Figure 5. Influence of CRF₁ knockdown and overexpression in RBL-2H3 MCs on IgE-mediated MC degranulation.

(A and B) IgE/DNP induced β-hex percentage of release from RBL-2H3 cells transfected with CRF₁ shRNA (A) or siRNA (B) (and scrambled controls). (C) IgE/DNP-induced histamine percentage of release into cell culture supernatants from RBL-2H3 cells transfected with siRNA. (D and E) IgE/DNP-induced β-hex (D) and histamine (E) percentage of release from RBL-2H3 cells transfected with lentiviral CRF₁ overexpressing plasmid (and lentiviral empty plasmid control). (F) Western blots showing reduction in CRF₁ protein bands at ∼55 and 37 kDa in CRF₁-shRNA RBL-2H3 MCs and combined densitometric analysis (55 and 37 kDa bands) of CRF₁ expression (relative to β-actin expression) presented as a histogram. (G) Western blot analysis showing enhanced expression of CRF₁ in CRF₁-overexpressed RBL-2H3 MCs and associated densitometric analysis. Data shown are the means ± se and are representative of 3 independent experiments performed in triplicate. Data were analyzed using either a t test (A and C) or 2-way ANOVA (B and D) with a post hoc Tukey’s test. *P < 0.05; **P < 0.001; ***P < 0.0001; *^# P < 0.05 vs. other treatment groups (ANOVA).