The Study Of Wnt-10a During Erythroid Differentiation

Erythroid differentiation is one of the complicated and precise regulatory processes in organism, which occurs during embryonic development, stem cell differentiation and regulation disorder in cancer cells. Erytoroid differentiation is combinatively regulated by small molecules such as piRNA and microRNA, numerous transcription factors and signaling pathways. Once one of these factors becomes abnormal, it always to be a disorder in differentaiation of hematopoisis, uncntrolled cell proliferation and cancer. In previous studies about leukemia, cancerous states can be reversed by series of small moleculeus such as Hemin, DMSO, Sodium butyrate and so on. So far,analyzing of the regulatory mechanism in erythroid differentiation are mainly through the discovery and study of differentiation-related proteins(such as cell differentiation factors).Proteomics and gene chip technology has given key supports for these investigation, also provides experimental basis for clinical inducers which can normalize leukemia cells to controllable normal differentiation and apoptosis process.In previous studies, we used 2-D electrophoresis technology to evaluate the proteins differential expressed in induced K562 and got hundreds of proteins which are possibly involved in erythroid differentiation regulation. In theses identified proteins, we found Wnt-10 a, which has a apparent expression change in induce differentiation process. Wnt is a large secreted protein family which is always relate to embryonic development, maintenence and differentiation of stem cells, cell proliferation, migration, polarition, cell death and cancer. Because Wnt signaling pathway is also involved in the regulation of stem cell differentiation process, and Wnt-10 a was foud in erythroid differentiation proteome, this paper intendes to investigate the function of Wnt-10 a in induced differentiation, and further more discove how wnt signaling pathway affects erythroid differentiation. By using the mehods of immunocytochemistry, RT-PCR, Real Time PCR, Western Bloting, MTT and other cell study technology, we initiatively study the protein expression and location changes in K562 induced by Hemin and transfect Wnt-10 a overexpression recombinant vector to K562 cell. We also use CHIR90021, an pharmacological activator of Wnt signaling pathway to identify the affect of Wnt signaling pathway during the erythroid differentiaton of K562 cell induced by Hemin.The results of RT-PCR, Real Time PCR and Western Blot reveals that the expresion level of Wnt-10 a has momently declined during induced differentiation of K562 cells. Meanwhile, Wnt-10 a has migrated to cell membrance when K562 was induced by Hemin. Theses experiments also confirmed three Wnt signaling pathways have all changed during induced erythroid differentiation. By using the Wnt signaling pathway activator the proliferation activity of K562 has raised during Hemin inducement, also suggested that Wnt signaling pathway has significant impact on the generation of leukemia.The results above show that Wnt-10 a and even Wnt signaling pathway has values worth further investigation in erythroid differentiation of mammalian cells. For this reason we expects to do more research to recovery the inherent mechanism of erythroid differntiation and give a better therapeutic approach for leukemia.