Objective： Although the kinase insert domain-containing receptor （KDR） gene play an very important role in the metastasis of cancer and is also as one of the molecular targets used in cancer therapy, mutation in th...Objective： Although the kinase insert domain-containing receptor （KDR） gene play an very important role in the metastasis of cancer and is also as one of the molecular targets used in cancer therapy, mutation in the tyrosine kinase （TK） domain of the KDR gene has not been reported. Here we detected the mutations and polymorphisms in the TK domain of KDR gene in human lung cancer patients and to give the basic evidence and clue for cancer prevention and target therapy. Methods： The entire sequence of exons 21, 22, 23 and 27 （which contain the coding sequence of tyrosine phosphorylation） in the TK domain of KDR gene in the patients with lung cancer and control healthy individuals were assayed by PCR and DNA sequencing. We also analyzed one non-coding single nucleotide polymorphisms （SNPs） in the KDR gene. Results： No mutations were found in exon 22, 23 and 27. One heterozygous mutation of c.＋2837 in exon 21 was found at a frequency of 2.08% （2/96） in the patients with lung cancer and none were detected in the healthy control individuals. The mutation was from a G to a A resulting in substitution of arginine with histidine residue. Conclusion： Our data suggested that we should focus on the mutation or SNP in the other regions or the expression levels of KDR gene, and the function of c.＋2837 mutation of KDR .qene may be needed further study in the future.展开更多
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:The aim of this study was to detect and localize the expression of the vascular endothelial growth factor receptors(Fms-like tyrosine kinase,Flt-1 and Kinase insert domain-containing receptor,KDR)in human ec...Objective:The aim of this study was to detect and localize the expression of the vascular endothelial growth factor receptors(Fms-like tyrosine kinase,Flt-1 and Kinase insert domain-containing receptor,KDR)in human ectopic and eutopic endometrium of patients with endometriosis.Methods:Endometrial biopsies from thirty three healthy and thirty seven endometriosis,thirty four ovarian chocolate cyst,fifteen red peritoneal lesions,and four endometriosis lesions form abdominal wall were used for immunohistochemistry,western blot and reverse transcript-polymerase chain reaction(RT-PCR)assessments to examine Flt-1 and KDR mRNA and protein expression for their relative level in the above tissues.Results:Flt-1 and KDR were expressed in the endometrial glandular epithelium and stroma cells besides the endometrial blood vessels.Among endometriotic lesions,the ectopic endometrium expressed higher level of Flt-1(94.3%)and KDR(91.4%)than ovarian endometrioma(74.3%,77.1%,P<0.05),and the same as eutopic endometriosis(93.8%,90.6%,P>0.05).Flt-1(mRNA level:2.41±1.21,protein level:31.1±17.4)and KDR(mRNA:3.02±1.42,protein level:35.6±23.6)in the eutopic endometrium expressed higher level than those in the ovarian endometrioma(mRNA level:1.51±0.92,1.83±1.03;protein level:16.8±6.1,19.7±10.5,P<0.05),and the same as those in the eutopic endometrium(mRNA level:1.91±0.83,2.26±1.27;protein level:24.2±17.8,25.0±16.5,P>0.05).Conclusions:VEGF may have some effects on the development of endometriosis by VEGFR(Flt-1 and KDR).The expressions of Flt-1 and KDR in endometriotic tissue appear to be associated with its neovascularization.展开更多
基金Supported by the grants from the National Natural Science Foundation of China (No. 30772531)Guangdong Provincial Medical Science and Technology Research Foundation (No. B2006001)China Postdoctoral Science Foundation (No. 20060400212)
文摘Objective： Although the kinase insert domain-containing receptor （KDR） gene play an very important role in the metastasis of cancer and is also as one of the molecular targets used in cancer therapy, mutation in the tyrosine kinase （TK） domain of the KDR gene has not been reported. Here we detected the mutations and polymorphisms in the TK domain of KDR gene in human lung cancer patients and to give the basic evidence and clue for cancer prevention and target therapy. Methods： The entire sequence of exons 21, 22, 23 and 27 （which contain the coding sequence of tyrosine phosphorylation） in the TK domain of KDR gene in the patients with lung cancer and control healthy individuals were assayed by PCR and DNA sequencing. We also analyzed one non-coding single nucleotide polymorphisms （SNPs） in the KDR gene. Results： No mutations were found in exon 22, 23 and 27. One heterozygous mutation of c.＋2837 in exon 21 was found at a frequency of 2.08% （2/96） in the patients with lung cancer and none were detected in the healthy control individuals. The mutation was from a G to a A resulting in substitution of arginine with histidine residue. Conclusion： Our data suggested that we should focus on the mutation or SNP in the other regions or the expression levels of KDR gene, and the function of c.＋2837 mutation of KDR .qene may be needed further study in the future.
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:The aim of this study was to detect and localize the expression of the vascular endothelial growth factor receptors(Fms-like tyrosine kinase,Flt-1 and Kinase insert domain-containing receptor,KDR)in human ectopic and eutopic endometrium of patients with endometriosis.Methods:Endometrial biopsies from thirty three healthy and thirty seven endometriosis,thirty four ovarian chocolate cyst,fifteen red peritoneal lesions,and four endometriosis lesions form abdominal wall were used for immunohistochemistry,western blot and reverse transcript-polymerase chain reaction(RT-PCR)assessments to examine Flt-1 and KDR mRNA and protein expression for their relative level in the above tissues.Results:Flt-1 and KDR were expressed in the endometrial glandular epithelium and stroma cells besides the endometrial blood vessels.Among endometriotic lesions,the ectopic endometrium expressed higher level of Flt-1(94.3%)and KDR(91.4%)than ovarian endometrioma(74.3%,77.1%,P<0.05),and the same as eutopic endometriosis(93.8%,90.6%,P>0.05).Flt-1(mRNA level:2.41±1.21,protein level:31.1±17.4)and KDR(mRNA:3.02±1.42,protein level:35.6±23.6)in the eutopic endometrium expressed higher level than those in the ovarian endometrioma(mRNA level:1.51±0.92,1.83±1.03;protein level:16.8±6.1,19.7±10.5,P<0.05),and the same as those in the eutopic endometrium(mRNA level:1.91±0.83,2.26±1.27;protein level:24.2±17.8,25.0±16.5,P>0.05).Conclusions:VEGF may have some effects on the development of endometriosis by VEGFR(Flt-1 and KDR).The expressions of Flt-1 and KDR in endometriotic tissue appear to be associated with its neovascularization.
