CA1 Modulates the Osteogenic Differentiation of Dental Follicle Stem Cells by Activating the BMP Signaling Pathway In Vitro.

BACKGROUND: Carbonic anhydrase 1 (CA1) has been found to be involved in osteogenesis and osteoclast in various human diseases, but the molecular mechanisms are not completely understood. In this study, we aim to use siRNA and lentivirus to reduce or increase the expression of CA1 in Dental follicle stem cells (DFSCs), in order to further elucidate the role and mechanism of CA1 in osteogenesis, and provide better osteogenic growth factors and stem cell selection for the application of bone tissue engineering in alveolar bone fracture transplantation. METHODS: The study used RNA interference and lentiviral vectors to manipulate the expression of the CA1 gene in DFSCs during in vitro osteogenic induction. The expression of osteogenic marker genes was evaluated and changes in CA1, alkaline phosphatase (ALP), Runt-related transcription factor 2 (RUNX2), and Bone morphogenetic proteins (BMP2) were measured using quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting (WB). The osteogenic effect was assessed through Alizarin Red staining. RESULTS: The mRNA and protein expression levels of CA1, ALP, RUNX2, and BMP2 decreased distinctly in the si-CA1 group than other groups (p < 0.05). In the Lentivirus-CA1 (LV-CA1) group, the mRNA and protein expressions of CA1, ALP, RUNX2, and BMP2 were amplified to varying degrees than other groups (p < 0.05). Apart from CA1, BMP2 (43.01%) and ALP (36.69%) showed significant upregulation (p < 0.05). Alizarin red staining indicated that the LV-CA1 group produced more calcified nodules than other groups, with a higher optical density (p < 0.05), and the osteogenic effect was superior. CONCLUSIONS: CA1 can impact osteogenic differentiation via BMP related signaling pathways, positioning itself upstream in osteogenic signaling pathways, and closely linked to osteoblast calcification and ossification processes.

Chemical constituents from the stem of Ficus pumila.

A new biphenyl compound: 5-aldehyde-4'-hydroxy-2,2'-dimethoxybiphenyl (1), along with 11 known phenolic compounds (2-12) were isolated from the stem of Ficus pumila Linn. Their structures were established by spectroscopic methods, including MS, UV, IR, 1 D and 2 D NMR. Compounds 4 and 10 showed weakly cytotoxicity against MCF-7, with IC50 values of 32.5 and 37.3 µM, respectively.

Inhibition of p38 mitogen-activated protein kinases may attenuate scar proliferation after cleft lip surgery in rabbits via Smads signaling pathway.

BACKGROUND: Cleft lip repair surgery always results in visible scarring. It has been proved that scar formation can be reduced by inhibiting the p38 mitogen-activated protein kinases (p38MAPKs) signaling pathway. However, the interaction between p38MAPK and Smads in scar formation is still controversial. METHODS: This study was designed to investigate whether inhibition of p38MAPK reduces postoperative scar formation of cleft lips on rabbits via the Smads signaling pathway. Scar models in rabbits after cleft lip surgery were created and their fibroblasts were extracted. Then the expression of p38MAPK was disturbed by adenovirus in vitro and Vivo. The scar thickness was measured and scar tissues were excised for Sirius red staining and immunohistochemistry to detect the expression of type I collagen (col I), type III collagen (col III), and α-smooth muscle actin (α-SMA). The underlying mechanisms of p38MAPK knockdown on the extracellular matrix and Smad signaling pathway were invested in vitro using the EdU assay, Western blot, RT PCR, and immunofluorescence. RESULTS: p38MAPK knockdown suppresses the expression of p-smad3 and p-smad2 in fibroblasts, modulating the expression of its target genes, such as α-SMA, col I, and col III. When Ad-P38MAPK-1 was injected into lip scar, it reduced the expression of scar-related genes and scar thickness when compared to the negative control groups. CONCLUSIONS: In rabbits, inhibiting p38MAPK expression prevents scar proliferation through inhibiting the Smad signaling pathway after cleft lip surgery.

p38MAPK silencing attenuates scar proliferation after cleft palate repair surgery in rats via MRTF-A/SRF pathway.

