METTL3 suppresses anlotinib sensitivity by regulating m6A modification of FGFR3 in oral squamous cell carcinoma.

BACKGROUND: N6-methyladenosine (m6A) is an abundant nucleotide modification in mRNA, but there were few studies on its role in cancer drug sensitivity and resistance. Anlotinib has been proved to have effective antitumor effects in oral squamous cell carcinoma (OSCC) in our previous study. Here, we sought to investigate the treatment target of anlotinib and the function and mechanisms of m6A modification in regulating anlotinib effect in OSCC. METHODS: Anlotinib treatment in a dose-dependent manner, western blotting, qRT-PCR and cell lost-of-function assays were used to study the treatment target of anlotinib in OSCC. RNA m6A dot blot assays, the m6A MeRIP-seq and MeRIP-qPCR, RNA and protein stability assays were used to explore the m6A modification of the treatment target of anlotinib. Cell lost-of-function assays after METTL3 depletion were conducted to investigate the effect of m6A modification level on the therapeutic effect of anlotinib in OSCC. Patient-derived tumor xenograft (PDX) models and immunohistochemistry staining were performed to study the relationship of METTL3 and antitumor sensitivity of anlotinib in vivo. RESULTS: Anlotinib targeted FGFR3 in the treatment of OSCC and inhibited tumor cell proliferation and promoted apoptosis by inactivating the FGFR3/AKT/mTOR signaling pathway. METTL3 was identified to target and modify FGFR3 m6A methylation and then decrease the stability of mRNA. METTL3 expression level was related to the anlotinib sensitivity in OSCC cells in vitro and METTL3 knockdown promoted anlotinib sensitivity of OSCC cells by inhibiting the FGFR3 expression. PDX models samples furthermore showed that METTL3 and FGFR3 levels were tightly correlated with the anlotinib efficacy in OSCC. CONCLUSIONS: In summary, our work revealed that FGFR3 was served as the treatment target of anlotinib and METTL3-mediated FGFR3 m6A modification played a critical function in the anlotinib sensitivity in OSCC.

SOD2 confers anlotinib resistance via regulation of mitochondrial damage in OSCC.

OBJECTIVE: Previous studies had revealed that anlotinib had outstanding anti-tumor efficacy on oral squamous cell carcinoma. However, the underlying mechanism is still unclear. MATERIALS AND METHODS: Anlotinib resistant OSCC cells were established and analyzed by RNA-sequencing. The correlations between SOD2 expression and anlotinib resistance were investigated in OSCC cells and PDX models. Functional assays were performed to verify the SOD2 expression and anlotinib resistance in OSCC cells. RESULTS: Anlotinib resistant genes were enriched in the biological processes of mitochondrion organization and the gene pathway of reactive oxygen species. SOD2 expression level was positively correlated with the resistance of anlotinib in OSCC cells and PDX models. Higher SOD2 expression of OSCC cells was more resistant to anlotinib. Anlotinib induced ROS generation, apoptosis and mitochondrial damage in OSCC cells, which can be enhanced by SOD2 knockdown and decreased by SOD2 overexpression. Mitochondrial damage was identified as swelling and cristae disappearance morphology under TEM, decreased mitochondrial membrane potential and lower MFN2 expression. CONCLUSIONS: SOD2 may be capable of protecting mitochondria by downregulating ROS generation, which contributes to the resistance of anlotinib in OSCC cells. SOD2 can be utilized as a potential therapeutic target to improve the anti-cancer efficacy of anlotinib in OSCC.

Arecoline induces epithelial-mesenchymal transformation and promotes metastasis of oral cancer by SAA1 expression.

