Journal: Oncotarget

Article Title: A novel pan-Nox inhibitor, APX-115, protects kidney injury in streptozotocin-induced diabetic mice: possible role of peroxisomal and mitochondrial biogenesis

doi: 10.18632/oncotarget.18540

Figure Lengend Snippet: Diabetes was induced in mice by intraperitoneal injection of STZ (50 mg/kg). Then APX-115 (60 mg/kg/day) or losartan (1.5 mg/kg/day) was administered orally for 12 weeks to diabetic mice. After 12 weeks, urine and blood samples were collected for analysis of ( A ) urinary albumin excretion, ( B ) albumin/creatinine ratio, ( C ) creatinine clearance rate, and ( D ) plasma cystatin C. ( E ) Kidneys were fixed in paraffin and cut into 3 μm sections that were subsequently stained with PAS reagent. Scale bar: 10 μm; original magnification: 630×. After PAS staining, ( F ) glomerular volume, ( G ) mesangial area, and ( H ) tuft area were analyzed using Image-Pro Plus 4.5.1. DM, STZ-induced diabetic mice. Data are presented as means ± SE of 10–12 mice/group; * p < 0.05 vs. control, † p < 0.05 vs. DM.

Article Snippet: A murine mesangial cell line (MES-13, cloned from mice transgenic for the early region of SV-40 virus, passage 25) was obtained from American Type Culture Collection (ATCC, Rockville, MD, USA).

Techniques: Injection, Clinical Proteomics, Staining, Control

Journal: Oncotarget

Article Title: A novel pan-Nox inhibitor, APX-115, protects kidney injury in streptozotocin-induced diabetic mice: possible role of peroxisomal and mitochondrial biogenesis

doi: 10.18632/oncotarget.18540

Figure Lengend Snippet: ( A ) Plasma LPO, ( B ) urinary LPO, ( C ) kidney tissue LPO, ( D ) Nox1, ( E ) Nox2, and ( F ) Nox4 mRNA expression levels in kidneys were measured using real-time PCR. ( G and H ) Frozen kidney sections were stained with DHE at 5 µM (original magnification: 400×; scale bar: 20 μm). (A–H) Data are presented as means ± SE of 10–12 mice/group; * p < 0.05 vs. control, † p < 0.05 vs. DM. ( I ) Mesangial cells were incubated with or without APX-115 (1 µM) for 30 min and stimulated with or without 30 mM high glucose (HG) for 24 h followed by angII for 30 min. After that cells were incubated with 10 µM DCF-DA for 10 min and the fluorescence intensity was measured with a Zeiss vision system. Data are presented as means ± SE of at least 2 independent experiments; * p < 0.05 vs. control, † p < 0.05 vs. angII or angII+HG in DMSO.

Article Snippet: A murine mesangial cell line (MES-13, cloned from mice transgenic for the early region of SV-40 virus, passage 25) was obtained from American Type Culture Collection (ATCC, Rockville, MD, USA).

Techniques: Clinical Proteomics, Expressing, Real-time Polymerase Chain Reaction, Staining, Control, Incubation, Fluorescence

Journal: American journal of physiology. Renal physiology

Article Title: Antagonistic effects of bone morphogenetic protein-4 and -7 on renal mesangial cell proliferation induced by aldosterone through MAPK activation.

doi: 10.1152/ajprenal.00402.2006

Figure Lengend Snippet: Fig. 1. Expression of receptors for angiotensin II (ANG II) and aldosterone and the bone morphogenetic protein (BMP) system in mouse mesangial cells. The expression of mRNAs encoding ANG II type 1 receptor (AT1R), miner- alocorticoid receptor (MR), BMPs, BMP receptor (BMPR) types I and II, Smads, and follistatin (FLS) was examined in total cellular RNAs extracted from MES13 cells and mouse whole kidney tissues (mKidney) by RT-PCR analysis. Results shown are representative of those obtained from 3 indepen- dent experiments. MM, molecular weight marker.

Article Snippet: The mouse glomerular mesangial cell line SV40 MES13 (40) was from American Type Culture Collection (Manassas, VA).

