Identification of immunogenic cell death-related damage-related molecular patterns (DAMPs) to predict outcomes in patients with head and neck squamous cell carcinoma.

PURPOSE: Head and neck cancer is the sixth most common type of cancer worldwide, wherein the immune responses are closely associated with disease occurrence, development, and prognosis. Investigation of the role of immunogenic cell death-related genes (ICDGs) in adaptive immune response activation may provide cues into the mechanism underlying the outcome of HNSCC immunotherapy. METHODS: ICDGs expression patterns in HNSCC were analyzed, after which consensus clustering in HNSCC cohort conducted. A 4-gene prognostic model was constructed through LASSO and Cox regression analyses to analyze the prognostic index using the TCGA dataset, followed by validation with two GEO datasets. The distribution of immune cells and the response to immunotherapy were compared between different risk subtypes through multiple algorithms. Moreover, immunohistochemical (IHC) analyses were conducted to validate the prognostic value of HSP90AA1 as a predictor of HNSCC patient prognosis. In vitro assays were performed to further detect the effect of HSP90AA1 in the development of HNSCC. RESULTS: A novel prognostic index based on four ICDGs was constructed and proved to be useful as an independent factor of HNSCC prognosis. The risk score derived from this model grouped patients into high- and low-risk subtypes, wherein the high-risk subtype had worse survival outcomes and poorer immunotherapy response. IHC analysis validated the applicability of HSP90AA1 as a predictor of prognosis of HNSCC patients. HSP90AA1 expression in tumor cells promotes the progression of HNSCC. CONCLUSIONS: Together, these results highlight a novel four-gene prognostic signature as a valuable tool to assess survival status and prognosis of HNSCC patients.

Neutrophil extracellular traps promote invasion and metastasis via NLRP3-mediated oral squamous cell carcinoma pyroptosis inhibition.

Neutrophil extracellular traps (NETs) are reticular structures composed of neutrophil elastase (NE), cathepsin G (CG) and DNA-histone enzyme complexes. Accumulating evidence has revealed that NETs play important roles in tumor progression, metastasis, and thrombosis. However, our understanding of its clinical value and mechanism of action in oral squamous cell carcinoma (OSCC) is limited and has not yet been systematically described. Here, we aimed to investigate the clinical significance of NETs in OSCC and the mechanisms by which they affect its invasive and metastatic capacity. Our results demonstrated that high enrichment of NETs is associated with poor prognosis in OSCC, and mechanistic studies have shown that NE in NETs promotes invasion and metastasis via NLRP3-mediated inhibition of pyroptosis in OSCC. These findings may provide a new therapeutic approach for OSCC.

A novel fluorescent traceable carbon quantum dots with selective antibacterial activity against Porphyromonas gingivalis.

Antibiotics can kill bacteria, but their continued use can easily lead to drug resistance, particularly the main pathogenic bacteria of periodontitis, Porphyromonas gingivalis. However, to avoid drug resistance, carbon quantum dots (CDs) have great potential as a bioactive material in antimicrobial therapy. Herein, we use ornidazole as raw material to prepare CDs of different sizes by microwave irradiation and screen CDs with fluorescence and bacteriostatic properties. The inhibition experiments and live/dead assays of P. gingivalis exhibited outstanding antibacterial effects. This research aimed to develop nano-level antibacterial active materials that also have fluorescence traceability. This study offers a different method for the development of multifunctional CDs, provides valuable strategies for the treatment of diseases associated with P. gingivalis, and predicts great application prospects in the field of biomedicine.

An outcomes-based module education via flipped classroom enhances undergraduate oral histopathology learning.

