Microbiota-derived corisin accelerates kidney fibrosis by promoting cellular aging

Abstract

The increasing global prevalence of diabetic nephropathy poses substantial health and economic burdens. Currently, effective anti-fibrotic therapies for managing kidney fibrosis associated with chronic kidney disease are lacking. This study reveals corisin, a microbiota-derived peptide, as a central driver in the progression of diabetic kidney fibrosis. Corisin levels were found to be markedly elevated in the serum of diabetic chronic kidney disease patients relative to healthy controls, with strong correlations to advanced disease stages and declining renal function. In a murine model of kidney fibrosis, corisin levels were similarly heightened, directly contributing to increased inflammation and worsening fibrosis and renal impairment. Notably, the use of a monoclonal anti-corisin antibody significantly reduced nephropathy severity in diabetic mice. Through molecular dynamics simulations and experimental validation, we demonstrated that corisin interacts with human serum albumin, potentially enhancing its renal accumulation and pathological impact. The pathogenic mechanism of corisin involves the acceleration of cellular senescence and the induction of epithelial-mesenchymal transition and apoptosis in kidney cells. These findings underscore the critical role of corisin in progressive diabetic nephropathy and suggest a promising new target for therapeutic intervention.

Fig. 1. Increased serum levels of corisin in diabetic chronic kidney disease patients.

A Thirty-five patients were enrolled in the study. Thirteen patients were excluded due to ongoing anticancer therapy, incomplete datasets, or suboptimal sample quality. B Data from 20 healthy subjects served as controls. Diabetic chronic kidney disease patients were allocated into two groups: one with stage G1 (n = 16) and another with stages G2, G3, and G4 (n = 19). Five patients with non-diabetic chronic kidney disease were also included in the study. Data are expressed as mean ± SD. Statistical significance was determined using one-way ANOVA followed by the Newman–Keuls post hoc test for comparisons among three groups. C Human corisin and corisin-like peptide sequences identified in the urine of patients with diabetic chronic kidney disease (DM-CKD) and healthy controls (HC). Staphylococcus species are the primary source of corisin in diabetic patients with chronic kidney disease. Sequences were aligned using DECIPHER, including two reference sequences (S. nepalensis: GenBank CP120099.1; S. haemolyticus: GenBank CP071512.1). Sequence IDs represent the closest species-level match, accompanied by a unique identifier corresponding to the original sequence. D A heatmap displaying read counts from sequence IDs, representing unique peptide sequences from the original amplicons, was used to normalize relative abundance across patients. The data are plotted on a log scale using phyloseq. The source data are available in the Source Data file.

Fig. 2. The administration of corisin induces acute kidney injury and exacerbation of kidney fibrosis in TGFβ1 transgenic mice with pre-existing renal injury.

A Experimental plan for inducing acute kidney injury in TGFβ1 transgenic (TG) mice with pre-existing renal dysfunction. One group of transforming growth factor β1 (TGFβ1) TG mice (TGFβ1 TG/corisin; n = 6) received 5 mg/kg of body weight of synthetic corisin by intraperitoneal injection every two days for two weeks, and another group (TGFβ1 TG/scr. peptide; n = 6) received a similar dose of scrambled peptide following the same schedule and route of administration. B Urinary corisin, albumin, and creatinine were measured as described under materials and methods. Number of mice: TGFβ1 TG/corisin, n = 6; TGFβ1 TG/scr. peptide, n = 6. Data are presented as mean ± SEM. Statistical significance was assessed using ANOVA with the Newman-Keuls test for longitudinal data and an unpaired one-sided t-test for comparisons between two groups. *p < 0.05 vs week 0; †p = 0.02 and ‡p = 0.01 vs TGFβ1 TG/scrambled peptide group. C The urinary levels of kidney injury molecule-1 (KIM-1), liver-type fatty acid-binding protein (L-FABP), blood urea nitrogen (BUN), and creatinine were measured as described under material and methods. Number of mice: TGFβ1 TG/corisin, n = 6; TGFβ1 TG/scr. peptide, n = 6. Data are presented as mean ± SD. Statistical significance was determined using a two-sided unpaired t-test. D Plasma levels of lipopolysaccharide-induced CXC chemokine (LIX/CXCL5), macrophage inflammatory protein-2 (MIP-2), interleukin-1β (IL-1β), platelet-derived growth factor (PDGF), tissue factor (TF), and plasminogen activator inhibitor-1 (PAI-1) were measured using enzyme immunoassays. Normally distributed data are presented as mean ± SD, while skewed data are expressed as the median with interquartile range. Number of mice: TGFβ1 TG/corisin, n = 6; TGFβ1 TG/scr. peptide, n = 6. Statistical significance was assessed using a two-sided unpaired t-test for normally distributed data and the two-sided Mann-Whitney U test for skewed data. E–I Masson’s trichrome and periodic acid–Schiff staining of renal tissues from both groups of mice. Renal fibrosis was quantified using WinROOF imaging software. Scale bars indicate 100 µm in (D) and 50 µm in (F). Number of mice: TGFβ1 TG/corisin, n = 6; TGFβ1 TG/scr. peptide, n = 6. Data are presented as mean ± SD. Statistical significance was determined using a two-sided unpaired t-test. The source data are available in the Source Data file.

