DJ-1-mediated p62 degradation delays intervertebral disc degeneration by inhibiting apoptosis of nucleus pulposus cells.

Intervertebral disc degeneration (IDD) is the most important pathological basis of degenerative spinal diseases, for which effective interventions are still lacking. Oxidative stress is considered to be one of the leading pathological mechanisms contributing to IDD. However, the exact role of DJ-1 as an essential member of the antioxidant defense system in IDD is still unclear. Therefore, the aim of this study was to investigate the role played by DJ-1 in IDD and to reveal its potential molecular mechanisms. Western blot and immunohistochemical staining assays were performed to detect the expression of DJ-1 in degenerative nucleus pulposus cells (NPCs). After overexpression of DJ-1 in NPCs by lentiviral transfection, DCFH-DA and MitoSOX fluorescent probes were used to evaluate the levels of reactive oxygen species (ROS); while western blot, TUNEL staining, and Caspase-3 activity were used to assess apoptosis. Immunofluorescence staining was used to demonstrate the relationship between DJ-1 and p62. After inhibition of lysosomal degradation function with chloroquine, p62 degradation and apoptosis in DJ-1 overexpressing NPCs were further examined. In vivo, we assessed the therapeutic effect of upregulated DJ-1 on IDD by X-ray, MRI and Safranin O-Fast green staining. The protein expression of DJ-1 was significantly decreased in degenerated NPCs, accompanied by increased apoptosis. However, overexpression of DJ-1 significantly inhibited the elevated ROS levels and apoptosis in NPCs under oxidative stress. Mechanistically, our results showed that upregulation of DJ-1 promoted p62 degradation via the autophagic lysosomal pathway and that the protective effect of DJ-1 on NPCs under oxidative stress was partially mediated by promoting lysosomal pathway degradation of p62. Moreover, intradiscal injection of adeno-associated virus for overexpression of DJ-1 mitigated the progression of IDD in rats. This study reveals that DJ-1 maintains the homeostasis of NPCs by promoting the degradation of p62 through the autophagic lysosomal pathway, suggesting that DJ-1 is a promising new target for IDD intervention.

Ehm2 transcript variant 1 inhibits breast cancer progression and increases E-cadherin stability.

Ehm2/1, an Ehm2 transcript variant, regulates the cytoskeleton by binding to plasma membrane proteins. However, the role of Ehm2/1 in breast cancer development remains poorly understood. This study shows that, the expression of Ehm2/1 was decreased in breast cancer and that patients with low Ehm2/1 expression had a significantly poorer prognosis than those with high expression of Ehm2/1. Overexpression of Ehm2/1 in MCF-7 breast cancer cells inhibited cell migration and invasion. Ehm2/1 markedly increased the stability and half-life of E-cadherin. Moreover, Ehm2/1 was collocated with E-cadherin in the plasma membrane of MCF-7 cells. Furthermore, downregulation of Ehm2/1 promoted ubiquitination of E-cadherin, whereas overexpression of Ehm2/1 inhibited ubiquitination of E-cadherin. These results suggest that Ehm2/1 could suppress the migration and invasion of breast cancer cells by increasing E-cadherin stability.

2021 revised algorithm for the management of knee osteoarthritis-the Chinese viewpoint.

AIM: The European Society for Clinical and Economic Aspects of Osteoporosis, Osteoarthritis and Musculoskeletal Diseases (ESCEO) algorithm for the management of knee osteoarthritis (OA) is available worldwide from 2014, but in 2019 an update was published. Based on this algorithm, a Working Group (WG), including ESCEO members and Chinese experts, wished to see how the new ESCEO algorithm was perceived by Chinese experts in knee OA and how it was integrated into their clinical practice. METHODS: A WG was held between members of the international ESCEO task force and a group of Chinese experts. RESULTS: Non-pharmacological approach should be combined with pharmacological interventions. In step 1, symptomatic slow-acting drugs for osteoarthritis (SYSADOA) are the most important background drugs. Evidence, supported by high-quality research, is available only for crystalline glucosamine sulfate (pCGS) and chondroitin sulfate. Topical NSAIDs could be used as an additional option. In step 2, oral NSAIDs could be useful, but cardiovascular/renal/gastrointestinal profiles of the patients should be considered. Intra-articular hyaluronic acid and corticosteroids are alternative to oral NSAIDs, but the evidence is still limited. If steps 1 and 2 are not sufficient, weak opioids could be used. Overall, the conclusions of the ESCEO algorithm are accepted in China for products available in this country. The WG suggests the importance of economic studies, specifically made in China. CONCLUSION: This work provides evidence-based advice to establish a treatment algorithm in knee OA, for practical implementation in clinical practice in China.

