Citation: Barth, J.; Loeffler, I.; Bondeva, T.; Liebisch, M.; Wolf, G. The Role of Hypoxia on the Trimethylation of H3K27 in Podocytes. Biomedicines 2023, 11, 2475. https://doi.org/10.3390/ biomedicines11092475 Academic Editor: Balazs Varga Received: 1 August 2023 Revised: 28 August 2023 Accepted: 4 September 2023 Published: 7 September 2023 Copyright: © 2023 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/). biomedicines Article The Role of Hypoxia on the Trimethylation of H3K27 in Podocytes Johanna Barth, Ivonne Loeffler , Tzvetanka Bondeva, Marita Liebisch * ,† and Gunter Wolf † Department of Internal Medicine III, University Hospital Jena, Am Klinikum 1, 07747 Jena, Germany; johanna.barth@uni-jena.de (J.B.); ivonne.loeffler@med.uni-jena.de (I.L.); tzvetanka.bondeva@med.uni-jena.de (T.B.); gunter.wolf@med.uni-jena.de (G.W.) * Correspondence: marita.liebisch@med.uni-jena.de † These authors contributed equally to this work. Abstract: Epigenetic alterations contribute to the pathogenesis of chronic diseases such as diabetes mellitus. Previous studies of our group showed that diabetic conditions reduce the trimethylation of H3K27 in podocytes in a NIPP1- (nuclear inhibitor of protein phosphatase 1) and EZH2- (enhancer of zeste homolog 2) dependent manner. It has been previously reported that in differentiated podocytes, hypoxia decreases the expression of slit diaphragm proteins and promotes foot process effacement, thereby contributing to the progression of renal disease. The exact mechanisms are, however, not completely understood. The aim of this study was to analyze the role of hypoxia and HIFs (hypoxia- inducible factor) on epigenetic changes in podocytes affecting NIPP1, EZH2 and H3K27me3, in vitro and in vivo. In vivo studies were performed with mice exposed to 10% systemic hypoxia for 3 days or injected with 3,4-DHB (dihydroxybenzoate), a PHD (prolyl hydroxylase) inhibitor, 24 h prior analyses. Immunodetection of H3K27me3, NIPP1 and EZH2 in glomerular podocytes revealed, to the best of our knowledge for the first time, that hypoxic conditions and pharmacological HIFs activation significantly reduce the expression of NIPP1 and EZH2 and diminish H3K27 trimethylation. These findings are also supported by in vitro studies using murine-differentiated podocytes. Keywords: hypoxia; podocyte; H3K27me3; epigenetic; NIPP1; nuclear inhibitor of protein phosphatase 1; EZH2; enhancer of zeste homolog 2 1. Introduction Chronic kidney disease (CKD) affects more than 10% of the general population world- wide [1]. Diabetic kidney disease (DKD) is the most common cause of CKD and end-stage renal disease (ESRD) that occurs in 30–40% of diabetic patients [2]. Podocyte dysfunction is a major factor in the development of DKD and correlates with proteinuria [2]. Laser scanning microscopy analyses of podocyte architecture in type 2 diabetic patients with microalbuminuria showed markers of injury, including podocyte hypertrophy, diffuse foot process effacement, as well as sites of initial detachment from the basement membrane; and in patients with late stages of proteinuria, podocyte loss and extensively denuded glomerular basement membranes [3]. Animal models revealed that extensive podocyte loss causes irreversible glomerular damage and progresses to tubulointerstitial fibrosis and ESRD [2,4–6]. Epigenetic changes mean variations in gene expression induced by environmental factors without variation in the DNA nucleotide sequence, such as histone methylation. EZH2 (enhancer of zeste homolog 2) catalyzes the trimethylation of lysine 27 of histone 3 (H3K27) [7]. Major clinical trials of patients with diabetes mellitus showed an increased development of DKD caused by a relatively short period of poor glucose control many years ago despite an excellent glycemic control thereafter. This phenomenon is called metabolic memory [2]. Biomedicines 2023, 11, 2475. https://doi.org/10.3390/biomedicines11092475 https://www.mdpi.com/journal/biomedicines