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.

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.

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.

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.

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.

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.

Cancer-associated fibroblasts promote oral squamous cell carcinoma progression through LOX-mediated matrix stiffness.

BACKGROUND: Cancer-associated fibroblasts (CAFs), the most abundant cells in the tumor microenvironment, have prominent roles in the development of solid tumors as stromal targets. However, the underlying mechanism of CAFs' function in oral squamous cell carcinoma (OSCC) development remains unclear. Here, we investigated the role of lysyl oxidase (LOX) expression in CAFs in tumor stromal remodeling and the mechanism of its effect on OSCC progression. METHODS: Multiple immunohistochemistry (IHC) staining was performed to detect the correlation of CAFs and LOX in the stroma of OSCC specimens, as well as the correlation with clinicopathological parameters and prognosis. The expression of LOX in CAFs were detected by RT-qPCR and western blot. The effects of LOX in CAFs on the biological characteristics of OSCC cell line were investigated using CCK-8, wound-healing and transwell assay. CAFs were co-cultured with type I collagen in vitro, and collagen contraction test, microstructure observation and rheometer were used to detect the effect of CAFs on remodeling collagen matrix. Then, collagen with different stiffness were established to investigate the effect of matrix stiffness on the progression of OSCC. Moreover, we used focal adhesion kinase (FAK) phosphorylation inhibitors to explored whether the increase in matrix stiffness promote the progression of OSCC through activating FAK phosphorylation pathway. RESULTS: LOX was colocalized with CAFs in the stroma of OSCC tissues, and its expression was significantly related to the degree of malignant differentiation and poor prognosis in OSCC. LOX was highly expressed in CAFs, and its knockdown impaired the proliferation, migration, invasion and EMT process of OSCC cells. The expression of LOX in CAFs can catalyze collagen crosslinking and increase matrix stiffness. Furthermore, CAFs-derived LOX-mediated increase in collagen stiffness induced morphological changes and promoted invasion and EMT process in OSCC cells by activating FAK phosphorylation pathway. CONCLUSIONS: Our findings suggest that CAFs highly express LOX in the stroma of OSCC and can remodel the matrix collagen microenvironment, and the increase in matrix stiffness mediated by CAFs-derived LOX promotes OSCC development through FAK phosphorylation pathway. Thus, LOX may be a potential target for the early diagnosis and therapeutic treatment of OSCC.

LncRNA HOTAIR promotes the invasion and metastasis of oral squamous cell carcinoma through metastasis-associated gene 2.

Despite therapeutic advancements, there has been little improvement in the survival status of patients with oral squamous cell carcinoma (OSCC). HOX antisense intergenic RNA (HOTAIR) has been shown to be dysregulated in several cancer types. However, the roles of HOTAIR in OSCC remain largely unknown. In this study, we investigated the association of HOTAIR expression with clinicopathological features in OSCC patients and the crucial roles of HOTAIR in the modulation of tumor progression. Our results showed that HOTAIR was highly expressed both in OSCC tissue samples and cell lines compared with corresponding normal oral mucosa tissues and human oral keratinocytes. Its overexpression was positively correlated with TNM (tumor-node-metastases) stage, histological grade, and regional lymph node metastasis. The knockdown of HOTAIR by short hairpin RNA significantly decreased the migration, invasion, and epithelial-mesenchymal transition of OSCC cells in vitro. Moreover, there was a negative correlation between HOTAIR and microRNA-326 expression in OSCC tissue samples and cell lines. Luciferase reporter and loss-of-function assays revealed that HOTAIR acted as a competitive endogenous RNA effectively sponging miR-326, thereby regulating the derepression of metastasis-associated gene 2 (MTA2). Finally, the expression and clinical significance of MTA2 were analyzed in another cohort of OSCC tissue samples. High MTA2 expression was significantly correlated with clinicopathological features of advanced OSCC and poor prognosis for patients with OSCC. Collectively, HOTAIR overexpression promoted the progression of OSCC. The HOTAIR-miR-326-MTA2 axis may contribute to a better understanding of OSCC pathogenesis and be a potential therapeutic target for OSCC.

Zoledronic acid promotes TLR-4-mediated M1 macrophage polarization in bisphosphonate-related osteonecrosis of the jaw.

