A WAP-Based Concept Acquisition Teaching Model in Cleft Lip and Palate Phenotype and Embryonic Development: Functionality and Usability Study.

OBJECTIVE: Taking advantage of the broad coverage of Wireless Application Protocol (WAP), we developed a Content Management System (CMS)-programmed mobile learning application. This application can help the undergraduate to obtain a comprehensive understanding of concepts in Cleft lip and palate Phenotype, and Embryonic development (CPE). The present study aims to evaluate the feasibility and efficacy of the concept acquisition teaching model on the basis of WAP in a practical undergraduate course of CPE. DESIGN: The concept acquisition teaching model based on WAP was programmed by CMS, covering definitions of various cleft lip and palate, the mechanisms underlying the phenotypes, practical medical cases, as well as corresponding tests after learning. SETTING: The CPE concept acquisition teaching model was distributed to a total of 524 undergraduate students and 46 tutors participated in CPE teaching from seven highly ranked schools of stomatology in China since April 2022. PARTICIPANTS: 524 undergraduate students and 46 tutors from seven highly ranked schools of stomatology in China. INTERVENTIONS: The CPE concept acquisition teaching model. MAIN OUTCOME MEASURES: The effectiveness of the CPE teaching model. RESULTS: The response rate to the survey was 100%. The grading of the questionnaires indicated that the students were satisfied with the usability, practicality, and outcome, whereas the tutors were more positive with the contents, cooperation, and outcome. CONCLUSIONS: The present study demonstrated the feasibility and efficacy of the WAP-based concept acquisition teaching model of CPE and a high level of satisfaction among undergraduate students and tutors who major in Stomatology.

Soft palate dysplasia: properties and surgical techniques.

OBJECTIVES: We aimed to report a hitherto undescribed class of patients with the obvious phenotype of a novel soft palate dysplasia combining unilateral soft palate hypoplasia with a fully developed uvula. We also aimed to investigate and evaluate the corresponding surgical approaches. MATERIALS AND METHODS: Twelve patients were clinically diagnosed with soft palate dysplasia. Clinical examination, including radiographic tests was performed to characterize the congenital deformity. The effectiveness of velopharyngeal closure and speech were tested pre- and post-operation. RESULTS: Soft palate dysplasia was featured with velopharyngeal insufficiency, food regurgitation, and speech disorders. It was commonly manifested as structural deformities of the soft palate, tongue palatine arch, pharyngeal palatine arch, and pterygomandibular fold, but complete uvula shape. According to radiographic analysis, in five patients the lateral pterygoid processes were poorly developed and other malformations were present. Velopharyngoplasty based on the unilateral posterior pharyngeal flap can well restore the velopharyngeal closure and speech intelligibility without respiration obstruction. CONCLUSIONS: Soft palate dysplasia is characterized as congenital velopharyngeal insufficiency manifested as a primary soft palate defect. It is highly associated with other physical deformities but independent of conventionally known syndromes. The cause may be an abnormal development of the pterygoid process. Unilateral velopharyngoplasty based on the posterior pharyngeal flap is a great technique to repair soft palate dysplasia (SPD).Clinical Relevance For soft palate muscle defects without cleft palate, we proposed a surgical technique by which personalized design of the posterior pharyngeal flap could be fulfilled according to the degree of deformity. It can restore the symmetry of the soft palate.

Real-Time Image-Guided Navigation in the Management of Alveolar Cleft Repair.

