siRNA干扰PDGFRβ促进胃癌MGC-803/VCR细胞的凋亡
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摘要
为研究血小板衍生生长因子受体β(PDGFRβ)在胃癌组织和长春新碱(vincristine, VCR)耐药胃癌细胞MGC-803/VCR中的表达,并探讨PDGFRβ沉默对MGC-803/VCR细胞增殖和凋亡的影响,本研究分别通过免疫组化和蛋白质印迹法(Western blotting)检测PDGFRβ蛋白在胃癌组织和耐药细胞株中的表达水平。通过Lipofectamine 2000将PDGFRβ小干扰RNA (si-PDGFRβ)转染到MGC-803/VCR细胞,Western blotting检测转染后PDGFRβ蛋白表达水平,Cell Counting Kit-8 (CCK-8)和流式细胞术分别检测si-PDGFRβ对VCR诱导人胃癌MGC-803细胞增殖和凋亡的影响。研究结果显示:PDGFRβ蛋白在胃癌组织中的表达显著高于正常胃组织;PDGFRβ蛋白在MGC-803/VCR细胞中的表达极显著高于MGC-803细胞,并且转染si-PDGFRβ后MGC-803/VCR细胞的PDGFRβ蛋白表达水平显著降低;1μmol/L、2μmol/L、4μmol/L、8μmol/L的VCR诱导MGC-803/VCR细胞后,si-PDGFRβ组细胞增殖抑制率分别为(21.97±0.84)%、(37.63±1.32)%、(55.77±1.39)%和(72.17±1.16)%,与对照组的(13.60±0.49)%、(22.33±1.01)%、(38.30±1.56)%和(52.90±1.08)%分别比较有极显著的差异(p<0.01);流式检测结果显示,与对照组的细胞凋亡率(13.61±0.49)%比较,发现si-PDGFRβ组胃癌MGC-803/VCR细胞凋亡率为(29.80±0.64)%,说明两者差异极显著(p<0.01)。本研究初步结论表明,si RNA干扰PDGFRβ能够促进VCR诱导的胃癌MGC-803/VCR细胞凋亡。
In order to study the expression of platelet derived growth factor receptor β(PDGFRβ) in gastric cancer tissues and vincristine(VCR) resistance gastric cancer cells MGC-803/VCR and investigate the effect of PDGFRβsilence on proliferation and apoptosis of MGC-803/VCR cells, in this research, the expression levels of PDGFRβprotein in gastric cancer tissues and drug-resistant cells lines were detected by immunohistochemistry and Western blotting. si-PDGFRβ were transfected into MGC-803/VCR cells by Lipofectamine 2000, and then Western blotting was used to detect the PDGFRβ protein expression after transfection. The effects of si-PDGFRβ on the proliferation and apoptosis of human gastric cancer MGC-803 cells induced by VCR were detected by Cell Counting Kit-8(CCK-8) and flow cytometry. The results showed that the expression of PDGFRβ protein in gastric cancer tissues was significantly higher than that in normal gastric tissues; the expression of PDGFRβ protein in MGC-803/VCR cells was significantly higher than that in MGC-803 cells, and the expression of PDGFRβ protein decreased significantly after transfection of si-PDGFRβ. After MGC-803/VCR cells were induced by 1 μmol/L, 2 μmol/L,4 μmol/L and 8 μmol/L VCR, the cell proliferation inhibition rates of si-PDGFRβ group were(21.97±0.84)%,(37.63±1.32)%,(55.77±1.39)% and(72.17±1.16)%. There were significant differences from(13.60±0.49)%,(22.33±1.01)%,(38.30±1.56)%, and(52.90±1.08)% of the control group(p<0.01). The results of flow cytometry showed that the apoptosis rate of MGC-803/VCR cells in the si-PDGFRβ group was(29.80±0.64)% compared with that in the control group(13.61±0.49)%, indicating that there was a significant difference between the two groups(p<0.01). The preliminary findings of this study indicated that interference with PDGFRβ by siRNA could promote the apoptosis of gastric cancer MGC-803/VCR cells induced by VCR.