The prevalence and clinical characteristics of thoracic spinal stenosis: a systematic review.

PURPOSE: Thoracic spinal stenosis (TSS) is a rare disease secondary to multiple pathological changes that differ in prevalence and clinical characteristics. The epidemiological characteristics of these pathologies are largely unknown due to the limited case samples and regional differences. Therefore, a systematic review was conducted to elucidate the prevalence and clinical characteristics of TSS. METHODS: Case series and case reports on the ossification of the posterior longitudinal ligaments (OPLL), ossification of the ligamentum flavum (OLF) and thoracic disk herniation (TDH) were screened from PubMed, Embase and Web of Science databases and systematically reviewed. Epidemiological, demographic and segmental distribution data were extracted and analyzed. RESULTS: A total of 129 studies including 1935 subjects were selected, of which 361 (18.7%) were diagnosed with OPLL, 804 (41.5%) with OLF, 143 (7.4%) with OPLL + OLF and 627 (32.4%) with TDH. Most reports were from China, Japan and USA. Thoracic OPLL occurred mostly at the middle-thoracic spine (43.4%), while OLF predominately occurred at the lower-thoracic spine (63.1%). TDH was mainly localized in the middle (46.0%) and lower-thoracic (50.3%) spine. Thirty-two studies involving 524 patients described tandem spinal stenosis, of which 52.1% had accompanying cervical diseases and 35.9% lumbar diseases. CONCLUSIONS: There are significant differences in the age, sex and segment distribution characteristics of different pathologies leading to TSS. Tandem spinal stenosis is not uncommon and should be considered when diagnosing TSS. Our findings provide new insights into the prevalence and clinical characteristics of TSS and can help reduce misdiagnosis.

MiR-199b-5p inhibits osteogenic differentiation in ligamentum flavum cells by targeting JAG1 and modulating the Notch signalling pathway.

Ossification of the ligamentum flavum (OLF) is a pathology almost only reported in East Asian countries. The leading cause of OLF is thoracic spinal canal stenosis and myelopathy. In this study, the role of miR-199b-5p and jagged 1 (JAG1) in primary ligamentum flavum cell osteogenesis was examined. MiR-199b-5p was found to be down-regulated during osteogenic differentiation in ligamentum flavum cells, while miR-199b-5p overexpression inhibited osteogenic differentiation. In addition, JAG1 was found to be up-regulated during osteogenic differentiation in ligamentum flavum cells, while JAG1 knockdown via RNA interference caused an inhibition of Notch signalling and osteogenic differentiation. Moreover, target prediction analysis and dual luciferase reporter assays supported the notion that JAG1 was a direct target of miR-199b-5p, with miR-199b-5p found to down-regulate both JAG1 and Notch. Further, JAG1 knockdown was demonstrated to block the effect of miR-199b-5p inhibition. These findings imply that miR-199b-5p performs an inhibitory role in osteogenic differentiation in ligamentum flavum cells by potentially targeting JAG1 and influencing the Notch signalling pathway.

MiR-132-3p Regulates the Osteogenic Differentiation of Thoracic Ligamentum Flavum Cells by Inhibiting Multiple Osteogenesis-Related Genes.

