Department of Urology, Huashan Hospital, Fudan University, Shanghai, 200040, People's Republic of China.
Clinical Research Center of Urolithiasis, Shanghai Medical College, Fudan University, Shanghai, 200040, People's Republic of China.
Int Urol Nephrol. 2024 Jul;56(7):2165-2177. doi: 10.1007/s11255-024-03948-3. Epub 2024 Feb 19.
Calcium-sensing receptor (CASR) influences the expression pattern of multiple genes in renal tubular epithelial cells. The objective of this inquiry was to explore the molecular mechanisms of CASR in renal tubular epithelial cells and nephrolithiasis.
HK-2 cells were transfected with lentiviruses carrying either CASR (named CASR) or an empty vector negative control (named NC), as well as shRNA intended to target CASR (named shCASR) or its corresponding negative control (named shNC). CCK-8 assay was used to detect the effect of CASR on the proliferation of HK-2 cells. RNA-Sequencing was applied to explore potential pathways regulated by CASR in HK-2 cells.
PCR and western blot results showed that CASR expression was significantly increased in CASR cells and was decreased in shCASR cells when compared to their corresponding negative control, respectively. CCK-8 assay revealed that CASR inhibited the proliferation of HK-2 cells. RNA-Sequencing results suggested that the shCASR HK-2 cells exhibited a significant up-regulation of 345 genes and a down-regulation of 366 genes. These differentially expressed genes (DEGs) were related to cell apoptosis and cell development. In CASR HK-2 cells, 1103 DEGs primarily functioned in mitochondrial energy metabolism, and amino acid metabolism. With the Venn diagram, 4 DEGs (Clorf116, ENPP3, IL20RB, and CLDN2) were selected as the hub genes regulated by CASR. Enrichment analysis revealed that these hub genes were involved in cell-cell junction, and epithelial cell development.
In summary, our investigation has the potential to offer novel perspectives on CASR regulating cell-cell junction in HK-2 cells.
钙敏感受体（CASR）影响肾小管上皮细胞中多种基因的表达模式。本研究旨在探讨 CASR 在肾小管上皮细胞和肾结石中的分子机制。
用携带 CASR（命名为 CASR）或空载体阴性对照（命名为 NC）的慢病毒转染 HK-2 细胞，以及靶向 CASR 的 shRNA（命名为 shCASR）或其相应的阴性对照（命名为 shNC）。CCK-8 测定法用于检测 CASR 对 HK-2 细胞增殖的影响。RNA 测序用于探索 CASR 在 HK-2 细胞中调节的潜在途径。
PCR 和 Western blot 结果表明，CASR 细胞中 CASR 表达明显增加，shCASR 细胞中 CASR 表达明显降低。CCK-8 测定法显示 CASR 抑制 HK-2 细胞的增殖。RNA 测序结果表明，shCASR HK-2 细胞中 345 个基因显著上调，366 个基因下调。这些差异表达基因（DEGs）与细胞凋亡和细胞发育有关。在 CASR HK-2 细胞中，1103 个 DEG 主要参与线粒体能量代谢和氨基酸代谢。通过 Venn 图，选择了 4 个 DEG（Clorf116、ENPP3、IL20RB 和 CLDN2）作为受 CASR 调节的枢纽基因。富集分析表明，这些枢纽基因参与细胞-细胞连接和上皮细胞发育。
综上所述，我们的研究为 CASR 调节 HK-2 细胞细胞-细胞连接提供了新的视角。