Scarring is the primary factor of maxilla growth restriction among people who have undergone cleft palate repair surgery. p38 mitogen-activated protein kinase (p38MAPK) promotes fibrosis in a variety of organs. However, its role in post-surgery scarring on the hard palate has not been fully understood. This study is designed to investigate the role of p38MAPK in scar formation and maxilla growth of rats. We removed the mucosa on the hard palate of rats and applied the p38MAPK silencing adenovirus vector on it two weeks after surgery. Then the scarring tissue and maxilla growth were evaluated by histological and morphological examination. The effect of p38MAPK silencing on scarring-related genes in fibroblasts was also studied. We found that local injection of Ad-p38MAPK-1 in vivo effectively reduces the expression of p38MAPK and scarring-related proteins and weakens the impact of scarring on the width of the hard palate. Mechanistically, p38MAPK silencing inhibits the expression of α-smooth muscle actin (α-SMA) via mediating the production and nuclear localization of myocardin-related transcription factor A (MRTF-A) in fibroblasts. These results reveal a molecular pathway of scar formation involving p38MAPK/MRTF-A stimulation and support targeting p38MAPK as a potentially effective treatment for post-surgery scarring on the hard palate.

Effect of a digital guide on the positional accuracy of intermaxillary fixation screw implantation in orthognathic surgery.

BACKGROUND: Intermaxillary fixation screw (IMFS) implantation is a common procedure in orthognathic surgery (OGS) performed to the temporary maxillary-mandibular fixation and stable bite relationships. The study aims to assess the accuracy of IMFS implantation with a digital guide to reduce the occurrence of root damage. METHODS: This prospective study involved 40 patients undergoing OGS at the Affiliated Hospital of Qingdao University from August 2017 to May 2021. The patients were randomly divided into two groups according to whether the IMFS implantation was with or without digital guide (20 patients in the experimental group and 20 controls). The digital guides used in the experimental group were designed according to a virtual implantation plan and printed using stereolithography. In the control group, IMFSs were directly implanted by a surgeon based on clinical experience. Postoperatively, cone-beam computed tomography was performed to compare root proximity of IMFSs between the two groups and verify the accuracy of IMFS placement. RESULTS: In the experimental group, there was no case of root damage, the incidence of the periodontal ligament (PDL) injured was 22.1%, and 77.9% IMFSs were placed without contacting adjacent anatomic structures. In the control group, the incidence of root damage had been up to 20.8%, 31.7% IMFSs injured the PDL, and only 47.5% IMFSs were placed between the roots (P < 0.001). CONCLUSION: IMFSs can be placed more accurately with surgical guides, reducing the incidence of root and PDL damages.

Clinical study on obstructive sleep apnea following pharyngeal flap surgery.

OBJECTIVES: This study aims to investigate the incidence and severity of obstructive sleep apnea (OSA) in cleft patients with velopharyngeal insufficiency (VPI) after pharyngeal flap surgery (PFS) and explore the influence of operation age. METHODS: A retrospective study was conducted in 82 cleft patients after PFS. The patients were divided into two groups according to their age at the time of surgery. The incidence and severity of OSA were assessed at least 1.2 years (mean 6.0 years) postoperatively by polysomnography (PSG). RESULTS: The incidence rates of OSA were 20% in the adult group and 31% in the child group. No significant difference was found between the two groups (P=0.289). Patients with OSA in the adult and child groups were classified into different levels of severity (mild, moderate, severe) according to the apnea hypoventilation index (AHI). No statistically significant difference in the severity of OSA was found between the two groups (P=0.079). CONCLUSIONS: Some patients still have OSA average of 6.0 years after PFS, and operation ageis unrelated to the incidence and severity of OSA.

Virtual Surgical Planning and Three-Dimensional Printing to Aid the Anatomical Reduction of an Old Malunited Fracture of the Mandible.

