Genome-wide gene expression profiling of tongue squamous cell carcinoma by RNA-seq.

OBJECTIVE: Tongue squamous cell carcinoma (TSCC) is significantly more malignant than other type of oral squamous cell carcinoma (OSCC). In this study, we aimed to identify specific global gene expression signatures of TSCC to investigate the more invasive behavior of the deeply infiltrating cancer. METHODS: Using RNA-seq technology, we detected gene expression of 20 TSCCs, 20 matched paratumor tissues, and 10 healthy normal mucosa tissues. Enrichment analysis of gene ontology (GO) and pathway was conducted using online tools DAVID for the dysregulated genes. Additionally, we performed the quantitative real-time RT-PCR (qRT-PCR) to validate the findings of RNA-Seq in 10 samples of TSCC, matched paratumor, and normal mucosa, respectively. RESULTS: We detected 252 differentially expressed genes (DEGs) between TSCC and matched paratumor tissue, including 117 up-regulated and 135 down-regulated genes. For comparison between TSCC and normal mucosa, 234 DEGS were identified, consisting of 67 up-regulated and 167 down-regulated genes. For both two comparisons, GO categories of muscle contraction (GO: 0006936), epidermis development (GO: 0008544), epithelial cell differentiation (GO: 0030855), and keratinization (GO: 0031424) were commonly enriched. Altered gene expression affected some cancer-related pathways, such as tight junction. The qRT-PCR validation showed that gene expression patterns of FOLR1, NKX3-1, TFF3, PIGR, NEFL, MMP13, and HMGA2 were fully in concordance with RNA-Seq results. CONCLUSION: Findings in this study demonstrated the genetic and molecular alterations associated with TSCC, providing new clues for understanding the molecular mechanisms of TSCC pathogenesis.

The anterolateral thigh flap for soft tissue reconstruction in patients with tongue squamous cell carcinoma.

BACKGROUND: Surgery remains the first choice of treatment for tongue cancer. Immediate reconstruction should be performed after wide resection of tumour. The aim of this study was to evaluate the anterolateral thigh flap for reconstruction of lingual defects. METHODS: We report 39 consecutive oral tongue squamous cell carcinoma patients who underwent lingual reconstruction with the anterolateral thigh flap between 2009 and 2010. RESULTS: The width of the skin island was 4 to 5 cm and the length of the skin island was 6 to 8 cm in 31 patients with T2 tumour, while the width of the skin island was 5.5 to 6 cm and the length of the skin island was 9 to 12 cm in 8 patients with T3/4 tumours. The all flap survival rate was 97.5 % in our series. CONCLUSIONS: We could obtain sufficient flap volume using the anterolateral thigh flap for tongue reconstruction. The single perforator-based anterolateral thigh flap is a good option for soft tissue reconstruction in patients with oral tongue squamous cell carcinoma.

The impact of clinical prognostic factors on the survival of patients with oral squamous cell carcinoma.

PURPOSE: This study investigated prognostic factors and useful predictors for survival in patients with oral squamous cell carcinoma. PATIENTS AND METHODS: The medical records of all patients with oral squamous cell carcinoma who underwent curative surgery with or without adjuvant radiation from 2009 through 2011 were retrospectively reviewed. RESULTS: Seventy-seven patients (63 men and 14 women) were enrolled. The 2-year disease-free and overall survival rates were 67.5% and 74%, respectively. Multivariate analyses showed that a diagnostic delay longer than 2 months (hazard ratio [HR]=4.43; 95% confidence interval [CI], 1.26-15.51; P=.02), T3 or T4 tumor (HR=4.40; 95% CI, 1.91-10.12; P=.001), neck metastasis (HR=1.96; 95% CI, 1.21-4.37; P=.01), and stage III or IV disease (HR=3.94; 95% CI, 1.64-9.47; P=.002) were independent adverse factors for survival rate. CONCLUSION: Oral squamous cell carcinoma is an important health issue associated with poor survival. A diagnostic delay longer than 2 months, T3 or T4 tumor, neck metastasis, and stage III or IV disease were independent adverse factors for subsequent survival rate and locoregional recurrence in patients with oral squamous cell carcinoma.

