FBXO11 is the substrate-recognition component of a ubiquitin ligase complex called SKP1-cullin-F-boxes. The role of FBXO11 in bone development is unexplored. In this study, we reported a novel mechanism of how bone development is regulated by FBXO11. FBXO11 gene knockdown by lentiviral transduction in mouse pre-osteoblast MC3T3-E1 cells leads to reduced osteogenic differentiation, while overexpressing FBXO11 acceler-ates their osteogenic differentiation in vitro. Furthermore, we generated two osteoblastic-specific FBXO11 conditional knockout mouse models, Col1a1-ERT2-FBXO11KO and Bglap2-FBXO11KO mice. In both conditional FBXO11KO mouse models, we found FBXO11 deficiency inhibits normal bone growth, in which the osteogenic activity in FBXO11cKO mice is reduced, while osteoclastic activity is not significantly changed. Mechanistically, we found FBXO11 deficiency leads to Snail1 protein accumulation in osteoblasts, leading to suppression of osteogenic activity and inhibition of bone matrix mineralization. FBXO11 knockdown in MC3T3-E1 cells reduced Snail1 protein ubiquitination and increased Snail1 protein accumulation in the cells, which eventually inhibited osteogenic differentiation. In conclusion, FBXO11 deficiency in osteoblasts inhibits bone formation through Snail1 accumulation, inhibiting osteogenic activity and bone mineralization.

Small airway fibrosis is a common pathology of chronic obstructive pulmonary disease (COPD) and contributes to airflow obstruction. However, the underlying fibrogenic mechanism is poorly understood. Epithelial-mesenchymal transition (EMT) has been proposed as a driver of fibrosis. EMT occurs in the airways of COPD patients and smokers, but it remains elusive whether EMT may contribute to airway fibrosis. We previously reported that FBXO11 is a critical suppressor of EMT and Fbxo11 deficiency in mice causes neonatal lethality and EMT in epidermis. Here, we found that Fbxo11 -deficient mouse embryonic lungs showed impaired epithelial differentiation, excess fibroblast cells surrounding the airways, and thickened interstitial mesenchyme. We further generated conditional mutant mice to ablate Fbxo11 selectively in the club airway epithelial cells in adult mice, which induced partial EMT in the airways. To determine the effect of EMT on airway fibrosis, Fbxo11 conditional mutant mice were exposed to cigarette smoke. Airway-specific loss of Fbxo11 markedly enhanced smoking-induced airway fibrotic remodeling and collagen deposition. Taken together, our study suggests that EMT in the airway epithelium exacerbates cigarette smoke-induced airway fibrosis.

Epithelial–mesenchymal transition (EMT) is implicated in tumor metastasis and therapeutic resistance. It remains a challenge to target cancer cells that have undergone EMT. The Snail family of key EMT-inducing transcription factors directly binds to and transcriptionally represses not only epithelial genes but also a myriad of additional genomic targets that may carry out significant biological functions. Therefore, we reasoned that EMT inherently causes various concomitant phenotypes, some of which may create targetable vulnerabilities for cancer treatment. In the present study, we found that Snail transcription factors bind to the promoters of multiple genes encoding subunits of the AMP-activated protein kinase (AMPK) complex, and expression of AMPK genes was markedly downregulated by EMT. Accordingly, high AMPK expression in tumors correlated with epithelial cell markers and low AMPK expression in tumors was strongly associated with adverse prognosis. AMPK is the principal sensor of cellular energy status. In response to energy stress, AMPK is activated and critically reprograms cellular metabolism to restore energy homeostasis and maintain cell survival. We showed that activation of AMPK by energy stress was severely impaired by EMT. Consequently, EMT cancer cells became hypersensitive to a variety of energy stress conditions and primarily underwent pyroptosis, a regulated form of necrotic cell death. Collectively, the study suggests that EMT impedes the activation of AMPK signaling induced by energy stress and sensitizes cancer cells to pyroptotic cell death under energy stress conditions. Therefore, while EMT promotes malignant progression, it concurrently induces collateral vulnerabilities that may be therapeutically exploited.

