Bone biology
Abstract
Osteoarthritis (OA) is the most common joint disease. Controlling the complex pathogenesis is challenging, thus disease-modifying OA drugs are not available. Forkhead box O (FOXO) transcription factors contribute to cartilage homeostasis through autophagy and oxidative stress resistance. Here, we sought to discover FOXO activators and found that cyproheptadine, a histamine H1 receptor (HRH1) inverse agonist, promoted FOXO3 nuclear translocation and increased FOXO target genes while suppressing inflammation. In a murine OA model, cyproheptadine reduced structural joint tissue damage and pain behaviors. Mechanistically, the inhibition of HRH1 constitutive activity mediated the effects of cyproheptadine on calcium balance between endoplasmic reticulum (ER) and cytoplasm, and FOXO activation was part of this mechanism. The anti-inflammatory effect of cyproheptadine involved the inhibition of protein kinase C/NF-κB pathway. HRH1 inhibition also suppressed osteogenesis in mesenchymal stem cells and nerve growth factor expression, which are mechanisms of osteophyte formation and pain behaviors. Moreover, cyproheptadine suppressed ER stress-induced lipogenesis by upregulating insulin-induced gene 1. Our findings suggest that HRH1 constitutive activity controls important OA-promoting mechanisms and indicate that HRH1 inverse agonists are promising drug repurposing candidates for structure and pain improvement in OA.
Authors
Ichiro Kurakazu, Merissa Olmer, Hannah Swahn, Kevin Myers, Chelsea Kenvisay, Yukio Akasaki, Yasuharu Nakashima, Martin K. Lotz
Abstract
Inflammatory diseases contribute to secondary osteoporosis. Hypertension is a highly prevalent inflammatory condition that is clinically associated with reduced bone mineral density and increased risk for fragility fracture. In this study, we showed that a significant loss in bone mass and strength occurs in two pre-clinical models of hypertension. This accompanied increases in immune cell populations, including monocytes, macrophages, and IL-17A-producing T cell subtypes in the bone marrow of hypertensive mice. Neutralizing IL-17A in angiotensin (ang) II-infused mice blunted hypertension-induced loss of bone mass and strength due to decreased osteoclastogenesis. Likewise, the inhibition of the CSF-1 receptor blunted loss of bone mass and prevented loss of bone strength in hypertensive mice. In an analysis of UK Biobank data, circulating bone remodeling markers exhibited striking associations with blood pressure and bone mineral density in > 27,000 humans. These findings illustrate a potential mechanism by which hypertension activates immune cells in the bone marrow, encouraging osteoclastogenesis and eventual loss in bone mass and strength.
Authors
Elizabeth M. Hennen, Sasidhar Uppuganti, Néstor de la Visitación, Wei Chen, Jaya Krishnan, Lawrence A. Vecchi III, David M. Patrick, Mateusz Siedlinski, Matteo Lemoli, Rachel Delgado, Mark P. de Caestecker, Wenhan Chang, Tomasz J. Guzik, Rachelle W. Johnson, David G. Harrison, Jeffry S. Nyman
Abstract
Birth defects are the leading cause of infant mortality, and most inborn errors of development are multifactorial in origin, resulting from complex gene-environment interactions. Defining specific gene-environment interactions in the etiology and pathogenesis of congenital disorders is critically needed in the absence of genotype-phenotype correlation but is challenging. This is particularly true for congenital craniofacial anomalies, which account for approximately one-third of all birth defects, as they typically exhibit considerable inter-familial and intra-familial variability. A classic example of this is Treacher Collins Syndrome (TCS), which, although primarily caused by mutations in TCOF1, is characterized by considerable variability in the severity of mandibulofacial dysostosis. Here, we describe the genetic and environmental factors with converging effects that mechanistically contribute to the etiology and pathogenesis of craniofacial variation in this rare congenital disorder. We discovered in Tcof1+/- mouse models of TCS, that the combination of different endogenous levels of Tcof1/Treacle protein and reactive oxygen species (ROS) within distinct genetic backgrounds correlates with TCS phenotype severity. Furthermore, geometric morphometric analyses revealed that genotype largely determines the craniofacial shape, but redox status determines the size of individual bones. Taken together, our results highlight the roles of ROS and genomic instability in modulating the variability and phenotypic severity of craniofacial anomalies.
