Roles And Mechanism Of SHP-2 Tyrosine Phosphatase In Myeloid Cytotoxicity Induced By DNA Damaging Chemotherapy And Radiotherapy

AIM: To demonstrated potential roles and signal mechanism for SHP-2 tyrosine phosphatase in myeloid cytotoxicity induced by DNA damaging chemotherapy and radiotherapy.METHODS: Mouse embryonic yolk sac hematopoietic progenitor cell were derived from day 9.0–9.5 embryos of SHP-2 homozygous mutant mouse (SHP-2-/-),SHP-2 heterozygosis mutant mouse(SHP-2+/-) and wild type (WT) mouse. Then they were committed to granulocyte-macrophage (GM) differentiation after incubated with IL-3(0.5ng/ml)and SCF(0.5ng/ml). These three kinds of GM cell were then treated with cisplatin(CDDP) or 60Coγ-irradiation and subjected to trypan blue staining and colony forming units analysis (CFU). Haemacytometry was applied to detect the levels of leucocyte, akaryocyte and haematoglobin in WT and SHP-2+/- mice. SHP-2 Rescued cell lines were generated by transduction of WT SHP-2 cDNA into SHP-2-/- GM hematopoietic cell through retroviral-mediated gene transfer. And then Fluorescence- activated Cell Sorting (FACS) was used to Cell Cycle Analysis in the WT cell, Rescued cell and SHP-2-/- cell.Lastly, western bolt was applied to exam the expression and phosphorylation of Cdc2, Chk1, Erk and p38 in WT and SHP-2-/- mutation hematopoietic cell.RESULTS: Trypan blue staining showed that following DNA damage induced by cisplatin or 60Coγ-irradiation, cell reproductive activity of SHP-2-/- hematopoietic cells were much better than SHP+/- and WT hematopoietic cells. Levels of leucocyte, akaryocyte and haematoglobin in blood of SHP-2+/- mutant mice were also higher than WT mice. Colony Forming Units Analysis showed an increased numbers of colony units of SHP-2-/- hematopoietic cell compared with SHP+/- cell and WT cell. Cell Cycle analysis showed the G2 (but not S) arrest response was diminished in hematopoietic cells lacking functional SHP-2. Notably, reintroduction of wild-type SHP-2 into the mutant cells fully restored the DNA damage-induced G2 arrest response, suggesting a direct role of SHP-2 in the G2/M checkpoint. Further biochemical analysis revealed that SHP-2 constitutively associated with Cdc2 phosphorylation. Additionally, we showed that following CDDP treated activation of p38 kinase was significantly elevated, while Erk kinase activation was decreased in the SHP-2-/- mutant hematopoietic cells.CONCLUSIONS: SHP-2 is involved in regulation of myeloid cytotoxicity induced by DNA damaging chemotherapy and radiotherapy.This effect may be base on regulation of G2/M arrest.SHP-2 enhances the DNA damage-induced G2/M arrest by inhibiting Cdc2 and by differentially regulating the MAP kinase pathways.