PMID- 38201227 OWN - NLM STAT- MEDLINE DCOM- 20240112 LR - 20240201 IS - 2073-4409 (Electronic) IS - 2073-4409 (Linking) VI - 13 IP - 1 DP - 2023 Dec 21 TI - NLRX1 Prevents M2 Macrophage Polarization and Excessive Renal Fibrosis in Chronic Obstructive Nephropathy. LID - 10.3390/cells13010023 [doi] LID - 23 AB - BACKGROUND: Chronic kidney disease often leads to kidney dysfunction due to renal fibrosis, regardless of the initial cause of kidney damage. Macrophages are crucial players in the progression of renal fibrosis as they stimulate inflammation, activate fibroblasts, and contribute to extracellular matrix deposition, influenced by their metabolic state. Nucleotide-binding domain and LRR-containing protein X (NLRX1) is an innate immune receptor independent of inflammasomes and is found in mitochondria, and it plays a role in immune responses and cell metabolism. The specific impact of NLRX1 on macrophages and its involvement in renal fibrosis is not fully understood. METHODS: To explore the specific role of NLRX1 in macrophages, bone-marrow-derived macrophages (BMDMs) extracted from wild-type (WT) and NLRX1 knockout (KO) mice were stimulated with pro-inflammatory and pro-fibrotic factors to induce M1 and M2 polarization in vitro. The expression levels of macrophage polarization markers (Nos2, Mgl1, Arg1, and Mrc1), as well as the secretion of transforming growth factor beta (TGFbeta), were measured using RT-PCR and ELISA. Seahorse-based bioenergetics analysis was used to assess mitochondrial respiration in naive and polarized BMDMs obtained from WT and NLRX1 KO mice. In vivo, WT and NLRX1 KO mice were subjected to unilateral ureter obstruction (UUO) surgery to induce renal fibrosis. Kidney injury, macrophage phenotypic profile, and fibrosis markers were assessed using RT-PCR. Histological staining (PASD and Sirius red) was used to quantify kidney injury and fibrosis. RESULTS: Compared to the WT group, an increased gene expression of M2 markers-including Mgl1 and Mrc1-and enhanced TGFbeta secretion were found in naive BMDMs extracted from NLRX1 KO mice, indicating functional polarization towards the pro-fibrotic M2 subtype. NLRX1 KO naive macrophages also showed a significantly enhanced oxygen consumption rate compared to WT cells and increased basal respiration and maximal respiration capacities that equal the level of M2-polarized macrophages. In vivo, we found that NLRX1 KO mice presented enhanced M2 polarization markers together with enhanced tubular injury and fibrosis demonstrated by augmented TGFbeta levels, fibronectin, and collagen accumulation. CONCLUSIONS: Our findings highlight the unique role of NLRX1 in regulating the metabolism and function of macrophages, ultimately protecting against excessive renal injury and fibrosis in UUO. FAU - Liu, Ye AU - Liu Y AD - Department of Critical Care Medicine, Zhongnan Hospital of Wuhan University, Wuhan 430071, China. AD - Department of Pathology, Amsterdam Infection & Immunity, Amsterdam UMC, University of Amsterdam, 1105 AZ Amsterdam, The Netherlands. FAU - Kors, Lotte AU - Kors L AD - Department of Pathology, Amsterdam Infection & Immunity, Amsterdam UMC, University of Amsterdam, 1105 AZ Amsterdam, The Netherlands. FAU - Butter, Loes M AU - Butter LM AD - Department of Pathology, Amsterdam Infection & Immunity, Amsterdam UMC, University of Amsterdam, 1105 AZ Amsterdam, The Netherlands. FAU - Stokman, Geurt AU - Stokman G AUID- ORCID: 0000-0002-3280-6311 AD - Department of Pathology, Amsterdam Infection & Immunity, Amsterdam UMC, University of Amsterdam, 1105 AZ Amsterdam, The Netherlands. FAU - Claessen, Nike AU - Claessen N AD - Department of Pathology, Amsterdam Infection & Immunity, Amsterdam UMC, University of Amsterdam, 1105 AZ Amsterdam, The Netherlands. FAU - Zuurbier, Coert J AU - Zuurbier CJ AUID- ORCID: 0000-0001-8361-2448 AD - Department of Anesthesiology, Amsterdam UMC, University of Amsterdam, 1105 AZ Amsterdam, The Netherlands. FAU - Girardin, Stephen E AU - Girardin SE AD - Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON M5S 1A1, Canada. FAU - Leemans, Jaklien C AU - Leemans JC AD - Department of Pathology, Amsterdam Infection & Immunity, Amsterdam UMC, University of Amsterdam, 1105 AZ Amsterdam, The Netherlands. FAU - Florquin, Sandrine AU - Florquin S AD - Department of Pathology, Amsterdam Infection & Immunity, Amsterdam UMC, University of Amsterdam, 1105 AZ Amsterdam, The Netherlands. FAU - Tammaro, Alessandra AU - Tammaro A AUID- ORCID: 0000-0003-3128-5259 AD - Department of Pathology, Amsterdam Infection & Immunity, Amsterdam UMC, University of Amsterdam, 1105 AZ Amsterdam, The Netherlands. LA - eng GR - 457002002/Netherlands Organisation for Health Research and Development/ PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20231221 PL - Switzerland TA - Cells JT - Cells JID - 101600052 RN - 0 (Transforming Growth Factor beta) RN - 0 (NLRX1 protein, mouse) RN - 0 (Mitochondrial Proteins) SB - IM MH - Animals MH - Mice MH - *Renal Insufficiency, Chronic MH - Macrophages MH - Genes, Regulator MH - Fibrosis MH - Transforming Growth Factor beta MH - Mitochondrial Proteins PMC - PMC10778504 OTO - NOTNLM OT - NLRX1 OT - OXPHOS OT - macrophage polarization OT - renal fibrosis COIS- The authors declare no conflict of interest. EDAT- 2024/01/11 07:41 MHDA- 2024/01/12 06:43 PMCR- 2023/12/21 CRDT- 2024/01/11 01:04 PHST- 2023/09/22 00:00 [received] PHST- 2023/12/04 00:00 [revised] PHST- 2023/12/12 00:00 [accepted] PHST- 2024/01/12 06:43 [medline] PHST- 2024/01/11 07:41 [pubmed] PHST- 2024/01/11 01:04 [entrez] PHST- 2023/12/21 00:00 [pmc-release] AID - cells13010023 [pii] AID - cells-13-00023 [pii] AID - 10.3390/cells13010023 [doi] PST - epublish SO - Cells. 2023 Dec 21;13(1):23. doi: 10.3390/cells13010023.