TGF-ß2 enhances glycolysis in chondrocytes via TßRI/p-Smad3 signaling pathway.

Chondrocytes rely heavily on glycolysis to maintain the metabolic homeostasis and cartilage matrix turnover. Glycolysis in chondrocytes is remodeled by diverse biochemical and biomechanical factors due to the sporty joint microenvironment. Transforming growth factor-ß2 (TGF-ß2), one of the most abundant TGF-ß superfamily members in chondrocytes, has increasingly attracted attention in cartilage physiology and pathology. Although previous studies have emphasized the importance of TGF-ß superfamily members on cell metabolism, whether and how TGF-ß2 modulates glycolysis in chondrocytes remains elusive. In the current study, we investigated the effects of TGF-ß2 on glycolysis in chondrocytes and explored the underlying biomechanisms. The results showed that TGF-ß2 could enhance glycolysis in chondrocytes by increasing glucose consumption, up-regulating liver-type ATP-dependent 6-phosphofructokinase (Pfkl) expression, and boosting lactate production. The TGF-ß2 signal entered chondrocytes via TGF-ß receptor type I (TßRI), and activated p-Smad3 signaling to regulate the glycolytic pathway. Subsequent experiments employing specific inhibitors of TßRI and p-Smad3 further substantiated the role of TGF-ß2 in enhancement of glycolysis via TßRI/p-Smad3 axis in chondrocytes. The results provide new understanding of the metabolic homeostasis in chondrocytes induced by TGF-ß superfamily and might shed light on the prevention and treatment of related osteoarticular diseases.

Crosstalk between ALK3(BMPR1A) deficiency and autophagy signaling mitigates pathological bone loss in osteoporosis.

Postmenopausal osteoporosis is recognized to be one of the major skeleton diseases strongly associated with impaired bone formation. Previous reports have indicated that the importance of bone morphogenetic protein (BMP) signaling of osteoblast lineage in bone development via classical Smad signaling, however, its critical role in osteoporosis is still not well understood. In the current study, we aim to investigate the pathological role of BMPR1A, a key receptor of BMPs, in osteoporosis and its underlying mechanism. We first found that knockdown of BMPR1A by using Col1a1-creER in osteoblasts mitigated early bone loss of osteoporosis in mice, yet along with late bone maturation defects by reducing mineral adherence rate and bone formation rate in vivo. At the cellular level, we then observed that BMPR1A deficiency promoted the proliferation of pre-osteoblasts under osteoporotic conditions but hindered their late-stage mineralization. We finally elucidated that BMPR1A deficiency compensatorily triggered mTOR-autophagy perturbation by a higher level in early osteoporotic pre-osteoblasts thus resulting in the enhancement of transient cell proliferation but impairment of final mineralization. Taken together, this study indicated the significance of BMPR1A-mTOR/autophagy axis, as a double-edged sword, in osteoporotic bone formation and provided new cues for therapeutic strategies in osteoporosis.

PDGF-AA guides cell crosstalk between human dental pulp stem cells in vitro via the PDGFR-α/PI3K/Akt axis.

AIM: To explore the influence of PDGF-AA on cell communication between human dental pulp stem cells (DPSCs) by characterizing gap junction intercellular communication (GJIC) and its potential biomechanical mechanism. METHODOLOGY: Quantitative real-time PCR was used to measure connexin family member expression in DPSCs. Cell migration and CCK-8 assays were utilized to examine the influence of PDGF-AA on DPSC migration and proliferation. A scrape loading/dye transfer assay was applied to evaluate GJIC triggered by PDGF-AA, a PI3K/Akt signalling pathway blocker (LY294002) and a PDGFR-α blocker (AG1296). Western blotting and immunofluorescence were used to test the expression and distribution of the Cx43 and p-Akt proteins in DPSCs. Scanning electron microscopy (SEM) and immunofluorescence were used to observe the morphology of GJIC in DPSCs. RESULTS: PDGF-AA promoted gap junction formation and intercellular communication between human dental pulp stem cells. PDGF-AA upregulates the expression of Cx43 to enhance gap junction formation and intercellular communication. PDGF-AA binds to PDGFR-α and activates PI3K/Akt signalling to regulate cell communication. CONCLUSIONS: This research demonstrated that PDGF-AA can enhance Cx43-mediated GJIC in DPSCs via the PDGFR-α/PI3K/Akt axis, which provides new cues for dental pulp regeneration from the perspective of intercellular communication.

Hydrogel platform with tunable stiffness based on magnetic nanoparticles cross-linked GelMA for cartilage regeneration and its intrinsic biomechanism.

