Fig. 4

Organoid glomeruli display a maturing GBM matrisome. a Serial chemical fractionation of organoid glomeruli (OrgGlom) and organoid proximal tubules (OrgPT) to derive fractions enriched for cellular and extracellular matrix (ECM) components, where C1 and C2 are predominantly cellular proteins, C3 nuclear proteins, and C4 enriched for ECM proteins. Representative blot showing one of the three biological replicates for each cell type. b Principal component analysis of mass spectrometry data from C1 and C4 fractions of both isolated glomeruli and tubules. c Mapping of organoid proteomic data onto the human matrisome database and the identification of 60 ECM proteins (Human matrisome resource: MatrisomeDB). d, e The expression profile of matrix proteins detected in OrgGloms and OrgPTs in the ECM enriched fraction (C4) and cellular fraction (C1). Gene ontology (GO) classifications (GO terms are based on the MatrisomeDB Gene Ontology divisions and categories) were used to group those detected into core matrix proteins (d), or proteoglycans and matrix-associated proteins (e). Error bars = SEM, n = 3 biological replicates. f Immunostaining of isolated OrgGloms shows basal expression of the GBM protein Laminin-α5. Representative image from >3 biological replicates. Expression levels of single channels shown in greyscale to preserve maximum contrast, merged image shown in colour. Scale bar 10 µm. g A comparison of matrix proteins identification in human organoid glomeruli (OrgGlom) versus: human glomerular matrix (GM), immortalised human glomerular endothelial cells (ciGEnC) (doi: 10.1681/ASN.2013070795), immortalised human podocytes (ciPod) (doi: 10.1681/ASN.2013030233) and GEnC and podocyte co-culture (doi: 10.1681/ASN.2013070795) based on previously reported data. Red bar denotes presence of this protein in the sample