protein markers.The proteins were then electro-transferred from the gel onto a nitrocellulose membrane,and left for about150min under350mA.The membranes were blocked with5%milk in TBST buffer(0.05% Tween-20/TBS)for1h at room temperature,and then probed with corresponding primary antibodies diluted in 2.5%milk-TBST solution overnight at48C.The membranes were washed with TBST solution3times(5min each)and then incubated with corresponding HRP-conjugated secondary antibodies for another1h.Immunoreactive bands were visualized using an ECL detection kit(Invitrogen,USA)following the manufacturer’s instruction.2.9.Real-time quantitative polymerase chain reaction analysisTotal RNA was extracted with Trizol reagent(Invitrogen,USA), and reverse-transcripted with oligo-dT using M-MLV reverse transcriptase(Promega,USA)according to the manufacturer’s protocol.The primers used were all synthesized by invitrogen.The sequences were as follows:human MMP-2:forward:50-GCTCA-GATCCGTGGTGAGAT-30;reverse:50-GGTGCTGGCTGAGTAGATCC-30;human MMP-9:forward:50-CAACATCACCTATTGGATCC-30; reverse:50-TGGGTGTAGAGTCTCTCCCT-30;internal control GAPDH:forward:50-CTGCACCACCAACTGCTTAGC-30;reverse:50-CTTCACCACCTTCTTGATGTC-30.Quantitative real-time PCR was performed using SYBR green reaction mixture in the ViiA7Real Time PCR System(Applied Biosystems,USA).The PCR cycling profile was as follows:one cycle at508C for2min and958C for10min,40cycles of958C for15s, 608C for1min.Thefluorescent signals were detected using the ViiA7Real Time PCR System.The gene expression data were normalized to the endogenous control GAPDH.The relative expression levels of genes were measured according to the formula2ÀD Ct,where D Ct is the difference in threshold cycle values between the targets and GAPDH.H.-H.Cao et al./Biochemical Pharmacology87(2014)424–434 4262.10.Gelatin zymographyActivities of matrix metalloproteinase(MMP-2)and MMP-9 were measured by gelatin zymography,an extremely sensitive technique that can detect picogram quantities.Cells were treated for24h with quercetin or vehicle control in serum-free DMEM medium,and then the supernatant was collected and concentrated using centricon YM-30concentrator(Millipore,USA).After the protein concentrations were determined,the samples were diluted in sample buffer(50mM,Tris–HCl(pH 6.8),2%SDS,0.1% bromophenol blue and10%glycerol),and applied to10%SDS-PAGE containing0.1%gelatin and then subjected to electrophore-sis.The gels were then washed with denaturing buffer(50mM Tris–HCl,pH7.5,2.5%Triton X-100)for30min to remove the SDS and incubated overnight in developing buffer(50mM Tris–HCl,pH 7.5,5mM CaCl2,200mM NaCl,0.2%Brij-35)in378C to allow the gelatinases to digest the gelatin structure.Gels were stained for3h in a mixture of30%methanol,10%acetic acid and0.5%Coomassie Brillant Blue and,subsequently,destained until bands became clear in30%methanol and10%acetic acid.The gelatinase activity was indicated by the clear bands on blue background,the width of the band is considered to be proportional to the enzyme activity.2.11.Plasmid transient transfectionThe constitutive activated STAT3expression construct Stat3-C Flag pRc/CMV was obtained from Addgene(USA).Transfection of STAT3plasmid into melanoma cells was conducted by using Lipofectamine2000(Invitrogen,USA)following manufacturer’s protocol.Cells were transfected with plasmids for48h before functional assays were carried out.2.12.Luciferase assayHuman melanoma A375and A2058cells were seeded in24-well plates and transiently transfected together with STAT3 reporter plasmid4ÂM67pTATA TK-Luc(0.2m g/per well, Addgene,USA)and Renilla luciferase-expressing plasmid PRL-CMV(0.1m g/per well,promega,USA)by use of lipofectamine2000 (Invitrogen,USA).48h later,cells were treated with various concentrations of quercetin for24h,and then lysed in100m l of passive lysis buffer(Promega,USA)in each well.A25m l aliquot of cell lysate was subjected to a luciferase assay by using the dual-luciferase assay kit(Promega,USA).STAT3luciferase activity was measured by EnVision Mutilabel Reader(Perkin Elmer,USA). Relative luciferase activity was calculated after the activity of STAT3-dependentfirefly luciferase had been normalized to that of Renilla luciferase.All values