[Research Progress in the Regulatory Role of circRNA-miRNA Network in Bone Remodeling]. / circRNA-miRNAç½‘ç»œè°ƒæŽ§éª¨é‡å¡‘çš„ç ”ç©¶è¿›å±•.

The dynamic balance between bone formation and bone resorption is a critical process of bone remodeling. The imbalance of bone formation and bone resorption is closely associated with the occurrence and development of various bone-related diseases. Under both physiological and pathological conditions, non-coding RNAs (ncRNAs) play a crucial regulatory role in protein expression through either inhibiting mRNAs translation or promoting mRNAs degradation. Circular RNAs (circRNAs) are a type of non-linear ncRNAs that can resist the degradation of RNA exonucleases. There is accumulating evidence suggesting that circRNAs and microRNAs (miRNAs) serve as critical regulators of bone remodeling through their direct or indirect regulation of the expression of osteogenesis-related genes. Additionally, recent studies have revealed the involvement of the circRNAs-miRNAs regulatory network in the process by which mesenchymal stem cells (MSCs) differentiate towards the osteoblasts (OB) lineage and the process by which bone marrow-derived macrophages (BMDM) differentiate towards osteoclasts (OC). The circRNA-miRNA network plays an important regulatory role in the osteoblastic-osteoclastic balance of bone remodeling. Therefore, a thorough understanding of the circRNA-miRNA regulatory mechanisms will contribute to a better understanding of the regulatory mechanisms of the balance between osteoblastic and osteoclastic activities in the process of bone remodeling and the diagnosis and treatment of related diseases. Herein, we reviewed the functions of circRNA and microRNA. We also reviewed their roles in and the mechanisms of the circRNA-miRNA regulatory network in the process of bone remodeling. This review provides references and ideas for further research on the regulation of bone remodeling and the prevention and treatment of bone-related diseases.

An Additional Lrp4 High Bone Mass Mutation Mitigates the Sost-Knockout Phenotype in Mice by Increasing Bone Remodeling.

Pathogenic variants disrupting the binding between sclerostin (encoded by SOST) and its receptor LRP4 have previously been described to cause sclerosteosis, a rare high bone mass disorder. The sclerostin-LRP4 complex inhibits canonical WNT signaling, a key pathway regulating osteoblastic bone formation and a promising therapeutic target for common bone disorders, such as osteoporosis. In the current study, we crossed mice deficient for Sost (Sost-/-) with our p.Arg1170Gln Lrp4 knock-in (Lrp4KI/KI) mouse model to create double mutant Sost-/-;Lrp4KI/KI mice. We compared the phenotype of Sost-/- mice with that of Sost-/-;Lrp4KI/KI mice, to investigate a possible synergistic effect of the disease-causing p.Arg1170Trp variant in Lrp4 on Sost deficiency. Interestingly, presence of Lrp4KI alleles partially mitigated the Sost-/- phenotype. Cellular and dynamic histomorphometry did not reveal mechanistic insights into the observed phenotypic differences. We therefore determined the molecular effect of the Lrp4KI allele by performing bulk RNA sequencing on Lrp4KI/KI primary osteoblasts. Unexpectedly, mostly genes related to bone resorption or remodeling (Acp5, Rankl, Mmp9) were upregulated in Lrp4KI/KI primary osteoblasts. Verification of these markers in Lrp4KI/KI, Sost-/- and Sost-/-;Lrp4KI/KI mice revealed that sclerostin deficiency counteracts this Lrp4KI/KI effect in Sost-/-;Lrp4KI/KI mice. We therefore hypothesize that models with two inactivating Lrp4KI alleles rather activate bone remodeling, with a net gain in bone mass, whereas sclerostin deficiency has more robust anabolic effects on bone formation. Moreover, these effects of sclerostin and Lrp4 are stronger in female mice, contributing to a more severe phenotype than in males and more detectable phenotypic differences among different genotypes.

Association of Bone Mineral Density and Bone Turnover Markers with the Risk of Diabetes: Hong Kong Osteoporosis Study and Mendelian Randomization.