INTRODUCTION: Oral histopathology is a bridge course connecting oral basic medicine and clinical dentistry. However, the application of outcomes-based education via flipped classroom (FC) in oral histopathology has not been well explored. This study has assessed the efficacy of outcomes-based education via FC in undergraduate oral histopathology module learning in Nanjing Medical University of China. MATERIALS AND METHODS: A total of 214 third-year students were enrolled and assigned to the FC group of the batch 2022-23 (n = 110) and the traditional classroom (TC) group of the batch 2021-22 (n = 104) to participate the oral histopathology sessions respectively in the study. The FC group were required to preview the online course materials pre-class, followed by in-class quizz, in-class interactive group discussion, and slides microscopic observation. The outcomes-based formative and summative assessments for FC were designed. The TC group attended traditional laboratory class for the same glass slides microscopic observation. In addition, a questionnaire was performed to investigate the satisfaction of learning. Along with this, the performances of FC group in written theory tests and oral histopathology slide tests were compared with TC group. RESULTS: Students in the FC group gained significantly final higher scores of the course than those in the TC group (score: 83.79 ± 11 vs. 76.73 ± 10.93, P<0.0001). Data from the student questionnaires indicated a preference for outcomes-based module education via FC. In the questionnaires, most students considered outcomes-based module education via FC to be beneficial to learning motivation, knowledge comprehension, critical thinking and teamwork. FC group had a higher level of satisfaction with oral histopathology teaching than TC group (satisfaction score: 4.599 ± 0.1027 vs. 4.423 ± 0.01366, P<0.01). CONCLUSION: An outcomes-based module education via FC has a promising effect on undergraduate oral histopathology education.

Impaired Efferocytosis Enables Apoptotic Osteoblasts to Escape Osteoimmune Surveillance During Aging.

Macrophage efferocytosis of apoptotic osteoblasts (apoOBs) is a key osteoimmune process for bone homeostasis. However, apoOBs frequently accumulate in aged bone marrow, where they may mount proinflammatory responses and progressive bone loss. The reason why apoOBs are not cleared during aging remains unclear. In this study, it is demonstrated that aged apoOBs upregulate the immune checkpoint molecule CD47, which is controlled by SIRT6-regulated transcriptional pausing, to evade clearance by macrophages. Using osteoblast- and myeloid-specific gene knockout mice, SIRT6 is further revealed to be a critical modulator for apoOBs clearance via targeting CD47-SIRPα checkpoint. Moreover, apoOBs activate SIRT6-mediated chemotaxis to recruit macrophages by releasing apoptotic vesicles. Two targeting delivery strategies are developed to enhance SIRT6 activity, resulting in rejuvenated apoOBs clearance and delayed age-related bone loss. Collectively, the findings reveal a previously unknown linkage between immune surveillance and bone homeostasis and targeting the SIRT6-regulated mechanism can be a promising therapeutic strategy for age-related bone diseases.

AuNP-Loaded Electrospinning Membrane Cooperated with CDs for Periodontal Tissue Engineering.

BACKGROUND: Guided bone regeneration (GBR) is commonly used to regenerate periodontal tissue. However, the bone inductivity and antibacterial properties of the GBR membranes currently in use are severely limited. This issue can be resolved by loading growth factors and antibiotics. Bioactive substitutes, such as Au nanoparticles (AuNPs) and carbon quantum dots (CDs), were proposed to prevent the denaturation of osteogenic growth factors and the induction of antibacterial drug resistance. METHODS: Ornidazole was initially used as the raw material to prepare the CDs, followed by the incorporation of an optimal ratio of nanoparticles to produce the electrospun membrane doped with AuNPs and novel traceable antibacterial CDs. The morphology of the membrane was characterized. The adhesion, proliferation, and osteogenic differentiation of cells on the membrane were evaluated in vitro. The antimicrobial characteristics of the membrane were also investigated. The electrospun membrane was implanted into a rat skull defect model in vivo to investigate its osteogenic potential. RESULTS: The blending of nanomaterials did not affect the micro morphology of the fiber, resulting in enhanced mechanical properties. Membranes doped with AuNPs and CDs exhibited excellent biocompatibility, increased ALP activity, improved calcified nodules, and increased expression of osteogenic-associated proteins, in addition to pronounced antibacterial effects. The membrane also demonstrated excellent osteogenic characteristics in rat models. CONCLUSION: The synergistic effect of loaded AuNPs electrospun fiber membrane with CDs can promote periodontal bone regeneration and exert antibacterial activity.

Oral pathogen aggravates atherosclerosis by inducing smooth muscle cell apoptosis and repressing macrophage efferocytosis.

