A novel approach in biomedical engineering: The use of polyvinyl alcohol hydrogel encapsulating human umbilical cord mesenchymal stem cell-derived exosomes for enhanced osteogenic differentiation and angiogenesis in bone regeneration.

Developing effective methods for alveolar bone defect regeneration is a significant challenge in orthopedics. Exosomes from human umbilical cord mesenchymal stem cells (HUMSC-Exos) have shown potential in bone repair but face limitations due to undefined application methods and mechanisms. To address this, HUMSC-Exos were encapsulated in polyvinyl alcohol (PVA) hydrogel (Exo@PVA) to create a novel material for alveolar bone repair. This combination enhanced the osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) and human umbilical vein endothelial cells (HUVECs) more effectively than Exos alone. Additionally, Exo@PVA significantly improved alveolar bone regeneration and defect repair in rats. The microRNA-21-5p (miR-21-5p) in Exo@PVA, identified through the GEO database and analyzed via in silico methods, played a crucial role. miR-21-5p promoted BMSC osteogenic differentiation by inhibiting WWP1-mediated KLF5 ubiquitination and enhanced HUVEC angiogenesis by targeting ATP2B4. These findings underscore the potential of an Exo-based approach with PVA hydrogel scaffolds for bone defect repair, operating through the miR-21-5p/WWP1/ATP2B4 signaling axis.

Accuracy of dynamic navigation compared to static surgical guides and the freehand approach in implant placement: a prospective clinical study.

BACKGROUND: Computer-guided implant surgery has improved the quality of implant treatment by facilitating the placement of implants in a more accurate manner. This study aimed to assess the accuracy of implant placement in a clinical setting using three techniques: dynamic navigation, static surgical guides, and freehand placement. We also investigated potential factors influencing accuracy to provide a comprehensive evaluation of each technique's advantages and disadvantages. MATERIALS AND METHODS: Ninety-four implants in 65 patients were included in this prospective study. Patients were randomly assigned to one of three groups: dynamic navigation, static surgical guides, or freehand placement. Implants were placed using a prosthetically oriented digital implant planning approach, and postoperative CBCT scans were superimposed on preoperative plans to measure accuracy. Seven deviation values were calculated, including angular, platform, and apical deviations. Demographic and consistency analyses were performed, along with one-way ANOVA and post-hoc tests for deviation values. RESULTS: The mean global platform, global apical, and angular deviations were 0.99 mm (SD 0.52), 1.14 mm (SD 0.56), and 3.66° (SD 1.64°) for the dynamic navigation group; 0.92 mm (SD 0.36), 1.06 mm (SD 0.47), and 2.52° (SD 1.18°) for the surgical guide group; and 1.36 mm (SD 0.62), 1.73 mm (SD 0.66), and 5.82° (SD 2.79°) for the freehand group. Both the dynamic navigation and surgical guide groups exhibited statistically significant differences in all values except depth deviations compared to the freehand group (p < 0.05), whereas only the angular deviation showed a significant difference between the dynamic navigation and surgical guide groups (p = 0.002). CONCLUSION: Our findings highlight the superior accuracy and consistency of dynamic navigation and static surgical guides compared to freehand placement in implant surgery. Dynamic navigation offers precision and flexibility. However, it comes with cost and convenience considerations. Future research should focus on improving its practicality. TRIAL REGISTRATION: This study was retrospectively registered at the Thai Clinical Trials Register-Medical Research Foundation of Thailand (MRF) with the TCTR identification number TCTR20230804001 on 04/08/2023. It was also conducted in accordance with the Declaration of Helsinki and approved by the institutional ethics committee at the Xian Jiaotong University Hospital of Stomatology, Xian, China (xjkqII[2021] No: 043). Written informed consent was obtained from all participants.

PF127 Hydrogel-Based Delivery of Exosomal CTNNB1 from Mesenchymal Stem Cells Induces Osteogenic Differentiation during the Repair of Alveolar Bone Defects.

