Preliminary Study On COL11A1 Promotes Tumorigenesis,Metastasis And Related Mechanisms Of Colon Cancer In Nude Mice

Objective:To explore the role of type XI collagen α1 chain(COL11A1)in the tumorigenesis and metastasis of colon cancer in nude mice,and to explore the regulation of COL11A1 expression by TGF-β 1 / Smad2 pathwayMethods:The cell line HCT116 with strong invasiveness and high expression of COL11A1 content was screened from colon cancer cells HCT116 and SW480 by protein immunoblotting(Western Blot),real-time fluorescence quantitative PCR(RT-PCR),cell migration,invasion and proliferation in vitro.Lentivirus was transfected into HCT116 cells to construct stable transformants of silencing / overexpressing COL11A1.According to the groups of NC group(saline group),no load group,ov-COL11A1 group(overexpression COL11A1 group)and sh-COL11A1 group(silent COL11A1 group),colon cancer subcutaneous transplantation tumor and caudal vein pulmonary metastasis model were established by selecting the third / fourth generation stable transplant strain in HCT116 group.There were 12 Balb/cu nude mice with 4-6 weeks old in each group.0.1ml2x106HCT116 cell suspension was injected subcutaneously into the right anterior axilla and caudal vein,and the animals were killed after 30 days and 45 days of anesthesia,respectively.the role of COL11A1 in colon cancer tumorigenesis and metastasis in nude mice was analyzed by RT-PCR,Western Blot and HE staining.HCT116 group was treated with TGF-β 1stimulant and TGF-β R1 receptor inhibitor LY2157299 respectively.The expression of p-Smad2 and COL11A1 and the changes of EMT related proteins E-cadherin and FAP-a were detected by Western blotting.The invasion and migration ability of HCT116 cells were detected by Transwell migration test and invasion test in vitro.Results:1.Western Blot,q RT-PCR and in vitro cell experiments showed that compared with SW480,the expression of COL11A1 in HCT116 was higher,and its proliferation,migration and invasion were stronger.2.Lentivirus transfection and successful construction of stable transgenic strain HCT116,ov-COL11A1 with silencing /overexpression of COL11A1 could significantly promote subcutaneous tumorigenesis and lung metastasis of colon cancer cell line HCT116 in nude mice.On the contrary,sh-COL11A1 could inhibit subcutaneous tumorigenesis and lung metastasis.It indirectly reflects that COL11A1 can promote the proliferation of colon cancer cell HCT116 and lung metastasis of II of Guizhou Medical University.3.After the stimulation of HCT116 cells with TGF-β1,the expression of COL11A1 was up-regulated and the phosphorylation process was promoted,and the EMT-related protein-epithelial marker E-cadherin was down-regulated and the stromal marker FAP-a was up-regulated.On the contrary,the phosphorylation process was blocked by the addition of,TGF-βR1 receptor inhibitor LY2157299,TGF-βR1 and COL11A1.EMT-related protein-epithelial marker E-cadherin was up-regulated and interstitial marker FAP-a was down-regulated.Conclusions:1.COL11A1 promotes tumorigenesis and Metastasis of Colon Cancer in Nude mice2.COL11A1 may induce EMT process by participating in the activation of TGF-β1/Smad2 signal pathway,thus affecting the growth,invasion and metastasis of colorectal cancer cells.