Hexane Fraction of Turbo brunneus Inhibits Intermediates of RANK-RANKL Signaling Pathway and Prevent Ovariectomy Induced Bone Loss.

Sachin Chaugule,Shubhada Chiplunkar,Madhavi Indap,C Murali Krishna,Pradip Chaudhari,Shalini Kashipathi Sureshbabu,Suresh Dakave

doi:10.3389/fendo.2019.00608

Abstract

Osteoporosis is a “silent disease” characterized by fragile and impaired bone quality. Bone fracture results in increased mortality and poor quality of life in aged people particularly in postmenopausal women. Bone is maintained through the delicate balance between osteoclast-mediated bone resorption and osteoblast-mediated bone formation. The imbalance is caused most often by overly active osteoclasts due to estrogen deficiency. Natural products have long been used to prevent and treat osteoporosis since they have fewer side effects. The marine environment is a potential source of biologically and structurally novel biomolecules with promising biological activities but is less explored for the treatment of bone-related diseases. The present study aims to evaluate the antiosteoporotic effect of Hexane fraction of Turbo brunneus methanolic extract (HxTME) and to investigate its role in RANK-RANKL signaling pathway using in vitro osteoclasts cultures and in vivo ovariectomized (OVX) Swiss mice model. The present study demonstrated that the HxTME significantly inhibited RANKL induced osteoclast differentiation and maturation in vitro. HxTME completely downregulated the mRNA expression of key transcription factors such as NFATc1, c-FOS, and osteoclasts related genes involved in osteoclastogenesis. In vivo studies also depicted the effectiveness of HxTME in ovariectomized mice by preserving bone microarchitecture, mineral content, and inhibiting bone loss in treated mice as analyzed by Histomorphometry, MicroCT, and Raman spectroscopy. Oral administration of HxTME fraction resulted in the decreased percentage of F4/80+, CD11b+, and CD4+ RANKL+ T cells in OVX mice whereas pro-osteoclastic cytokine, IL6 was markedly reduced upon treatment with HxTME. On stimulation with PMA/Io and PHA, a significant decrease in proliferative response in the splenocytes of HxTME treated OVX mice was observed. Fatty acid profiling revealed that HxTME is rich in ω3 and ω6 polyunsaturated fatty acids (PUFAs), which have high nutraceutical properties and are known to play important role in growth, development and maintenance of health. Therefore, HxTME may be a good source of nutraceutical in the treatment of bone-related diseases particularly in postmenopausal osteoporosis and may be pursued as a potential candidate for treatment and management of osteoporosis.

Highlights

Bone is a dynamic tissue of the vertebrate body that provides mechanical support and serves as a major storage site for minerals and growth factors
We examined the inhibitory effects of HxTME on RANKL-induced osteoclastogenesis in the Bone Marrow Macrophage cells (BMMs) cells
The BMM cells were cultured in α MEM medium with various concentrations of HxTME fraction ranging from 3.125 to 25 μg/ml in presence of MacrophageColony Stimulating Factor (M-CSF) and RANKL for 7–8 days

Summary

Introduction

Bone is a dynamic tissue of the vertebrate body that provides mechanical support and serves as a major storage site for minerals and growth factors. The activation and formation of mature osteoclasts are achieved through binding of RANKL to its cognate receptor RANK present on the cell surface of osteoclast precursors [7]. Binding of RANK-RANKL triggers trimerization of TNF receptor-associated factor 6 (TRAF6) in osteoclast precursor cells which activates downstream signaling pathways such as p38 mitogen-activated protein kinase (MAPK), c-Jun-N-terminal kinase (JNK), extracellular signal-regulated kinase (ERK), and NF-kB. Activation of MAPKs and NF-kB results in expression of key transcriptional factors such as c-Fos and NFATc1 which are essential for osteoclasts differentiation [8]. Due to the pivotal role of these transcription factors and signaling molecules in osteoclastogenesis, it is important to regulate their activation and expression in the treatment of bone-related diseases

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Conclusion