Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The . gov means it’s official. Federal government websites often end in . gov or VSports app下载. mil. Before sharing sensitive information, make sure you’re on a federal government site. .

Https

The site is secure. The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely. V体育官网.

. 2017 Nov 6;7(1):14603.
doi: 10.1038/s41598-017-14906-8.

Puerarin attenuates diabetic kidney injury through the suppression of NOX4 expression in podocytes

Xueling Li  1 , Weijing Cai  2 , Kyung Lee  2 , Bohan Liu  2 , Yueyi Deng  1 , Yiping Chen  1 , Xianwen Zhang  1 , John Cijiang He  3   4 , Yifei Zhong  5
Affiliations

Puerarin attenuates diabetic kidney injury through the suppression of NOX4 expression in podocytes

"VSports最新版本" Xueling Li et al. Sci Rep. .

Erratum in

"VSports最新版本" Abstract

Radix puerariae, a traditional Chinese herbal medication, has been used to treat patients with diabetic nephropathy (DN). Several studies demonstrated that puerarin, the active compound of radix puerariae, reduces diabetic injury in streptozotocin (STZ)-induced diabetic rodent models VSports手机版. However, as STZ injection alone results in mild kidney injury, the therapeutic benefit afforded by puerarin in DN remained inconclusive. Thus we sought to clarify the role of puerarin by employing an accelerated DN model, STZ-induced diabetes in the endothelial nitric oxide synthase-null (eNOS-/-) mice. Puerarin treatment of diabetic eNOS-/- mice significantly attenuated albuminuria and diabetic kidney injury, which were associated with reduced oxidative stress and reduced NAPDH oxidase 4 (NOX4) in glomeruli of diabetic eNOS-/- mice. Puerarin treatment of murine podocytes culture in high glucose conditions led to reduced superoxide production and NOX4 expression. We further determined that that puerarin treatment increased both mRNA and protein levels of SIRT1 in podocytes and that puerarin led to SIRT1-mediated deacetylation of NF-κB and suppression of NOX4 expression. Our findings confirm the renoprotective effects of puerarin in an experimental model of advanced DN and provide a molecular mechanism by which puerarin exerts the anti-oxidative effects in podocytes in the diabetic milieu. .

PubMed Disclaimer

Conflict of interest statement

The authors declare that they have no competing interests.