Ossification of the ligamentum flavum (OLF) is a disorder of heterotopic ossification of spinal ligaments and is the main cause of thoracic spinal canal stenosis. Previous studies suggested that miR-132-3p negatively regulates osteoblast differentiation. However, whether miR-132-3p is involved in the process of OLF has not been investigated. In this study, we investigated the effect of miR-132-3p and its target genes forkhead box O1 (FOXO1), growth differentiation factor 5 (GDF5) and SRY-box 6 (SOX6) on the osteogenic differentiation of ligamentum flavum (LF) cells. We demonstrated that miR-132-3p was down-regulated during the osteogenic differentiation of LF cells and negatively regulated the osteoblast differentiation. Further, miR-132-3p targeted FOXO1, GDF5 and SOX6 and down-regulated the protein expression of these genes. Meanwhile, FOXO1, GDF5 and SOX6 were up-regulated after osteogenic differentiation and the down-regulation of endogenous FOXO1, GDF5 or SOX6 suppressed the osteogenic differentiation of LF cells. In addition, we also found FOXO1, GDF5 and SOX6 expression in the ossification front of OLF samples. Overall, these results suggest that miR-132-3p inhibits the osteogenic differentiation of LF cells by targeting FOXO1, GDF5 and SOX6.

Understanding the impact of nanoplastics on reproductive health: Exposure pathways, mechanisms, and implications.

Microplastic pollution is a pressing global environmental concern with particular urgency surrounding the issue of nanoplastic particles. Plastic products exhibit a remarkable persistence in natural ecosystems, resisting easy degradation. Nanoplastics, characterized by their diminutive size, possess distinct properties when compared to their larger counterparts, which could potentially render them more ecologically detrimental. Microplastics themselves serve as carriers for toxic and hazardous substances, such as plastic additives, that enter and persist in the environmental cycle. Importantly, nanoplastics exhibit enhanced bioavailability upon entering the food chain. Notably, studies have demonstrated the adverse effects of nanoplastics on the reproductive function of aquatic organisms, and evidence of micro- and nanoplastics have emerged within human reproductive organs, including the placenta. However, a knowledge gap persists regarding the impacts of nanoplastics on the reproductive systems of mammals and, indeed, humans. This paper aims to elucidate the less frequently discussed sources and distribution of nanoplastics in the environment, along with the pathways of human exposure. We also emphasize the extent to which nanoplastics accumulate within the reproductive systems of organisms. Subsequently, we present an in-depth analysis of the effects of nanoplastics and their associated contaminants on mammalian and human reproductive health. The mechanisms through which nanoplastics contribute to reproductive disorders are comprehensively explored, highlighting their potential to disrupt endocrine levels in mammals and humans. Additionally, we scrutinize and discuss studies on biotoxicity of nanoplastics, offering insights into potential areas for future research.

Circular RNA CircFOXO3 Functions as a Competitive Endogenous RNA for Acid-Sensing Ion Channel Subunit 1 Mediating Oxeiptosis in Nucleus Pulposus.

Oxeiptosis is a reactive oxygen species (ROS)-induced pathway of cell death. The involvement of circular RNAs (circRNAs) has been confirmed in the incidence and progression of intervertebral disc degeneration (IVDD). However, whether oxeiptosis occurs in IVDD and how circRNAs regulate oxeiptosis is still unclear. In this study, we discovered that oxeiptosis could be induced in nucleus pulposus cells (NPCs), and circFOXO3 was significantly upregulated after oxeiptosis induction. Transfection using circFOXO3 small interfering RNA (siRNA) significantly inhibited oxeiptosis in NPCs. Mechanistically, circFOXO3 upregulated acid-sensing ion channel subunit 1 (ASIC1) expression by functioning as a molecular sponge for miR-185-3p and miR-939-5p. Subsequent rescue experiments validated that circFOXO3 could regulate oxeiptosis in NPCs via the miR-185-3p/miR-939-5p-ASIC1 axis. Further research on ASIC1 functions indicated that this regulation was achieved by affecting the Calcium ion (Ca2+) influx mediated by ASIC1. A mouse IVDD model was established, and silencing circFOXO3 in vivo was found to inhibit IVDD development and the activation of the oxeiptosis-related pathway. Overall, circFOXO3 is one of the factors contributing to the progression of IVDD by mediating oxeiptosis.

Genetic insights into the 'sandwich fusion' subtype of Klippel-Feil syndrome: novel FGFR2 mutations identified by 21 cases of whole-exome sequencing.