Cartilage injury affects numerous individuals, but the efficient repair of damaged cartilage is still a problem in clinic. Hydrogel is a potent scaffold candidate for tissue regeneration, but it remains a big challenge to improve its mechanical property and figure out the interaction of chondrocytes and stiffness. Herein, a novel hybrid hydrogel with tunable stiffness was fabricated based on methacrylated gelatin (GelMA) and iron oxide nanoparticles (Fe2O3) through chemical bonding. The stiffness of Fe2O3/GelMA hybrid hydrogel was controlled by adjusting the concentration of magnetic nanoparticles. The hydrogel platform with tunable stiffness modulated its cellular properties including cell morphology, microfilaments and Young's modulus of chondrocytes. Interestingly, Fe2O3/GelMA hybrid hydrogel promoted oxidative phosphorylation of mitochondria and facilitated catabolism of lipids in chondrocytes. As a result, more ATP and metabolic materials generated for cellular physiological activities and organelle component replacements in hybrid hydrogel group compared to pure GelMA hydrogel. Furthermore, implantation of Fe2O3/GelMA hybrid hydrogel in the cartilage defect rat model verified its remodeling potential. This study provides a deep understanding of the bio-mechanism of Fe2O3/GelMA hybrid hydrogel interaction with chondrocytes and indicates the hydrogel platform for further application in tissue engineering.

FGF19 increases mitochondrial biogenesis and fusion in chondrocytes via the AMPKα-p38/MAPK pathway.

Fibroblast growth factor 19 (FGF19) is recognized to play an essential role in cartilage development and physiology, and has emerged as a potential therapeutic target for skeletal metabolic diseases. However, FGF19-mediated cellular behavior in chondrocytes remains a big challenge. In the current study, we aimed to investigate the role of FGF19 on chondrocytes by characterizing mitochondrial biogenesis and fission-fusion dynamic equilibrium and exploring the underlying mechanism. We first found that FGF19 enhanced mitochondrial biogenesis in chondrocytes with the help of ß Klotho (KLB), a vital accessory protein for assisting the binding of FGF19 to its receptor, and the enhanced biogenesis accompanied with a fusion of mitochondria, reflecting in the elongation of individual mitochondria and the up-regulation of mitochondrial fusion proteins. We then revealed that FGF19-mediated mitochondrial biogenesis and fusion required the binding of FGF19 to the membrane receptor, FGFR4, and the activation of AMP-activated protein kinase alpha (AMPKα)/peroxisome proliferator-activated receptor-gamma coactivator 1 alpha (PGC-1α)/sirtuin 1 (SIRT1) axis. Finally, we demonstrated that FGF19-mediated mitochondrial biogenesis and fusion was mainly dependent on the activation of p-p38 signaling. Inhibition of p38 signaling largely reduced the high expression of AMPKα/PGC-1α/SIRT1 axis, decreased the up-regulation of mitochondrial fusion proteins and impaired the enhancement of mitochondrial network morphology in chondrocytes induced by FGF19. Taking together, our results indicate that FGF19 could increase mitochondrial biogenesis and fusion via AMPKα-p38/MAPK signaling, which enlarge the understanding of FGF19 on chondrocyte metabolism. Video Abstract.

IL-10 enhances cell-to-cell communication in chondrocytes via STAT3 signaling pathway.

Gap junction intercellular communication (GJIC) allows the transfer of material, message and energy between cells, which influences cell behaviors including cell proliferation, migration, differentiation and apoptosis and determines cell fate. Interleukin-10 (IL-10), a versatile cytokine, attracts more and more attention in the cartilage pathology such as osteoarthritis (OA) due to its potential in anti-inflammation and wound repair. However, whether IL-10 can mediate GJIC in chondrocytes remains elusive. In the current study, we aimed to explore the role of IL-10 on GJIC and its underlying mechanism. We found that IL-10 can promote GJIC in living chondrocytes. IL-10-enhanced GJIC in chondrocytes was dependent on the up-regulation of connexin 43 (Cx43). Knockdown experiment based on siRNA interference then confirmed that IL-10-enhanced GJIC required participation of IL-10 receptor 1 (IL-10R1). IL-10 activated signal transducer and activator of transcription 3 (STAT3) signaling and promoted the nuclear accumulation of p-STAT3 through IL-10 receptor 1. Inhibitor experiment further confirmed the importance of STAT3 signaling in IL-10-mediated GJIC. Taking together, our results provided a thorough process of IL-10-modulated cell-to-cell communication in chondrocytes and established a bridge between inflammatory factor, IL-10, and GJIC, which can increase our understanding about the physiology and pathology of cartilage.

[Latest Research Findings on the Regulation of Bone Homeostasis by ALK3].

