Figure 2

Activation of cofilin in resting CD4 T cells of HIV-1-infected patients. (A) Resting CD4 T cells were purified by antibody-mediated negative depletion using Dynalbeads. Cells purified from HIV-1-infected and uninfected donors were cultured overnight in the absence of cytokines or activation, and then lysed and analyzed by western blot using antibodies against P-cofilin or total cofilin. The relative ratios of P-cofilin to total cofilin were measured and plotted. HIV-1-infected patients had statistically-significant lower ratios of P-cofilin/cofilin (0.535 versus 1.142, p = 0.002), suggesting higher levels of active cofilin. For statistic analysis, a two-tailed Student's t-test on the means resulted in a p-value of p = 0.002. At a pre-determined significance level of 0.05, this shows that the difference in the mean ratios of the two sample groups is statistically significant. A standard power computation showed that the t-test was very well powered (95.2%) for this study. (B) Shown are the longer exposures of the western blots used in (A). The results were confirmed by NEPHGE-western blot to directly separate P-cofilin to active cofilin, and then probed with an anti-cofilin antibody [20]. Shown are the absolute ratios of P-cofilin to active cofilin in HIV-1 negative donors (C) and HIV-1-infected donors (D).