Bisphosphonate-related osteonecrosis of the jaw (BRONJ) is a detrimental side effect of the long-term administration of bisphosphonates. Although macrophages were reported to be an important mediator of BRONJ, the detailed potential mechanism of BRONJ remains unclear. Here, we reported an elevated TLR-4 expression in macrophages under action of zoledronic acid (ZA), resulting in enhanced M1 macrophage polarization and decreased M2 macrophage polarization both in vitro and in vivo. After inhibiting the TLR-4 signaling pathway, the activation of the TLR-4/NF-κB signaling pathway and the induction of NF-κB nuclear translocation and production of proinflammatory cytokines by ZA were suppressed in macrophages, thereby inhibiting M1 macrophage polarization. By utilizing the TLR-4-/- mice, development of BRONJ was markedly ameliorated, and M1 macrophages were significantly attenuated in the extraction socket tissues in the TLR-4-/- mice. Importantly, the systemic administration of the TLR-4 inhibitor TAK-242 improved the wound healing of the extraction socket and decreased the incidence rate of BRONJ. Taken together, our findings suggest that TLR-4-mediated macrophage polarization participates in the pathogenesis of BRONJ in mice, and TLR-4 may be a potential target for the prevention and therapeutic treatment of BRONJ.-Zhu, W., Xu, R., Du, J., Fu, Y., Li, S., Zhang, P., Liu, L., Jiang, H. Zoledronic acid promotes TLR-4-mediated M1 macrophage polarization in bisphosphonate-related osteonecrosis of the jaw.

Upregulated LOX and increased collagen content associated with aggressive clinicopathological features and unfavorable outcome in oral squamous cell carcinoma.

OBJECTIVES: Collagen is a core protein that maintains the homeostasis of the extracellular matrix (ECM), and its dysregulation in human cancers has attracted increasing attention. In tumors, increased lysyl oxidase (LOX)-catalyzed collagen cross-linking plays a critical role in collagen dysregulation. However, the expression patterns of LOX and collagen and their clinicopathological significance in oral squamous cell carcinoma (OSCC) have not been well established. METHODS: The LOX mRNA expression in OSCC was measured by RT-PCR and bioinformatics analysis. LOX protein expression and total collagen content were identified by immunohistochemistry or Masson's trichrome staining in a retrospective cohort of primary OSCC samples, respectively. Moreover, the associations between LOX and collagen expression and various clinicopathological parameters or patient survival were assessed. RESULTS: LOX mRNA was overexpressed in OSCC samples. Higher expression of LOX, collagen content or co-overexpression of LOX and collagen was significantly associated with aggressive clinicopathological features. Importantly, aberrant expression of LOX, collagen content, or both were markedly correlated with decreased overall and disease-free survival (P < 0.05). Moreover, univariate and multivariate Cox models analyses indicated that LOX, collagen content or their combination could serve as an independent prognostic predictor for OSCC patients. ROC analysis further revealed that the combination of LOX and collagen was superior to parameter alone as a prognostic predictor. CONCLUSIONS: Our findings reveal that elevated LOX and collagen content significantly corelate with aggressiveness and worse prognosis in OSCC.

Heat shock factor 1 in cancer-associated fibroblasts is a potential prognostic factor and drives progression of oral squamous cell carcinoma.

Heat shock factor 1 (HSF1) is highly expressed in various malignancies and is a potential modulator of tumor progression. Emerging evidence suggests that HSF1 activation in stromal cells is closely related to poor patient prognosis. However, the role of HSF1 in oral squamous cell carcinoma (OSCC) remains elusive. We aimed to investigate the function of HSF1 in cancer-associated fibroblasts (CAFs) of the tumor microenvironment (TME) and in tumor development. In the present study, we found that HSF1 was highly expressed in both CAFs and tumor cells, and was significantly correlated with poor prognosis and overall survival. Moreover, HSF1 overexpression in CAFs resulted in a fibroblast-like phenotype of Cal27 cells, induced epithelial-mesenchymal transition (EMT), and promoted proliferation, migration and invasion in Cal27 cells. HSF1 knockdown attenuated features of CAFs and reduced EMT, proliferation, migration and invasion in Cal27 cells. Furthermore, HSF1 in CAFs promoted tumor growth in nude mice. Taken together, these data suggest that HSF1 expression in CAFs drive OSCC progression, and could serve as an independent prognostic marker of patients with OSCC. Thus, HSF1 is a potent mediator of OSCC malignancy.

A novel Notch1 missense mutation (C1133Y) in the Abruptex domain exhibits enhanced proliferation and invasion in oral squamous cell carcinoma.

