CAJ | 학술논문

目的：观察转化生长因子β1（TGF-β1）对腹膜间皮细胞上皮-间质表型转化的调控及对胃癌细胞腹膜转移的影响。方法 TGF-β1作用人腹膜间皮细胞系HMrSV5，倒置显微镜下观察间皮细胞形态学变化。采用Western-blot检测上皮细胞表型蛋白（细胞角蛋白和E-钙黏素）、间皮细胞表型蛋白（α-SMA和弹性蛋白）及Smad2蛋白水平的表达情况。采用黏附试验观察表型转化间皮细胞对胃癌细胞系HSC-39黏附的影响。间皮细胞（8×104／孔）与胃癌细胞系HSC-39（4×104／孔）共培养，采用体外Transwell侵袭实验观察腹膜微环境变化对胃癌细胞侵袭能力的影响。结果 TGF-β1作用腹膜间皮细胞24 h后部分细胞转变为狭长型，72 h后间皮细胞转变为典型纤维细胞样外观。TGF-β1作用间皮细胞可诱导弹性蛋白和α-SMA表达上调，细胞角蛋白和 E-钙黏素表达下降，并且呈现时间依赖性变化（P＜0．05）。 TGF-β1作用15 min后，间皮细胞内磷酸化Smad2表达开始升高，30 min达到顶峰，较对照组增加432％（P＜0．01）；但总Smad2表达则无明显变化（P＞0．05）。 TGF-β1作用72 h后，间皮细胞与HSC-39胃癌细胞的黏附率较对照组增加（146±17）％（P＜0．05）。胃癌细胞与TGF-β1刺激的间皮细胞共培养48 h后，平均每视野转移癌细胞数目为61．1±11．4，较对照组（31．9±8．1）明显增多（P＜0．05）。结论 TGF-β1能够诱导间皮细胞向成纤维细胞样转化，Smad2信号转导通路在间皮细胞表型转化中发挥重要作用；而这种腹膜微环境变化可增强胃癌细胞的黏附和侵袭能力，为癌细胞转移播散提供适宜的“土壤”环境。
목적：관찰전화생장인자β1（TGF-β1）대복막간피세포상피-간질표형전화적조공급대위암세포복막전이적영향。방법 TGF-β1작용인복막간피세포계HMrSV5，도치현미경하관찰간피세포형태학변화。채용Western-blot검측상피세포표형단백（세포각단백화E-개점소）、간피세포표형단백（α-SMA화탄성단백）급Smad2단백수평적표체정황。채용점부시험관찰표형전화간피세포대위암세포계HSC-39점부적영향。간피세포（8×104／공）여위암세포계HSC-39（4×104／공）공배양，채용체외Transwell침습실험관찰복막미배경변화대위암세포침습능력적영향。결과 TGF-β1작용복막간피세포24 h후부분세포전변위협장형，72 h후간피세포전변위전형섬유세포양외관。TGF-β1작용간피세포가유도탄성단백화α-SMA표체상조，세포각단백화 E-개점소표체하강，병차정현시간의뢰성변화（P＜0．05）。 TGF-β1작용15 min후，간피세포내린산화Smad2표체개시승고，30 min체도정봉，교대조조증가432％（P＜0．01）；단총Smad2표체칙무명현변화（P＞0．05）。 TGF-β1작용72 h후，간피세포여HSC-39위암세포적점부솔교대조조증가（146±17）％（P＜0．05）。위암세포여TGF-β1자격적간피세포공배양48 h후，평균매시야전이암세포수목위61．1±11．4，교대조조（31．9±8．1）명현증다（P＜0．05）。결론 TGF-β1능구유도간피세포향성섬유세포양전화，Smad2신호전도통로재간피세포표형전화중발휘중요작용；이저충복막미배경변화가증강위암세포적점부화침습능력，위암세포전이파산제공괄의적“토양”배경。
Objective To elucidate the role of transforming growth factor-beta1 (TGF-β1) in epithelial-mesenchymal transition of mesothelial cells and peritoneal metastasis of gastric cancer. Methods HMrSV5 cells, a human peritoneal mesothelial cell line, were incubated with TGF-β1, and their morphological changes were observed by phase contrast microscopy. Expressions of α-smooth muscle actin (α-SMA), vimentin, cytokeratin, E-cadherin, phosphorylated-Smad2 and Smad2 were examined by Western blotting. After fibroblastic-like mesothelial cells were co-incubate with HSC-39 cells (gastric cancer cell line), the adhesion and invasion potential of HSC-39 were evaluated by adhesion and invasion assay in vitro. Results Few mesothelial cells converted to spindle fibroblast-like morphology for 24 h, and remarkable phenotypic changes were observed at 72 h of TGF-β1 activation. TGF-β1 could induce α-SMA and vimentin expression, and down-regulate cytokeratin and E-cadherin expression in mesothelial cells (P<0.05). TGF-β1 induced phosphorylation of Smad2 within 15 min of stimulation, reached a maximum at 30 min after treatment and remained high level during the experiment without affecting total Smad2 expression (P>0.05). The percentage of HSC-39 gastric cancer cells adhered were significantly increased as compared to the control. When the mesothelial cells were treated by TGF-β1 for 72 h, the increased adhesion percentage was (146 ±17)%(P<0.05). After fibroblastic-like mesothelial cells co-incubated with HSC-39 cells for 48 h , more cancer cells [(61.1 ±11.4) cells/view field] invaded the coated membrane as compared to the control group [(31.9±8.1) cells/view field] (P<0.05). Conclusion TGF-β1 can induce the transition of mesothelial cells into myofibroblasts and Smad2 signal pathway may play a role in this transition , which is associated with increased adhesion and invasiveness of gastric cancer cells , and provides favorable environment for the dissemination of gastric cancer.