The competitive mechanism of EZH1 and EZH2 in promoting oral squamous cell carcinoma.

Enhancer of Zeste Homolog 1 (EZH1) and Enhancer of Zeste Homolog 2 (EZH2) are the key components of polycomb repressive complex 2 (PRC2); however, the roles of these proteins in oral squamous cell carcinoma (OSCC) have yet to be elucidated. In this study, we aimed to determine the respective roles of these proteins in OSCC by investigating the expression levels of EZH1 and EZH2 in OSCC tissues (N = 63) by immunohistochemistry. In addition, we used lentiviruses to construct stable OSCC cell lines that overexpressed EZH1 and EZH2. Then, we investigated these cell lines for cell viability, colony formation capacity, stemness, and epithelial-mesenchymal transition (EMT). Binding competition between EZH1 and EZH2 with PRC2 was further evaluated using Co-immunoprecipitation (Co-IP). Compared with normal tissues, the expression levels of EZH2 in OSCC tissues was up-regulated, while the expression of EZH1 was down-regulated. EZH2 enhanced cell viability, colony formation capacity, stemness, and EMT, while EZH1 did not. Furthermore, analysis indicated that EZH1 and EZH2 bound competitively to PRC2 and influenced the methylation status of H3K27. In conclusion, our findings verified that EZH1 and EZH2 play opposing roles in OSCC and that EZH1 and EZH2 compete as the key component of PRC2, thus affecting the characteristics of OSCC via the methylation of H3K27.

Phenotype remodelling of HNSCC cells in the muscle invasion environment.

BACKGROUND: Tumour invading muscle in head and neck squamous cell carcinoma (HNSCC) is often associated with destructive growth and poor prognosis. However, the phenotypic functions and pathological mechanisms of muscle-invasive cancer cells in tumour progress remains unknown. In this study, we aimed to investigate the phenotypic functions of muscle-invasive cancer cells of HNSCC and their potential crosstalk with tumour microenvironment. METHODS: We obtained scRNA-seq data (SC) from GSE103322 (N = 18) and GSE181919 (N = 37), spatial RNA-seq data (ST) from GSE208253 and GSE181300 (N = 4), transcriptomics of human HNSCC samples from GSE42743 (N = 12) and GSE41613 (N = 97). Utilizing the TCGA-HNSC dataset, we conducted univariate and multivariate Cox analyses to investigate the prognostic impact of muscle-invasion in HNSCC, with validation in an additional cohort. Through Stutility and AUCell approaches, we identified and characterized muscle-invasive cancer cell clusters, including their functional phenotypes and gene-specific profiles. Integration of SC and ST data was achieved using Seurat analysis, multimodal intersection analysis, and spatial deconvolution. The results were further validated via in vitro and in vivo experiments. RESULTS: Our analyses of the TCGA-HNSC cohort revealed the presence of muscle-invasion was associated with a poor prognosis. By combining ST and SC, we identified muscle-invasive cancer cells exhibiting epithelial-to-mesenchymal transition (EMT) and myoepithelial-like transcriptional programs, which were correlated with a poor prognosis. Furthermore, we identified G0S2 as a novel marker of muscle-invasive malignant cells that potentially promotes EMT and the acquisition of myoepithelium-like phenotypes. These findings were validated through in vitro assays and chorioallantoic membranes experiments. Additionally, we demonstrated that G0S2-overexpressing cancer cells might attract human ECs via VEGF signalling. Subsequent in vitro and in vivo experiments revealed G0S2 plays key roles in promoting the proliferation and invasion of cancer cells. CONCLUSIONS: In this study, we profiled the transcriptional programs of muscle-invasive HNSCC cell populations and characterized their EMT and myoepithelial-like phenotypes. Furthermore, our findings highlight the presence of muscle-invasion as a predictive marker for HNSCC patients. G0S2 as one of the markers of muscle-invasive cancer cells is involved in HNSCC intravasation, probably via VEGF signalling.

TCL1A-expressing B cells are critical for tertiary lymphoid structure formation and the prognosis of oral squamous cell carcinoma.