OBJECTIVE Our study aimed to determine the relationship of antidepressant medicine use with periodontal diseases, exploring the association of different pharmacological classes of antidepressant with observations of clinical attachment loss (CAL) and alveolar bone level (BL) in patients with periodontitis. BACKGROUND Existing evidence on the impact of antidepressant medication on periodontal tissues has focused on some classes only and is still unclear. Therefore, this retrospective study evaluated the association of different antidepressant classes with clinical attachment loss (CAL) and alveolar bone level (BL). METHODS This study was carried out in a population of patients aged ≥ 30 years old with periodontitis who sought treatment at the University of Florida from 2014 to 2018. The following variables were obtained from patients' records; usage of antidepressant medications and their pharmacological classes (selective serotonin reuptake inhibitors [SSRI], serotonin-norepinephrine reuptake inhibitors [SNRI], tricyclic, atypical, and monoamine oxidase inhibitors [MAO]), age, gender, smoking habit, mild systemic diseases, CAL, and cement-enamel junction (CEJ) and alveolar bone crest (BC) distance, defined as BL, in the Ramfjord index teeth. RESULTS Five hundred and eighty-two periodontitis patients were evaluated, of which 113 (19.4%) were antidepressant users. Antidepressant users exhibited significantly lower BL and fewer sites with severe CAL (≥5 mm), than non-users (p < .05). Among all single-class antidepressant users, the SSRI users showed significantly less CAL and lower BL than non-users (p < .05). Patients taking combinations of the different classes of antidepressants also showed better CAL and BL than non-users. Generalized linear models, including variables such as gender, age, systemic diseases, and smoking, demonstrated that antidepressant users were more likely to have lower mean BL and fewer sites with severe bone loss (i.e. BL > 3 and >5 mm) than non-users (p < .05). CONCLUSIONS Antidepressant medications were associated with higher alveolar bone level and less clinical attachment loss in patients with periodontitis. When the different classes of antidepressants were analyzed individually, only the SSRI class users and the multiple-class users showed significantly less periodontal breakdown than non-users.

Titanium hypersensitivity is rare, but it may exist among patients who need dental implant treatment. It could contribute to mild or severe peri-implant tissue hypersensitivity reactions and affect patients' oral and general health according to some clinical reports. In extreme cases, it may lead to implant failure or extraction. In this case report, a 64-year-old Caucasian female patient received a titanium implant placement on #5. Right after the implant placement, she reported the symptoms of pain, eczema, and slight extraoral swelling, along with significant burning sensation occurring intra- and extraorally. The symptoms were not released after the systemic intervention of antibiotics for six days. On retrieving her medical history, the patient reported a previous allergic reaction to jewelry-like earrings in her childhood. The diagnosis of titanium hypersensitivity was made based on the rapid onset of symptoms and her metal allergy history. Therefore, the dental implant was removed after diagnosis, and a bone allograft was used to preserve the alveolar bone volume. Immediately after implant extraction, the swelling and burning sensation were receded. A complete recovery was achieved three weeks later. The dimension of the alveolar ridge bone was found well maintained in 10 weeks follow-up visit. Conclusion. Rapid onset of peri-implant gingival swelling may indicate a hypersensitivity to titanium implant in the clinic. For patients with a history of allergy to jewelry, the hypersensitivity tests to titanium alloy such as patch test or lymphocyte transformation test should be recommended as part of the dental implant treatment plan.

Insufficient nutrition exchange and limited transportation of blood supply in a porous only scaffold often hinder bone formation, even though the porous scaffold is loaded with cells or growth factors. To overcome these issues, we developed a cell- and growth factor-free approach to induce bone formation in a critical-size bone defect by using an interconnected porous beta-tricalcium phosphate (β-TCP) scaffold with multiple channels. In vitro cell experimental results showed that multiple channels significantly promoted cell attachment and proliferation of human bone marrow mesenchymal stem cells, stimulated their alkaline phosphatase activity, and up-regulated the osteogenic gene expression. Multiple channels also considerably stimulated the expression of various mechanosensing markers of the cells, such as focal adhesion kinase, filamentous actin, and Yes-associated protein-1 at both static and dynamic culturing conditions. The in vivo bone defect implantation results demonstrated more bone formation inside multiple-channeled scaffolds compared to non-channeled scaffolds. Multiple channels prominently accelerated collagen type I, bone sialoprotein and osteocalcin protein expression. Fluorochrome images and angiogenic marker CD31 staining exhibited more mineral deposition and longer vasculature structures in multiple-channeled scaffolds, compared to non-channeled scaffolds. All the findings suggested that the creation of interconnected multiple channels in the porous β-TCP scaffold is a very promising approach to promote bone tissue regeneration.

