{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,11,2]],"date-time":"2026-11-02T23:40:13Z","timestamp":1793662813280,"version":"4.5.1"},"reference-count":30,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2023,3,2]],"date-time":"2023-03-02T00:00:00Z","timestamp":1677715200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Cell Dev. Biol."],"abstract":"<jats:p>Alport syndrome (AS) is a rare disease characterized by defective glomerular basement membranes, caused by mutations in COL4A3, COL4A4, and COL4A5, which synthesize collagen type IV. Patients present with progressive proteinuria, hematuria and podocyte loss. There is currently no cure for Alport syndrome, and this is mainly due to its complex and variable pathogenesis, as well as the lack of models that can faithfully mimic the human phenotype. Here we have developed a novel human culture model of Alport syndrome and used it to study the effects of different mutations on podocyte development and biology. First, we established a differentiation protocol that allowed us to generate podocyte spheroids from patient-derived human induced pluripotent stem cells (hiPSCs). We have then carried out discovery proteomics and demonstrated that a total of 178 proteins were differentially expressed between Alport (AS1 and AS3) and control (LT) podocytes. GO analysis indicated alterations in several metabolic pathways, such as oxidative phosphorylation, RNA maturation, chromatin condensation, and proliferation. Although functional assays showed no changes in lactate production and mitochondrial potential compared to healthy controls, immunofluorescence and electron microscopy analysis showed key morphological changes related to the phenotypical maturation of Alport podocytes. Moreover, the studied mutations led to persistent proliferation, increased reactive oxygen species (ROS) production and the concomitant expression of peroxisome proliferator-activated receptors \u03b1 and \u03b3 (PPAR\u03b1 and PPAR\u03b3) in podocytes. These data on patient-derived podocytes provide evidence that collagen mutations, in addition to playing a central role in the defective development of the glomerular filtration barrier, cause significant alterations in podocyte development and metabolism very early in development, even before the formation of the filtering apparatus. In conclusion, our study provides a new methodological platform for the differentiation of podocytes and to study human podocytopathies in a personalized manner, and reveals new insights into the etiopathogenesis and pathobiology of Alport syndrome.<\/jats:p>","DOI":"10.3389\/fcell.2023.1111424","type":"journal-article","created":{"date-parts":[[2023,3,2]],"date-time":"2023-03-02T06:53:46Z","timestamp":1677740026000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":8,"title":["Patient-derived podocyte spheroids reveal new insights into the etiopathogenesis of Alport syndrome"],"prefix":"10.3389","volume":"11","author":[{"given":"Ricardo","family":"Romero-Guevara","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Orthodoxia","family":"Nicolaou","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Benedetta","family":"Petracca","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sadr","family":"Shaheed","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Christopher","family":"Sutton","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Eleni","family":"Frangou","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Marina","family":"Afami","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kyriacos","family":"Kyriacou","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Adonis","family":"Ioannides","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Christodoulos","family":"Xinaris","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2023,3,2]]},"reference":[{"key":"B1","doi-asserted-by":"publisher","first-page":"130","DOI":"10.1016\/j.scr.2016.06.001","article-title":"Generation of functional podocytes from human induced pluripotent stem cells","volume":"17","author":"Ciampi","year":"2016","journal-title":"Stem Cell Res."},{"key":"B2","doi-asserted-by":"publisher","first-page":"e94818","DOI":"10.1172\/jci.insight.94818","article-title":"Osteopontin deficiency ameliorates Alport pathology by preventing tubular metabolic deficits","volume":"3","author":"Ding","year":"2018","journal-title":"JCI Insight"},{"key":"B3","doi-asserted-by":"publisher","first-page":"785","DOI":"10.1681\/ASN.2011100988","article-title":"Imaging of podocyte foot processes by fluorescence microscopy","volume":"23","author":"Grgic","year":"2012","journal-title":"J. Am. Soc. Nephrol."