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J. M. Harder, L. Gan, R. T. Libby; Analysis of Pro-Apoptotic Bcl2 Family Member Initiators BIM, BID, and BBC3 in Retinal Ganglion Cell Death. Invest. Ophthalmol. Vis. Sci. 2010;51(13):2118.
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During development and after axonal insult, retinal ganglion cell (RGC) death is dependent on the Bcl2 family member BAX. Three pro-death Bcl2 family members, BID, BIM, and BBC3 are thought to be critical for BAX activation. Here, we investigate which of these molecules are responsible for activating BAX during developmental and axonal injury induced RGC death.
Null alleles of the Bax, Bim, Bbc3, and Bid on the C57BL/6J genetic background were used. Cleaved CASP3 and POU4F2 positive cells or RGC layer neurons were counted at various times in retinal development, in the adult, or after control optic nerve crush (CONC) to determine the role of these genes in RGC death.
Deficiency in Bim or Bid did not alter the number of adult RGC layer neurons (n>5; P>0.05). In contrast, Bbc3 deficiency significantly increased the number of RGCs that survived into the adult, as judged by the number of RGC layer neurons (% +/+ ± SEM, 157 ± 3.0%, n=7, P<0.001) and POU4F2 positive cells (174 ± 11%, n=3, P<0.01). This increase corresponded to a decrease in cleaved CASP3+ cells during RGC development. The increase in RGC layer neurons in Bbc3-/- mice approaches the rescue of RGC layer neurons in Bax-/- mice (176 ± 5.3%, n=4) but is significantly different (P=0.009). Deficiency in either Bim or Bbc3, but not Bid, delayed RGC death 7 days after CONC (% unmanipulated eye ± SEM: +/+, 78 ± 2.3%, n=5; Bbc3-/-, 86 ± 1.8%, n=5, P=0.006; Bim-/-, 94 ± 2.8%, n=4, P<0.001), but, unlike in Bax-/- mice, there was no protection by 60 days (P>0.05).
During development BBC3 plays a major role in RGC apoptosis. However, after axonal injury, a single deficiency in the three most potent pro-apoptotic Bcl2 family members, including the known direct activators of BAX, did not reproduce the complete long-term protection provided by BAX deficiency. Thus, either multiple pro-death members of the Bcl2 family are important in RGC death after axonal injury or other mechanisms of BAX activation are involved.
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