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MiR‑30a‑5p mitigates autophagy by regulating the Beclin‑1/ATG16 pathway in renal ischemia/reperfusion injury.

ABSTRACT: Renal ischemia/reperfusion (I/R) injury often occurs during multiple organ failure and sepsis, and autophagy may serve a role in I/R injury. The aim of the present study was to explore the effect of microRNA (miR)‑30a‑5p on autophagy in renal I/R injury. miR‑30a‑5p and autophagy‑related protein expression levels in renal I/R injury mouse models and in hypoxia/re‑oxygenation HK‑2 cell models were determined using reverse transcription‑quantitative PCR or western blotting; apoptosis was analyzed using flow cytometry. The effects of miR‑30a‑5p, Beclin‑1 and autophagy‑related gene 16 (ATG16) on the proliferation and autophagy of HK‑2 cells were analyzed through gain‑ and loss‑of‑function studies. miR‑30a‑5p expression was significantly decreased after renal I/R injury in the in vivo and in vitro experiments. Renal I/R injury led to upregulated expression of autophagy‑related proteins microtubule‑associated protein light chain 3 (LC3)‑Ⅱ and Beclin‑1, and downregulated expression of p62. miR‑30a‑5p overexpression decreased the number of LC3 punctae, decreased HK‑2 cell apoptosis, increased p62 expression and decreased LC3‑Ⅱ and Beclin‑1 expression. Inhibition of miR‑30a‑5p exhibited the opposite effects. A luciferase reporter assay demonstrated that miR‑30a‑5p targeted Beclin‑1. Beclin‑1 overexpression led to a significant increase in LC3‑Ⅱ expression and a decrease in p62 expression, as well as a significant increase in apoptosis. Beclin‑1 overexpression also increased the protein expression level of ATG16. Downregulation of Beclin‑1 decreased the expression of LC3‑Ⅱ, elevated the p62 level and decreased apoptosis. ATG16 knockdown showed similar effects as those of Beclin‑1 downregulation. In conclusion, miR‑30a‑5p was increased in renal I/R injury and might mitigate autophagy by regulating the Beclin‑1/ATG16 pathway.

SUBMITTER: Fang Y

PROVIDER: S-EPMC8175063 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Penehyclidine hydrochloride (PHC) suppresses renal ischemia and reperfusion (I/R) injury (IRI); however, the underlying mechanism of action that achieves this function remains largely unknown. The present study aimed to investigate the potential role of autophagy in PHC?induced suppression of renal IRI, as well as the involvement of cell proliferation and apoptosis. A rat IRI model and a cellular hypoxia/oxygenation (H/R) model were established; PHC, 3?methyladenine (3?MA) and rapamycin (Rapa) were administered to the IRI model rats prior to I/R induction and to H/R cells following reperfusion. Serum creatinine was measured using a biochemistry analyzer, whereas aspartate aminotransferase (ASAT) and alanine aminotransferase (ALAT) expression levels were detected using ELISA kits. Renal tissue injury was evaluated by histological examination. In addition, microtubule?associated protein light chain 3B (LC3B) expression, autophagosome formation, cell proliferation and apoptosis were detected in the cellular H/R model. The results demonstrated that I/R induced renal injury in IRI model rats, upregulated serum creatinine, ALAT and ASAT expression levels, and increased autophagic processes. In contrast, pretreatment with PHC or Rapa significantly prevented these I/R?induced changes, whereas the administration of 3?MA enhanced I/R?induced injuries through suppressing autophagy. PHC and Rapa increased LC3B and Beclin?1 expression levels, but decreased sequestome 1 (p62) expression in the cellular H/R model, whereas 3?MA prevented these PHC?induced changes. PHC and Rapa promoted proliferation and autophagy in the cellular H/R model; these effects were accompanied by increased expression levels of LC3B and Beclin?1, and reduced p62 expression levels, whereas these levels were inhibited by 3?MA. Furthermore, PHC and Rapa inhibited apoptosis in the cellular H/R model through increasing Bcl?2 expression levels, and suppressing Bax and caspase?3 expression levels; the opposite effect was induced by 3?MA. In conclusion, PHC suppressed renal IRI through the induction of autophagy, which in turn promoted proliferation and suppressed apoptosis in renal cells.

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MiR‑30a‑5p mitigates autophagy by regulating the Beclin‑1/ATG16 pathway in renal ischemia/reperfusion injury.

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