BACKGROUND: Klippel-Feil syndrome (KFS) is a rare congenital disorder characterized by the fusion of two or more cervical vertebrae during early prenatal development. This fusion results from a failure of segmentation during the first trimester. Although six genes have previously been associated with KFS, they account for only a small proportion of cases. Among the distinct subtypes of KFS, "sandwich fusion" involving concurrent fusion of C0-1 and C2-3 vertebrae is particularly noteworthy due to its heightened risk for atlantoaxial dislocation. In this study, we aimed to investigate novel candidate mutations in patients with "sandwich fusion." METHODS: We collected and analyzed clinical data from 21 patients diagnosed with "sandwich fusion." Whole-exome sequencing (WES) was performed, followed by rigorous bioinformatics analyses. Our focus was on the six known KFS-related genes (GDF3, GDF6, MEOX1, PAX1, RIPPLY2, and MYO18). Suspicious mutations were subsequently validated through in vitro experiments. RESULTS: Our investigation revealed two novel exonic mutations in the FGFR2 gene, which had not previously been associated with KFS. Notably, the c.1750A > G variant in Exon 13 of FGFR2 was situated within the tyrosine kinase domain of the protein, in close proximity to several established post-translational modification sites. In vitro experiments demonstrated that this certain mutation significantly impacted the function of FGFR2. Furthermore, we identified four heterozygous candidate variants in two genes (PAX1 and MYO18B) in two patients, with three of these variants predicted to have potential clinical significance directly linked to KFS. CONCLUSIONS: This study encompassed the largest cohort of patients with the unique "sandwich fusion" subtype of KFS and employed WES to explore candidate mutations associated with this condition. Our findings unveiled novel variants in PAX1, MYO18B, and FGFR2 as potential risk mutations specific to this subtype of KFS.

Involvement of DJ-1 in the pathogenesis of intervertebral disc degeneration via hexokinase 2-mediated mitophagy.

Intervertebral disc degeneration (IDD) is an important pathological basis for degenerative spinal diseases and is involved in mitophagy dysfunction. However, the molecular mechanisms underlying mitophagy regulation in IDD remain unclear. This study aimed to clarify the role of DJ-1 in regulating mitophagy during IDD pathogenesis. Here, we showed that the mitochondrial localization of DJ-1 in nucleus pulposus cells (NPCs) first increased and then decreased in response to oxidative stress. Subsequently, loss- and gain-of-function experiments revealed that overexpression of DJ-1 in NPCs inhibited oxidative stress-induced mitochondrial dysfunction and mitochondria-dependent apoptosis, whereas knockdown of DJ-1 had the opposite effect. Mechanistically, mitochondrial translocation of DJ-1 promoted the recruitment of hexokinase 2 (HK2) to damaged mitochondria by activating Akt and subsequently Parkin-dependent mitophagy to inhibit oxidative stress-induced apoptosis in NPCs. However, silencing Parkin, reducing mitochondrial recruitment of HK2, or inhibiting Akt activation suppressed DJ-1-mediated mitophagy. Furthermore, overexpression of DJ-1 ameliorated IDD in rats through HK2-mediated mitophagy. Taken together, these findings indicate that DJ-1 promotes HK2-mediated mitophagy under oxidative stress conditions to inhibit mitochondria-dependent apoptosis in NPCs and could be a therapeutic target for IDD.

ß-Receptor blocker enhances the anabolic effect of PTH after osteoporotic fracture.

The autonomic nervous system plays a crucial role in regulating bone metabolism, with sympathetic activation stimulating bone resorption and inhibiting bone formation. We found that fractures lead to increased sympathetic tone, enhanced osteoclast resorption, decreased osteoblast formation, and thus hastened systemic bone loss in ovariectomized (OVX) mice. However, the combined administration of parathyroid hormone (PTH) and the ß-receptor blocker propranolol dramatically promoted systemic bone formation and osteoporotic fracture healing in OVX mice. The effect of this treatment is superior to that of treatment with PTH or propranolol alone. In vitro, the sympathetic neurotransmitter norepinephrine (NE) suppressed PTH-induced osteoblast differentiation and mineralization, which was rescued by propranolol. Moreover, NE decreased the PTH-induced expression of Runx2 but enhanced the expression of Rankl and the effect of PTH-stimulated osteoblasts on osteoclastic differentiation, whereas these effects were reversed by propranolol. Furthermore, PTH increased the expression of the circadian clock gene Bmal1, which was inhibited by NE-ßAR signaling. Bmal1 knockdown blocked the rescue effect of propranolol on the NE-induced decrease in PTH-stimulated osteoblast differentiation. Taken together, these results suggest that propranolol enhances the anabolic effect of PTH in preventing systemic bone loss following osteoporotic fracture by blocking the negative effects of sympathetic signaling on PTH anabolism.

