In Vitro Evaluation of the Effects of Cadmium on Endocytic Uptakes of Proteins into Cultured Proximal Tubule Epithelial Cells
In Vitro Evaluation of the Effects of Cadmium on Endocytic Uptakes of Proteins into Cultured Proximal Tubule Epithelial Cells
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DOI：
10.3390/toxics8020024
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发表时间：
2020-06-01
期刊：
影响因子：
4.6
通讯作者：
Himeno, Seiichiro
中科院分区：
文献类型：
--
作者：
Fujishiro, Hitomi;Yamamoto, Hazuki;Himeno, Seiichiro
镉（Cd）是一种环境污染物，已知会导致肾脏中生物分子的肾小管重吸收功能障碍。低分子量蛋白质如β 2-微球蛋白（β 2-MG）的尿排泄水平升高已被用作镉诱导的肾小管功能障碍的指标。然而，很少有研究探讨镉对蛋白质重吸收效率的直接影响，使用培养的肾细胞。在这里，我们开发了一种体外测定系统，用于定量的荧光标记的蛋白质的内吞摄取的流式细胞术在S1和S2细胞来自小鼠肾近端小管。通过荧光成像和流式细胞术证实了荧光标记的白蛋白、转铁蛋白、β（2）-MG和金属硫蛋白进入S1细胞的内吞摄取。暴露的S1和S2细胞镉在1和3 μ M的3天导致β（2）-MG和金属硫蛋白的摄取显着下降，但在白蛋白或转铁蛋白。这些结果表明，镉影响肾小管重吸收的低分子量蛋白质，即使在非致死浓度。在这项研究中开发的体外测定系统，以评估蛋白质的内吞摄取可能作为一个有用的工具，用于检测有毒物质，导致肾小管功能障碍。
Cadmium (Cd) is an environmental pollutant known to cause dysfunctions of the tubular reabsorption of biomolecules in the kidney. Elevated levels of urinary excretion of low-molecular-weight proteins such as beta(2)-microglobulin (beta(2)-MG) have been used as an indicator of Cd-induced renal tubular dysfunctions. However, very few studies have examined the direct effects of Cd on the reabsorption efficiency of proteins using cultured renal cells. Here, we developed an in vitro assay system for quantifying the endocytic uptakes of fluorescent-labeled proteins by flow cytometry in S1 and S2 cells derived from mouse kidney proximal tubules. Endocytic uptakes of fluorescent-labeled albumin, transferrin, beta(2)-MG, and metallothionein into S1 cells were confirmed by fluorescence imaging and flow cytometry. The exposure of S1 and S2 cells to Cd at 1 and 3 mu M for 3 days resulted in significant decreases in the uptakes of beta(2)-MG and metallothionein but not in those of albumin or transferrin. These results suggest that Cd affects the tubular reabsorption of low-molecular-weight proteins even at nonlethal concentrations. The in vitro assay system developed in this study to evaluate the endocytic uptakes of proteins may serve as a useful tool for detecting toxicants that cause renal tubular dysfunctions.