BACKGROUND Previous studies have suggested an association between taking antidepressants and dental implant failure. This study aimed to investigate the association of different antidepressant classes with dental implant failure. METHODS This retrospective study included patients that received dental implants at the University of Florida from 2011 to 2016. The variables of implant failure, antidepressant use and classes (selective serotonin reuptake inhibitors [SSRI], serotonin-norepinephrine reuptake inhibitors [SNRI], tricyclic antidepressants [TCA], atypical antidepressants [AA], and monoamine oxidase inhibitors [MAOI]), age, gender, smoking, mild systemic diseases and implant location were obtained from patients' records. Odds ratio (OR) and confidence intervals (CI) of implant failure in patients taking different antidepressant classes, in relation to non-antidepressant users, were estimated, and the influence of multiple variables on implant failure were investigated. RESULTS 771 patients and 1820 implants were evaluated. The statistically significant predictors for implant failure included: Smoking (OR = 5.221), use of antidepressants (OR = 4.285), posterior maxilla location (OR = 2.911), mild systemic disease (OR = 2.648), and age (OR = 1.037) (P < 0.05). The frequency of implant failure was 33.3% in TCA users, 31.3% in SNRI users, 6.3% in SSRI users, 5.2% in Atypical antidepressant users, and 3.9% in non-users. Significant associations were observed between the use of SNRI (OR: 11.07; 95% CI: 3.265-33.82) and TCA (OR: 12.16; 95% CI: 1.503-71.58) and implant failure (P < 0.05). CONCLUSION Users of antidepressants were at higher risk of implant failure than non-users. Patients taking SNRI and TCA were at the highest risk of implant loss, when compared to non-users. Conclusions about TCA, however, are based on a limited number of cases. This article is protected by copyright. All rights reserved.

The review aimed at assessing the osteopromotive potential as well as soft tissue and temporomandibular joint (TMJ) cartilage healing properties of simvastatin by summarizing its efficacy on the current dental treatment of periodontal bone and soft tissue defects, and temporomandibular joint (TMJ) arthritis from the available animals and human studies. An electronic search was performed on MEDLINE, Scopus, and Cochrane Central Register of Controlled Trials (CENTRAL) using a combination of keywords. A hand search was undertaken on seven oral surgery journals. No limitation of publication year in the English language was placed. Controlled randomized animal and human clinical trials, as well as prospective comparative studies, were included. Data on the comparison of topical/systemic simvastatin on bone healing in intrabony and furcation defects, extraction sockets, distraction osteogenesis, as well as soft tissue healing in mucogingival grafting procedures and cartilage protection in TMJ arthritis were extracted from all the eligible studies. Studies with a minimum of ten participants and follow up at least 6 months were included. Ten animal studies and six clinical studies were included in this study. All the animal studies included a minimum of eight sites per group assessed clinically, histologically, and radiographically. All human studies included clinical and radiological evaluation. The results of the review show that simvastatin administration displays positive treatment outcomes in the full range of therapies investigated in the oral regions such as periodontal infection control, periodontal and alveolar bone regeneration, soft tissue grafting, TMJ inflammation reduction, and cartilage repair. Its mechanism includes stimulating bone formation, promoting soft tissue healing, increasing articular and condylar cartilage thickness, as well as reducing inflammation at surgical sites in TMJ disorders. Simvastatin administration is beneficial to the healing of oral bone and cartilage. More studies are desired to determine its potential in soft tissue healing.

