Effects of renal tubular epithelial cell microenvironments on proliferation, migration, and differentiation of metanephric mesenchymal cells

doi:10.3877/cma.j.issn.2095-3216.2025.04.002

Abstract

Abstract:

Objective

To investigate the effects of normal and injured microenvironments derived from renal tubular epithelial cell line TCMK-1 cells on the proliferation, migration, and differentiation of metanephric mesenchymal cells (MMCs), and to identify the signaling pathways involved.

Methods

The normal microenvironment supernatant was prepared using the TCMK-1 cells, while the injured microenvironment supernatant was generated by treating TCMK-1 cells with 5 μg/mL cisplatin. The epithelial induction medium was prepared by using 50 ng/ml basic fibroblast growth factor and 10 ng/ml transforming growth factor α. In the proliferation experiment, the MMCs were divided into control group (regular culture medium), normal microenvironment group, and injured microenvironment group. Cell proliferation was detected by the cell counting kit-8 method. In the migration experiment, the MMCs were divided into control group, cisplatin group (regular culture medium+ 5 μg/ml cisplatin), normal microenvironment group, and injured microenvironment group. The cell migration ability was assessed by means of the transwell chamber assay. In the differentiation experiment, the MMCs were divided into control group, epithelial induction group (epithelial induction medium), epithelial induction plus normal microenvironment group, and epithelial induction plus injured microenvironment group. After 10 days of culture, Western blotting was used to detect the protein expression of E-cadherin, mesenchymal marker α-smooth muscle actin, and renal tubular epithelial-specific marker aquaporin 2. In the pathway verification experiment, Western blotting was used to detect the expression levels of canonical wingless/integrated (Wnt) pathway proteins, including Wnt3A, β-catenin, and T-cell factor 4. The small molecule inhibitor MSAB of the canonical Wnt pathway was added to assess its effect on epithelial differentiation.

Results

Compared with the control group, the proliferative activity of MMCs was inhibited in both the normal microenvironment group and the injured microenvironment group (both P<0.05), while the migration ability of the two groups increased (both P<0.01). Compared with the control group and the epithelial induction group, the expression of E-cadherin in the MMCs of both the normal microenvironment group and the injured microenvironment group increased, while the expression of α-smooth muscle actin decreased. Additionally, the expression of aquaporin 2 in the injured microenvironment group also increased. The expression of canonical Wnt pathway-related proteins as Wnt3A, β-catenin, and T-cell factor 4 was higher in the injured microenvironment group (all P<0.05), and the addition of MSAB could inhibit the effect of the injured microenvironment in promoting the epithelial differentiation of MMCs (P<0.05).

Conclusion

Both the normal and injured microenvironments of the renal tubular epithelial cells slightly inhibited the proliferation of the MMCs, but promoted their migration. The injured microenvironment of the renal tubular epithelial cells activated the canonical Wnt pathway and promoted the differentiation of MMCs into renal tubular epithelial cells.

Key words: Microenvironment, Metanephric mesenchymal cells, Canonical Wnt pathway, Epithelial differentiation

Pengcheng Ji, Wenkai Guo, Jingru Bi, Bing Han, Bo Fu, Yuansheng Xie. Effects of renal tubular epithelial cell microenvironments on proliferation, migration, and differentiation of metanephric mesenchymal cells[J]. Chinese Journal of Kidney Disease Investigation(Electronic Edition), 2025, 14(04): 188-195.

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Figures/Tables 5

References 26

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