Figure 5.

(A) Analysis of effects of the compounds on the mRNA expression of cell survival and apoptotic genes. An upregulation was observed in genes of caspases family and proapoptotic genes ie, HRK, BAX and BAK, while anti-apoptotic BCL-2L1 and p53 genes and cell survival VEGF and COX-2 genes significantly downregulated. Interestingly, initiator caspase-8 of death receptor pathway is also inhibited after the treatment with betulin. GAPDH was used as reference gene. (B) Quantitative analysis of expression of apoptotic genes compared to control. *P<0.05 compared to control. ^**P<0.01; ^***P<0.001. (C) Immunocytochemical analysis of major apoptotic markers after 48 hours treatment with betulin observed under fluorescent microscope at 20× magnification. The caspase-3 and caspase-6 are highly expressed in treated cells while caspase-8 is downregulated, suggesting activation of intrinsic pathway of apoptosis. Furthermore, anti-apoptotic BCL-2L1 show marked inhibition posttreatment.

Abbreviation: Int Den, integrated density.