Modulation of prostate cancer genetic risk by omega-3 and omega-6 fatty acids
J. Clin. Invest. Isabelle M. Berquin, et al. 117:1866 doi:10.1172/JCI31494 [Go to this article.]

Figure 7
Dependence on Bad for omega-3 PUFA–induced cell death. (A) PC3 cells were infected with lentivirus expressing scrambled shRNA (C shRNA) or Bad-specific shRNA (Bad shRNA) or mock infected (no shRNA) for 2 days. Cells were seeded and incubated with FAs for 6 days, then photographed using a fluorescence microscope. The presence of lentivirus is evidenced by GFP expression. Western blotting was performed to confirm the successful knockdown of Bad in both PC3 and LNCaP cells. Day 0, prior to FA treatment; day 6, after incubation with media containing FA for 6 days. C, control. (B) Live and dead cells were enumerated by the trypan blue exclusion method using a hemocytometer. P values were determined by Student’s t test. LNCaP cells have higher background apoptosis compared with PC3 cells, but omega-3 FA treatment increased apoptosis approximately 4-fold in both cell lines.(C) LNCaP cells were transfected with control vector or mouse Bad expression vector. G418-resistant colonies were isolated, verified for successful expression of the HA-tagged mouse Bad by Western blotting, and pooled. Vector control– (V) and mouse Bad–expressing (mBad) LNCaP cells were infected with shRNA lentivirus as described above. Cells were incubated with FAs for 6 days, and live and dead cells were enumerated as described above. Cell lysates were used for confirmation of successful knockdown of the endogenous Bad but not the exogenous mouse Bad (whose sequence varies from human Bad in the region targeted by the shRNA).