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Molecular Weight, Marker

Journal: American journal of physiology. Renal physiology

Article Title: Antagonistic effects of bone morphogenetic protein-4 and -7 on renal mesangial cell proliferation induced by aldosterone through MAPK activation.

doi: 10.1152/ajprenal.00402.2006

Figure Lengend Snippet: Fig. 10. Effects of ANG II and Aldo on mesangial BMP systems. A: total cellular RNAs were extracted from MES13 cells cultured for 24 h in the absence (Control) or presence of ANG II (30 nM) and Aldo (100 nM). Extracted RNAs (2.0 g) were used as templates for GEArray analysis (SuperArray Bioscience). The signal intensities of the spots on the membranes obtained from 2 separate experiments were analyzed with the GEArray analyzer program (SuperArray Bioscience) after subtraction of the background levels of pUC18 DNA. Cyclophilin A (PPIA) is used as a positive control to compare the membranes. ALK, activin-like kinase. B: after preculture, the cells were treated with or without ANG II (30 nM) and Aldo (100 nM) for 24 h. Total cellular RNA was extracted and ALK-3, ActRII, BMPRII, Smad6, BMP-4, and BMP-7 mRNA levels were examined by real-time RT-PCR. The expression levels of target genes were standardized by RPL19 level in each sample. Results are shown as means SE of data from at least 3 separate experiments, each performed with triplicate samples. *P 0.05, **P 0.01 vs. each control.

Article Snippet: The mouse glomerular mesangial cell line SV40 MES13 (40) was from American Type Culture Collection (Manassas, VA).

Techniques: Cell Culture, Control, Positive Control, Quantitative RT-PCR, Expressing

Journal: Oxidative Medicine and Cellular Longevity

Article Title: Short-Chain Fatty Acids Ameliorate Diabetic Nephropathy via GPR43-Mediated Inhibition of Oxidative Stress and NF- κ B Signaling

doi: 10.1155/2020/4074832

Figure Lengend Snippet: SCFAs prevented the renal dysfunction and kidney injury. Urine ACR (a) of T2D mice were measured every 2 weeks, and blood urea nitrogen (BUN) (b), serum creatinine (SCr) (c), and serum cystatin C (d) were assayed at the 20th week of the experiment. Histopathological examination of renal tissues was by H&E, PAS, and Masson's trichrome staining (400x) (e). Mesangial expansion (f), glomerular tuft (g), and the accumulation of collagen (h) were measured. Ac: acetate; Pr: propionate; But: butyrate; ∗ p < 0.05 compared with the NC group; # p < 0.05 compared with the T2D group; & p < 0.05 compared with the Ac or Pr group.

Article Snippet: Mouse glomerular mesangial cells (SV-40 MES 13) were obtained from the China Center for Type Culture Collection and cultured in Dulbecco's modified Eagle's medium (Gibco, Waltham, MA, USA) containing 5.6 mM glucose and 10% fetal bovine serum (Gibco) at 37°C and 5% CO 2 .

Techniques: Staining

Journal: Oxidative Medicine and Cellular Longevity

Article Title: Short-Chain Fatty Acids Ameliorate Diabetic Nephropathy via GPR43-Mediated Inhibition of Oxidative Stress and NF- κ B Signaling

doi: 10.1155/2020/4074832

Figure Lengend Snippet: SCFA treatment partially inhibited oxidative stress and NF- κ B activation in high glucose-induced GMCs. (a) The effects of a concentration range of SCFAs or GPR43 agonist on GMC proliferation were analyzed by MTT assay. GMCs were stimulated with 30 mM high glucose in the presence of the indicated concentration of SCFAs or GPR43 agonist for 24 h. ROS (b), MDA (c), MCP-1 (d), and IL-1 β (e) in the cell culture supernatant were evaluated by kit. The protein expression of I- κ B α , NF- κ Bp65, p-NF- κ Bp65, and MCP-1 was assayed by western blotting (f). Ac: acetate group; Pr: propionate group; But: butyrate group; ∗ p < 0.05 compared with the NC group; # p < 0.05 compared with the HG group.

Article Snippet: Mouse glomerular mesangial cells (SV-40 MES 13) were obtained from the China Center for Type Culture Collection and cultured in Dulbecco's modified Eagle's medium (Gibco, Waltham, MA, USA) containing 5.6 mM glucose and 10% fetal bovine serum (Gibco) at 37°C and 5% CO 2 .