ABSTRACT: Mandible fracture is a common injury in maxillofacial surgery. It causes not only maxillofacial dysfunction but also facial deformities. Malunited fractures of the mandible have been a vast challenge in clinical treatment due to the misalignment of the broken ends and the occurrence of occlusal disorders. This case report describes using virtual surgical planning and three-dimensional printing to treat a patient with malunited fracture of the mandible. Failing to perform mandibular surgery due to severe brain trauma after the car accident, the patient got malunited healing of mandible. The authors applied virtual surgical planning to perform preoperative analysis and surgical design on this patient, three-dimensional printing to fabricate occlusal plate, and models of the preoperative and postoperative mandible to guide the operation. Finally, the authors achieved the reduction and reconstruction of the mandible with satisfactory clinical results.

Treatment of oral lichen planus by surgical excision and acellular dermal matrix grafting: Eleven case reports and review of literature.

BACKGROUND: Oral lichen planus (OLP) is a chronic inflammatory disorder, and it can affect normal oral function. The conventional treatments for OLP are not always effective, and relapse easily occurs. Therefore, treatment of OLP is difficult and challenging. In this study, we evaluated over a long period the clinical efficacy of surgical excision and acellular dermal matrix (ADM) grafting in patients with refractory OLP. CASE SUMMARY: Eleven patients with refractory OLP underwent a standardized protocol of surgical excision and ADM grafting. The condition of the area of the grafted wound, the intraoperative maximum mouth opening, pain, and clinical healing were assessed at postoperative follow-up visits. All patients had a flat surgical area with similar mucosal tissue coverage and local scar formation. Patients had no irritation and pain in their mucous membranes when eating acidic and spicy food. All patients' mouth openings returned to normal within 2-6 mo after surgery. During follow-up, none of the patients had recurrence of OLP after surgery. The longest follow-up was 11 yr and the shortest was 6 mo, and none of the patients relapsed during follow-up. CONCLUSION: Surgical excision and ADM grafting could be an effective method to treat refractory OLP.

Outcomes of bone morphogenetic protein-2 and iliac cancellous bone transplantation on alveolar cleft bone grafting: A meta-analysis.

BACKGROUND: To systematically evaluate the effect of bone morphogenetic protein-2 (BMP-2) and iliac cancellous bone graft (ICBG) on alveolar cleft bone grafting (ACBG) in cleft lip and palate. METHOD: Online databases were searched for case-control studies related to the application of BMP-2 and ICBG in ACBG. RESULT: Meta-analysis showed no significant statistical difference in the filling rate (OR = 4.1, 95% CI (0.06, 2.63)), the volume of bone graft area (OR=-0.42, 95% CI (-1.44, 0.60)), the height of bone graft area (OR = -21.38, 95% CI (-23.00, -19.76)), the density of bone graft area (OR = 0.43, 95% CI (-0.79, 1.64)), the failure rate of bone graft (OR = 0.02, 95% CI (-0.03, 0.06)), infection after operation, and the rate (OR = 0.20, 95% CI (0.05, 0.73)) and the incidence of postoperative oronasal fistula (OR = 4.1, 95% CI (0.06, 2.63)) between BMP-2 and ICBG in ACBG. However, there were obvious statistical differences in operative time (OR = -3.64, 95% CI (-7.35, 0.06)) and the length of hospital stay (OR = -1.97, 95% CI (-2.41, -1.53)). CONCLUSION: The meta-analysis shows that there is no significant difference between BMP-2 and ICBG in filling rate, volume, density, failure rate, and the occurrence of oronasal fistula after ACBG. There were significant differences between BMP-2 and ICBG in the operation time and hospitalization time of ACBG. Compared with ICBG bone graft, BMP-2 has more advantages in ACBG such as remaining area height, postoperative infection rate, operative time, and length of hospital stay.

[Association between cervical vertebral maturation stages and dental calcification stages in patients with unilateral complete cleft lip and palate].