Stereology study of oral verrucous carcinoma.

PURPOSE: Squamous cell carcinoma (SCC) is the most prevalent malignant neoplasm of the oral cavity and oral verrucous carcinoma (OVC) is a verrucous variant of SCC. The purpose of this study was to examine the clinical classification of OVC and see for any difference in the biological behavior between OVC and CSS. METHODS: OVC and SCC were divided into 5 groups: the exogenic type of OVC (eOVC), cystoid type of OVC (cOVC) and infiltrative type of OVC (iOVC); well differentiated SCC (wdSCC), and medium/poorly differentiated SCC (m/pdSCC). A normal mucosa (NM) group was also created and studied. Stereology was used to measure and describe the morphological parameters of the nucleus to cytoplasm ratio (Vnp), desmosomes, mitochondria, etc. Analysed were also the nucleus volume density (Vv), Vnp, desmosomes and intracellular desmosomes number density (Nv), which were observed by stereology. RESULTS: We noticed some statistically significant differences in the morphological parameters among the 6 groups including the Vv (p<0.05), the Vnp (p<0.05), the number density of desmosomes (p<0.05), and the Nv (p<0.05). CONCLUSION: This study provides a theoretical basis for the clinical diagnosis and therapy of OVC.

[Rat model with oral submucous fibrosis induced by arecoline and mechanical stimulation].

OBJECTIVE: The aim of this study was to induce oral submucous fibrosis (OSF) in Sprague-Dawley(SD) rat models by arecoline and mechanical stimulation. METHODS: Two factors factorial design was used to divide 48 rats into 8 groups (n=6). Different concentrations of arecoline (0, 0.5, 2, and 8 mg·mL⁻¹) and mechanical stimulation (with or without brush) were treated. After 16 weeks of treatment, the mouth opening was measured, the pathological changes of the buccal mucosa were observed, and the expressions of type Ⅲ collagen, transforming growth factor ß1 (TGF-ß1), and interferon-γ (IFN-γ) were detected. RESULTS: In rats with moderate and high concentrations of arecoline, typical OSF pathological features were observed in the buccal mucosa, the mouth openings were significantly reduced, and the expression levels of type Ⅲ colla-gen and TGF-ß1 were significantly increased (P<0.05). Although mechanical stimulation can increase the three indexes of mucosa (P<0.05), no pathological change and difference in the mouth opening was observed (P>0.05). CONCLUSIONS: Moderate and high concentrations of arecoline can induce OSF in SD rats, but mechanical stimulation cannot induce OSF.

[Anterolateral thigh flap for reconstruction of defects after en bloc resection of buccal cancer : a retrospective study of 23 cases].

PURPOSE: This study was aimed to observe the effects of anterolateral thigh flap for reconstruction of tissue defects after en bloc resection of buccal cancer. METHODS: Twenty-three patients with soft tissue defects after en bloc resection of buccal cancer underwent simultaneous reconstruction with anterolateral thigh flap from May, 2013 to May, 2015 were observed. Anterolateral thigh flaps were designed and harvested in form of single or multiple islands to restore the defect in buccal region after surgery. The appearance and function of both the oral and maxillofacial region and the donor site were recorded and evaluated. RESULTS: All the 23 flaps survived. Only 3 of them experienced vascular crisis within 24 hours after surgery, and recovered gradually after salvage. The success rate was 100%. One to three years of follow-up showed satisfying morphology and function for both the receipt sites and the donor sites. Buccal abscess was observed in 1 patient and recovered after rinsing and drainage. Two patients died of recurrence. CONCLUSIONS: Good effects can be achieved using anterolateral thigh flap to reconstruct buccal defects after en bloc resection of cancer, which is suitable for application in clinical practice.

Trend Analysis of Betel Nut-associated Oral Cancer and Health Burden in China.