Bone remodeling, which is well orchestrated by osteogenesis of osteoblasts and osteoclastogenesis of osteoclasts, maintains the homeostasis of osteal development and metabolism under physiological conditions. Bone morphogenetic protein receptor type 1A, also known as activin receptor-like kinase 3 (ALK3), which exists on cytomembrane, is one of the key receptors of BMP factors, and is an important "gateway" that regulates the entrance of BMP signaling into cells in order to perform biological functions. The roles of BMP signaling in bone remodeling have been extensively studied. Many new discoveries have been reported in recent years through research based on transgenic mice models and focused on ALK3 as targets, shedding new light on the regulations of bone remodeling, cartilage and joint development, and the occurrence and treatment of bone-related diseases. Established understanding has been expanded, but new challenges on existing clinical application of BMPs also appeared. Hence, we reviewed recent studies on ALK3's involvement in bone formation and bone resorption, analyzed its mechanism of action in bone regulation, summarized the roles of ALK3 in the development of cartilage and temporomandibular joint, and reported the latest progress in treatment in preclinical studies, intending to provide references for subsequent studies and clinical applications in the future.

Vascular Calcification: New Insights Into BMP Type I Receptor A.

Vascular calcification (VC) is a complex ectopic calcification process and an important indicator of increased risk for diabetes, atherosclerosis, chronic kidney disease, and other diseases. Therefore, clarifying the pathogenesis of VC is of great clinical significance. Numerous studies have shown that the onset and progression of VC are similar to bone formation. Members of the bone morphogenetic protein (BMP) family of proteins are considered key molecules in the progression of vascular calcification. BMP type I receptor A (BMPR1A) is a key receptor of BMP factors acting on the cell membrane, is widely expressed in various tissues and cells, and is an important "portal" for BMP to enter cells and exert their biological effect. In recent years, many discoveries have been made regarding the occurrence and treatment of ectopic ossification-related diseases involving BMP signaling targets. Studies have confirmed that BMPR1A is involved in osteogenic differentiation and that its high expression in vascular endothelial cells and smooth muscle cells can lead to vascular calcification. This article reviews the role of BMPR1A in vascular calcification and the possible underlying molecular mechanisms to provide clues for the clinical treatment of such diseases.

TGF-ß2 increases cell-cell communication in chondrocytes via p-Smad3 signalling.

Connexin 43 (Cx43)-mediated gap junction intercellular communication (GJIC) plays a crucial role in the pathology and physiology of joint tissues. Transforming growth factor-ß2 (TGF-ß2), one of the potent regulatory factors in chondrocytes, plays a key role in the regulation of cell cycle and development of joint diseases. However, it is still unknown how TGF-ß2 mediates GJIC in chondrocytes. The aim of this study was to explore the potential mechanism by which TGF-ß2 regulates GJIC in chondrocytes. CCK-8 assays and scratch assays were performed to define the role of TGF-ß2 on cell proliferation and migration. The scrape loading/dye transfer assay and scanning electron microscopy (SEM) were used to verify the effect of TGF-ß2 on GJIC between chondrocytes. qPCR was performed to analyse the expression of genes in the gap junction protein family in chondrocytes. The expression of the Cx43 protein and phosphorylated Smad3 (p-Smad3) was evaluated by western blot assay. Immunofluorescence staining was used to explore p-Smad3 signalling pathway activation and Cx43 distribution. From these experiments, we found that the Cx43 protein was the most highly expressed member of the gap junction protein family in chondrocytes. We also found that TGF-ß2 facilitated cell-to-cell communication in chondrocytes by upregulating Cx43 expression in chondrocytes. Finally, we found that TGF-ß2 activated Smad3 signalling and promoted the nuclear aggregation of p-Smad3. Inhibition experiments by SIS3 also confirmed that TGF-ß2-mediated GJIC through p-Smad3 signalling. For the first time, this study confirmed that TGF-ß2 could regulate the formation of Cx43-mediated GJIC in chondrocytes via the canonical p-Smad3 signalling pathway.

Distribution of bacteria infected by metagenomic sequencing technology in maxillofacial space.

