rat renal interstitial fibroblast nrk49f cells  (ATCC)

Journal: Renal Failure

Article Title: Ubiquitin-specific protease 11 facilitates the activation and proliferation of renal interstitial fibroblasts through epidermal growth factor receptor signaling pathways

doi: 10.1080/0886022X.2026.2666452

Figure Lengend Snippet: USP11 is highly upregulated in UA-stimulated RIF. In vitro , we stimulated NRK49F with UA in dose-dependent manner (0, 200, 400, 800 μM) and in time-dependent manner (0, 12, 24, 36 hours). Western blot analysis of USP11, α-SMA and Collagen I protein expressions in each group were conducted, with normalization to GAPDH (A, C). Quantitative analysis of USP11, α-SMA and Collagen I protein levels in each group (B, D). Data were expressed as mean± SD ( n = 4 for each group). N.S.: no significant difference. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: Rat renal interstitial fibroblast (NRK49F) cells were obtained from ATCC (Manassas, VA).

Techniques: In Vitro, Western Blot

Journal: Renal Failure

Article Title: Ubiquitin-specific protease 11 facilitates the activation and proliferation of renal interstitial fibroblasts through epidermal growth factor receptor signaling pathways

doi: 10.1080/0886022X.2026.2666452

Figure Lengend Snippet: USP11 contributes to RIF activation in the UA-stimulated NRK49F. NRK49F cells were seeded to 70–80% confluence in the antibiotic-free medium and grown followed by transfection with USP11 siRNA or USP11-pcDNA 3.0 plasmid. After transfection, the medium was changed to DMEM with F12 containing 0.5% FBS for starvation and then cells were incubated with or without uric acid (800 μM) for an additional 36 hours before being harvested for analysis. Western blot analysis of USP11, α-SMA and Collagen I protein expressions in each group were conducted, with normalization to GAPDH (A, F, H). Quantitative analysis of USP11, α-SMA and Collagen I protein levels in each group (B, G, I). Representative images and quantitative analysis of immunofluorescence staining for Fibronectin and Vimentin with DAPI nuclear counterstaining in each group (C). Then NRK49F cells were starved for 24 h with DMEM containing 0.5% FBS before they were exposed to uric acid in the presence or absence of MTX (1, 5, and 10 μM). Western blot analysis of USP11, α-SMA and Collagen I protein expressions in each group were conducted, with normalization to GAPDH (D). Quantitative analysis of USP11, α-SMA and Collagen I protein levels in each group (E). Data were expressed as mean± SD ( n = 4 for each group). N.S.: no significant difference. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. Scale bars = 100μm.

Article Snippet: Rat renal interstitial fibroblast (NRK49F) cells were obtained from ATCC (Manassas, VA).

Techniques: Activation Assay, Transfection, Plasmid Preparation, Incubation, Western Blot, Immunofluorescence, Staining

Journal: Renal Failure

Article Title: Ubiquitin-specific protease 11 facilitates the activation and proliferation of renal interstitial fibroblasts through epidermal growth factor receptor signaling pathways

doi: 10.1080/0886022X.2026.2666452

Figure Lengend Snippet: USP11 directly interacts with EGFR and stabilizes its protein level. Immunofluorescence co-staining of USP11 and EGFR in the UA-stimulated NRK49F (A). Co-IP assay in NRK49F cells. Flag-tagged USP11 and Myc-tagged EGFR were immunoprecipitated using an anti-Flag/Myc antibody, and the presence of Myc-tagged EGFR and Flag-tagged USP11 in the precipitates was detected by immunoblotting (B, C). Western blot analysis of USP11 and EGFR protein expressions in each group were conducted, with normalization to GAPDH (D). Quantitative analysis of EGFR protein levels in each group (E). Data were expressed as mean± SD ( n = 4 for each group). N.S.: no significant difference, *** p < 0.001. Scale bars = 50μm.

Article Snippet: Rat renal interstitial fibroblast (NRK49F) cells were obtained from ATCC (Manassas, VA).

Techniques: Immunofluorescence, Staining, Co-Immunoprecipitation Assay, Immunoprecipitation, Western Blot

Journal: Renal Failure

Article Title: Ubiquitin-specific protease 11 facilitates the activation and proliferation of renal interstitial fibroblasts through epidermal growth factor receptor signaling pathways

doi: 10.1080/0886022X.2026.2666452

Figure Lengend Snippet: USP11 activates EGFR signaling pathway in the UA-stimulated NRK49F. Western blot analysis of p-EGFR and EGFR protein expressions in each group were conducted, with normalization to GAPDH (A, C, E). Quantitative analysis of p-EGFR and EGFR protein levels in each group (B, D, F). Data were expressed as mean ± SD (n = 4 for each group). N.S.: no significant difference. *** p < 0.001, **** p < 0.0001.

