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2,3,5,6-Tetramethylpyrazine (TMP) down-regulated arsenic-induced heme oxygenase-1 and ARS2 expression by inhibiting Nrf2, NF-κB, AP-1 and MAPK pathways in human proximal tubular cells

  • Inorganic Compounds
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Abstract

Our recent study demonstrated that sodium arsenite at a clinically relevant dose induced nephrotoxicity in human renal proximal tubular epithelial cell line HK-2, which could be inhibited by natural product 2,3,5,6-tetramethylpyrazine (TMP) with antioxidant activity. The present study demonstrated that arsenic exposure resulted in protein and enzymatic induction of heme oxygenase-1 (HO-1) in dose- and time-dependent manners in HK-2 cells. Blocking HO-1 enzymatic activity by zinc protoporphyrin (ZnPP) augmented arsenic-induced apoptosis, ROS production and mitochondrial dysfunction, suggesting a critical role for HO-1 as a renal protectant in this procession. On the other hand, TMP, upstream of HO-1, inhibited arsenic-induced ROS production and ROS-dependent HO-1 expression. TMP also prevented mitochondria dysfunction and suppressed activation of the intrinsic apoptotic pathway in HK-2 cells. Our results revealed that the regulation of arsenic-induced HO-1 expression was performed through multiple ROS-dependent signal pathways and the corresponding transcription factors, including p38 MAPK and JNK (but not ERK), AP-1, Nrf2 and NF-κB. TMP inhibited arsenic-induced activations of JNK, p38 MAPK, ERK, AP-1 and Nrf2 and block HO-1 protein expression. The present study, furthermore, demonstrated arsenic-induced expression of arsenic response protein 2 (ARS2) that was regulated by p38 MAPK, ERK and NF-κB. To our knowledge, this is the first report showing that ARS2 involved in arsenic-induced nephrotoxicity, while TMP pretreatment prevented such an up-regulation of ARS2 in HK-2 cells. Given ARS2 and HO-1 sharing the similar regulation mechanism, we speculated that ARS2 might also mediate cell survival in this procession. In summary, our study highlighted a role of HO-1 in the protection against arsenic-induced cytotoxicity downstream from the primary targets of TMP and further indicated that TMP may be used as a potential therapeutic agent in the treatment of arsenic-induced nephrotoxicity.

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Abbreviations

AG:

Aminoguanidine

AKI:

Acute kidney injury

ARS2:

Arsenic response protein 2

As:

Arsenic

Bay:

Bay 11-7082

CKD:

Chronic kidney disease

DHE:

Dihydroethidium

FACS:

Fluorescence-activated cell sorter

HO-1:

Heme oxygenase-1

MAPK:

Mitogen-activated protein kinase

Nrf2:

Nuclear factor erythroid derived-2

NAC:

N-acetylcysteine

NF-κB:

Nuclear factor-κB

PARP:

Poly ADP-ribose polymerase

PI:

Propidium iodide

ROS:

Reactive oxygen species

SB:

SB203580

SDH:

Sucinate dehydrogenase

SP:

SP600125

TMP:

Tetramethylpyrazine

TUNEL:

Terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling

U0:

U0126

ZnPP:

Zinc protoporphyrin

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Acknowledgments

The authors would like to thank Drs. Preety Sharma, Erik Young, Bo Zhang, Gangkun Lin, Wupeng Liao, Hongning Zhou and Winsome Walker for advice and helpful discussion. This work was supported by Environmental Center Grant P30 ES009089, NIH Grant P01 CA049062 and National Natural Science Foundation of China Grant 81573936 and 81373614.

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Correspondence to Xuezhong Gong.

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Gong, X., Ivanov, V.N. & Hei, T.K. 2,3,5,6-Tetramethylpyrazine (TMP) down-regulated arsenic-induced heme oxygenase-1 and ARS2 expression by inhibiting Nrf2, NF-κB, AP-1 and MAPK pathways in human proximal tubular cells. Arch Toxicol 90, 2187–2200 (2016). https://doi.org/10.1007/s00204-015-1600-z

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