Authors
Sharien Fitriasari, Roberta Fiorino, Thoa H.K. Truong, Mary C. McKinney, Jill Dixon, Michael J. Dixon, Paul A. Trainor
Abstract
Achondroplasia, the most prevalent short-stature disorder, is caused by missense variants overactivating the fibroblast growth factor receptor 3 (FGFR3). As current surgical and pharmaceutical treatments only partially improve some disease features, we sought to explore a genetic approach. We show that an enhancer located 29 kb upstream of mouse Fgfr3 (–29E) is sufficient to confer a transgenic mouse reporter with a domain of expression in cartilage matching that of Fgfr3. Its CRISPR/Cas9-mediated deletion in otherwise WT mice reduced Fgfr3 expression in this domain by half without causing adverse phenotypes. Importantly, its deletion in mice harboring the ortholog of the most common human achondroplasia variant largely normalized long bone and vertebral body growth, markedly reduced spinal canal and foramen magnum stenosis, and improved craniofacial defects. Consequently, mouse achondroplasia is no longer lethal, and adults are overall healthy. These findings, together with high conservation of –29E in humans, open a path to develop genetic therapies for people with achondroplasia.
Authors
Marco Angelozzi, Arnaud Molin, Anirudha Karvande, Ángela Fernández-Iglesias, Samantha Whipple, Andrew M. Bloh, Véronique Lefebvre
Abstract
ATP-dependent chromatin remodeling protein ATRX is an essential regulator involved in maintenance of DNA structure and chromatin state and regulation of gene expression during development. ATRX was originally identified as the monogenic cause of X-linked α-thalassemia mental retardation (ATR-X) syndrome. Affected individuals display a variety of developmental abnormalities and skeletal deformities. Studies from others investigated the role of ATRX in skeletal development by tissue-specific Atrx knockout. However, the impact of ATRX during early skeletal development has not been examined. Using preosteoblast-specific Atrx conditional knockout mice, we observed increased trabecular bone mass and decreased osteoclast number in bone. In vitro coculture of Atrx conditional knockout bone marrow stromal cells (BMSCs) with WT splenocytes showed impaired osteoclast differentiation. Additionally, Atrx deletion was associated with decreased receptor activator of nuclear factor κ-B ligand (Rankl)/ osteoprotegerin (Opg) expression ratio in BMSCs. Notably, Atrx-deficient osteolineage cells expressed high levels of the neuropeptide cocaine- and amphetamine-regulated transcript prepropeptide (Cartpt). Mechanistically, ATRX suppresses Cartpt transcription by binding to the promoter, which is otherwise poised for Cartpt expression by RUNX2 binding to the distal enhancer. Finally, Cartpt silencing in Atrx conditional knockout BMSCs rescued the molecular phenotype by increasing the Rankl/Opg expression ratio. Together, our data show a potent repressor function of ATRX in restricting Cartpt expression during skeletal development.