Preclinical studies demonstrated that bone plays a central role in energy metabolism. However, how bone metabolism is related to the risk of diabetes in humans is unknown. We investigated the association of bone health (bone mineral density [BMD] and bone turnover markers) with incident type-2 diabetes mellitus (T2DM) based on the Hong Kong Osteoporosis Study (HKOS). A total of 993 and 7160 participants from the HKOS were studied for the cross-sectional and prospective analyses, respectively. The cross-sectional study evaluated the association of BMD and bone biomarkers with fasting glucose and glycated hemoglobin (HbA1c ) levels, whereas the prospective study examined the associations between BMD at study sites and the risk of T2DM by following subjects a median of 16.8 years. Body mass index (BMI) was adjusted in all full models. Mendelian randomization (MR) was conducted for causal inference. In the cross-sectional analysis, lower levels of circulating bone turnover markers and higher BMD were significantly associated with increased fasting glucose and HbA1c levels. In the prospective analysis, higher BMD (0.1 g/cm2 ) at the femoral neck and total hip was associated with increased risk of T2DM with hazard ratios (HRs) of 1.10 (95% confidence interval [CI], 1.03 to 1.18) and 1.14 (95% CI, 1.08 to 1.21), respectively. The presence of osteoporosis was associated with a 30% reduction in risk of T2DM compared to those with normal BMD (HR = 0.70; 95% CI, 0.55 to 0.90). The MR results indicate a robust genetic causal association of estimated BMD (eBMD) with 2-h glucose level after an oral glucose challenge test (estimate = 0.043; 95% CI, 0.007 to 0.079) and T2DM (odds ratio = 1.064; 95% CI, 1.036 to 1.093). Higher BMD and lower levels of circulating bone biomarkers were cross-sectionally associated with poor glycemic control. Moreover, higher BMD was associated with a higher risk of incident T2DM and the association is probably causal. © 2023 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).

Association of PFN1 Gene Polymorphisms with Bone Mineral Density, Bone Turnover Markers, and Osteoporotic Fractures in Chinese Population.

Recent studies have discovered an association between the PFN1 gene and Paget's disease. However, it is currently unknown whether the PFN1 gene is related to osteoporosis. This study was performed to investigate the association of Single-Nucleotide Polymorphisms (SNPs) in the PFN1 gene with Bone Mineral Density (BMD) as well as bone turnover markers and osteoporotic fractures in Chinese subjects. A total of 2836 unrelated Chinese subjects comprising 1247 healthy subjects and 1589 osteoporotic fractures patients (Fracture group) were enrolled in this study. Seven tagSNPs (rs117337116, rs238243, rs6559, rs238242, rs78224458, rs4790714, and rs13204) of the PFN1 gene were genotyped. The BMD of the lumbar spine 1-4 (L1-4), femoral neck, and total hip as well as bone turnover markers, such as ß-C-Terminal telopeptide of type 1 collagen (ß-CTX) and Procollagen type 1 N-terminal Propeptide (P1NP), were measured. The association between 7 tagSNPs and BMD and bone turnover markers was analyzed in 1247 healthy subjects only. After age matching, we selected 1589 osteoporotic fracture patients (Fracture group) and 756 nonfracture controls (Control group, selected from 1247 healthy subjects) for a case-control study, respectively. For the case-control study, we used logistic regression to investigate the relationship between 7 tagSNPs and osteoporotic fractures risk. In the All group, the PFN1 haplotype GAT was associated with the ß-CTX (P = 0.007). In the Female group, the PFN1 haplotype GAT was associated with the ß-CTX (P = 0.005). In the Male group, the rs13204, the rs78224458, and the PFN1 haplotype GAC were associated with the BMD of the L1-4 (all P = 0.012); the rs13204, the rs78224458, and the PFN1 haplotype GAC were associated with the BMD of the femoral neck (all P = 0.012); the rs13204 and rs78224458 were associated with the BMD of the total hip (both P = 0.015); and the PFN1 haplotype GAT was associated with the ß-CTX (P = 0.013). In the subsequent case-control study, the rs13204 and rs78224458 in the male group were associated with the risk of L1-4 fracture (P = 0.016 and 0.010, respectively) and total hip fracture (P = 0.013 and 0.016, respectively). Our study reveals that PFN1 gene polymorphisms are associated with BMD in Chinese males and ß-CTX in Chinese people and confirmed the relationship between PFN1 gene polymorphisms and Chinese male osteoporotic fractures in a case-control study.

Dynamic changes in transcriptome during orthodontic tooth movement.