Vascularization is essential for tissue regeneration. Despite extensive efforts in the past decades, sufficient and rapid vascularization remains a major challenge in tissue engineering. Many studies have shown that the addition of channels in a porous scaffold can provide the ability to promote cell growth and rapid vascularization, thus leading to better outcomes in new tissue formation. Large size scaffolds lack perfusable channel networks and negatively impair the survival of transplanted cells and tissue function development, leading to necrotic core formation and the failure of functional tissue formation. Presently, there are many methods to produce channels in porous scaffolds for vascularization. Here, we review the function of channels in porous scaffolds and the approaches to produce those channels.

The bone marrow contains a heterogeneous milieu of cells, including macrophages, which are key cellular mediators for resolving infection and inflammation. Macrophages are most well known for their ability to phagocytose foreign bodies or apoptotic cells to maintain homeostasis; however, little is known about their function in the bone microenvironment. In the current study, we investigated the in vitro interaction of murine macrophages and bone marrow stromal cells (BMSCs), with focus on the juxtacrine induction of IL-6 signaling and the resultant effect on BMSC migration and growth. The juxtacrine interaction of primary mouse macrophages and BMSCs activated IL-6 signaling in the co-cultures, which subsequently enhanced BMSC migration and increased BMSC numbers. BMSCs and macrophages harvested from IL-6 knockout mice revealed that IL-6 signaling was essential for enhancement of BMSC migration and increased BMSC numbers via juxtacrine interactions. BMSCs were the main contributor of IL-6 signaling, and hence activation of the IL-6/gp130/STAT3 pathway. Meanwhile, macrophage derived IL-6 remained important for the overall production of IL-6 protein in the co-cultures. Taken together, these findings show the function of macrophages as co-inducers of migration and growth of BMSCs, which could directly influence bone formation and turnover.

Wnt4 promotes bone growth while inhibiting bone loss to prevent osteopenia in a mouse model. Aging-related bone loss and osteoporosis affect millions of people worldwide. Chronic inflammation associated with aging promotes bone resorption and impairs bone formation. Here we show that Wnt4 attenuates bone loss in osteoporosis and skeletal aging mouse models by inhibiting nuclear factor-κB (NF-κB) via noncanonical Wnt signaling. Transgenic mice expressing Wnt4 from osteoblasts were significantly protected from bone loss and chronic inflammation induced by ovariectomy, tumor necrosis factor or natural aging. In addition to promoting bone formation, Wnt4 inhibited osteoclast formation and bone resorption. Mechanistically, Wnt4 inhibited NF-κB activation mediated by transforming growth factor-β–activated kinase-1 (Tak1) in macrophages and osteoclast precursors independently of β-catenin. Moreover, recombinant Wnt4 alleviated bone loss and inflammation by inhibiting NF-κB in vivo in mouse models of bone disease. Given its dual role in promoting bone formation and inhibiting bone resorption, our results suggest that Wnt4 signaling could be an attractive therapeutic target for treating osteoporosis and preventing skeletal aging.

Mesenchymal stem cell (MSC)-based transplantation is a promising therapeutic approach for bone regeneration and repair. In the realm of therapeutic bone regeneration, the defect or injured tissues are frequently inflamed with an abnormal expression of inflammatory mediators. Growing evidence suggests that proinflammatory cytokines inhibit osteogenic differentiation and bone formation. Thus, for successful MSC-mediated repair, it is important to overcome the inflammation-mediated inhibition of tissue regeneration. In this study, using genetic and chemical approaches, we found that proinflammatory cytokines TNF and IL-17 stimulated IκB kinase (IKK)–NF-κB and impaired osteogenic differentiation of MSCs. In contrast, the inhibition of IKK–NF-κB significantly enhanced MSC-mediated bone formation. Mechanistically, we found that IKK–NF-κB activation promoted β-catenin ubiquitination and degradation through induction of Smurf1 and Smurf2. To translate our basic findings to potential clinic applications, we showed that the IKK small molecule inhibitor, IKKVI, enhanced osteogenic differentiation of MSCs. More importantly, the delivery of IKKVI promoted MSC-mediated craniofacial bone regeneration and repair in vivo. Considering the well established role of NF-κB in inflammation and infection, our results suggest that targeting IKK–NF-κB may have dual benefits in enhancing bone regeneration and repair and inhibiting inflammation, and this concept may also have applicability in many other tissue regeneration situations.

