Abstract
PurposeWe sought to investigate the role of PAX2 in renal epithelial-to-mesenchymal transition (EMT), examining the influence of PAX2 on ADAM10 expression during renal EMT and ADAM10 expression in fibrotic kidneys.MethodsA rat renal tubular epithelial cell line, NRK52E, was transfected with lentivirus carrying PAX2, and E-cadherin and α-SMA expressions were measured. The influence of PAX2 on ADAM10 promoter activity was evaluated using chromatin immunoprecipitation (CHIP) and dual-luciferase reporter assay. We also treated NRK52E with ADAM10-specific over-expression vector and inhibitors and measured E-cadherin and α-SMA expression. In vivo, Wistar rats (n = 36) were subjected to unilateral ureteral obstruction (UUO) (n = 18) or sham surgery (n = 18), with tissues from post-operative day 3, 7, and 14 days examined, and PAX2/ADAM10 activity measured. ADAM10 expression was also assessed in kidneys from patients with chronic kidney disease (CKD).ResultsIn NRK52E overexpressing PAX2, ADAM10 and α-SMA levels were increased, while E-cadherin levels were decreased. CHIP and dual-luciferase reporter assay showed that PAX2 directly bound to a specific site within the ADAM10 promoter, and over-expression of PAX2 significantly activated ADAM10 transcription. NRK52E with ADAM10 over-expression also significantly decreased E-cadherin and increased α-SMA levels. In the fibrotic kidneys of rats with UUO, E-cadherin levels were increased and α-SMA levels were decreased, and expression of PAX2 and ADAM10 increased. ADAM10 expression also elevated in the renal tissues of CKD patients.ConclusionsPAX2 directly increased expression of ADAM10, the presence of which contributed to EMT in renal tubular epithelia and hence plays an important role in renal fibrosis.
Highlights
Chronic kidney disease (CKD), characterized by renal fibrosis, is increasingly recognized as a global health issue due to its high prevalence and the devastating sequelae of endstage renal disease, accompanied by a higher risk of cardiovascular events and premature death [1]
We investigated the influence of ADAM10 on paired box 2 (PAX2)-induced Epithelial-to-mesenchymal transition (EMT) in renal tubular epithelia and the effect of ADAM10 in renal fibrosis
NRK52E cells were transfected with pGC–LV–PAX2 and intracellular PAX2 expression was observed via fluorescent microscope
Summary
Chronic kidney disease (CKD), characterized by renal fibrosis, is increasingly recognized as a global health issue due to its high prevalence and the devastating sequelae of endstage renal disease, accompanied by a higher risk of cardiovascular events and premature death [1]. Epithelial-to-mesenchymal transition (EMT), an important process related to renal fibrosis [2], refers to the pathogenic changes in epithelial lineage cells, including loss of structural polarity and the basement membrane contact, followed by morphological alterations producing spindle-shaped cells, similar to mesenchymal/myofibroblast cells [3]. EMT involves four pivotal stems: (1) loss of epithelial adhesion markers, such as the E-cadherin adhesion complex, (2) de novo expression of α-smooth muscle cell actin (SMA) and actin re-organization, (3) matrix metalloproteinasesinduced basement membrane disruption, and (4) increased cell migration and invasion into the nearby interstitium [4]. The activation of mesenchymal markers is accompanied by epithelial marker suppression [5]. This is followed by actin architecture re-organization,
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