are expressed as–fold induction relative to basal activity.2.13.Nude mice xenograft modelMale nu/nu BALB/c mice(6weeks old)were obtained from the Laboratory Animal Services Centre,Chinese University of Hong Kong.The nude mice were maintained in individual ventilated cages in a specific animal handling room of Hong Kong Baptist University.All care and handling of animals were performed with the approval of the Government of The Hong Kong Special Administrative Region Department of Health.A375(5Â106)cells were resuspended in0.1ml PBS and inoculated subcutaneously into the backs of nude mice and allowed to grow for7days.After that,mice were randomly assigned to2groups(n=6for each group)and treated by intragastric administration with200m l0.5% CMC-Na solution as the vehicle control or100mg/kg quercetin daily for21consecutive days.Body weight and tumor volume were measured every3days.Tumor volumes were determined by a caliper and calculated according to the following formula: (width2Âlength)/2.At the end of the experiment,mice were sacrificed and tumor xenografts were collected and weighed. Tumor tissues were stored atÀ808C for Western blot analysis.2.14.In vivo model of lung metastasisTo produce experimental metastasis model,male C57BL/6mice (6weeks old)were used.B16F10cells(5Â105/0.1ml)were injected into the lateral tail vein of C57BL/6mice.Mice were randomly divided into2groups of12mice each.Each mouse was given a daily intragastric injection of either200m l0.5%CMC-Na solution as the vehicle control or100mg/kg quercetin.Treatment was started1day before cell injection and lasted for consecutive24 days.At the end of experiment,mice were sacrificed.Lungs were removed andfixed in4%formalin and black tumor cell colonies were counted.2.15.Statistical analysisAll data are expressed as meanÆstandard deviation.Statistical analysis was performed by the Student’s t-test,using the statistical software Graphpad Prism5.0.P<0.05was considered statistically significant.3.Results3.1.Quercetin reduced viability and induced apoptosis in melanoma cellsThe effect of quercetin on melanoma cell growth was investigated using MTT assay on human melanoma A375and A2058cell lines.The cell viability curves indicated that quercetin decreased the survival of both A375and A2058cells in a time-and dose-dependent manner(Fig.1B),with IC50values of99.59Æ10 and118.1Æ14.2m M after48h treatment,respectively.The crystal violet staining assay showed that after48h treatment by quercetin, the viability of B16F10decreased in a dose-dependent manner (Fig.1C).To determine whether quercetin induced apoptosis,Annexin V/ PI double staining assay was conducted.Phosphatidylserine(PS) translocated to the surface of the cellular membrane was considered as the hallmark of apoptosis in the early phase,which can be determined by PI/Annexin V double staining assay.Both A375and A2058cells were exposed to increasing concentrations of quercetin for48h,followed by PI/Annexin V double staining and flow cytometric measurement.It was found that quercetin induced apparent apoptosis in A375and A2058cells(Fig.1D).The rate of Annexin V positive cells was22.1Æ3.2%in quercetin-treated A375 cells versus6.35Æ0.25%in control cells,while the rate in A2058cells was24.2Æ3.4%versus6.35Æ0.85%in control cells.To further assess the pro-apoptotic effect of quercetin,we evaluated the cleavage of PARP in A375and A2058cells after24h treatment with quercetin. The active form of poly(ADP-ribose)polymerase(PARP),a protein associated with DNA repair,is considered as another hallmark of apoptosis.Western blot data showed that quercetin dose-depen-dently induced PARP cleavage in A375and A2058cells(Fig.1E), which provide further evidence that quercetin induced apoptosis. These results indicated that quercetin inhibited cell growth and induced apoptosis in melanoma cells.3.2.Quercetin impaired the migratory and invasive capacities of melanoma cellsMetastasis,which involves migration of tumor cells and invasion of normal tissues,is an important process for melanomaH.