Buccal Mucosal Tissue Research Articles

STRO-1 confers myofibroblast transdifferentiation in fibroblasts derived from oral submucous fibrosis.

STRO-1 is a mesenchymal cell marker present on all clonogenic stromal precursors. Current evidence has indicated that the pathogenesis of fibrotic changes may be mediated by stemness properties. The aim of this study was to investigate the role of STRO-1 in areca quid chewing-associated oral submucous fibrosis (OSF). Thirty OSF specimens and ten normal buccal mucosae were examined by immunohistochemistry. The activity of STRO-1 from fibroblasts cultured from normal buccal mucosa (BMFs) and OSF (OSFFs) were measureed and the effect of arecoline, a major areca nut alkaloid, on STRO-1 in BMFs was evaluated. Compared the activities between sorted STRO-1+ cells and STRO-1- cells from OSFF were measured by collagen gel contraction, migration, invasion abilities, and the expression of α-smooth muscle actin (α-SMA) and pro-α1 (I) chain of type I collagen. Our results first showed that the expression of STRO-1 was more evident in areca quid chewing-associated OSF than normal buccal mucosa tissues (P < .05). Arecoline dose-dependently activated the level of STRO-1 in BMFs (P < .05). The relative expression of STRO-1 was significantly higher in OSFFs compared with BMFs (P < .05). In addition, the sorted STRO-1+ cells from OSFFs exhibited higher collagen gel contraction, migration, and invasion abilities as well as elevated expression of α-SMA and pro-α1 (I) chain of type I collagen than the negative subset (P < .05). These findings suggested that the stemness marker STRO-1 may be a crucial factor in the pathogenesis of areca quid chewing-associated OSF.

Use of oral mucosal cell sheets for accelerated oral surgical wound healing.

We developed a highly efficient in vitro-engineered mucosa equivalent using completely autologous mucosa and blood and investigated its feasibility and efficacy for oral surgical wound healing. Small oral mucosa samples were obtained from surgical patients, and keratinocytes and fibroblasts were primarily grown in media without animal products for generating 3D cell sheets. Morphological characteristics of the cell sheets were comparable to those of human mucosa, although p63-positive cells were more numerous in cell sheets. In addition, cell sheets were flexible, expandable, and easy to handle or transfer. In further in vivo rat experiments with deep wounding of the buccal mucosa and soft tissues, controls had significantly thinner epithelium and thicker collagen densities than those with cell sheets. Autologous cell sheets can be engineered in vitro from oral keratinocytes, fibroblasts, and fibrin, and can be used clinically to accelerate healing of oral soft tissue defects.

Chemopreventive effect of syringic acid on 7,12-dimethylbenz(a)anthracene induced hamster buccal pouch carcinogenesis

Oral squamous cell carcinoma is most prevalent and refractory cancers worldwide. Recently, chemoprevention could be a promising approach in developing countries. The present study investigates the mechanism of syringic acid (SA), a phenolic constituent of plant Alpiniacalcarata Roscoe, and their leaves are used as traditional Indian Ayurveda medicines, mediated chemoprevention on 7,12-dimethylbenz(a)anthracene (DMBA)-induced hamster buccal pouch carcinogenesis (HBPC). Lipid peroxidation and antioxidants were measured in the plasma and buccal tissues in experimental hamsters. Modulating effect of SA on the expression pattern of PCNA, Cyclin D1, and mutant p53 markers was used for immunoexpression and western blotting analysis. In the present study, 100% tumor formation with marked abnormalities in the biochemical parameters of lipid peroxidation and antioxidants through up-regulation of molecular markers like PCNA, Cyclin D1, and mutant p53 was accompanied with tumor-bearing hamsters. Oral administration of SA at the doses of 50 and 100 mg/kg body weight (bw) to DMBA-treated hamsters significantly inhibited adverse changes in the biochemical parameters of the plasma and buccal mucosal tissues and also down-regulation of molecular marker expression (PCNA, Cyclin D1, and mutant p53). The present study thus suggests that SA has potent anti-lipid peroxidative, antioxidant, anti-cell proliferative, and apoptosis-inducing properties during DMBA-induced HBPC.

The In Vivo Effect of Ytterbium-Doped Fiber Laser on Rat Buccal Mucosa as a Simulation of Its Effect on the Urinary Tract: A Preclinical Histopathological Evaluation.