OBJECTIVE: The purpose of the study is to investigate the relationship between dental calcification stages (DCS) and cervical vertebral maturation stages (CVMS) in patients with unilateral complete cleft lips and palates (UCLP) and to provide a theoretical basis for the treatment time selection of cleft lip and palate (CLP) patients. METHODS: A total of 123 UCLP patients and 215 non-CLP subjects were selected. The DCS of the left mandibular canine, premolar, and second molar in non-CLP subjects and on both cleft sides of UCLP patients were assessed utilizing the Demirjian method. CVMS was observed utilizing the Baccetti method. The results were analyzed by Spearman rank correlation, and the correlation coefficients were compared. RESULTS: There was a correlation between the CVMS and the DCS of the left mandibular canine, the first premolar, the second premolar, and the second molar in the non-CLP subjects and on both cleft sides of the UCLP patients (r=0.762-0.864, P<0.05; r=0.809-0.914, P<0.05, respectively). The correlation between the CVMS and the DCS of the mandibular first premolar was highest among the UCLP patients. Except for the first and the second premolars of UCLP females, the correlation between the DCS and the CVMS of the other teeth did not differ among the non-CLP subjects (P>0.05). CONCLUSIONS: DCS can be utilized as a biological index to determine the growth development statuses. The correlation between the CVMS and the DCS of the mandibular first premolar was the highest.

Longitudinal Study of Scar Hyperplasia Formation Following Cleft Lip Wound Healing.

The purpose of this study was to observe the hyperplasia trend of scar after the cleft lip surgery in a rabbit animal model, and determine the time-point of the highest hypertrophic degree of scar after cleft lip repair. Forty New Zealand white rabbits from the same offspring were used to establish a cleft lip wound healing model using Millard surgery procedure. The scar volumes were measured and granulation tissues were observed visually in the 2, 3, 4, and 5 weeks after operation. The scar tissues were harvested at the indicated time-points. Immunohistochemical (IHC) and Western Blot analyses were performed to detect the expression level of α-smooth muscle actin (α-SMA) in the scar tissue. The scars shrunk and the volumes reduced at 3 to 4 weeks after surgery; however, at 5 weeks postsurgery, the volumes increased. IHC and Western blot analyses indicated the expression of α-SMA was significantly enhanced 3 to 4 weeks, but reduced in the 5 weeks after surgery. Overall, the degree of scar hyperplasia after cleft lip surgery in rabbits was normally distributed and the scarring was most severe in the 3 to 4 weeks after cleft lip surgery. The study confirms a novel animal model for the assessment of therapies for the treatment of scar hyperplasia of human cleft lip in future.

IRF6 is the mediator of TGFß3 during regulation of the epithelial mesenchymal transition and palatal fusion.

Mutation in interferon regulatory factor 6 (IRF6) is known to cause syndromic and non-syndromic cleft lip/palate in human. In this study, we investigated the molecular mechanisms related to IRF6 during palatal fusion using palatal shelves organ culture. The results showed that ablation of Irf6 resulted in a delay in TGFß3-regulated palatal fusion. Ectopic expression of IRF6 was able to promote palatal fusion and rescue shTgfß3-induced fusion defect. These findings indicate that IRF6 is involved in TGFß3-mediated palatal fusion. Molecular analysis revealed that ectopic expression of IRF6 increased the expression of SNAI2, an epithelial mesenchymal transition (EMT) regulator, and diminished the expression of various epithelial markers, such as E-cadherin, Plakophilin and ZO-1. In addition, knockdown of Irf6 expression decreased SNAI2 expression, and restored the expression of ZO-1 and Plakophilin that were diminished by TGFß3. Blocking of Snai2 expression delayed palatal fusion and abolished the IRF6 rescuing effect associated with shTgfß3-induced fusion defect. These findings indicate that TGFß3 increases IRF6 expression and subsequently regulates SNAI2 expression, and IRF6 appears to regulate EMT during palatal fusion via SNAI2. Taken together, this study demonstrates that IRF6 is a mediator of TGFß3, which regulates EMT and fusion process during the embryonic palate development.

Evaluation of Bone Height and Bone Mineral Density Using Cone Beam Computed Tomography After Secondary Bone Graft in Alveolar Cleft.