Fig. 3. Treatment with the anticorisin antibody inhibits the progression of kidney fibrosis under diabetic conditions.

A Experimental plan. Diabetic TGFβ1 TG mice were divided into the following groups. One group (DM TG/anticorisin) received 10 mg/kg of body weight of anticorisin mAb by intraperitoneal injection three times a week for eight weeks, while another group (DM TG/control IgG) received a similar dose of IgG control. Following the same sampling schedule, a wild-type (WT) (n = 4) group was also included. B Blood glucose levels throughout the experiment, intraperitoneal glucose tolerance test (IPGTT), plasma creatinine, blood urea nitrogen (BUN), urine albumin/creatinine ratio, and urinary liver-type fatty acid-binding protein (L-FABP) levels. n = 5 for DM TG/control IgG and DM TG/anti-corisin; n = 4 for WT/SAL. Normally distributed data are presented as the mean, while skewed data are presented as the median. Statistical significance was assessed using ANOVA followed by either the Newman-Keuls or Dunn’s post hoc test, as appropriate; all tests were two-sided. †p < 0.05 vs WT/SAL across the same week. C, D Paraffin-embedded kidney tissue samples for collagen staining with trichrome acid. n = 5 in DM TG/control IgG and DM TG/anticorisin groups, and n = 4 in the WT/SAL group. Renal fibrosis was quantified using WinROOF. Scale bars represent 500 µm. Data are presented as mean ± SD. Statistical significance was assessed using ANOVA and the Newman-Keuls test; all tests were two-sided. E, F Paraffin-embedded kidney tissue samples for Periodic acid-Schiff (PAS) staining. Scale bars represent 50 µm. n = 5 in DM TG/control IgG and DM TG/anticorisin groups, and n = 4 in the WT/SAL group. Data are presented as mean ± SD. Statistical significance was assessed using ANOVA and the Newman-Keuls test; all tests were two-sided. G, H The plasma levels of lipopolysaccharide-induced CXC chemokine/C-X-C motif chemokine 5 (LIX/CXCL5), interleukin-1β (IL-1β), matrix metalloproteinase-2 (MMP-2), platelet-derived growth factor (PDGF), total and active transforming growth factor β1 (TGFβ1), connective tissue growth factor (CTGF), and collagen I were measured by enzyme-linked immune assays. n = 11 in DM TG/control IgG, n = 10 in DM TG/anticorisin groups, and n = 4 in the WT/SAL group. Normally distributed data are presented as mean ± SD, while skewed data are expressed as the median with interquartile range. Statistical significance was assessed using ANOVA followed by the Newman-Keuls or Dunn’s post hoc test, as appropriate; all tests were two-sided. The source data are available in the Source Data file.

Fig. 4. Corisin interacts with serum human albumin.

