Composition and function of oral microbiota between gingival squamous cell carcinoma and periodontitis.

OBJECTIVES: Previous studies have proved that periodontitis is an independent risk factor of oral squamous cell carcinoma (OSCC) epidemiologically. Along with the important role of microbiota in the cancer process and the specific anatomical position, our study explored the microbial composition and functions in periodontitis and gingival squamous cell carcinoma (GSCC). MATERIALS AND METHODS: GSCC patients (n = 10), matched periodontitis patients (n = 15), and healthy individuals (n = 15) were recruited. Saliva, subgingival plaque, tongue dorsum, buccal mucosa, cancerous tissue, and paracancerous tissue samples were collected. 16S rDNA amplicon sequencing and functional prediction were applied for the taxonomic analysis. RESULTS: Periodontal pathogens occupied 46% in GSCC. Besides, the mutual operational taxonomy unites (OTU) generated from the subgingival plaque occupied 38.36% and 44.13% from saliva. Fusobacterium, Peptostreptococcus, and Prevotella were more abundant in cancerous tissues, while Streptococcus, Neisseria, and Haemophilus were more enriched in saliva or soft mucosa. PCoA exhibited similar cluster between tongue dorsum and saliva in GSCC. GSCC showed lower richness than periodontitis. In saliva and subgingival plaque, Atopobium was more prevalent in GSCC than periodontitis and controls in descending order. Lipopolysaccharide (LPS) biosynthesis increased in subgingival plaque of GSCC compared with the other two groups. CONCLUSION: Periodontal pathogens were abundant in GSCC. Cancerous tissues harbor enriched periodontal pathogens while saliva or soft mucosa harbored more periodontal health related bacteria. A high level of Atopobium in saliva and LPS biosynthesis have the potential for increasing the risk of suffering from GSCC in individuals with periodontitis, which needs more evidence to clarify it.

MicroRNA-532-3p Suppresses Malignant Behaviors of Tongue Squamous Cell Carcinoma via Regulating CCR7.

To provide better therapeutic avenues for treating tongue squamous cell carcinoma (TSCC), a series of experiments about the effects of microRNA (miR)-532-3p on TSCC malignant behaviors were carried out. The result showed that miR-532-3p was down-regulated and C-C chemokine receptor 7 (CCR7) was up-regulated in the tumor tissues compared with those in the paired paratumor tissues. Further, expression of miR-532-3p was detected in four TSCC cell lines, TSCCA, TCA8113, CAL-27, and SCC-25. The miR-532-3p mimics and inhibitor were transfected into the CAL-27 and TCA8113 cell lines which were the relatively lowest and highest miR-532-3p expressions, respectively. It was found that the overexpression of miR-532-3p suppressed TSCC cell proliferation, migration, invasion, and promoted apoptosis in vitro, whilst the knockdown of miR-532-3p reversed these behaviors. The bioinformatics predicted that CCR7 was a downstream gene of miR-532-3p, which was confirmed via luciferase assay. Following, the decline of CCR7 in the miR-532-3p mimics group and the rise of CCR7 in the miR-532-3p inhibitor group were also verified. In addition, enhanced cell proliferation, migration and invasion induced by CCR7 were partly restrained by miR-532-3p in TSCC cell. Meanwhile, miR-532-3p attenuated tumourigenesis in vivo due to the reduction of tumor volume and Ki-67 positive rate and the increase of apoptotic cells. Taken together, these findings reveal a pivotal role for the miR-532-3p/CCR7 axis in regulating TSCC, and this novel axis could be suitable for therapeutic intervention in TSCC disease.

Association between polymorphisms in pre-miRNA genes and risk of oral squamous cell cancer in a Chinese population.

