8-Nitro-cGMP suppresses mineralization by mouse osteoblasts

Kotaro Kaneko; Yoichi Miyamoto; Tomoaki Ida; Masanobu Morita; Kentaro Yoshimura; Kei Nagasaki; Kazuki Toba; Risa Sugisaki; Hozumi Motohashi; Takaaki Akaike; Daichi Chikazu; Ryutaro Kamijo

doi:10.3164/jcbn.21-129

8-Nitro-cGMP suppresses mineralization by mouse osteoblasts

J Clin Biochem Nutr. 2022 Nov;71(3):191-197. doi: 10.3164/jcbn.21-129. Epub 2022 Jul 27.

Authors

Kotaro Kaneko^{1

2}, Yoichi Miyamoto¹, Tomoaki Ida³, Masanobu Morita³, Kentaro Yoshimura¹, Kei Nagasaki^{1

4}, Kazuki Toba^{1

5}, Risa Sugisaki^{1

2}, Hozumi Motohashi⁶, Takaaki Akaike³, Daichi Chikazu², Ryutaro Kamijo¹

Affiliations

¹ Department of Biochemistry, Showa University School of Dentistry, 1-5-8 Hatanodai, Shinagawa, Tokyo 142-8555, Japan.
² Department of Oral and Maxillofacial Surgery, Tokyo Medical University, 6-7-1 Nishishinjuku, Shinjuku, Tokyo 160-0023, Japan.
³ Department of Environmental Medicine and Molecular Toxicology, Tohoku University Graduate School of Medicine, 2-1 Seiryo-cho, Aoba-ku, Sendai 980-8575, Japan.
⁴ Department of Orthopedics, Showa University School of Medicine, 1-5-8 Hatanodai, Shinagawa, Tokyo 142-8555, Japan.
⁵ Department of Oral and Maxillofacial Surgery, Showa University School of Dentistry, 1-5-8 Hatanodai, Shinagawa, Tokyo 142-8555, Japan.
⁶ Department of Gene Expression Regulation, Institute of Development, Aging and Cancer, Tohoku University, 4-1 Seiryo-cho, Aoba-ku, Sendai 980-8575, Japan.

Abstract

Nitric oxide and reactive oxygen species regulate bone remodeling, which occurs via bone formation and resorption by osteoblasts and osteoclasts, respectively. Recently, we found that 8-nitro-cGMP, a second messenger of nitric oxide and reactive oxygen species, promotes osteoclastogenesis. Here, we investigated the formation and function of 8-nitro-cGMP in osteoblasts. Mouse calvarial osteoblasts were found to produce 8-nitro-cGMP, which was augmented by tumor necrosis factor-α (10 ng/ml) and interleukin-1β (1 ng/ml). These cytokines suppressed osteoblastic differentiation in a NO synthase activity-dependent manner. Exogenous 8-nitro-cGMP (30 μmol/L) suppressed expression of osteoblastic phenotypes, including mineralization, in clear contrast to the enhancement of mineralization by osteoblasts induced by 8-bromo-cGMP, a cell membrane-permeable analog of cGMP. It is known that reactive sulfur species denitrates and degrades 8-nitro-cGMP. Mitochondrial cysteinyl-tRNA synthetase plays a crucial role in the endogenous production of RSS. The expression of osteoblastic phenotypes was suppressed by not only exogenous 8-nitro-cGMP but also by silencing of the Cars2 gene, indicating a role of endogenous 8-nitro-cGMP in suppressing the expression of osteoblastic phenotypes. These results suggest that 8-nitro-cGMP is a negative regulator of osteoblastic differentiation.

Keywords: 8-nitro-cGMP; differentiation; mineralization; nitric oxide; osteoblasts.