Authors
Yi-Ting Chen, Ming-Ming Jiang, Carolina Leynes, Mary Adeyeye, Camilla F. Majano, Barakat Ibrahim, Urszula Polak, George Hung, Zixue Jin, Denise G. Lanza, Lan Liao, Brian Dawson, Yuqing Chen-Evenson, Oscar E. Ruiz, Richard J. Gibbons, Jason D. Heaney, Yangjin Bae, Brendan Lee
Endothelial BMAL1 decline during aging leads to bone loss by destabilizing extracellular fibrillin-1
Abstract
The occurrence of aging is intricately associated with alterations in circadian rhythms that coincide with stem cell exhaustion. Nonetheless, the extent to which the circadian system governs skeletal aging remains inadequately understood. Here, we noticed that skeletal aging in male mice was accompanied by a decline in a core circadian protein, BMAL1, especially in bone marrow endothelial cells (ECs). Using male mice with endothelial KO of aryl hydrocarbon receptor nuclear translocator–like protein 1 (Bmal1), we ascertained that endothelial BMAL1 in bone played a crucial role in ensuring the stability of an extracellular structural component, fibrillin-1 (FBN1), through regulation of the equilibrium between the extracellular matrix (ECM) proteases thrombospondin type 1 domain–containing protein 4 (THSD4) and metalloproteinase with thrombospondin motifs 4 (ADAMTS4), which promote FBN1 assembly and breakdown, respectively. The decline of endothelial BMAL1 during aging prompted excessive breakdown of FBN1, leading to persistent activation of TGF-β/SMAD3 signaling and exhaustion of bone marrow mesenchymal stem cells. Meanwhile, the free TGF-β could promote osteoclast formation. Further analysis revealed that activation of ADAMTS4 in ECs lacking BMAL1 was stimulated by TGF-β/SMAD3 signaling through an ECM-positive feedback mechanism, whereas THSD4 was under direct transcriptional control by endothelial BMAL1. Our investigation has elucidated the etiology of bone aging in male mice by defining the role of ECs in upholding the equilibrium within the ECM, consequently coordinating osteogenic and osteoclastic activities and retarding skeletal aging.
Authors
Ying Yin, Qingming Tang, Jingxi Yang, Shiqi Gui, Yifan Zhang, Yufeng Shen, Xin Zhou, Shaoling Yu, Guangjin Chen, Jiwei Sun, Zhenshuo Han, Luoying Zhang, Lili Chen
Abstract
Local immunoinflammatory events instruct skeletal stem cells (SSCs) to repair/regenerate bone after injury, but mechanisms are incompletely understood. We hypothesized that specialized Regulatory T (Treg) cells are necessary for bone repair and interact directly with SSCs through organ-specific messages. Both in human patients with bone fracture and mouse model of bone injury, we identified a bone injury-responding Treg subpopulation with bone-repair capacity marked by CCR8. Local production of CCL1 induced a massive migration of CCR8+ Treg cells from periphery to the injury site. Depending on secretion of progranulin (PGRN), a protein encoded by the granulin (Grn) gene, CCR8+ Treg cells supported the accumulation and osteogenic differentiation of SSCs, and thereby bone repair. Mechanistically, we revealed that CCL1 enhanced expression level of basic leucine zipper ATF-like transcription factor (BATF) in CCR8+ Treg cells, which bound to Grn promoter and increased Grn translational output and then PGRN secretion. Together, our work provides a new perspective in osteoimmunology and highlights possible ways of manipulating Treg cell signaling to enhance bone repair and regeneration.
Authors
Ruiying Chen, Xiaomeng Zhang, Bin Li, Maurizio S. Tonetti, Yijie Yang, Yuan Li, Beilei Liu, Shujiao Qian, Yingxin Gu, Qingwen Wang, Kairui Mao, Hao Cheng, Hongchang Lai, Junyu Shi
Abstract
Translation of mRNA to protein is tightly regulated by tRNAs, which are subject to various chemical modifications that maintain the structure, stability and function. Deficiency of tRNA N7-methylguanosine (m7G) modification in patients causes a type of primordial dwarfism, but the underlying mechanism remains unknown. Here we report the loss of m7G rewires cellular metabolism, leading to the pathogenesis of primordial dwarfism. Conditional deletion of the catalytic enzyme Mettl1 or missense mutation of the scaffold protein Wdr4 severely impaired endochondral bone formation and bone mass accrual. Mechanistically, Mettl1 knockout decreased abundance of m7G-modified tRNAs and inhibited translation of mRNAs relating to cytoskeleton and Rho GTPase signaling. Meanwhile, Mettl1 knockout enhanced cellular energy metabolism despite of incompetent proliferation and osteogenic commitment. Further exploration revealed that impaired Rho GTPase signaling upregulated branched-chain amino acid transaminase 1 (BCAT1) level that rewired cell metabolism and restricted intracellular α-ketoglutarate (αKG). Supplementation of αKG ameliorated the skeletal defect of Mettl1-deficient mice. In addition to the selective translation of metabolism-related mRNAs, we further revealed that Mettl1 knockout globally regulated translation via integrated stress response (ISR) and mammalian target of rapamycin complex 1 (mTORC1) signaling. Restoring translation either by targeting ISR or mTORC1 aggravated bone defects of Mettl1-deficient mice. Overall, our study unveils a critical role of m7G tRNA modification in bone development by regulating cellular metabolism, and indicates that suspension of translation initiation as quality control mechanism in response to tRNA dysregulation.
