Fig. 8: The ROCK1/2 inhibitor, Fasudil, blocks PEG-IFNα induced ROCK1/2 activation and enhances PEG-IFNα effects in a MPN mouse model.

a–j CD45.1 mice were transplanted with Jak2^V617F/+ KI bone marrow cells isolated from Jak2^V617F/+VavCre+ CD45.2 donor mice. Five weeks after transplantation and confirmation of engraftment (see Supplementary Fig. 10a–e), CD45.1 recipient mice were treated with vehicles (PBS, CTRL), or PEG-IFNα (IFNα, 600 ng/mouse, subcutaneous injection) once a week, and/or Fasudil (FAS, 25 mg/kg, intraperitoneal injection) four times per week for 4 weeks. a Schematic illustration of the MPN mouse model, treatment regimen per week, and analyses performed. Gy: gray units of ionizing radiation dose. b Scatter dot plots show median fluorescence of phosphorylated MYPT1 (p-MYPT1 A647) in peripheral blood mononuclear cells collected 26 h after the third dose of PEG-IFNα and 2 h after the eleventh dose of Fasudil and/or respective vehicles (CTRL, n = 3; FAS, n = 4; IFNα, n = 4; FAS + IFNα, n = 4). Data were assessed by flow cytometry and the gating strategy used is shown in Supplementary Fig. 7h. Results are representative of two independent experiments. c–j Scatter dot plots show effects of Fasudil (FAS) and/or PEG-IFNα (IFNα) treatments on (c) hematocrit (HCT) percentage and (d) hemoglobin (HB) levels in peripheral blood, (e) spleen weight (images are representative of spleen size per treatment group), f–g percentage of Ter119^highCD71^high basophilic erythroblasts (R2) in f bone marrow (BM) and g spleen, and on the h–j percentage of bone marrow h myeloid progenitors, i granulocyte-monocyte progenitors (GMP) and j megakaryocyte-erythroid progenitors (MEP). c–j Data shown are means ± SEM pooled from four independent studies (CTRL n = 15, FAS n = 17, IFNα n = 15, FAS + IFNα n = 18); each independent study performed using 3–6 mice per treatment group. The flow cytometry gating strategy used is shown in Supplementary Figs. S11a and S11f. b–j Statistical analyses were performed using one-way ANOVA followed by Tukey’s multiple comparisons test. Adjusted p-values are reported. See also Supplementary Figs. 10 and 11. Source data are provided as a Source Data file.