Article Snippet: Rat renal interstitial fibroblast (NRK49F) cells were obtained from ATCC (Manassas, VA).

Techniques: Western Blot

Journal: Renal Failure

Article Title: Ubiquitin-specific protease 11 facilitates the activation and proliferation of renal interstitial fibroblasts through epidermal growth factor receptor signaling pathways

doi: 10.1080/0886022X.2026.2666452

Figure Lengend Snippet: USP11 is involved in the activation of EGFR signaling pathway in the UA-stimulated NRK49F. To fully demonstrate the relationship between USP11 and EGFR signaling, we stimulated starved NRK49F cells with EGF (5 ng/ml) in the presence or absence of USP11 siRNA or treated with gefitinib (1 nM and 5 nM), a highly selective EGFR inhibitor in the presence of USP11 pcDNA 3.0 plasmid for an additional 36 hours. Western blot analysis of p-EGFR and EGFR protein expressions in each group were conducted, with normalization to GAPDH (A, E). Quantitative analysis of p-EGFR and EGFR protein levels in each group (B, F). Western blot analysis of USP11, α-SMA and Collagen I protein expressions in each group were conducted, with normalization to GAPDH (C, G). Quantitative analysis of USP11, α-SMA and Collagen I protein levels in each group (D, H). Data were expressed as mean ± SD ( n = 4 for each group). N.S.: no significant difference. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: Rat renal interstitial fibroblast (NRK49F) cells were obtained from ATCC (Manassas, VA).

Techniques: Activation Assay, Plasmid Preparation, Western Blot

Journal: Renal Failure

Article Title: Ubiquitin-specific protease 11 facilitates the activation and proliferation of renal interstitial fibroblasts through epidermal growth factor receptor signaling pathways

doi: 10.1080/0886022X.2026.2666452

Figure Lengend Snippet: Inhibition of USP11 attenuates the proliferation and migration in the UA-stimulated NRK49F. Photomicrographs of migrating cells in wound healing assay were taken at 0h and 36h (A). The migratory rate was calculated as (A-B)/A*100%, where A and B reflect the width of the wound at 0h and 36h respectively (B). The CCK-8 proliferation kit was used according to the manufacturer’s instructions and the final optical density values were read at 450 nm (C). Representative images and quantitative analysis of immunofluorescence staining for Ki67 with DAPI nuclear counterstaining in each group (D, E). Western blot analysis of PCNA and Cyclin E protein expressions in each group were conducted, with normalization to GAPDH (F, H). Quantitative analysis of PCNA and Cyclin E protein levels in each group (G, I). Data were expressed as mean± SD ( n = 4 for each group). N.S., no significant difference. * p < 0.05, *** p < 0.001, **** p < 0.0001. Scale bars = 500μm (A) and 50μm (D).

Article Snippet: Rat renal interstitial fibroblast (NRK49F) cells were obtained from ATCC (Manassas, VA).

Techniques: Inhibition, Migration, Wound Healing Assay, CCK-8 Assay, Immunofluorescence, Staining, Western Blot

Journal: Journal of Advanced Research

Article Title: Direct pharmacological targeting of Piezo1 by Paeoniflorin: a novel therapeutic approach for renal fibrosis