Authors
Qiwen Li, Shuang Jiang, Kexin Lei, Hui Han, Yaqian Chen, Weimin Lin, Qiuchan Xiong, Xingying Qi, Xinyan Gan, Rui Sheng, Yuan Wang, Yarong Zhang, Jieyi Ma, Tao Li, Shuibin Lin, Chenchen Zhou, Demeng Chen, Quan Yuan
Abstract
The periosteum contains skeletal stem/progenitor cells that contribute to bone fracture healing. However, the in vivo identity of periosteal skeletal stem cells (P-SSCs) remains unclear, and membrane protein markers of P-SSCs that facilitate tissue engineering are needed. Here, we identified integral membrane protein 2A (Itm2a) enriched in SSCs using single-cell transcriptomics. Itm2a+ P-SSCs displayed clonal multipotency and self-renewal and sat at the apex of their differentiation hierarchy. Lineage-tracing experiments showed that Itm2a selectively labeled the periosteum and that Itm2a+ cells were preferentially located in the outer fibrous layer of the periosteum. The Itm2a+ cells rarely expressed CD34 or Osx, but expressed periosteal markers such as Ctsk, CD51, PDGFRA, Sca1, and Gli1. Itm2a+ P-SSCs contributed to osteoblasts, chondrocytes, and marrow stromal cells upon injury. Genetic lineage tracing using dual recombinases showed that Itm2a and Prrx1 lineage cells generated spatially separated subsets of chondrocytes and osteoblasts during fracture healing. Bone morphogenetic protein 2 (Bmp2) deficiency or ablation of Itm2a+ P-SSCs resulted in defects in fracture healing. ITM2A+ P-SSCs were also present in the human periosteum. Thus, our study identified a membrane protein marker that labels P-SSCs, providing an attractive target for drug and cellular therapy for skeletal disorders.
Authors
Wenhui Xing, Heng Feng, Bo Jiang, Bo Gao, Jiping Liu, Zaiqi Xie, Yazhuo Zhang, Xuye Hu, Jun Sun, Matthew B. Greenblatt, Bo O. Zhou, Weiguo Zou
Abstract
Chronic low back pain (LBP) can severely affect daily physical activity. Aberrant osteoclast-mediated resorption leads to porous endplates for the sensory innervation to cause LBP. Here, we report that the expression of proton-activated chloride (PAC) channel is induced during osteoclast differentiation in the porous endplates via a RANKL-NFATc1 signaling pathway. Extracellular acidosis evokes robust PAC currents in osteoclasts. An acidic environment of porous endplates and elevated PAC activation-enhanced osteoclast fusion provoke LBP. Further, we find that genetic knockout of PAC gene Pacc1 significantly reduces endplate porosity and spinal pain in a mouse LBP model, but it does not affect bone development or homeostasis of bone mass in adult mice. Moreover, both osteoclast bone resorptive compartment environment and PAC traffic from the plasma membrane to endosomes to form an intracellular organelle Cl channel have low pH around 5.0. The low pH environment activates PAC channel to increase sialyltransferase St3gal1 expression and sialylation of TLR2 in initiation of osteoclast fusion. Aberrant osteoclast-mediated resorption is also found in most skeletal disorders, including osteoarthritis, ankylosing spondylitis, rheumatoid arthritis, heterotopic ossification, enthesopathy. Thus, elevated Pacc1 expression and PAC activity could be a potential therapeutic target for LBP and osteoclast-associated pain.
Authors
Peng Xue, Weixin Zhang, Mengxi Shen, Junhua Yang, Jiachen Chu, Shenyu Wang, Mei Wan, Junying Zheng, Zhaozhu Qiu, Xu Cao
Copyright © 2025 American Society for Clinical Investigation
ISSN: 0021-9738 (print), 1558-8238 (online)