In order to study the expression of platelet derived growth factor receptor β(PDGFRβ) in gastric cancer tissues and vincristine(VCR) resistance gastric cancer cells MGC-803/VCR and investigate the effect of PDGFRβsilence on proliferation and apoptosis of MGC-803/VCR cells, in this research, the expression levels of PDGFRβprotein in gastric cancer tissues and drug-resistant cells lines were detected by immunohistochemistry and Western blotting. si-PDGFRβ were transfected into MGC-803/VCR cells by Lipofectamine 2000, and then Western blotting was used to detect the PDGFRβ protein expression after transfection. The effects of si-PDGFRβ on the proliferation and apoptosis of human gastric cancer MGC-803 cells induced by VCR were detected by Cell Counting Kit-8(CCK-8) and flow cytometry. The results showed that the expression of PDGFRβ protein in gastric cancer tissues was significantly higher than that in normal gastric tissues; the expression of PDGFRβ protein in MGC-803/VCR cells was significantly higher than that in MGC-803 cells, and the expression of PDGFRβ protein decreased significantly after transfection of si-PDGFRβ. After MGC-803/VCR cells were induced by 1 μmol/L, 2 μmol/L,4 μmol/L and 8 μmol/L VCR, the cell proliferation inhibition rates of si-PDGFRβ group were(21.97±0.84)%,(37.63±1.32)%,(55.77±1.39)% and(72.17±1.16)%. There were significant differences from(13.60±0.49)%,(22.33±1.01)%,(38.30±1.56)%, and(52.90±1.08)% of the control group(p<0.01). The results of flow cytometry showed that the apoptosis rate of MGC-803/VCR cells in the si-PDGFRβ group was(29.80±0.64)% compared with that in the control group(13.61±0.49)%, indicating that there was a significant difference between the two groups(p<0.01). The preliminary findings of this study indicated that interference with PDGFRβ by siRNA could promote the apoptosis of gastric cancer MGC-803/VCR cells induced by VCR.
引文
Appiah-Kubi K.,Wang Y.,Qian H.,Wu M.,Yao X.,Wu Y.,and Chen Y.,2016,Platelet-derived growth factor receptor/platelet-derived growth factor(PDGFR/PDGF)system is a prognostic and treatment response biomarker with multifarious therapeutic targets in cancers,Tumour Biol.,37(8):10053-10066
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Gramer I.,Killick D.,and Scase T.,2017,Expression of VEGFRand PDGFR-alpha/-beta in 187 canine nasal carcinomas,Vet.Comp.Oncol.,15(3):1041-1150
Kerr D.A.,Busarla S.V.P.,Gimbel D.C.,Sohani A.R.,and Nazarian R.M.,2017,m TOR,VEGF,PDGFR,and c-kit signaling pathway activation in Kaposi sarcoma,Hum.Pathol.,65:157-165
Ma S.Y.,Li M.Y.,Liu N.,Li Y.,Li Z.P.,Yang Y.,Yu F.L.,Hu X.Q.,Liu C.,and Mei X.Q.,2017,Vincristine liposomes with smaller particle size have stronger diffusion ability in tumor and improve tumor accumulation of vincristine significantly,Oncotarget,8(50):87276-87291
Maia A.L.C.,Ferreira C.A.,Barros A.L.B.,Silva E.A.T.M.,Ramaldes G.A.,Júnior S.C.A.D.,Oliveira D.C.P.,Fernandes C.,and Soares F.D.C.,2018,Vincristine-loaded hydroxyapatite nanoparticles as a potential delivery system for bone cancer therapy,J.Drug Target,26(7):592-603