doi: 10.1016/j.jare.2025.07.015

Figure Lengend Snippet: PF inhibits matrix stiffness-induced acceleration of EndMT through Piezo1 activation (A–F) Matrix stiffness-dependent protein modulation in HUVECs. HUVECs were cultured on polyacrylamide hydrogels with soft (1.00 ± 0.31 kPa) or stiff (40.40 ± 2.39 kPa) for 48 h, with or without PF. (A) Western blot analysis of (B) Piezo1, (C) VE-Cadherin, (D) eNOS, (E) Vimentin, and (F) TGF-β1. Quantification normalized to GAPDH. (n = 3; *p < 0.05, **p < 0.01 vs. soft; #p < 0.05 vs. stiff without PF). (G-L) Piezo1 knockdown reverses stiffness-induced EndMT. HUVECs transfected with Piezo1 siRNA (50 nM, 24 h) or scramble siRNA (control) were cultured on stiffness hydrogels (40.40 ± 2.39 kPa) ± PF (400 μM). (G) Western blot analysis of (H) Piezo1, (I) VE-Cadherin, (J) eNOS, (K) Vimentin, and (L) TGF-β1. Quantification normalized to GAPDH (n = 3; *p < 0.05, **p < 0.01 vs. scramble siRNA control; ns vs. Piezo1 siRNA without PF).(M) Schematic of co-culture model. HUVECs and NRK-49F fibroblasts were co-cultured on stiffness-tunable hydrogels using a transwell system (0.4 μm pore size) for 5 days to assess paracrine signaling. (N) TGF-β1 levels in HUVEC supernatants from the co-culture system were quantified by ELISA. (n = 3; *p < 0.05 vs. softness; #p < 0.05 vs. stiffness without PF). (O) Fibrotic gene expression in NRK-49F cells. RT-qPCR analysis of Fibronectin , COL1A1 , Vimentin , and TGF-β1 mRNA levels. Data normalized to 18 s ( n = 3). (P-S) Fibrotic protein expression in NRK-49F cells. (P) Western blot analysis of (Q) Fibronectin, (R) COL1, (S) Vimentin, and (T) TGF-β1. Quantification normalized to GAPDH (n = 3). (U–V) Immunofluorescence of Fibronectin (red) in NRK-49F cells. Nuclei stained with DAPI (blue). (U) Representative images. (V) Quantification of fluorescence intensity using ImageJ (n = 3). Scale bar: 20 μm. Data presented as mean ± SEM. *p < 0.05, **p < 0.01 vs . softness co-culture; # p < 0.05 vs . stiffness co-culture without PF . (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: Human umbilical vein endothelial cells (HUVECs) and rat kidney fibroblast cell line (NRK-49F) were purchased from ATCC (American Type Culture Collection, https://www.atcc.org ).

Techniques: Activation Assay, Cell Culture, Western Blot, Knockdown, Transfection, Control, Co-Culture Assay, Pore Size, Enzyme-linked Immunosorbent Assay, Gene Expression, Quantitative RT-PCR, Expressing, Immunofluorescence, Staining, Fluorescence

Journal: Journal of Advanced Research

Article Title: Direct pharmacological targeting of Piezo1 by Paeoniflorin: a novel therapeutic approach for renal fibrosis

doi: 10.1016/j.jare.2025.07.015

Figure Lengend Snippet: PF inhibits EndMT through the Piezo1-mediated HIF-1α signaling pathway (A–C) Renal HIF-1α expression analysis. (A) Representative RT-qPCR analysis of HIF-1α mRNA levels in kidney tissues. Data normalized to 18 s . (B-C) Western blot and quantification of HIF-1α protein expression in renal tissues. Data normalized to GAPDH (n = 5–6) . (D–H) Effects of PF or HIF-1α inhibitor BAY 87-2243 (10 μM, 24 h) on endothelial markers in HUVECs cultured with or without Piezo1 activation by Yoda1 (5 μM, 12 h). (D) Western blot analysis of (E) Piezo1, (F) HIF-1α, (G) VE-Cadherin, and (H) eNOS. Quantification normalized to GAPDH. Quantification showing Yoda1-induced Piezo1 upregulation and HIF-1α/VE-Cadherin/eNOS downregulation, reversed by PF or BAY 87-2243 (n = 3). (I-K) PF or BAY 87-2243 inhibits Yoda1-induced EndMT in HUVECs. (I) Western blot analysis of (J) Vimentin and (K) TGF-β1. Yoda1 increased Vimentin and TGF-β1, suppressed by PF or BAY 87-2243 (n = 3). (L) Schematic of HUVEC-NRK-49F co-culture. HUVECs pre-treated with/without Yoda1 (5 μM, 6 h) were co-cultured with NRK-49F fibroblasts for 48 h. (M) TGF-β1 levels in HUVEC supernatants from the co-culture system were quantified by ELISA. (N) RT-qPCR analysis of Fn1 , COL1A1 , and Vimentin mRNA in NRK-49F cells co-cultured with Yoda1-treated HUVECs. PF attenuated Yoda1-induced fibrotic marker expression (n = 3). (O-S) Western blot validation of (P) Fibronectin, (Q) COL1, (R) Vimentin, and (S) TGF-β1 in NRK-49F cells. PF reduced Yoda1-induced protein expression (n = 3). Data presented as mean ± SEM. *p < 0.05, ** p < 0.01, *** p < 0.001 vs. CTL/control; # p < 0.05, ## p < 0.01, ### p < 0.001 vs. CRF/Yoda1.