Arecoline, the main alkaloid of areca nut, is well known for its role in inducing submucosal fibrosis and oral squamous cell carcinoma (OSCC), however the mechanism remains unclear. The aim of this study was to establish an arecoline-induced epithelial-mesenchymal transformation (EMT) model of OSCC cells and to investigate the underlying mechanisms. CAL33 and UM2 cells were induced with arecoline to establish an EMT cell model and perform RNA-sequence screening. Luminex multiplex cytokine assays, western blot, and RT-qPCR were used to investigate the EMT mechanism. Arecoline at a concentration of 160 µg/ml was used to induce EMT in OSCC cells, which was confirmed using morphological analysis, transwell assays, and EMT marker detection. RNA-sequence screening and Luminex multiplex cytokine assays showed that many inflammatory cytokines (such as serum amyloid A1 [SAA1], interleukin [IL]-6, IL-36G, chemokine [CCL]2, and CCL20) were significantly altered during arecoline-induced EMT. Of these cytokines, SAA1 was the most highly upregulated. SAA1 overexpression induced EMT and promoted the migration and invasion of CAL33 cells, while SAA1 knockdown attenuated arecoline-induced EMT. Moreover, arecoline enhanced cervical lymph node metastasis in an orthotopic xenograft model of the tongue established using BALB/c nude mice. Our findings revealed that arecoline induced EMT and enhanced the metastatic capability of OSCC by the regulation of inflammatory cytokine secretion, especially that of SAA1. Our study provides a basis for understanding the mechanism of OSCC metastasis and suggests possible therapeutic targets to prevent the occurrence and development of OSCC associated with areca nut chewing.

METTL3 Promotes Tumorigenesis and Metastasis through BMI1 m6A Methylation in Oral Squamous Cell Carcinoma.

RNA modification plays an essential function in regulating gene expression and diverse biological processes. RNA modification enzyme methyltransferase-like 3 (METTL3) affects tumor progression by regulating the N6-methyladenosine (m6A) modification in the mRNAs of critical oncogenes or tumor suppressors, but its effect in oral squamous cell carcinoma (OSCC) remains unknown. In this study, we revealed that METTL3 was consistently upregulated in two OSCC cohorts, and high METTL3 expression was associated with poor prognosis. Functionally, cell proliferation, self-renewal, migration, and invasion ability in vitro and tumor growth and metastasis in vivo were decreased after METTL3 knockdown in OSCC cells. In contrast, the opposite results were obtained after METTL3 overexpression. In addition, the results obtained with the Mettl3 genetically modified mouse model validated the essential role of Mettl3 in chemical-induced oral carcinogenesis. In mechanism, methylated RNA immunoprecipitation sequencing (MeRIP-seq), MeRIP-quantitative real-time PCR, and luciferase reporter and mutagenesis assays identified that METTL3 mediates the m6A modification in the 3' UTR of BMI1 mRNA. METTL3 promotes BMI1 translation in OSCC under the cooperation with m6A reader IGF2BP1. Our findings revealed that METTL3 promotes OSCC proliferation and metastasis through BMI1 m6A methylation, suggesting that the METTL3-m6A-BMI1 axis may serve as a prognostic biomarker or therapeutic target in patients with OSCC.

The serum biomarker chemerin promotes tumorigenesis and metastasis in oral squamous cell carcinoma.

Chemerin, which is encoded by retinoic acid receptor responder 2 (RARRES2), has been found to be related to malignant tumours, but its role in the development of oral squamous cell carcinoma (OSCC) is largely unexplored. In the present study, a higher serum level of chemerin was evident in patients with OSCC than in healthy individuals, and this high level of chemerin significantly decreased after tumour resection. In addition, high chemerin levels were positively associated with advanced tumour stage and lymph node metastasis. The expression levels of chemerin and Chemerin Receptor 23 (ChemR23) were positively correlated with the migration and invasion of OSCC cell lines. Recombinant chemerin (R-chemerin) enhanced the in vitro migration, invasion and proliferation of OSCC cells in a concentration-dependent manner, and short hairpin RNAs (shRNAs) targeting RARRES2 decreased chemerin expression and inhibited OSCC cell metastasis and proliferation both in vitro and in vivo Additionally, R-chemerin activated manganese superoxide dismutase (SOD2) and increased the amount of intracellular hydrogen peroxide (H2O2), leading to a significant decrease in E-cadherin expression and dramatic increase in the expression of phosphorylated ERK1/2 (p-ERK1/2), Slug, Vimentin and N-cadherin, but shRNAs targeting RARRES2 reversed these effects. Moreover, knockdown of ChemR23 with small interfering RNAs (siRNA) significantly inhibited chemerin-induced OSCC cell migration/invasion and SOD2 activity. Our results revealed that chemerin is a novel biomarker for OSCC. Chemerin/ChemR23 promotes tumorigenesis and metastasis in OSCC and may be a new therapeutic target for OSCC.