Sahu N.,Chan E.,Chu F.,Pham T.,Koeppen H.,Forrest W.,Merchant M.,and Settleman J.,2017,Cotargeting of MEKand PDGFR/STAT3 pathways to treat pancreatic ductal adenocarcinoma,Mol.Cancer Ther.,16(9):1729-1738
Taniyama Y.,Katanoda K.,Charvat H.,Hori M.,Ohno Y.,Sasazuki S.,and Tsugane S.,2017,Estimation of lifetime cumulative incidence and mortality risk of gastric cancer,Jpn.J.Clin.Oncol.,47(11):1097-1102
Yu B.Q.,Gu D.S.,Zhang X.L.,Liu B.Y.,and Xie J.W.,2017,The role of GLI2-ABCG2 signaling axis for 5Fu resistance in gastric cancer,J.Genet.Genomics,44(8):375-383
Zha L.,He L.,Xie W.,Cheng J.,Li T.,Mohsen M.O.,Lei F.,Storni F.,Bachmann M.,Chen H.,and Zhang Y.,2017,Therapeutic silence of pleiotrophin by targeted delivery of si RNA and its effect on the inhibition of tumor growth and metastasis,PLo S One,12(5):e0177964
Ciesielski M.,Kruszewski W.J.,Walczak J.,Szajewski M.,Szefel J.,Wydra J.,Buczek T.,and Czerepko M.,2017,Analysis of postoperative morbidity and mortality following surgery for gastric cancer.S urgeon volume as the most sig nificant prognostic fact,Prz.Gastroenterol,12(3):215-221
Drinane M.C.,Yaqoob U.,Yu H.,Luo F.,Greuter T.,Arab J.P.,Kostallari E.,Verma V.K.,Maiers J.,De Assuncao T.M.,Simons M.,Mukhopadhyay D.,Kisseleva T.,Brenner D.A.,Urrutia R.,Lomberk G.,Gao Y.,Ligresti G.,Tschumperlin D.J.,Revzin A.,Cao S.,and Shah V.H.,2017,Synectin promotes fibrogenesis by regulating PDGFR isoforms through distinct mechanisms,JCI Insight,2(24):92821
Gramer I.,Killick D.,and Scase T.,2017,Expression of VEGFRand PDGFR-alpha/-beta in 187 canine nasal carcinomas,Vet.Comp.Oncol.,15(3):1041-1150
Kerr D.A.,Busarla S.V.P.,Gimbel D.C.,Sohani A.R.,and Nazarian R.M.,2017,m TOR,VEGF,PDGFR,and c-kit signaling pathway activation in Kaposi sarcoma,Hum.Pathol.,65:157-165
Ma S.Y.,Li M.Y.,Liu N.,Li Y.,Li Z.P.,Yang Y.,Yu F.L.,Hu X.Q.,Liu C.,and Mei X.Q.,2017,Vincristine liposomes with smaller particle size have stronger diffusion ability in tumor and improve tumor accumulation of vincristine significantly,Oncotarget,8(50):87276-87291
Maia A.L.C.,Ferreira C.A.,Barros A.L.B.,Silva E.A.T.M.,Ramaldes G.A.,Júnior S.C.A.D.,Oliveira D.C.P.,Fernandes C.,and Soares F.D.C.,2018,Vincristine-loaded hydroxyapatite nanoparticles as a potential delivery system for bone cancer therapy,J.Drug Target,26(7):592-603
Sahu N.,Chan E.,Chu F.,Pham T.,Koeppen H.,Forrest W.,Merchant M.,and Settleman J.,2017,Cotargeting of MEKand PDGFR/STAT3 pathways to treat pancreatic ductal adenocarcinoma,Mol.Cancer Ther.,16(9):1729-1738
Taniyama Y.,Katanoda K.,Charvat H.,Hori M.,Ohno Y.,Sasazuki S.,and Tsugane S.,2017,Estimation of lifetime cumulative incidence and mortality risk of gastric cancer,Jpn.J.Clin.Oncol.,47(11):1097-1102
Yu B.Q.,Gu D.S.,Zhang X.L.,Liu B.Y.,and Xie J.W.,2017,The role of GLI2-ABCG2 signaling axis for 5Fu resistance in gastric cancer,J.Genet.Genomics,44(8):375-383
Zha L.,He L.,Xie W.,Cheng J.,Li T.,Mohsen M.O.,Lei F.,Storni F.,Bachmann M.,Chen H.,and Zhang Y.,2017,Therapeutic silence of pleiotrophin by targeted delivery of si RNA and its effect on the inhibition of tumor growth and metastasis,PLo S One,12(5):e0177964