Pluronic F127 (PF127) hydrogel has been highlighted as a promising biomaterial for bone regeneration, but the specific molecular mechanism remains largely unknown. Herein, we addressed this issue in a temperature-responsive PF127 hydrogel loaded with bone marrow mesenchymal stem cells (BMSCs)-derived exosomes (Exos) (PF127 hydrogel@BMSC-Exos) during alveolar bone regeneration. Genes enriched in BMSC-Exos and upregulated during the osteogenic differentiation of BMSCs and their downstream regulators were predicted by bioinformatics analyses. CTNNB1 was predicted to be the key gene of BMSC-Exos in the osteogenic differentiation of BMSCs, during which miR-146a-5p, IRAK1, and TRAF6 might be the downstream factors. Osteogenic differentiation was induced in BMSCs, in which ectopic expression of CTNNB1 was introduced and from which Exos were isolated. The CTNNB1-enriched PF127 hydrogel@BMSC-Exos were constructed and implanted into in vivo rat models of alveolar bone defects. In vitro experiment data showed that PF127 hydrogel@BMSC-Exos efficiently delivered CTNNB1 to BMSCs, which subsequently promoted the osteogenic differentiation of BMSCs, as evidenced by enhanced ALP staining intensity and activity, extracellular matrix mineralization (p < 0.05), and upregulated RUNX2 and OCN expression (p < 0.05). Functional experiments were conducted to examine the relationships among CTNNB1, microRNA (miR)-146a-5p, and IRAK1 and TRAF6. Mechanistically, CTNNB1 activated miR-146a-5p transcription to downregulate IRAK1 and TRAF6 (p < 0.05), which induced the osteogenic differentiation of BMSCs and facilitated alveolar bone regeneration in rats (increased new bone formation and elevated BV/TV ratio and BMD, all with p < 0.05). Collectively, CTNNB1-containing PF127 hydrogel@BMSC-Exos promote the osteogenic differentiation of BMSCs by regulating the miR-146a-5p/IRAK1/TRAF6 axis, thus inducing the repair of alveolar bone defects in rats.

A Pilot Investigation Generating an Alternative Therapeutic Strategy Combining Deproteinized Bovine Bone and Gelatin Sponge for a Novel Implant Material in Sinus Floor Elevation Operation.

BACKGROUND Implant placement in the posterior maxilla is typically complicated by a shortage of bone. Gelatin sponge could be combined with an appropriate material to enhance mechanical strength and maintain stability of an implant. This study aimed to evaluate the clinical application of bone grafting with bovine bone mixed with gelatin sponge. MATERIAL AND METHODS Fifty-four patients were divided into a control group (deproteinized bovine bone, n=26) and a test group (deproteinized bovine bone combined with gelatin sponge, n=28). Implants were placed in patients simultaneously after surgery (operation). Cone-beam computed tomography examination was carried out immediately and 6 months after surgery. Space with grafting materials was measured with Mimics software (version 16.0). RESULTS No remarkable differences were found for simultaneous placement, height of residual bone, delayed placement, width of residual bone, graft volume immediately after surgery (V1), graft volume 6 months after surgery (V2), or volumetric change rate between the test group and the control group (P>0.05). Graft volume V2 was remarkably decreased compared with V1 in the control and test groups (P=0.01). There were no significant differences for bone height immediately after surgery (H1) and bone height at 6 months after surgery (H2) between the 2 groups. Bone height H2 was markedly decreased compared with H1 (P<0.05). At 1 year after implantation, there was 1 implant loss in the control group and 2 in the test group. The implant survival rate in the control group was 97.62% and 95.24% in the test group. CONCLUSIONS Absorbable gelatin sponge combined with bovine bone particles was an effective and economical material for use in routine sinus floor elevation surgery.

Animal Models of Temporomandibular Disorder.

Temporomandibular disorders (TMD) are a group of diseases in the oral and maxillofacial region that can manifest as acute or chronic persistent pain, affecting millions of people worldwide. Although hundreds of studies have explored mechanisms and treatments underlying TMD, multiple pathogenic factors and diverse clinical manifestations make it still poorly managed. Appropriate animal models are helpful to study the pathogenesis of TMD and explore effective treatment measures. At present, due to the high cost of obtaining large animals, rodents and rabbits are often used to prepare TMD animal models. Over the past decade, various animal models have been intensively developed to understand neurobiological and molecular mechanisms of TMD, and seek effective treatments. Although these models cannot carry out all clinical features, they are valuable in revealing the mechanisms of TMD and creating curative access. Currently, there are multitudinous animal models of TMD research. They can be constructed in different means and summarized into four ways according to the various causes and symptoms, including chemical induction (intra-articular injection of ovalbumin, collagenase, formalin, vascular endothelial growth factor, intramuscular injection of complete Freund's adjuvant, etc.), mechanical stress stimulation (passive mouth opening, change of chewing load), surgical operation (partial disc resection, joint disc perforation) and psychological stress induction. Here, we summarize and discuss different approaches of animal models for determining neurophysiological and mechanical mechanisms of TMD and assess their advantages and limitations, respectively.

