Figure 5

P. vulgaris extracts have a modest effect on HIV-1 binding to permissive cells. A) Pre-incubation of extract with cells. HeLa37 cells were pre-incubated with concentrations of aqueous extracts noted in the panel in DMEM with 10% FCS for 1 hour at 37°C. Unbound extracts were removed and cells were suspended in fresh media and 200 infectious units of HIV-1 NL4-3 added to the culture (MOI = 0.01). Cells were fixed and immunostained for HIV antigens 40 hours after infection. Pre-exposure of cells to extract reduced the level of virus infectivity by 20%, which was statistically significantly different from the control. *, p < 0.05. B) Pre-incubation of extracts with virions. Approximately 4 × 10⁴ infectious HIV-1 NL4-3 particles were combined with extracts and incubated at room temperature for 10 minutes. The mixture was diluted 100 fold with media to reduce extract concentrations to irrelevant levels and added to 2 × 10⁴ cells/well of HeLa37 cells in a 48-well format, resulting in a final MOI of 0.02. Cells were fixed at 40 hours following infection and immunostained as described above. **, p < 0.01; ***, p < 0.001. C) Ability of extracts to inhibit HIV-1 binding to cells. Increasing concentrations of P. vulgaris extract were incubated with HIV-1 NL4-3 (3.8 × 10⁵ infectious particles) (MOI = 19) and 2 × 10⁴ HeLa37 cells at 4°C for 2 hours in DMEM with 10% FCS. Unbound virus was removed with several washes of media and cells were then lysed. Cell lysates were immunoblotted for HIV-1 p24 and cellular β-actin. All experiments were performed in triplicate and independently performed three times.