Human and simian immunodeficiency viruses deregulate early hematopoiesis through a Nef/PPARγ/STAT5 signaling pathway in macaques
J. Clin. Invest. Stéphane Prost, et al. 118:1765 doi:10.1172/JCI33037 [Go to this article.]

Figure 5
Nef action on hematopoietic cells depends on the presence of PPARγ and its downregulation of STAT5. (A) Clonogenic capacities of CD34⁺ BM cells were assayed following preincubation for 2 days with culture medium alone (control) or supplemented with SIV-rNef (rNef, 0.15 μM) or PPARγ agonists troglitazone (T) or rosiglitazone (R) or PGJ₂ (25 μM) in the presence or absence of PPARγ antagonist GW9662 (antag, 10 μM). Samples from 5 macaques were analyzed, each represented by 1 symbol. The number of colonies scored was expressed as percentage of control values. (B) Clonogenic capacities of CD34⁺ BM cells were assayed on cells transfected with or without PPARγ siRNA or irrelevant siRNA (ir) and incubated for 2 days in culture medium in the absence or presence of rNef before CFC assays. Samples from 5 macaques were analyzed, each represented by 1 symbol. CFC numbers were expressed relative to the CFC scored in the absence of siRNA PPARγ and rNef. (C) Clonogenic capacities of hematopoietic progenitors isolated from mice (n = 8, left) or macaques (n = 2, right), pretreated with or without rosiglitazone (agonist) during 2 weeks. (D) Real-time RT-PCR of STAT5B mRNA from murine BM sca-1⁺ c-kit⁺ CD3^– cells or macaques CD34⁺ BM cells, treated with or without rosiglitazone (agonist) during 2 weeks. STAT5B mRNA were normalized to GAPDH mRNA. (E) K562 cell proliferation, evaluated by [³H]thymidine incorporation, in cells incubated for 7 days in the presence of various concentrations of rNef or troglitazone. (F) Real-time RT-PCR of STAT5B mRNA from K562 cells incubated with or without siRNA directed against PPARγ and troglitazone (25 μM) or rNef (0.15 μM). STAT5B mRNA were normalized to GAPDH mRNA and were expressed relative to untreated cells. Efficiency of the siRNA transfection has assessed with a fluorescent control siRNA (25 nM) was over 98%; inhibition of PPARγ mRNA was 98% ± 2% (n = 5).