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  2. Prolonged Cadmium Exposure and Osteoclastogenesis: A Mechanistic Mouse and in Vitro Study

Prolonged Cadmium Exposure and Osteoclastogenesis: A Mechanistic Mouse and in Vitro Study

  • Environ Health Perspect. 2024 Jun;132(6):67009. doi: 10.1289/EHP13849.
Zhiyuan Liu 1 2 3 Jinzhi Wu 1 2 3 Zhe Dong 1 2 3 Yanshuai Wang 1 2 3 Gang Wang 1 2 4 Chengjie Chen 1 2 3 Huihui Wang 1 2 5 Yang Yang 6 Yongxin Sun 6 Maowei Yang 7 Jingqi Fu 1 2 3 Jiliang Li 8 Qiang Zhang 9 Yuanyuan Xu 1 2 5 Jingbo Pi 1 2 3
Affiliations

Affiliations

  • 1 Key Laboratory of Environmental Stress and Chronic Disease Control and Prevention, Ministry of Education, China Medical University, Shenyang, Liaoning, P.R. China.
  • 2 Key Laboratory of Liaoning Province on Toxic and Biological Effects of Arsenic, China Medical University, Shenyang, Liaoning, P.R. China.
  • 3 Program of Environmental Toxicology, School of Public Health, China Medical University, Shenyang, Liaoning, P.R. China.
  • 4 Experimental and Teaching Center, School of Public Health, China Medical University, Shenyang, Liaoning, P.R. China.
  • 5 Group of Chronic Disease and Environmental Genomics, School of Public Health, China Medical University, Shenyang, Liaoning, P.R. China.
  • 6 Department of Rehabilitation Medicine, First Hospital of China Medical University, Shenyang, Liaoning, P.R. China.
  • 7 Department of Orthopedics, First Hospital of China Medical University, Shenyang, Liaoning, P.R. China.
  • 8 Department of Biology, Indiana University Indianapolis, Indianapolis, Indiana, USA.
  • 9 Gangarosa Department of Environmental Health, Rollins School of Public Health, Emory University, Atlanta, Georgia, USA.
Abstract

Background: Cadmium (Cd) is a highly toxic and widespread environmental oxidative stressor that causes a myriad of health problems, including osteoporosis and bone damage. Although nuclear factor erythroid 2-related factor 2 (NRF2) and its Cap 'n' Collar and basic region Leucine Zipper (CNC-bZIP) family member nuclear factor erythroid 2-related factor 1 (NRF1) coordinate various stress responses by regulating the transcription of a variety of antioxidant and cytoprotective genes, they play distinct roles in bone metabolism and remodeling. However, the precise roles of both transcription factors in bone loss induced by prolonged Cd exposure remain unclear.

Objectives: We aimed to understand the molecular mechanisms underlying Cd-induced bone loss, focusing mainly on the roles of NRF2 and NRF1 in osteoclastogenesis provoked by Cd.

Methods: Male wild-type (WT), global Nrf2-knockout ( N r f 2 - / - ) and myeloid-specific Nrf2 knockout [Nrf2(M)-KO] mice were administered Cd (50 or 100 ppm ) via drinking water for 8 or 16 wk, followed by micro-computed tomography, histological analyses, and plasma biochemical testing. Osteoclastogenesis was evaluated using bone marrow-derived osteoclast progenitor cells (BM-OPCs) and RAW 264.7 cells in the presence of Cd (10 or 20 nM ) with a combination of genetic and chemical modulations targeting NRF2 and NRF1.

Results: Compared with relevant control mice, global N r f 2 - / - or Nrf2(M)-KO mice showed exacerbated bone loss and augmented osteoclast activity following exposure to 100 ppm Cd in drinking water for up to 16 wk. In vitro osteoclastogenic analyses suggested that Nrf2-deficient BM-OPCs and RAW 264.7 cells responded more robustly to low levels of Cd (up to 20 nM ) with regard to osteoclast differentiation compared with WT cells. Further mechanistic studies supported a compensatory up-regulation of long isoform of NRF1 (L-NRF1) and subsequent induction of nuclear factor of activated T cells, cytoplasmic, Calcineurin dependent 1 (NFATc1) as the key molecular events in the Nrf2 deficiency-worsened and Cd-provoked osteoclastogenesis. L-Nrf1 silenced (via lentiviral means) Nrf2-knockdown (KD) RAW cells exposed to Cd showed dramatically different NFATc1 and subsequent osteoclastogenesis outcomes compared with the cells of Nrf2-KD alone exposed to Cd, suggesting a mitigating effect of the Nrf1 silencing. In addition, suppression of Reactive Oxygen Species by exogenous antioxidants N -acetyl-l-cysteine ( 2 mM ) and mitoquinone mesylate (MitoQ; 0.2 μ M ) mitigated the L-NRF1-associated effects on NFATc1-driven osteoclastogenesis outcomes in Cd-exposed Nrf2-KD cells.

Conclusions: This in vivo and in vitro study supported the authors' hypothesis that Cd exposure caused bone loss, in which NRF2 and L-NRF1 responded to Cd and osteoclastogenic stimuli in a cooperative, but contradictive, manner to coordinate Nfatc1 expression, osteoclastogenesis and thus bone homeostasis. Our study suggests a novel strategy targeting NRF2 and L-NRF1 to prevent and treat the bone toxicity of Cd. https://doi.org/10.1289/EHP13849.

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