Periodontitis imparting the increased risk of atherosclerotic cardiovascular diseases is partially due to the immune subversion of the oral pathogen, particularly the Porphyromonas gingivalis (P. gingivalis), by inducing apoptosis. However, it remains obscure whether accumulated apoptotic cells in P. gingivalis-accelerated plaque formation are associated with impaired macrophage clearance. Here, we show that smooth muscle cells (SMCs) have a greater susceptibility to P. gingivalis-induced apoptosis than endothelial cells through TLR2 pathway activation. Meanwhile, large amounts of miR-143/145 in P.gingivalis-infected SMCs are extracellularly released and captured by macrophages. Then, these miR-143/145 are translocated into the nucleus to promote Siglec-G transcription, which represses macrophage efferocytosis. By constructing three genetic mouse models, we further confirm the in vivo roles of TLR2 and miR-143/145 in P. gingivalis-accelerated atherosclerosis. Therapeutically, we develop P.gingivalis-pretreated macrophage membranes to coat metronidazole and anti-Siglec-G antibodies for treating atherosclerosis and periodontitis simultaneously. Our findings extend the knowledge of the mechanism and therapeutic strategy in oral pathogen-associated systemic diseases.

Mirror training device improves dental students' performance on virtual simulation dental training system.

INTRODUCTION: Clinical practice of dentistry entails the use of indirect vision using a dental mirror. The Mirrosistant is a device that helps dental students become proficient with use of indirect vision mirror operation. This study aimed to explore the role of the Mirrosistant on students' performance with the virtual simulation dental training system. MATERIALS AND METHODS: A total of 72 dental students were equally assigned to the Control group and the Experimental group. Subsequently, Mirrosistant was used to conduct a series of mirror training exercises in the Experimental group. The training consisted of tracing the edge and filling in the blank of the prescribed shape, as well as preparing the specified figure on raw eggs using indirect vision via Mirrosistant. Next, both groups were examined using the SIMODONT system, a virtual reality dental trainer, for mirror operation. In addition, a five-point Likert scale questionnaire was used to assess student feedback by using Mirrosistant. RESULTS: The mirror operation examination conducted by the SIMODONT system revealed that mirror training using Mirrosistant had statistically improved students' performances (score: 80.42 ± 6.43 vs. 69.89 ± 15.98, P = 0.0005) and shorten their performance time of mirror operation (time of seconds: 243.28 ± 132.83 vs. 328.53 ± 111.89, P = 0.0013). Furthermore, the questionnaire survey indicated that the participants had positive attitudes toward the mirror training using Mirrosistant. Most students believed that the mirror training device could improve their perceptions of direction and distance, as well as their sensations of dental operation and dental fulcrum. CONCLUSION: Mirror training using Mirrosistant can enhance dental students' mirror perceptual and operational skills on virtual simulation dental training system.

Prognostic role of CD11b+ myeloid-derived suppressor cells in oral squamous cell carcinoma.

Introduction: Myeloid-derived suppressor cells (MDSCs) are critically involved in cancer immune suppression and MDSC density has been recognized as a robust prognostic biomarker. Here, we sought to characterize the densities and locations of CD11b+ MDSCs in primary oral squamous cell carcinoma (OSCC) and determine their prognostic significance. Material and methods: A total of 144 eligible OSCC samples from a tertiary referral oral cancer center were retrospectively collected. Intensities of CD11b+ MDSCs at the tumor center (CT) and invasive margin (IM) in OSCC samples were detected by immunohistochemistry and automatically quantified using Image J software. The optimal cutoff values for CD11b CT and CD11b IM were determined by X-tile based on overall survival. The associations between CD11b+ MDSCs and clinicopathological parameters were assessed by the χ2 test. The prognostic value of CD11b+ MDSCs was evaluated by Kaplan-Meier plots, Cox regression analyses and receiver operating characteristics curves. Results: High density of CD11b+ MDSCs at CT or IM was significantly associated with inferior overall and disease-free survival (Kaplan-Meir, p < 0.05, log-rank test). CD11b CT and CD11b IM were identified as independent prognostic predictors for patient survival. The prediction accuracy and specificity of CD11b CT and CD11b IM were superior to other prognostic parameters. Conclusions: Our data indicated that increased densities of CD11b+ MDSCs in CT and IM regions were significantly associated with poor prognoses, which might be novel prognostic factors for OSCC.

Tertiary lymphoid structures in head and neck squamous cell carcinoma improve prognosis by recruiting CD8+ T cells.

