gms | German Medical Science

Objectives: Cigarette smoke (CS) is a known risk factor for osteoporosis. Heated tobacco products are considered potential reduced-risk alternatives to cigarettes. However, the impact of these products on bone cells remains to be elucidated. Our aim was to establish a 2D osteoblast/osteoclast coculture that mimicked a CS-induced osteoporotic environment, as observed in vivo, and to evaluate the effects of reduced-risk products on bone homeostasis in comparison to CS.

Methods: A human immortalized mesenchymal stem cell line (SCP-1) and human monocytic cell line (THP-1), in a 1:8 ratio, were cultured as osteoblast and osteoclast precursors on a 47°C-heat-decellularized Saos-2 cell matrix. The cocultures were exposed daily to total particular matter (TPM) from reference CS (1R6F; at 0.08, 0.4, or 2 µg nicotine/mL) alone or in combination with anti-osteoporotic drugs (50 µM zoledronate/alendronate) for 14 days. The cocultures were also treated acutely with TPM from 1R6F CS or Tobacco Heating System (THS) 2.4 aerosol (Philip Morris Products S.A.) at 0.4, 2, or 10 µg nicotine/mL. Cell viability and function were evaluated by mitochondrial activity (resazurin conversion), adenosine triphosphate and lactate dehydrogenase (LDH) content, alkaline phosphatase (AP), tartrate-resistant acid phosphatase (TRAP), and carbonic anhydrase II (CAII) activity, and cell morphology (phalloidin-TRITC staining) on days 3, 7, and 14.

Results and Conclusion: Alendronate/zoledronate treatment improved the negative impact of continuous exposure to 0.4 µg/mL 1R6F TPM on coculture cell viability, function, and calcium matrix deposition, demonstrating that our coculture could represent the clinical observation in orthopedic patients who are smokers. Interestingly, cocultures treated acutely with 10 µg/mL 1R6F TPM showed a significant decrease in metabolic activity, ATP content, and osteoblast and osteoclast cell number relative to THS TPM on days 3, 7, and 14. Additionally, cocultures exposed to 1R6F TPM showed greater LDH release than those exposed to THS TPM at comparable nicotine concentrations. Moreover, 10 µg/mL 1R6F TPM treatment caused significant downregulation of TRAP and CAII activity relative to the untreated and THS TPM-treated condition. Nevertheless, no changes in AP activity were observed with 1R6F TPM treatment. These results suggest that THS 2.4 TPM had less harmful effects than 1R6F TPM on the osteoblast/osteoclast coculture at equal nicotine concentrations. Further studies are required to confirm whether THS is a less harmful substitute for orthopedic patients who are smokers.