OBJECTIVE: To forecast the future trend of betel nut-associated oral cancer and the resulting burden on health based on historical oral cancer patient data in Hunan province, China. METHODS: Oral cancer patient data in five hospitals in Changsha (the capital city of Hunan province) were collected for the past 12 years. Three methods were used to analyse the data; Microsoft Excel Forecast Sheet, Excel Trendline, and the Logistic growth model. A combination of these three methods was used to forecast the future trend of betel nut-associated oral cancer and the resulting burden on health. RESULTS: Betel nut-associated oral cancer cases have been increasing rapidly in the past 12 years in Changsha. As of 2016, betel nuts had caused 8,222 cases of oral cancer in Changsha and close to 25,000 cases in Hunan, resulting in about ¥5 billion in accumulated financial loss. The combined trend analysis predicts that by 2030, betel nuts will cause more than 100,000 cases of oral cancer in Changsha and more than 300,000 cases in Hunan, and more than ¥64 billion in accumulated financial loss in medical expenses. CONCLUSION: The trend analysis of oral cancer patient data predicts that the growing betel nut industry in Hunan province will cause a humanitarian catastrophe with massive loss of human life and national resources. To prevent this catastrophe, China should ban betel nuts and provide early oral cancer screening for betel nut consumers as soon as possible.

Effect of Capsaicin Cream on the Secretion of the Submandibular and Parotid Gland in the General Population with Different Chilli-eating Habits.

OBJECTIVE: To investigate the effect of capsaicin cream on the secretion of the submandibular gland (SMG) and the parotid gland (PG) in the general population, with different chilli-eating habits. METHODS: In two groups with different chilli-eating habits, the salivary flow rate of the SMG and the PG was detected at statics and different times, after application of capsaicin cream. RESULTS: In both groups, the topical application of capsaicin cream could significantly increase the salivary secretion of SMG (P < 0.05), but the increase in the salivary flow rate of the SMG between the two groups had no significant difference (P > 0.05). On the other hand, although the salivary flow rate of PG also increased after stimulation, the increase had no statistical difference (P > 0.05). CONCLUSION: The application of capsaicin cream can effectively promote the secretion of the SMG and the PG, and its effect is independent of chilli-eating habits, which indicates that topical application of capsaicin cream can be considered as a potential treatment for the hypofunction of the salivary gland.

Investigation of the association between miR­181b, Bcl­2 and LRIG1 in oral verrucous carcinoma.

Abnormal expression of microRNAs (miRNAs) is involved in the development of and anti­apoptotic effects in various types of human cancer. However, miRNA­mediated regulation of oral verrucous carcinoma (OVC) remains to be elucidated. The present study aimed to investigate the expression of miR­181b in OVC and oral squamous cell carcinoma (OSCC). The expression levels of miR­181b were determined using reverse transcription­quantitative polymerase chain reaction. The expression levels of B­cell lymphoma 2 (Bcl­2) and leucine rich repeats and immunoglobulin like domains 1 (LRIG1), were evaluated using immunohistochemical staining. The correlation between Bcl­2 and LRIG1 expression was determined using a Pearson correlation analysis. The expression levels of miR­181b and Bcl­2 in OVC were significantly higher compared with normal mucosal tissue (NM); however, lower compared with the OSCC. The key target of miR­181b was LRIG1 and it was significantly lower in OVC tissues compared with NM tissue; however this was higher when compared with OSCC tissue. The expression levels of Bcl­2 were correlated with expression levels of LRIG1 in OVC tissues. Therefore, LRIG1 may be associated with anti­apoptotic function in OVC tissues.

[Expression of matrix metalloproteinase 2 in oral verruvous carcinoma and squamous cell carcinoma].