Public awareness and anxiety during COVID-19 epidemic in China: A cross-sectional study.

OBJECTIVE: The study aims to investigate public awareness of coronavirus disease 2019 (COVID-19) and measure levels of anxiety during the outbreak. METHOD: A total of 2115 subjects from 34 provinces in China were evaluated. A questionnaire was designed, which covers demographic characteristics, knowledge of COVID-19, and factors that influenced anxiety during the outbreak to test public awareness and determine the impact of the outbreak on people's lives. In addition, a generalized anxiety disorder (GAD) scale was utilized to assess anxiety levels during the outbreak. Lastly, the chi-square test and multiple logistic regression analysis were used to identify factors associated with levels of public anxiety. RESULTS: A majority of respondents reported high levels of awareness of COVID-19. A total of 1107 (52.3%), 707 (33.4%), 154 (7.3%), and 147 (7%) respondents exhibited no, mild, moderate, and severe levels of anxiety, respectively. Results of the chi-square test and multiple logistic regression analysis demonstrated that respondents (a) with no college education, (b) are unaware of neighbors who may have been infected, (c) who spent considerable time collecting information and browsing negative information related to the virus, (d) are unhealthy, and (e) displayed low levels of awareness of the transmission routes were highly likely to be anxious. CONCLUSION: During the outbreak, the majority of people exhibited high levels of awareness and knowledge regarding preventive measures from COVID-19. The absence of psychological anxiety was observed in more than half of the respondents. Adaptive responses to anxiety and high levels of awareness about COVID-19 may have protected the public during the outbreak.

miR­144­3p inhibits tumor cell growth and invasion in oral squamous cell carcinoma through the downregulation of the oncogenic gene, EZH2.

Accumulating evidence demonstrates that microRNAs (miRNAs or miRs) play important roles in the development and progression of human malignancies, including oral squamous cell carcinoma (OSCC); however, the unique roles of miRNAs are not yet fully understood in OSCC. The present study aimed to identify novel miRNAs associated with OSCC and to elucidate their functions. Based on a microarray analysis, miR­144­3p was found to be one of the most significantly downregulated miRNAs in OSCC tissues. Its low expression was closely associated with tumor size, differentiation and lymph node metastasis. Functionally, miR­144­3p overexpression suppressed proliferation, promoted apoptosis, and suppressed the invasion and migration of OSCC cells. In addition, enhancer of zeste homolog 2 (EZH2), a well­known oncogene, was proven to be a direct target of miR­144­3p, and its protein expression was negatively regulated by miR­144­3p. Moreover, EZH2 expression was increased, and inversely correlated with the miR­144­3p level in OSCC tissues. Notably, EZH2 knockdown inhibited cell proliferation, promoted cell apoptosis, and suppressed the invasion and migration of OSCC cells, whereas EZH2 overexpression partially reversed the anticancer effects mediated by miR­144­3p overexpression. On the whole, the findings of the present study suggest that miR­144­3p functions as a tumor suppressor by targeting the EZH2 oncogene, and may thus be considered as a potential diagnostic and therapeutic target for OSCC.

Acute Synovitis after Trauma Precedes and is Associated with Osteoarthritis Onset and Progression.

Osteoarthritis (OA) is a whole-joint disease characterized by cartilage destruction, subchondral bone sclerosis, osteophyte formation, and synovitis. However, it remains unclear which part of the joint undergoes initial pathological changes that drives OA onset and progression. In the present study, we investigated the longitudinal alterations of the entire knee joint using a surgically-induced OA mouse model. Histology analysis showed that synovitis occurred as early as 1 week after destabilization of the medial meniscus (DMM), which preceded the events of cartilage degradation, subchondral sclerosis and osteophyte formation. Importantly, key pro-inflammatory cytokines such as IL-1ß, IL-6, TNFα, and Ccl2, major matrix degrading enzymes including Adamts4, Mmp3 and Mmp13, as well as nerve growth factor (NGF), all increased significantly in both synovium and articular cartilage. It is notable that the inductions of these factors in synovium are far more extensive than those in articular cartilage. Results from behavioral tests demonstrated that sensitization of knee joint pain developed after 8 weeks, later than histological and molecular changes. In addition, the nanoindentation modulus of the medial tibiae decreased 4 weeks after DMM surgery, simultaneous with histological OA signs, which is also later than appearance of synovitis. Collectively, our data suggested that synovitis precedes and is associated with OA, and thus synovium may be an important target to intervene in OA treatment.

The microRNAs miR-204 and miR-211 maintain joint homeostasis and protect against osteoarthritis progression.