Up-regulation of INSR/IGF1R by C-myc promotes TSCC tumorigenesis and metastasis through the NF-κB pathway.

The insulin receptor (INSR) and insulin-like growth factor 1 receptor (IGF1R) have been reported to be involved in the tumorigenesis and metastasis of various malignancies. The aim of our study was to investigate and compare the effects of INSR and IGF1R on the tumorigenesis and metastasis of tongue squamous cell carcinoma (TSCC) and explore the possible mechanism(s) involved. We found that INSR had the same up-regulated expression pattern as IGF1R in TSCC tissues. INSR and IGF1R up-regulation were correlated with each other and associated with lymph node metastasis and poor prognosis. Functional studies established that knocking down either INSR or IGF1R dramatically impeded TSCC cell proliferation, migration, and invasion in vitro and tumorigenesis and tumor metastasis in vivo, whereas ectopic overexpression of INSR or IGF1R enhanced these activities. Both INSR and IGF1R directly targeted p65 and activated the NF-κB pathway; furthermore, C-myc was observed to directly bind to the INSR and IGF1R promoters and up-regulates INSR and IGF1R expression in TSCC. Thus, our current data demonstrate that both INSR and IGF1R are directly targeted by C-myc and exert similar effects to promote the tumorigenesis and metastasis of TSCC through the NF-κB pathway. Therefore, INSR and IGF1R may be therapeutic target genes and potential prognostic factors for TSCC.

Dysregulation of AKT1, a miR-138 target gene, is involved in the migration and invasion of tongue squamous cell carcinoma.

BACKGROUND: AKT1, also known as PKBα, is abnormally expressed in various malignancies. In this study, we aimed to evaluate the role of AKT1 in the tongue squamous cell carcinoma (TSCC) and further clarify the mechanisms of AKT1 in the migration and invasion of TSCC. METHODS: At first, immunohistochemistry (IHC) was conducted to detect the expression of AKT1 in TSCC. Then, we determined the role of AKT1 in the migration and invasion of TSCC and further investigated whether AKT1 was the target gene of miR-138 using dual luciferase reporter assays and Western blot. RESULTS: Immunohistochemistry results suggested that AKT1 dysregulation was a frequent event in TSCC, and upregulation of AKT1 was correlated with lymph node metastasis and associated with reduced overall survival. UM1 cells with higher migratory and invasive abilities had more robust AKT1 protein expression than UM2 cells with lower migratory and invasive abilities. The migration and invasion abilities were inhibited in UM1 cells upon AKT1 knockdown, meanwhile resulted in a decline of metastasis-related proteins (vimentin, slug, and pERK1/2), and upregulation of E-cadherin. Luciferase assays revealed that AKT1 was directly targeted by miR-138, and ectopic transfection of miR-138 reduced the expression of AKT1 protein. CONCLUSIONS: Our results confirm that upregulation of AKT1, a miR-138 target gene, is a frequent event in TSCC and contributes to the aggressive behaviors and poor prognosis of TSCC.

Nanotubular topography enhances the bioactivity of titanium implants.