Figures

Figure 1
Figure 1
Puerarin reduced albuminuria and kidney injury in STZ-induced diabetic eNOS knockout mice. (A) Both diabetic and non-diabetic mice were treated with either vehicle or puerarin when mice developed albuminuria for a total of 9 weeks. Urine samples were collected weekly after starting the treatments for determination of urinary albumin/creatinine ratio. Significant differences were observed starting at 2 weeks after puerarin treatment compared with vehicle-treated diabetic mice. *p < 0.05 and *p < 0.001 compared to vehicle-treated diabetic mice, n = 6. (B) Representative image of periodic acid-Schiff (PAS) stained kidneys. Scale bar: 20 μm. (C,D) Morphometric analysis was performed to determine the glomerular area (C) and % of mesangial area (D) in the glomerular cross sections. **p < 0.01 and ***p < 0.001, n = 6.
Figure 2
Figure 2
Puerarin improved oxidative stress in diabetic glomeruli. (A) Representative image of nitro-tyrosine immunostaining in paraffin-embedded kidney sections. Scale bar: 100 μm. (B) Representative images of 8-oxoG (green) and podocyte marker WT1 (red) immunofluorescence. DAPI was used as a counterstain. Scale bar: 20 μm. (C) Quantification of 8-oxoG immunofluorescence are shown. ***p < 0.001, n = 6 mice.
Figure 3
Figure 3
Treatment of puerarin reduced expression of NOX4 in the glomeruli of diabetic mice. (A) Real-time PCR analysis of NOX4 mRNA levels from isolated glomeruli. **p < 0.01 and ***p < 0.001, n = 6 mice. (B) Representative western blot analyses showing NOX4 expression in glomerular lysates. (C) Quantification of NOX4 protein expression normalized to GAPDH. *p < 0.05, n = 6 mice.
Figure 4
Figure 4
Puerarin inhibited oxidative stress in murine podocytes cultured in high glucose. (A,B) Immortalized mouse podocytes were cultured in either normal glucose (NG: 5 mM glucose + 25 mM mannitol) or high glucose (HG: 30 mM glucose) treated with DMSO vehicle or 10 μM puerarin for 24 hours. Superoxide production was determined by incubation of cells with dichlorofluorescin diacetate (DCFDA) fluorogenic dye and by detection of oxidized 2′, 7′-dichlorofluorescein (DCF). Puerarin reduced superoxide generation in podocytes cultured in high glucose media. Representative image is shown in (A) and ratio of DCF+ cells/total (DAPI+) cells per field are shown in (B) (***p < 0.001, n = 3, 7 fields per group). (C) Real-time PCR analysis of NOX4 mRNA levels in podocytes cultured with NG or HG treated with vehicle or 10 μM puerarin for 4 hours. GAPDH mRNA was used as an internal control. ***p < 0.001, n = 3. (D) Representative western blot analyses of NOX4 expression in podocytes cultured in NG or HG conditions for 24 hours. The representative blots of three individual experiments are shown here. (E) Quantification of NOX4 protein expression normalized to GAPDH is shown. **p < 0.01 and ***p < 0.001, n = 3.
Figure 5
Figure 5
Puerarin downregulated NOX4 expression through activation of SIRT1 in murine podocytes exposed to high glucose. (A,B) Murine podocytes cultured in NG or HG were treated with different amounts of puerarin as indicated. The cells were lysed for western blot analysis for SIRT1, NOX4, acetyl-p65, total-p65, and β-actin. Representative blots of three independent experiments are shown in A, and quantification is shown in (B). (C,D) Podocytes transfected with SIRT1 overexpression vector or control vector were cultured in NG and HG for 24 hours. The cells were lysed for western blot analysis for SIRT1, NOX4, acetyl-p65, total-p65, and β-actin. Representative blots of three independent experiments are shown in (C), and quantification is shown in (D).
Figure 6
Figure 6
Puerarin downregulated NOX4 expression through deacetylation of p65NF-κB in murine podocytes exposed to high glucose. (A,B) Podocytes transfected with either wildtype p65 (WT), p65 with mutation of K310 acetyl-residue (K310R) or control empty vector (EV) were treated with TNF-α for 24 hours. Cells were lysed for western blot analysis for p65, NOX4 and β-actin. Representative blots of three independent experiments are shown in A, and quantification is shown in (B). (C) Podocytes cultured in NG or HG were treated with 10 μM puerarin or 1 μM SIRT1 agonist SRT1720 for 24 hours. Superoxide production was then measured by the DCF fluorescence method. *p < 0.05 compared between groups n = 3. (D) Real-time PCR analysis of Sirt1 mRNA in podocytes treated with vehicle or puerarin in NG or HG conditions show increased Sirt1 expression by puerarin. *p < 0.05 compared between groups n = 3.
See this image and copyright information in PMC

References

    1. USRDS. Annual Data Report: Atlas of End-Stage-Renal-Disease in the United States. (2011).
    1. World Health Organization. Global status report on noncommunicable diseases. volumes (World Health Organization, Geneva, Switzerland).
    1. Zhang L, et al. Trends in Chronic Kidney Disease in China. The New England journal of medicine. 2016;375:905–906. doi: 10.1056/NEJMc1602469. - "V体育官网" DOI - PubMed
    1. Yang W, et al. Prevalence of diabetes among men and women in China. The New England journal of medicine. 2010;362:1090–1101. doi: 10.1056/NEJMoa0908292. - DOI - PubMed
    1. de Zeeuw D. Unmet need in renal protection–do we need a more comprehensive approach? Contrib Nephrol. 2011;171:157–160. doi: 10.1159/000327337. - DOI - PubMed

Publication types

MeSH terms