OBJECTIVES: The objective of this study was to determine global changes in gene expression with next generation sequencing (NGS) in order to assess the biological effects of orthodontic tooth movement (OTM) on alveolar bone in a rat model. MATERIALS AND METHODS: Thirty-five Wistar rats (age 14 weeks) were used in the study. The OTM was performed using closed coil Nickel-Titanium spring to apply a mesial force on maxillary first molars of 8-10 g. Three hours, 1, 3, 7 and 14 days after the placement of the appliance, rats were killed at each time point respectively. The alveolar bone, around left maxillary first molar, were excised on compression side. The samples were immediately frozen in liquid nitrogen for subsequent RNA extraction. Total RNA samples were prepared for mRNA sequencing using the Illumina kit. RNA-Seq reads were aligned to the rat genomes using the STAR Aligner and bioinformatic analysis was performed. RESULTS: A total of 18 192 genes were determined. Day 1 has the highest number of differentially expressed genes (DEGs) observed with more upregulated than downregulated genes. A total of 2719 DEGs were identified to use as input for the algorithm. Six distinct clusters of temporal patterns were observed representing proteins that were differentially regulated indicating different expression kinetics. Principal component analysis (PCA) showed distinct clustering by time points and days 3, 7 and 14 share similar gene expression pattern. CONCLUSIONS: Distinct gene expression pattern was observed at different time points studied. Hypoxia, inflammation and bone remodelling pathways are major mechanisms behind OTM.

HERC1 deficiency causes osteopenia through transcriptional program dysregulation during bone remodeling.

Bone remodeling is a continuous process between bone-forming osteoblasts and bone-resorbing osteoclasts, with any imbalance resulting in metabolic bone disease, including osteopenia. The HERC1 gene encodes an E3 ubiquitin ligase that affects cellular processes by regulating the ubiquitination of target proteins, such as C-RAF. Of interest, an association exists between biallelic pathogenic sequence variants in the HERC1 gene and the neurodevelopmental disorder MDFPMR syndrome (macrocephaly, dysmorphic facies, and psychomotor retardation). Most pathogenic variants cause loss of HERC1 function, and the affected individuals present with features related to altered bone homeostasis. Herc1-knockout mice offer an excellent model in which to study the role of HERC1 in bone remodeling and to understand its role in disease. In this study, we show that HERC1 regulates osteoblastogenesis and osteoclastogenesis, proving that its depletion increases gene expression of osteoblastic makers during the osteogenic differentiation of mesenchymal stem cells. During this process, HERC1 deficiency increases the levels of C-RAF and of phosphorylated ERK and p38. The Herc1-knockout adult mice developed imbalanced bone homeostasis that presented as osteopenia in both sexes of the adult mice. By contrast, only young female knockout mice had osteopenia and increased number of osteoclasts, with the changes associated with reductions in testosterone and dihydrotestosterone levels. Finally, osteocytes isolated from knockout mice showed a higher expression of osteocytic genes and an increase in the Rankl/Opg ratio, indicating a relevant cell-autonomous role of HERC1 when regulating the transcriptional program of bone formation. Overall, these findings present HERC1 as a modulator of bone homeostasis and highlight potential therapeutic targets for individuals affected by pathological HERC1 variants.

Cellular voids in the pathogenesis of otosclerosis.

BACKGROUND: Otosclerosis is a common ear disease that causes fixation of the stapes and conductive hearing impairment. However, the pathogenesis of otosclerosis is still unknown. Otosclerosis could be associated with the unique bony environment found in the otic capsule. Normal bone remodelling is almost completely absent around the inner ear after birth allowing degenerative changes and dead osteocytes to accumulate. High levels of inner ear anti resorptive osteoprotegerin (OPG) is most likely responsible for this capsular configuration. Studies have demonstrated how osteocyte lifespan variation creates occasional clusters of dead osteocytes, so-called cellular voids, at otosclerotic predilection sites in the human otic capsule. These cellular voids have been suggested as possible starting points of otosclerosis. AIM: To describe the cellular viability in otosclerotic lesions and compare it to that of cellular voids. MATERIALS AND METHODS: The study was based on unbiased stereological quantifications in undecalcified human temporal bones with otosclerosis. RESULTS: Osteocyte viability was found to vary within the otosclerotic lesions. Furthermore, the results presented here illustrate that inactive otosclerotic lesions consist of mainly dead interstitial bone, much like cellular voids. CONCLUSIONS AND SIGNIFICANCE: Focal degeneration in the otic capsule may play an important role in the pathogenesis of otosclerosis.

The effects of epigenetic modifications on bone remodeling in age-related osteoporosis.