OBJECTIVE: To determine the expression of MMP2 mRNA in oral verruvous carcinoma and squamous cell carcinoma. METHODS: Thirty cases were divided into 3 groups: verruvous carcinoma (n = 10), well-differentiated squamous cell carcinoma (n = 15) and moderately or poorly differentiated squamous cell carcinoma (n = 5). Reverse transcription polymerase chain reaction (RT-PCR) was used to test the expression of MMP2 mRNA in the carcinoma tissues and matched normal tissues from 3 groups above. RESULTS: The expression of MMP2 mRNA in the carcinoma tissues was significantly higher than that in their matched normal tissues (P < 0.05). The expression of MMP2 mRNA in verruvous carcinoma was significantly higher than that in well-differentiated and moderately or poorly differentiated squamous cell carcinoma (P < 0.05). However, the expression of MMP2 mRNA was not obviously different between well-differentiated and moderately or poorly differentiated squamous cell carcinoma (P > 0.05). CONCLUSION: The expression of MMP2 mRNA in oral verruvous carcinoma and squamous cell carcinoma tissues was significantly higher than that in their matched normal tissues. The expression of MMP2 mRNA in verruvous carcinoma was significantly higher than that in squamous cell carcinoma.

Gene profiling analysis for patients with oral verrucous carcinoma and oral squamous cell carcinoma.

Oral verrucous carcinoma (OVC) is one malignant tumor which was carved out from the oral squamous cell carcinoma (OSCC). However, the clinical and pathological features as well as the treatment strategies of OVC are different from OSCC. Here, global transcript abundance of tumor tissues from five patients with primary OVC and six patients with primary OSCC including their matched adjacently normal oral mucosa were profiled using the Affymetrix HGU133 Plus 2.0. Ingenuity Systems IPA software was used to analyse the gene function and biological pathways. There were 109 differentially expressed genes (more than 2-fold) between OVC and the adjacently normal tissue, among them 66 were up-regulated and 43 were down-regulated; 1172 differentially expressed genes (2-fold) between OSCC and the adjacently normal tissue, among them 608 were up-regulated and 564 were down-regulated. There were 39 common differentially expressed genes in OVC and OSCC compared with their matched normal oral mucosa, among them 22 up-regulated and 17 down-regulated, and 8 of them different between OVC and OSCC. In addition, the gene expression profile was further validated by quantitative real-time PCR (Q-RT-PCR) analysis for four of those 39 selected genes.

[Study of αB-crystallin and its possible role of anti-apoptosis in oral verrucous carcinoma].

PURPOSE: To investigate the expression of αB-crystallin and its possible role of anti-apoptosis in oral verrucous carcinoma. METHODS: The expression of αB-crystallin and activated caspase-3 was detected in oral verrucous carcinoma, oral squamous carcinoma and normal mucosa by immunohistochemistry, and their relationship was investigated. SPSS 16.0 software package was used for statistical analysis. Nonparametric test and spearman correlation test were performed. RESULTS: The expression of αB-crystallin in oral verrucous carcinoma and oral squamous carcinoma was significantly higher than that in normal mucosa(P<0.05). And in oral verrucous carcinoma, the increase of expression of αB-crystallin coincided with the decrease of expression of activated caspase-3(P<0.05). CONCLUSION: αB-crystallin may play a role of anti-apoptosis by inhibiting the activation of caspase-3 in oral verrucous carcinoma.

[Myoepithelial carcinoma of salivary glands: a clinical analysis of 13 cases].

PURPOSE: This study retrospectively analyzed the clinical data of 13 patients with myoepithelial carcinoma of salivary glands for improving the accuracy of diagnosis and treatment outcome. METHODS: Thirteen cases with myoepithelial carcinoma of the salivary glands in Xiangya Hospital from January 1992 to September 2010 were reviewed, including the clinical biological behavior, diagnosis,treatment and prognosis. RESULTS: Thirteen patients included 6 men and 7 women, aged from 14 to 71 years (median 40 years).The tumor occurred predominantly in the parotid gland (53.8%).Among the 13 cases,7 were clinically misdiagnosed as benign tumors and 2 were misdiagnosed pathologically. All cases underwent operation. Two cases received surgery plus adjuvant chemotherapy; five cases underwent surgery plus adjuvant radiotherapy. 4(30.8%) had cervical lymph node metastasis and 2 cases(15.4%) developed distant metastasis. Follow-up time ranged from 3 months to 6 years. Six cases died of local recurrence or distal metastasis. CONCLUSIONS: Myoepithelial carcinoma of the salivary glands is a rare tumor. The diagnosis is depended on histology and immunohistochemistry. The tumor has a high rate of distant metastasis and high rate of lymph node metastasis in T3 to T4 cases. Radical surgery is the treatment of choice. Elective neck dissection should be considered in T3 to T4 cN0 cases. The effect of chemotherapy and radiotherapy needs to be investigated.

