PROTECTIVE AUTOPHAGY IS UP-REGULATED IN HIV RESERVOIR CELLS

P. N. FERNÁNDEZ LARROSA; D. GRASSO; M.I. VACCARO; L. A. MARTÍNEZ PERALTA

Cape Town - SouthAfrica

Conferencia; 5th IAS Conference on HIV Pathogenesis, Treatment and Prevention; 2009

IAS

Background: HIV reservoir cells were shown to be resistant to cell death by different molecular mechanisms. Our group has already published that HIV-persistently-infected cells were resistant to apoptosis by downmodulating Bax at mitochondria in a Bcl-2-independent manner. However, one possible explanation to this resistance could be that HIV upregulates autophagy in persistently infected cells.Methods: In order to confirm this hypotesis, lymphoid cell line H9 and its HIV-persistently-infected clone, H9HTLVIIIB, were treated during 24 hours with 10µM H2O2 and 0,1µM Staurosporine (STS) as apoptotic inducers, and with complete medium as control. Alternatively, cells were incubated in starvation conditions for 6 hours. After incubation, cells were collected and incubated with 0.15 mM Monodansyl Cadaverine (MDC) for 10 minutes at 37ºC. After this, cells were washed with PBS 1X and observed using a fluorescent microscope in order to look for autophagosomes. Results: Both cell lines tested equally positive by 50-60% when cells were in starvation, though negative when incubated with complete medium. When cells treated with H2O2 or STS were analyzed, necrotic cells showed a non-specific stain; so, only viable cells were observed following analysis. H9 cells had only 15% of positive cells with a typic autophagosome stain. In contrast, 50% of viable H9HTLVIIIB cells tested positive for MDC in a specific manner (p< 0.01).Conclusions: These results clearly suggest that persistently infected cells showed higher levels of autophagy when treated with apoptotic inducers than uninfected cells. Protective autophagy could explain the Bax decrease at mitochondria since it could be responsible for eradicating damaged mitochondria before apoptosis breaks out. This is the first time that protective autophagy was shown to prevent apoptosis in a viral infection, and contribute to the survival of HIV reservoir cells.