| Background::Costimulatory molecule B7-H3(B7 homolog 3,B7-H3)is a widely concerned immune checkpoint molecule involved in tumor progression.At present,it has been found that B7-H3 is involved in many tumor biological processes such as immune escape and glycolysis in colorectal cancer(Colorectal cancer,CRC),but the molecular mechanism of B7-H3 regulating angiogenesis and metastasis of CRC remains to be further studied.More and more evidence indicates that tumor-derived exosomes can participate in angiogenesis and tumor metastasis by affecting the functional changes of endothelial cells in the microenvironment.Therefore,the role of exocrine transport-related molecules in tumor microenvironment affecting the progression of colorectal cancer is an important direction in the diagnosis and treatment of CRC.Methods:Collect 55 CRC tissue samples and 6 CRC patients’ urine samples combined with high-throughput open data set of colorectal cancer to comprehensively analyze the correlation between B7-H3 expression and CRC progression.The expression of B7-H3 protein in exosomes derived from colorectal cancer cells was detected by Western blot,TEM,NTA and PKH67 fluorescence tracing,which acted on HUVECs cells.A colorectal cancer cell model with stable overexpression of B7-H3 was constructed,and the exosomes were collected.The effects of B7-H3 on HUVECs cells and the progression of intestinal cancer cells were analyzed by MTT,scratching,invasion and migration and angiogenesis.The effect of B7-H3 on HUVECs through VEGFA/VEGFR2/AKT1/m TOR pathway was detected by Western blot,and the exosome combined with AKT1 pathway inhibitor(MK-2206)was used to explore the possible mechanism of B7-H3 in exosome affecting the progression of colorectal cancer cells.An in vivo model was established to verify the effect of exocrine carrying B7-H3 on CRC metastasis.Results:Immunohistochemical analysis of B7-H3 in colorectal cancer tissue and highthroughput open data set analysis of colorectal cancer showed that B7-H3 was significantly overexpressed in CRC tissue,and positively correlated with the expression of angiogenesis marker molecule CD31.Compared with healthy controls,the expression of B7-H3 in urine samples of patients with CRC was significantly higher than that in exocrine bodies.The supernatant of overexpression B7-H3(oe B7-H3)HCT116 cells could significantly upregulate the expression of B7-H3 in HUVECs protein.In vitro,it was found that the exosomes derived from oe B7-H3 HCT116 could be uptake by HUVECs and significantly increase the level of B7-H3 protein in HUVECs.Functionally,there was no significant difference in the effect of oe B7-H3-exo on endothelial cell proliferation measured by MTT,while cell scratch test and Transwell test showed that exocrine B7-H3 could significantly improve the ability of endothelial cell migration,invasion and angiogenesis.Oe B7-H3-exo can promote the protein level of VEGFA secreted by HUVECs.In mechanism,intestinal cancer-derived exosomes transfer B7-H3 to promote the activation of VEGFR2/Akt1/m TOR signal pathway of HUVECs,and the use of Akt1 inhibitor MK-2206 can inhibit the activation of HUVECs and the ability of migration,invasion and angiogenesis.In vivo studies have shown that cancer-derived exosomes carry B7-H3 to promote angiogenesis and lung metastasis in colorectal cancer microenvironment.Conclusion: In this study,we found that B7-H3 secreted by CRC can be transferred into vascular endothelial cells through exosomes,and oe B7-H3-exo can promote angiogenesis of CRC by activating VEGFA/VEGFR2 and Akt1/m TOR signaling pathways.In vivo,experiments have shown that oe B7-H3-exo can promote the transfer of CRC.To sum up,this study proved that exocrine B7-H3 plays an important role in angiogenesis and metastasis of colorectal cancer. |
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