Journal: Kidney International Reports

Article Title: Vincristine Treatment Protects Against Podocyte Damage in Focal Segmental Glomerulosclerosis

doi: 10.1016/j.ekir.2025.08.002

Figure Lengend Snippet: Vincristine alters the podocyte transcriptome in FSGS. (a) Schematic diagram demonstrating the experimental set up for all experiments. Top 5 gene ontology results derived from PANTHER Reactome Pathways with the input of downregulated podocyte genes compared with cells exposed to presentation serum in (b) cells exposed to serum taken during vincristine treatment, and in (c) cells exposed to serum taken when the patient was in remission. (d) Venn diagram summarizing differentially expressed genes analyses. (e) Normalized cell counts from all 3 treatment groups to show differential expression of (i) tubulin alpha 1b (TUBA1B) , (ii) tubulin alpha 1c (TUBA1C) , (iii) tubulin beta 3 class III (TUBB3) , (iv) tubulin beta 6 class VI (TUBB6), (v) angiopoietin-like 4 ( ANGPTL4 ), (vi) GrpE like 1, mitochondrial (GRPEL1) , (vii) perilipin 2 ( PLIN2) , (viii) cluster of differentiation 109 ( CD109) , and (ix) family with sequence similarity 84, member B (FAM84B) . Data are shown as the mean ± SD of 4 independent experiments. For PANTHER Reactome Pathway analyses, the false discovery rate test was used to identify significantly enriched Reactome Pathways. For individual gene plots, 1- way analysis of variance with Tukey’s post hoc test was used. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001. FSGS, focal segmental glomerulosclerosis; GO, gene ontology.

Article Snippet: Cells then underwent magnetic activated cell sorting to isolate podocytes (using biotin anti-podoplanin antibody, BioLegend, 337015, conjugated to antibiotin microbeads, Miltenyi Biotec, 130-090-485) and glomerular endothelial cells (gECs, using anti-CD31 microbeads, Miltenyi Biotec, 130-091-935) before expansion in VRADD (RPMI-1640, ThermoFisher, 218750910 supplemented with 10% FBS, 1% penicillin/streptomycin, 100 nM 1,25(OH)2D3, 1 μM all trans retinoic acid and 100 nM dexamethasone) and gECs (Cell Biologics, H1168) media, respectively.

Techniques: Derivative Assay, Quantitative Proteomics, Sequencing

Journal: Kidney International Reports

Article Title: Vincristine Treatment Protects Against Podocyte Damage in Focal Segmental Glomerulosclerosis

doi: 10.1016/j.ekir.2025.08.002

Figure Lengend Snippet: Vincristine protects the podocyte cytoskeleton and cell area in FSGS. (a) Representative images of podocyte (i) centrally concentrated tubulin (arrow), (ii) dispersed tubulin (arrowhead), (iii) cortical actin fibers (arrow). Prevalence of (b) centrally concentrated tubulin, (c) dispersed tubulin, (d) cortical actin fibers, and quantification of (e) podocyte cell area relative to FBS from biological repeat after exposure to either FBS, healthy control, presentation, treatment or remission serum as well as presentation serum with 5, 100, and 330 ng/ml of vincristine. One-way analysis of variance with Dunnett’s multiple comparison tests were performed. For (b), aaa = P < 0.001 versus healthy control, bb = P < 0.01 versus presentation, bbb = P < 0.001 versus presentation. For (c–e) aaa = P < 0.001 versus FBS, bbb = P < 0.001 versus healthy control c = P < 0.05 versus presentation, ccc = P < 0.001 versus presentation. Each data point shows the mean of 50 cells. Data are shown as the mean ± SD of 4 to 8 independent experiments. FBS, fetal bovine serum; FSGS, focal segmental glomerulosclerosis; VCR, vincristine.

Article Snippet: Cells then underwent magnetic activated cell sorting to isolate podocytes (using biotin anti-podoplanin antibody, BioLegend, 337015, conjugated to antibiotin microbeads, Miltenyi Biotec, 130-090-485) and glomerular endothelial cells (gECs, using anti-CD31 microbeads, Miltenyi Biotec, 130-091-935) before expansion in VRADD (RPMI-1640, ThermoFisher, 218750910 supplemented with 10% FBS, 1% penicillin/streptomycin, 100 nM 1,25(OH)2D3, 1 μM all trans retinoic acid and 100 nM dexamethasone) and gECs (Cell Biologics, H1168) media, respectively.

Techniques: Control, Comparison

Journal: Kidney International Reports

Article Title: Vincristine Treatment Protects Against Podocyte Damage in Focal Segmental Glomerulosclerosis

doi: 10.1016/j.ekir.2025.08.002

Figure Lengend Snippet: Podocyte F-actin reorganization following presentation serum exposure was not due to IgG in patient serum. (a) Experimental design. Serum was depleted of IgG before treatment of human immortalized podocytes with either presentation serum or IgG-depleted presentation serum. (b) Representative western blot of full serum, IgG depleted serum elutions and IgG elutions after 3 repetitions of protein G spin column IgG depletion. Quantification of (c) podocyte cell area (unpaired t test). Quantification of percentage of cells displaying (d) cortical F-actin stress fibers (1-way analysis of variance with Tukey post hoc test). Each data point shows the mean of 50 cells. Data are shown as the mean ± SD of 4 independent experiments. ∗ P < 0.05. FBS, fetal bovine serum.

