Vascular endothelial growth factors（VEGFs） respectively bind to each of three receptor tyrosine kinases（RTKs）,known as Flt-1,KDR and Flt-4.Since VEGFs and their respective families of receptor tyrosine kinases（VE...Vascular endothelial growth factors（VEGFs） respectively bind to each of three receptor tyrosine kinases（RTKs）,known as Flt-1,KDR and Flt-4.Since VEGFs and their respective families of receptor tyrosine kinases（VEGFRs） are critical proteins which can regulate vascular development during angiogenesis,we decided to explore the inhibitory effects of soluble kinase insert domain-containing receptor（sKDR） on endothelial cells and angiogenesis.Total RNA was extracted from human umbilical vein endothelial cells（HUVEC）,and cDNA of extracellular domains 1―4 was amplified and recombined with pQE40 vector.After being expressed,affinity purified,renatured and analyzed by Western blot,the sKDR was assayed for its effects on endothelial cells by [3-（4,5-dimethylthiazol-2-yl）2,5-diphenyltetrazolium bromide]（MTT）,and on angiogenesis by chick chorioallantoic membrane（CAM） experiment.sKDR cDNA of 1150 bp was obtained via real-time polymerase chain reaction（RT-PCR）,and sKDR was expressed by pQE40 procaryotic expression system,purified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis（SDS-PAGE） analysis with only one band and proved by Western blot.MTT assay demonstrateds that sKDR could inhibit the VEGF-stimulated HUVEC from proliferation,and CAM experiment showed sKDR could block the VEGF-induced angiogenesis.sKDR has the biological activity to bind with VEGF ligands and is a potential target for tumor anti-angiogenesis therapy.展开更多
Objective: To detect the expression of KDR in different originated carcinomas and to explore its expressed ways and the relationship with tumor progression. Methods : KDR cDNA (V VII domains) fragment wa...Objective: To detect the expression of KDR in different originated carcinomas and to explore its expressed ways and the relationship with tumor progression. Methods : KDR cDNA (V VII domains) fragment was cloned from human umbilical vein with RT PCR and was expressed in Ecoli.Jm109. The fusion protein of GST KDR was used for immunizing Balb/c mice to prepare monoclonal antibodies against KDR. The different tumor tissues and related normal tissues were examined with KDR McAb by S P immuno histochemistry. Results: the rate and intensification of KDR expression among different originated cancers are very different, bladder cancers from transmigrated epidermis are 100% positive and highest intensification. The expression of KDR in breast cancer and intestinal cancer lie in the second rate, the weakest expression of KDR is in lung squamous carcinoma. Moreover, expression of KDR in tumor tissues lie both in endothelial cells (EC) of tumor blood vessels and tumor cells. Conclusion: VEGF may be not only the para secretory factor making EC proliferation but also auto secretory factor stimulating the proliferation of tumor cells to benefit the growth and metastasis of malignant tumors. The different expression of KDR in different originated carcinomas may relate with malignant degree of tumor.展开更多
文摘Vascular endothelial growth factors（VEGFs） respectively bind to each of three receptor tyrosine kinases（RTKs）,known as Flt-1,KDR and Flt-4.Since VEGFs and their respective families of receptor tyrosine kinases（VEGFRs） are critical proteins which can regulate vascular development during angiogenesis,we decided to explore the inhibitory effects of soluble kinase insert domain-containing receptor（sKDR） on endothelial cells and angiogenesis.Total RNA was extracted from human umbilical vein endothelial cells（HUVEC）,and cDNA of extracellular domains 1―4 was amplified and recombined with pQE40 vector.After being expressed,affinity purified,renatured and analyzed by Western blot,the sKDR was assayed for its effects on endothelial cells by [3-（4,5-dimethylthiazol-2-yl）2,5-diphenyltetrazolium bromide]（MTT）,and on angiogenesis by chick chorioallantoic membrane（CAM） experiment.sKDR cDNA of 1150 bp was obtained via real-time polymerase chain reaction（RT-PCR）,and sKDR was expressed by pQE40 procaryotic expression system,purified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis（SDS-PAGE） analysis with only one band and proved by Western blot.MTT assay demonstrateds that sKDR could inhibit the VEGF-stimulated HUVEC from proliferation,and CAM experiment showed sKDR could block the VEGF-induced angiogenesis.sKDR has the biological activity to bind with VEGF ligands and is a potential target for tumor anti-angiogenesis therapy.
文摘Objective: To detect the expression of KDR in different originated carcinomas and to explore its expressed ways and the relationship with tumor progression. Methods : KDR cDNA (V VII domains) fragment was cloned from human umbilical vein with RT PCR and was expressed in Ecoli.Jm109. The fusion protein of GST KDR was used for immunizing Balb/c mice to prepare monoclonal antibodies against KDR. The different tumor tissues and related normal tissues were examined with KDR McAb by S P immuno histochemistry. Results: the rate and intensification of KDR expression among different originated cancers are very different, bladder cancers from transmigrated epidermis are 100% positive and highest intensification. The expression of KDR in breast cancer and intestinal cancer lie in the second rate, the weakest expression of KDR is in lung squamous carcinoma. Moreover, expression of KDR in tumor tissues lie both in endothelial cells (EC) of tumor blood vessels and tumor cells. Conclusion: VEGF may be not only the para secretory factor making EC proliferation but also auto secretory factor stimulating the proliferation of tumor cells to benefit the growth and metastasis of malignant tumors. The different expression of KDR in different originated carcinomas may relate with malignant degree of tumor.
