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4 Interestingly, lack of TNF-α signal transduction reduced UCB-induced cell death for short periods of incubation, in contrast, inhibition of IL-1 β cascade produced a sustained blockade of astrocyte injury by UCB.
Astrocyte Reactivity to Unconjugated Bilirubin Requires TNF-α and IL-1β Receptor Signaling
Pathways
GLIA 59:14–25 (2011) SCI(2010):5.19
周康康 2012-7-2
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3 Cell Treatment :50μM UCB plus 100μM human serum albumin (HSA) (UCB to HSA molar ratio of 0.5), from15 min to 24 h, at 370C.
4 Western Blot :The protein expression of TNFR1, TNFR2, and IL-1R1were determined by Western blot analysis.
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5 Measurement of Cytokine Release :TNF-α, IL-1β, and IL-6 with specific DuoSetR ELISA Development kits.
6 Detection of NF-κB Activation — immunofluorescence detection :rabbit anti-p65 NF- κB subunit antibody (1:200) as the primary antibodies, a FITC-labeled goat anti-rabbit antibody (1:160) as the secondary antibodieastrocytes to UCB increased the expression of both TNF- α receptor TNFR1 and IL-1 β receptor IL-1R1, but not TNFR2, as well as their activation, observed by augmented binding of receptors’ molecular adaptors, TRAF2 and TRAF6, respectively.