Tertiary lymphoid structures (TLSs) are formed in long-term chronic inflammation, promoting the local recruitment of lymphocytes, antigen presentation and regulation of immune response, correlated with a better prognosis for cancer patients. Although studies have been conducted to explore methods that accelerate the establishment of TLSs, related research in head and neck squamous cell carcinoma (HNSCC) is still lacking. In this study, we analysed data from The Cancer Genome Atlas and performed immunohistochemical staining analyses of 188 patient samples. The results showed that TLSs promoted the infiltration of immune cells. Patients with TLSs with high infiltration of CD8+ cells showed the best prognosis. Since lymphotoxin α (LTα) was significantly increased in tissues with TLSs, we overexpressed LTα in SCC7 cells (a mouse-derived HNSCC cell line) and established tongue-tumour-bearing models. The polychromatic observation of tissue sections showed that T-cell aggregation increased in the LTα cell group, and a grade 1 TLS was formed on the 12th day after inoculating the cells. Moreover, the tumour volume in the LTα group was significantly less than that of the control group, whereas both the number and the proportion of infiltrated CD8+ T cells were increased. The peripheral CD8+ cells in mice were removed, and no difference was observed in tumour size or TLS formation. Remarkably, we found that TLS led to an increase in the antitumour effect by recruiting CD8+ T cells in HNSCC, showing a CD8+ T-cell-dependent antitumour effect. Moreover, LTα overexpression in the tumour promoted the formation of TLSs.

circFNDC3B Accelerates Vasculature Formation and Metastasis in Oral Squamous Cell Carcinoma.

Emerging evidence has demonstrated that circular RNAs (circRNA) are involved in cancer metastasis. Further elucidation of the role of circRNAs in oral squamous cell carcinoma (OSCC) could provide insights into mechanisms driving metastasis and potential therapeutic targets. Here, we identify a circRNA, circFNDC3B, that is significantly upregulated in OSCC and is positively associated with lymph node (LN) metastasis. In vitro and in vivo functional assays showed that circFNDC3B accelerated the migration and invasion of OSCC cells and the tube-forming capacity of human umbilical vein endothelial cells and human lymphatic endothelial cells. Mechanistically, circFNDC3B regulated ubiquitylation of the RNA-binding protein FUS and the deubiquitylation of HIF1A through the E3 ligase MDM2 to promote VEGFA transcription, thereby enhancing angiogenesis. Meanwhile, circFNDC3B sequestered miR-181c-5p to upregulate SERPINE1 and PROX1, which drove epithelial-mesenchymal transition (EMT) or partial-EMT (p-EMT) in OSCC cells and promoted lymphangiogenesis to accelerate LN metastasis. Overall, these findings uncovered the mechanistic role of circFNDC3B in orchestrating cancer cell metastatic properties and vasculature formation, suggesting circFNDC3B could be a potential target to reduce OSCC metastasis. SIGNIFICANCE: Dual functions of circFNDC3B in enhancing the metastatic ability of cancer cells and promoting vasculature formation through regulation of multiple pro-oncogenic signaling pathways drive lymph node metastasis of OSCC.

SIRT6-regulated macrophage efferocytosis epigenetically controls inflammation resolution of diabetic periodontitis.

Rationale: Diabetes exacerbates the prevalence and severity of periodontitis, leading to severe periodontal destruction and ultimately tooth loss. Delayed resolution of inflammation is a major contributor to diabetic periodontitis (DP) pathogenesis, but the underlying mechanisms of this imbalanced immune homeostasis remain unclear. Methods: We collected periodontium from periodontitis with or without diabetes to confirm the dysfunctional neutrophils and macrophages in aggravated inflammatory damage and impaired inflammation resolution. Our in vitro experiments confirmed that SIRT6 inhibited macrophage efferocytosis by restraining miR-216a-5p-216b-5p-217 cluster maturation through ''non-canonical'' microprocessor complex (RNA pulldown, RIP, immunostaining, CHIP, Luciferase assays, and FISH). Moreover, we constructed m6SKO mice that underwent LIP-induced periodontitis to explore the in vitro and in vivo effect of SIRT6 on macrophage efferocytosis. Finally, antagomiR-217, a miRNA antagonism, was delivered into the periodontium to treat LIP-induced diabetic periodontitis. Results: We discovered that insufficient SIRT6 as a histone deacetylase in macrophages led to unresolved inflammation and aggravated periodontitis in both human and mouse DP with accumulated apoptotic neutrophil (AN) and higher generation of neutrophil extracellular traps. Mechanistically, we validated that macrophage underwent high glucose stimulation resulting in disturbance of the SIRT6-miR-216/217 axis that triggered impeded efferocytosis of AN through targeting the DEL-1/CD36 axis directly. Furthermore, we demonstrated the inhibitory role of SIRT6 for MIR217HG transcription and identified a non-canonical action of microprocessor that SIRT6 epigenetically hindered the splicing of the primary miR-216/217 via the complex of hnRNPA2B1, DGCR8, and Drosha. Notably, by constructing myeloid-specific deletion of SIRT6 mice and locally delivering antagomir-217 in DP models, we strengthened the in vivo effect of this axis in regulating macrophage efferocytosis and inflammation resolution in DP. Conclusions: Our findings delineated the emerging role of SIRT6 in mediating metabolic dysfunction-associated inflammation, and therapeutically targeting this regulatory axis might be a promising strategy for treating diabetes-associated inflammatory diseases.