A, B The predicted interaction structure of human serum albumin-corisin and bovine serum albumin-corisin by molecular dynamic simulations. Human serum albumin and bovine serum albumin are shown in surface representation, and domains I, II, and III are shown in pink, yellow, and ice blue, respectively. The corisin is shown in licorice representation. C, D Western blotting showing the interaction of corisin with recombinant human albumin (rhAlb) in vitro. A fixed concentration of rhAlb was combined with concentrations of synthetic corisin or scrambled peptide and incubated at room temperature for 10 min. The mixture underwent sodium dodecyl-sulfate polyacrylamide gel electrophoresis (SDS-PAGE), followed by Western blotting using anti-corisin monoclonal antibody (mAb) or anti-rhAlb antibody. The experiment was independently repeated three times with similar results. E Co-immunoprecipitation of corisin with rhAlb. Synthetic corisin was incubated with rhAlb, treated with an anti-corisin mAb, and immunoprecipitated using protein G agarose beads. rhAlb was not immunoprecipitated by the anti-corisin mAb and protein G agarose beads alone (lane 5). In the presence of corisin, rhAlb was successfully co-immunoprecipitated by the anti-corisin mAb and protein G agarose beads (lane 6). The experiment was independently repeated three times with similar results. Ig, immunoglobulin; MW, molecular weight. F Corisin interacts with human and mouse serum albumin. A predetermined concentration of human serum albumin (HSA) or mouse serum albumin (MSA) was incubated with synthetic corisin. The mixture was subjected to SDS-PAGE and Western blotting using an anti-corisin monoclonal antibody. The experiment was independently repeated three times with similar results. G Corisin interacts with human serum albumin derived from both patients with diabetic nephropathy and healthy individuals. Serum samples were diluted (1:20) and incubated for 10 min with corisin (4 µg) diluted in saline. As a control, a mixture containing corisin (4 µg) and rhAlb (5 µg) was prepared. Each mixture was then subjected to electrophoresis, followed by Western blot analysis using mAb specific for corisin or human albumin. The experiment was independently repeated three times with similar results. H, I Complex formation between corisin and albumin in the urine of diabetic CKD patients. Urine samples from 3 diabetic patients with chronic kidney disease (CKD) and 3 healthy subjects were concentrated 5 times, and 5 µL of each sample were subjected to SDS-PAGE, followed by Western blotting using an anti-corisin mAb (left panel) or anti-human albumin antibody (right panel). DM diabetes mellitus.

Fig. 5. Corisin penetrates kidney cells via the human albumin receptor cubilin to target mitochondria.

A–D, Human Caki-2 cell lines and normal human primary podocytes were cultured to near confluence, then treated with fluorescein isothiocyanate (FITC)-labeled corisin (corisin-FITC) or FITC-labeled scrambled peptide for 4 h. MitoTracker was added 30 min before fixation to label mitochondria, and cells were subsequently imaged via microscopy. The fluorescence intensities of F-actin and DAPI were quantified using ImageJ. Scale bars represent 20 µm. n = 8 replicates. Data are presented as mean ± SD. Statistical analysis was conducted using a two-sided unpaired t-test. E–G, Normal human primary podocytes were cultured for 48 h with siRNA targeting CD36, megalin, cubilin, SPARC, and FCGRT, and the relative expression of albumin receptors was evaluated. n = 4 replicates. Data are expressed as the mean ± SD. Statistical comparison between each receptor-specific siRNA and its corresponding control siRNA was performed using a two-sided unpaired t-test. In a separate experiment, cells were cultured under the same conditions and then treated with corisin-FITC, MitoTracker, and DAPI and observed using fluorescence microscopy. The fluorescence intensities of F-actin and DAPI were quantified using ImageJ, an open-source software from the National Institutes of Health (NIH). Scale bars represent 20 µm. n = 8 replicates. n = 12 in control and n = 6 in remaining groups. Data are presented as mean ± SD. Statistical analysis was conducted using ANOVA followed by Dunnett’s test; all tests were two-sided. The source data are available in the Source Data file.