PurposeThe aim of this study was to perform a histological analysis of the effect of a ytterbium-doped fiber (YDF) laser on oral buccal mucosa tissue in vivo to simulate its effect on the mucosa of the lower urinary tract.MethodsA total of 90 8-week-old Sprague-Dawley rats were anesthetized with urethrane (1.2 g/kg intraperitoneally). A prespecified inner buccal mucosal site was irradiated with a YDF master-oscillator power amplifier (MOPA) system for 60 seconds, with output power settings of 0.5, 1, and 2 W, respectively, in 3 treatment groups. Specimens of irradiated tissue were harvested at 2 hours, 24 hours, 2 weeks, and 4 weeks after irradiation. The tissue specimens were stained with hematoxylin and eosin for histological analysis. ResultsIn the group treated with 0.5 W, basal cell elongation and vacuolization were observed at 2 hours and 24 hours after treatment, respectively. No evident injury was observed after 2 or 4 weeks. The group treated with 1 W presented partial basal layer separation, and even complete epidermal ablation, within 2 hours. At 24 hours after laser treatment, new capillaries on an edematous background of fibroblasts and myofibroblasts, as well as profuse infiltration of the neutrophils to the basal layer, were observed. Collagen deposition and reepithelization were observed in specimens taken 2 weeks and 4 weeks after treatment. The group treated with 2 W presented bigger and deeper injuries at 2 hours after irradiation. Meanwhile, subepidermal bullae with full-thickness epidermal necrosis and underlying inflammatory infiltrate were observed 24 hours after treatment. The presence of fibrous connective tissue and collagen deposition were observed 2 weeks and 4 weeks after the treatment.ConclusionsTo our knowledge, this is the first report regarding the effect of a YDF laser on living tissue. Our study demonstrated that the typical histological findings of the tissue reaction to the YDF MOPA apparatus were very similar to those associated with thermal injuries. The extent and degree of tissue damage increased proportionally to the output power.

Assessment of Lipid Peroxidation and Antioxidant Status in Vanillic Acid Treated 7,12-Dimethylbenz[a]anthracene Induced Hamster Buccal Pouch Carcinogenesis.

Vanillic acid, a naturally occurring bioactive substance, possesses diverse pharmacological potential including free radical scavenging and anticancer properties. Excessive generation of Reactive Oxygen Species (ROS) and insufficient antioxidant potential has been involved in numerous pathological disorders, including cancer. To explore the anti-lipid peroxidative and antioxidant efficacy of vanillic acid in Dimethylbenz[a]Anthracene (DMBA) induced oral carcinogenesis. Topical application of DMBA for 14 weeks in the buccal pouch of hamsters resulted in well developed oral squamous cell carcinoma. Vanillic acid at a dosage of 200 mg/kg body weight was orally administrated to the hamsters for 14 weeks. The status of lipid peroxidation and antioxidants were measured in the plasma and buccal mucosa of hamsters using specific colorimetric methods. Altered levels of lipid peroxidation by-products {Thiobarbituric Acid Reactive Substances (TBARS)} and disturbances in antioxidants status {Superoxide Dismutase (SOD), Catalase (CAT), Glutathione Peroxidase (GPx), vitamin E, vitamin C and reduced Glutathione (GSH)} were observed in the plasma and buccal mucosa tissues of hamsters treated with DMBA alone. Vanillic acid (200 mg/kg bw p.o) significantly restored the above mentioned plasma and buccal mucosa biochemical variables to near normal range in DMBA treated hamsters. Present findings thus confirm the anti-lipid peroxidative and antioxidant efficacy of vanillic acid in DMBA induced hamster buccal pouch carcinogenesis.

Abstract 1631: FGF2 activation of FGFR1 in head and neck squamous cell carcinoma is associated with more invasive disease and can be attenuated by FGFR inhibition