Article Snippet: Cells then underwent magnetic activated cell sorting to isolate podocytes (using biotin anti-podoplanin antibody, BioLegend, 337015, conjugated to antibiotin microbeads, Miltenyi Biotec, 130-090-485) and glomerular endothelial cells (gECs, using anti-CD31 microbeads, Miltenyi Biotec, 130-091-935) before expansion in VRADD (RPMI-1640, ThermoFisher, 218750910 supplemented with 10% FBS, 1% penicillin/streptomycin, 100 nM 1,25(OH)2D3, 1 μM all trans retinoic acid and 100 nM dexamethasone) and gECs (Cell Biologics, H1168) media, respectively.

Techniques: Western Blot

Journal: Kidney International Reports

Article Title: Vincristine Treatment Protects Against Podocyte Damage in Focal Segmental Glomerulosclerosis

doi: 10.1016/j.ekir.2025.08.002

Figure Lengend Snippet: Vincristine addition to presentation serum from patients with FSGS reduced albumin permeability. Representative images of GOAC seeded with primary human podocytes and gECs from 23-week postconceptional kidneys exposed to (a) FBS, (b) vincristine, 1% presentation serum from 4 different patients with FSGS (c) without and (d) with 5 ng/ml vincristine. (e) Quantification of absorbance (485 nm) of FITC-albumin flow through into urinary space channel. Each data point represents a GOAC with 4 chips used for each condition. Scale bar = 750 μm. Data are shown as mean ± SD. Kruskal-Wallis test with Dunn’s multiple comparisons post hoc test were used for statistical testing. a, P < 0.05 versus FBS; aaa, P < 0.001 versus FBS; b, P < 0.05 versus FBS + 5 ng/ml; bbb, P < 0.001 versus FBS + 5 ng/ml; c, P < 0.05 versus patient 1; ddd, P < 0.001 versus patient 2. FBS, fetal bovine serum; FSGS, focal segmental glomerulosclerosis; gECs, glomerular endothelial cells; GOAC, glomerulus-on-a-chip; VCR, vincristine.

Article Snippet: Cells then underwent magnetic activated cell sorting to isolate podocytes (using biotin anti-podoplanin antibody, BioLegend, 337015, conjugated to antibiotin microbeads, Miltenyi Biotec, 130-090-485) and glomerular endothelial cells (gECs, using anti-CD31 microbeads, Miltenyi Biotec, 130-091-935) before expansion in VRADD (RPMI-1640, ThermoFisher, 218750910 supplemented with 10% FBS, 1% penicillin/streptomycin, 100 nM 1,25(OH)2D3, 1 μM all trans retinoic acid and 100 nM dexamethasone) and gECs (Cell Biologics, H1168) media, respectively.

Techniques: Permeability

Journal: Kidney International Reports

Article Title: Vincristine Treatment Protects Against Podocyte Damage in Focal Segmental Glomerulosclerosis

doi: 10.1016/j.ekir.2025.08.002

Figure Lengend Snippet: Podocyte tubulin disorganization was found following exposure to serum from multiple patients with FSGS. Quantification of (a) centrally concentrated tubulin, (b) dispersed tubulin, (c) cortical actin fibers from 3 other patients with FSGS with 5 ng/ml of vincristine. Each data point represents the mean of 50 cells and data are shown as the mean ± SD of 4 independent experiments. One way analysis of variance with Tukey’s post hoc test was used for all statistical testing. a, P < 0.05 versus healthy control; aa, P < 0.01 versus healthy control; b, P < 0.05 versus healthy control + 5ng/ml vincristine; bb, P < 0.01 versus healthy control + 5ng/ml vincristine; c, P < 0.05 versus patient 2 without vincristine; d, P < 0.05 versus patient 3 without vincristine; dd, P < 0.05 versus patient 3 without vincristine; e, P < 0.05 versus patient 4 without vincristine; ee, P < 0.05 versus patient 4 without vincristine.

Article Snippet: Cells then underwent magnetic activated cell sorting to isolate podocytes (using biotin anti-podoplanin antibody, BioLegend, 337015, conjugated to antibiotin microbeads, Miltenyi Biotec, 130-090-485) and glomerular endothelial cells (gECs, using anti-CD31 microbeads, Miltenyi Biotec, 130-091-935) before expansion in VRADD (RPMI-1640, ThermoFisher, 218750910 supplemented with 10% FBS, 1% penicillin/streptomycin, 100 nM 1,25(OH)2D3, 1 μM all trans retinoic acid and 100 nM dexamethasone) and gECs (Cell Biologics, H1168) media, respectively.

Techniques: Control