Catharanthine tartrate ameliorates osteoclastogenesis by destabilizing HIF-1α

AUTHORS

Luqiong Cai, Chenxin Yu, Binli Zhao, Qihang Wu, Haibo Liang, Meng Zhou, Jiansen Miao, Jiangtao Luo, Jiake Xu, Haiming Jin, Youjin Pan

ABSTRACT

With the aging population, postmenopausal osteoporosis (PMOP), clinically manifested by reduced bone density, weakened skeletal strength, and compromised skeletal microstructure, has become the most prevalent type. The decline in estrogen levels fosters oxidative stress and osteoclastogenesis, which significantly enhance the activity of osteoclasts. Current treatments prefer to adopt relevant strategies to inactivate osteoclasts but come with unavoidable side effects. In our study, Catharanthine Tartrate (CAT), a derivative of the alkaloid catharanthine found in Catharanthus roseus, promised to be an effective therapy for PMOP. CAT inhibited RANKL-induced osteoclast differentiation and bone resorption in vitro. Moreover, CAT inhibited osteoclast activity by enhancing the ubiquitination-mediated proteasomal degradation of HIF-1α, which reduced oxidative stress and subsequently suppressed osteoclast activity. The inhibitory effects of CAT on osteoclast function and oxidative stress were reversed by DMOG, a known inhibitor of HIF-1α degradation. Next, an in vivo mouse experiment using the Ovariectomized (OVX) model to induce osteoporosis indicated that CAT enhanced bone mass density, bone structure, and bone remodeling. Our findings revealed that CAT inhibits PMOP through facilitating HIF-1α ubiquitination and degradation, suggesting a promising therapeutic approach for this disorder.

Dimeric R25CPTH(1–34) activates the parathyroid hormone-1 receptor in vitro and stimulates bone formation in osteoporotic female mice

AUTHORS

Minsoo Noh, Xiangguo Che, Xian Jin, Dong-Kyo Lee, Hyun-Ju Kim, Doo Ri Park, Soo Young Lee, Hunsang Lee, Thomas J Gardella, Je-Yong Choi, Sihoon Lee

ABSTRACT

Osteoporosis, characterized by reduced bone density and strength, increases fracture risk, pain, and limits mobility. Established therapies of parathyroid hormone (PTH) analogs effectively promote bone formation and reduce fractures in severe osteoporosis, but their use is limited by potential adverse effects. In the pursuit of safer osteoporosis treatments, we investigated R25CPTH, a PTH variant wherein the native arginine at position 25 is substituted by cysteine. These studies were prompted by our finding of high bone mineral density in a hypoparathyroidism patient with the R25C homozygous mutation, and we explored its effects on PTH type-1 receptor (PTH1R) signaling in cells and bone metabolism in mice. Our findings indicate that R25CPTH(1–84) forms dimers both intracellularly and extracellularly, and the synthetic dimeric peptide, R25CPTH(1–34), exhibits altered activity in PTH1R-mediated cyclic AMP (cAMP) response. Upon a single injection in mice, dimeric R25CPTH(1–34) induced acute calcemic and phosphaturic responses comparable to PTH(1–34). Furthermore, repeated daily injections increased calvarial bone thickness in intact mice and improved trabecular and cortical bone parameters in ovariectomized (OVX) mice, akin to PTH(1–34). The overall results reveal a capacity of a dimeric PTH peptide ligand to activate the PTH1R in vitro and in vivo as PTH, suggesting a potential path of therapeutic PTH analog development.

