Fhl2 Regulates The Function Of Hematopoietic Stem/Progenitor Cells

Fhl2 is a member of four and one half LIM domain protein family. It acts as a transcriptional modulator, regulating proliferation and apoptosis in a cell-context dependent manner. The role of endogenous Fhl2 in hematopoietic stem/progenitor cells (HSCs/HPCs) is still unknown. To determine the in vivo function of Fhl2 in hematopoiesis, we characterized the HSCs/HPCs in Fhl2-null mice. At age of 3 months, Fhl2-null mice revealed normal hematopoietic parameters. As shown by flow cytometric analysis, Fhl2-null mice had a normal distribution of Mac-1⁺Gr⁺ myeloid cells, B220⁺ B cells and CD3⁺ T cells in bone marrow and spleen, suggesting that Fhl2 loss does not affect mature cell differentiation in young mice. The frequency and total number of stem cell enriched population LSK (Lin^-Sca⁺c-Kit⁺) and Long-term HSCs (LT-HSCs) (Lin^-Sca⁺c-Kit⁺CD48^-CD150⁺) were marginally increased in Fhl2^-/- mice as compared to wildtype mice, whereas the frequency and the number of hematopoietic progenitor cells was reduced in Fhl2^-/- mice as compared to Fhl2^+/+ mice.

Transplantation exposes HSCs to various stresses including replicative and oxidative stresses. To examine whether Fhl2 regulates the function of HSCs/HPCs under stress, we transplanted bone marrow cells from Fhl2^-/- or control mice into lethally-irradiated wildtype mice to generate chimeric mice. 7 months after transplantation, the Fhl2^-/- chimeric mice developed MDS-like disease with a significant decrease in platelet, red blood cell (RBC) counts and Hemoglobin (Hb) level as compared to control chimeric mice. The number of LT-HSC, LSK and HPCs in Fhl2^-/- chimeric mice were significantly less than they were in control chimeric mice, as determined by flow cytometric analysis. However, the frequency of Mac-1⁺Gr-1⁺ myeloid cells, B220⁺ B cells and CD3⁺ T cells were comparable in Fhl2^-/- and Fhl2^+/+ chimeric mice, suggesting that Fhl2 loss reduces repopulation capacity of LT-HSCs but does not affect HPCs differentiation in recipient mice. As evidenced by the reduced G₀ population of HSCs and LSKs in Fhl2-null chimeric mice as compared to control chimeric mice, Fhl2 may regulate HSCs self-renewal under stress by controlling HSC quiescence. To further determine the function of HSCs, we performed the competitive repopulation assay. In agreement with the observation that Fhl2^-/- HSCs have a reduced repopulation capacity, the ratio of Fhl2^-/- derived total Peripheral Blood (PB) cells vs. competitor-derived PB cells was significantly decreased as compared to Fhl2^+/+-derived recipients at secondary transplantation, and the ratio gradually decreased up to 4-5-fold as compared to Fhl2^+/+-derived recipients at tertiary transplantation.

Previously, we showed that FHL2 is down-regulated in subsets of Therapy-related Myelodyplastic Syndrome or Acute Myeloid Leukemia (t-MDS/t-AML patients). Together, these data suggest that Fhl2 is required for maintenance of HSCs/HPCs, and its downregulation may contribute to the development of t-MDS/t-AML in patients.