血小板衍生生长因子受体样基因的表达与功能特征。
Expression and functional characterization of platelet-derived growth factor receptor-like gene.
机构信息
Cancer Research Institute, Xiangya Medical School, Central South University, Changsha 410078, Hunan Province, China.
出版信息
World J Gastroenterol. 2010 Mar 28;16(12):1465-72. doi: 10.3748/wjg.v16.i12.1465.
AIM
To investigate the role of platelet-derived growth factor receptor-like gene (PDGFRL) in the anti-cancer therapy for colorectal cancers (CRC).
METHODS
PDGFRL mRNA and protein levels were measured by reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry in CRC and colorectal normal tissues. PDGFRL prokaryotic expression vector was carried out in Escherichia coli (E. coli), and purified by immobilized metal affinity chromatography. The effect of PDGFRL protein on CRC HCT-116 cells was detected by 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT), clone counting, cell cycle, and wound healing assay.
RESULTS
Both RT-PCR and immunohistochemistry showed that the expression of PDGFRL in colorectal normal tissues was higher than in cancer tissues. Recombinant pET22b-PDGFRL prokaryotic expression vector was successfully expressed in E. coli, and the target protein was expressed in the form of inclusion bodies. After purification and refolding, recombinant human PDGFRL (rhPDGFRL) could efficiently inhibit the proliferation and invasion of CRC HCT-116 cells detected by MTT, clone counting and wound healing assay. Moreover, rhPDGFRL arrested HCT-116 cell cycling at the G0/G1 phase.
CONCLUSION
PDGFRL is a potential gene for application in the anti-cancer therapy for CRC.
目的
探讨血小板衍生生长因子受体样基因（PDGFRL）在结直肠癌（CRC）抗癌治疗中的作用。
方法
采用逆转录-聚合酶链反应（RT-PCR）和免疫组织化学法检测CRC及结直肠正常组织中 PDGFRLmRNA 和蛋白水平。在大肠杆菌（E. coli）中进行 PDGFRL 原核表达载体，并通过固定化金属亲和层析进行纯化。通过 3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐（MTT）、克隆计数、细胞周期和划痕愈合试验检测 PDGFRL 蛋白对 CRC HCT-116 细胞的影响。
结果
RT-PCR 和免疫组织化学均显示，PDGFRL 在结直肠正常组织中的表达高于癌组织。成功在大肠杆菌中表达了重组 pET22b-PDGFRL 原核表达载体，目的蛋白以包涵体形式表达。经纯化和复性后，重组人 PDGFRL（rhPDGFRL）可有效抑制 CRC HCT-116 细胞的增殖和侵袭，MTT、克隆计数和划痕愈合试验均证实这一点。此外，rhPDGFRL 使 HCT-116 细胞周期停滞在 G0/G1 期。
结论
PDGFRL 是 CRC 抗癌治疗中具有应用潜力的基因。
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