Abstract Introduction. Head and neck squamous cell carcinomas (HNSCCs) account for nearly 600,000 deaths worldwide annually and have limited treatment options. Approximately 20% of HNSCCs harbor amplifications of fibroblast growth factor receptor 1 (FGFR1) on chromosome 8p, however FGFR1 amplification by itself does not predict clinical response to FGFR inhibitors. We hypothesized that FGF2, or basic FGF, ligand expression is a better marker of FGFR activation and predictor of response to FGFR inhibitors. Results. A tissue micro array (TMA) of HNSCC patient biopsies was stained and quantitated for FGF2 expression by Aperio ImageScope software. FGF2 was significantly increased in recurrent tissue samples (p = 0.04). We examined a number of immortalized HNSCC cell lines and found that overexpression of both FGF2 and FGFR1 predicted response to the selective FGFR inhibitor PD173074. FGFR inhibition did not cause apoptosis, but rather induced a G0/G1 arrest and growth inhibition. FGFR inhibition also induced a change in cell morphology, with a significant increase in cell size and adherence. The expression of epithelial-to-mesenchymal transition (EMT) proteins was examined and FGF2-FGFR1 activation was associated with a more mesenchymal phenotype. Accordingly, FGFR inhibition reversed invasiveness as measured using the Incucyte WoundMaker scratch assay, suggesting that HNSCCs with FGF2-FGFR1 activation have more metastatic potential. Invasiveness of these cells in vivo was confirmed using orthotopic injection into the buccal pad of NSG mice. Once primary tumors reached 0.8 cm in size, mice were sacrificed and buccal mucosa, lung, liver, and neck tissue were examined post-mortem. All of the injected animals developed local invasion, and distant metastases in the lungs. 5/7 mice also had metastases in the liver and this model is being used to test the ability of FGFR inhibition to prevent metastasis. The mechanism of autocrine FGF2-FGFR1 activation was further explored and FGF2 was found to be secreted in association with extracellular vesicles (ECVs). Interestingly, inhibition of FGFR reduced secretion of ECVs and FGF2, providing a novel approach to target autocrine and paracrine FGFR1 activation within the tumor. We further tested a number of small molecule inhibitors in combination with PD173074 to look for synergistic combinations of kinase inhibitors and found significant synergy between EGFR and FGFR inhibitors suggesting this combination may be most effective in patients with HNSCC. Conclusions. Increased FGF2 in HNSCC patient samples is correlated with recurrent disease. FGF2-FGFR1 activation increases invasiveness through activation of EMT genes both in vitro, and in an orthotopic model. Inhibition of FGF2-FGFR1 reversed the invasive phenotype in vitro and may be an effective therapeutic strategy to reduce metastases in HNSCC patients. Citation Format: Isabel A. English, Jacqueline Martinez, Edward El Rassi, Mark Schmidt, Ellen Langer, Sophia Bornstein, John Gleysteen, Melissa Wong, Brian Druker, Elie Traer. FGF2 activation of FGFR1 in head and neck squamous cell carcinoma is associated with more invasive disease and can be attenuated by FGFR inhibition. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1631.

Biointerfacial phenomena of amlodipine buccomucosal tablets of HPMC matrix system containing polyacrylate polymer/β-cyclodextrin: Correlation of swelling and drug delivery performance

ABSTRACTObjectives: This study focuses on the development of amlodipine bilayer buccal tablets of hydroxypropyl methylcellulose (HPMC) matrix system containing polyacrylate polymer (Carbopol®)/β-cyclodextrin as the drug layer and ethylcellulose as the non-swellable backing layer, and their biointerfacial phenomena.Methods: Tablets were evaluated for swelling, erosion and mucoadhesion using buccal mucosal tissue ex vivo. In vitro drug release and ex vivo drug transport across mucosal tissue were also performed in phosphate buffer (pH 6.8). The relationship of swelling with buccoadhesion and buccal permeation of various bilayer tablet formulations containing HPMC alone and in combination with Carbopol or drug-β-cyclodextrin complex has been prepared.Results: Overall buccoadhesion of the tablet with combination of HPMC and Carbopol was increased significantly compared with that of HPMC alone. Presence of cyclodextrin did not change bioadhesion force and swelling behavior significantly. Ex vivo permeation was increased with the increase of HPMC proportion in other formulations as observed in in vitro dissolution.Conclusion: Drug–cyclodextrin complexes in the tablet improved permeation due to its improved dissolution at the site of biointerface of tablet and buccomucosa. Correlations of ex vivo and in vitro data have been established to predict the buccomucosal permeation from the swelling index or drug release alone.