BACKGROUND: MicroRNAs play important roles in the development of human cancers. This case-control study is to evaluate the roles of the polymorphisms in pre-miRNAs on risk of oral cancer in a Chinese population. METHODS: The genotypes of three polymorphisms were determined in 340 patients with oral squamous cell cancer and 340 healthy controls who were frequency matched for age and sex. Odds ratios (ORs) and 95% confidence intervals (95%CIs) were calculated to assess the association. All analyses were performed using the SPSS software. 3.154() 0.001. RESULTS: For miR-499 rs3746444, individuals carrying homozygous CC genotype had increased risks of oral cancer compared with the homozygous wild TT genotype (adjusted OR was 3.154, 95%CI was 1.555-6.397, P value was 0.001). The C allele of miR-499 rs3746444 was associated with a higher risk of oral cancer with significant odds ratio of 1.453. In the stratified analyses by sex, the associations between miR-499 rs3746444 and miR-146a rs2910164 polymorphisms with the susceptibility of oral squamous cell cancer were significant in males. However, with 1/4 as many subjects there were no significant associations between the three polymorphisms and oral cancer risks in females. The joint effects of miRNA polymorphisms and smoking on the risk of OSCC were analyzed and the results suggested that the association between microRNA genetic variants and OSCC risk was modified by smoking. CONCLUSIONS: These findings suggest that miR-499 rs3746444 and miR-146a rs2910164 polymorphisms may contribute to genetic susceptibility to oral squamous cell cancer.

The association between miR-499 polymorphism and cancer susceptibility: a meta-analysis.

BACKGROUND: MicroRNAs are a class of new noncoding RNA that play important roles in the pathogenesis of tumor. Rs3746444 in miR-499 is suggested to be associated with cancer susceptibility. In the present study, we assess the association between miR-499 rs3746444 polymorphism and cancer susceptibility through a meta-analysis. METHODS: We searched relevant articles from the PubMed and Embase databases. We screened all the resulting articles for adherence to the inclusion and exclusion criteria. The associations between miR-499 polymorphism and cancer susceptibility were estimated by computing the odds ratios (ORs) and 95% confidence intervals (CIs). All analyses were performed using Stata software. RESULTS: There are 18 datasets included in the analysis. Statistically significant associations were found between the miR-499 rs3746444 polymorphism and susceptibility to cancer (GG versus AA: OR =1.24, 95% CI: 1.01-1.52; G versus A: OR =1.11, 95% CI: 1.01-1.23). A subsequent analysis, on the basis of ethnicity for the population characteristic, showed that Asians had increased susceptibility to cancer (GG versus AA: OR =1.32, 95% CI: 1.09-1.59; GG + AG versus AA: OR = 1.17, 95% CI: 1.01-1.37). In the subgroup analysis of tumor type, none of the genetic models had statistically significant results. The meta-regression suggested that race and cancer types are not the source of heterogeneity in the present meta-analysis. No publication bias was detected by either the inverted funnel plot or Egger's test. CONCLUSION: Rs3746444 in miR-499 might be related to susceptibility to cancer.

Frizzled2 mediates the migration and invasion of human oral squamous cell carcinoma cells through the regulation of the signal transducer and activator of transcription-3 signaling pathway.

Frizzled2 (Fzd2) is a receptor for wingless-type MMTV integration site family members (Wnts), the aberrant overexpression of which has been noted to contribute to cancer metastasis. The present study was performed to characterize the role of Fzd2 in the migration and invasion of oral squamous cell carcinomas (OSCC) in vitro. Using TSCCa cells (a tongue SCC cell line) for loss- or gain-of-function of Fzd2, we found that a forced overexpression of Fzd2 promoted TSCCa cell migration and invasion, decreased the expression of epithelial­cadherin (E-cadherin, an epithelial marker) and increased that of vimentin, Snail Slug, matrix metalloproteinases (MMPs)-2/-9/-13 and a-disintegrin and metalloproteinase with thrombospondin motifs-5 (ADAMTS5). By contrast, RNA interference (RNAi)-mediated knockdown of Fzd2 had opposite effects on OSCC cells. In addition, we found that the phosphorylation of signal transducer and activator of transcription-3 (STAT3) was enhanced by Fzd2 overexpression, but suppressed by Fzd2 depletion, and that STAT3­specific shRNA attenuated Fzd2 overexpression­induced cell invasion. In summary, the present study demonstrated that Fzd2 contributes to the migration and invasion of OSCC cells, at least partly through regulation of the STAT3 pathway. These results suggest Fzd2 as a novel therapeutic target for OSCC.