Human bone marrow mesenchymal stem/stromal cells (MSCs) are multipotent progenitor cells with multilineage differentiation potentials including osteogenesis and adipogenesis. While significant progress has been made in understanding transcriptional controls of MSC fate, little is known about how MSC differentiation is epigenetically regulated. Here we show that the histone demethylases KDM4B and KDM6B play critical roles in osteogenic commitment of MSCs by removing H3K9me3 and H3K27me3. Depletion of KDM4B or KDM6B significantly reduced osteogenic differentiation and increased adipogenic differentiation. Mechanistically, while KDM6B controlled HOX expression by removing H3K27me3, KDM4B promoted DLX expression by removing H3K9me3. Importantly, H3K27me3- and H3K9me3-positive MSCs of bone marrow were significantly elevated in ovariectomized and aging mice in which adipogenesis was highly active. Since histone demethylases are chemically modifiable, KDM4B and KDM6B may present as therapeutic targets for controlling MSC fate choices and lead to clues for new treatment in metabolic bone diseases such as osteoporosis.

The role of the transcription factor NFκB in osteoclasts and bone degradation is well understood. In this Perspectives article, the authors discuss its newly described inhibitory function in osteoblasts and bone formation, and how therapies that target NFκB might be beneficial in osteoporosis and other inflammatory bone diseases. NFκB is a family of transcription factors involved in immunity and the normal functioning of many tissues. It has been well studied in osteoclasts, and new data indicate an important role for NFκB in the negative regulation of bone formation. In this article, we discuss how NFκB activation affects osteoblast function and bone formation. In particular, we describe how reduced NFκB activity in osteoblasts results in an increase in bone formation via enhanced c-Jun N-terminal kinase (JNK) activity, which regulates FOSL1 (also known as Fra1) expression. Furthermore, we discuss how estrogen and NFκB crosstalk in osteoblasts acts to oppositely regulate bone formation. Future NFκB-targeting treatments for osteoporosis, rheumatoid arthritis and other inflammatory bone diseases could lead to increased bone formation concurrent with decreased bone resorption.

Control of organ size by cell proliferation and survival is a fundamental developmental process, and its deregulation leads to cancer. However, the molecular mechanism underlying organ size control remains elusive in vertebrates. In Drosophila, the Hippo (Hpo) signaling pathway controls organ size by both restricting cell growth and proliferation and promoting cell death. Here we investigated whether mammals also require the Hpo pathway to control organ size and adult tissue homeostasis. We found that Mst1 and Mst2, the two mouse homologs of the Drosophila Hpo, control the sizes of some, but not all organs, in mice, and Mst1 and Mst2 act as tumor suppressors by restricting cell proliferation and survival. We show that Mst1 and Mst2 play redundant roles, and removal of both resulted in early lethality in mouse embryos. Importantly, tumors developed in the liver with a substantial increase of the stem/progenitor cells by 6 months after removing Mst1 and Mst2 postnatally. We show that Mst1 and Mst2 were required in vivo to control Yap phosphorylation and activity. Interestingly, apoptosis induced by TNFα was blocked in the Mst1 and Mst2 double-mutant cells both in vivo and in vitro. As TNFα is a pleiotropic inflammatory cytokine affecting most organs by regulating cell proliferation and cell death, resistance to TNFα-induced cell death may also contribute significantly to tumor formation in the absence of Mst1 and Mst2.

An imbalance in bone formation relative to bone resorption results in the net bone loss that occurs in osteoporosis and inflammatory bone diseases. Although it is well known how bone resorption is stimulated, the molecular mechanisms that mediate impaired bone formation are poorly understood. Here we show that the time- and stage-specific inhibition of endogenous inhibitor of kappa B kinase (IKK)-nuclear factor-kappa B (NF-kappa B) in differentiated osteoblasts substantially increases trabecular bone mass and bone mineral density without affecting osteoclast activities in young mice. Moreover, inhibition of IKK-NF-kappa B in differentiated osteoblasts maintains bone formation, thereby preventing osteoporotic bone loss induced by ovariectomy in adult mice. Inhibition of IKK-NF-kappa B enhances the expression of Fos-related antigen-1 (Fra-1), an essential transcription factor involved in bone matrix formation in vitro and in vivo. Taken together, our results suggest that targeting IKK-NF-kappa B may help to promote bone formation in the treatment of osteoporosis and other bone diseases.