Osteoarthritis (OA) is a common, painful disease. Currently OA is incurable, and its etiology largely unknown, partly due to limited understanding of OA as a whole-joint disease. Here we report that two homologous microRNAs, miR-204 and miR-211, maintain joint homeostasis to suppress OA pathogenesis. Specific knockout of miR-204/-211 in mesenchymal progenitor cells (MPCs) results in Runx2 accumulation in multi-type joint cells, causing whole-joint degeneration. Specifically, miR-204/-211 loss-of-function induces matrix-degrading proteases in articular chondrocytes and synoviocytes, stimulating articular cartilage destruction. Moreover, miR-204/-211 ablation enhances NGF expression in a Runx2-dependent manner, and thus hyper-activates Akt signaling and MPC proliferation, underlying multiplex non-cartilaginous OA conditions including synovial hyperplasia, osteophyte outgrowth and subchondral sclerosis. Importantly, miR-204/-211-deficiency-induced OA is largely rescued by Runx2 insufficiency, confirming the miR-204/-211-Runx2 axis. Further, intraarticular administration of miR-204-expressing adeno-associated virus significantly decelerates OA progression. Collectively, miR-204/-211 are essential in maintaining healthy homeostasis of mesenchymal joint cells to counteract OA pathogenesis.

Deletion of Runx2 in condylar chondrocytes disrupts TMJ tissue homeostasis.

Runt-related transcription factor-2 (Runx2) is essential for chondrocyte maturation during cartilage development and embryonic mandibular condylar development. The process that chondrocytes, especially a subgroup of hypertrophic chondrocytes (HC), could transform into bone cells in mandibular condyle growth makes chondrocytes crucially important for normal endochondral bone formation. To determine whether Runx2 regulates postnatal condylar cartilage growth and tissue homeostasis, we deleted Runx2 in chondrocytes in postnatal mice and assessed the consequences on temporomandibular joint (TMJ) cartilage growth and remodeling. The cell lineage tracing data provide information demonstrating the role of chondrocytes in subchondral bone remodeling. The histologic and immunohistochemical data showed that Runx2 deficiency caused condylar tissue disorganization, including loss of HC and reduced hypertrophic zone, reduced proliferative chondrocytes, and decreased cartilage matrix production. Expression of Col10a1, Mmp13, Col2a1, Aggrecan, and Ihh was significantly reduced in Runx2 knockout mice. The findings of this study demonstrate that Runx2 is required for chondrocyte proliferation and hypertrophy in TMJ cartilage and postnatal TMJ cartilage growth and homeostasis, and that Runx2 may play an important role in regulation of chondrocyte-derived subchondral bone remodeling.

Runx2 is required for postnatal intervertebral disc tissue growth and development.

Runx2 plays an essential role in embryonic disc tissue development in mice. However, the role of runt-related transcription factor 2 (Runx2) in postnatal disc tissue growth and development has not been defined. In the present studies, we generated Runx2 conditional knockout (KO) mice (Runx2Agc1ER ), in which Runx2 was deleted in Aggrecan-expressing cells in disc tissue at postnatal 2-weeks of age. We then analyzed changes in disc tissue growth and development using histology and immunohistochemical methods in 3-month-old mice. We found that large vacuolated notochordal cells were accumulated in the nucleus pulposus (NP) in Runx2 KO mice. The growth plate cartilage tissue in the disc was thicker in Runx2 KO mice. We also found a significant upregulation of Indian hedgehog (Ihh) expression in the cells in NP cells and in annulus fibrosus cells of Runx2 KO mice. These results demonstrated that Runx2 may play an important role in postnatal disc tissue development through interacting with Ihh signaling.

Activation of ß-catenin signaling in aggrecan-expressing cells in temporomandibular joint causes osteoarthritis-like defects.