Surface modification on titanium implants plays an important role in promoting mesenchymal stem cell (MSC) response to enhance osseointegration persistently. In this study, nano-scale TiO2 nanotube topography (TNT), micro-scale sand blasted-acid etched topography (SLA), and hybrid sand blasted-acid etched/nanotube topography (SLA/TNT) were fabricated on the surfaces of titanium implants. Although the initial cell adherence at 60 min among TNT, SLA and TNT/SLA was not different, SLA and SLA/TNT presented to be rougher and suppressed the proliferation of MSC. TNT showed hydrophilic surface and balanced promotion of cellular functions. After being implanted in rabbit femur models, TNT displayed the best osteogenesis inducing ability as well as strong bonding strength to the substrate. These results indicate that nano-scale TNT provides favorable surface topography for improving the clinical performance of endosseous implants compared with micro and hybrid micro/nano surfaces, suggesting a promising and reliable surface modification strategy of titanium implants for clinical application.

MicroRNA-21 regulates prostaglandin E2 signaling pathway by targeting 15-hydroxyprostaglandin dehydrogenase in tongue squamous cell carcinoma.

BACKGROUND: Oral tongue squamous cell carcinoma (OTSCC) is one of the most aggressive forms of head and neck/oral cancer (HNOC), and is a complex disease with extensive genetic and epigenetic defects, including microRNA deregulation. Identifying the deregulation of microRNA-mRNA regulatory modules (MRMs) is crucial for understanding the role of microRNA in OTSCC. METHODS: A comprehensive bioinformatics analysis was performed to identify MRMs in HNOC by examining the correlation among differentially expressed microRNA and mRNA profiling datasets and integrating with 12 different sequence-based microRNA target prediction algorithms. Confirmation experiments were performed to further assess the correlation among MRMs using OTSCC patient samples and HNOC cell lines. Functional analyses were performed to validate one of the identified MRMs: miR-21-15-Hydroxyprostaglandin Dehydrogenase (HPGD) regulatory module. RESULTS: Our bioinformatics analysis revealed 53 MRMs that are deregulated in HNOC. Four high confidence MRMs were further defined by confirmation experiments using OTSCC patient samples and HNOC cell lines, including miR-21-HPGD regulatory module. HPGD is a known anti-tumorigenic effecter, and it regulates the tumorigenic actions of Prostaglandin E2 (PGE2) by converts PGE2 to its biologically inactive metabolite. Ectopic transfection of miR-21 reduced the expression of HPGD in OTSCC cell lines, and the direct targeting of the miR-21 to the HPGD mRNA was confirmed using a luciferase reporter gene assay. The PGE2-mediated upregulation of miR-21 was also confirmed which suggested the existence of a positive feed-forward loop that involves miR-21, HPGD and PGE2 in OTSCC cells that contribute to tumorigenesis. CONCLUSIONS: We identified a number of high-confidence MRMs in OTSCC, including miR-21-HPGD regulatory module, which may play an important role in the miR-21-HPGD-PGE2 feed-forward loop that contributes to tumorigenesis.

PRL-3 promotes migration and invasion and is associated with poor prognosis in salivary adenoid cystic carcinoma.

BACKGROUND: PRL-3 had been found to be involved in tumorigenesis in various malignancies. In this study, we investigated the role of PRL-3 in the development, migration, and invasion of salivary adenoid cystic carcinoma (SACC). METHODS: Immunohistochemistry (IHC) was used to analyze the role of PRL-3 in the development and prognosis of SACC. Then, we overexpressed or inhibited the expression of PRL-3 in paired SACC cells to analyze the role of PRL-3 in the migration and invasion of SACC. In vitro migration and invasion assays were used. Western blotting was used to detect metastasis-related protein levels. RESULTS: IHC results confirmed that the deregulation of PRL-3 was a frequent event in SACC; the upregulation of PRL-3 was related to clinical stages, vital status, and distant metastasis, which was associated with reduced overall survival and disease-free survival. SACC-LM cells with higher migratory and invasive abilities had more robust PRL-3 protein expression than SACC-83 cells with lower migratory and invasive abilities. PRL-3 overexpression promoted cell migration, invasion, and proliferation, led to simultaneous upregulation of phosphorylated PRL-3, pERK1/2, Slug, vimentin, and downregulation of E-cadherin in SACC-83 cells. However, the inhibition of PRL-3 by PRL-3 inhibitor or PRL-3 siRNA in SACC-LM cells inhibited cell migration, invasion, and proliferation, resulted in simultaneous downregulation of phosphorylated PRL-3, pERK1/2, Slug, vimentin, and upregulation of E-cadherin. CONCLUSIONS: Our results confirm that PRL-3 plays an important role in the development of SACC and contributes to the migratory and invasive abilities of SACC.

