白蛋白对近端肾小管上皮细胞增殖与凋亡影响.docVIP

白蛋白对近端肾小管上皮细胞增殖与凋亡的影响 【摘要】 【目的】 探讨白蛋白对近端肾小管上皮细胞(PTCs)增殖与凋亡的影响。【方法】 用不同剂量(0.1,0.5,1,5,10,30 mg/mL)去脂牛血清白蛋白(dBSA)超载体外培养的NRK52E,dBSA 0 mg/mL作为对照。细胞增殖活性用MTS比色法检测和显微镜观察。细胞凋亡检测:荧光染料DAPI染核,DNA梯带实验,原位末端标记(TUNEL)实验和苔盼蓝染色计数。【结果】 高剂量dBSA超载未完全融合NRK52E细胞144 h后细胞变得更密集。MTS比色结果发现,高剂量dBSA(5、10、30 mg/mL)超载可明显诱导NRK52E细胞增殖,增殖活性分别为(0.700 ± 0.045)A、(1.021 ± 0.029) A和(1.273 ± 0.173) A,较对照细胞(0.441 ± 0.020) A显着升高,P均 0.05。高剂量dBSA超载的NRK52E细胞,有较多出现核碎裂和核固缩,有较强DNA梯带和原位末端标记信号。苔盼蓝染色计数发现,高剂量dBSA(5、10、30 mg/mL)超载NRK52E细胞72 h的死亡细胞数[(680 000 ± 28 618)/mL, (970 000 ± 52 915)/mL和(1 132 500 ± 88 954)/mL较对照的死亡细胞数[(312 500 ± 28 395)/mL]显着升高, P均 0.05;而且死亡细胞数呈剂量依赖性。【结论】白蛋白超载NRK52E细胞既可诱导细胞增殖,又可诱导细胞凋亡。 【关键词】 白蛋白; 肾小管上皮细胞; 细胞增殖; 细胞凋亡 Abstract: 【Objective】 To study the effects of albumin overload on proliferation and apoptosis in proximal tubular epithelial cells (PTCs). 【Methods】 Different concentration of delipidated bovine serum albumin (dBSA) ranged from 0.1-30 mg/mL were added to NRK52E cells cultured in vitro; dBSA 0 mg/mL as control. Cell proliferation was observed under microscope and measured by One Solution Cell Proliferation Assay Kit (MTS assay). Apoptosis were detected by following methods: DNA laddering assay, In situ terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick-end labeling (TUNEL assay), nuclei changes were observed under immunofluorecent microscope after stained with DAPI (4′,6-diamidino -2-phenylindole) and total death cells were counted with trypan blue staining under light microscope. 【Results】Under light microscope, sub-confluent NRK52E cells became more confluent after dBSA overload 144 h. MTS assay revealed that higher dosages of dBSA (5, 10, and 30 mg/mL) overload induced significant proliferation (0.700 ± 0.045A, 1.021 ± 0.029 A, and 1.273 ± 0.173 A, respectively) 48 hours later in NRK52E, and were higher than that of control cells (0.441 ± 0.020 A), P 0.05. In higher dosages of dBSA (5, 10, and 30 mg/mL) overload NRK52E cells, there were more condensed