ß-Catenin plays a critical role in cartilage formation and development. To further understand the role of ß-catenin in osteoarthritis (OA) development in temporomandibular joint (TMJ), we have generated ß-catenin conditional activation mice (ß-cat(ex3) Agc1CreER ) by breeding Agc1-CreER mice with ß-cateninflox(ex3)/+ mice. Results of histologic analysis showed the progressive TMJ defects in 3- and 6-month-old ß-cat(ex3) Agc1CreER mice (tamoxifen induction was performed at 2 weeks of age), including decreased chondrocyte numbers in the superficial layer associated with less Alcian blue staining, increased numbers of hypertrophic chondrocytes in deep layers, and rough articular surface. Compared to the TMJ phenotype of ß-cat(ex3) Col2CreER mice, ß-cat(ex3) Agc1CreER mice showed much severe morphological defects in the superficial layer of TMJ. This may reflect that Agc1-CreER mice could efficiently target cells in the superficial layer of TMJ. Results of immunostaining showed significantly increased expression of MMP13, Col-X, Adamts4, and Adamts5 in TMJ of ß-cat(ex3) Agc1CreER mice. Results of proliferating cell nuclear antigen (PCNA), Ki67, and terminal deoxinucleotidyl transferase-mediated dUTP-fluorescein nick end labeling (TUNEL) staining further demonstrated that cell proliferation was decreased and cell apoptosis was increased in condylar cartilage of ß-cat(ex3) Agc1CreER mice. Our findings indicate that abnormal upregulation of ß-catenin in TMJ leads to defects assembling to OA-like phenotype, further demonstrating that ß-catenin plays a critical role in TMJ pathogenesis.

Deletion of Runx2 in Articular Chondrocytes Decelerates the Progression of DMM-Induced Osteoarthritis in Adult Mice.

Runx2 may play an important role in development of osteoarthritis (OA). However, the specific role of Runx2 in articular chondrocyte function and in OA development in adult mice has not been fully defined. In this study, we performed the destabilization of the medial meniscus (DMM) surgery at 12-week-old mice to induce OA in adult Runx2 Agc1CreER mice, in which Runx2 was specifically deleted in Aggrecan-expressing chondrocytes by administering tamoxifen at 8-weeks of age. Knee joint samples were collected 8- and 12-weeks post-surgery and analyzed through histology, histomorphometry and micro-computed tomography (µCT). Our results showed that severe OA-like defects were observed after DMM surgery in Cre-negative control mice, including articular cartilage degradation and subchondral sclerosis, while the defects were significantly ameliorated in Runx2 Agc1CreER KO mice. Immunohistochemical (IHC) results showed significantly reduced expression of MMP13 in Runx2 Agc1CreER KO mice compared to that in Cre-negative control mice. Results of quantitative reverse-transcription PCR (qRT-PCR) demonstrated that expression of the genes encoding for matrix degradation enzymes was significantly decreased in Runx2 Agc1CreER KO mice. Thus, our findings suggest that inhibition of Runx2 in chondrocytes could at least partially rescue DMM-induced OA-like defects in adult mice.

TNF Accelerates Death of Mandibular Condyle Chondrocytes in Rats with Biomechanical Stimulation-Induced Temporomandibular Joint Disease.

OBJECTIVE: To determine if temporomandibular joint chondrocyte apoptosis is induced in rats with dental biomechanical stimulation and what a role TNF takes. METHODS: Thirty-two rats were divided into 4 groups (n = 8/group) and exposed to incisor mal-occlusion induced by unilateral anterior crossbite biomechanical stimulation. Two groups were sampled at 2 or 4 weeks. The other two groups were treated with local injections of a TNF inhibitor or PBS into the temporomandibular joints area at 2 weeks and then sampled at 4 weeks. Twenty-four rats either served as unilateral anterior crossbite mock operation controls (n = 8/group) with sampling at 2 or 4 weeks or received a local injection of the TNF inhibitor at 2 weeks with sampling at 4 weeks. Chondrocytes were isolated from the temporomandibular joints of 6 additional rats and treated with TNF in vitro. Joint samples were assessed using Hematoxylin&eosin, Safranin O, TUNEL and immunohistochemistry staining, real-time PCR, fluorogenic activity assays and Western blot analyses. The isolated chondrocytes were also analyzed by flow cytometry. RESULTS: Unilateral anterior crossbite stimulation led to temporomandibular joint cartilage degradation, associated with an increase in TUNEL-positive chondrocytes number, caspase-9 expression levels, and the release of cytochrome c from mitochondria at 2 weeks without changes in TNF and caspase-8 levels until after 4 weeks. TNF stimulated apoptosis of the isolated chondrocytes and up-regulated caspase-8 expression, but did not change caspase-9 expression levels. Local injection of TNF inhibitor down-regulated caspase-8 expression and reduced TUNEL-positive cell number, but did not reverse cartilage thickness reduction, caspase-9 up-regulation or cytochrome c release. CONCLUSIONS: Unilateral anterior crossbite stimulation induces mitochondrion-mediated apoptosis of articular chondrocytes. TNF accelerated the unilateral anterior crossbite induced chondrocytes apoptosis via death-receptor pathway. However, anti-TNF therapy does not prevent cartilage loss in this model of temporomandibular joint.