Hydroxyurea blunts mitochondrial energy metabolism and osteoblast and osteoclast differentiation exacerbating trabecular bone loss in sickle cell mice

AUTHORS

Ashish Kumar Tripathi, Sadaf Dabeer, Jun Song, Tatyana Vikulina, Susanne Roser-Page, Jessica A. Alvarez, David. R. Archer, M. Neale Weitzmann

ABSTRACT

Sickle cell disease (SCD) is a severe hematological disorder characterized by erythrocyte sickling that causes significant morbidity and mortality. Skeletal complications of SCD include a high incidence of bone loss, especially in vertebrae, leading to fragility fractures that contribute to disease burden. Whether hydroxyurea (HU), a front-line therapy for SCD ameliorates bone disease has not been established. To investigate HU action on SCD-related vertebral defects, we used HU-treated “Townes” mice, an SCD animal model and performed high-resolution micro-computed tomography (µCT) imaging to resolve bone volume and micro-architectural structure of cortical and trabecular bone, the two major compartments contributing to bone mass and strength. Our data revealed that cortical bone was significantly diminished in the vertebrae of skeletally mature (representing adults) and immature (representing children) SCD mice, while only mature mice lost trabecular bone mass. Administration of HU ameliorated cortical bone loss in mature SCD mice, but paradoxically promoted trabecular bone decline in both groups. We further investigated the mechanisms of HU action in wild-type C57BL6/J mice. HU caused dose-dependent trabecular bone loss due to diminished osteoclast and osteoblast function, indicative of a low bone turnover state. Mechanistic investigations in vitro revealed that HU impeded osteoblast-progenitor proliferation and early differentiation, and diminished osteoclastogenic cytokine production, blunting osteoclast formation as well as the activity of mature osteoclasts. HU further, suppressed mitochondrial, but not glycolytic energy metabolism in both differentiating osteoblasts and differentiated osteoclasts. Collectively, these findings reveal that despite ameliorating cortical bone loss, HU inhibits trabecular bone formation and resorption, by suppressing mitochondrial energy metabolism and blunting the differentiation and/or activity of osteoblasts and osteoclasts. Together HU drives a low bone turnover state culminating in trabecular bone loss. Further investigation into HU’s impact on bone in SCD patients is warranted for understanding and managing skeletal complications in this population.

Secreted protein TNA: a promising biomarker for understanding the adipose-bone axis and its impact on bone metabolism

AUTHORS

Shaobo Wu, Zhihao Xia, Liangliang Wei, Jiajia Ji, Yan Zhang & Dageng Huang

ABSTRACT

Background

Osteoporosis (OP) is a systemic bone disease characterized by reduced bone mass and deterioration of bone microstructure, leading to increased bone fragility. Platelets can take up and release cytokines, and a high platelet count has been associated with low bone density. Obesity is strongly associated with OP, and adipose tissue can influence platelet function by secreting adipokines. However, the biological relationship between these factors remains unclear.

Methods

We conducted differential analysis to identify OP platelet-related plasma proteins. And, making comprehensive analysis, including functional enrichment, protein-protein interaction network analysis, and Friends analysis. The key protein, Tetranectin (TNA/CLEC3B), was identified through screening. Then, we analyzed TNA’s potential roles in osteogenic and adipogenic differentiation using multiple RNA-seq data sets and validated its effect on osteoclast differentiation and bone resorption function through in vitro experiments.

Results

Six OP-platelet-related proteins were identified via differential analysis. Then, we screened the key protein TNA, which was found to be highly expressed in adipose tissue. RNA-seq data suggested that TNA may promote early osteoblast differentiation. In vitro experiments showed that knockdown of TNA expression significantly increased the expression of osteoclast markers, thereby promoting osteoclast differentiation and bone resorption.

Conclusions

We identified TNA as a secreted protein that inhibits osteoclast differentiation and bone resorption. While, it potentially promoted early osteoblast differentiation from bioinformatic results. TNA may play a role in bone metabolism through the adipose-bone axis.