WAVE3 Induces EMT and Promotes Migration and Invasion in Intrahepatic Cholangiocarcinoma.

BACKGROUND: Wiskott-Aldrich syndrome protein family verprolin-homologous protein 3 (WAVE3) plays a critical role in cancer progression and metastasis. However, the specific role of WAVE3 in intrahepatic cholangiocarcinoma (ICC) has not been studied. AIMS: This study aimed to explore the role and mechanism of WAVE3 in the progression and metastasis of ICC. METHODS: The expression of WAVE3 in ICC tissues and adjacent non-cancerous tissues was detected by immunohistochemistry. Western blot analysis was utilized to detect the expression of WAVE3 in ICC cells. A transwell assay was used to assess the potential for migration and invasion. The expression of WAVE3 in CC-LP-1 cells was knocked down by small interfering RNA (siRNA) interference. RESULTS: The expression of WAVE3 in ICC tissues was significantly higher than that in adjacent non-cancerous tissues. The overall survival was lower in the subgroup of ICC patients with higher WAVE3 expression compared to the subgroup with a lower level of WAVE3 expression. WAVE3 expression was an adverse prognostic factor for ICC patients. CC-LP-1 cells expressed higher levels of WAVE3 protein compared to RBE cells and human intrahepatic biliary epithelial cells, which correlated with greater migration and invasion capabilities compared with the RBE cells. After the transfection of CC-LP-1 cells with WAVE3 siRNA, the level of WAVE3 protein was significantly decreased, accompanied by a marked reduction in migration, invasion and proliferation. Moreover, after the knockdown of WAVE3 expression in CC-LP-1 cells, the protein levels of Slug and Vimentin were significantly decreased, while that of E-cadherin was significantly increased. CONCLUSIONS: WAVE3 may represent a new adverse prognostic factor for patients with ICC. This protein enhances migration and invasion capabilities in ICC, most likely through the induction of an epithelial-mesenchymal transition.

Deregulation of Bmi-1 is associated with enhanced migration, invasion and poor prognosis in salivary adenoid cystic carcinoma.

BACKGROUND: Bmi-1 had been found to involve in self renewal of stem cells and tumorigenesis in various malignancies. In this study, we investigated the role of Bmi-1 in the development of salivary adenoid cystic carcinoma (SACC). METHODS: At first, we confirmed that the deregulation of Bmi-1 was a frequent event in SACC; up-regulation of Bmi-1 was correlated with clinical stages, vital status and distant metastasis and associated with reduced overall survival and disease free survival. SACC-LM cells, higher migration and invasion abilities, elevated the expression of Bmi-1 protein, epithelial-mesenchymal transition (EMT) related proteins (Snail, Slug and Vimentin) and cancer stem cells (CSCs) related proteins (ABCG2, Notch, ALDH-1, Oct-4, Nanog and Epcam) compared to the SACC-83 cells (lower migration and invasion abilities). The migration and invasion abilities were inhibited in SACC-LM cells upon Bmi-1 knockdown. Meanwhile, Bmi-1 knockdown resulted in simultaneous loss of stem cell markers and EMT markers in SACC-LM cells. CONCLUSION: Our studies confirm that Bmi-1 deregulation plays an important role in the development of SACC and contributes to the migration and the invasion abilities of SACC, which is involved in EMT and CSCs. GENERAL SIGNIFICANCE: To our knowledge, this is the first study revealing that Bmi-1 deregulation is associated with enhanced migration, invasion and poor prognosis in salivary adenoid cystic carcinoma.

MTUS1/ATIP3a down-regulation is associated with enhanced migration, invasion and poor prognosis in salivary adenoid cystic carcinoma.