Transverse cervical vessels as recipient vessels in oral and maxillofacial microsurgical reconstruction after former operations with or without radiotherapy.

BACKGROUND: The purpose of this study was to investigate the reliability and outcome of using the transverse cervical vessel (TCV) as a recipient vessel for microvascular reconstruction in patients whose vessels in the neck region are unavailable because of previous surgery or radiotherapy. METHODS: Between January 2012 and August 2014, secondary head and neck reconstruction was performed using the TCV as a recipient vessel in eight patients who had undergone previous neck dissection and radiation therapy (n = 5). Five patients had a recurrent carcinoma, one had undergone an operation for scar release and two had been treated surgically for a second primary cancer. The anterolateral thigh flap (ALT), anteromedial thigh flap (AMT), and fibular flap were used for the reconstruction. Clinical data were recorded for each patient. RESULTS: All of the ipsilateral transverse cervical arteries were found to be free of disease. The second free flap was revascularized using the TCVs (n = 6) or the external (n = 1) or internal (n = 1) jugular vein. The free flaps used for the reconstruction included the ALT flap (n = 6), AMT flap (n = 2), and fibular flap (n = 1). All of the flaps survived without vascular events, and the patients healed without major complications. The mean follow-up time was 11 months. One patient died of distant metastases during follow-up. CONCLUSIONS: In patients who have previously undergone neck surgery with or without radiotherapy, the TCVs are reliable and easily accessible recipient vessels for microsurgical reconstruction in the oral and maxillofacial region. If the transverse cervical vein is unavailable, the internal or external jugular vein should be dissected carefully to serve as an alternative for microvascular anastomoses.

Chemokine receptor 7 promotes tumor migration and invasiveness via the RhoA/ROCK pathway in metastatic squamous cell carcinoma of the head and neck.

Metastatic squamous cell carcinoma of the head and neck (SCCHN) has been shown to express chemokine receptor 7 (CCR7), which can activate signaling pathways to promote invasion and survival of SCCHN cells. We hypothesized that the RhoA/Rho-associated kinase (ROCK) pathway is involved in the CCR7-induced invasion and migration of metastatic SCCHN cells. Thus, using migration, matrigel invasion and scrape wound-healing assays, we elucidated the role of RhoA in mediating CCR7-associated cellular mobility. Pull-down assays and western blotting were used to measure RhoA and its downstream expression. Immunohistochemical staining and analysis were useful in identifying the correlation between CCR7 and RhoA expression and clinicopathological factors. The results showed that inhibition of RhoA/ROCK reduced the tumor cell migration and invasiveness induced by CCL19. Activated RhoA, proline-rich tyrosine kinase-2 (Pyk2) and cofilin induced by CCL19 were elevated, and increased RhoA, Pyk2 and cofilin activity was eliminated by CCR7mAb, RhoA/ROCK and Pyk2 inhibitors, indicating involvement of the RhoA/ROCK-Pyk2-cofilin cascade. In summary, CCR7 via RhoA/ROCK-Pyk2 cofilin pathway promotes invasion and migration of metastatic SCCHN cells.

Association between polymorphism of TGFA Taq I and cleft lip and/or palate: a meta-analysis.

BACKGROUND: Cleft lip and palate (CL/P) is one of the most common malformations in humans. Transforming growth factor alpha (TGFA) is a well characterized mammalian growth factor which might contribute to the development of CL/P. This meta-analysis aimed to summarize the association between the TGFA Taq I polymorphisms and CL/P. METHODS: We retrieved the relevant articles from PubMed, EMBASE, ISI Web of Science and SCOPUS databases. Studies were selected using specific inclusion and exclusion criteria. The odds ratios (ORs) and their 95% confidence intervals (95% CIs) were calculated to assess the association between TGFA Taq I polymorphism and CL/P risk. Meta-analyses were performed on the total data set and separately for the major ethnic groups, disease type and source of control. All analyses were performed using the Stata software. RESULTS: Twenty articles were included in the present analysis. There is a significant association between the TGFA Taq I polymorphism and CL/P (C1C2 vs C1C1: OR = 1.67, 95% CI = 1.23-2.25, C2C2 + C1C2 vs C1C1C1: OR = 1.52, 95% CI = 1.15-2.01; C2 vs C1:OR = 1.41, 95% CI = 1.12-1.78). Stratified analyses suggested that the TGFA Taq I polymorphism was significantly associated with CL/P in Caucasians (C1C2 vs C1C1: OR = 1.95, 95% CI = 1.34-2.86; C2C2 + C1C2 vs C1C1: OR = 1.68, 95% CI = 1.18-2.38; C2 vs V1: OR = 1.52, 95% CI = 1.14 -2.02). CONCLUSION: TGFA Taq I polymorphism may be associated with the risk of CL/P.