-H.Cao et al./Biochemical Pharmacology87(2014)424–434427progression.To test the effects of quercetin on melanoma cell migration and invasion,wound healing assay and transwell invasion chamber assay were performed,respectively.For the wound healing assay,a single scratch was created in the 80–90%confluent monolayer of cells by using a 10m l pipette tip.The scratches were photographed at 0h and 16h after treatment with indicated dosages of quercetin or vehicle control.Cells were found to migrate to the wound area after incubation for 16h,but quercetin at a concentration of 40m M strongly inhibited A375and A2058cell migration (Fig.2A).Similar effects were found with B16F10cells.Also,those B16F10cells treated with a higher dosage of quercetin showed a decrease in the wound closure rates compared to vehicle control (Fig.2B).For the invasion chamber assay,cells were treated with 40m M quercetin or vehicle control for 16h.After that,those cells that had invaded to the lower surface of the membrane were fixed,stained and photographed.Quercetin significantly reduced A375and A2058cell invaded to the lower surface of the chamber (Fig.2C).Under all the conditions,quercetin did not affect cell viability.Taken together,these results indicate that quercetin inhibits melanoma cells migration and invasion.3.3.Quercetin reduced constitutive STAT3phosphorylation in melanoma cellsIt has been well recognized that constitutive phosphorylation/activation of STAT3contributes to the development of human melanoma by favoring cancer cell growth,survival,migration and invasion [10,11,12].Therefore,we investigated whether quercetin modulates the constitutive STAT3phosphorylation/activation.It was found that the levels of phosphorylated STAT3at the tyrosine 705(Tyr705)site in both A375and A2058cells were reduced in a dose-dependent manner after 24h treatment and in a time-dependent manner after 60m M treatment by quercetin (Fig.3A).It has been reported that constitutive STAT3phosphorylation/activation is usually mediated by upstream tyrosine kinases such as JAK2and Src.Both JAK2and Src have been found to beconstitutively phosphorylated/activated and to contribute to the development of various cancers including melanoma [10,15,16,32].Next,we sought to find out if quercetin affects the upstream JAK2and Src activities of STAT3.The data showed that quercetin dose-and time-dependently suppressed the constitutive phosphorylation of JAK2(Y1007/1008)and Src (Tyr416)in both A375and A2058cells (Fig.3B).Moreover,quercetin was also found to be able to dose-dependently decrease constitutive phosphorylation of STAT3,JAK2and Src in murine melanoma B16F10cells (Fig.3C).It has been reported that the ERK and AKT pathways are two of the major constitutively activated signaling pathways in human melanoma [35],so we want to know if quercetin showed some impacts on these two pathways,in contrast to the inhibition of STAT3phosphorylation,we found that quercetin dose-dependent-ly activated AKT and ERK phosphorylation in human melanoma A375and A2058cells (Fig.3D).These findings strongly suggested that quercetin effectively and selectively suppresses the constitutive activation of STAT3in melanoma cells and this suppression can be attributed to the reduction of phosphorylation of JAK2and Src.3.4.Quercetin suppressed STAT3nuclear localization and STAT3-luciferase reporter activitySTAT3dimerization can be induced by tyrosine phosphoryla-tion at site tyrosine 705,which then leads to nuclear translocation.Hence we examined whether quercetin can inhibit the nuclear localization of STAT3.After quercetin treatment,the cytoplasmic and nuclear proteins of A375and A2058cells were extracted and expressions of STAT3in both fractions were determined by Western blotting.As shown in Fig.4A,the levels of STAT3in nuclear fractions were reduced significantly by quercetin in a dose-dependent manner,while those in cytoplasmic fractions were slightly increased.Next,we examined whether quercetin affected the STAT3reporter gene transcription.A375and A2058cells werecotrans-Fig.2.Quercetin impaired the migratory and invasive capacities of melanoma cells.(A)A375,A2058and (B)B16F10cells were plated in 6-well plates,and the next day,a single scratch was made in the confluent monolayer followed by vehicle control or quercetin treatment for 16h,each scratch was photographed after 16h treatment.