BACKGROUND: Microtubule-associated tumor suppressor gene (MTUS1) has been identified as tumor suppressor gene in many malignant tumors. In this study, we investigated the role of MTUS1 in the development of salivary adenoid cystic carcinoma (SACC) and its functional effect on the migration and invasion of SACC. METHODS: Archival clinical samples including 49 primary SACC were examined for MTUS1 expression by immunohistochemistry. Statistical analyses were performed to evaluate the correlation between MTUS1 with histopathological features and survival. The expression of MTUS1/ATIP (AT2 receptor-interacting protein) isoforms was determined in SACC tissue samples and cell lines using quantitative RT-PCR assays. Then we investigated whether the migration and invasion of SACC were mediated by MTUS1/ATIP3a using in vitro cell migration and invasion assay. RESULTS: We confirmed that the down-regulation of MTUS1 was a frequent event in SACC, and was correlated with distant metastasis and associated with reduced overall survival and disease free survival. Isoform specific quantitative RT-PCR assays revealed that ATIP1, ATIP3a and ATIP3b were the major isoforms of the MTUS1 gene products in SACC, and were significant down-regulation in SACC as compared to matching normal tissues. For functional analyses, we found that SACC-LM cells (SACC cell line with higher migration and invasion ability) possessed a lower expression level of ATIP3a compared to SACC-83 cells (lower migration and invasion ability). Restoration of ATIP3a expression in SACC-LM cells induced anti-proliferative activity and inhibited the migration and invasion ability. Knockdown of ATIP3a promoted the proliferation, migration and invasion ability of SACC-83 cells. Restoration of ATIP3a inhibited the phosphorylation of ERK (extracellular-regulated kinase) 1/2, the expression of Slug and Vimentin in SACC-LM cells, while knockdown of ATIP3a increased the phosphorylation of ERK1/2, the expression of Slug and Vimentin in SACC-83 cells. CONCLUSIONS: Our studies confirm that MTUS1 plays an important role in the progression of SACC, and may serve as a biomarker or therapeutic target for patients with SACC. MTUS1/ATIP3a down-regulation contributes to the proliferation, migration and the invasion abilities of SACC.

MicroRNA-181a suppresses salivary adenoid cystic carcinoma metastasis by targeting MAPK-Snai2 pathway.

BACKGROUND: To date microRNAs and their contribution to the onset and propagation of salivary adenoid cystic carcinoma (SACC) are limited. The objective of this study was to identify miR-181a and its mechanism in the metastasis of SACC. METHODS: At first microarray and quantitative RT-PCR were used to investigate microRNA profiles and miR-181a in paired SACC cell lines with different metastatic potential. Then the effect of miR-181a on metastatic potential of SACC was investigated. MiR-181a target genes and Snai2 promoter activity were investigated using luciferase reporter gene assays. Western blot was used to detect MAPK-Snai2 pathway-related protein level. RESULTS: A panel of deregulated microRNAs (including miR-181a) was identified in paired of SACC cell lines. Functional analysis indicated that miR-181a inhibited SACC cell migration, invasion and proliferation in vitro, and it suppressed tumor growth and lung metastasis in vivo. Direct targeting of miR-181a to MAP2K1, MAPK1 and Snai2 was confirmed by luciferase reporter gene assays. MiR-181a mimic inhibited the expression of MAP2K1, MAPK1 and Snai2 in SACC cells. MAP2K1 or MAPK1 siRNA suppressed Snai2 gene promoter activity and reduced Snai2 expression and the metastatic potential of SACC cells. CONCLUSIONS: Our results indicate that miR-181a plays an important role in the metastasis of SACC, and may serve as a novel therapeutic target for SACC. MiR-181a regulates the MAPK-Snai2 pathway both through direct cis-regulatory mechanism and through indirect trans-regulatory mechanism. GENERAL SIGNIFICANCE: To our knowledge, this is the first study revealing that miR-181a deregulation mediated the metastasis of SACC by regulating MAPK-Snai2 pathway.