Techniques: Activation Assay, Concentration Assay, MTT Assay, Cell Culture, Expressing, Western Blot

Journal: Oxidative Medicine and Cellular Longevity

Article Title: Short-Chain Fatty Acids Ameliorate Diabetic Nephropathy via GPR43-Mediated Inhibition of Oxidative Stress and NF- κ B Signaling

doi: 10.1155/2020/4074832

Figure Lengend Snippet: SCFA-mediated antioxidant and anti-inflammatory effects were significantly facilitated by GPR43 overexpression. Plasmids expressing GPR43 with pCD513B-1 (an N-terminal GFP tag) and pCD513B-1 control plasmid that expresses GFP but cannot overexpress GPR43 were constructed to determine the effect of overexpressed GPR43 and GFP protein in GMCs according to fluorescence images (200x) (a) and western blotting (b). Following 30 mM high glucose for 24 h, 5 mM But-mediated inhibition of p-NF- κ Bp65 and MCP-1 protein expression (c) was significantly facilitated by GPR43 overexpression. The But-mediated inhibition of ROS (d) and p-NF- κ Bp65 (e) and MCP-1 and IL-1 β release (f) were reversed by siRNA-GPR43 but facilitated by GPR43 overexpression. But butyrate: ∗ p < 0.05 compared with the NC group, # p < 0.05 compared with the HG group, and & p < 0.05 compared with the But group.

Article Snippet: Mouse glomerular mesangial cells (SV-40 MES 13) were obtained from the China Center for Type Culture Collection and cultured in Dulbecco's modified Eagle's medium (Gibco, Waltham, MA, USA) containing 5.6 mM glucose and 10% fetal bovine serum (Gibco) at 37°C and 5% CO 2 .

Techniques: Over Expression, Expressing, Control, Plasmid Preparation, Construct, Fluorescence, Western Blot, Inhibition

Journal: Oxidative Medicine and Cellular Longevity

Article Title: Short-Chain Fatty Acids Ameliorate Diabetic Nephropathy via GPR43-Mediated Inhibition of Oxidative Stress and NF- κ B Signaling

doi: 10.1155/2020/4074832

Figure Lengend Snippet: Interaction between β -arrestin-2 and I- κ B α was induced by SCFAs via GPR43. (a) GMCs were treated with 30 mM high glucose for 6, 12, and 24 h. RT-PCR was performed to detect β -arrestin-2 and β -arrestin-1 mRNA levels. (b) The effects of indicated concentrations of SCFAs or an GPR43 agonist on β -arrestin-2 and β -arrestin-1 expression were analyzed by RT-PCR. (c) Western blot assay for the expression of β -arrestin-2 after 30 mM high glucose challenge for 6, 12, and 24 h. (d) High glucose-induced β -arrestin-2 expression was significantly reversed by SCFAs or GPR43 agonist. (e) The interaction between β -arrestin-2 and I- κ B α under physiological conditions was detected by immunoprecipitation (IP) with anti- β -arrestin-2 antibody or normal mouse IgG antibody (negative control), followed by western blotting with an anti-I- κ B α antibody. β -Arrestin-2 was conjugated with I- κ B α in vitro . (f) The interaction between β -arrestin-2 and I- κ B α was decreased by 30 mM high glucose but was reversed by 5 mM butyrate. And these butyrate-mediated effects were significantly reversed by siRNA-GPR43 but were facilitated by overexpressed GPR43. IgG-H marks the IgG heavy chain. But butyrate: ∗ p < 0.05 compared with the NC group, # p < 0.05 compared with the HG group, and & p < 0.05 compared with the HG+But group. Ac: acetate; Pr: propionate.

Article Snippet: Mouse glomerular mesangial cells (SV-40 MES 13) were obtained from the China Center for Type Culture Collection and cultured in Dulbecco's modified Eagle's medium (Gibco, Waltham, MA, USA) containing 5.6 mM glucose and 10% fetal bovine serum (Gibco) at 37°C and 5% CO 2 .

Techniques: Reverse Transcription Polymerase Chain Reaction, Expressing, Western Blot, Immunoprecipitation, Negative Control, In Vitro