Aldosterone-Induced Renal Lymphangiogenesis and Endothelial-To-Mesenchymal Transformation to Promote Renal Interstitial Fibrosis Through the MR/TGF-β1 Pathway in Mice

Background: Lymphangiogenesis and phenotypic transformation of endothelial cells are closely associated with the progression of renal interstitial fibrosis. Inflammatory injury triggered by Mineralocorticoid Receptor (MR) activation serves as the initial stimulus for lymphangiogenesis.

Methods: Thirty specific pathogen-free (SPF) male C57BL/6 mice were assigned to three groups randomly: the control group (CON), aldosterone-treated group (ALD group, in which aldosterone was infused at a rate of 0.75 μg/h via mini-osmotic pumps for 12 weeks), and esaxerenone-treated group (ESA group, administered at a dosage of 1 mg/kg/day via diet). The expression levels of lymphatic markers (lymphatic vessel endothelial hyaluronan receptor 1 (LYVE-1), vascular endothelial growth factor receptor 3 (VEGFR3), podoplanin, and VEGFC) were assessed using immunohistochemistry, immunofluorescence, and western blot analysis. Inflammatory injury markers (CD68, F4/80, IL-1β, TNF-α and TGF-β1) and endothelial-to-mesenchymal transition (EndMT, LYVE-1⁺ vimentin/α smooth muscle actin (α-SMA)⁺) were evaluated. In vitro, the effects of aldosterone on the migration, tube formation, and phenotypic transformation of human lymphatic endothelial cells (HLECs) in the presence of TGF-β1 or VEGFC were investigated.

Results: In the ALD group, significant increases in lymphangiogenesis, macrophage infiltration, and the expression of TGF-β1, TNF-α, IL-1β and VEGFC were observed. Immunofluorescence double staining revealed that VEGFC was predominantly secreted by macrophages, and that lymphatic endothelial cells exhibited expression of vimentin and α-SMA. In vitro experiments demonstrated that aldosterone promoted HLECs migration and tube formation, as well as the activation of inflammatory cytokines and MR. Flow cytometry analysis indicated that HLECs underwent myofibroblastic transformation, which could be attenuated by MR blocker esaxerenone.

Conclusions: Aldosterone induces inflammatory injury, thereby promoting renal lymphangiogenesis and EndMT.

EndMT; TGF-β1; VEGFC; aldosterone; lymphangiogenesis; mineralocorticoid receptor.