Correlation between clinicopathological characteristics and the clinical prognosis of patients with gastroenteropancreatic neuroendocrine tumors.

Gastroenteropancreatic (GEP) neuroendocrine tumors (NETs) are one of the most common types of NETs, accounting for 65-75% of all NETs. However, epidemiological characteristics of patients with GEP-NETs in China are still lacking. The present retrospective study aimed to investigate the local epidemiology of GEP-NETs and assess the prognostic factors in China. The data of 267 patients with GEP-NETs who were admitted to the First Affiliated Hospital of Bengbu Medical College (Bengbu, China) and the Affiliated Hospital of West Anhui Health Vocational College (Lu'an, China) were retrospectively reviewed. The clinical and pathological characteristics of the patients, as well as follow-up information, were collected, and the 5-year survival rate was calculated. Kaplan-Meier curves and log-rank analysis were used to analyze the prognostic factors. The stomach (100/267; 37.5%) was the most common site of GEP-NETs and the liver (25/39; 64.1%) was the most common metastatic site. A total of 166 (62.2%) and 219 (82.0%) patients had positive results for chromogranin A (CgA) and synaptophysin (Syn), respectively. The percentage of patients with tumor grade G1, G2 and G3 was 33.3, 21.0 and 45.7%, respectively. The 5-year overall survival rate was 79.7%, and the age, tumor site, distant metastasis and tumor grading upon diagnosis were all prognostic factors. In conclusion, the present case series investigated the epidemiology and prognostic factors of GEP-NETs in China. CgA and Syn could be used as diagnostic markers for NETs and the stomach was the most common primary tumor site. Lymph node metastasis, tumor site, distant metastasis and tumor grading were important prognostic factors.

Interleukin-17A Promotes Proliferation and Osteogenic Differentiation of Human Ligamentum Flavum Cells Through Regulation of ß-Catenin Signaling.

STUDY DESIGN: A basic experimental study. OBJECTIVE: To elucidate the role and mechanism of interleukin (IL)-17A in thoracic ossification of the ligamentum flavum (TOLF). SUMMARY OF BACKGROUND DATA: TOLF is characterized by the replacement of the thoracic ligamentum flavum with ossified tissue and is one of the leading causes of thoracic spinal stenosis. IL-17A is an important member of the IL-17 family that has received widespread attention for its key contributions to the regulation of bone metabolism and heterotopic ossification. However, it is unclear whether IL-17A is involved in TOLF. MATERIALS AND METHODS: Cell counting kit-8 assay and 5-ethynyl-2'-deoxyuridine staining were performed to assess the proliferation of ligamentum flavum cells (LFCs). Alkaline phosphatase activity assay, Alizarin red staining, and protein level expression of osteogenic-related genes were used to evaluate the osteogenic differentiation potential of LFCs. The effect of IL-17A on the proliferation and osteogenic differentiation of LFCs was further assessed after silencing ß-catenin by transfection with small interfering RNA. In addition, the possible source of IL-17A was further demonstrated by coculture assays of T helper 17 (Th17) cells with LFCs. Student t test was used for comparisons between groups, and the one-way analysis of variance, followed by the Tukey post hoc test, was used for comparison of more than two groups. RESULTS: IL-17A was elevated in TOLF tissue compared with normal ligamentum flavum. IL-17A stimulation promoted the proliferation and osteogenic differentiation of LFCs derived from patients with TOLF. We found that IL-17A promoted the proliferation and osteogenic differentiation of LFCs by regulating the ß-catenin signaling. Coculture of Th17 cells with LFCs enhanced ß-catenin signaling-mediated proliferation and osteogenic differentiation of LFCs. However, these effects were markedly attenuated after the neutralization of IL-17A. CONCLUSIONS: This is the first work we are aware of to highlight the importance of IL-17A in TOLF. IL-17A secreted by Th17 cells in the ligamentum flavum may be involved in the ossification of the microenvironment by regulating ß-catenin signaling to promote the proliferation and osteogenic differentiation of LFCs.

