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首页> 美国卫生研究院文献>Molecular Medicine Reports >Apolipoprotein M increases the expression of vitamin D receptor mRNA in colorectal cancer cells detected with duplex fluorescence reverse transcription-quantitative polymerase chain reaction
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Apolipoprotein M increases the expression of vitamin D receptor mRNA in colorectal cancer cells detected with duplex fluorescence reverse transcription-quantitative polymerase chain reaction

机译：载脂蛋白M通过双向荧光逆转录-定量聚合酶链反应检测到的大肠癌细胞中维生素D受体mRNA的表达增加

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Apolipoprotein M (ApoM) and the vitamin D receptor (VDR) are apolipoproteins predominantly presenting in high-density lipoprotein (HDL) and a karyophilic protein belonging to the steroid-thyroid receptor superfamily, respectively. Previous studies have demonstrated that ApoM and VDR are associated with cholesterol metabolism, immune and colorectal cancer regulation. In order to investigate whether ApoM affected the expression of VDR in colorectal cancer cells, a single-tube duplex fluorescence reverse transcription-quantitative polymerase chain reaction (RT-qPCR) system was developed to simultaneously detect the mRNA levels of VDR and GAPDH in HT-29 cells overexpressing ApoM. The results demonstrated that the amplification products were confirmed as the specific fragment of VDR/GAPDH using the DNA sequencing instrument. The sensitivity, linear range, correlation coefficient, amplification efficiency, intra-assay and inter-assay coefficients of variation were 40 copies/µl, 4.00×101-4.00×105 copies/µl, 0.999, 92.42%, 0.09–0.34% and 0.32–0.65% for VDR, and 40 copies/µl, 4.00×101-4.00×105 copies/µl, 0.999, 98.07%, 0.19–0.43% and 0.40–0.75% for GAPDH, respectively. The results indicated that the expression of VDR mRNA was significantly higher in HT-29 cells overexpressing ApoM, compared with the negative control group (P<0.05). In conclusion, the current study successfully developed the single-tube duplex RT-qPCR to simultaneously detect VDR and GAPDH expression in colorectal cancer cells. The methodology results demonstrated that the duplex RT-qPCR system with high sensitivity and specificity could ensure the objectivity and credibility of the detection. The present study confirmed that ApoM significantly increased the expression of VDR in HT-29 cells. In addition, it was hypothesized that ApoM may be involved in antineoplastic activity via the upregulation of VDR expression, which may provide novel directions for the investigation of ApoM in cancer.
机译：载脂蛋白M（ApoM）和维生素D受体（VDR）分别是主要存在于高密度脂蛋白（HDL）和类固醇-甲状腺受体超家族的嗜碱性蛋白中的载脂蛋白。先前的研究表明，ApoM和VDR与胆固醇代谢，免疫和结直肠癌调节有关。为了研究ApoM是否会影响大肠癌细胞中VDR的表达，开发了一种单管双工荧光逆转录定量聚合酶链反应（RT-qPCR）系统，以同时检测HT-细胞中VDR和GAPDH的mRNA水平。 29个过表达ApoM的细胞。结果表明，使用DNA测序仪证实扩增产物为VDR / GAPDH的特异性片段。灵敏度，线性范围，相关系数，扩增效率，批内和批内变异系数分别为40个拷贝/微升，4.00×101-4.00×105个拷贝/微升，0.999、92.42％，0.09-0.34％和0.32对于VDR为–0.65％，对于GAPDH，分别为40拷贝/微升，4.00×101-4.00×105拷贝/微升，0.999、98.07％，0.19–0.43％和0.40–0.75％。结果表明，与阴性对照组相比，过表达ApoM的HT-29细胞中VDR mRNA的表达显着升高（P <0.05）。总之，当前的研究成功开发了单管双工RT-qPCR，可同时检测大肠癌细胞中的VDR和GAPDH表达。方法学结果表明，具有高灵敏度和特异性的双重RT-qPCR系统可以确保检测的客观性和可信性。本研究证实，ApoM可显着增加HT-29细胞中VDR的表达。另外，假设ApoM可能通过上调VDR表达而参与抗肿瘤活性，这可能为研究ApoM在癌症中提供新的方向。

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