A retrospective study of iRoot BP Plus pulpotomy compared with Vitapex pulpectomy for irreversible pulpitis of primary molars with the presence of coronal pulp tissue.

BACKGROUND: Pulpotomy has been successfully performed in immature and mature permanent teeth with irreversible pulpitis but rarely in primary teeth. AIM: To evaluate the outcomes of iRoot BP Plus pulpotomy and Vitapex pulpectomy in primary molars with irreversible pulpitis. DESIGN: We selected 130 primary molars of 99 patients, aged 3-7 years, diagnosed with irreversible pulpitis with coronal pulp tissue and treated with iRoot BP Plus pulpotomy or Vitapex pulpectomy (median follow-up period: 18 months). They were divided into the pulpotomy (n = 88) and pulpectomy (n = 42) groups according to treatment procedure. The pulpotomy group was further divided into asymptomatic (n = 46) and symptomatic (n = 42) subgroups according to preoperative symptoms. The chi-squared test and Cox regression were performed to analyze the outcomes. RESULTS: Clinical and radiographic success rates were significantly higher in the pulpotomy group (98.9% and 95.5%) than in the pulpectomy group (88.1% and 54.8%) and did not differ significantly between asymptomatic and symptomatic pulpotomy subgroups. CONCLUSION: Irreversible pulpitis of primary molars with coronal pulp tissue can be successfully treated with iRoot BP Plus pulpotomy. Early intraradicular resorption of materials is the main adverse outcome of Vitapex pulpectomy.

Tertiary lymphoid structures in oral lichen planus and oral epithelial dysplasia with lichenoid features: A comparative study.

OBJECTIVE: Tertiary lymphoid structures (TLSs) provide sites for antigen presentation and activation of lymphocytes, promoting their infiltration; thus, enhancing specific immune responses. The aim of this comparative cross-sectional study was to reveal the characteristics and influence of TLSs in oral lichen planus (OLP) and oral epithelial dysplasia (OED) with lichenoid features. METHODS: Clinical information and samples of 51 OLP and 19 OED with lichenoid features were collected. Immunohistochemistry was performed, and the structures where CD20+ B cells and CD3+ T cells aggregated with peripheral lymph node addressin positive (PNAd+) vessels were defined as TLSs. The results and clinical information were analysed. RESULT: TLS were found in 44 (86.3%) patients with OLP and 19 (100%) patients with OED. The TLS score was higher in OED group (p = 0.023), accompanied by an increased number of PNAd+ vessels. The TLS was significantly correlated with PNAd+ vessels (p = 0.027), CD20+ B (p < 0.001) and CD208+ dendritic cells (p = 0.001). Foxp3+ Treg cells but not CD8+ T cells infiltrated more severely in OED (p = 0.003) and increased when TLS score was high (p = 0.002). CONCLUSIONS: This study revealed the widespread development of TLSs in the OLP and OED. The presence of TLSs showed a close relationship with dysplasia and may increase malignant potency by over-inducing Treg cells.

Influence of PD-1/PD-L1 on immune microenvironment in oral leukoplakia and oral squamous cell carcinoma.