Bone mass loss in chronic heart failure is associated with sympathetic nerve activation.

PURPOSE: Chronic heart failure causes osteoporosis, but the mechanism remains unclear. The sympathetic nerve plays an important role in both bone metabolism and cardiovascular function. METHODS: Thirty-six adult male SD rats were randomly divided into the following four groups: sham surgery (Sham) group, guanethidine (GD) group, abdominal transverse aorta coarctation-induced heart failure + normal saline (TAC) group, and TAC + guanethidine (TAC + GD) group. Normal saline (0.9 % NaCl) or guanethidine (40 mg/kg/ml) was intraperitoneally injected daily for 5 weeks. Then, DXA, micro-CT, ELISA and RT-PCR analyses were performed 12 weeks after treatment. RESULTS: The bone loss in rats subjected to TAC-induced chronic heart failure and chemical sympathectomy with guanethidine was increased. Serum norepinephrine levels were increased in rats with TAC-induced heart failure but were decreased in TAC-induced heart failure rats treated with guanethidine. The expression of α2A adrenergic receptor, α2C adrenergic receptor, osteoprotegerin (OPG), and osteocalcin in the tibia decreased in the TAC-induced heart failure group, and the expression of ß1 adrenergic receptor, ß2 adrenergic receptor, receptor activator of nuclear factor-κ B ligand (RANKL), and RANKL/OPG in the tibia increased in the heart failure group. In addition, these changes in gene expression levels were rescued by chemical sympathectomy with guanethidine. CONCLUSIONS: TAC-induced chronic heart failure is associated with bone mass loss, and the sympathetic nerve plays a significant role in heart failure-related bone mass loss. MINI ABSTRACT: The present study supports the hypothesis that heart failure is related to bone loss, and the excessive activation of sympathetic nerves participates in this pathophysiological process. The present study suggests a potential pathological mechanism of osteoporosis associated with heart failure and new perspectives for developing strategies for heart failure-related bone loss.

Herbal Formula Modified Bu-Shen-Huo-Xue Decoction Attenuates Intervertebral Disc Degeneration via Regulating Inflammation and Oxidative Stress.

OBJECTIVE: This study aims to clarify the potential mechanism of modified Bu-Shen-Huo-Xue decoction (MBSHXD) in treating intervertebral disc degeneration (IDD) with methods of network pharmacology and molecular docking. METHODS: An MBSHXD and IDD-related common target gene set was established through TCMSP, UniProt, and two disease gene databases. GO and KEGG enrichment analysis and protein-protein interaction (PPI) networks were performed through the R platform and STRING to discover the potential mechanism. Molecular docking between the active ingredients and the core genes is used to calculate the binding energy. RESULTS: A total of 147 active ingredients and 79 common genes (including 10 core genes, TNF, VEGFA, IL6, MAPK3, AKT1, MAPK8, TP53, JUN, MMP9, and CXCL8) were identified. The results of GO and KEGG enrichment analysis showed that MBSHXD plays an essential role in regulating inflammation and oxidative stress. The meaningful pathways are the AGE-RAGE signaling pathway in diabetic complications, the IL-17 signaling pathway, the TNF signaling pathway, the PI3K-Akt signaling pathway, the MAPK signaling pathway, and apoptosis. In addition, the PPI network and molecular docking further demonstrated the roles that nine bioactive ingredients of MBSHXD play in IDD treatment through their interference with core target proteins. CONCLUSION: This study reveals that MBSHXD has the characteristics of a "multi-component, multi-target, and multi-pathway" in the treatment of IDD by regulating inflammation and oxidative stress, and network pharmacology may provide a feasible method to verify the molecular mechanism of MBSHXD for IDD by combining with molecular docking.

Proximity to major roads and the incidence of osteoporotic fractures in elderly women: The BONE study in Beijing.