Article Snippet: Human umbilical vein endothelial cells (HUVECs) and rat kidney fibroblast cell line (NRK-49F) were purchased from ATCC (American Type Culture Collection, https://www.atcc.org ).

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Cell Culture, Activation Assay, Co-Culture Assay, Enzyme-linked Immunosorbent Assay, Marker, Biomarker Discovery, Control

Journal: Research

Article Title: Epigenetic Suppression of RASAL1 by HDAC3 and Cofactor YY1 Promotes Fibroblast–Myofibroblast Transition and Renal Fibrosis

doi: 10.34133/research.1073

Figure Lengend Snippet: Fibrotic kidneys induced by UUO or AAI display aberrant up-regulation of HDAC3 and suppression of RASAL1 in renal fibroblasts. C57BL/6 mice were subjected to UUO for 7 d or AAI (5 mg/kg, intraperitoneal injection every other day) for 14 d (6 mice in each group). (A) Representative photomicrographs of kidney sections stained with Masson’s trichrome and Sirius Red from sham-, UUO-, control-, and AAI-treated mice. The arrows indicate collagen-stained fibrotic areas. (B) Quantitation of renal fibrosis in (A). (C) Western blots of renal tissues for Colla 1, α-SMA, HDAC3, and RASAL1. Two representative samples from each group were shown. β-Actin served as loading control. (D) Quantifications of (C). (E) RT-PCR of renal tissues from sham-, UUO-, control-, and AAI-treated mice ( n = 6) for Rasal1 mRNAs. Gapdh served as the internal control. Two random samples from each group were shown. The right panel was the quantification. bp, base pairs. Data were presented as means ± SEM based on 6 renal samples. * P = 0.05, 2-tailed unpaired t test for (B), (D), and (E). (F) Representative photomicrographs of kidney sections from sham-, UUO-, control-, and AAI-treated mice stained for RASAL1 by IHC staining. Positive staining was indicated by arrows. (G) Western blotting. NRK-49F and mouse primary renal fibroblast (PRF) cells were treated with AAI (30 μM) for 24 h, and then the cell lysates were tested for HDAC3 and RASAL1. The quantitation was on the right side. Data were presented as means ± SD of 6 repeated cell assays. *P < 0.05, 2-tailed unpaired t test. (H) Representative photomicrographs of kidney sections from renal patients (mild injury and CKD) stained by Masson’s trichrome for renal fibrosis and IHC for HDAC3 and RASAL1. The arrows indicate collagen-stained fibrotic areas and positively stained fibroblast-rich areas. (I) Quantifications of (H) presented as means ± SEM of 6 mild injury and 10 CKD samples. Data were presented as means ± SEM based on 6 renal samples. * P < 0.05, 2-tailed unpaired t test.

Article Snippet: Human embryonic kidney (HEK) 293T cells (American Type Culture Collection, USA) were cultured in DMEM, and rat renal fibroblast NRK-49F cells (Procell Life Science & Technology, Wuhan, China) and mouse primary renal fibroblasts were cultured in DMEM/F12 medium.

Techniques: Injection, Staining, Control, Quantitation Assay, Western Blot, Reverse Transcription Polymerase Chain Reaction, Immunohistochemistry

Journal: Research

Article Title: Epigenetic Suppression of RASAL1 by HDAC3 and Cofactor YY1 Promotes Fibroblast–Myofibroblast Transition and Renal Fibrosis

