RESISTANCE TO APOPTOSIS BY H2O2 AND STAUROSPORINE IN AN HIV-1 PERSISTENTLY INFECTED CELL LINE

P. N. FERNÁNDEZ LARROSA; D. A. RIVA; M. BIBINI; S. E. MERSICH; L. A. MARTÍNEZ PERALTA

Toronto, Canada

Conferencia; XVI International AIDS Conference; 2006

Background: HIV-1 is known to modify redox state in infected T cells. Besides, high sensitivity of acutely HIV-infected T cells to different apoptosis inducers, as well as resistance to apoptosis in HIV-infected reservoir cells have been described. In order to better understand this phenomenon, persistently infected and non infected human lymphoblastoid cell lines were treated with two different apoptosis inducers and results were compared. Methods: Non infected H9 and persistently infected H9HTLVIIIB T cell lines were treated with different concentrations of H2O2 or Staurosporine (STS) and collected at 24 hours post treatment. Cellular viability was assayed by measuring mitochondrial succynate dehydrogenase functionality (MTT reactive) and normalized to the respective cell control (CC) as percentage. Apoptosis was evaluated by dual stain with annexin-V-FITC and Propidium Iodure (PI) and revealed by FACS. Antigen p24 production was measured with a commercial kit. Significance was analyzed by T Student test. Results: H9HTLVIIIB cells showed 32% higher viability when treated with 10uM H2O2 (P=0.004) and 45% with 20uM H2O2 (P<0.001) than H9 cells. Treatment with 0.1 uM STS showed similar results: 16% higher viability at H9HTLVIIIB than H9 cells (P<0.001). These results were confirmed significantly by annexin-V/PI staining. H9 presented 20% higher levels of early apoptosis (P=0.01) when treated with 10uM H2O2 and 18% when treated with 0.1uM STS (P=0.014) than H9HTLVIIIB. When H9HTLVIIIB cells were treated with H2O2, cell death seemed to occur mainly by necrosis than apoptosis; but even in this case, total cell death was lower in these cells than in H9. p24 levels were significantly decreased in both treatments. Conclusions: Non infected H9 cell line is more sensitive to H2O2 and STS than the persistently infected H9HTLVIIIB. This in vitro resistance to apoptosis could be related to the in vivo HIV reservoir cells persistence.