In vitro effect of iASPP on cell growth of oral tongue squamous cell carcinoma.

iASPP is an inhibitory member of the apoptosis-stimulating proteins of P53 (ASPP) family. iASPP is over expressed in several malignant tumors and potentially affects cancer progression. However, the expression and potential role of iASPP in oral tongue squamous cell carcinoma (OTSCC) have not been addressed. In our study, we detected iASPP expression in OTSCC by immunohistochemistry. iASPP expression is up-regulated in OTSCC tissues. Moreover, in clinical pathology specimens, we found that increased iASPP expression correlates with poor differentiation and lymph node metastasis. Using multicellular tumor spheroids (MTS) and flow cytometry, we demonstrated that iASPP down-regulation arrests OTSCC cells at the G0/G1 phase, induces OTSCC cell apoptosis and inhibits OTSCC cell proliferation. These results indicate that iASPP plays a significant role in the progression of OTSCC and may serve as a biomarker or therapeutic target for OTSCC patients.

Mitochondrial outer membrane protein MTUS1/ATIP1 exerts antitumor effects through ROS-induced mitochondrial pyroptosis in head and neck squamous cell carcinoma.

We showed that microtubule-associated tumor suppressor gene (MTUS1/ATIP) downregulation correlated with poor survival in head and neck squamous cell carcinoma (HNSCC) patients and that MTUS1/ATIP1 was the most abundant isoform in HNSCC tissue. However, the location and function of MTUS1/ATIP1 have remain unclear. In this study, we confirmed that MTUS1/ATIP1 inhibited proliferation, growth and metastasis in HNSCC in cell- and patient-derived xenograft models in vitro and in vivo. MTUS1/ATIP1 localized in the outer mitochondrial membrane, influence the morphology, movement and metabolism of mitochondria and stimulated oxidative stress in HNSCC cells by directly interacting with MFN2. MTUS1/ATIP1 activated ROS, recruiting Bax to mitochondria, facilitating cytochrome c release to the cytosol to activate caspase-3, and inducing GSDME-dependent pyroptotic death in HNSCC cells. Our findings showed that MTUS1/ATIP1 localized in the outer mitochondrial membrane in HNSCC cells and mediated anticancer effects through ROS-induced pyroptosis, which may provide a novel therapeutic strategy for HNSCC treatment.

Metabolic reprogramming driven by METTL1-mediated tRNA m7G modification promotes acquired anlotinib resistance in oral squamous cell carcinoma.

Tyrosine kinase inhibitors (TKIs) are frequently utilized in the management of malignant tumors. Studies have indicated that anlotinib has a significant inhibitory effect on oral squamous cell carcinoma (OSCC). However, the mechanisms underlying the development of resistance with long-term anlotinib treatment remain obscure. Our research found that METTL1 expression was heightened in anlotinib-resistant OSCC cells. We observed that METTL1 played a role in fostering resistance to anlotinib in both transgenic mouse models and in vitro. Mechanistically, the elevated METTL1 levels in anlotinib-resistant OSCC cells contributed to enhanced global mRNA translation and stimulated oxidative phosphorylation (OXPHOS) through m7G tRNA modification. Bioenergetic profiling demonstrated that METTL1 drived a metabolic shift from glycolysis to OXPHOS in anlotinib-resistant OSCC cells. Additionally, inhibition of OXPHOS biochemically negated METTL1's impact on anlotinib resistance. Overall, this study underscores the pivotal role of METTL1-mediated m7G tRNA modification in anlotinib resistance and lays the groundwork for novel therapeutic interventions to counteract resistance in OSCC.

The fusiform skin paddle in fibula free flap: a fusiform-designed skin paddle for maxillofacial soft defect reconstruction and reducing leg wound tension.