C392T polymorphism of the Wnt10a gene in non-syndromic oral cleft in a northeastern Chinese population.

Non-syndromic oral cleft is one of the most common congenital malformations, and more than 40 genes may be involved in its aetiology. Recent studies have shown that the Wnt10a gene may also contribute. We recruited 198 patients with non-syndromic oral clefts, comprising 96 elementary families (restricted to the patients and their parents) and 187 controls, to investigate their associations with the risk of such clefts and their subgroups in a Chinese Han population. The variant evaluated in this study was a single nucleotide polymorphism - specifically, a missense mutation C392T of Wnt10a. Polymerase chain reaction (PCR) restriction fragment length polymorphism (RFLP) was used to genotype the marker, and case-control and family-based associations were analysed. Although in the case-control study there were no significant differences in frequency distributions of genotypes or alleles between cases and controls in the groups with cleft palate and cleft lip and palate, the genotypic and allelic frequencies of C392T in the total groups and the group with cleft lip alone differed significantly from those in the controls (p=0.04, and 0.01, respectively). A transmission disequilibrium test showed a transmitted disequilibrium in C392T. In conclusion, we found an association between the C392T variant and non-syndromic oral clefts.

Pituitary tumor-transforming gene 1 (PTTG1) is overexpressed in oral squamous cell carcinoma (OSCC) and promotes migration, invasion and epithelial-mesenchymal transition (EMT) in SCC15 cells.

Pituitary tumor-transforming gene 1 (PTTG1) is an important oncogenic transcription factor implicated in various malignancies, including oral squamous cell carcinoma (OSCC), a common malignancy of head and neck. Although PTTG1 is reportedly overexpressed in OSCC tissues, its role in human OSCC remains elusive. Thus, this study was conducted to explore the correlation between PTTG1 expression and tumorigenesis of OSCC. We first examined PTTG1 mRNA and protein expression in 28 pairs of OSCC tissues and adjacent non-tumor tissues. PTTG1 protein levels in 98 OSCC specimens were also evaluated by using immunohistochemistry. Our data showed that both mRNA and protein expression levels of PTTG1 in OSCC tissue specimens were markedly higher than that in the corresponding non-tumor tissue samples. A high level of PTTG1 protein expression was found in 74 out of 98 cases (75.51 %) and it was correlated with lymph node metastasis (P = 0.002) and tumor-node-metastasis (TNM) stage (P = 0.007) of patients with OSCC. Moreover, forced overexpression of PTTG1 enhanced SCC15 cell migration and invasion, whereas knockdown of PTTG1 resulted in reverse phenomena. In addition, elevated PTTG1 also increased the activities and expressions of matrix metalloproteinase (MMP)-2, and enhanced epithelial-mesenchymal-transition (EMT) process in SCC15 cells. The EMT changes were accompanied by downregulation of epithelial cadherin (E-cadherin) protein expression and upregulation of snail and vimentin. In summary, our results illustrate that PTTG1 may contribute to the development and progression of human OSCC.

Overexpression of Wnt11 promotes chondrogenic differentiation of bone marrow-derived mesenchymal stem cells in synergism with TGF-ß.