BACKGROUND: Notch1 has been regarded as a fundamental regulator in tissue differentiation and stem cell properties. Recently, Notch1 mutations have been reported intensively both in solid tumors and in hematopoietic malignancies. However, little is known about the biological effect and the clinical implication of these reported mutations. Previously, we discovered several missense mutations in the Notch1 receptor in a Chinese population with oral squamous cell carcinoma (OSCC). METHODS: We selected a 'hotspot' mutation in the Abruptex domain (C1133Y). The expression of Notch1 was determined by western blot and real-time qPCR in OSCC cell lines transfected with pcDNA3.1-Notch1WT, pcDNA3.1-Notch1C1133Y, or pcDNA3.1 empty vector. CCK-8 assays were used to assess cell proliferation. Flow cytometry and western blot were used to confirm the alteration of cell cycle after transfection. Transwell assays and the detection of Epithelial-to-mesenchymal transition (EMT) markers were used to determine the invasive ability. The effects of Notch1 C1133Y mutation were analyzed by Immunofluorescence staining and the expression of EGFR-PI3K/AKT signaling. RESULTS: We demonstrated that Notch1C1133Y mutation inactivated the canonical Notch1 signaling. We identified an oncogenic phenotype of this mutation by promoting cell proliferation, invasion and by inducing EMT in OSCC cell lines. We found that the Notch1C1133Y mutation exhibited a decreased S1-cleavage due to the impaired transport of Notch1 protein from the endoplasmic reticulum (ER) to the Golgi complex, which was consistent with the observation of the failure of the Notch1C1133Y mutated receptor to present at the cell surface. Importantly, the mutated Notch1 activated the EGFR-PI3K/AKT signaling pathway, which has been confirmed as an overwhelming modulator in OSCC. CONCLUSIONS: Taken together, our findings revealed for the first time a novel Notch1 mutation that enhances proliferation and invasion in OSCC cell lines. The Notch1 C1133Y mutation impairs the processing of notch1 protein and the critical links between the mutated Notch1 and the activated EGFR-PI3K/AKT signaling pathway.

MiR-876-5p modulates head and neck squamous cell carcinoma metastasis and invasion by targeting vimentin.

BACKGROUND: Local or distant metastasis remains the main course of death in head and neck squamous cell carcinoma (HNSCC) patients. MicroRNAs (miRNAs) have been implicated in metastasis of HNSCC, but the mechanisms of their action are mainly undocumented. Through public head and neck cancer miRNA expression datasets, we found that miR-876-5p was a novel potential tumor suppressor targeting HNSCC metastasis. METHODS: Clinical significance and mechanism of miR-876-5P was systematically analyzed in HNSCC. Quantitative RT-PCR was used to evaluate miR-876-5p levels in HNSCC cell lines and in 20 pairs of HNSCC with associated regional nodal metastases and HNSCC without metastatic primary tumors. Scratch and invasion assays were evaluated to determine the role of miR-876-5p in the regulation of HNSCC cell migration and invasion, respectively. Western blotting was used to investigate the mechanism by which miR-876-5p suppresses HNSCC cell invasion and migration. Luciferase assays were performed to assess miR-876-5p binding to the vimentin gene. The animal model was used to support the in vitro experimental findings. RESULTS: MiR-876-5p mimics inhibited HNSCC cell migration and invasion. Vimentin protein and mRNA levels were decreased in the miR-876-5p mimics group but increased in the miR-876-5p inhibitors group, which demonstrated that miR-876-5p inhibits vimentin expression in HNSCC cells. By directly targeting the vimentin 3'-UTR, we used dual-luciferase reporter assays to verify that vimentin is a functional downstream target of miR-876-5p. Importantly, increased vimentin expression promoted cell migration and invasion, and co-transfection with miR-876-5p mimics and vimentin restored cell aggressiveness to the original level. Moreover, miR-876-5p overexpression significantly downregulated vimentin expression level and inhibited the distal metastasis of HNSCC cells in vivo. CONCLUSIONS: miR-876-5p, which functions as a tumor suppressor in HNSCC, inhibits metastasis by targeting vimentin and provides a novel therapeutic target for HNSCC treatment.

Density and location of CD3+ and CD8+ tumor-infiltrating lymphocytes correlate with prognosis of oral squamous cell carcinoma.