BACKGROUND: Oral squamous cell carcinoma (OSCC) is a malignant tumor with a poor prognosis. Traditional treatments have limited effectiveness. Regulation of the immune response represents a promising new approach for OSCC treatment. B cells are among the most abundant immune cells in OSCC. However, the role of B cells in OSCC treatment has not been fully elucidated. METHODS: Single-cell RNA sequencing analysis of 13 tissues and 8 adjacent normal tissues from OSCC patients was performed to explore differences in B-cell gene expression between OSCC tissues and normal tissues. We further investigated the relationship between differentially expressed genes and the immune response to OSCC. We utilized tissue microarray data for 146 OSCC clinical samples and RNA sequencing data of 359 OSCC samples from The Cancer Genome Atlas (TCGA) to investigate the role of T-cell leukemia 1 A (TCL1A) in OSCC prognosis. Multiplex immunohistochemistry (mIHC) was employed to investigate the spatial distribution of TCL1A in OSCC tissues. We then investigated the effect of TCL1A on B-cell proliferation and trogocytosis. Finally, lentiviral transduction was performed to induce TCL1A overexpression in B lymphoblastoid cell lines (BLCLs) to verify the function of TCL1A. RESULTS: Our findings revealed that TCL1A was predominantly expressed in B cells and was associated with a better prognosis in OSCC patients. Additionally, we found that TCL1A-expressing B cells are located at the periphery of lymphatic follicles and are associated with tertiary lymphoid structures (TLS) formation in OSCC. Mechanistically, upregulation of TCL1A promoted the trogocytosis of B cells on dendritic cells by mediating the upregulation of CR2, thereby improving antigen-presenting ability. Moreover, the upregulation of TCL1A expression promoted the proliferation of B cells. CONCLUSION: This study revealed the role of B-cell TCL1A expression in TLS formation and its effect on OSCC prognosis. These findings highlight TCL1A as a novel target for OSCC immunotherapy.

The interaction of CD300lf and ceramide reduces the development of periodontitis by inhibiting osteoclast differentiation.

AIM: The regulation of osteoclasts (OCs) by inhibitory immunoreceptors maintains bone homeostasis and is considered an important determinant of the extent of periodontal pathology. The aim of this study was to investigate the role of the inhibitory immunoreceptor CD300lf and its ligand ceramide in osteoclastogenesis in periodontitis. MATERIALS AND METHODS: The expression of CD300lf was measured in vitro and in a ligature-induced periodontitis model. The effect of CD300lf ablation on osteoclastogenesis was examined in ligature-retained and ligature removal periodontitis models. The effect of ceramide, the ligand of CD300lf, was examined in osteoclastogenesis in vitro and in vivo by smearing 20 µg of ceramide dissolved in carboxymethylcellulose on teeth and gingiva every other day in an experimental periodontitis model and ligature removal model. RESULTS: CD300lf expression was downregulated during osteoclastogenesis. Ablation of CD300lf in the ligature-induced periodontitis model increased the number of OCs and exacerbated bone damage. Bone resorption caused by CD300lf ablation was reversible following ligature removal. CD300lf-ceramide binding suppressed osteoclastogenesis in vitro and inhibited alveolar bone loss in a mouse periodontitis model. CONCLUSIONS: Our findings reveal that CD300lf-ceramide binding plays a critical negative role in alveolar bone loss in periodontitis by inhibiting OCs differentiation.

Intratumoral platelet microthrombi in oral squamous cell carcinoma: A marker of lymph node metastasis.

OBJECTIVE: A hypercoagulable state exists in patients with oral squamous cell carcinoma (OSCC), but the role of platelets in the tumour microenvironment has not been explored. This study revealed the status of intratumoral plateletmicrothrombi (PLT-MT) and their clinicopathological relevance and predictive value in OSCC. STUDY DESIGN: This study retrospectively evaluated 106 OSCC patients. Tumour and tumour-adjacent tissue specimens were used to stain PLT-MT. Clinicopathological information, patient follow-ups and outcomes and preoperative coagulation and inflammatory hematologic indicators were collected, and their correlation with PLT-MT was analysed. RESULTS: Intratumoral PLT-MT was present in 35 of 106 patients with OSCC who had higher preoperative D-dimer, CRP, FIB and PT levels and lower TT levels. PLT-MT was an independent correlative factor of lymph node metastasis and suggested worse OS in N0 patients. CONCLUSIONS: Intratumoral PLT-MT was found in OSCC and was correlated with a hypercoagulable inflammatory state. PLT-MT was an independent marker of lymph node metastasis and showed potential in prognosis prediction.

Dysfunctional role of elevated TIGIT expression on T cells in oral squamous cell carcinoma patients.

OBJECTIVE: This study was aimed to analyze the role of T-cell immunoreceptor with immunoglobulin and tyrosine-based inhibitory motif domains (TIGIT) expression on T cells in patients with oral squamous cell carcinoma (OSCC). MATERIALS AND METHODS: Peripheral blood mononuclear cells (PBMC) and tumor-infiltrating lymphocytes (TILs) were collected from OSCC patients. The correlation between TIGIT expression and clinicopathologic features was analyzed by chi-square test. Phenotypic and functional study of TIGIT+ T cells were performed by flow cytometry. RESULTS: TIGIT was highly expressed on T cells from PBMC and TILs. High expression of TIGIT on CD4+ T cells (19.0%) and CD8+ T cells (35.9%) was also associated with higher T stage and nodal invasion. Moreover, TIGIT+ CD4+ and TIGIT+ CD8+ T cells sorted from OSCC patients showed a dysfunctional phenotype (low cell proliferation and low secretion of IL-2, TNF-α and IFN-γ), and TIGIT+ CD4+ T cells exhibited inhibitory function (high expression of Foxp3 and high amounts of IL-10). Importantly, TIGIT blockade can enhance the proliferation ability and effective cytokine production (IL-2, TNF-α, and IFN-γ) of CD4+ and CD8+ T cells from OSCC patients in vitro. CONCLUSIONS: TIGIT-expressing T cells exhibit a lower effector cytokine-releasing phenotype in OSCC patients.