Fig. 6. Corisin induces increased expression of senescence markers in kidney cells.

A–F Induction of senescence-associated β-galactosidase (SAβGal) activity by corisin. Human Caki-2 cells (left panels), human primary renal tubular epithelial cells (middle panels), and normal human primary podocytes (right panels) were cultured in the absence or presence of 20 or 40 µg/mL of corisin, diluted in 0.5% recombinant human albumin (rhAlb), or 40 µg/mL of corisin with anti-corisin antibody. SAβGal activity was measured, and %SAβGal-positive cells was counted across multiple high-power fields using immunofluorescence microscopy. Scale bars represent 50 µm. Control, n = 6; corisin (20 µg/mL), n = 6; corisin (40 µg/mL), n = 6; corisin (40 µg/mL) + anticorisin (200 µg/mL), n = 6. Data are expressed as the mean ± SD. Statistical analysis by ANOVA followed by the Newman-Keuls test; all tests were two-sided. G–H Increased mRNA expression of senescence-associated factors in kidney cells by corisin. Normal human podocytes (n = 6) and normal human primary tubular epithelial cells (n = 5) were cultured in the absence or presence of 20 and 40 µg/mL corisin, diluted in 0.5% recombinant human albumin, or 40 µg/mL of corisin with anti-corisin antibody for 48 h. mRNA expression levels of cyclin-dependent kinase inhibitors p15 (CDKN2), p16 (CDKN2A), p21 (CDKN1A), p27 (CDKN1B), p53 (TP53), Ki-67 (MKI67), matrix metalloproteinase-12 (MMP12), and secreted phosphoprotein 1 (SPP1, also known as osteopontin) were evaluated by RT-PCR. Normally distributed data are presented as mean ± SD, while skewed data are expressed as the median with interquartile range. Statistical significance was assessed using ANOVA with the Newman-Keuls or Dunn’s test; all tests were two-sided. I, J Anticorisin antibody inhibits the expression of senescence markers. Diabetes mellitus (DM) was induced in transforming growth factor β1 (TGFβ1) transgenic (TG) mice by streptozotocin. The mice were divided into a DM TG/control IgG group (n = 5) and a DM TG/anticorisin group (n = 5). Wild-type (WT) mice injected intraperitoneally with saline (SAL, n = 4) served as controls. Paraffin-embedded kidney tissue sections were prepared for p21 immunofluorescent staining. Green immunofluorescent signals (red arrows) indicate p21 expression. The p21-positive area was quantified using WinROOF. Scale bars represent 100 µm. Data are presented as mean ± SD. Statistical significance was evaluated by ANOVA followed by the Newman-Keuls test; all tests were two-sided. The source data are available in the Source Data file.

Fig. 7. Corisin induces epithelial-mesenchymal transition in podocytes and renal tubular epithelial cells.

A–D Normal human primary podocytes or primary renal proximal tubular epithelial cells (RPTEC) were cultured for 48 h under conditions without corisin (medium containing 0.5% recombinant human albumin) and with corisin at concentrations of 20 and 40 µg/mL, dissolved in 0.5% recombinant human albumin, or 40 µg/mL of corisin with anti-corisin antibody. Subsequently, cells were stained with Phalloidin-iFluor™ 488 and 4’,6-diamidino-2-phenylindole (DAPI). Fluorescence intensity of F-actin and cell counts were quantified using ImageJ, a public domain software from the National Institutes of Health (NIH). n = 4 in experiments using podocytes and n = 6 in experiments using RPTEC. Scale bars indicate 20 µm. Data are expressed as the mean intensity per cell ratio ± SD. Statistical analysis was performed using ANOVA followed by the Newman-Keuls test; all tests were two-sided. E, F Cells were cultured under the same conditions to collect total RNA from each treatment group and assess the mRNA expression of α-smooth muscle actin (ACTA2), fibronectin (FN1), collagen I (COL1a1), and transforming growth factor β1 (TGFB1). n = 5 in (E) and n = 6 in (F). Normally distributed data are presented as mean ± SD, while skewed data are expressed as the median with interquartile range. Statistical significance was assessed using ANOVA, followed by the Newman-Keuls or Dunn’s test; all tests were two-sided. The source data are available in the Source Data file.