Do you mind if I vape? Immediate effects of electronic cigarettes on perfusion in buccal mucosal tissue - a pilot study

The association between smoking and postoperative complications is compounded in patients who have oral and maxillofacial operations by an additional local effect, and patients often continue to smoke after operation despite advice to stop. Recent studies have suggested that nicotine may reduce inflammation and improve angiogenesis, so topical application may be beneficial for smokers. The electronic cigarette is increasing in popularity and more patients ask whether they can vape after operation. We investigated the effect of electronic cigarettes (of which half contained nicotine and half did not) on blood flow in the buccal mucosa in 10 volunteers immediately after vaping. Smokers were excluded as this was considered an additional variable in a small pilot study and our Trust has a no-smoking policy. After vaping for 5minutes, capillary blood flow was measured in the buccal mucosa at 5-minute intervals using a laser Doppler probe, and the results were expressed as arbitrary perfusion units. There was a wide variation in results and a small but significant rise (p=0.008) as a result of nicotine vaping, but these fell to the same levels as before within 30minutes. Electronic cigarettes may have an effect on blood flow to the oral mucosa, although further studies are needed to show whether they improve healing time after operation. Additional work is also needed to compare them with cigarettes.

Bioinformatics Analysis of the Effects of Tobacco Smoke on Gene Expression.

This study was designed to explore the effects of tobacco smoke on gene expression through bioinformatics analyses. Gene expression profile GSE17913 was downloaded from the Gene Expression Omnibus database. The differentially expressed genes (DEGs) in buccal mucosa tissues between 39 active smokers and 40 never smokers were identified. Gene Ontology (GO) and pathway enrichment analyses of DEGs were performed, followed by protein-protein interaction (PPI) network, transcriptional regulatory network as well as miRNA-target regulatory network construction. In total, 88 up-regulated DEGs and 106 down-regulated DEGs were identified. Among these DEGs, cytochrome P450, family 1, subfamily A, polypeptide 1 (CYP1A1) and CYP1B1 were enriched in the Metabolism of xenobiotics by cytochrome P450 pathway. In the PPI network, tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein, zeta (YWHAZ), and CYP1A1 were hub genes. In the transcriptional regulatory network, transcription factors of MYC associated factor X (MAX) and upstream transcription factor 1 (USF1) regulated many overlapped DEGs. In addition, protein tyrosine phosphatase, receptor type, D (PTPRD) was regulated by multiple miRNAs in the miRNA-DEG regulatory network. CYP1A1, CYP1B1, YWHAZ and PTPRD, and TF of MAX and USF1 may have the potential to be used as biomarkers and therapeutic targets in tobacco smoke-related pathological changes.

Giant Panda (Ailuropoda melanoleuca) Buccal Mucosa Tissue as a Source of Multipotent Progenitor Cells.

Since the first mammal was cloned, the idea of using this technique to help endangered species has aroused considerable interest. However, several issues limit this possibility, including the relatively low success rate at every stage of the cloning process, and the dearth of usable tissues from these rare animals. iPS cells have been produced from cells from a number of rare mammalian species and this is the method of choice for strategies to improve cloning efficiency and create new gametes by directed differentiation. Nevertheless information about other stem cell/progenitor capabilities of cells from endangered species could prove important for future conservation approaches and adds to the knowledge base about cellular material that can be extremely limited. Multipotent progenitor cells, termed skin-derived precursor (SKP) cells, can be isolated directly from mammalian skin dermis, and human cheek tissue has also been shown to be a good source of SKP-like cells. Recently we showed that structures identical to SKPs termed m-SKPs could be obtained from monolayer/ two dimensional (2D) skin fibroblast cultures. Here we aimed to isolate m-SKPs from cultured cells of three endangered species; giant panda (Ailuropoda melanoleuca); red panda (Ailurus fulgens); and Asiatic lion (Panthera leo persica). m-SKP-like spheres were formed from the giant panda buccal mucosa fibroblasts; whereas dermal fibroblast (DF) cells cultured from abdominal skin of the other two species were unable to generate spheres. Under specific differentiation culture conditions giant panda spheres expressed neural, Schwann, adipogenic and osteogenic cell markers. Furthermore, these buccal mucosa derived spheres were shown to maintain expression of SKP markers: nestin, versican, fibronectin, and P75 and switch on expression of the stem cell marker ABCG2. These results demonstrate that giant panda cheek skin can be a useful source of m-SKP multipotent progenitors. At present lack of sample numbers means that we can only postulate why we were unable to obtain m-SKPs from the lion and red panda cultures. However the giant panda observations point to the value of archiving cells from rare species, and the possibilities for later progenitor cell derivation.

