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目的:建立稳定表达人滋养层细胞表面抗原(Trop-2)NIH3T3细胞,分析过表达Trop-2对NIH3T3细胞的生长、增殖和侵袭特性的影响。方法:将Trop-2基因克隆到真核表达载体pcDNA3.1,转染NIH3T3细胞,通过G418筛选及RT-PCR鉴定获得稳定表达Trop-2的NIH3T3细胞(NIH3T3-Trop-2)。用MTS法检测NIH3T3-Trop-2细胞的增殖能力,软琼脂集落形成实验检测NIH3T3-Trop-2细胞的克隆形成能力,明胶酶谱法检测NIH3T3-Trop-2细胞的基质金属蛋白酶(MMP)-2和MMP-9的分泌及细胞划痕实验检测NIH3T3-Trop-2细胞的迁移能力。结果:稳定表达Trop-2的NIH3T3细胞在生长增殖、克隆形成及侵袭能力均较NIH3T3细胞强,细胞培养上清中的MMP-2和MMP-9增多。结论:Trop-2对细胞的增殖与迁移能力具有明显的促进作用。
OBJECTIVE: To establish NIH3T3 cells stably expressing human trophoblast cell surface antigen (Trop-2) and analyze the effect of Trop-2 overexpression on the growth, proliferation and invasiveness of NIH3T3 cells. Methods: Trop-2 gene was cloned into eukaryotic expression vector pcDNA3.1 and transfected into NIH3T3 cells. NIH3T3 cells (NIH3T3-Trop-2) stably expressing Trop-2 were obtained by G418 screening and RT-PCR. The proliferation of NIH3T3-Trop-2 cells was detected by MTS assay, the colony formation ability of NIH3T3-Trop-2 cells was detected by soft agar colony formation assay, and the matrix metalloproteinase (MMP) 2 and MMP-9 secretion and cell scratch assay NIH3T3-Trop-2 cell migration. Results: NIH3T3 cells stably expressing Trop-2 were stronger than NIH3T3 cells in proliferation, clonogenicity and invasion, and MMP-2 and MMP-9 were increased in the cell culture supernatant. Conclusion: Trop-2 can significantly promote the proliferation and migration of cells.