The protective effect of klotho against contrast-associated acute kidney injury via the antioxidative effect

Hyung Jung Oh, Hyewon Oh, Bo Young Nam, Je Sung You, Dong Ryeol Ryu, Shin Wook Kang, Yong Eun Chung

Research output: Contribution to journalArticle

Abstract

As oxidative stress is one major factor behind contrast-associated acute kidney injury (CA-AKI), we investigated the protective effect of klotho against CA-AKI via the antioxidative effect. In in vitro experiments, cells (NRK-52E) were divided into the following three groups: control, iopamidol, or iopamidol + recombinant klotho (rKL) groups. Moreover, cell viability was measured with the Cell Counting Kit-8 assay, and oxidative stress was examined with 2',7'-dichlorodihydrofluorescein diacetate fluorescence intensity. RT-PCR and Western blot analysis were performed to assess propidium iodide klotho expression, and Bax-to-Bcl-2 and apoptosis ratios were evaluated with annexin V/Hoechst 33342 staining. Furthermore, we knocked down the klotho gene using siRNA to verify the endogenous effect of klotho. In our in vivo experiments, oxidative stress was evaluated with the thiobarbituric acid-reactive substance assay, and apoptosis was evaluated with the Bax-to-Bcl-2 ratio and cleaved caspase-3 immunohistochemistry. Additionally, cell and tissue morphology were investigated with transmission electron microscopy. In both in vitro and in vivo experiments, mRNA and protein expression of klotho significantly decreased in CA-AKI mice compared with control mice, whereas oxidative stress and apoptosis markers were significantly increased in CA-AKI mice. However, rKL supplementation mitigated the elevated apoptotic markers and oxidative stress in the CA-AKI mouse model and improved cell viability. In contrast, oxidative stress and apoptotic markers were more aggravated when the klotho gene was knocked down. Moreover, we found more cytoplasmic vacuoles in the CA-AKI mouse model using transmission electron microscopy but fewer cytoplasmic vacuoles in rKL-supplemented cells. The present study shows that klotho in proximal tubular cells can protect against CA-AKI via an antioxidative effect.

Original languageEnglish
Pages (from-to)F881-F889
JournalAmerican journal of physiology. Renal physiology
Volume317
Issue number4
DOIs
Publication statusPublished - 2019 Oct 1

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Acute Kidney Injury
Oxidative Stress
Iopamidol
Apoptosis
Vacuoles
Transmission Electron Microscopy
Cell Survival
Thiobarbituric Acid Reactive Substances
Propidium
Annexin A5
Caspase 3
Small Interfering RNA
Genes
Fluorescence
Western Blotting
Immunohistochemistry
Staining and Labeling
Polymerase Chain Reaction
Control Groups
Messenger RNA

All Science Journal Classification (ASJC) codes

  • Physiology
  • Urology

Cite this

Oh, Hyung Jung ; Oh, Hyewon ; Nam, Bo Young ; You, Je Sung ; Ryu, Dong Ryeol ; Kang, Shin Wook ; Chung, Yong Eun. / The protective effect of klotho against contrast-associated acute kidney injury via the antioxidative effect. In: American journal of physiology. Renal physiology. 2019 ; Vol. 317, No. 4. pp. F881-F889.
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abstract = "As oxidative stress is one major factor behind contrast-associated acute kidney injury (CA-AKI), we investigated the protective effect of klotho against CA-AKI via the antioxidative effect. In in vitro experiments, cells (NRK-52E) were divided into the following three groups: control, iopamidol, or iopamidol + recombinant klotho (rKL) groups. Moreover, cell viability was measured with the Cell Counting Kit-8 assay, and oxidative stress was examined with 2',7'-dichlorodihydrofluorescein diacetate fluorescence intensity. RT-PCR and Western blot analysis were performed to assess propidium iodide klotho expression, and Bax-to-Bcl-2 and apoptosis ratios were evaluated with annexin V/Hoechst 33342 staining. Furthermore, we knocked down the klotho gene using siRNA to verify the endogenous effect of klotho. In our in vivo experiments, oxidative stress was evaluated with the thiobarbituric acid-reactive substance assay, and apoptosis was evaluated with the Bax-to-Bcl-2 ratio and cleaved caspase-3 immunohistochemistry. Additionally, cell and tissue morphology were investigated with transmission electron microscopy. In both in vitro and in vivo experiments, mRNA and protein expression of klotho significantly decreased in CA-AKI mice compared with control mice, whereas oxidative stress and apoptosis markers were significantly increased in CA-AKI mice. However, rKL supplementation mitigated the elevated apoptotic markers and oxidative stress in the CA-AKI mouse model and improved cell viability. In contrast, oxidative stress and apoptotic markers were more aggravated when the klotho gene was knocked down. Moreover, we found more cytoplasmic vacuoles in the CA-AKI mouse model using transmission electron microscopy but fewer cytoplasmic vacuoles in rKL-supplemented cells. The present study shows that klotho in proximal tubular cells can protect against CA-AKI via an antioxidative effect.",
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The protective effect of klotho against contrast-associated acute kidney injury via the antioxidative effect. / Oh, Hyung Jung; Oh, Hyewon; Nam, Bo Young; You, Je Sung; Ryu, Dong Ryeol; Kang, Shin Wook; Chung, Yong Eun.

In: American journal of physiology. Renal physiology, Vol. 317, No. 4, 01.10.2019, p. F881-F889.

Research output: Contribution to journalArticle

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AU - You, Je Sung

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