BACKGROUND: Tumor-infiltrating lymphocytes (TILs) are regarded as adaptive immune response of the host to cancer cells and valuable prognostic factors. Here, we sought to characterize the densities and locations of CD3+ and CD8+ TILs in primary oral squamous cell carcinoma (OSCC) samples and assess their clinicopathological and prognostic significance. METHODS: A total number of 169 OSCC samples from 2 independent patient cohorts (Nanjing cohort, 93 cases; Wuxi cohort, 76 cases) were retrospectively collected. The numbers of CD3+ and CD8+ TILs at tumor center (CT) and invasive margin (IM) of OSCC were identified by immunohistochemistry and calculated. The optimal cutoff values for CD3+ and CD8+ TILs to stratify patients were determined by X-tile software in Nanjing cohort and further utilized in Wuxi cohort. The associations between CD3+ /CD8+ TILs and clinicopathological parameters or patient survival were assessed. The prognostic values of CD3+ / CD8+ TILs were evaluated by Cox regression analyses. RESULTS: CD3+ and CD8+ TILs were identified at both CT and IM and enriched at IM. High density of CD3+ TILs at IM (CD3 IM) was significantly associated with increased overall and disease-specific survival (P < .05). High density of CD8+ TILs at CT (CD8 CT) was significantly associated with increased overall but not disease-specific survival. Moreover, CD3 IM and CD8 CT were identified as independent prognostic factors for patient survival. CONCLUSIONS: Our findings provide further evidence to support the prognostic values of CD3+ and CD8+ TILs for OSCC, suggesting that TIL subsets might be viable biomarkers and therapeutic targets with translational significance.

Toll-like receptor 4 deficiency affects the balance of osteoclastogenesis and osteoblastogenesis in periodontitis.

Toll-like receptor 4 (TLR4) acts as a double-edged sword in the occurrence and development of periodontitis. While the activation of TLR4 in macrophages aids in clearing local pathogens, it can also disrupt innate immune responses, upsetting microecological balance and accelerating the destruction of periodontal bone tissues. To date, the effects of TLR4 on osteogenesis and osteoclastogenesis in periodontitis have not been comprehensively studied. In this study, we investigated the development of periodontitis in the Tlr4-/- mice by ligating their second molars with silk threads. Compared to wild-type (WT) mice, Tlr4-/- mice demonstrated increased resistance to periodontitis-associated bone destruction, as evidenced by decreased bone resorption and enhanced bone regeneration. Mechanistically, the deletion of Tlr4 not only inhibited osteoclast formation by reducing the expression of NFATc1, CTSK and TRAP, but also enhanced osteogenic abilities through increased expression of OCN, OPN and RUNX2. In conclusion, TLR4 tips the balance of osteoclastogenesis and osteogenesis, thereby promoting periodontal bone destruction in periodontitis.

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.

Membrane Type 1 Matrix Metalloproteinase induces an epithelial to mesenchymal transition and cancer stem cell-like properties in SCC9 cells.

BACKGROUND: Tissue invasion and metastasis are acquired abilities of cancer and related to the death in oral squamous cell carcinoma (OSCC). Emerging observations indicate that the epithelial-to-mesenchymal transition (EMT) is associated with tumor progression and the generation of cells with cancer stem cells (CSCs) properties. Membrane Type 1 Matrix Metalloproteinase (MT1-MMP) is a cell surface proteinase, which is involved in degrading extracellular matrix components that can promote tumor invasion and cell migration. METHODS: In the current study, we utilized SCC9 cells stably transfected with an empty vector (SCC9-N) or a vector encoding human MT1-MMP (SCC9-M) to study the role of MT1-MMP in EMT development. RESULTS: Upon up-regulation of MT1-MMP, SCC9-M cells underwent EMT, in which they presented a fibroblast-like phenotype and had a decreased expression of epithelial markers (E-cadherin, cytokeratin18 and ß-catenin) and an increased expression of mesenchymal markers (vimentin and fibronectin). We further demonstrated that MT1-MMP-induced morphologic changes increased the level of Twist and ZEB, and were dependent on repressing the transcription of E-cadherin. These activities resulted in low adhesive, high invasive abilities of the SCC9-M cells. Furthermore, MT1-MMP-induced transformed cells exhibited cancer stem cell (CSC)-like characteristics, such as low proliferation, self-renewal ability, resistance to chemotherapeutic drugs and apoptosis, and expression of CSCs surface markers. CONCLUSIONS: In conclusion, our study indicates that overexpression of MT1-MMP induces EMT and results in the acquisition of CSC-like properties in SCC9 cells. Our growing understanding of the mechanism regulating EMT may provide new targets against invasion and metastasis in 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.