Extended Data Fig. 4: Loss of Ecm1 in fibroblasts alleviates kidney fibrosis after IRI and UUO. | Nature Metabolism

Extended Data Fig. 4: Loss of Ecm1 in fibroblasts alleviates kidney fibrosis after IRI and UUO.

From: Early-activated extracellular matrix proteins shape the metabolic and spatial dynamics of the kidney fibrotic microenvironment

Extended Data Fig. 4: Loss of Ecm1 in fibroblasts alleviates kidney fibrosis after IRI and UUO.The alternative text for this image may have been generated using AI.

(a) PCR-based genotyping of mice genomic DNA. Lane 1: Col1a2+ fibroblast-specific Ecm1 conditional knockout (cKO) mice (genotype: Ecm1fl/fl, Cre), designated as Col1α2-Ecm1-/-. Lane 2: wild type (WT) mice (genotype: Ecm1fl/fl), designated as Col1α2-Ecm1+/+. (b) PCR genotyping of Pdgfrb+ fibroblast-specific Ecm1 knockout mice. Lane 1: Pdgfrb-Ecm1+/+ (WT) mice (genotype: Ecm1fl/fl). Lane 2: Pdgfrb-Ecm1-/- (cKO) mice (genotype: Ecm1fl/fl, Cre). (c) Scr levels in normal Col1α2-Ecm1+/+ (n = 5) versus Col1α2-Ecm1-/- mice (n = 5) (upper) and Pdgfrb-Ecm1+/+ (n = 5) versus Pdgfrb-Ecm1-/- mice (lower) (n = 4). Age- (8–10 weeks) and sex-matched littermates were used. (d) Kidney weight (KW) to body weight (BW) ratio in Col1α2-Ecm1+/+ (n = 5) versus Col1α2-Ecm1-/- (n = 5) mice (upper) and Pdgfrb-Ecm1+/+ (n = 5) versus Pdgfrb-Ecm1-/- (n = 4) mice (lower). (e) PAS staining of both strains of WT and cKO kidneys. (f) qPCR of Ecm1 mRNA in whole kidneys from both strains at 2 weeks after IRI. (g) Immunohistochemistry of kidney ECM1 expression in both strains after unilateral IRI at 2 weeks. (h) Quantitative data of fibronectin (FN) and α-SMA protein levels in Col1α2-Ecm1+/+ and Col1α2-Ecm1-/- kidneys following unilateral IRI (upper, n = 6) or UUO (lower, n = 5). (i) Quantitative data of MTS and immunohistochemistry (COL1A1, FN, CD45) in Col1α2-Ecm1+/+ and Col1α2-Ecm1-/- kidneys after unilateral IRI and UUO (Sham, n = 3; IRI, n = 5; UUO, n = 5). (j) Quantitative data of ATP5A, UQCRC2, MTCO1, SDHB, and NDUFB8 protein levels in Col1α2-Ecm1+/+ and Col1α2-Ecm1-/- kidneys after unilateral IRI (n = 6). (k) Western blot analysis of mitochondrial OXPHOS complexes I-V in Col1α2-Ecm1+/+ and Col1α2-Ecm1-/- kidneys after UUO. Numbers represent individual animals in each group. (l) Quantitative data of SDHB and NDUFB8 protein levels in Col1α2-Ecm1+/+ and Col1α2-Ecm1-/- kidneys after UUO (n = 5). (m) Quantitative data of FN and α-SMA protein levels in Pdgfrb-Ecm1+/+ and Pdgfrb-Ecm1-/- kidneys after unilateral IRI (n = 5). (n, o) Representative micrographs of COL1A1 staining (n) and quantitative analysis (o) in Pdgfrb-Ecm1+/+ and Pdgfrb-Ecm1-/- kidneys after unilateral IRI (Sham, n = 3; IRI, n = 5). (p, q) Quantitative data of MTS and CD45 staining in Pdgfrb-Ecm1+/+ and Pdgfrb-Ecm1-/- kidneys after unilateral IRI (Sham, n = 3; IRI, n = 5). (r) Quantitative data of UQCRC2, MTCO1, and SDHB protein levels in Pdgfrb-Ecm1+/+ and Pdgfrb-Ecm1-/- kidneys after IRI (n = 5). (s-v) Western blot analysis of FN in male (s) and female (t) Col1α2-Ecm1+/+ and Col1α2-Ecm1-/- kidneys and male (u) and female (v) Pdgfrb-Ecm1+/+ and Pdgfrb-Ecm1-/- kidneys after bilateral IRI at 28 days. Quantitative data is shown on the right of each panel (n = 6). Scale bar, 25 µm. Data are presented as mean ± s.e.m. Statistical differences were assessed using two-sided unpaired t-tests or one-way ANOVA followed by the Student-Newman-Keuls test.

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