Study On The Interaction Between Malignant Transformation Of Human Pulmonary Epithelial Cells And Activation Of Fibroblasts Induced By Yunnan Tin Mine Dust And The Mechanisms

【Objectives】To study the interaction between transformation of human pulmonary epithelial cells and activation of fibroblasts induced by Yunnan tin mine dust and the mechanism.【Methods】(1)Immortalized human bronchial epithelial cells BEAS-2B and human embryonic pulmonary fibroblast cells WI-38 were treated with Yunnan tin mine dust for 72h on every other generation until the 9th generation.From the 11th generation,the cells were cocultured.The groups were divided into:B,W,B100,W100,B(W),B(W100),B100(W), B100(W100),W(B),W(B100),W100(B)and W100(B100)groups.(2)The cell morphology was observed with inverted microscopy.(3)The cell cycles were analyzed through flow cytometry.(4)The transformation of epithelial cells was identified by coagulation with ConA and anchorage independent growth in soft agar.(5)The expressions ofα-SMA in fibroblasts, FHIT and Ki-67 in epithelia were investigated by the method of immunocytochemistry.(6) The TGF-β1 level in the supernatant of epithelial cells was examined with ELISA.(7)The levels of FAP,TβRⅡ,HGF and Smad2/3 phosphorylation in fibroblasts and TβRⅡ,c-met phosphorylation in epithelia were detected through western blot.【Results】(1)The Tin mine dust could induce malignant transformation of human pulmonary epithelial cells BEAS-2B.The time of transformation of epithelia was shorted after cocultured with fibroblasts WI-38.(2)The expression ofα-SMA in fibroblast cells WI-38 was negative in the 16th generation.The expression ofα-SMA became positive in WI-38 cells treated with the mine dust in the 26th and 36th generations,especially after cocultured with fibroblasts.(3)The epithelial cells exhibited loss of FHIT protein expression in the early stage of transformation induced by the mine dust.The epithelial cells expressed FHIT protein again in the late stage of transformation.This molecular event could be promoted by the fibroblasts.(4)The expression of ki-67 was similar in different epithelial groups untreated with mine dust.The proliferation index of epithelia treated with mine dust was increased(P＜0.05).(5)The expression of FAP in fibroblast cells WI-38 was negative in the 16th generation.The expression of FAP became positive in WI-38 cells treated with the mine dust in the 26th and 36th generations,especially after cocultured with fibroblasts.(6) The content of TGF-β1 is elevated significantly in culture supernatant of mine dust-induced BEAS-2B cells,especially in the early transformation stage.And a relative consistent level is maintained following.(7)The expression of TβRⅡin the epithelial cells is decreased until lost during mine dust-induced malignant transformation.(8)The expression of TβRⅡin the fibroblasts does not change during mine dust-induced activation.(9)The levels of Smad2/3 phosphorylation in fibroblasts are increased after coclutured with epithelia treated with mine dust.(10)The expression of HGF protein in the fibroblasts in the developing process of induced lung cancer by Yunnan tin mine dust increased.And the expression of c-met phosphorylation increased in epithelial groups coclutured with fibroblasts.【Conclusions】(1)The Tin mine dust can induce malignant transformation of epithelia and activation of fibroblasts.(2)The malignant transformation of epithelia and activation of fibroblasts induced by Yunnan tin mine dust interact and co-evolve.(3)The epithelial cells are major target in carcinogenesis induced by Yunnan tin mine dust.(4)FHIT could be a target that Yunnan tin mine dust induced lung cancer,the decrease or loss of its protein expression counld be a valuable biologic marker to predict one risk of lung cancer.(5)The expression ofα-SMA and FAP in fibroblasts counld be a early molecular event in the developing process of induced lung cancer by Yunnan tin mine dust.(6)The role of TGF-β1 could be inhibition in early stage of carcinogenesis,but improvement in late stage.(7)The interaction between the malignant transformation of epithelial cells and activation of fibroblasts is related with TGF-β1/Smad and HGF/c-met signal pathway.