[A retrospective study of oral squamous cell carcinomas originated from oral submucous fibrosis].

OBJECTIVE: To investigate the clinical characteristics, pathological and biological behaviors of oral squamous cell carcinomas (OSCC) originated from oral submucous fibrosis (OSF). METHODS: A total of 528 patients with OSCC treated at Department of Oral and Maxillofacial Surgery, Xiangya Hospital, Central South University from 2002 to 2010 were divided into two groups, OSCC originated from OSF and OSCC not originated from OSF. The clinical data regarding age, gender, location, pathological type, metastasis and recurrence rate of these patients were analyzed retrospectively. RESULTS: The mean age of the group of OSCC originated from OSF (45.8 years) was younger than that of the group of OSCC not originated from OSF (55.9 years). The male female ratio of the group of OSCC originated from OSF (32.3:1) was higher than that of the group of OSCC not originated from OSF (2.3:1). The metastasis and recurrence rate of the group of OSCC originated from OSF (13.5%, 39.1%) was higher than that of the group of OSCC not originated from OSF (7.6%, 27.8%.). CONCLUSIONS: OSCC originated from OSF occurs at younger age and more in male, and is clinically more invasive and metastatic.

[Reconstruction of oral and maxillofacial defects using free anterolateral thigh flaps in patients with malignant tumors].

PURPOSE: To explore the value of free anterolateral thigh flaps in the reconstruction of oral and maxillofacial defects following radical resection of malignant tumors. METHODS: Seventy-six cases of oral and maxillofacial malignant tumor from December 2007 to June 2009, including 32 tongue carcinomas, 16 buccal carcinomas, 10 carcinomas of floor of mouth, 9 gingival carcinomas, 8 oropharyngeal carcinomas and 1 skin carcinoma, underwent radical resection. Free anterolateral thigh flaps were used to reconstruct the defects. The postoperative effect and flap success rate were evaluated. RESULTS: Among 76 cases, 74 flaps survived completely with satisfied configuration, only 2 flaps had complete necrosis. The survival rate was 97.3%. No complications were observed in the donor site. CONCLUSIONS: The free anterolateral thigh flap is an ideal choice for reconstruction of oral and maxillofacial defects, as it has many advantages such as reliable blood supply, large size available, and little donor site morbidity.

[Clinical application of heterogeneous acellular dermal matrix in the surgical treatment of oral submucous fibrosis].

PURPOSE: The aim of this study was to evaluate the clinical effect of heterogeneous acellular dermal matrix in the surgical treatment of advanced oral submucous fibrosis (OSF). METHODS: There were eight patients who had undergone surgical treatment of trismus caused by OSF. Surgery was performed under general anaesthesia given through a nasoendotracheal tube using a fibreoptic bronchoscope. All the fibrous bands on the buccal mucosa were incised and bluntly dissected to stretch the mouth opening. Based on the defect, heterogeneous acellular dermal matrix graft was applied directly on the bilateral wounds. A tie-over dressing technique was used to hold the graft firmly to its bed.The sutures and dressing material were removed on the tenth to fourteenth day after the operation. All the patients were asked to stretch the mouth opening 3 times daily and were followed up for at least 6 months. A modality of medical treatment including multiple micronutrient supplements and steroid injection therapy were carried out if necessary. The changes in the interincisal distances (IDs) were analyzed in the preoperative, intraoperative, and postoperative stages. The data was analyzed with SPSS16.0 software package for one-way ANOVA. RESULTS: The overall success rate was 100% without any or only partial graft loss. There were no immunologic reactions or significant complications.The mean preoperative, intraoperative, and final follow-up interincisal distances were 12.04, 35.46, and 29.33 mm, respectively. Evaluation of the changes in the IDs revealed statistically significant differences between the values recorded at the different stages.(P<0.05). CONCLUSIONS: Heterogeneous acellular dermal matrix may be a valuable biomaterial for repairing oral mucosal defects after surgery on the fibrous bands in patients with advanced OSF through covering and protecting wound surface early,which shortens the healing time of wound, decreases relapse of fibrosis and improve the restricted mouth opening.

