HMGB1促进人骨髓间充质干细胞迁移的实验研究

HMGB1 induces migration of human bone marrow-derived mesenchymal stem cells

目的:构建人高迁移率族蛋白(HMGB1)全长编码基因的原核表达载体,诱导和纯化重组蛋白,分析其对人骨髓间充质干细胞的迁移作用。方法:RT-PCR方法从人的单个核细胞中扩增出HMGB1全长编码基因,克隆于原核表达载体pET-24 a-d(+),经IPTG诱导表达,通过亲和层析方法纯化得到携带(H is)6标签的HMGB1融合蛋白;RT-PCR方法检测间充质干细胞HMGB1受体的表达;观察HMGB1对人骨髓来源间充质干细胞的迁移作用。结果:构建了融合蛋白HMGB1的原核表达载体,获得了高度纯化的HMGB1融合蛋白。RT-PCR方法检测到间充质干细胞表达HMGB1的某些受体如RAGE和TLR-4。HMGB1在体外能够诱导人骨髓来源的间充质干细胞的迁移。结论:重组HMGB1蛋白能够诱导骨髓间充质干细胞的迁移,此作用有可能通过RAGE和(或)TLR-4介导。

Objective： To observe the effect of recombinant human high mobility group box 1 （HMGB1） protein on the migration of human bone marrow derived-mesenchymal stem cells （MSC） in vitro. Methods： Total RNA was extracted from the mononuclear cells of normal adult, and the fall-length HMGB1 cDNA obtained by RT-PCR was cloned onto expression vector pET-24a-d （ ＋ ）. Expression of the （His）6-HMGB1 fusion protein was induced by addition of IPTG. The bacterial lysate was subjected to affinity purification using HiTrap Chelating column. The effect of recombinant HMGB1 on the migration of MSC from human bone marrow in vitro was observed, and the receptors for HMGB1 on MSC were detected by RT-PCR. Results： The recombinant HMGB1 was expressed and purified. Human bone marrow-derived MSC express the two of known-receptors for HMGB1, i.e. receptor for advanced glycation end product （RAGE） and Toll-like receptor 4 （TLR-4）. Furthermore, HMGB1 induces the migration of human bone marrow derived-MSC in vitro. Conclusion： The recombinant HMGB1 protein could induce the migration of MSC, which might be mediated by RAGE and/or TLR-4.