Bone marrow-derived mesenchymal stem cells (MSCs), the most widely used cell source for cartilage tissue engineering, are multipotent cells which have been shown to differentiate into various mesenchyme-lineage cell types including chondrocytes. However, the molecular mechanisms controlling the chondrogenic differentiation of MSCs remain to be fully elucidated. It has been demonstrated that Wnt signaling involves regulating chondrogenesis and MSC differentiation. The aim of the present study was to investigate the role of Wnt11, a member of noncanonical Wnts, in MSCs during chondrogenic differentiation. We observed that overexpression of Wnt11 inhibited proliferation of MSCs and caused a G0/G1 cell cycle arrest. The expression level of chondrogenic markers, aggrecan and Collagen II, was significantly increased in MSCs transduced with Wnt11 as compared with non-transduced cells or MSCs transduced with the empty lentiviral vector. Furthermore, ectopic expression of Wnt11 stimulated gene expression of chondrogenic regulators, SRY-related gene 9, Runt-related transcription factor 2, and Indian hedgehog. Finally, Wnt11 overexpression promoted chondrogenic differentiation of MSCs in synergism with TGF-ß. Collectively, these results indicate that Wnt11 plays a crucial role in regulating MSC chondrogenic differentiation.

Lack of influence of an XRCC3 gene polymorphism on oral cancer susceptibility: meta-analysis.

BACKGROUND: To systematically summarize the association between the X-ray repair cross complementing 3 (XRCC3) gene polymorphism and oral cancer susceptibility by meta-analysis. MATERIALS AND METHODS: Databases including PubMed, EMbase, CNKI, VIP and WanFang Data were searched to identify case-control studies concerning the association between an XRCC3 gene polymorphism and the risk of oral cancer from the inception to June 2014. Two reviewers independently screened the literature according to the criteria, extracted the data and assessed the quality. Then meta-analysis was performed using Stata 11.0 software. RESULTS: Seven published case-control studies including 775 patients with oral cancer and 1922 controls were selected. Associations between the rs861539 polymorphism and overall oral cancer risk were not statistically significant in all kinds of comparison models (CT vs CC: OR=0.94, 95%CI=0.74-1.18; TT vs CC: OR=0.94, 95%CI=0.64- 1.38; dominant model: OR=0.95, 95%CI=0.76-1.18; recessive model: OR=0.94, 95%CI=0.69-1.29; allele T vs C: OR=0.97, 95%CI=0.84-1.11). In the stratified analysis by ethnicity, no significant associations were found among Asians and Caucasians. On stratification by tumor type, no significant associations were found for cancer and oral premalignant lesions. CONCLUSIONS: The XRCC3 gene polymorphism was not found to be associated with the risk of oral cancer. Considering the limited quality of the included case-control studies, more high quality studies with large sample size are needed to verify the above conclusion.

Association between polymorphisms in ERCC2 gene and oral cancer risk: evidence from a meta-analysis.

BACKGROUND: Excision repair cross-complementing group 2 (ERCC2) plays important roles in the repair of DNA damage and adducts. Single nucleotide polymorphisms (SNPs) of ERCC2 gene are suspected to influence the risks of oral cancer. We performed a meta-analysis to systematically summarize the possible association of ERCC2 rs1799793 and rs13181 polymorphisms with oral cancer risks. METHODS: We retrieved the relevant articles from PubMed and Embase databases. Studies were selected using specific criteria. ORs and 95% CIs were calculated to assess the association. All analyses were performed using the Stata software. RESULTS: Six studies were included in this meta-analysis. There were no significant associations between ERCC2 rs1799793 and rs13181 polymorphism with overall oral cancer risk. In the stratified analysis by ethnicity, no significant associations were found. In the stratified analysis by tumor type, the risk of oral leukoplakia was significant associated with rs13181 polymorphism (AC vs. AA: OR = 1.28, 95% CI = 1.01-1.62, P = 0.546 for heterogeneity, I² = 0.0%; CC vs. AA: OR = 1.94, 95% CI = 0.99-3.79, P = 0.057 for heterogeneity, I² = 60.1%; dominant model AC + CC vs. AA: OR = 1.35, 95% CI = 1.08-1.69, P = 0.303 for heterogeneity, I² = 17.6%; allele C vs. A: OR = 1.38, 95% CI = 1.04-1.82. P = 0.043 for heterogeneity, I² = 56.4%). CONCLUSION: Rs13181 in ERCC2 gene might be associated with oral leukoplakia risk.