OBJECTIVES: This study aimed to compare and analyze the consistency and difference between metageno-mic next-generation sequencing (mNGS) and conventional bacterial culture in the detection of pathogenic microorganisms in maxillofacial space infection, as well as to provide a new detection method for the early clinical identification of pathogenic bacteria in maxillofacial space infection. METHODS: The clinical data of 16 patients with oral and maxillofacial space infections in the First Affiliated Hospital of Zhengzhou University from March 2020 to June 2020 were collected. mNGS and conventional bacterial culture methods were used to detect pus. We then analyzed and compared the test results of the two methods, including the test cycle, positive detection rate, anaerobic bacteria, facultative anaerobes and aerobic bacteria detection rates, distribution of pathogenic bacteria, relative species abundance, and resistance genes. RESULTS: The average inspection period of mNGS was (18.81±3.73) h, and the average inspection period of bacterial culture was (83.25±11.64) h, the former was shorter than the latter (P<0.05). The positive detection rate of mNGS was 100% (16/16), and the positive detection rate of conventional bacterial culture was 31.25% (5/16), the former was higher than the latter (P<0.05). The detection rate of mNGS anaerobic bacteria was 93.75% (15/16), the detection rate of bacterial culture anaerobes was 0 (0/16), the former was higher than the latter (P<0.05). Using mNGS, the detection rate of facultative anaerobes in bacterial culture was 75.00% (12/16), and the detection rate of facultative anaerobes in bacterial culture was 25.00% (4/16), the former was higher than the latter (P<0.05). The detection rate of aerobic bacteria in bacterial culture was 12.50% (1/16), the former was higher than the latter (P>0.05). mNGS detected 15 kinds of pathogenic bacteria, among which 3 were Gram positive, 12 were Gram negative, 49 were non-pathogenic, 16 were Gram positive, and 32 were Gram negative, 1 was fungus. CONCLUSIONS: Compared with conventional bacterial culture, mNGS has the characteristics of short test time, high sensitivity, and high accuracy. Thus, it is a new detection method for the early identification of pathogenic bacteria in maxillofacial space infection and is beneficial to the early clinical diagnosis and treatment of the disease.

Study on clinical and biological characteristics of ameloblastic carcinoma.

BACKGROUND: Ameloblastic carcinoma (AC) is an odontogenic malignant tumor which is closely related to benign ameloblastoma. Because of its rarity, diagnosis and treatment are difficult. In this study, we summarized and analyzed the clinical and biological characteristics of AC. RESULTS: Fifteen patients with AC and a median age of 53 years were identified. Among of them, five patients who were tested carried a BRAF-V600E mutation. Two patients presented with cervical lymph nodes and lung metastases. Primary AC was more invasive, and the bone destruction ability of the primary type was more radical than that of the secondary type. CONCLUSIONS: This study revealed that the BRAF-V600E mutation was related to the aggressive behavior of AC, and early radical resection is crucial. Moreover, targeted therapy may be a new direction in the future.

[Clinical value of oral repair membrane and ß-tricalcium phosphate in the treatment of the postoperative bone defect of jaw cyst].

OBJECTIVE: This study aims to evaluate the clinical effect of oral repair membrane and ß-tricalcium phosphate (ß-TCP) on the treatment of jaw cyst. METHODS: A retrospective analysis was performed on 81 cases of jaw cysts, and clinical data were collected for the comparison of traditional surgical curettage (group A, 27 cases), biofilm covering bone wounds after curettage (group B, 27 cases), and ß-TCP filling combined with biofilm covering. RESULTS: No recurrence occurred in 81 patients, and no significant difference in preoperative CT value among the three groups (P<0.05). Follow-up CT reexamination 3, 6, and 12 months after operation showed significant differences among the three groups of CT values (P<0.05). Group C was better than Group B or Group A (P<0.05). CONCLUSIONS: In traditional jaw cyst curettage, the application of biofilm exhibited good osteogenesis effect, and the combined application of ß-TCP and biofilm exerted a better effect.

Downregulation of MicroRNA-551b Correlates With Dissemination of Human Oral Squamous Cell Carcinoma.

PURPOSE: Altered expression of microRNAs contributes to invasion and metastasis of many human cancers; however, the importance of microRNAs in head and neck cancers remains to be elucidated. In this study, we examined whether altered microRNA (miR)-551b expression correlated with invasive phenotypes of human oral squamous cell carcinoma (OSCC) in vivo and in vitro. MATERIALS AND METHODS: Real-time polymerase chain reaction was used to detect the expression level of miR-551b in 71 OSCC tissues with lymph node metastasis and 50 nonmetastatic OSCC tissues. We also constructed miR-551b mimic-transfected cell lines HN4 and HN12. The effects of overexpressing miR-551b on the proliferation, migration, and invasion of OSCC cells were examined using Cell Counting Kit 8 (Dojindo, Kumamoto, Japan), plate clone formation, wound healing, and Transwell invasion experiments (Corning, Corning, NY). The association between clinical pathologic parameters and the expression level of miR-551b was analyzed using Kaplan-Meier survival analysis. RESULTS: The expression of miR-551b measured 0.33 ± 0.11 in the 71 OSCC tissues with lymph node metastasis versus 0.54 ± 0.06 in the 50 tissues with non-lymph node metastasis (P = .021). Regarding OSCC patients, the expression of miR-551b negatively correlated with patients' overall survival (P = .035). The ectopic expression of miR-551b inhibited the invasion and migration of OSCC cells. CONCLUSIONS: This is the first report showing that reduced miR-551b expression may be an event leading to OSCC invasion and metastasis.

