Betulinic Acid Protects From Bone Loss in Ovariectomized Mice and Suppresses RANKL-Associated Osteoclastogenesis by Inhibiting the MAPK and NFATc1 Pathways.

Jiyong Wei,Kun Tian,Yanni Lan,Jiake Xu,Jinmin Zhao,Chengming Wei,Chunbo Lin,Qian Liu,Liwei Wu,Yicheng Li,Yuan Yang

doi:10.3389/fphar.2020.01025

Abstract

Osteoclasts with elevated bone resorption are commonly present in postmenopausal osteoporosis, and other osteolytic pathologies. Therefore, suppressing osteoclast generation and function has been the main focus of osteoporosis treatment. Betulinic acid (BA) represents a triterpenoid mainly purified from the bark of Betulaceae. BA shows multiple biological activities, including antitumor and anti-HIV properties, but its effect on osteolytic conditions is unknown. Here, BA suppressed receptor activator of nuclear factor‐κB ligand (RANKL)‐associated osteoclastogenesis and bone resorptive function, as assessed by tartrate‐resistant acid phosphatase (TRAP) staining, fibrous actin ring generation, and hydroxyapatite resorption assays. Mechanistically, BA downregulated the expression of osteoclastic-specific genes. Western blot analysis revealed that BA significantly interrupted ERK, JNK and p38 MAPK activation as well as intracellular reactive oxygen species (ROS) production, thus altering c-Fos and NFATc1 activation. Corroborating the above findings in cell-based assays, BA prevented ovariectomy-associated bone loss in an animal model. In conclusion, these findings suggest that BA can inhibit osteoclast generation and function as well as the RANKL signaling pathway, and might be used for treating osteoclast-related osteoporosis.

Highlights

Osteoporosis features decreased bone mass and strength, and characteristic bone microstructure degradation with increasing bone brittleness and fractures (Xie et al, 2019)
Many signaling molecules contributing to osteoclast induction and proliferation play positive or negative regulatory roles; the most critical two factors include macrophage colony stimulating factor (M-CSF) and receptor activator of nuclear factor-kB ligand (RANKL), which contribute to osteoclast differentiation (Boyle et al, 2003; Boyce, 2013)
At non-cytotoxic doses, various Betulinic acid (BA) amounts were supplemented to bone marrow macrophages (BMMs) cultures alongside RANKL induction to assess the effects on osteoclastogenesis

Summary

Introduction

Osteoporosis features decreased bone mass and strength, and characteristic bone microstructure degradation with increasing bone brittleness and fractures (Xie et al, 2019). TRAF6 induces a series of downstream molecular networks such as the nuclear factor-kB (NF-kB) and mitogen-activated protein kinase (MAPK) pathways, involving extracellular signal-regulated kinase (ERK), c-Jun Nterminal kinases (JNK) and p38 mitogen-activated protein kinase (p38) (Yuan et al, 2015). These networks upregulate and activate transcription factors, including c-Fos and NFATc1, in a synergistic manner; NFATc1 constitutes the key molecule controlling the differentiation of osteoclasts (Takayanagi et al, 2002). RANKL induced signaling pathway is a therapeutic target for abnormal bone resorption

Methods

Results

Conclusion