doi: 10.34133/research.1073

Figure Lengend Snippet: HDAC3 inhibits RASAL1 transcription in renal fibroblasts. (A) NRK-49F cells were transfected with either a control plasmid (vector) or a plasmid overexpressing Flag-tagged HDAC3 (F-HD3). Twenty-four hours later, the cell lysates were assayed for HDAC3, RASAL1, and Colla 1. β-Actin served as the internal control. (B) NRK-49F cells were treated with TGFβ (10 ng/ml) in the presence or absence of RG (10 μM) for 24 h, and then the cell lysates were tested for HDAC3, RASAL1, and Colla 1. (C) Quantitation of Western blots in (B). (D) Western blotting. NRK-49F cells transfected with vector or Flag-tagged HDAC3 plasmids for 24 h, followed by treatment with TGFβ (10 ng/ml) and/or RG (10 μM) for another 24 h. Cell lysates were assayed with antibody against HDAC3, RASAL1, and α-SMA. (E) Quantitation of (D). (F) NRK-49F cells were treated with TGFβ (10 ng/ml) in presence or absence of RG (10 μM) for 24 h, and the mRNA of Rasal1 was analyzed by RT-PCR. Gapdh served as the internal control. The bottom panel is the quantitation. (G) Luciferase assay. HEK293T cells were transfected with either a positive control plasmid 3TP-luc or the Rasal1 promoter reporter Rasal1 p-luc together with a Renilla luciferase reporter and then treated with TGFβ (10 ng/ml) with or without RG (10 μM) for 24 h. Cell lysates were assayed for luciferase activities. (H) Schematic representation of the mouse Rasal1 promoter region spanning −300 to +243 relative to the transcriptional start site. The CpG island (gray-shaded area) and positions of qMSP primers (boxed regions) are depicted. (I) qMSP analysis of the Rasal1 promoter methylation in renal tissues from sham and UUO mice (7 d) (left) and in NRK-49F renal fibroblasts treated with TGFβ (10 ng/ml; 24 h) (right) with or without RG (10 mg/kg in mice; 10 μM in cell culture). (J) Representative agarose gel analysis of methylated (Met), unmethylated (unMet) and input PCR products for mouse tissue and NRK-49F cells mentioned above. (K) Quantitation of (J). Data were presented as means ± SD of 3 repeated cell assays or means ± SEM based on 6 renal samples. * P < 0.05, 2-tailed unpaired t test (3TP-luc) or 2-way ANOVA ( Rasal1 p-luc) (C, E, F, I, and K).

Article Snippet: Human embryonic kidney (HEK) 293T cells (American Type Culture Collection, USA) were cultured in DMEM, and rat renal fibroblast NRK-49F cells (Procell Life Science & Technology, Wuhan, China) and mouse primary renal fibroblasts were cultured in DMEM/F12 medium.

Techniques: Transfection, Control, Plasmid Preparation, Quantitation Assay, Western Blot, Reverse Transcription Polymerase Chain Reaction, Luciferase, Positive Control, Methylation, Cell Culture, Agarose Gel Electrophoresis

Journal: Research

Article Title: Epigenetic Suppression of RASAL1 by HDAC3 and Cofactor YY1 Promotes Fibroblast–Myofibroblast Transition and Renal Fibrosis

doi: 10.34133/research.1073

Figure Lengend Snippet: HDAC3 inhibition of Rasal1 transcription involves YY1. (A) Western blots of renal tissues from sham-, UUO (7 d)-, control-, and AAI (14 d)-treated mice ( n = 6) for YY1. The quantitation was on the right side. (B) Molecular docking between HDAC3 and YY1. The binding energy was analyzed by MM-GBSA. (C) Co-IP assay. Renal tissue homogenates from sham-, RG-, UUO-, or RG/UUO mice ( n = 6) were assayed for HDAC3 and YY1 as input controls (the bottom 3 blots). Subsequently, the same tissue lysates were immunoprecipitated with isoform-matched immunoglobulin (Ig) or antibodies (IPab) specific to HDAC3 (HD3) or YY1, and then immunoprecipitants were reciprocally assessed for HDAC3 and YY1 by Western blotting (the top 2 blots). (D) Quantification of Co-IP in (C). Data were presented as means ± SEM based on 3 renal samples from each group. (E) NRK-49F cells were transfected with control lentivirus (siC) and 3 YY1 knockdown lentiviruses (si1, si2, and si3) for 48 h. Western blotting was performed on cell lysates to measure YY1 protein levels. The bottom panel was the quantification of protein. (F) NRK-49F cells were transfected with control lentivirus (siC) and YY1 knockdown lentivirus (siR-Yy1, si3) for 24 h, followed by treatment with TGFβ (10 ng/ml) for additional 24 h. Cell lysates were assayed for RASAL1 and YY1 expression by Western blotting. (G) The top panel shows the YY1 binding motif sequence logo from JASPAR, while the bottom panel depicts a schematic diagram of YY1 binding to the Rasal1 promoter. TSS, transcription start site. (H) ChIP assay. The control, UUO, or AAI mice were treated with or without RG (10 mg/kg). The renal tissues indicated above were immunoprecipitated with antibodies to HDAC3, YY1 and Ac-H3, and then the precipitated genomic DNA (input) and the antibody-bound DNAs were PCR amplified with primers covering the YY1 motif on Rasal1 promoter. The RT-PCR products were analyzed on an agarose gel (left). RT-qPCR (right) was normalized to input DNA and presented as fold changes relative to sham. (I) Luciferase assay. HEK293T cells were transfected with the murine Rasal1 promoter reporter ( Rasal1 p-luc) or the m Rasal1 p-luc plus a Renilla luciferase plasmid. After 24 h of YY1 knockdown and control lentivirus treatment, cells were treated with TGFβ (10 ng/ml) for additional 24 h, and the luciferase activities were measured and normalized to Renilla luciferase activities. Data were presented as means ± SEM based on 6 renal samples or means ± SD of 3 repeated cell assays. * P < 0.05, one-way ANOVA (A and E), 2-way ANOVA (D, F, and H), or 3-way ANOVA (I).