Expression of TMPRSS4 in patients with salivary adenoid cystic carcinoma: correlation with clinicopathological features and prognosis.

Although there is growing evidence supporting the hypothesis that TMPRSS4 is linked with cancer susceptibility, the precise role of TMPRSS4 expression in salivary adenoid cystic carcinoma (SACC) is still unknown. The aim of this study was to examine TMPRSS4 expression in SACC and determine its associations with clinicopathological features and survival. TMPRSS4 expression in 125 SACC tissue and adjacent non-cancerous tissues was analyzed by immunohistochemistry and Western blotting. In addition, the correlation of TMPRSS4 expression with clinicopathological variables was evaluated. The prognostic value of TMPRSS4 for overall survival (OS) and disease-free survival (DFS) was determined by Kaplan-Meier estimates, and the significance of differences between curves was evaluated by the log-rank test. We found that high TMPRSS4 expression was predominantly observed in SACC tissues, but not in the adjacent normal salivary gland tissues. High TMPRSS4 expression in SACC tissues was correlated significantly with tumor TNM stage (P = 0.016), lymph node metastasis (P = 0.002) and distant metastasis (P < 0.001). While high TMPRSS4 expression was associated with poor OS (P = 0.019) and DFS (P = 0.031), Cox regression analysis also revealed that TMPRSS4 was an independent predictor of OS and DFS. These findings suggested that TMPRSS4 was involved in the pathogenesis of SACC and might indicate a poor prognosis for SACC patients.

Clinical anatomical study and evaluation of the use of the free anteromedial thigh perforator flaps in reconstructions of the head and neck.

The anteromedial thigh (AMT) perforator flap is usually thin, pliable, and nearly hairless, making it particularly suitable to repair defects of the head and neck. We studied the topography and outcomes of AMT perforator flaps in such defects after excision of tumours. We retrospectively reviewed the casenotes of 11 consecutive patients who had had reconstructions of the head and neck with the initial intent of using an AMT perforator flap from January 2010 to July 2011. For each patient we recorded the size and thickness of the flap; the length of the pedicle; and the number, external diameters, anatomical types, source vessels, and sites of the sizeable perforators. Of the 11 patients, 10 had successful reconstruction using AMT perforator flaps, but one had no AMT perforator big enough. The mean (range) number of sizeable perforators/flap was 1.3 (1-2), length of pedicle 10.6 (7-13) cm, and diameter of the artery 1.1(1.0-1.5) mm. Of the 13 sizeable perforators, 3 were direct and septocutaneous. The remaining ones were all musculocutaneous. Most of them were located in the middle third of the thigh. Primary closure of the donor site was achieved in all patients. One flap was successfully revised after compression of the perforator. All flaps survived with good functional and aesthetic outcomes. The free AMT perforator flap is suitable for reconstructions of the head and neck if a sizeable perforator can be found. The AMT flap may be used as a primary flap rather than as an alternative to the anterolateral thigh flap or a component of a chimeric flap.

[Perforator vascular anatomy and clinical application of the anteromedial thigh flap for head and neck reconstruction].

PURPOSE: A clinical study was undertaken to define the vascular anatomy of anteromedial thigh perforator flap (AMT) and evaluate the outcomes of the flap in head and neck reconstruction. METHODS: The sizable perforators of AMT flaps and their origins were prospectively explored in 54 patients. For each patient, we recorded the sizable perforators' location, diameter, source vessel, numbers and anatomical types. Among them, 14 cases underwent head and neck reconstruction with AMT flaps. The complications and functions of donor and recipient sites were recorded and the operative techniques of AMT were described. Statistical analysis was performed with SPSS 13.0 software package. RESULTS: Eight of fifty-four thighs had no sizable AMT perforators. AMT flap was based on the medial branch of descending branch of lateral circumflex femoral artery (d-LCFA) and shared the same vascular pedicle with anterolateral thigh flap (ALT). The total sizable perforators were 56. Among them, 40.9%(25/61) were direct septocutaneous perforators, the remaining perforators were all musculocutaneous. Most of the sizable perforators (58/61, 95.1%) were located in the middle one-third of the thigh, with an average of (3.9±0.72) cm medial to a line connecting the anterior superior iliac spine and the superolateral patella and an average of (22.5±2.38) cm to anterior superior iliac spine. There was an negtive relationship between the number of sizable perforators of AMT and ALT flaps (P<0.01). 14 flaps survived completely. No complications were observed in recipient and donor site. CONCLUSIONS: The pedicle of AMT flap is the medial branch of d-LCFA. The AMT flap may be useful if ALT flap is without sizable perforators. AMT flap may be as a primary or an alternative choice of anterolateral thigh flap for head and neck reconstruction.