Estrogen deficiency induces changes in bone matrix bound water that do not closely correspond with bone turnover

AUTHORS

Corinne E. Metzger, Peter Olayooye, Landon Y. Tak, Oli Culpepper, Alec N. LaPlant, Peter Jalaie, Pearl-Marie Andoh, Wikum Bandara, Olivia N. Reul, Andrew A. Tomaschke, Rachel K. Surowiec

ABSTRACT

Postmenopausal osteoporosis, marked by estrogen deficiency, is a major contributor to osteoporotic fractures, yet early prediction of fractures in this population remains challenging. Our goal was to explore the temporal changes in bone-specific inflammation, oxidative stress, bone turnover, and bone-matrix water, and their relationship with estrogen deficiency-induced modifications in bone structure and mechanical properties. Additionally, we sought to determine if emerging clinically translatable imaging techniques could capture early bone modifications prior to standard clinical imaging.

Two-month-old female Sprague Dawley rats (n = 48) underwent ovariectomy (OVX, n = 24) or sham operations (n = 24). A subgroup of n = 8 rats per group was sacrificed at 2-, 5-, and 10-weeks post-surgery to assess the temporal relationships of inflammation, oxidative stress, bone turnover, bone matrix water, mechanics, and imaging outcomes.

OVX rats exhibited higher body weight compared to sham rats at all time points. By 5-weeks, OVX animals showed elevated markers of inflammation and oxidative stress in cortical bone, which persisted throughout the study, while cortical bone formation rate did not differ from sham until 10-weeks. DXA outcomes did not reveal differences between OVX and sham at any time point. Bound water, assessed using ultrashort echo time magnetic resonance imaging (UTE MRI), was lower in OVX at the earliest time point (2-weeks) and reduced again at 10-weeks with no difference at 5-weeks.

These data demonstrate that bound water assessment using novel UTE MRI technology was lower at the earliest time point following OVX. However, no temporal relationship with bone turnover, inflammation, or oxidative stress was observed at the time points assessed in this study. These findings underscore both the increased need to understand bone hydration changes and highlight the usefulness of UTE MRI for non-invasive bone hydration measurements.

Epimedin A inhibits the PI3K/AKT/NF-κB signalling axis and osteoclast differentiation by negatively regulating TRAF6 expression

AUTHORS

Jun Li, Jia J. Wei, Cen H. Wu, Tao Zou, Hong Zhao, Tian Q. Huo, Cheng J. Wei, Ting Yang

ABSTRACT

Background

Epimedin A (EA) has been shown to suppress extensive osteoclastogenesis and bone resorption, but the effects of EA remain incompletely understood. The aim of our study was to investigate the effects of EA on osteoclastogenesis and bone resorption to explore the corresponding signalling pathways.

Methods

Rats were randomly assigned to the sham operation or ovariectomy group, and alendronate was used for the positive control group. The therapeutic effect of EA on osteoporosis was systematically analysed by measuring bone mineral density and bone biomechanical properties. In vitro, RAW264.7 cells were treated with receptor activator of nuclear factor kappa-B ligand (RANKL) and macrophage colony-stimulating factor (M-CSF) to induce osteoclast differentiation. Cell viability assays, tartrate-resistant acid phosphatase (TRAP) staining, and immunofluorescence were used to elucidate the effects of EA on osteoclastogenesis. In addition, the expression of bone differentiation-related proteins or genes was evaluated using Western blot analysis or quantitative polymerase chain reaction (PCR), respectively.

Results

After 3 months of oral EA intervention, ovariectomized rats exhibited increased bone density, relative bone volume, trabecular thickness, and trabecular number, as well as reduced trabecular separation. EA dose-dependently normalized bone density and trabecular microarchitecture in the ovariectomized rats. Additionally, EA inhibited the expression of TRAP and NFATc1 in the ovariectomized rats. Moreover, the in vitro results indicated that EA inhibits osteoclast differentiation by suppressing the TRAF6/PI3K/AKT/NF-κB pathway. Further studies revealed that the effect on osteoclast differentiation, which was originally inhibited by EA, was reversed when the TRAF6 gene was overexpressed.

Conclusions

The findings indicated that EA can negatively regulate osteoclastogenesis by inhibiting the TRAF6/PI3K/AKT/NF-κB axis and that ameliorating ovariectomy-induced osteoporosis in rats with EA may be a promising potential therapeutic strategy for the treatment of osteoporosis.