Protective autophagy is up-regulated in HIV reservoir cells

P.N. FERNANDEZ LARROSA, D. GRASSO, M.I. VACCARO, L. MARTINEZ PERALTA.

Cape Town, Sudáfrica, 19-22 de Julio de 2009.

Congreso; 5th IAS CONFERENCE ON HIV PATHOGENESIS, TREATMENT AND PREVENTION,; 2009

International Aids Society

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 10uM H2O2 and 0,1uM Staurosporine (STS) as apoptotic inducers, and with complete medium as control. Alternatively, cells were incubated in starvation with medium without aminoacids and serum 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 at a fluorescent microscopy in order to look for autophagosomes. Results: Both cell lines showed to be equally positive in a 50-60% when cells were in starvation, but negative when incubated with complete medium. When cells treated with H2O2 or STS were analysed, necrotic cells showed an inespecific stain; so, only viable cells were observed for following analysis. H9 cells presented only 15% of positive cells with a typic autophagosome stain. In contrast, 50% of viable H9HTLVIIIB cells resulted to be positive for MDC in a specific manner. (p