[Overexpression of Twist1 promotes tumor invasion in human tongue squamous cell carcinoma cell line Tca8113].

AIM: To construct an eukaryotic expression vector of human Twist1 and investigate the relationship between Twist1 overexpression and tumor invasion in human tongue squamous cell carcinoma cell line Tca8113. METHODS: Total mRNA isolated from Tca8113 cells were reversely transcribed to cDNA. Human Twist1 was amplified using specific PCR primers and then subcloned into the pcDNA3.1-myc-hisA vector. The fusion expression plasmid was named Myc-Twist1. Myc-Twist1 was transfected into Tca8113 cells and examined by Western blotting. The localization of Twist1 in Tca8113 cells was observed using confocal laser scanning microscopy. E-cadherin promoter activity in response to Myc-Twist1 overexpression was measured by the dual luciferase reporter assay system. Transwell cell migration assay was performed to detect the invasive capacity of Tca8113 cells stably expressing Myc-Twist1. RESULTS: The fusion protein Myc-Twist1 was successfully constructed into eukaryotic expression vector. Western blotting showed that Myc-Twist1 was stably expressed in Tca8113 cells and it was localized mainly in the nucleus and a little in the cytoplasm. The Twist1 significantly inhibited the E-cadherin promoter activity and enhanced the cell invasion. CONCLUSION: Twist1 promotes tumor invasion by down-regulating E-cadherin expression in Tca8113 cells.

[Perforator-based chimeric anterolateral thigh flap for head and neck reconstruction after en bloc resection].

PURPOSE: To discuss and report the operative techniques for harvesting perforator-based chimeric flap in anterolateral thigh region and the advantages for head and neck reconstruction after en bloc resection. METHODS: A retrospective review was performed of perforator-based chimeric anterolateral thigh (ALT) flap for head and neck reconstruction since December of 2007 to March of 2011. 66 perforator-based chimeric flaps were harvested including a skin paddle and a muscular flap supplied by one mother pedicle-descending branch of lateral circumflex femoral artery(d-LCFA). 32 flaps were used for the mobile tongue and floor of mouth reconstruction, 30 flaps for base of the tongue and parapharyngeal walls, two for the buccal skin, one for hemimandible and one for parotid. The muscular flap were used to eliminate the dead space of submandibular area. Flaps size ranged from 7cm±4cm to 16cm±7cm and muscular flap was 3cm±4cm approximately. The complications and functions of both donor and recipient sites were recorded and the operative techniques of perforator-based chimeric flap elevation were generalized. RESULTS: All 65 flaps survived completely and the total survival percentage was 98.5%. Only one flap failed and was removed 5 days postoperatively. No complications(fistula, infection, hematoma, seroma et al) were observed in recipient and donor sites. Two anteromedial thigh flaps (AMT) were used for reconstruction due to no sizable perforators in the ALT region. All cases were followed up for 0.5-3 years. The flaps didn't atrophy after six months and the contour was satisfactory. The functions of speech and swallow were recovered well. All the donor sites were closed primarily and the scar was not obvious. The leg's function recovered well. CONCLUSIONS: Using a combination of retrograde and antegrade dissection is a safe and versatile method for harvesting a perforator-based chimeric flap. A chimeric flap including multiple components can meet the requirements of three-dimensional reconstruction. Perforator-based chimeric anterolateral thigh flap is one of the best choices for complex head and neck reconstruction after en bloc resection.