},{"key":"B4","doi-asserted-by":"publisher","first-page":"577","DOI":"10.1016\/j.kint.2018.09.023","article-title":"ATF6\u03b1 downregulation of PPAR\u03b1 promotes lipotoxicity-induced tubulointerstitial fibrosis","volume":"95","author":"Jao","year":"2019","journal-title":"Kidney Int."},{"key":"B5","doi-asserted-by":"publisher","first-page":"103162","DOI":"10.1016\/j.ebiom.2020.103162","article-title":"Discoidin domain receptor 1 activation links extracellular matrix to podocyte lipotoxicity in Alport syndrome","volume":"63","author":"Kim","year":"2020","journal-title":"EBioMedicine"},{"key":"B6","doi-asserted-by":"publisher","first-page":"183108","DOI":"10.1155\/2008\/183108","article-title":"PPARgamma in kidney physiology and pathophysiology","volume":"2008","author":"Kiss-T\u00f3th","year":"2008","journal-title":"PPAR Res."},{"key":"B7","doi-asserted-by":"publisher","first-page":"291","DOI":"10.1016\/j.scr.2017.08.019","article-title":"Generation of integration-free induced pluripotent stem cell lines derived from two patients with X-linked Alport syndrome (XLAS)","volume":"25","author":"Kuebler","year":"","journal-title":"Stem Cell Res."},{"key":"B8","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1016\/j.scr.2017.08.021","article-title":"Integration-free induced pluripotent stem cells derived from a patient with autosomal recessive Alport syndrome (ARAS)","volume":"25","author":"Kuebler","year":"","journal-title":"Stem Cell Res."},{"key":"B9","doi-asserted-by":"publisher","first-page":"854998","DOI":"10.3389\/fcell.2022.854998","article-title":"\u2018Cell hypertrophy: A \u201cbiophysical roadblock\u201d to reversing kidney injury\u2019","volume":"10","author":"Lavecchia","year":"2022","journal-title":"Front. Cell Dev. Biol."},{"key":"B10","doi-asserted-by":"publisher","first-page":"937","DOI":"10.1111\/nep.13764","article-title":"Alport syndrome: Proteomic analysis identifies early molecular pathway alterations in Col4a3 knock out mice","volume":"25","author":"Nicolaou","year":"2020","journal-title":"Nephrology"},{"key":"B11","doi-asserted-by":"publisher","first-page":"59","DOI":"10.1016\/j.bbrc.2017.06.094","article-title":"Mitochondrial dynamics coordinate cell differentiation","volume":"500","author":"Noguchi","year":"2018","journal-title":"Biochem. Biophysical Res. Commun."},{"key":"B12","doi-asserted-by":"publisher","first-page":"260","DOI":"10.1681\/ASN.2012121217","article-title":"Evidence for activation of the unfolded protein response in collagen IV nephropathies","volume":"25","author":"Pieri","year":"2014","journal-title":"J. Am. Soc. Nephrol. JASN"},{"key":"B13","doi-asserted-by":"publisher","first-page":"563981","DOI":"10.3389\/fphys.2020.563981","article-title":"Kidney organoids as disease models: Strengths, weaknesses and perspectives","volume":"11","author":"Romero-Guevara","year":"2020","journal-title":"Front. Physiology"},{"key":"B14","doi-asserted-by":"publisher","first-page":"166166","DOI":"10.1016\/j.bbadis.2021.166166","article-title":"Mutations in cis that affect mRNA synthesis, processing and translation","volume":"1867","author":"Roos","year":"2021","journal-title":"Biochimica Biophysica Acta (BBA) - Mol. Basis Dis."},{"key":"B15","doi-asserted-by":"publisher","first-page":"504","DOI":"10.1038\/sj.ki.5002041","article-title":"The normal kidney filters nephrotic levels of albumin retrieved by proximal tubule cells: Retrieval is disrupted in nephrotic states","volume":"71","author":"Russo","year":"2007","journal-title":"Kidney Int."},{"key":"B16","doi-asserted-by":"publisher","first-page":"1755","DOI":"10.1016\/j.bbamcr.2006.09.006","article-title":"Peroxisomes and oxidative stress","volume":"1763","author":"Schrader","year":"2006","journal-title":"Biochimica Biophysica Acta (BBA) - Mol. Cell Res."