A new approach for treatment of precancerous lesions with curcumin solid–lipid nanoparticle-loaded gels: in vitro and clinical evaluation

Objective: Preparation and characterization of curcumin solid–lipid nanoparticle (CurSLN)-loaded mucoadhesive gel for local treatment of oral precancerous lesions with low dose.Methodology: The formulated CurSLNs were dispersed in a mucoadhesive gel matrix to be applied to the buccal mucosa. Conventional mucoadhesive gel using binary system was adopted. The prepared gels were evaluated for in vitro drug dialysis, ex vivo mucoadhesion test and ex vivo permeation study using chicken buccal mucosa. Short-term clinical evaluation was carried out on 10 patients suffering oral erythroplakia in terms of pain index and lesion size measurement.1Results: The results showed that the loaded gel with CurSLN showed good mucoadhesion property and 25 min in vivo residence time. In addition to stability enhancement for the Cur powder. All formulae did not show any drug permeated, however, significant amount of Cur was retained within the chicken buccal mucosal tissue confirmed by histological examination. Significant reduction in pain, and complete healing was observed after 6 weeks of treatment.Conclusion: The local use of Cur in low dose is a promising option for treatment of precancerous lesions. The lack of local anti-inflammatory compounds with reduced side effects intensifies the importance of studying natural products for this purpose.

Circadian rhythm variation of the clock genes Per1 and cell cycle related genes in different stages of carcinogenesis of buccal mucosa in animal model

To investigate the expression and circadian rhythm variation of biological clock gene Per1 and cell cycle genes p53, CyclinD1, cyclin-dependent kinases (CDK1), CyclinB1 in different stages of carcinogenesis in buccal mucosa and its relationship with the development of buccal mucosa carcinoma. Ninety golden hamsters were housed under 12 hours light-12 hours dark cycles, and the model of buccal squamous cell carcinoma was established by using the dimethylbenzanthracene (DMBA) to smear the golden hamster buccal mucosa. Before the DMBA was used and after DMBA was used 6 weeks and 14 weeks respectively, the golden hamsters were sacrificed at 6 different time points (5 rats per time point) within 24 hour, including 4, 8, 12, 16, 20 and 24 hour after lights onset (HALO), and the normal buccal mucosa, precancerous lesions and cancer tissue were obtained, respectively. HE stained sections were prepared to observe the canceration of each tissue. Real time RT-PCR was used to detect the mRNA expression of Per1, p53, CyclinD1, CDK1 and CyclinB1, and a cosine analysis method was applied to determine the circadian rhythm variation of Per1, p53, CyclinD1, CDK1 and CyclinB1 mRNA expression, which were characterized by median, amplitude and acrophase. The expression of Per1, p53, CDK1 and CyclinD1 mRNA in 6 different time points within 24 hours in the tissues of three different stages of carcinogenesis had circadian rhythm, respectively. However, the CyclinB1 mRNA was expressed with circadian rhythm just in normal and cancer tissue (P < 0.05), while in precancerous lesions the circadian rhythm was in disorder (P > 0.05). As the development of carcinoma, the median of Per1 and p53 mRNA expression were significantly decreased (P < 0.05), yet the median of CDK1, CyclinB1 and CyclinD1 mRNA expression were significantly increased (P < 0.05). The amplitude of Per1, p53 and CyclinD1 mRNA expression was significantly decreased as the development of carcinoma (P < 0.05), however the amplitude of CDK1 mRNA expression was significantly increased (P < 0.05). In addition, there was no significant difference in the amplitude of CyclinB1 mRNA expression. The time that the peak expression value of Per1 and CDK1 mRNA appeared (Acrophase) in precancerous lesions was remarkably earlier than that in normal tissues, but the acrophase of p53 and CyclinD1 mRNA expression was remarkably delayed. Moreover, the acrophase of CDK1 and CyclinB1 mRNA expression in cancer tissues was obviously earlier than that in normal tissues, yet there was no significant variation in acrophase of Per1, p53, CyclinD1 mRNA expression between normal tissues and cancer tissues. The circadian rhythm of clock gene Per1 and cell cycle genes p53, CyclinD1, CDK1, CyclinB1 expression remarkably varied with the occurrence and development of carcinoma. Further research into the interaction between circadian and cell cycle of two cycle activity and relationship with the carcinogenesis may providenew ideas and methods of individual treatment and the mechanism of carcinogenesis.

Pharmacokinetic and pharmacodynamic interactions between palifermin and heparin.