Background: There is growing evidence to suggest that living near major roads (and suffering from the air pollution of urban streets) can have an adverse effect on bone health. However, little is known about its relationship to fractures caused by osteoporosis. Objective: This study was designed to investigate the relationship between residents living near major roads and the incidence of osteoporotic fractures. Methods: A retrospective cohort of 529 subjects was established based on community populations in older women aged 65-91. All participants lived in Beijing between September 27, 2007 and September 26, 2017. The distance between the residential sites of the subjects and the main roads was determined by the authors. Osteoporotic fracture diagnosis was based on medical histories and imaging examinations (DXA and X-rays). The Cox proportional hazard model was used to assess the association between traffic proximity and osteoporotic fractures, with suitable adjustments for individual and background factors. Results: The age range of all participants was 65-91 years, with an average age of 75.8 years (and a standard deviation 6.8 years). Of these, 19 (3.59%) suffered from diabetes, and 48 (9%) had hypertension; 85 (14%) families had annual incomes below US $30,000 and 402 (76%) had received a secondary school education or higher. Nearly 25% of people lived within 50 m of a main road, while 50% lived within 300 m. Between 2007 and 2017, a total of 96 osteoporotic fractures were observed. For people living <50 m from a main road, the adjusted hazard ratio (HR) for osteoporotic fractures was 2.509 (95% CI 1.345-4.680), while it was 1.830 (95% CI 1.029-3.255) for those living at a distance of 50-300 m from a main road vs. those living further than 300 m away. Conclusion: In this community-based cohort, living near a major road was associated with a higher incidence of osteoporotic fractures.

Reduction effect of oral pravastatin on the acute phase response to intravenous zoledronic acid: protocol for a real-world prospective, placebo-controlled trial.

INTRODUCTION: Zoledronic acid (ZA) has been used as a first-line treatment in patients with osteoporosis (OP) who receive an annual injection of 5 mg. However, side effects of bone pain and fever, known as the acute phase response (APR), have often been observed after clinical usage. A meta-analysis reported that the incidence of APR was 49.4% among patients with OP who received ZA for the first time and that 30% of patients with these adverse effects refused treatment in the following year. As a clinically used hypolipidaemic drug, statins can inhibit 3-hydroxy-3-methylglutaryl coenzyme A reductase to block the pathway upstream of farnesyl pyrophosphate synthase. This process can decrease the accumulation of isopentenyl pyrophosphate to prevent γδT-cell activation and inflammatory factor production, blocking APR occurrence. The aim of this study is to determine the reduction effect of oral pravastatin on APR and investigate the possible mechanisms underlying the effect in vivo. METHODS AND ANALYSIS: This will be a single-centre, placebo-controlled trial. Female participants will be allocated at a 1:1 ratio to receive either oral pravastatin or a placebo at 1-hour predose and 24 and 48 hours post-administration of ZA. The primary outcome will be the incidence of APR within 72 hours after ZA infusion. The secondary outcomes will include the occurrence time and severity of APR and the frequency and amount of acetaminophen usage within 72 hours after ZA infusion. This study will determine the preventive effect of oral pravastatin on APR in Chinese patients with OP, supporting the clinical application of ZA to alleviate concerns regarding safety and increase patient compliance. ETHICS AND DISSEMINATION: This study protocol has been registered with ClinicalTrials.gov. This study protocol was reviewed and approved by the Peking University Third Hospital Medical Science Research Ethics Committee. The results will be published in scientific peer-reviewed journals. TRIAL REGISTRATION NUMBER: NCT04719481.

Deciphering Obesity-Related Gene Clusters Unearths SOCS3 Immune Infiltrates and 5mC/m6A Modifiers in Ossification of Ligamentum Flavum Pathogenesis.