Objectives: To present the use of dynamic navigation system in the repair of alveolar cleft. Patients and Participants: A total of three non-syndromic patients with unilateral alveolar cleft were involved in this study. Real-time computer-aided navigation were used to achieve restoration and reconstruction with standardized surgical technique. Methods: With the individual virtual 3-dimensional (3-D) modeling based on computed tomography (CT) data, preoperative planning and surgical simulation were carried out with the navigation system. During preoperative virtual planning, the defect volume or the quantity of graft is directly assessed at the surgical region. With the use of this system, the gingival periosteum flap incision can be tracked in real-time, and the bone graft can be navigated under the guidance of the 3-D views until it matches the preoperatively planned position. Results: Three patients with alveolar cleft were successfully performed under navigation guidance. Through the model alignment procedure, accurate matches between the actual intraoperative position and the CT images were achieved within the systematic error of 0.3 mm. The grafted bone was implanted according to the preoperative plan with the aid of instrument- and probe-based navigation. All the patients were healed well without serious complications. Conclusions: These findings suggest that image-guided surgical navigation, including preoperative planning, surgical simulation, postoperative assessment, and computer-assisted navigation was feasible and yielded good clinical outcomes. Clinical relevance: This dynamic navigation could be proved to be a valuable option for this complicated surgical procedure in the management of alveolar cleft repair.

Mutations in the TBX15-ADAMTS2 pathway associate with a novel soft palate dysplasia.

We reported de novo variants in specific exons of the TBX15 and ADAMTS2 genes in a hitherto undescribed class of patients with unique craniofacial developmental defects. The nine unrelated patients represent unilateral soft palate hypoplasia, lost part of the sphenoid bone in the pterygoid process, but the uvula developed completely. Interestingly, these clinical features are contrary to the palate's anterior-posterior (A-P) developmental direction. Based on developmental characteristics, we suggested that these cases correspond to a novel craniofacial birth defect different from cleft palate, and we named it soft palate dysplasia (SPD). However, little is known about the molecular mechanism of the ADAMTS2 and TBX15 genes in the regulation of soft palate development. Phylogenetic analysis showed that the sequences around these de novo mutation sites are conserved between species. Through cellular co-transfections and chromatin immunoprecipitation assays, we demonstrate that TBX15 binds to the promoter regions of the ADAMTS2 gene and activates the promoter activity. Furthermore, we show that TBX15 and ADAMTS2 are colocalization in the posterior palatal mesenchymal cells during soft palate development in E13.5 mice embryos. Based on these data, we propose that the disruption of the TBX15-ADAMTS2 signaling pathway during embryogenesis leads to a novel SPD.

The Protective Effects of Sulforaphane on High-Fat Diet-Induced Obesity in Mice Through Browning of White Fat.

Background: Sulforaphane (SFN), an isothiocyanate naturally occurring in cruciferous vegetables, is a potent indirect antioxidant and a promising agent for the control of metabolic disorder disease. The glucose intolerance and adipogenesis induced by diet in rats was inhibited by SFN. Strategies aimed at induction of brown adipose tissue (BAT) could be a potentially useful way to against obesity. However, in vivo protective effect of SFN against obesity by browning white adipocyte has not been reported. Our present study is aimed at evaluation the efficacy of the SFN against the high-fat induced-obesity mice and investigating the potential mechanism. Methods: High-Fat Diet-induced obese female C57BL/6 mice were intraperitoneally injected with SFN (10 mg/kg) daily. Body weight was recorded every 3 days. 30 days later, glucose tolerance test (GTT) and insulin tolerance test (ITT) were performed. At the end of experiment, fat mass were measured and the adipogenesis as well as browning associated genes expression in white adipose tissue (WAT) were determined by RT-qPCR and western blot. Histological examination of the adipose tissue samples were carried out with hematoxylin-eosin (HE) staining and immunofluorescence staining method. In vitro, pre-adipocytes C3H10T1/2 were treated with SFN to investigate the direct effects on adipogenesis. Results: SFN suppressed HFD-induced body weight gain and reduced the size of fat cells in mice. SFN suppressed the expression of key genes in adipogenesis, inhibited lipid accumulation in C3H10T1/2 cells, increased the expression of brown adipocyte-specific markers and mitochondrial biogenesis in vivo and in vitro, and decreased cellular and mitochondrial oxidative stress. These results suggested that SFN, as a nutritional factor, has great potential role in the battle against obesity by inducing the browning of white fat. Conclusion: SFN could significantly decrease the fat mass, and improve glucose metabolism and increase insulin sensitivity of HFD-induced obese mice by promoting the browning of white fat and enhancing the mitochondrial biogenesis in WAT. Our study proves that SFN could serve as a potential medicine in anti-obesity and related diseases.