Oral mucositis, a severe complication during chemo- and/or radiotherapy, is prevented with palifermin treatment, a recombinant human keratinocyte growth factor (KGF/FGF-7). The FGF family belongs to the larger family of heparin-binding growth factors. Because it has been shown that heparin modulates binding of KGF to the KGF receptor and subsequently affects cellular proliferation induced by the KGF mitogenic signal, it is critical to understand the drug-drug interactions between palifermin and heparin, particularly because of heparin's narrow therapeutic margin. Two studies were performed in healthy subjects to characterize the effect of palifermin on the pharmacodynamics of heparin (activated partial thromboplastin time) and evaluate the impact of heparin on the pharmacokinetics and pharmacodynamics (Ki67 staining of buccal mucosal tissue) of palifermin. Results demonstrated a pronounced pharmacokinetic interaction; heparin coadministration increased the palifermin AUC 4- to 5-fold and decreased its half-life by 40%-45%, suggesting an approximate 70%-80% decrease in palifermin clearance and volume of distribution. These changes in the pharmacokinetics of palifermin during coadministration of heparin, however, did not affect the pharmacodynamic effect of palifermin, or the anticoagulant activity of heparin, and did not lead to increased safety findings. Therefore, these results suggest that dose adjustments for heparin and palifermin are not warranted when administered concurrently.

MiR-203 inhibits arecoline-induced epithelial-mesenchymal transition by regulating secreted frizzled-related protein 4 and transmembrane-4 L six family member 1 in oral submucous fibrosis

Oral submucous fibrosis (OSF) is a potentially malignant disease predominantly found in Asian people. The areca nut has been implicated in this disease. Arecoline, one of the areca alkaloids, induces epithelial-mesenchymal transition (EMT)-related factors in primary human buccal mucosal fibroblasts. Yet, the mechanisms of the underlying arecoline-induced EMT in OSF remain unknown. In the present study, we aimed to investigate the role of microRNAs (miRNAs) in arecoline-induced EMT in HaCaT cells. We found that miR-203 was significantly downregulated in OSF tissues compared to that in normal buccal mucosa tissues, and that miR-203 negatively regulated secreted frizzled-related protein 4 (SFRP4) and positively regulated transmembrane-4 L six family member 1 (TM4SF1). We observed that upregulation of miR-203 significantly decreased the cell proliferation of HaCaT cells, and significantly upregulated the expression of cytokeratin 19 (CK19) and E-cadherin proteins, whereas it significantly downregulated the expression of N-cadherin and vimentin compared to these levels in the vehicle control cells. Thus, we provide evidence to illustrate that miR-203 plays a role in the pathogenesis of OSF, which may be a target for OSF management.

Hypoxic regulation of plasminogen activator inhibitor-1 expression in human buccal mucosa fibroblasts stimulated with arecoline.

Oral submucous fibrosis (OSF) is regarded as a pre-cancerous condition with fibrosis in oral subepithelial connective tissue. Hypoxia-inducible factor (HIF)-1α regulates a wide variety of profibrogenic genes, which are closely associated with tissue fibrosis. The aim of this study was to compare HIF-1α expression in normal buccal mucosa tissues and OSF specimens and further explore the potential mechanisms that may lead to the induction of HIF-1α expression. Twenty-five OSF specimens and six normal buccal mucosa were examined by immunohistochemistry. The expression of HIF-1α from fibroblasts cultured from OSF and normal buccal mucosa was measured by Western blot. Arecoline, a major areca nut alkaloid, was challenged to normal buccal mucosa fibroblasts (BMFs) to elucidate whether HIF-1α expression could affect by arecoline. In addition, the effects of arecoline on plasminogen activator inhibitor (PAI)-1 expression were evaluated in environmental hypoxia. HIF-1α expression was significantly higher in OSF specimens and expressed mainly by fibroblasts, epithelial cells, and inflammatory cells. Fibroblasts derived from OSF were found to exhibit higher HIF-1α protein expression than BMFs (P<0.05). Arecoline was found to upregulate HIF-1α protein in a dose-dependent manner (P<0.05). Hypoxia increased arecoline-induced PAI-1 protein expression than normoxic conditions (P<0.05). These results suggest that HIF-1α expression is significantly upregulated in OSF tissues from areca quid chewers, implying a potential role as a biomarker for local tissue hypoxia. The activation of HIF-1α may promote fibrogenesis by an increase of PAI-1 expression and a subsequent elevation of extracellular matrix production in oral submucosa leading to fibrosis.

