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目的观察人类新血小板反应素R-spondin 3(Rspo 3)的细胞定位,并探讨其在肿瘤发生发展中的可能作用。方法利用荧光显微成像法观察EGFP-Rspo 3在HEK293细胞中的分布。将重组质粒pcDNA-Rspo 3转染结肠癌细胞HT-29、LoVo,采用流式细胞术观察Rspo 3基因过表达对肿瘤细胞周期与凋亡的影响;采用Matrigel、Transwell实验分别检测Rspo 3基因过表达对肿瘤细胞黏附及侵袭能力的影响。结果荧光显微成像法观察发现,EGFP-Rspo 3融合蛋白在细胞核表达,呈弥散点状分布。流式细胞术观察发现,Rspo 3基因过表达对肿瘤细胞生长周期没有明显影响;Rspo 3基因过表达不影响低恶性的HT-29细胞凋亡,但诱导高恶性的LoVo细胞凋亡(P<0.01)。Rspo 3基因过表达增强肿瘤细胞黏附性(P<0.01),降低肿瘤细胞的侵袭力(P<0.01),对肿瘤细胞移行有一定抑制作用。结论 Rspo 3有核定位功能,能诱导部分肿瘤细胞凋亡并抑制其转移扩散。
Objective To observe the cellular localization of human R-spondin 3 (Rspo 3) and to explore its possible role in tumorigenesis. Methods The distribution of EGFP-Rspo 3 in HEK293 cells was observed by fluorescence microscopy. The recombinant plasmid pcDNA-Rspo 3 was transfected into colon cancer cells HT-29 and LoVo, and the effect of Rspo 3 gene overexpression on cell cycle and apoptosis was observed by flow cytometry. Matrigel and Transwell assay were used to detect the expression of Rspo 3 Effect of Expression on Adhesion and Invasion Ability of Tumor Cells. Results Fluorescence microscopy showed that the EGFP-Rspo 3 fusion protein was distributed in the nucleus. Flow cytometry showed that overexpression of Rspo 3 had no significant effect on the growth of tumor cells. Overexpression of Rspo 3 did not affect the apoptosis of low-grade HT-29 cells but induced the apoptosis of high-grade LoVo cells (P < 0.01). Overexpression of Rspo 3 enhanced the adhesion of tumor cells (P <0.01), reduced the invasiveness of tumor cells (P <0.01), and inhibited the migration of tumor cells. Conclusion Rspo 3 has nuclear localization function, which can induce some tumor cell apoptosis and inhibit its metastasis and diffusion.