Cell cycle delay and apoptosis are induced by high salt and urea in renal medullary cells
Cell cycle delay and apoptosis are induced by high salt and urea in renal medullary cells
复制标题
DOI：
10.1152/ajprenal.2000.278.2.f209
复制
发表时间：
2000-02-01
影响因子：
4.2
通讯作者：
Burg, MB
中科院分区：
文献类型：
--
作者：
Michea, L;Ferguson, DR;Burg, MB
我们研究了高渗对小鼠肾集合管细胞亚融合培养存活和增殖的影响。用3-(4，5-二甲基噻唑-2-基)-2，5-二苯基四氮唑溴化法测定，高浓度的氯化钠和/或尿素(但不是甘油)会减少活细胞的数量。用流式细胞仪测定，加入氯化钠和/或尿素将渗透压从正常水平(300momoL/kg)提高到550-1000momoL/kg，可显著增加细胞在8h内处于G(2)M期的细胞比例。当浓度达到600momoL/kg时，这种作用只是暂时的，当浓度达到550momoL/kg时，作用12h后效应逆转，多数细胞处于G(1)期。用5-溴脱氧尿嘧啶核苷(BrdU)脉冲追逐的流式细胞术显示，细胞周期的第8个时相的移动变慢，这取决于氯化钠和/或尿素的浓度，而G(2)M的持续时间显著增加(从300momoL/kg的2.5h增加到高渗透压下的16h以上)。在总渗透压为550 momoL/kg或更高的情况下，加入氯化钠和/或尿素也可诱导细胞凋亡，表现为特征性的电子显微镜形态变化、流式细胞仪中出现亚二倍体峰以及caspase-3的激活。亚二倍体DNA和caspase-3激活的细胞数在8-12h达到高峰，caspase-3在细胞周期的所有时相都有激活，但在G(0)/G(1)和8个周期中激活程度不成比例。我们的结论是，高浓度的氯化钠和/或尿素通过减缓通过8期的过渡，通过延迟G(2)M和G(1)的细胞周期，并通过诱导细胞凋亡来减少增殖的mIMCD3细胞的数量。
We investigated the effects of hyperosmolality on survival and proliferation of subconfluent cultures of mIMCD3 mouse renal collecting duct cells. High NaCl and/or urea (but not glycerol) reduces the number of viable cells, as measured with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT). Raising osmolality from a normal level (300 mosmol/kg) to 550-1,000 mosmol/kg by adding NaCl and/or urea greatly increases the proportion of cells in the G(2)M phase of the cell cycle within 8 h, as measured by flow cytometry. Up to 600 mosmol/kg the effect is only transient, and by 12 h at 550 mosmol/kg the effect reverses and most cells are in G(1). Flow cytometry with 5-bromodeoxyuridine (BrdU) pulse-chase demonstrates that movement through the 8 phase of the cell cycle slows, depending on the concentrations of NaCl and/or urea, and that the duration of G(2)M increases greatly (from 2.5 h at 300 mosmol/kg to more than 16 h at the higher osmolalities). Addition of NaCl and/or urea to total osmolality of 550 mosmol/kg or more also induces apoptosis, as demonstrated by characteristic electron microscopic morphological changes, appearance of a subdiploid peak in flow cytometry, and caspase-3 activation. The number of cells with subdiploid DNA and activated caspase-3 peaks at 8-12 h. Caspase-3 activation occurs in all phases of the cell cycle, but to a disproportionate degree in G(0)/G(1) and 8 phases. We conclude that elevated NaCl and/or urea reduces the number of proliferating mIMCD3 cells by slowing the transit through the 8 phase, by cell cycle delay in the G(2)M and G(1), and by inducing apoptotic cell death.