Transforming growth factor beta 1 in oral submucous fibrosis: An immunohistochemical study - Understanding the pathogenesis

Background: The development of fibrosis is pathognomic in the potentially malignant oral disorder, oral submucous fibrosis (OSF). Strong evidence exists to implicate the chewing of areca nut in the pathogenesis of the lesion. The constituents of areca nut activate several pro-fibrotic cytokines, chiefly transforming growth factor beta 1 (TGF-β1), which leads to an increased deposition and decreased degradation of extracellular matrix and collagen. TGF-β1 probably represents the major pathway in the deposition of collagen fibers in this condition. The present study aims to identify and correlate the expressions of TGF-β1 immunohistochemically on paraffin sections of various stages of OSF. Materials and Methods: The expression of TGF-β1 antibody was detected immunohistochemically using the anti-TGF-β1 mouse monoclonal antibodies (8A11-NovusBio USA) on paraffin sections of 58 cases of OSF, 10 cases of normal buccal mucosa tissue and 5 cases of scar tissue. The site, extent, and intensity of expression and quantification of TGF-β1 were noted and a comparative evaluation between various grades of OSF. Scar tissue and normal oral mucosa was made using image analysis software (Jenoptik optical system-ProReg ® Capture Pro 2.8.8 software [2011]). Results: Cells of spinous layer of the epithelium showed more intense staining in all grades of OSF, Grade II showed the highest percentage of expression, same as that of keloid (17%) but less than that of normal mucosa (12%). Positive staining was seen around blood vessels, muscles, fibers in the submucosa and perimuscle fibers. Highest expression was in the muscle in Grade III (80%) compared with normal oral mucosa (37%). Conclusion: These results suggest that the pathogenesis of OSF and scar/keloid could be linked through the TGF-β1 pathway. Interventions directed at the TGF-beta pathway may hold the key in the future management of this oral potentially malignant condition.

N-Succinyl Chitosan as Buccal Penetration Enhancer for Delivery of Herbal Agents in Treatment of Oral Mucositis

Oral mucositis is one of the major side effects of cancer chemotherapy (30-76%) and radiotherapy (over 50%). Current palliative treatments of oral mucositis include specialized agents like pelifermin, platelet derived factors etc. or oral hygienic agents which suffered from various drawbacks like systemic side effect, least effect owing to fast wash out of buccal mucosa, patient unfriendly delivery systems, and mere symptomatic relief. In this research work, N-succinyl chitosan gel delivery system of microemulsified eugenol, honey and sodium hyaluronate was prepared to explore their multiple and synergistic effects on various pathological factors of oral mucositis. N-succinyl chitosan was synthesized in our laboratory and loaded with microemulsified eugenol (10% v/v), honey (10% v/v) and sodium hyaluronate (0.2% w/v) to prepare orogel with optimum pH, spreadability, mucoadhesion strength, and viscosity. In vitro eugenol release from N-succinyl chitosan gel after 8 hours in PBS (pH-6.4) was found to be 87.45±0.14%, which was better in comparison to that released from chitosan gel. Ex vivo penetration studies using rat buccal mucosal tissue also suggested better J-efflux of eugenol through N-succinyl chitosan in comparison to chitosan gel with enhancement ratio (ER) of 1.71. The antimicrobial effect of N-succinyl chitosan based orogel against S. aureus and C. albicans efficacy was found to be statistically high in comparison to chitosan based orogel as well as marketed formulation of chlorhexidine (p<0.05). The N-succinyl chitosan orogel in 5-fluoro uracil induced oral mucositis animal (Wistar rats) model showed enhanced survival ratio, weight gain and high tissue regeneration activity than chitosan gel formulation within 15 days. The formulation was successful in elevating the survival and reducing the inflammation in the oral mucosa of animals compared to disease control (p<0.05) and hence suggesting the potential of N-succinyl chitosan orogel in the treatment of oral mucositis.