The purpose of this study was to assess the optimal timing of subsequent dental implant placement and orthodontics after alveolar bone grafting (ABG) in patients with unilateral complete clefts of the alveolar process. Iliac bone graft surgery was performed on 60 patients. Bone mineral density (BMD) and height of the ABG areas were assessed using cone beam computed tomography at 3 and 6 months postoperatively. The heights of the labial and palatal bone graft areas were classified using the modified Bergland classification. The study found that there was no change in BMD between 3 months (mean ±â€ŠSD: 406.51 ±â€Š71.28 Hounsfield units [HU]) and 6 months (409.53 ±â€Š46.37 HU; P = 0.381). Significant changes in the distribution of bone height classifications were observed in the labial and palatal sides of the ABG between 3 and 6 months (P = 0.025 for labial bone height, P = 0.008 for palatal bone height). These results indicate that the alveolar density remained stable between 3 and 6 months, whereas bone height level declined during that period after ABG, the latter indicating bone graft absorption over time. It is, therefore, suggested that subsequent orthodontic or dental implants be placed 3 months after ABG rather than at 6 months or later.

Intermittent Stretching and Osteogenic Differentiation of Bone Marrow Derived Mesenchymal Stem Cells via the p38MAPK-Osterix Signaling Pathway.

AIMS: The relationship between the p38MAPK signaling pathway and osterix in osteogenic differentiation of BMMSCs subjected to intermittent stretching was investigated. METHODS: BMMSCs derived from C57BL/6J mice were divided into the following groups: 1) control, 2) stretch, and 3) SB203580+stretch (SB203580 is a p38MAPK signal pathway inhibitor). BMMSCs were exposed to an intermittent mechanical strain of 0.8% (8000µ strain) at 0.5 Hz, twice a day for 30 min each application. BMMSCs were harvested on days 1, 3, and 5 post-treatment. The expression of ALP, COL I, OCN, and osterix mRNA was assessed utilizing RT-PCR while the expression of P-p38MAPK and osterix protein was assessed by Western blot analysis. The osterix gene in mouse BMMSCs was knocked down using RNAi technology and its protein expression was also assessed by Western blot. RT-PCR was used to detect ALP, COL I, and OCN mRNA expression. RESULTS: Intermittent stretching was found to promote expression of ALP, COL I, OCN, and osterix mRNA. Silencing the osterix gene was found to reduce levels of ALP, COL I, and OCN mRNA. Western blot analysis demonstrated that the levels of osterix and P-p38MAPK proteins in the stretch group were significantly higher than in the control group (P<0.05). There was less expression of ALP, COL I, OCN, and osterix mRNA in the SB203580+stretch group than in the control and stretch groups. CONCLUSIONS: Data demonstrate that intermittent stretching promotes osteogenic differentiation of BMMSCs, and the p38MAPK-osterix pathway has an important role in the control of osteogenesis-related gene expression.

[Preliminary report of botulinum toxin type A injection at trigger point for treatment of trigeminal neuralgia: experiences of 16 cases].

PURPOSE: To investigate the effects of injection of botulium toxin type A at trigger point for treatment of patients with primary trigeminal neuralgia. METHODS: Sixteen patients with primary Trigeminal Neuralgia were treated with injection of botulium toxin type A. Visual analog scores(VAS) at 1 week, 2 weeks, 1 month, 3 months and 6 months after treatment and Barrow Neurological Institute (BNI) pain evaluation criteria were utilized to measure the degree of pain. The data was analyzed with SPSS 10.0 software package. RESULTS: The VAS score was 9.12±0.65 before botulium toxin type A injection while the scores were 2.8±1.36, 2.2±1.26, 1.3±1.45, 1.3±1.45 and 1.2±2.52 at 1 week, 2 weeks, 1 month,3 months and 6 months after treatment. There was significant difference in VAS compared with before treatment. VAS score was lower and stable at 1 month, 3 months and 6 months after treatment, but no significant difference was found at 1-week and 2-week after treatment. BNI evaluation results showed good therapeutic effect 1 week after treatment, while the best therapeutic effect was noted 1-3 months after treatment. 6 months later, 1 patient had recurrence and 11 patients had complete relief of pain. CONCLUSIONS: Botulium toxin type A injection is an effective way for treatment of patients with primary trigeminal neuralgia.