Sulforaphane Inhibits Osteoclastogenesis via Suppression of the Autophagic Pathway.

Previous studies have demonstrated that sulforaphane (SFN) is a promising agent against osteoclastic bone destruction. However, the mechanism underlying its anti-osteoclastogenic activity is still unclear. Herein, for the first time, we explored the potential role of autophagy in SFN-mediated anti-osteoclastogenesis in vitro and in vivo. We established an osteoclastogenesis model using receptor activator of nuclear factor kappa-ß ligand (RANKL)-induced RAW264.7 cells and bone marrow macrophages (BMMs). Tartrate-resistant acid phosphatase (TRAP) staining showed the formation of osteoclasts. We observed autophagosomes by transmission electron microscopy (TEM). In vitro, we found that SFN inhibited osteoclastogenesis (number of osteoclasts: 22.67 ± 0.88 in the SFN (0) group vs. 20.33 ± 1.45 in the SFN (1 µM) group vs. 13.00 ± 1.00 in the SFN (2.5 µM) group vs. 6.66 ± 1.20 in the SFN (2.5 µM) group), decreased the number of autophagosomes, and suppressed the accumulation of several autophagic proteins in osteoclast precursors. The activation of autophagy by rapamycin (RAP) almost reversed the SFN-elicited anti-osteoclastogenesis (number of osteoclasts: 22.67 ± 0.88 in the control group vs. 13.00 ± 1.00 in the SFN group vs. 17.33 ± 0.33 in the SFN+RAP group). Furthermore, Western blot (WB) analysis revealed that SFN inhibited the phosphorylation of c-Jun N-terminal kinase (JNK). The JNK activator anisomycin significantly promoted autophagy, whereas the inhibitor SP600125 markedly suppressed autophagic activation in pre-osteoclasts. Microcomputed tomography (CT), immunohistochemistry (IHC), and immunofluorescence (IF) were used to analyze the results in vivo. Consistent with the in vitro results, we found that the administration of SFN could decrease the number of osteoclasts and the expression of autophagic light chain 3 (LC3) and protect against lipopolysaccharide (LPS)-induced calvarial erosion. Our findings highlight autophagy as a crucial mechanism of SFN-mediated anti-osteoclastogenesis and show that the JNK signaling pathway participates in this process.

Genome-wide analyses of non-syndromic cleft lip with palate identify 14 novel loci and genetic heterogeneity.

Non-syndromic cleft lip with palate (NSCLP) is the most serious sub-phenotype of non-syndromic orofacial clefts (NSOFC), which are the most common craniofacial birth defects in humans. Here we conduct a GWAS of NSCLP with multiple independent replications, totalling 7,404 NSOFC cases and 16,059 controls from several ethnicities, to identify new NSCLP risk loci, and explore the genetic heterogeneity between sub-phenotypes of NSOFC. We identify 41 SNPs within 26 loci that achieve genome-wide significance, 14 of which are novel (RAD54B, TMEM19, KRT18, WNT9B, GSC/DICER1, PTCH1, RPS26, OFCC1/TFAP2A, TAF1B, FGF10, MSX1, LINC00640, FGFR1 and SPRY1). These 26 loci collectively account for 10.94% of the heritability for NSCLP in Chinese population. We find evidence of genetic heterogeneity between the sub-phenotypes of NSOFC and among different populations. This study substantially increases the number of genetic susceptibility loci for NSCLP and provides important insights into the genetic aetiology of this common craniofacial malformation.

MYBL2 guides autophagy suppressor VDAC2 in the developing ovary to inhibit autophagy through a complex of VDAC2-BECN1-BCL2L1 in mammals.