Assessment of the effect of local application of amifostine on acute radiation-induced oral mucositis in guinea pigs

The aim of present study was to assess the radioprotective effects of the local application of amifostine to treat acute buccal mucositis in guinea pigs. A total of 32 guinea pigs were randomized into four groups: (Group A) topically administered 50 mg of amifostine plus radiotherapy (RT); (Group B) 100 mg amifostine plus RT; (Group C) normal saline plus RT; and (Group D) normal saline plus sham RT. The opportunity for administration was 15 min before irradiation. When administered, the cotton pieces that had been soaked with 0.5 ml amifostine solution or saline were applied gently on the buccal mucosa of each guinea pig for 30 min. The animals in Groups A, B and C were irradiated individually with a single dose of 30 Gy to the bilateral buccal mucosa. Eight days after irradiation, the animals were scored macroscopically; they were then euthanized, and the buccal mucosal tissues were processed for hematoxylin–eosin staining and ICAM-1 immunohistochemical analysis. In Groups A and B, the mean macroscopic scores were 2.9 ± 0.6 and 2.4 ± 1.1, respectively. There was no significant difference between the two groups (P > 0.05). However, when they were separately compared with Group C (4.4 ± 0.7), a noticeable difference was obtained (P < 0.05). No mucositis was observed in Group D. Comparisons of the expression of ICAM-1 were in agreement with the macroscopic data. Histologically, superficial erosion, exudate and ulcer formation were all observed in the RT groups; only the severity and extent were different. The microscopic observations in the amifostine-treated groups were better than in Group C. The results demonstrated that topical administration of amifostine to the oral mucosa is effective treatment of acute radiation-induced mucositis.

Forehead Flap for Simultaneous Reconstruction After Head and Neck Malignant Tumor Resection.

Oncologic resection of the head and neck malignant tumor often results in complex reconstructive problems that require a reliable flap to restore both the form and function in single-stage operation. The free flap transfer has revolutionized head and neck reconstruction. However, forehead flap pedicled with superficial temporal artery (STA) or supraorbital artery (SOA)/supratrochlear artery (STrA) is a new option for reconstruction of head and neck with its own specific advantages. The forehead flap was applied to reconstruct the defects of the posterior hypopharyngeal wall, external perioral skin and buccal mucosal, and nose and periorbital tissues after malignant tumor resection. Nine of the patients were reconstructed with forehead flap pedicled with STA, 7 of whom underwent postoperative radiation. Eight of the patients were pedicled with SOA (and/or STrA), 1 of whom underwent postoperative radiation. We applied this method in a total of 17 male patients. The mean age of patients was 61 years (range, 47-69 years) with a follow-up time of 12 to 180 months. The biggest defect covered by forehead flap pedicled with STA was measured 10.0 × 6.5 cm. The biggest defect covered by forehead flap pedicled with SOA and/or STrA was measured 9.0 × 7.0 cm. In all 17 patients, no vascular crisis happened and no partial or total flap necrosis was observed. All skin graft area and donor site healed well. No patient experienced flap strictures or fistulas after operation or after radiotherapy. No local recurrence developed during the follow-up time. Functions were assessed by specialist as optimal as no limitation of eyes or mouth opening, and no limitation of food swallowing. The forehead flap provides thin pliable tissue with reliable blood supply and high tolerance to radiotherapy. These advantages make it a valuable option to simultaneously reconstruct the defects after head and neck malignant tumor resection in certain selected patients.

In vivo preventive effects of insect tea on buccal mucosa cancer in ICR mice.

Insect tea is a particular drink or health product in China and it is also used as Chinese medicine now. Its functional effects need to be proved. The ICR mice buccal mucosa cancer model was established by injecting the mice with U14 cells and mice was treated with insect tea. Tumor volumes and lymph node metastasis rates were determined. And the buccal mucosa tissues and cancer cervical lymph node were also checked by histology test, real-time polymerase chain reaction, and western blot assays. The tumor volumes for the group treated with insect tea mice was smaller than those from the control mice. The sections of buccal mucosa cancer tissue showed that the canceration of insect tea mice was weaker than control mice. Insect tea significantly induced apoptosis in buccal mucosa tissues by upregulating Bax, caspases, and downregulating Bcl-2. Nuclear factor-κB, inducible nitric oxide synthase, and COX-2 gene, was significantly downregulated by insect tea, demonstrating its anti-inflammatory properties. Insect tea also exerted a great anti-metastasis effect on tissues as demonstrated by decreased expression of Matrix metalloproteinases genes and increased expression of tissue inhibitors of matrix metalloproteinases. The highest concentration of 1600 mg/kg oral gavage and 400 mg/mL smear insect tea showed the best anticancer effects. Based on the results, insect tea showed the strong in vivo buccal mucosa cancer preventive effects.