论文部分内容阅读
为研究乳腺癌微环境中前脂肪细胞和成熟脂肪细胞对乳腺癌MDA-MB-231细胞增殖和迁移能力的影响,该研究将前脂肪细胞3T3-L1诱导为成熟脂肪细胞,再将前脂肪细胞和成熟脂肪细胞分别与乳腺癌MDA-MB-231细胞共培养,通过显微镜成像、油红O染色实验、MTT实验、Transwell实验分别观察肿瘤细胞形态、增殖及迁移能力的改变。Western blot和ELISA检测前脂肪细胞和成熟脂肪细胞瘦素(leptin)的表达水平。细胞免疫荧光法和Western blot分别检测肿瘤细胞中瘦素受体(leptin recepter,Ob-R)、瘦素信号通路关键分子及下游靶因子的表达水平变化。结果显示,共培养后,肿瘤细胞形态变得更加纤长,增殖能力增加(P<0.05),穿过小室的细胞数明显增多(P<0.05)。在成熟脂肪细胞共培养组的肿瘤细胞中还出现了明显的脂质累积。Western blot和ELISA检测发现,前脂肪细胞和成熟脂肪细胞均有瘦素的表达。与空白对照组相比,两个共培养组中肿瘤细胞的p-Akt、p-STAT3、cyclin D1和MMP9蛋白质水平均明显上调(P<0.05),而p-ERK1/2仅在前脂肪细胞共培养组中上调(P<0.001),在成熟脂肪细胞共培养组中没有明显变化。和共培养组相比,瘦素中和抗体处理后可以抑制肿瘤细胞中瘦素下游信号通路的激活。该研究表明,前脂肪细胞和成熟脂肪细胞均能促进乳腺癌MDA-MB-231细胞的增殖和迁移,且这一促进作用和瘦素信号通路有关。
To investigate the effect of preadipocytes and mature adipocytes on the proliferation and migration of breast cancer MDA-MB-231 cells in breast cancer microenvironment, the study induced the preadipocyte 3T3-L1 to mature adipocytes and then preadipocytes And mature adipocytes were co-cultured with breast cancer MDA-MB-231 cells respectively. Morphological changes, proliferation and migration of tumor cells were observed by microscope imaging, oil red O staining, MTT assay and Transwell assay. Western blot and ELISA were used to detect the expression of leptin in preadipocytes and mature adipocytes. Immunofluorescence and Western blot were used to detect the expression of leptin receptor (leptin receptor, Ob-R), key molecules of leptin signaling pathway and downstream target genes in tumor cells respectively. The results showed that after co-culture, the morphology of tumor cells became more slender, the proliferation ability increased (P <0.05), and the number of cells passing through the cell increased significantly (P <0.05). Significant lipid accumulation also occurred in the tumor cells of the mature adipocyte co-culture group. Western blot and ELISA showed that both preadipocytes and mature adipocytes had leptin expression. Compared with the blank control group, the protein levels of p-Akt, p-STAT3, cyclin D1 and MMP9 in the two co-cultured groups were significantly increased (P <0.05), while p-ERK1 / 2 only in preadipocytes Co-culture group increased (P <0.001), in the mature adipocyte co-culture group did not change significantly. Compared with the co-culture group, leptin neutralizing antibody treatment can inhibit the activation of leptin downstream signaling pathway in tumor cells. The study shows that both preadipocytes and mature adipocytes can promote the proliferation and migration of breast cancer MDA-MB-231 cells, and this promotion is related to the leptin signaling pathway.