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目的探讨甲状旁腺素相关肽在高糖诱导大鼠肾小管导管上皮细胞株NRK-52E转分化作用。方法应用RT-PCR及Western blot检测无糖组(无糖培养基培养),正常葡萄糖组(含5mmol/L葡萄糖DMEM培养基培养)组及高浓度葡萄糖组(含16.7mmol/L葡萄糖DMEM培养基培养)中甲状旁腺素相关肽(parathyroid hormone-related peptide,PTHrP)及其受体1在大鼠肾小管导管上皮细胞株NRK-52E表达。10pmol/L PTHrP及16.7mmol/L葡萄糖分别干预72h,应用Western blot检测转化因子β、金属蛋白酶2、Ⅰ型胶原及α平滑肌肌动蛋白表达情况。ROS含量应用CM2H2DCFDA试剂盒检测。结果与空白组和正常葡萄糖组相比,高浓度葡萄糖能显著上调PTHrP(P<0.05,P<0.01)和甲状旁腺激素受体1(Type1receptor parathyroid hormone-related protein,PTH1R)(P<0.01,P<0.05)蛋白及PTHrP mRNA(P<0.01)表达,PTH1R mRNA在高浓度及正常葡萄糖浓度组的表达高于空白组(P<0.01)。对比空白组,PTHrP能显著上调NRK-52E细胞转化因子β1(P<0.01)、Ⅰ型胶原(P<0.01)及α平滑肌肌动蛋白(P<0.01)表达,而在高浓度葡萄糖状态下,其表达上调更显著(P<0.05,P<0.01)。金属蛋白酶2在PTHrP及高浓度葡萄糖共同干预下表达上调(P<0.01)。PTHrP及高血糖均可以升高NRK-53E细胞ROS含量(P<0.01,P<0.01)。结论高浓度葡萄糖可能通过调控PTHrP/PTH1R的水平从而影响肾小管导管上皮转分化。
Objective To investigate the transdifferentiation of parathyroid hormone-related peptide (NRH) in NRK-52E induced by high glucose in rat renal tubular epithelial cell line. Methods RT-PCR and Western blot were used to detect glucose-free medium (glucose-free medium), normal glucose group (medium containing 5mmol / L glucose DMEM) and high glucose group (16.7mmol / L glucose DMEM medium (PTHrP) and its receptor 1 in rat renal tubular epithelial cell line NRK-52E. 10 pmol / L PTHrP and 16.7 mmol / L glucose for 72 h respectively. Western blot was used to detect the expression of transforming growth factor β, metalloproteinase 2, collagen Ⅰ and α-smooth muscle actin. ROS content using CM2H2DCFDA kit detection. Results Compared with blank group and normal glucose group, high concentration glucose could up-regulate PTHrP (P <0.05, P <0.01) and type 1 receptor parathyroid hormone-related protein (PTH1R) PTH1R mRNA and PTHrP mRNA (P <0.01), PTH1R mRNA expression in high concentration and normal glucose concentration group was higher than that in blank group (P <0.01). Compared with blank group, PTHrP up-regulated the expression of transforming growth factor β1 (P <0.01), type I collagen (P <0.01) and α-smooth muscle actin (P <0.01) in NRK-52E cells, The expression was significantly increased (P <0.05, P <0.01). Metalloproteinase 2 was up-regulated by PTHrP and high glucose (P <0.01). Both PTHrP and hyperglycemia increased ROS content in NRK-53E cells (P <0.01, P <0.01). Conclusion High concentration of glucose may affect tubular epithelial-mesenchymal transition by regulating the level of PTHrP / PTH1R.