Piwi-Interacting RNA1037 Enhances Chemoresistance and Motility in Human Oral Squamous Cell Carcinoma Cells.

BACKGROUND: Piwi-interacting RNAs (piRNAs) are thought to silence transposable genetic elements. However, the functional roles of piRNAs in oral squamous cell carcinoma (OSCC) remain unelucidated. In the present study, we aimed to investigate the role of Piwi-interacting RNA 1037 (piR-1037) in chemoresistance to cisplatin (CDDP)-based chemotherapy and the oncogenic role of piR-1037 in OSCC cells. METHODS: RT-PCR was used to evaluate the levels of piR-1037 and X-linked Inhibitor of apoptosis protein (XIAP) mRNA in OSCC cell lines or tumor xenografts. Transfection of piR-1037 DNA antisense and piR-1037 RNA oligonucleotides was performed to suppress and overexpress piR-1037 in OSCC cells, respectively. A CCK8 assay was used to measure the viability or proliferation of OSCC cells. Apoptosis in OSCC cells and xenografts was determined using a TUNEL assay kit. The activity of caspase-3, caspase-8 and caspase-1 in OSCC cells was measured with colorimetric caspase assay kits. Western blot analysis was conducted to analyze XIAP expression in OSCC cells and xenograft samples. Immunoprecipitation (IP) and RNA pull-down assays were utilized to analyze the piR-1037 - XIAP interaction. Transwell assays were performed to evaluate migration and invasion of OSCC cells. RESULTS: CDDP treatment upregulated piR-1037 expression in OSCC cells and OSCC xenografts. Suppression of the CDDP-induced upregulation of piR-1037 expression enhanced the sensitivity of OSCC cells to CDDP. piR-1037 promoted protein expression and directly bound XIAP, a key apoptotic inhibitor that is implicated in chemoresistance. The relationship between piR-1037 and XIAP suggested that piR-1037 enhanced OSCC cell chemoresistance to CDDP at least partially through XIAP. Moreover, targeting the basal expression of piR-1037 inhibited cell motility by affecting epithelial-mesenchymal transition (EMT). CONCLUSION: piR-1037 enhances the chemoresistance and motility of OSCC cells. piR-1037 promotes chemoresistance by interacting with XIAP and regulates the motility of OSCC cells by driving EMT.

[Clinical value of vacuum sealing drainage in the treatment of oral and maxillofacial space infection].

OBJECTIVE: This study aims to observe the efficacy of vacuum sealing drainage (VSD) by continuous negative pressure drainage and saline irrigation in the treatment of oral and maxillofacial space infection. METHODS: Retrospective analysis was conducted on 116 cases of maxillofacial space infection, and clinical data were collected to compare the therapeutic effects of routine incision with drainage treatment (traditional treatment group, 58 cases) and VSD treatment (VSD group, 58 cases). RESULTS: The length of hospital stay, white blood cell count, scar length, frequency of dressing change, and pain degree of patients in the VSD group were all lower than those in the traditional treatment group. Moreover, the improvement degree of mouth opening in the VSD groups was better than that in the traditional treatment group (P<0.05). CONCLUSIONS: VSD is a more effective method for the treatment of oral and maxillofacial space infection.

Diagnosis and treatment of a carotid body tumor: A case report of a rare bilateral tumor.

In the present case report, a rare bilateral carotid body tumor (CBT) and the imaging and pathological features of a CBT are described. In the present report, a rare case of bilateral carotid body tumor, which developed in the bifurcation of the common carotid artery, and the clinical manifestations, imaging and pathological features of this CBT are summarized. The imaging cannot validate the diagnosis; however, imaging identified that the tumor exhibited an intact envelope. Immunohistochemical staining revealed that the tumor cells were strongly positive for cluster of differentiation 56, Syn and protein S-100, moderately positive for transcription factor E3, negative for cytokeratin and epithelial membrane antigen, and partial cells were weakly positive for Desmir (<5%). In view of the clinical and pathological features of the carotid body tumor, surgery is hypothesized to be the optimal treatment and may enable the tumor to be resected completely. Refined surgical techniques provide the security of safe resection and decrease the risk of complications occurring.