},{"key":"B17","doi-asserted-by":"publisher","first-page":"175","DOI":"10.1007\/s00412-018-0659-8","article-title":"Ki-67: More than a proliferation marker","volume":"127","author":"Sun","year":"2018","journal-title":"Chromosoma"},{"key":"B18","doi-asserted-by":"publisher","first-page":"53","DOI":"10.1016\/j.stem.2013.11.010","article-title":"Redefining the in vivo origin of metanephric nephron progenitors enables generation of complex kidney structures from pluripotent stem cells","volume":"14","author":"Taguchi","year":"2014","journal-title":"Cell Stem Cell"},{"key":"B19","doi-asserted-by":"publisher","first-page":"730","DOI":"10.1016\/j.stem.2017.10.011","article-title":"Higher-order kidney organogenesis from pluripotent stem cells","volume":"21","author":"Taguchi","year":"2017","journal-title":"Cell Stem Cell"},{"key":"B20","doi-asserted-by":"publisher","first-page":"861","DOI":"10.1016\/j.cell.2007.11.019","article-title":"Induction of pluripotent stem cells from adult human fibroblasts by defined factors","volume":"131","author":"Takahashi","year":"2007","journal-title":"Cell"},{"key":"B21","doi-asserted-by":"publisher","first-page":"118","DOI":"10.1038\/ncb2894","article-title":"Directing human embryonic stem cell differentiation towards a renal lineage generates a self-organizing kidney","volume":"16","author":"Takasato","year":"2014","journal-title":"Nat. Cell Biol."},{"key":"B22","doi-asserted-by":"publisher","first-page":"564","DOI":"10.1038\/nature15695","article-title":"Kidney organoids from human iPS cells contain multiple lineages and model human nephrogenesis","volume":"526","author":"Takasato","year":"2015","journal-title":"Nature"},{"key":"B23","doi-asserted-by":"publisher","first-page":"394","DOI":"10.1016\/j.semnephrol.2012.06.011","article-title":"How many ways can a podocyte die?","volume":"32","author":"Tharaux","year":"2012","journal-title":"Seminars Nephrol."},{"key":"B24","doi-asserted-by":"publisher","first-page":"e20914","DOI":"10.1371\/journal.pone.0020914","article-title":"Energy metabolism in human pluripotent stem cells and their differentiated counterparts","volume":"6","author":"Varum","year":"2011","journal-title":"PLOS ONE"},{"key":"B25","doi-asserted-by":"publisher","first-page":"413","DOI":"10.1186\/s13023-022-02553-w","article-title":"A novel missense mutation in GREB1L identified in a three-generation family with renal hypodysplasia\/aplasia-3","volume":"17","author":"Wu","year":"2022","journal-title":"Orphanet J. Rare Dis."},{"key":"B26","doi-asserted-by":"publisher","first-page":"1400","DOI":"10.1681\/ASN.2015030316","article-title":"Functional human podocytes generated in organoids from amniotic fluid stem cells","volume":"27","author":"Xinaris","year":"2016","journal-title":"J. Am. Soc. Nephrol."},{"key":"B27","doi-asserted-by":"publisher","first-page":"1857","DOI":"10.1681\/ASN.2012050505","article-title":"In vivo maturation of functional renal organoids formed from embryonic cell suspensions","volume":"23","author":"Xinaris","year":"2012","journal-title":"J. Am. Soc. Nephrol."},{"key":"B28","doi-asserted-by":"publisher","first-page":"1756","DOI":"10.1038\/sj.ki.5000336","article-title":"Peroxisome proliferator-activated receptor-gamma agonist is protective in podocyte injury-associated sclerosis","volume":"69","author":"Yang","year":"2006","journal-title":"Kidney Int."},{"key":"B29","doi-asserted-by":"publisher","first-page":"C1207","DOI":"10.1152\/ajpcell.00253.2012","article-title":"A tale of two collagen receptors, integrin \u03b21 and discoidin domain receptor 1, in epithelial cell differentiation","volume":"303","author":"Yeh","year":"2012","journal-title":"Am. J. Physiology-Cell Physiology"},{"key":"B30","doi-asserted-by":"publisher","first-page":"304","DOI":"10.1681\/ASN.2018070747","article-title":"Manipulation of nephron-patterning signals enables selective induction of podocytes from human pluripotent stem cells","volume":"30","author":"Yoshimura","year":"2019","journal-title":"J. Am. Soc. Nephrol. JASN"}],"container-title":["Frontiers in Cell and Developmental Biology"],"original-title":[],"link":[{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fcell.2023.1111424\/full","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2023,3,2]],"date-time":"2023-03-02T06:53:50Z","timestamp":1677740030000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fcell.2023.1111424\/full"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2023,3,2]]},"references-count":30,"alternative-id":["10.3389\/fcell.2023.1111424"],"URL":"https:\/\/doi.org\/10.3389\/fcell.2023.1111424","relation":{},"ISSN":["2296-634X"],"issn-type":[{"value":"2296-634X","type":"electronic"}],"subject":[],"published":{"date-parts":[[2023,3,2]]},"article-number":"1111424"}}