机构地区:[1]福建医科大学附属第一医院泌尿外科二区,福州350000
出 处:《中国临床药理学杂志》2018年第2期137-140,共4页The Chinese Journal of Clinical Pharmacology
基 金:2016年福建省自然科学基金资助项目(2016J01528)
摘 要:目的研究钙敏感受体(CasR)抑制药NPS-2390对肾小管上皮细胞钙超载及黏附晶体能力的影响。方法在对数生长期的NRK-52E细胞中转染过表达CasR载体及空白载体,分别作为模型组及对照组。实验组在模型组的基础上加入5μg·mL^(-1)NPS-2390。蛋白免疫印迹(Western blot)及荧光定量聚合酶链式反应(q PCR)技术检测各组细胞CasR的表达情况。激光共聚焦技术检测各组细胞中钙离子浓度的变化,用噻唑蓝(MTT)法检测细胞活性,用红色荧光标记的一水草酸钙晶体(Alexa Fluor350COM)刺激细胞,荧光显微镜观察细胞荧光强度。结果模型组、对照组及实验组CasR的蛋白表达量分别为0.73±0.06,0.33±0.04,0.29±0.03,模型组CasR的蛋白表达量显著高于对照组及实验组,差异均有统计学意义(均P<0.05)。模型组、对照组及实验组CasR的mRNA表达量分别为0.58±0.05,0.15±0.01及0.10±0.01,模型组CasR的mRNA表达量显著高于对照组及实验组,差异均有统计学意义(均P<0.05)。模型组、对照组及实验组钙离子荧光强度增加率分别为-34.8%,9.0%,4.3%,模型组荧光强度增加率显著低于对照组及实验组,差异均有统计学意义(均P<0.05)。模型组、对照组及实验组细胞内晶体荧光强度分别为(166±22),(241±29),(255±32)cd,模型组晶体荧光强度显著低于对照组及实验组,差异有统计学意义(P<0.05)。模型组、对照组及实验组细胞活力在48h时分别为3.42±0.35,2.74±0.32,2.55±0.28,模型组细胞活力显著高于对照组及实验组,差异均有统计学意义(均P<0.05)。模型组、对照组及实验组细胞活力在72 h时分别为7.16±0.63,5.64±0.51,5.24±0.58,模型组细胞活力显著高于对照组及实验组,差异均有统计学意义(均P<0.05)。结论 NPS-2390可抑制CasR的表达,进而加重肾小管上皮细胞钙超载并引起细胞活性减弱,促进细胞黏附晶体。Objective To investigate the effects of calcium sensing re- ceptor (CasR) inhibitor NPS - 2390 on hesion in renal tubular epithelial cells. calcium overload and crystal ad- Methods NRK- 52E cell was transfered CasR expression vector and blank vector in logarithmic growth phase, respectively in the model group and the control group, and the inhibition expression of CasR in model group by NPS -2390 regard as test group. Western blot and fluorescence quantitative PCR (qPCR) were used to detect the expression of CasR in the cells. Confocal laser scanning technique was used to detect the changes of calcium ion concentration inthe cells. MTF assay was used to detect the cell viability. Alexa F1uor350COM was used to stimulate cells, watching the fluorescence intensity of cells by fluorescence microscopy. Results The protein expression of CasR in model group, control group and test group were 0. 73 + 0. 06, 0. 33 + 0.04 and 0. 29 + 0.03, and the expression of CasR protein in model group was significantly higher than those in control group and test group, and the difference was statis- tically significant (P 〈 0. 05 ). The mRNA expression of CasR in model group, control group and test group were 0. 58 +0. 05, 0. 15 +0. 01 and 0. 10 +0. 01, and the expression of CasR mRNA in model group was significantly high- er than those in control group and test group, and the diffe-rence was statistically significant (P 〈 0. 05). The calcium fluorescence intensity increase rate in model group, control group and test group were respectively -34. 8%, 9.0% and 4. 3%, and the calcium fluorescence intensity increase rate in model group was significantly lower than those in control group and test group, and the difference was statistically significant (P 〈 0.05 ). The crystal fluorescence inten- sity in model group, control group and test group were (166 +22), (241 +29) and (255 +32) cd, and the crystal fluorescence intensity in model group was significantly lower than those in control
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