Fig. 8. Corisin induces the expression of senescence-associated factors in primary renal proximal tubular epithelial cells.

A Renal proximal tubular epithelial cells (RPTEC) were cultured for 24 h in the presence of corisin or scrambled peptides and subsequently subjected to single-cell RNA sequencing analysis. B Violin plots and Uniform Manifold Approximation and Projection (UMAP) plots depict the expression of senescence markers and senescence-associated secretory phenotype (SASP) components in corisin- and scrambled peptide-treated cells. Statistical analysis was conducted using a two-sided Mann–Whitney U test. C Violin plots and UMAP plots illustrating the expression of epithelial-mesenchymal transition (EMT) markers in corisin- and scrambled peptide-treated cells. Statistical analysis was conducted using a two-tailed Mann-Whitney U test. D Additional senescence, SASP, and EMT markers in corisin- and scrambled peptide-treated cells. Statistical analysis was conducted using a two-sided Mann-Whitney U test. For panels B, C, and D, single-cell RNA sequencing was performed using three independent RPTEC cultures per stimulant, yielding 9327 high-quality cells for the corisin-treated group and 8186 cells for the scrambled peptide (control) group. Violin plots in (B–D), represent the distribution of gene expression values. The width of each violin corresponds to the kernel density estimation. The thick vertical bar within each violin denotes the interquartile range (25th-5th percentile), and the thin line (whisker) extends to the minimum and maximum values within 1.5 times the interquartile range. The median is not explicitly displayed.

Fig. 9. Corisin stimulation in primary renal proximal tubular epithelial cells promotes pathways associated with the DNA damage response, cellular senescence, the senescence-associated secretory phenotype (SASP), and myofibroblast-like differentiation.

A Heatmap showing gene expression in cells stimulated with corisin or a scrambled peptide control. B Volcano plots showing differentially expressed genes between corisin- and scrambled peptide-treated renal proximal tubular epithelial cells (RPTEC), based on single-cell RNA sequencing. Each point represents a gene, with the x-axis showing the log₂ fold change and the y-axis showing the -log₁₀ adjusted p-value. Differential expression analysis was performed using a two-tailed Mann–Whitney U test. Since gene expression was assessed across thousands of genes, p-values were adjusted for multiple comparisons using the Benjamini-Hochberg method to control for the false discovery rate. C Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis reveals significant overrepresentation of pathways related to cellular senescence in corisin-treated renal proximal tubular epithelial cells. Enrichment analysis was performed using the hypergeometric test, and p-values were adjusted for multiple comparisons using the Benjamini-Hochberg method to control the false discovery rate. D Categorization of differentially expressed genes by their biological roles, demonstrating their involvement in various cellular processes.

Fig. 10. Corisin-induced kidney cell damage and fibrosis in diabetic chronic kidney disease and the protective potential of anticorisin monoclonal antibody.

Diabetes-associated dysbiosis increases corisin release from the microbiome into systemic circulation, where it binds to serum albumin. The corisin-albumin complex reaches the glomeruli and proximal tubular epithelial cells, binding to cubilin, an albumin receptor, and thereby facilitating corisin entry into podocytes and proximal tubular epithelial cells. Within these cells, corisin induces a senescence-associated secretory phenotype, resulting in the elevated secretion of inflammatory cytokines, chemokines, matrix metalloproteinases, and growth factors that promote inflammation, epithelial-mesenchymal transition, apoptosis of podocytes and tubular epithelial cells, myofibroblast recruitment, and extracellular matrix deposition (e.g., collagen I). Areas of the glomeruli and tubules affected by increased apoptosis are subsequently replaced by fibrotic tissue, accelerating disease progression and leading ultimately to a fatal outcome. The anti-corisin monoclonal antibody binds to corisin peptides, blocking their pro-senescence activity and mitigating disease progression. mAb, monoclonal antibody.