Figure 5.
Jak2V617F-mutant Mk-derived IL-6 enhances erythropoiesis. (A) Multiplexed serum profiling (Luminex) of 32 cytokines/chemokines in sera from 5-month-old Jak2 V617F/+; Pf4-Cre+ mice and littermate controls showed significant increases in Ccl11 (Eotaxin), Cxcl1 (KC), Cxcl2 (MIP-2), and IL-6. Data shown are representative of 2 independent experiments. (B) ELISA confirming the increase in serum IL-6 levels observed in Jak2 V617F/+; Pf4-Cre+ mice. (C) IL-6 mRNA expression is elevated in CD41+ megakaryocytes sorted from Jak2 V617F/+; Pf4-Cre+ mice relative to littermate controls. IL-6 transcript levels were normalized to Gapdh. (A-C) N = 4 mice per group. (B-C) Results are representative of 3 independent experiments. (D-E) IL-6 increased BFU-E formation from wild type HSPCs, (D) LSKs and (E) MEPs, in a dose-dependent manner. LSK/MEP populations were FACS sorted from 6–8-week-old WT C57BL/6 mice. (F-H) Three pairs of 16–20-week-old male or female Jak2 V617F/+; Pf4-Cre+ mice were treated with either an antibody against IL-6 or PBS for 6 weeks. (F) IL-6 blockade markedly reduced BFU-E formation by splenocytes in vitro. Concordant decreases in the splenic erythroid progenitors (G) Pre-CFU-E and Pre-MegE populations and (H) immature erythroid (R1 and R2) populations were also observed by flow cytometry. (D-H) Results are representative of 2 independent experiment. * P < 0.05, ** P < 0.01 by unpaired two-sided Student’s T-test.