Pterostilbene-isothiocyanate impedes RANK/TRAF6 interaction to inhibit osteoclastogenesis, promoting osteogenesis in vitro and alleviating glucocorticoid induced osteoporosis in rats

Biochem Pharmacol. 2022 Dec:206:115284. doi: 10.1016/j.bcp.2022.115284. Epub 2022 Oct 7.

Abstract

Prolonged glucocorticoid treatment often leads to glucocorticoid-induced osteoporosis (GIOP), a common iatrogenic complication. This study has explored the anti-osteoporotic potential of semi-synthetic compound, pterostilbene isothiocyanate (PTER-ITC) in GIOP rat model and bone formation potential in vitro. Dysregulated bone-remodelling leads to osteoporosis. PTER-ITC has shown anti-osteoclastogenic activity in vitro. However, its molecular target remains unidentified, which has been explored in this study through in silico and experimental approaches. Alizarin Red S and von-Kossa staining, and alkaline phosphatase (ALP) activity showed the osteogenic differentiation potential of PTER-ITC in pre-osteoblastic mouse MC3T3-E1 and human hFOB 1.19 cells, further, confirmed through the expressions of osteogenic markers at transcriptional (RT-qPCR) and translational (immunoblotting) levels. The anti-osteoclastogenic property of PTER-ITC was confirmed through inhibition of actin ring formation in mouse RAW 264.7 and human THP-1 macrophagic cells. Molecular docking and molecular dynamic simulation showed that PTER-ITC inhibited the crucial osteoclastogenic RANK/TRAF6 interaction, which was further confirmed biochemically through co-immunoprecipitation assay. Osteoporotic bone architecture [validated through scanning electron microscopy (SEM), X-ray radiography, and micro-computed tomography (µ-CT)], physiology (confirmed through compression testing, Young's modulus and stress versus strain output) and histology (verified through hematoxylin-eosin, Alizarin Red S, von-Kossa and Masson-trichrome staining) of PTER-ITC-treated GIOP female Wistar rats were assuaged. Osteoporotic amelioration through PTER-ITC treatment was further substantiated through serum biomarkers, like, parathyroid hormone (PTH), ALP, calcium (Ca2+), Procollagen type I N-terminal propeptide (P1NP), and 25-hydroxy vitamin D. In conclusion, this study identifies the molecular target of PTER-ITC in impeding osteoclastogenesis and facilitating osteogenesis to ameliorate osteoporosis.

Keywords: Dexamethasone; Glucocorticoid-induced osteoporosis; Micro-computed tomography; Pterostilbene-isothiocyanate (PTER-ITC); Scanning electron microscopy; Ultimate compressive strength; Young’s modulus.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Differentiation
  • Female
  • Glucocorticoids / adverse effects
  • Humans
  • Isothiocyanates* / pharmacology
  • Mice
  • Molecular Docking Simulation
  • Osteoblasts / metabolism
  • Osteogenesis*
  • Osteoporosis* / chemically induced
  • Osteoporosis* / drug therapy
  • Osteoporosis* / metabolism
  • Rats
  • Rats, Wistar
  • Receptor Activator of Nuclear Factor-kappa B
  • Stilbenes* / pharmacology
  • TNF Receptor-Associated Factor 6 / metabolism
  • X-Ray Microtomography

Substances

  • Glucocorticoids
  • Isothiocyanates
  • isothiocyanic acid
  • pterostilbene
  • TNF Receptor-Associated Factor 6
  • Stilbenes
  • Receptor Activator of Nuclear Factor-kappa B