Down-regulation of TGF beta receptor I (TBRI) can inhibit smad2 activation in hematopoietic cells and stimulate MDS hematopoiesis. GFP-expressing lentiviral-based shRNA against TBRI (A) was used to knock down TBRI in hematopoietic cells. qPCR shows decrease in TBRI mRNA expression after lentiviral shRNA-TBRI infection in bone marrow stromal (HS-5) and leukemia cells (K562) compared with scrambled control (B). K562 cells with stable expression of TBRI-shRNA lentivirus show decreased smad2 phosphorylation after TGF stimulation (C). Primary CD34⁺ progenitors were electroporated with GFP coexpressing anti–TBRI-shRNA construct and sorted after 48 hours. GFP-positive cells were grown in methylcellulose with cytokines, and erythroid colonies were counted after 14 days. TBRI-shRNA–transfected progenitors were less inhibited by TGF-β compared with cells transfected with scrambled control shRNA (31% colonies/control vs 16% colonies/control). Expressed as means (± SEM) of 4 independent experiments (P = .03, 2-tailed t test) (D). CD34⁺ cells transfected with anti–TBRI-shRNA also formed bigger colonies in the presence of TGF-β1 compared with controls (E; Nikon, 40×). Primary bone marrow–derived mononuclear cells from 5 patients with MDS were transfected with shRNA targeting TBRI and control, and equal numbers of cells (for each individual patient) were grown in methylcellulose with cytokines. Erythroid (BFU-E) and myeloid (CFU-GM) colonies were counted after 14 days of culture and demonstrated an increase after anti–TBRI-shRNA transfection (significance between means calculated by 2-tailed t test) (F).