PURPOSE: As the population ages, there is an increased risk of fracture and morbidity diseases associated with aging, such as age-related osteoporosis and other bone diseases linked to aging skeletons. RESULTS: Several bone-related cells, including multipotent bone mesenchymal stem cells, osteoblasts that form bone tissue, and osteoclasts that break it down, are in symbiotic relationships throughout life. Growing evidence indicates that epigenetic modifications of cells caused by aging contribute to compromised bone remodeling and lead to osteoporosis. A number of epigenetic mechanisms are at play, including DNA/RNA modifications, histone modifications, microRNAs (miRNAs), and long noncoding RNAs (lncRNAs), as well as chromatin remodeling. CONCLUSION: In this review, we summarized the epigenetic modifications of different bone-related cells during the development and progression of osteoporosis associated with aging. Additionally, we described a compensatory recovery mechanism under epigenetic regulation that may lead to new strategies for regulating bone remodeling in age-related osteoporosis.

Altering osteoclast numbers using CTSK models in utero affects mice offspring craniofacial morphology.

BACKGROUND: Bone remodelling during development and growth is important for craniofacial integrity of offspring. The aim of this study was to investigate the changes in offspring adult skull morphology when the osteoclasts number was altered in utero, using three-dimensional (3D) geometric morphometric analysis (GMA). MATERIALS AND METHODS: We altered osteoclasts number in utero via two approaches. First, we generated heterozygous CtskCre ;DTAfl/+ (diphtheria toxin A) mice. Second, we altered Ctsk expression in vivo by injecting pregnant wild-type dams at embryonic day (E) 12.5 with in vivo siRNA specific for Ctsk. Mice were collected at 6 weeks and analysed using geometric morphometric analysis via computed tomography, histomorphometry and gene expression analysis. RESULTS: Altering osteoclasts number in utero affected the offspring adult skull morphology. Decreased Ctsk and osteoclast numbers were associated with a decrease in cranial vault height and an increase in mandibular body length. Changes in size and shape were observed with an increased number of osteoclasts in CtskCre ;DTAfl/+ mice, including an increase in cranial vault height, as well as a shortening of mandibular body length and ramus height. CONCLUSION: The findings of this study suggest that modulation of osteoclast numbers during pre- and post-natal development may be a previously unknown factor in the aetiology of skeletal malocclusions. An improved understanding of the factors affecting bone homeostasis during development and growth may help in the development of future therapies that would target the early intervention of skeletal malocclusion.

Histochemical assessment of accelerated bone remodeling and reduced mineralization in Il-6 deficient mice.

OBJECTIVES: Interleukin-6 (IL-6) contributes to the regulation of functions in various tissues and organs. Even though IL-6 has been reported to modulate bone metabolism in previous studies, this finding is controversial. This study aims to evaluate the possible involvement of IL-6 in bone metabolism by examining the histological activity of osteoblasts and osteoclasts in the femora of Il-6 deficient (Il-6-/-) mice. METHODS: Eight-week-old male Il-6-/- mice and their wild-type littermates were fixed with a paraformaldehyde solution, and their femora were extracted for micro-CT analysis, immunohistochemistry, and real-time PCR analysis. RESULTS: Il-6-/- femora showed an increased bone volume/tissue volume (TV) but a reduced bone mineral density compared with the wild-type. Furthermore, the tissue-nonspecific alkaline phosphatase positive area/TV ratio, the expression of Runx2, Osterix, and Rankl, and the number of tartrate-resistant acid phosphatase-positive osteoclasts were all increased in the Il-6-/- mice. A considerable number of unmineralized areas within the bone matrix and abundant sclerostin-reactive osteocytes were observed in Il-6-/- femoral metaphyses but not in the wild-type. Interestingly, the gene expression of Cd206 was elevated in Il-6-/- femora, and many F4/80-positive macrophages/monocytes and CD206-immunoreactive macrophages in the primary trabeculae had migrated closer to the growth plate, where intense RANKL immunoreactivity was detected. These results suggest that, in an IL-6-deficient state, CD206-positive macrophages may differentiate into osteoclasts when in contact with RANKL-reactive osteoblastic cells. CONCLUSION: In a state of IL-6 deficiency, the population and cell activities of osteoblast, osteoclasts, and macrophages seemed to be facilitated, except for the reduced mineralization in bone.

Aberrant paracrine signalling for bone remodelling underlies the mutant histone-driven giant cell tumour of bone.