Background: Ossification of ligamentum flavum (OLF) is an insidious and debilitating heterotopic ossifying disease with etiological heterogeneity and undefined pathogenesis. Obese individuals predispose to OLF, whereas the underlying connections between obesity phenotype and OLF pathomechanism are not fully understood. Therefore, this study aims to explore distinct obesity-related genes and their functional signatures in OLF. Methods: The transcriptome sequencing data related to OLF were downloaded from the GSE106253 in the Gene Expression Omnibus (GEO) database. The obesity-related differentially expressed genes (ORDEGs) in OLF were screened, and functional and pathway enrichment analysis were applied for these genes. Furthermore, protein-protein interactions (PPI), module analysis, transcription factor enrichment analysis (TFEA), and experiment validation were used to identify hub ORDEGs. The immune infiltration landscape in OLF was depicted, and correlation analysis between core gene SOCS3 and OLF-related infiltrating immune cells (OIICs) as well as 5mC/m6A modifiers in OLF was constructed. Results: Ninety-nine ORDEGs were preliminarily identified, and functional annotations showed these genes were mainly involved in metabolism, inflammation, and immune-related biological functions and pathways. Integrative bioinformatic algorithms determined a crucial gene cluster associated with inflammatory/immune responses, such as TNF signaling pathway, JAK-STAT signaling pathway, and regulation of interferon-gamma-mediated signaling. Eight hub ORDEGs were validated, including 6 down-regulated genes (SOCS3, PPARG, ICAM-1, CCL2, MYC, and NT5E) and 2 up-regulated genes (PTGS2 and VEGFA). Furthermore, 14 differential OIICs were identified by ssGSEA and xCell, and SOCS3 was overlapped to be the core gene, which was associated with multiple immune infiltrates (dendritic cells, macrophage, and T cells) and six m6A modifiers as well as four 5mC regulators in OLF. Reduced SOCS3 and FTO expression and up-regulated DNMT1 level in OLF were validated by Western blotting. Conclusion: This study deciphered immune/inflammatory signatures of obesity-related gene clusters for the first time, and defined SOCS3 as one core gene. The crosstalk between 5mC/m6A methylation may be a key mediator of SOCS3 expression and immune infiltration. These findings will provide more insights into molecular mechanisms and therapeutic targets of obesity-related OLF.

CMTM3 suppresses bone formation and osteogenic differentiation of mesenchymal stem cells through inhibiting Erk1/2 and RUNX2 pathways.

Osteoporosis, fracture, large-scale craniofacial defects and osteonecrosis are hot topics and are still underdiagnosed and undertreated in the clinic. It is urgent to understand the molecular mechanisms corresponding to the regulation of bone formation. CMTM3 (CKLF-like MARVEL transmembrane domain containing 3) connects the classic chemokine to the transmembrane 4 superfamily and plays an important role in intracellular vesicles transport, EGF receptor function maintenance and cancer development. However, its expression and function in bone remain unclear. In this paper, we found that the bone volume/total volume, trabecular number, trabecular thickness and bone surface area/bone volume of Cmtm3 KO mice increased significantly, and trabecular separation and trabecular pattern factor decreased in Cmtm3 KO mice compared with WT mice by microcomputed tomography. Moreover, the bone mineral content, bone mineral density, ultimate force and stiffness were also increased in Cmtm3 KO mice. Using in vitro analysis, we showed that CMTM3 expression decreases during the differentiation of hBMSCs to osteoblasts. Knockdown of CMTM3 promoted ALP and mineralization of hBMSCs and facilitated osteoblastic differentiation with increasing RUNX2 expression. However, overexpression of CMTM3 got the opposite results. These results proved that CMTM3 was essential for osteogenic differentiation. In addition, knockdown of CMTM3 enhanced p-Erk1/2, but had no significant effect on p-Akt or p-STAT3 in hBMSCs and MC3T3-E1 cells. Taken together, our results indicated that Erk1/2 and RUNX2 pathways mediated by CMTM3 were involved in the process of osteogenic differentiation, and CMTM3 might be a new potential target in the treatment of bone formation-related disease.

Sirtuins: Potential Therapeutic Targets for Defense against Oxidative Stress in Spinal Cord Injury.

Spinal cord injury (SCI) is one of the most incapacitating neurological disorders. It involves complex pathological processes that include a primary injury and a secondary injury phase, or a delayed stage, which follows the primary injury and contributes to the aggravation of the SCI pathology. Oxidative stress, a key pathophysiological event after SCI, contributes to a cascade of inflammation, excitotoxicity, neuronal and glial apoptosis, and other processes during the secondary injury phase. In recent years, increasing evidence has demonstrated that sirtuins are protective toward the pathological process of SCI through a variety of antioxidant mechanisms. Notably, strategies that modulate the expression of sirtuins exert beneficial effects in cellular and animal models of SCI. Given the significance and novelty of sirtuins, we summarize the oxidative stress processes that occur in SCI and discuss the antioxidant effects of sirtuins in SCI. We also highlight the potential of targeting sirtuins for the treatment of SCI.