Soft tissue perineurioma of the tongue: report of a case and review of the literature.

Perineurioma is a rare benign tumor of the peripheral nervous system distinct from schwannomas and neurofibromas. It may be intraneural or extraneural (in the soft tissue). Extraneural soft tissue perineuriomas are uncommon; rare cases have been reported in the oral cavity. We present a case of soft tissue perineurioma in the tip of the tongue. The tumor was characterized by slender spindle cells, arranged in short fascicles or whorls, and focal areas showing a distinct storiform pattern. Tumor cells showed the immunohistochemical profile of perineurial cells, including epithelial membrane antigen. Smooth muscle actin, S-100, and CD34 were not expressed by the tumor cells. The tumor was surgically excised and in 2 years there has been no recurrence. Knowledge of the tumor in the oral cavity is important to reach a correct diagnosis and to avoid unnecessary aggressive local excision.

[Preliminary study of palatal lengthening by levator veli palatini retropositioning according to Sommerlad palatoplasty].

PURPOSE: To research whether palate lengthening can be achieved after palatoplasty with levator veli palatini retropositioning according to Sommerlad palatoplasty. METHODS: Sixty-five cases with incomplete cleft palate were selected randomly, with the age ranged from 10 to 13 months. Sommerlad palatoplasty was performed by the same surgeon with operating loupes. A paper ruler was used to measure the straight-line and the curved distance of palatal length starting from the meeting point between the center of the 2 medial incisor teeth and the lingual gingiva up to the uvular tip while the patient was under general anesthesia before and immediately after palatoplasty. The data was analyzed with SPSS 19.0 software package. RESULTS: Through measurements, the straight-line distance of the palate was about (44.24±0.76) mm, and the curve-line distance of the palate was about (53.11±0.74) mm before Sommerlad palatoplasty. The straight-line distance of the palate was (48.81±0.72) mm, and the curve-line distance of the palate was (59.41±0.8) mm after Sommerlad palatoplasty. The straight-line elongation was about (4.56±0.27) mm, with the mean elongation ratio of 10.31% (P<0.01). The curve-line elongation was about (6.30±0.43) mm, with the mean elongation ratio of 11.86% (P<0.01). CONCLUSIONS: Significant elongation in palate length is obtained after Sommerlad palatoplasty, which may be beneficial to recover the patients' normal speech and improve velopharyngeal competence.

[Establishment of palatal organ culture model of C57BL/6J mouse embryos in vitro].

PURPOSE: To establish palatal organ culture model of C57BL/6J mouse embryos in vitro and provide platform for study of embryo palatal development. METHODS: The mouse palatal shelves were harvested under sterilization from a female mouse of gestation day(GD) 13.5 by stereoscopic microscope and cultured in vitro. Totally 36 pairs of palatal shelves were divided into three groups equally and cultured 6 h, 24 h and 48 h, respectively. Finally, all palatal shelves were embedded and stained by Hematoxylin and Eosin (HE) and subjected to scanning electron microscope (SEM) analysis. RESULTS: The results of HE dyeing showed that the palatal shelves did not fuse on 6 h group, and began to fuse on 24 h group, but still had some medial edge epithelial (MEE) cells remained. The palatal shelves completely fused and all the MEE cells disappeared on 48 h group. The results of SEM showed that there was a gap between palatal shelves on 6 h group. The palatal shelves began to contact and form the medial epithelial seam (MES) on 12 h group. Finally, palatal shelves completely fused and MES disappeared on 48 h group. CONCLUSION: This method provides an effective way for investigating the etiology of cleft palate in vitro.

[Parotid major duct cyst: a case report].

Parotid gland cyst can be congenital or acquired, which is rarely seen in clinic. Cysts in the main duct of the parotid gland are especially rare. We reported a 37-year-old man who presented with a painless mass on the right cheek. Superficial parotidectomy was performed and histopathologic examination of the specimen revealed a main parotid duct cyst. The clinical features, diagnosis, and treatment of the case were discussed and the relevant literatures were reviewed.