Study About The Mechanism Of Connexin43 Mediating The Formation Of Gap Junction Promotes The Cancer Proliferation In 3D Model

Gap junction is a protein channel that communicate the cytoplasm of adjacent cells,allowing small molecules of less than 1KDa to transfer between cells.The gap junction is composed of connexin(Cx),and Cx43 is the major isoform expressed on the intestinal epithelium.The traditional view is that connexin protein has inhibitory effect on tumor,while recent studies show that connexin has different roles in different tumors and stages,and its function of promoting or suppressing cancer requires further elucidation.The role of connexin proteins and gap junctions in colon cancer are poorly reported.Based on the analysis of TCGA cancer database,we find that the prognosis of the patients with high expression Cx43 is worse,suggesting that the connexin protein could promote colon tumorigenesis and development.However,under the condition of 2D culture,overexpression of Cx43 has no effect on the proliferation of intestinal cancer cells.We imitate the growing conditions of tumor in vivo by cultivating the tumor cells in 3D state,and surprisingly find that the expression Cx43 promotes the proliferation of intestinal cancer cells.However,neither over expression Cx43 mutant T154A,which cannot form the full channel or the hemichannel,nor the Cx43 mutant R76W which can form hemichannel but not full channel,has any effects on proliferation.Furthermore,the promoting effect of Cx43can be inhibited by using gap junction channel inhibitor CBX.These results suggest that Cx43 promotes tumor proliferation in the 3D state,which relies on the function of its gap junction channel.In order to further study the mechanism involved,we stain the 3D cell spheres with Ki67 and Tunel,and the results show that the central necrotic cells in the Cx43 group are significantly less than those of the control group,and the region of proliferation is more than the control groups.In addition,subcutaneous tumor forming in the nude mice assay confirms that the proliferation rate of Cx43 group is higher than other groups,and the biopsy results suggest that the necrosis of colon cancer cells after expression of Cx43 is significantly lower than those in the rest groups.These results suggest that Cx43 in colon cancer cells can help the cells in the center of the tumor to obtain nutrients and promotes the proliferation of colon cancer by forming a gap junction.In order to reveal what substance is passed through the gap junction of colon cancer cells,we use fluorescent glucose analogues and find that the Cx43 group takes in more glucose than others,suggesting that colon tumor may transfer glucose and other nutrients to the central cells by gap junction,thus promote the proliferation of cancer cells.This study reveals the new role of Cx43 and gap junction in the development of colon cancer,and provides a theoretical basis for the development of Cx43-targeted colon cancer therapy.