Oogenesis is essential for female gamete production in mammals. The total number of ovarian follicles is determined early in life and production of ovarian oocytes is thought to stop during the lifetime. However, the molecular mechanisms underling oogenesis, particularly autophagy regulation in the ovary, remain largely unknown. Here, we reveal an important MYBL2-VDAC2-BECN1-BCL2L1 pathway linking autophagy suppression in the developing ovary. The transcription factors GATA1 and MYBL2 can bind to and activate the Vdac2 promoter. MYBL2 regulates the spatiotemporal expression of VDAC2 in the developing ovary. Strikingly, in the VDAC2 transgenic pigs (Sus scrofa/Ss), VDAC2 exerts its function by inhibiting autophagy in the ovary. In contrast, Vdac2 knockout promotes autophagy. Moreover, VDAC2-mediated autophagy suppression is dependent on its interactions with both BECN1 and BCL2L1 to stabilize the BECN1 and BCL2L1 complex, suggesting VDAC2 as an autophagy suppressor in the pathway. Our findings provide a functional connection among the VDAC2, MYBL2, the BECN1-BCL2L1 pathway and autophagy suppression in the developing ovary, which is implicated in improving female fecundity.

Loss-of-function mutation in the X-linked TBX22 promoter disrupts an ETS-1 binding site and leads to cleft palate.

The cleft palate only (CPO) is a common congenital defect with complex etiology in humans. The molecular etiology of the CPO remains unknown. Here, we report a loss-of-function mutation in X-linked TBX22 gene (T-box 22) in a six-generation family of the CPO with obvious phenotypes of both cleft palate and hyper-nasal speech. We identify a functional -73G>A mutation in the promoter of TBX22, which is located at the core-binding site of transcription factor ETS-1 (v-ets avian erythroblastosis virus E26 oncogene homolog 1). Phylogenetic analysis showed that the sequence around the -73G>A mutation site is specific in primates. The mutation was detected in all five affected male members cosegregating with the affected phenotype and heterozygote occurred only in some unaffected females of the family, suggesting an X-linked transmission of the mutation in the family. The -73G>A variant is a novel single nucleotide mutation. Cell co-transfections indicated that ETS-1 could activate the TBX22 promoter. Moreover, EMSA and ChIP assays demonstrated that the allele A disrupts the binding site of ETS-1, thus markedly decreases the activity of the TBX22 promoter, which is likely to lead to the birth defect of the CPO without ankyloglossia. These results suggest that a loss-of-function mutation in the X-linked TBX22 promoter may cause the cleft palate through disruption of TBX22-ETS-1 pathway.

DNA demethylation and USF regulate the meiosis-specific expression of the mouse Miwi.

Miwi, a member of the Argonaute family, is required for initiating spermiogenesis; however, the mechanisms that regulate the expression of the Miwi gene remain unknown. By mutation analysis and transgenic models, we identified a 303 bp proximal promoter region of the mouse Miwi gene, which controls specific expression from midpachytene spermatocytes to round spermatids during meiosis. We characterized the binding sites of transcription factors NF-Y (Nuclear Factor Y) and USF (Upstream Stimulatory Factor) within the core promoter and found that both factors specifically bind to and activate the Miwi promoter. Methylation profiling of three CpG islands within the proximal promoter reveals a markedly inverse correlation between the methylation status of the CpG islands and germ cell type-specific expression of Miwi. CpG methylation at the USF-binding site within the E2 box in the promoter inhibits the binding of USF. Transgenic Miwi-EGFP and endogenous Miwi reveal a subcellular co-localization pattern in the germ cells of the Miwi-EGFP transgenic mouse. Furthermore, the DNA methylation profile of the Miwi promoter-driven transgene is consistent with that of the endogenous Miwi promoter, indicating that Miwi transgene is epigenetically modified through methylation in vivo to ensure its spatio-temporal expression. Our findings suggest that USF controls Miwi expression from midpachytene spermatocytes to round spermatids through methylation-mediated regulation. This work identifies an epigenetic regulation mechanism for the spatio-temporal expression of mouse Miwi during spermatogenesis.