(C)A375and A2058cells were allowed to migrate through matrigel coated membrane with 8m m pores for 24h in the presence of vehicle control or quercetin.Representative photographs of invasive cells (left)and quantification of invasive cells (right)were shown.Data were shown as mean ÆSD from three independent experiments,**P <0.01.H.-H.Cao et al./Biochemical Pharmacology 87(2014)424–434428fected with STAT3-TATA-Luc and Renilla -Luc for 48h and then treated with quercetin.Data showed that quercetin treatment caused an apparent reduction in STAT3reporter activity (Fig.4B).These data demonstrate clearly that quercetin is capable of inhibiting STAT3nuclear localization and STAT3luciferase activity in A375and A2058cells.3.5.Quercetin downregulated STAT3targeted genes expression levelsIn the nucleus,STAT3acts as a transcription factor;its target genes Mcl-1and Bcl-xL play important roles in melanoma cell growth and survival [36].Furthermore,vascular endothelial growth factor (VEGF),and matrix metalloproteinases (MMP)-2and MMP-9are essential for tumor invasion and metastasis and can be regulated by STAT3activation [12,13].We found that in both A375and A2058cells,quercetin dose-dependently decreased the expression of Mcl-1,while it did not affect the expression of Bcl-xL in both cell lines (Fig.5A).Quercetin also dose-dependently reduced MMP-2and MMP-9mRNA expression levels (Fig.5B),inhibited MMP-2and MMP-9enzymatic activities (Fig.5C),and suppressed VEGF expression (Fig.5D).3.6.Overexpression of STAT3partially prevented quercetin-induced cell growth inhibitionTo clarify whether quercetin-induced cell growth inhibition is mediated by STAT3inhibition in A375and A2058cells,we investigated whether STAT3overexpression can reduce growth inhibition caused by quercetin.A375and A2058cells were transiently transfected 48h with STAT3C,an oncogenic mutant form of Stat3that is constitutively activated without tyrosine phosphorylation [37],then the indicated concentrations of quercetin were added for 48h;cell viability was then determined by MTT assay.Western blotting showed that,compared to transient transfect with the empty vector –pcDNA3.0,transfection of STAT3-expressing construct in A375cell (A)and A2058cell (B)resulted in a remarkable increase in STAT3and p-STAT3levels.When exposed to different dosages of quercetin for 48h,there was no significant difference in cell viability between quercetin-treated parental cells and cells transfected with empty vector.For cells transfected with STAT3plasmid,under the 80m M quercetin condition,cell viability increased from 53.35Æ1.34%to 75.77Æ7.35%in A375cells and from 44.96Æ2.30%toFig.3.Quercetin selectively suppressed constitutive STAT3phosphorylation in melanoma cells.(A)A375and A2058cells were treated with various concentrations of quercetin for 24h or a fixed concentration (60m M)for various durations,and then total cell lysates were extracted for Western blot analysis by using antibodies specific to p-STAT3(Tyr705)and STAT3.The representative figures (left)and the relative expression levels analyzed by Image J software (right)were shown.(B)A375and A2058cells were treated with various concentrations of quercetin for 24h or a fixed concentration (60m M)for various durations,and then total cell lysates were extracted for Western blot analysis by using antibodies specific to p-Src (Tyr416),Src,p-JAK2(Y1007/1008)and JAK2.The representative figures (left)and the relative expression levels analyzed by Image J software (right)were shown (C)A375and A2058cells were exposed to indicated concentrations of quercetin for 24h and then total cell lysates were extracted for Western blot analysis by using antibodies specific to AKT,p-AKT,ERK,and p-ERK.The representative figures (upper)and the relative expression levels analyzed by Image J software (bottom)were shown.(D)Murine melanoma B16F10cells were treated with quercetin at various concentrations for 24h and then were lysed for Western blot analysis by using antibodies specific to STAT3,p-STAT3,JAK2,p-JAK2,Src and p-Src.The representative figures (upper)and the relative expression levels analyzed by Image J software (bottom)were shown.Data were shown as mean ÆSD from three independent experiments,*P <0.05,**P <0.01.H.-H.Cao et al./Biochemical Pharmacology 87(2014)424–434429。