Prediction of postoperative lower respiratory tract infections in tongue cancer patients based on pretreatment swallowing function.

OBJECTIVES: This study investigated the predictive role of pretreatment swallowing function and surgical factors on postoperative and nosocomial lower respiratory tract infections (PN-LRTIs). SUBJECTS AND METHODS: A retrospective study for predicting PN-LRTIs from January 2017 to December 2018 at Hospital of Stomatology, Sun Yat-sen University. Patients who were newly diagnosed with tongue squamous cell carcinoma (TSCC) were enrolled. Presurgical swallowing function was assessed using water swallow test (WST) and the M.D. Anderson Dysphagia Inventory (MDADI). RESULTS: A total of 83 patients were recruited to the study. Of which 54 were men (65.1%) and 29 were women (34.9%), with the mean age of 51 years old. Thirteen (15.7%) developed PN-LRTIs. On univariate analysis, the outcomes of WST, the MDADI scores, T stage, tongue resection range, operative time, segmental mandibulectomy, and type of neck dissection exhibited a statistical significance (p < .05). On multivariate analysis, abnormal group of WST (odds ratio [OR], 15.88; 95% CI, 2.13-118.64) and total glossectomy (OR, 12.20; 95% CI, 2.01-68.32) was demonstrated to be independent risk factors. CONCLUSIONS: The WST together with the resection range of tongue can predict the postoperative risk of PN-LRTIs collaboratively. Clinically, preventive measures and intensified care should be taken for patients with abnormal WST outcome before surgery and management of total glossectomy.

P53 Alleviates the Progression of Periodontitis by Reducing M1-type Macrophage Differentiation.

Our objective is to explore the effect of P53 on the progression of periodontitis by regulating macrophages differentiation both in vitro and in vivo. Eighteen normal and periodontitis gingival tissues were collected for detecting P53 expression and macrophages infiltration by immunofluorescence, real-time PCR (qPCR) and western-blot. The differentiation and the inflammatory cytokines (TNF-α and IL-6) expression of THP-1, RAW264.7 and bone marrow derived macrophage (BMDM) cells, treating with Pifithrin-α (P53 inhibitor) or Nutlin-3a (P53 activator) under lipopolysaccharide (LPS) stimulation, were observed by flow cytometry, qPCR and ELISA. The severity of periodontitis, inflammatory cytokines expression and macrophages infiltration were measured in experimental periodontitis wild-type mice and p53 gene conditional knocked-out (p53-CKO) mice, which were established by ligation and LPS injection. A higher number of P53-positive macrophages was found infiltrated in periodontitis tissues. In vitro experiments showed that compared with Nutlin-3a, the proportion of M1-type macrophages and the expression of TNF-α and IL-6 were higher in Pifithrin-α treated cells under LPS stimulation. In vivo experimental periodontitis mice, the Pifithrin-α intraperitoneal injection group showed greater alveolar bone loss, higher levels of TNF-α and IL-6 secretion and more M1-type macrophages infiltration, while the Nutlin-3a intraperitoneal injection group were observed mild symptoms compared with mice in the periodontitis group. P53-CKO mice exhibited more severe periodontitis and more M1-type macrophages infiltrated in local tissues compared with wild-type mice. The activation of p53 gene could alleviate periodontitis by reducing M1-type macrophage polarization. P53 may serve as keeper in the progression of periodontitis, providing new insights into periodontitis treatment.

Size-optimized nuclear-targeting phototherapy enhances the type I interferon response for "cold" tumor immunotherapy.

