FIG 2.

Long term persistence of HIV-1 latency in renal epithelial cells. Gating strategy for flow cytometry sort of latently infected and actively infected Hpt1-b renal epithelial cells (day 5 postinfection). Uninfected cells were used as negative control. Positive controls include Hpt1-b cells stably transduced with a lentiviral vector expressing either GFP or mCherry. (b) Flow-sorted primary renal cells (HRCEp cells) were reseeded and the expression of mCherry and GFP was analyzed at 2 weeks post-sort in both the latent (top panel) and active cells (bottom panel). (c) Flow-sorted mCherry only (latent) Hpt1-b cells were cultured for 12 weeks postinfection. mCherry expression could be detected in about half of the cells, while the other half did not express any fluorescent marker. No GFP expression (active infection) was detected at this late time point. Each panel represents a separate experiment. (d) The presence of the HIV-R7GEmC construct DNA was assessed in the four populations of flow-sorted Hpt1-b cells: GFP only, GFP/mCherry double positive (DP), latently infected (mCherry only) and double negative (DN). Serial dilutions of genomic DNA extracted from Hpt-1b cells stably transduced with a lentiviral vector expressing mCherry were used to generate a standard curve. The amount of DNA and the corresponding number of cells used to generate the standard curve are indicated. Uninfected Hpt1-b cells were included as negative control. NTC = no template control.