| It has been reported that chemokines and their receptors play important and complex roles in carcinogenesis and development of malignant tumors. Some kinds of chemokines can promote tumor growth and metastasis by accelerating proliferation and migration of tumor cells or angiogenesis in tumor tissue. However, some may inhibit the tumor growth and angiogenesis, enhance innate or specific host immunity against tumor implantation. Therefore, it is of great significance to study the roles of chemokines and their receptors in the process of carcinogenesis, invasion and angiogenesis.This study includes four parts: ⑴ The differentiation state of three glioma cell lines, U87-MG, SHG-44 and CHG-5, was determined. This expression of CXCR4 in three glioma cell lines, ECV304 cells and human astrocytomas sections were also detected by immunohistochemistry. ⑵ The role of CXCR4 in spontaneous motility and chemotaxis was studied in the four cell lines. ⑶ [Ca2+] mobility and VEGF expression were measured in the four cell lines after CXCR4 combined with its ligand, SDF1. ⑷ Effect of Nordy on the cell growth, CXCR4 expression, spontaneous motility and chemotaxis in these cells were studied. The main results and conclusions are as the following:1. The expression of GFAP and vimentin in the three human glioma cell lines were detected by immunocytochemistry. The result showed that differences existed in GFAP and vimentin expression among U87-MG, SHG-44 and CHG-5 cells. U87-MG expressed the lowest level of GFAP but the highest level of vimentin, which suggested that the cell was poorly differentiated with a high malignancy. In contrast, CHG-5 cells expressed the highest level of GFAP, but the lowest level of vimentin. SHG-44 cell line was between them in differentiation.2. RT-PCR technique was utilized to detect the mRNA expression of chemokine receptors (CXCR4, CXCR2, CXCR1 and CCR2) in three glioma cell lines and ECV304 cell line. The result showed that CXCR4 was highly expressed, while CXCR2, CXCR1 and CCR2 were expressed at low level by these cells. The mRNA of CXCR4 gene in the three human glioma cell lines and ECV304 cells were most obvious among the four types of chemoattractant receptors. Immunocytochemical staining showed that CXCR4 was expressed in cultured glioma cell lines. U87-MG stained most heavily for CXCR4, CHG-5 was less. Moreover, ECV304 expressed CXCR4. The results suggested that CXCR4 might be an important receptor for chemokines on glioma cells and vascular endothelial cells.3. Immunohistochemistry for CXCR4 was performed on paraffin-embedded glioma sections for the expression in human brain glioma. The results showed that CXCR4 was expressed in the glioma cells and increased along with the histological grades. The endothelial cells in the glioma tissues also expressed CXCR4. These data suggested the expression of CXCR4 might be related to the invasive ability and angiogenesis of glioma.4. Chemotaxis assay and spontaneous motility assay demonstrated that SDF1 could increase the spontaneous motility and the chemotasis of glioma cells and ECV304. The migration of glioma cells and ECV304 cells induced by SDF1(30 ng/ml) could be inhibited by an anti-CXCR4 antibody(1 and 20μg/ml) in a dose-independent manner.5. Ca2+ mobility and VEGF expression in glioma and enthelial cells activated by SDF1's binding to CXCR4 were measured by laser confocal microscopy and immunocytochemistry. The result showed that SDF1 could trigger the increase of [Ca2+]i and VEGF expression. The results suggested that the CXCR4 on glioma cells and ECV304 was functional and it might play an important role in promoting rapid glioma cell growth and angiogenesis. 6. After U87-MG, SHG-44, CHG-5 and ECV304 cells being treated with Nordy, their proliferation rates decreased obviously. The expressions of CXCR4 in U-87, SHG-44 and CHG-5 were reduced after treatment. Nordy significantly inhibited the chemotactic migration of glioma cells induced by SDF1. Nordy could induce a transient increase [Ca2+]i in these cells. These results m...
|
PDF Full Text Request