Article Snippet: Human embryonic kidney (HEK) 293T cells (American Type Culture Collection, USA) were cultured in DMEM, and rat renal fibroblast NRK-49F cells (Procell Life Science & Technology, Wuhan, China) and mouse primary renal fibroblasts were cultured in DMEM/F12 medium.

Techniques: Inhibition, Western Blot, Control, Quantitation Assay, Binding Assay, Co-Immunoprecipitation Assay, Immunoprecipitation, Transfection, Knockdown, Expressing, Sequencing, Amplification, Reverse Transcription Polymerase Chain Reaction, Agarose Gel Electrophoresis, Quantitative RT-PCR, Luciferase, Plasmid Preparation

Journal: Research

Article Title: Epigenetic Suppression of RASAL1 by HDAC3 and Cofactor YY1 Promotes Fibroblast–Myofibroblast Transition and Renal Fibrosis

doi: 10.34133/research.1073

Figure Lengend Snippet: RASAL1 derepression is essential for the antifibrosis function of HDAC3 inhibition in vitro and in vivo . (A) Top: NRK-49F cells were treated with siC, si1, or si2 for 48 h. Western blotting of the cell lysates was assayed for RASAL1 proteins. Bottom: C57BL/6 mice were treated with Ch-siC, Ch-si1, or Ch-si2 once by intravenous injection, 6 mice in each group. Seven days later, Western blotting was performed on mouse renal tissues to detect RASAL1 proteins. (B) Western blotting. NRK-49F cells were infected with siC or si2 for 48 h, and then the cells were treated with TGFβ (10 ng/ml) and/or RG (10 μM) for 24 h. Cell lysates were assayed with antibody against HDAC3, RASAL1, α-SMA, and Colla 1. (C) Quantifications of Western blots in (B). Data were presented as means ± SD of 3 repeated cell assays. * P < 0.05, 3-way ANOVA followed by Tukey’s post hoc test. (D) Mice receiving Ch-siC or Ch-si2 were subgrouped into sham, RG, UUO, or RG/UUO ( n = 6). Representative photomicrographs of kidney sections were stained by Sirius Red. The black arrows indicate fibrotic areas. (E) Quantification of (D). Data were presented as scatter plot. *P < 0.05, 3-way ANOVA followed by Tukey’s post hoc test. (F) Western blotting. The renal tissues were assayed for HDAC3, RASAL1, α-SMA, and Colla 1. Two samples from each group were shown. (G) Quantifications of (F). Data were presented as means ± SEM based on 6 renal samples. *P < 0.05, 2-way ANOVA. (H) A schematic diagram of sequential HDAC3 elevation, RASAL1 suppression, and FMT during renal fibrosis. Elevated HDAC3, in association with YY1, induces hypoacetylation and transcriptional suppression of the Rasal1 promoter, resulting in persistent FMT and development of renal fibrosis (red lines). Conversely, selective HDAC3 inhibition (HDAC3i) by RG mitigates Rasal1 suppression (black lines), FMT, and renal fibrosis.

Article Snippet: Human embryonic kidney (HEK) 293T cells (American Type Culture Collection, USA) were cultured in DMEM, and rat renal fibroblast NRK-49F cells (Procell Life Science & Technology, Wuhan, China) and mouse primary renal fibroblasts were cultured in DMEM/F12 medium.

Techniques: Inhibition, In Vitro, In Vivo, Western Blot, Injection, Infection, Staining