Total Flavones of Abelmoschus manihot Ameliorates Podocyte Pyroptosis and Injury in High Glucose Conditions by Targeting METTL3-Dependent m6A Modification-Mediated NLRP3-Inflammasome Activation and PTEN/PI3K/Akt Signaling

Abstract

Background: The total flavones of Abelmoschus manihot (TFA), a compound that is extracted from Abelmoschus manihot, has been widely used in China to reduce podocyte injury in diabetic kidney disease (DKD). However, the mechanisms underlying the therapeutic action of this compound have yet to be elucidated. Podocyte pyroptosis is characterized by activation of the NLRP3 inflammasome and plays an important role in inflammation-mediated diabetic kidneys. Regulation of the PTEN/PI3K/Akt pathway is an effective strategy for improving podocyte damage in DKD. Previous research has also shown that N6-methyladenosine (m6A) modification is involved in DKD and that m6A-modified PTEN regulates the PI3K/Akt pathway. In this study, we investigated whether TFA alleviates podocyte pyroptosis and injury by targeting m6A modification-mediated NLRP3-inflammasome activation and PTEN/PI3K/Akt signaling.Methods: We used MPC-5 cells under high glucose (HG) conditions to investigate the key molecules that are involved in podocyte pyroptosis and injury, including activation of the NLRP3 inflammasome and the PTEN/PI3K/Akt pathway. We detected alterations in the levels of three methyltransferases that are involved in m6A modification. We also investigated changes in the levels of these key molecules in podocytes with the overexpression or knockdown of methyltransferase-like (METTL)3.Results: Analysis showed that TFA and MCC950 protected podocytes against HG-induced pyroptosis and injury by reducing the protein expression levels of gasdermin D, interleukin-1β, and interleukin-18, and by increasing the protein expression levels of nephrin, ZO-1, WT1 and podocalyxin. TFA and 740Y-P inhibited activation of the NLRP3 inflammasome via the PI3K/Akt pathway by inhibiting the protein levels of NIMA-related kinase7, NLRP3, ASC, and caspase-1, and by increasing the protein expression levels of p-PI3K and p-Akt. TFA improved pyroptosis and injury in HG-stimulated podocytes by regulating METTL3-dependent m6A modification.Conclusion: Collectively, our data indicated that TFA could ameliorate pyroptosis and injury in podocytes under HG conditions by adjusting METTL3-dependent m6A modification and regulating NLRP3-inflammasome activation and PTEN/PI3K/Akt signaling. This study provides a better understanding of how TFA can protect podocytes in DKD.

WTAP-mediated N6-methyladenosine modification of NLRP3 mRNA in kidney injury of diabetic nephropathy

“…These data suggest that METTL14 and WTAP may play an important role in renal glomerulus and tubules, respectively. Moreover, the extensive m 6 A modification of NLRP3 mRNA by METTL3 in HG-afflicted podocytes has also been confirmed [ 54 ]. METTL14 also increased the m 6 A modification of NLRP3 mRNA in degenerative nucleus pulposus cells, while the reader decoding the m 6 A modification was different [ 55 ].…”

m6A: An Emerging Role in Programmed Cell Death

“…Pyroptosis is a cell death pathway that is characterized by the activation of various caspases including caspase-1 which is mediated by inflammasomes and causes cell perforation ( Mulay, 2019 ). Studies have suggested that m 6 A plays a key role in the regulatory pathway responsible for pyroptosis in cells ( Zha et al, 2020 ; Liu et al, 2021 ). NOD-like receptor protein 3 (NLRP3) is a key component of the inflammasome that causes pyroptosis and is known to promote pyroptosis and increase the levels of pro-inflammatory cytokines ( Mulay, 2019 ).…”

A bibliometric analysis of RNA methylation in diabetes mellitus and its complications from 2002 to 2022

“…The knockdown of WTAP inhibited pyroptosis and diminished the release of IL-18, IL-1b, caspase-1, and NLRP3 in high-glucoseinduced HK-2 cells (33). In clinical applications of regulators targeting RNA modification enzymes, a cell experiment demonstrated the total flavones of Abelmoschus Manihot, the main components of the Huangkui capsule and ameliorated pyroptosis and injury in podocytes under high-glucose conditions by adjusting METTL3-dependent m 6 A modification and regulating NLRP3-inflammasome activation and PTEN/PK13/Akt signaling (50).…”