Figure 5.

HIV, interferon (IFN)-γ, and vitamin D receptor (VDR) agonists induce apolipoprotein (APO) L1 and transition markers in parietal epithelial cells (PECs). A: To examine the effect of APOL1 stimulators on APOL1 induction and expression of PEC transition markers, PECs were incubated in media that contained vehicle [control (C)], VDR agonists (EB1089, 100 nmol/L), or IFN-γ (10 ng/mL) for 48 hours. Protein blots were probed for APOL1, Wilms' tumor 1 (WT1), podocalyxin (PDX), and α-actinin and reprobed for glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Gels from two different lysates are shown. B: To assess the effect of HIV on APOL1 induction and expression of PEC transition markers, PECs were transduced with vector (Vec) or HIV [NL4-3, 10³ green fluorescent protein (GFP)–expressing units (GEU)/mL]. Protein blots were probed for APOL1, WT1, podocalyxin, and α-actinin and reprobed for GAPDH. Gels from two different lysates are shown. C: Cumulative densitometric data from the cells treated with VDA and IFN-γ are shown. D: Cumulative densitometric data from the cells transduced with Vec or HIV are displayed. IFN-γ–, VDA receptor–, and HIV-induced APOL1 expression is associated with the expression of transition markers in PECs. E: To evaluate the effect of APOL1 induction on the transcription of PEC markers, RNAs were extracted from the lysates of HIV-, IFN-γ–, and VDR agonist–treated cells (A and B). cDNAs were amplified with specific primers for PAX2 and claudin 1. Cumulative data are shown in a bar diagram. APOL1 induction in PECs attenuates the expression of PEC markers. F: To examine the effect of APOL1 induction on the transcription of PEC transition markers, RNAs were extracted from the lysates of HIV-, IFN-γ–, and VDR agonist–-treated cells (A and B). cDNA was amplified with specific primers for APOL1, WT1, α-actinin, PDX, and CD2AP. Cumulative data are shown in bar graphs. APOL1 induction in PECs results in enhanced transcription of PEC transition markers. G: To visualize the expression of PEC transition markers in response to APOL1 inducers, PECs grown on coverslips were treated under similar conditions (as in A) and labeled for PEC transition markers. Representative fluoromicrographs are displayed. Expression of APOL1, α-actinin, and PDX is indicated by green fluorescence and of WT1 by red fluorescence. H: To visualize the expression of PEC transition markers in response to HIV, PECs grown on coverslips were transduced with VEC (GFP positive) or HIV (GFP positive) and labeled for PEC transition markers. Representative fluoromicrographs are displayed. Both Vec- and HIV-transduced cells are GFP positive (indicated by green fluorescence). HIV-transduced cells display an overt expression of APOL1, synaptopodin (SYNPT), α-actinin, and WT1 (red fluorescence). n = 4 (A and B); n = 3 (E and F). ^∗P < 0.05, ^∗∗P < 0.01, ^∗∗∗P < 0.001 versus respective control. Scale bars: 5 µm (G, all panels); 10 µm (H, all panels). VDA, VDR agonist.