目的:探讨膜结合型干细胞因子（membranebound stem cell factor，mSCF）在白血病细胞中的作用。方法：克隆并构建携可溶型干细胞因子（soluble stem cell factor，sSCF）前体的真核表达质粒pTARGETs，采用Overlap PCR法去除外显子6序列，进一步构建mSCF的表达质粒pTARGETm，DNA测序鉴定。通过脂质体介导分别将上述载体和空载体pTARGET转染白血病K562细胞，用G418筛选稳定表达细胞株，并用RTPCR、Western blotting法鉴定。通过CCK8细胞增殖实验以及集落形成实验观察不同细胞株体外增殖特点。结果：成功构建了sSCF和mSCF的真核表达载体，获得了稳定转染细胞株K562V、K562S、K562M。U底培养条件下，K562M增殖能力明显高于K562V和K562S（均P<0.01）。K562M集落形成率显著高于K562V 和K562S (均P<0.01)，且集落形态大于其他两种细胞。结论： mSCF与Ckit之间的并置性作用显著促进白血病细胞的增殖。

Objective: To explore the role of membranebound stem cell factor (mSCF) in the pathogenesis of leukemia. Methods: The eukaryotic expression plasmid of soluble stem cell factor（sSCF）precursor (pTARGETs) was constructed. Overlap PCR was used to obtain mSCF sequence with the deletion of exon 6, and pTARGETm was constructed. After verified by DNA sequencing, pTARGETm and control pTARGET vector were transfected into K562 cells via Lipofetamine 2000, and the positive cells were screened by G418. K562 cells stably transfected with pTARGETm were verified by RTPCR and Western blotting. Proliferation and colonyformation of these stably transfected cells were studied. Results: The sSCF and mSCF eukaryotic expression vectors were successfully constructed. The stable transfectants K562V, K562S, and K562M were obtained. Under Ubottom culture condition, proliferation ability of K562M cells was significantly stronger than those of K562V or K562S (both P<0.01). Colonyformation ability of K562M was significantly higher than those of K562V and K562S (both P<0.01). Furthermore, the colony size of K562M was larger than those of the other two kinds of cells. Conclusion: mSCF significantly enhances proliferation and colonyformation of leukemia cells by a juxtacrine mechanism.

国家重点基础研究发展规划（973）前期资助项目（2008CB517301）；美国中华医学基金会（CMB）资助项目（No.2007001）