Cloning of SDMF and role in vivo

1. 平滑肌细胞源性迁移因子（SDMF）的纯化：从100 L的大鼠主动脉平滑肌细胞条件培养基中纯化SDMF。SDMF cDNA的克隆：纯化后的SDMF经酸切成25kD和35kD两条肽段。从25kD肽中得到13个氨基酸序列。该序列与某种小鼠蛋白（蛋白X）高度同源。分别从小鼠心肌、大鼠心肌和人胎盘中合成了X蛋白的引物，并获得了PCR产物（cDNA）。利用这些cDNA作为探针，从大鼠培养平滑肌细胞和人胎盘的DNA文库中克隆出X蛋白的cDNA。从这些cDNA中得到的氨基酸序列包含上述序列。将来自平滑肌细胞的cDNA以pGM表达载体转染到COSl细胞中，在培养基中发现了与纯化后的SDMF大小相近的蛋白。对这种蛋白质的精确分析正在进行中。调节SDMF: tgf - β抑制大鼠平滑肌细胞合成/分泌SDGF，在1ng/ml时抑制50%。此外，动脉粥样硬化内膜的平滑肌细胞比正常介质分泌更多的SDMF。该研究表明，SDMF在动脉粥样硬化的发展中起作用，但在其开始过程中不起作用。

1. Purification of smooth muscle cell derived migration factor (SDMF) : SDMF was purified from 100 L of conditioned medium of rat aortic smooth muscle cells as previously described.2. Cloning of SDMF cDNA : Purified SDMF was digested by an acid into two peptides, 25kD and 35kD peptides. 13 amino acid sequence was obtained from the 25kD peptide. The sequence was highly homologous to a certain mouse protein (protein X). Primers for protein X were synthesized and PCR products (cDNA) were obtained from mouse heart muscle, rat heart muscle and human placenta. Using these cDNA as probe, . cDNA of protein X was cloned from DNA libraries of rat culture smooth muscle cells and human placenta. Amino acid sequence derived from these cDNA contained the sequence mentioned above. cDNA from smooth muscle cells was transfected into COSl cells with an expression vector, pGM,and the protein with similar size of purified SDMF was found in the culture medium. The precise analysis of this protein is going on.3. Regulation of SDMF : TGF-beta inhibited the synthesis/secretion of SDGF from rat smooth muscle cells, being 50% inhibition at 1ng/ml. Moreover, smooth muscle cells from atheroscerotic intima secreted more SDMF than those from normal media.4. The study suggested that SDMF plays a role in the development of atherosclerosis but not in its initiation.