Oncohistones represent compelling evidence for a causative role of epigenetic perturbations in cancer. Giant cell tumours of bone (GCTs) are characterised by a mutated histone H3.3 as the sole genetic driver present in bone-forming osteoprogenitor cells but absent from abnormally large bone-resorbing osteoclasts which represent the hallmark of these neoplasms. While these striking features imply a pathogenic interaction between mesenchymal and myelomonocytic lineages during GCT development, the underlying mechanisms remain unknown. We show that the changes in the transcriptome and epigenome in the mesenchymal cells caused by the H3.3-G34W mutation contribute to increase osteoclast recruitment in part via reduced expression of the TGFß-like soluble factor, SCUBE3. Transcriptional changes in SCUBE3 are associated with altered histone marks and H3.3G34W enrichment at its enhancer regions. In turn, osteoclasts secrete unregulated amounts of SEMA4D which enhances proliferation of mutated osteoprogenitors arresting their maturation. These findings provide a mechanism by which GCTs undergo differentiation in response to denosumab, a drug that depletes the tumour of osteoclasts. In contrast, hTERT alterations, commonly found in malignant GCT, result in the histone-mutated neoplastic cells being independent of osteoclasts for their proliferation, predicting unresponsiveness to denosumab. We provide a mechanism for the initiation of GCT, the basis of which is dysfunctional cross-talk between bone-forming and bone-resorbing cells. The findings highlight the role of tumour/microenvironment bidirectional interactions in tumorigenesis and how this is exploited in the treatment of GCT.

BMSC-derived exosomal miR-27a-3p and miR-196b-5p regulate bone remodeling in ovariectomized rats.

Background: In the bone marrow microenvironment of postmenopausal osteoporosis (PMOP), bone marrow mesenchymal stem cell (BMSC)-derived exosomal miRNAs play an important role in bone formation and bone resorption, although the pathogenesis has yet to be clarified. Methods: BMSC-derived exosomes from ovariectomized rats (OVX-Exo) and sham-operated rats (Sham-Exo) were co-cultured with bone marrow-derived macrophages to study their effects on osteoclast differentiation. Next-generation sequencing was utilized to identify the differentially expressed miRNAs (DE-miRNAs) between OVX-Exo and Sham-Exo, while target genes were analyzed using bioinformatics. The regulatory effects of miR-27a-3p and miR-196b-5p on osteogenic differentiation of BMSCs and osteoclast differentiation were verified by gain-of-function and loss-of-function analyses. Results: Osteoclast differentiation was significantly enhanced in the OVX-Exo treatment group compared to the Sham-Exo group. Twenty DE-miRNAs were identified between OVX-Exo and Sham-Exo, among which miR-27a-3p and miR-196b-5p promoted the expressions of osteogenic differentiation markers in BMSCs. In contrast, knockdown of miR-27a-3p and miR-196b-5p increased the expressions of osteoclastic markers in osteoclast. These 20 DE-miRNAs were found to target 11435 mRNAs. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses revealed that these target genes were involved in several biological processes and osteoporosis-related signaling pathways. Conclusion: BMSC-derived exosomal miR-27a-3p and miR-196b-5p may play a positive regulatory role in bone remodeling.

Runx2 and Nell-1 in dental follicle progenitor cells regulate bone remodeling and tooth eruption.

Dental follicles are necessary for tooth eruption, surround the enamel organ and dental papilla, and regulate both the formation and resorption of alveolar bone. Dental follicle progenitor cells (DFPCs), which are stem cells found in dental follicles, differentiate into different kinds of cells that are necessary for tooth formation and eruption. Runt-related transcription factor 2 (Runx2) is a transcription factor that is essential for osteoblasts and osteoclasts differentiation, as well as bone remodeling. Mutation of Runx2 causing cleidocranial dysplasia negatively affects osteogenesis and the osteoclastic ability of dental follicles, resulting in tooth eruption difficulties. Among a variety of cells and molecules, Nel-like molecule type 1 (Nell-1) plays an important role in neural crest-derived tissues and is strongly expressed in dental follicles. Nell-1 was originally identified in pathologically fused and fusing sutures of patients with unilateral coronal synostosis, and it plays indispensable roles in bone remodeling, including roles in osteoblast differentiation, bone formation and regeneration, craniofacial skeleton development, and the differentiation of many kinds of stem cells. Runx2 was proven to directly target the Nell-1 gene and regulate its expression. These studies suggested that Runx2/Nell-1 axis may play an important role in the process of tooth eruption by affecting DFPCs. Studies on short and long regulatory noncoding RNAs have revealed the complexity of RNA-mediated regulation of gene expression at the posttranscriptional level. This ceRNA network participates in the regulation of Runx2 and Nell-1 gene expression in a complex way. However, non-study indicated the potential connection between Runx2 and Nell-1, and further researches are still needed.

