The Effect Of Transforming Growth Factor-β1 In The Transformation Of Human Gastric Mucosa Fibroblasts And Its Expression In Gastric Cancer Tissues And Serum

Objective:In order to explore the relationship between the TGF-β1concentration in the culture medium and the transformation of primary gastric mucosal fibroblast;to study the expression level and clinical significance of TGF-β1in gastric cancer tissues and serum under different TNM staging.Methods:（1）Detected the expression of Desmin,Vimentin,TSP-1,FAP-a,Collagen III and Collagen I in primary cultured HGMF cells by flow cytometry.Treated the primary HGMF cells with different TGF-β1 concentrations as 5pg/ml,10pg/ml and 15pg/ml respectively and collected different generations of cells to detect:Cellular ultrastructure（with transmission electron microscope）,cell growth curve（by counting method）and cellular total protein content（by BSA assay）.（2）Detected and analyzed TGF-β1 expression in 40 cases of paraffin sections of gastric cancer by immunohistochemical SP method as well as in serum by ELISA.Results:（1）The expression of Desmin（negative）,Vimentin（positive）,TSP-1（negative）,FAP-a（negative）,Collagen III（positive）and Collagen I（positive）in primary cultured HGMF cells.The cytoplasma mitochondria of B2,B3,C2,C3 cells showed swelling,increased and other morphological changes under the electron microscopy.No significant difference appeared in the 8^th-generation cells.The growth rate of C2was significantly faster than that of B2 and P2（P<0.01）of 12^th-generation cells,while there was no significant difference between B2 and P2.In the 16^th-generation cells,the growth rate of both C3 and B3 were significantly faster than that of P3（P<0.01）,while there was no significant difference between C3 and B3.BSA assay results showed that the total protein contents（each generation couting 1×10⁶ cells）of12^th-generation B2,C2 and P2 cells were 0.3747±0.0308mg/ml,0.5292±0.0407mg/ml and 0.3697±0.0476mg/ml respectively（B2 vs.C2,P=0.000;B2 vs.P2,p=0.833;C2vs.P2,P=0.000）.The protein contents of 16^th-generation B3,C3 and P3 cells were respectively 0.5323±0.0413mg/ml,0.5415±0.0482mg/ml and 0.3704±0.0421mg/ml（B3 vs.P3,P=0.000;C3 and P3,P=0.000;B3 vs.C3,P=0.723）.（2）The positive rates of TGF-β1 in gastric cancer tissues showed as:with vs.without lymph node metastasis was 85%vs.45%,（p=0.008）;with vs.without distant metastases was100%vs.58.82%（p=0.137）.The positive rates of TGF-β1 in the tissues of different TNM stages was:T1 25%,T2 45.45%,T3 88.24%and T4 100%（p=0.000,comnbined T1T2 vs.combined T3T4）.The positive rates of TGF-β1 in poorly differentiated tissues,moderately differentiated tissues and highly differentiated tissues were 86.36%,36.36%,42.86%,respectively（p=0.002,poorly differentiated cases vs.combined moderately/highly differentiated cases）.The ELISA results of TGF-β1（pg/ml）expresseion in serum showed that:17.15±5.82 vs.10.81±4.34（with vs.without lymph node metastasis,P<0.01,t=4.22）;24.44±3.64 vs.12.43±4.50（with vs.without distant metastasis,P<0.01,t=6.172）.The ELISA results of TGF-β1（pg/ml）expresseion in serum in different TNM stages were:T1 8.91±3.04,T2 10.19±1.75,T3 17.11±5.14,T4 3.72±3.81（P<0.01,t=19.37）;poorly differentiated cases17.62±5.85,moderately differentiated cases 11.11±3.51,highly differentiated cases8.47±2.39（P<0.01,t=12.521）.The data indicated that there was an interaction between TGF-β1 positive rate in gastric cancer tissues and the increased expression of TGF-β1 in serum（P<0.01,t=7.094）.Conclusion:The transformation of primary culture cells could appeare at lower generation(such as 12^th generation)once TGF-β1 concentration in culture medium increased to 15pg/ml.Once the gastric cancer had lymph node metastasis,distant metastasis,the higher infiltration depth and the lower degree differentiation,the positive rate of TGF-β1 in tissues was significantly increased accompanied with the increased expression of TGF-β1 in serum.The up-regulation of TGF-β1 expression in both tissues and serum was closely related to the occurrence and the biological behaviors of gastric cancer.The expression of TGF-β1 can act as an important reference index to judge the clinical stages and differentiated degree of gastric cancer.