OBJECTIVE: To evaluate the relation between the expression of PD-1, PD-L1, CD3, CD8, Foxp3 and clinicopathological features in patients with oral leukoplakia (OLK) and oral squamous cell carcinomas (OSCC) as well as the malignant outcome in OLK patients, and to study the effect of PD-1 and PD-L1 on immune microenvironment in the progression of oral carcinogenesis. METHODS: We evaluated the expression of PD-1/PD-L1 and composition of CD3+ , CD8+ and Foxp3+ T lymphocytes in OLK and OSCC samples by immunohistochemical (IHC) staining and analyzed their relation with clinical information and malignant transformation in OLK patients. RESULTS: IHC staining demonstrated that the expression of PD-1 was significantly increased in the high-grade OLK group than in the low-grade OLK group, while PD-L1 was detected mainly in OSCC. The expression of CD3, CD8, and Foxp3 was found higher in the high-grade OLK group than in the low-grade OLK group, and the Foxp3+ cells were found more in the OSCC group than in the high-grade OLK group. PD-1 was significantly correlated with CD3 (p < 0.05, R = 0.52), CD8 (p < 0.05, R = 0.46), and Foxp3 (p < 0.05, R = 0.46), and the low PD-1-expression group showed a better malignant-free survival than high PD-1 expression group in the OLK (p < 0.05). CONCLUSION: The PD-1/PD-L1 may induce immune suppression in OLK and accelerate the progress of malignant transformation.

Capnocytophaga gingivalis is a potential tumor promotor in oral cancer.

OBJECTIVES: To investigate the role of oral microbiome in promoting oral squamous cell carcinoma (OSCC) development. MATERIALS AND METHODS: We investigated the salivary microbiome of 108 controls and 70 OSCC cases by16S rRNA gene sequencing and detected the fluorescence signal of OSCC-related pathological bacteria by fluorescence in situ hybridization assay (FISH). The invasion and migration assays were used to show the differences of invasive and migrative abilities between control and experimental groups. Quantitative real-time PCR and Western blotting were used to verify the epithelial-to-mesenchymal transition (EMT). RESULTS: In our study, the overall microbiome abundance and composition were richer in the 108 controls than in the 70 OSCC cases. We demonstrated that Streptococcus, Capnocytophaga, Peptostreptococcus, and Lactobacillus were highly abundant in the saliva of OSCC patients by 16S rDNA sequencing and FISH. Moreover, we found that Capnocytophaga gingivalis (C. gingivalis) was highly presented in OSCC tissues by FISH. We focused on C. gingivalis and found that its supernatant induced OSCC cells to undergo EMT, causing the cells to acquire a mesenchymal phenotype associated with highly invasive and metastatic properties. CONCLUSION: Taken together, these results indicated that C. gingivalis might invade OSCC tissues and played an important role in OSCC by promoting OSCC invasion and metastasis by inducing EMT. Hence, the role of C. gingivalis in cancer progression revealed a new direction for the research of OSCC.

Macrophage miR-149-5p induction is a key driver and therapeutic target for BRONJ.

Bisphosphonate-related (BP-related) osteonecrosis of the jaw (BRONJ) is one of the severe side effects of administration of BPs, such as zoledronic acid (ZA), which can disrupt the patient's quality of life. Although the direct target of skeletal vasculature and bone resorption activity by BPs has been phenomenally observed, the underlying mechanism in BRONJ remains largely elusive. Thus, it is urgently necessary to discover effective therapeutic targets based on the multifaceted underlying mechanisms in the development of BRONJ. Here, we determined the inhibitory role of ZA-treated macrophages on osteoclast differentiation and type H vessel formation during tooth extraction socket (TES) healing. Mechanistically, ZA activated the NF-κB signaling pathway and then induced p65 nuclear translocation in macrophages to promote miR-149-5p transcription, resulting in impaired osteoclast differentiation via directly binding to the Traf6 3'-UTR region. Moreover, we identified that miR-149-5p-loaded extracellular vesicles derived from ZA-treated bone marrow-derived macrophages could regulate biological functions of endothelial cells via the Rap1a/Rap1b/VEGFR2 pathway. Furthermore, local administration of chemically modified antagomiR-149-5p was proven to be therapeutically effective in BRONJ mice. In conclusion, our findings illuminate the dual effects of miR-149-5p on skeletal angiogenesis and bone remolding, suggesting it as a promising preventive and therapeutic target for BRONJ.

lncRNA CYTOR promotes aberrant glycolysis and mitochondrial respiration via HNRNPC-mediated ZEB1 stabilization in oral squamous cell carcinoma.