Objective: To investigate the feasibility of leg wound closure and reconstruction of maxillofacial soft defect by a fusiform-designed skin paddle in fibula free flap (FFF). Methods: Fifty patients who underwent FFF for reconstruction of maxillofacial soft defect were divided into two groups. The fusiform group (20 patients) was treated using a fusiform-designed skin paddle in FFF (skin paddle width less than 2 cm), and leg wound was closed using primary suturing. Reconstruction of the maxillofacial soft defect or filling of dead space was achieved by folding the fusiform skin paddle. The conventional group (30 patients) was treated using the conventional-designed skin paddle (skin paddle width no less than 2.5 cm). The leg wound was closed using mattress suturing or skin graft, while reconstruction of the maxillofacial soft defect or filling of dead space by conventional way. The average postoperative length of hospital stay, healing time of leg wound, and post-surgical complications were recorded at least 6 months after the surgery. Results: Compared with traditional method, the fusiform-designed skin paddle reduced the average healing time of the leg wound (fusiform group: 11.05 days, conventional group: 14.77 days, P < 0.05). The average length-to-width ratio in fusiform group was significantly greater than that of in conventional group (fusiform group: 5.85, conventional group: 2.93, P < 0.05), and no difference was observed on the graft size of skin paddle between two groups (fusiform group: 23.13, conventional group: 27.13, P > 0.05). The post-surgical early complications of the leg wound in the conventional group were higher than that of in the fusiform group (fusiform group: 0%, conventional group: 6.67%), while the post-surgical late complication of the donor site between the two groups showed no case. Healing disorders of maxillofacial soft reconstruction in the conventional group were higher than that of in the fusiform group (fusiform group: 5.26%, conventional group: 20.69%). Conclusions: Fusiform-designed skin paddle for closure of the leg wound and maxillofacial soft defect is a feasible alternative to the conventional- designed skin paddle. The fusiform- designed skin paddle resulted in the less postoperative length of hospital stay, shorter healing time of leg wound and less complication.

miR-181a-Twist1 pathway in the chemoresistance of tongue squamous cell carcinoma.

Although many researches have been undertaken to disclose the mechanisms of chemoresistance, the mechanisms remain unclear. The aim of this study is to elucidate the role of miR-181a-Twist1 pathway in the chemoresistance of tongue squamous cell carcinoma (TSCC). We found that cisplatin-induced chemoresistance in TSCC cell lines underwent EMT (epithelial-mesenchymal transition) and was accompanied by enhancing metastatic potential (migration and invasion in vitro), miR-181a downregulation and Twist1 upregulation. Functional analyses indicated that miR-181a reversed chemoresistance, inhibited EMT and metastatic potential in TSCC cells. Twist1 was confirmed as a direct miR-181a target gene by luciferase reporter gene assays. Twist1 knockdown by siRNA led to a reversal of the chemoresistance, inhibited EMT and metastatic potential in TSCC cells. Our study demonstrates that miR-181a-Twist1 pathway may play an important role in the development of cisplatin-chemoresistance, with EMT and an increase the metastatic potential of TSCC cells.

Polycomb group protein EZH2-mediated E-cadherin repression promotes metastasis of oral tongue squamous cell carcinoma.

Enhancer of Zeste homolog 2 (EZH2) is a critical component of the polycomb-repressive complex 2 (PRC2) that regulates many essential biological processes, including embryogenesis and many developmental events. The oncogenic role of EZH2 has recently been implicated in several cancer types. In this study, we first confirmed that the over-expression of EZH2 is a frequent event in oral tongue squamous cell carcinoma (OTSCC). We further demonstrated that EZH2 over-expression is correlated with advanced stages of the disease and is associated with lymph node metastasis. Statistical analysis revealed that EZH2 over-expression was correlated with reduced overall survival. Furthermore, over-expression of EZH2 was correlated with reduced expression of tumor suppressor gene E-cadherin. These observations were confirmed in vitro, in which knockdown of EZH2-induced E-cadherin expression and reduced cell migration and invasion. In contrast, ectopic transfection of EZH2 led to reduced E-cadherin expression and enhanced cell migration and invasion. Furthermore, EZH2 may act on cell migration in part by suppressing the E-cadherin expression. Taken together, these data suggest that EZH2 plays major roles in the progression of OTSCC, and may serve as a biomarker or therapeutic target for patients at risk of metastasis.