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Published on: February 28, 2017
8-Nitro-cGMP suppresses mineralization by mouse osteoblasts
Kotaro Kaneko1,2, Yoichi Miyamoto1, Tomoaki Ida3
1Department of Biochemistry, Showa University School of Dentistry, 1-5-8 Hatanodai, Shinagawa, Tokyo 142-8555, 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.
Insights
8-nitro-cGMP, a messenger molecule, is produced by osteoblasts and suppresses bone formation. This finding reveals 8-nitro-cGMP as a negative regulator of osteoblastic differentiation, impacting bone remodeling.
Area of Science:
- Bone biology
- Cell signaling
- Biochemistry
Background:
- Bone remodeling involves osteoblasts (formation) and osteoclasts (resorption).
- Nitric oxide (NO) and reactive oxygen species (ROS) are key regulators of bone remodeling.
- 8-nitro-cGMP acts as a second messenger for NO and ROS, previously shown to promote osteoclastogenesis.
Purpose of the Study:
- To investigate the formation and function of 8-nitro-cGMP in osteoblasts.
- To determine the role of 8-nitro-cGMP in osteoblastic differentiation.
- To elucidate the regulatory mechanisms of 8-nitro-cGMP in bone formation.
Main Methods:
- Primary mouse calvarial osteoblasts were cultured.
- 8-nitro-cGMP production was measured and modulated by cytokines (TNF-α, IL-1β).
- Osteoblastic differentiation and mineralization were assessed following treatment with 8-nitro-cGMP or 8-bromo-cGMP, and after Cars2 gene silencing.
Main Results:
- Osteoblasts produce 8-nitro-cGMP, with production increased by TNF-α and IL-1β.
- These cytokines suppressed osteoblastic differentiation via NO synthase activity.
- Exogenous 8-nitro-cGMP inhibited osteoblastic phenotypes and mineralization, unlike 8-bromo-cGMP.
- Silencing of Cars2, involved in reactive sulfur species production, mimicked the suppressive effects of 8-nitro-cGMP.
Conclusions:
- 8-nitro-cGMP is produced endogenously by osteoblasts.
- 8-nitro-cGMP acts as a negative regulator of osteoblastic differentiation and function.
- These findings reveal a novel mechanism controlling bone formation and remodeling.
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