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. Author manuscript; available in PMC: 2019 Apr 1.
Published in final edited form as: J Immunol. 2018 Feb 14;200(7):2291–2303. doi: 10.4049/jimmunol.1600924

Figure 10. HspA1A is required for alcohol-mediated inhibition of TLR4-MyD88 dependent TNFα but not TLR4-MyD88 independent RANTES.

Figure 10

(A) RAW macrophages were transfected with 10 nM siRNA targeting HspA1A or negative control (Neg ctrl) siRNA. Cells were subjected to heatshock (HS) (42°C for 45 min) and total RNA was used for HspA1A mRNA determination by qRT-PCR. Percent knockdown in HS cells was calculated with respect to untransfected. Graph depicts mean ± SD (n=3). Representative gels are shown to illustrate knockdown at protein level with HspA1A (top) and loading control, β-actin (bottom). (B,C) RAW macrophages were transfected with 10 nM siRNA targeting HspA1A or negative control (Neg ctrl) siRNA. 24 hours after transfection, macrophages were stimulated with 100 ng/ml LPS for 2 hours, 50 mM alcohol (EtOH) alone for 24 hours or pre-exposed to alcohol followed by LPS stimulation for indicated times. (B) TNFα and (C) RANTES mRNA was analyzed by qRT-PCR. Graph depicts mean ± SD of 3 independent experiments and statistical analysis was performed by one way ANOVA with Sidak’s test for multiple comparisons. (ns p>0.05, * p<0.05, ** p<0.01, *** p<0.001, ****p=0.0001)