The effect of osteoporotic and non-osteoporotic individuals' T cell-derived exosomes on osteoblast cells' bone remodeling related genes expression and alkaline phosphatase activity.

OBJECTIVES: Osteoporosis is a common skeletal disorder attributed to age and is defined as a systematic degradation of bone mass and the microarchitecture leading to bone fractures. Exosomes have been reported in almost all biological fluids and during the failure of bone remodeling. 20 ml of blood samples were obtained from osteoporotic and non-osteoporotic postmenopausal women. After the isolation of peripheral blood mononuclear cells (PBMCs), T cells were separated via the magnetic-activated cell sorting (MACS) technique. Exosomes were driven from T cells of non-osteoporotic and osteoporotic volunteers. Subsequently, normal osteoblasts were treated with obtained T cell exosomes to assess osteoblastic function and gene expression. RESULTS: Runx2, type I collagen, osteopontin, and osteocalcin expression decreased in osteoblasts treated by osteoporotic T cell exosomes. In contrast, an increased expression of the mentioned genes was observed following non-osteoporotic T cell exosome treatment. Additionally, osteoblast alkaline phosphatase (ALP) activity treated with non-osteoporotic T cell exosomes increased. However, this activity decreased in another group. Our data demonstrated that T cell exosomes obtained from osteoporotic and non-osteoporotic individuals could alter the osteoblastic function and gene expression by affecting the genes essential for bone remodeling.

Ablation of the miRNA cluster 24 in cartilage and osteoblasts impairs bone remodeling.

MicroRNAs (miRNAs) post-transcriptionally regulate cartilage and bone development and function, however, only few miRNAs have been described to play a role for cartilage to bone transition in vivo. Previously, we showed that cartilage-specific deletion of the Mirc24 cluster in newborn male mice leads to impaired growth plate cartilage development due to increased RAF/MEK/ERK signaling and affects the stability of the cartilage extracellular matrix on account of decreased SOX6 and SOX9 and increased MMP13 levels. Here, we studied how Mirc24 cluster inactivation in cartilage and osteoblasts leads to an increased bone density associated with defects in collagen remodeling in trabecular bone. No changes in osteoblast distribution were observed, whereas the number of osteoclasts was reduced and TRAP activity in osteoclasts decreased. Surprisingly, an increased level of cluster-encoded miR-322 or miR-503 raises Rankl gene expression and inactivation of the cluster in chondrocytes reduces Rankl expression. These results suggest that the Mirc24 cluster regulates Rankl expression in chondrocytes at the chondro-osseous border, where the cluster is mainly expressed to modulate osteoclast formation, bone remodeling and bone integrity.

STAT1 and CXCL10 involve in M1 macrophage polarization that may affect osteolysis and bone remodeling in extrapulmonary tuberculosis.

OBJECTIVE: This study was aimed to reveal the molecular mechanism of bone destruction due to macrophage polarization leading to during extrapulmonary tuberculosis (EPTB) infection. METHODS: The dataset GSE83456 was downloaded from the GEO database, and the xCell tool was used to obtain the 64 types of immune cells. The flow cytometry was performed to identified the differences between M1 and M2 macrophages between EPTB and the healthy controls (HCs). The enrichment analyses were performed on the differentially expressed genes (DEGs) and their functionally related modules. The hub genes were screened out, and their relationships with EPTB and the immune cell subtypes were further analyzed. RESULTS: The flow cytometric analysis validated this hypothesis of M1-macrophage polarization correlated with the pathogenesis of EPTB. Of the obtained 103 DEGs, 97 genes were upregulated, and 6 genes were downregulated. The GO and KEGG pathway analyses showed that the DEGs were particularly involved in the immune-related processes. The hub genes (STAT1 and CXCL10) might be involved in M1-macrophage polarization and correlated with the pathogenesis of EPTB. STAT1 and CXCL10 could also behave as biomarkers for EPTB. CONCLUSION: STAT1 and CXCL10 were involved in the M1-macrophage polarization and correlated with the pathogenesis of EPTB. Besides, both of them could also behave as biomarkers for EPTB diagnosis and provide the required clues for targeted therapy in the future.