Mesenchymal stem cells (MSCs) are multipotent cells that can be differentiated into osteoblasts and provide an excellent cell source for bone regeneration and repair. Recently, the canonical Wnt/β-catenin signaling pathway has been found to play a critical role in skeletal development and osteogenesis, implying that Wnts can be utilized to improve de novo bone formation mediated by MSCs. However, it is unknown whether noncanonical Wnt signaling regulates osteogenic differentiation. Here, we find that Wnt-4 enhanced in vitro osteogenic differentiation of MSCs isolated from human adult craniofacial tissues and promoted bone formation in vivo. Whereas Wnt-4 did not stabilize β-catenin, it activated p38 MAPK in a novel noncanonical signaling pathway. The activation of p38 was dependent on Axin and was required for the enhancement of MSC differentiation by Wnt-4. Moreover, using two different models of craniofacial bone injury, we found that MSCs genetically engineered to express Wnt-4 enhanced osteogenesis and improved the repair of craniofacial defects in vivo. Taken together, our results reveal that noncanonical Wnt signaling could also play a role in osteogenic differentiation. Wnt-4 may have a potential use in improving bone regeneration and repair of craniofacial defects.

Background: Micro-computed tomography (micro-CT) offers significant potential for identifying mineralized structures. However, three-dimensional (3-D) micro-CT of alveolar bone has not been adapted readily for quantification. Moreover, conventional methods are not highly sensitive for analyzing bone loss or bone gain following periodontal disease or reconstructive therapy. The objective of this investigation was to develop a micro-CT methodology for quantifying tooth-supporting alveolar bone in 3-D following experimental preclinical situations of periodontitis or reconstructive therapy.Methods: Experimental in vivo bone loss or regeneration situations were developed to validate the micro-CT imaging techniques. Twenty mature Sprague-Dawley rats were divided into two groups: bone loss (Porphyromonas gingivalis lipopolysaccharide-mediated bone resorption) and regenerative therapy. Micro-CT and software digitized specimens were reconstructed three-dimensionally for linear and volumetric parameter assessment of alveolar bone (linear bone height, bone volume, bone volume fraction, bone mineral content, and bone mineral density). Intra- and interexaminer reproducibility and reliability were compared for methodology validation.Results: The results demonstrated high examiner reproducibility for linear and volumetric parameters with high intraclass correlation coefficient (ICC) and coefficient of variation (CV). The ICC showed that the methodology was highly reliable and reproducible (ICC > 0.99; 95% confidence interval, 0.937 to 1.000; CV < 1.5%), suggesting that 3-D measurements may provide better alveolar bone analysis than conventional 2-D methods.Conclusions: The developed methods allow for highly accurate and reproducible static measurements of tooth-supporting alveolar bone following preclinical situations of bone destruction or regeneration. Future investigations should focus on using in vivo micro-CT imaging for real-time assessments of alveolar bone changes.

Mesenchymal stem cells (MSCs) are multipotent cells that can be differentiated into osteoblasts and provide an excellent cell source for bone regeneration and repair. Recently, the canonical Wnt/-catenin signaling pathway has been found to play a critical role in skeletal development and osteogenesis, implying that Wnts can be utilized to improve de novo bone formation mediated by MSCs. However, it is unknown whether noncanonical Wnt signaling regulates osteogenic differentiation. Here, we find that Wnt-4 enhanced in vitro osteogenic differentiation of MSCs isolated from human adult craniofacial tissues and promoted bone formation in vivo. Whereas Wnt-4 did not stabilize -catenin, it activated p38 MAPK in a novel noncanonical signaling pathway. The activation of p38 was dependent on Axin and was required for the enhancement of MSC differentiation by Wnt-4. Moreover, using two different models of craniofacial bone injury, we found that MSCs genetically engineered to express Wnt-4 enhanced osteogenesis and improved the repair of craniofacial defects in vivo. Taken together, our results reveal that noncanonical Wnt signaling could also play a role in osteogenic differentiation. Wnt-4 may have a potential use in improving bone regeneration and repair of craniofacial defects.