Oral squamous cell carcinoma (OSCC), the most common malignancy of the oral and maxillofacial region, severely affects human health. However, current treatments for OSCC commonly show only a ~60% 5-year survival rate of patients with distant metastases, indicating an urgent need for targeted treatments for patients with advanced metastases. Here, we report a survival-related long non-coding RNA, CYTOR, which is highly expressed in the lesions of oral cancer patients. We found that CYTOR can promote both migration and invasion in oral cancer cells as well as the epithelial-mesenchymal transition (EMT). RNA-sequencing of CYTOR-knockdown oral cancer cells revealed that CYTOR can regulate mitochondrial respiration and RNA splicing. Mechanistically, we found that nuclear-localized CYTOR interacts with HNRNPC, resulting in stabilization of ZEB1 mRNAs by inhibiting the nondegradative ubiquitination of HNRNPC. By synthesizing CYTOR-targeting small interfering RNAs (siRNAs) encapsulated in Nanoscale Metal Organic Frameworks (NMOFs), we demonstrate the targeted suppression of CYTOR to inhibit invasion and metastasis of oral cancer cells in a nude mouse model. Cumulatively, this study reveals the potential role of the CYTOR-HNRNPC-ZEB1 axis in regulating mitochondrial metabolism and glycolysis of oral cancer cells, and illustrates the effective use of lncRNA targeting in anti-metastatic cancer therapies.

The deubiquitinase USP7 promotes HNSCC progression via deubiquitinating and stabilizing TAZ.

Dysregulated abundance, location and transcriptional output of Hippo signaling effector TAZ have been increasingly linked to human cancers including head neck squamous cell carcinoma (HNSCC). TAZ is subjected to ubiquitination and degradation mediated by E3 ligase ß-TRCP. However, the deubiquitinating enzymes and mechanisms responsible for its protein stability remain underexplored. Here, we exploited customized deubiquitinases siRNA and cDNA library screen strategies and identified USP7 as a bona fide TAZ deubiquitinase in HNSCC. USP7 promoted cell proliferation, migration, invasion in vitro and tumor growth by stabilizing TAZ. Mechanistically, USP7 interacted with, deubiquitinated and stabilized TAZ by selectively removing its K48-linked ubiquitination chain independent of canonical Hippo kinase cascade. USP7 potently antagonized ß-TRCP-mediated ubiquitin-proteasomal degradation of TAZ and enhanced its nuclear retention and transcriptional output. Importantly, overexpression of USP7 correlated with TAZ upregulation, tumor aggressiveness and unfavorable prognosis in HNSCC patients. Pharmacological inhibition of USP7 significantly suppressed tumor growth in both xenograft and PDX models. Collectively, these findings identify USP7 as an essential regulator of TAZ and define USP7-TAZ signaling axis as a novel biomarker and potential therapeutic target for HNSCC.

Development of a prognostic pyroptosis-related gene signature for head and neck squamous cell carcinoma patient.

OBJECTIVE: Head and neck squamous cell carcinoma (HNSCC) is a major threat to public health. Pyroptosis is a form of inflammatory programmed cell death that is still incompletely understood. The role of pyroptotic cell death in HNSCC remains to be fully defined. As such, the present study was developed to explore the potential prognostic utility of a pyroptosis-related gene (PRG) signature in HNSCC. METHODS: PRG expression patterns and the associated mutational landscape in HNSCC were analyzed, after which a 6-gene prognostic model was constructed through least absolute shrinkage and selection operator (LASSO) and Cox regression analyses using the TCGA dataset, followed by validation with two GEO datasets (GSE41643 and GSE65858). The relative expression of the genes in the prognostic model was assessed via RT-qPCR in tumor and paired adjacent normal tissue samples from a 32-patient cohort. Potential predictors of patient outcomes associated with this 6-gene model were identified through topological degree analyses of a protein-protein interaction network. Moreover, the prognostic value of NLRP3 as a predictor of HNSCC patient prognosis was established through immunohistochemical (IHC) analyses of samples from 176 HNSCC patients. Lastly, in vitro studies were performed to further demonstrate the relevance of NLRP3 in the context of HNSCC development. RESULTS: Differentially expressed PRGs were able to readily differentiate between HNSCC tumors and normal tissues. Risk scores derived from the 6-gene PRG model were independent predictors of HNSCC patient prognosis, and genes that were differentially expressed between low- and high-risk groups were associated with tumor immunity. RT-qPCR assays also showed the potential protective role of NLRP3 in HNSCC patients. IHC analyses further supported the value of NLRP3 as a predictor of HNSCC patient outcomes. Invasion and migration assays demonstrated the potential role of NLRP3 in the inhibition of HNSCC development. CONCLUSIONS: Overall, these results highlight a novel prognostic gene signature that offers value in the context of HNSCC patient evaluation, although additional research will be essential to elucidate the mechanisms linking these PRGs to HNSCC outcomes.