Carfilzomib alleviated osteoporosis by targeting PSME1/2 to activate Wnt/ß-catenin signaling.

Osteoporosis (OP) is characterized by decreased bone mineral density and impaired bone strength. Carfilzomib (CFZ) is a new-generation proteasome inhibitor and has been found to affect bone metabolism. However, the effect and mechanism of CFZ on OP has not been investigated systematically. In this study, we found that protein levels of proteasome activator subunit 1/2 (PSME1/2) increased in OP, and accumulated mostly in osteoblasts and osteoclasts. Treatment with PSME1/2 recombinant protein inhibited osteogenesis and promoted osteoclast formation in vitro. Also, PSME1/2 inhibited the expression of ß-catenin protein, resulting in limitation of Wnt/ß-catenin signaling. CFZ inhibited PSME1 and PSME2 proteasome activities and increased ß-catenin protein level, resulting in the translocation of ß-catenin to the nucleus and activation of canonical Wnt/ß-catenin signaling, further promoting osteogenesis and inhibiting osteoclastic differentiation. In vivo, we conducted ovariectomy (OVX) to create a model of OVX-induced postmenopausal OP in mice. When analyzed by micro-CT scanning, enhancement of bone mineral density, bone volume, trabecular number, and thickness was seen in the CFZ-treated mice. Also, we noticed increased osteogenesis and decreased osteoclastogenesis, diminished expression of PSME1 and PSME2 and activated Wnt/ß-catenin signaling in bone sections from OP mice treated with CFZ. Overall, our data indicated that PSME1/2 may serve as new targets for the treatment of OP, and targeting PSME1/2 with CFZ provides a candidate therapeutic molecule for postmenopausal OP.

On-Bone Fixation of Free Gingival Graft Induces an Osteoinductive Effect in Human Alveolar Bone.

We examined alveolar bone samples in the area of on-bone fixation of a free gingival graft performed during surgery in patients aged 37-55 years with a diagnosis of secondary partial adentia of the upper and lower jaws. Six months after fixation of the graft in the alveolar bone, foci of neoosteogenesis were found in the contact zone. They were characterized by the appearance of appositional lines, cords of basophilic osteoblasts, and growing osteons. An immunohistochemical study revealed an increase in the number of CD44+, CD29+, and osteocalcin+ cells in the layer of the outer circumferential lamellae, primary osteons, and the lining of the Haversian canals. TGF-ß1+ cells were located in the intertrabecular reticular tissue and wall of microvessels. The results indicate activation of mesenchymal stem cells in the area of localization of the graft and differentiating osteoblasts. The observed osteoinductive effect of free gingival graft is associated with its participation in reorganization in MSC and induction of morphogenetic molecules.

X-Linked Osteogenesis Imperfecta Possibly Caused by a Novel Variant in PLS3.

Osteogenesis imperfecta (OI) represents a complex spectrum of genetic bone diseases that occur primarily due to mutations and deletions of the COL1A1 and COL1A2 genes. Recent molecular studies of the network of signaling pathways have contributed to a better understanding of bone remodeling and the pathogenesis of OI caused by mutations in many other genes associated with normal bone mineralization. In this paper, a case of a rare X-linked variant of OI with a change in the gene encoding plastin 3-a protein important for the regulation of the actin cytoskeleton, is presented. A 16-year-old patient developed ten bone fractures caused by minor trauma or injury, including a compression fracture of the second lumbar vertebra during his lifetime. Next-generation sequencing analysis did not show pathologically relevant deviations in the COL1A1 and COL1A2 genes. Targeted gene analyses (Skeletal disorder panel) of the patient, his father, mother and sister were then performed, detecting variants of uncertain significance (VUS) for genes PLS3, FN1 and COL11A2. A variant in the PLS3 gene were identified in the patient, his mother and sister. Since the PLS3 gene is located on the X chromosome, the mother and sister showed no signs of the disease. Although the variant in the PLS3 gene (c.685G>A (p.Gly229Arg)) has not yet been described in the literature, nor is its pathogenicity known, clinical findings combined with genetic testing showed that this variant may explain the cause of X-linked OI in our patient. This rare case of the PLS3 variant of X-linked OI might point to a novel target for personalized therapy in patients with this severe disease.