Kidney Week

Abstract: SA-PO854

Protein Carbamylation Exacerbates Vascular Calcification

Session Information

• Vascular Calcification
  November 04, 2017 | Location: Hall H, Morial Convention Center
  Abstract Time: 10:00 AM - 10:00 AM

Category: Mineral Disease

• 1205 Vascular Calcification

Authors

• Mori, Daisuke, Osaka University Graduate School of Medicine, Suita, Japan

Daisuke Mori,

• Matsui, Isao, Osaka University Graduate School of Medicine, Suita, Japan

Isao Matsui,

• Hashimoto, Nobuhiro, Osaka University Graduate School of Medicine, Suita, Japan

Nobuhiro Hashimoto,

• Matsumoto, Ayumi, Osaka University Graduate School of Medicine, Suita, Japan

Ayumi Matsumoto,

• Shimada, Karin, Osaka University Graduate School of Medicine, Suita, Japan

Karin Shimada,

• Yamaguchi, Satoshi, Osaka University Graduate School of Medicine, Suita, Japan

Satoshi Yamaguchi,

• Oka, Tatsufumi, Osaka University Graduate School of Medicine, Suita, Japan

Tatsufumi Oka,

• Kubota, Keiichi, Osaka University Graduate School of Medicine, Suita, Japan

Keiichi Kubota,

• Yonemoto, Sayoko, Osaka University Graduate School of Medicine, Suita, Japan

Sayoko Yonemoto,

• Sakaguchi, Yusuke, Department of Comprehensive Kidney Disease Research, Osaka University Graduate School of Medicine, Osaka, Japan

Yusuke Sakaguchi,

• Hamano, Takayuki, Department of Comprehensive Kidney Disease Research, Osaka University Graduate School of Medicine, Osaka, Japan

Takayuki Hamano,

• Isaka, Yoshitaka, Osaka University Graduate School of Medicine, Suita, Japan

Yoshitaka Isaka,

Background

Protein carbamylation is an irreversible posttranslational modification that can occur non-enzymatically in the presence of urea. Although carbamylation is recognized as a prognostic biomarker, the effects of protein carbamylation on organ dysfunction remain uncertain.

Methods

Using in vitro, ex vivo, in vivo models, we investigated the effects of carbamylation on vascular calcification (VC), a life-threatening pathological condition that is common under carbamylation-prone situations.

Results

Protein carbamylation exacerbated the calcification of human vascular smooth muscle cells (hVSMCs) by suppressing the expression of ectonucleotide pyrophosphate/phosphodiesterase 1 (ENPP1), a key enzyme in the generation of pyrophosphate. By using immunoprecipitation in combination with mass spectrometry, we determined that several mitochondrial proteins, including ATP synthase subunits α and β, were carbamylated, although ENPP1 itself was not identified as a carbamylated protein. Rather, protein carbamylation reduced mitochondrial membrane potential and exaggerated mitochondria-derived oxidative stress, which downregulated ENPP1. The effects of carbamylation on ectopic calcification were abolished in mitochondrial-DNA-depleted hVSMCs and in hVSMCs treated with Mito-TEMPO, which indicated that mitochondria played essential roles in carbamylation-mediated effects. We also evaluated the carbamylation effects by using ex vivo and in vivo models: Protein carbamylation suppressed enzymatic histochemical staining for cytochrome c oxidase and succinate dehydrogenase in rat aortae, exacerbated calcifying-medium-induced VC in aortic ring cultures, and exacerbated warfarin/vitamin-D-induced VC in rats.

Conclusion

Protein carbamylation exacerbates VC by exaggerating mitochondria-derived oxidative stress and the resultant suppression of ENPP1.