The Biological Function Of B7-H3 On Cancer-Associated Fibroblasts In Promoting The Progression Of Renal Cancer

The development of malignant tumors is not only an independent event of oncogene mutation and overgrowth,but also the result of interaction between malignant tumors and the surrounding tumor matrix microenvironment.The tumor microenvironment is a complex ecosystem.The microenvironment,high pressure,hypoxia,acidosis,and a large number of cytokines,chemokines,enzyme molecules and immunoinflammatory reactions constitute the biological characteristics of the metabolic environment of malignant tumor tissues.This feature together promotes immune escape,growth and metastasis of malignant tumors.Tumor microenvironment contains a large number of cancer-associated fibroblasts.Cancer-associated fibroblasts are fibroblasts in the tumor stroma."Tumors are like wounds that never heal." In the tumor microenvironment,activated fibroblasts are in a state of continuous activation,synthesizing and secreting large amounts of extracellular matrix proteins,metalloproteinases,cytokines,and chemokines.Tumor cell proliferation and invasion,remodeling of extracellular matrix,recruitment of immune cells,and production of new blood vessels.The B7-H3(CD276)molecule has become a research hotspot after being first discovered by Chapoval et al.in 2001.So far,B7-H3 molecules have been extensively studied in many types of tumor cells,and have made a gratifying result.These studies indicate that B7-H3 molecules play a key role in promoting cancer progression.Recently,the role of B7-H3 signaling in cell differentiation,invasion and metastasis and anti-apoptosis has gradually become a hot topic in current research.It has also been reported in the literature that B7-H3 signaling can significantly resist apoptosis induced by chemotherapeutic drugs,and it is also confirmed that B7-H3 does have anti-apoptotic effects.For this reason,we used shRNA-mediated gene silencing to study the function and signaling effects of B7-H3 molecules on tumor-associated fibroblast proliferation and survival,and observe the changes in biological functions of CAFs.The impact of cancer progression.Our in vitro analysis showed that decreased expression of B7-H3 could induce apoptosis of cancer-associated fibroblasts and arrest the cell cycle,which ultimately led to a decrease in the invasion and metastasis of renal cancer cells(A498).Part ? Establishment of cancer-associated fibroblasts inhibiting B7-H3 gene expression and its biological functionAbstract:To investigate the effect of B7-H3 on proliferation and survival of cancer-associated fibroblasts(CAFs).Method:(1)The expression of B7-H3 molecular protein on CAFs was detected by flow cytometry.(2)The cancer-associated fibroblasts were transfected with lentivirus B7-H3 targeting shRNA and identified by western blot and qPCR.(3)The gene chip method was used to screen the differentially expressed genes between the two groups.(4)The proliferation differences of CAFs-shB7-H3 and CAFs-NC cells were determined by CCK8 assay.(5)Apoptosis and cell cycle changes were detected by flow assay after Annexin V-PE/PI double staining.(6)The expression of apoptosis related molecules was detected by Western blot.Result:(1)High expression of B7-H3 protein in cancer-associated fibroblasts.(2)The stable expression of CAFs-shRNA in cell lines and its control cell lines were obtained(CAFs-NC).(3)B7-H3 signaling can lead to up-regulation of up to 4,531 genes and down-regulation of 3,677 genes.(4)The growth rate of cancer-associated fibroblasts decreased significantly after B7-H3 knockdown.(5)The percentage of early apoptosis cells of CAFs-shRNA was higher than that of CAFs-NC,but the difference of apoptosis rates between the two groups was not statistically significant,CAFs-shRNA cells are blocked during the G0/G1 phase of the cell cycle.(6)Bax,Caspase3 expression was up-regulated in CAFs-shRNA cells,Bcl-2,P-AKT,HGF,CXCL-12 expression was down-regulated and total AKT was not significantly changed.Conclusion:Knockdown of B7-H3 inhibited the proliferation of cancer-associated fibroblasts,promoted cell blocking at G0/G1 phase,and induced apoptosis.Part ? B7-H3 on cancer-associated fibroblasts promotes kidney Biological function of cancer progressionAbstract:To investigate the key role of B7-H3 on cancer-associated fibroblasts(CAFs)in the occurrence and metastasis of renal cancer.Method:(1)CAFs-shRNA and CAFs-NC cells co-cultured with renal cancer cells(A498),The effect of on the proliferation of renal cancer cells was determined by CCK8 assay.(2)The effects of CAFs-shRNA and CAFs-NC on the migration and invasion of A498 cells were evaluated by Transwell assay.(3)Western blot analysis of HGF and CXCL-12 protein expression.Result:(1)A498 cells in CAFs-NC group showed significantly higher activity than those in CAFs-shRNA group.(2)The ability of CAF-shRNA cells to migrate and invade A498 was significantly lower than that of CAFs-NC group.(3)Compared with the control group,HGF and CXCL-12 protein expressions were decreased.conclusion:B7-H3 on cancer-associated fibroblasts has a strong promoting effect on the growth and metastasis of renal tumors.