[Effects of clotrimazole on the growth of oral squamous cell carcinoma].

OBJECTIVE: to investigate the effects of clotrimazole on the growth of oral squamous cell carcinoma (OSCC). METHODS: OSCC-25 cells growing in log phase were treated with various doses of clotrimazole. The concentration of IC(50), cell cycle and cell cycle related protein were examined. RESULTS: the concentration of clotrimazole for inhibiting OSCC was IC(50) 8.51 µmol/L. Clotrimazole induced cell cycle arrest in the G(0)-G(1) cell cycle phase, with a concomitant decrease of cells in the G(2)-M and S-phase. Furthermore, clotrimazole significantly decreased the levels of cyclin D, cyclin E and CDK-4. CONCLUSIONS: clotrimazole inhibits the growth of OSCC.

[Establishment of xenografted model of oral verrucous carcinoma with focal squamous cell carcinoma metastasizing to the cervical lymph nodes in nude mice].

PURPOSE: To use the excised cervical lymph node tissue from oral verrucous carcinoma patient with focal squamous cell carcinoma subcutaneously to establish a xenografted model in nude mice. METHODS: The xenograft tumors were finally removed for histopathological study and the mice were laparotomized to examine metastatic tumors in livers, kidneys, lungs. RESULTS: The tumor formation rate was 87.5%(7/8),and the appearance of transplanted tumors was like that in human and HE staining showed that the cancer cells of those tumors models and mesenchymal components remained morphologically like the original tumor. The liver, renal, lung and lymph nodes didn't show obvious metastasis. CONCLUSION: The xenografted model is successfully established with a higher formation rate, and the model morphologically resembles the human tumor.

[Gene expression profile changes in oral verrucous carcinoma and oral squamous cell carcinoma].

OBJECTIVE: To determine the difference in the gene expression between human oral verrucous carcinoma (OVC) and oral squamous cell carcinoma (OSCC). METHODS: cDNA chip was used to detect the mRNA of cancer tissue from 4 OVC and 4 OSCC. After profile blotted and handled by bioinformation, the gene expression of these two kinds of lesions was examined. RESULTS: Using the BioStarH-40 profile, 593 different expression of genes was found. The rate of different genes was 15.2%, of which the expression of 283 genes increased (59 genes significantly increased) and 310 genes decreased (98 genes significantly decreased) in OVC tissue than that in OSCC. CONCLUSIONS: The gene expression of OVC and OSCC was different, which many contribute to the different biological behavior of these two kinds of lesions.

[The expression of Maspin and VEGF gene in oral squamous cell carcinoma and its significance].

PURPOSE: To study the expression of Maspin(marray serine proteinase inhibitor) and VEGF(vascular endothelial growth factor) gene in oral squamous cell cancers(OSCC) and the relationship between Maspin and VEGF. METHODS: mRNA expression of Maspin and VEGF gene was investigated by RT-PCR in every samples of 18 OSCC cases. F test and Person test were used for data analysis. RESULTS: mRNA expression of Maspin gene increased gradually in OSCC, paratumor and normal epithelium(P<0.01); mRNA expression of VEGF gene decreased gradually in OSCC,paratumor and normal epithelium(P<0.01); The expression of Maspin and VEGF gene was negatively correlated in OSCC(P<0.01) ; The expression of Maspin gene was negatively associated but VEGF gene was positively associated with lymph node metastasis in OSCC(P<0.05); The expression of Maspin and VEGF gene had no relation with the histological grade or tumor size in OSCC. CONCLUSION: Maspin and VEGF gene may play an important role in the development of OSCC and Maspin gene may down-regulate the expression of VEGF gene.