| Objective: The expression of platelet-derived growth factor(PDGF)-B facilitates the pericytes recruitment into blood vessels, which may be involved the stability of tumor blood vessels. As pericyte coverage may lead to resistant to antiangiogenic drugs, combined therapy targeting both endothelial cells and pericytes has been proposed. However, the interaction between pericytes and endothelial cells in regulating the stability, remodeling and function of angiogenesis in tumors is complicated. In this study, we aimed to evaluate the functions of pericyte recruitment induced by PDGF-B on tumor growth and progression in renal cell carcinoma(RCC).Methods: 1.ACHN cells, one of human kidney cancer cell lines, were transfected with a plasmid encoding PDGF-B gene and the expression of PDGF-B was verified by Western-blotting. 2.Proliferation analysis of transfected cells was performed by MTT assay. 3.Western-blotting analysis was used to gauge the phosphorylation PDGFR-β(p-PDGFR-β) expression level of VSMCs and HUVECs cells. 4.The effects of PDGF-B secreted by ACHN cells on VSMCs and HUVECs function in vitro were assessed by a variety of assays. FCM analysis was performed to detect the cell-cycle of VSMCs and HUVECs. Transwell cell culture chambers and tube formation assay were used to assess the migration and tube formation capability of VSMCs and HUVECs respectively. 5.PDGF-B-transfected and mock-transfected ACHN cells were injected onto the left groin of mice and the size of ACHN tumors were compared by measuring tumor diameter every 3 days. 6.Tumor cells proliferation and progression were assessed using immunohistochemistry and histomorphology. 7.Microarray analysis was performed to investigate the PDGF-B expression of cc RCC in the Van Andel Research Institute.Results: 1.Western-blotting analysis showed that, the expression of PDGF-B is up-regulated in ACHN cells transfected with PDGF-B gene. 2.MTT assay showed that there was no significant difference of OD value between PDGF-B-transfected and mock-transfected ACHN cells(P>0.05). 3.Up-expression of p-PDGFR-β was observed in VSMCs cultured in PDGF-B-overexpression conditioned medium. 4.In PDGF-B-overexpression group, FCM analysis showed that more VSMCs tend to be distributed into S and G2/M phases of the cell cycle. The migration of VSMCs was significantly greater(P<0.001) than in the other two conditioned media. VSMCs had the capability of tube formation only in PDGF-B-overexpression group. 5.In vivo, the mean volume of the PDGF-B-over-expression tumors was less than the control ones(P= 0.028). 6.The enhanced proliferation of tumor cells induced by PDGF-B overexpression was confirmed by Immunohistochemisty stained for Ki-67(P=0.019). In PDGF-B-overexpression tumors, only one mouse was found to have invasion in the fibrofatty tissue around the tumor as the control group had more metastasis or invasion. 7.Microarray analysis showed a better disease-specific survival for high PDGF-B expression group than low PDGF-B expression group(P<0.001).Conclusion: 1. PDGF-B up-regulates the expression of PDGFR-β on pericytes and increases the pericytes proliferation, migration and tube formation capability in vitro via PDGF-B/PDGFR-β signaling pathways. 2. Our animal studies revealed that PDGF-B overexpression inhibits tumor growth and tumor cell proliferation in vivo. The PDGF-B secreted by tumor cells may contribute to the tumor blood vessel normalization and stabilization by pericytes recruitment, which leads to the decrease of tumor metastasis and invasion events. 3. Microarray analysis showed high expression of PDGF-B is associated with decreased risks of cancer-specific mortality. Our current study suggested that therapy targeting pericytes in tumor is contentious and the function of pericytes in RCC warrants further investigation. |
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