There is growing interest in the effect of innate immune silencing in "cold" tumors, which always fail in the immune checkpoint blockade monotherapy using PD-L1 monoclonal antibodies (aPD-L1). Combination of aPD-L1 with photodynamic therapy, i.e., photoimmunotherapy, is a promising strategy to improve the mono immunotherapy. Nuclear-targeting nanoparticles could elicit a type I interferon (IFN)-mediated innate immune response and reverse the immunosuppressive microenvironment for long-term immunotherapy of "cold" tumors. Photosensitizers such as zinc phthalocyanine (ZnPc) have limited ability to target the nucleus and activate innate sensing pathways to minimize tumor recurrence. Additionally, the relationship between nanoparticle size and nuclear entry capacity remains unclear. Herein, graphene quantum dots (GQDs) were employed as aPD-L1 and ZnPc carriers. Three particle sizes (200 nm, 32 nm and 5 nm) of aPD-L1/ZnPc/GQD-PEG (PZGE) were synthesized and tested. The 5 nm nanoparticles achieved the best nuclear enrichment capacity contributing to their ultrasmall size. Notably, 5 nm PZGE-based photodynamic therapy enabled an amplification of the type I IFN-mediated innate immune response and could convert "immune-cold" tumors into "immune-hot" ones. Utilizing their size advantage to target the nucleus, 5 nm nanoparticles induced DNA damage and activated the type I IFN-mediated innate immune response, subsequently promoting cytotoxic T-lymphocyte infiltration and reversing negative PD-L1 expression. Furthermore, the nanoplatform we designed is promising for the effective suppression of distant oral squamous cell carcinoma. Thus, for the first time, this study presents a size design strategy for nuclear-targeted photo-controlled immune adjuvants and the nuclear-targeted phototherapy-mediated immunomodulatory functions of type I IFN innate immune signalling for "immune-cold" tumors. STATEMENT OF SIGNIFICANCE: The potential of commonly used photosensitizers to activate innate sensing pathways for producing type I IFNs is limited due to the lack of nuclear targeting. Facilitating the nuclear-targeting of photosensitizers to enhance innate immune response and execute long-term tumor killing effect would be a promising strategy for "cold" tumor photoimmunotherapy. Herein, we report an optimal size of PZGE nanoparticles that enable the nuclear-targeting of ZnPc, which reinforces the type I IFN-mediated innate immune response, synergistically reversing "cold tumors" to "hot tumors" for effective primary and distant tumor photoimmunotherapy. This work highlights the marked efficacy of ultrasmall nuclear-located nanocarriers and offers new insight into "immune-cold tumors" via prominent innate immune activation mediated by nuclear-targeting photoimmunotherapy.

A cuproptosis and copper metabolism-related gene prognostic index for head and neck squamous cell carcinoma.

Background: The purpose of this study was to identify the prognostic value of cuproptosis and copper metabolism-related genes, to clarify their molecular and immunological characteristics, and to elucidate their benefits in head and neck squamous cell carcinoma (HNSCC). Methods: The details of human cuproptosis and copper metabolism-related genes were searched and filtered from the msigdb database and the latest literature. To identify prognostic genes associated with cuproptosis and copper metabolism, we used least absolute shrinkage and selection operator regression, and this coefficient was used to set up a prognostic risk score model. HNSCC samples were divided into two groups according to the median risk. Afterwards, the function and immune characteristics of these genes in HNSCC were analyzed. Results: The 14-gene signature was constructed to classify HNSCC patients into low-risk and high-risk groups according to the risk level. In the The Cancer Genome Atlas (TCGA) cohort, the overall survival (OS) rate of the high-risk group was lower than that of the low-risk group (P < 0.0001). The area under the curve of the time-dependent Receiver Operator Characteristic (ROC) curve assessed the good performance of the genetic signature in predicting OS and showed similar performance in the external validation cohort. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment assays and Protein-Protein Interaction (PPI) protein networks have been used to explore signaling pathways and potential mechanisms that were markedly active in patients with HNSCC. Furthermore, the 14 cuproptosis and copper metabolism-related genes were significantly correlated with the immune microenvironment, suggesting that these genes may be linked with the immune regulation and development of HNSCC. Conclusions: Our results emphasize the significance of cuproptosis and copper metabolism as a predictive biomarker for HNSCC, and its expression levels seem to be correlated with immune- related features; thus, they may be a possible biomarker for HNSCC prognosis.

The niche-specialist and age-related oral microbial ecosystem: crosstalk with host immune cells in homeostasis.

Although characterization of the baseline oral microbiota has been discussed, the current literature seems insufficient to draw a definitive conclusion on the interactions between the microbes themselves or with the host. This study focuses on the spatial and temporal characteristics of the oral microbial ecosystem in a mouse model and its crosstalk with host immune cells in homeostasis. The V3V4 regions of the 16S rRNA gene of 20 samples from four niches (tongue, buccal mucosa, keratinized gingiva and hard palate) and 10 samples from two life stages (adult and old) were analysed. Flow cytometry (FCM) was used to investigate the resident immune cells. The niche-specialist and age-related communities, characterized based on the microbiota structure, interspecies communications, microbial functions and interactions with immune cells, were addressed. The phylum Firmicutes was the major component in the oral community. The microbial community profiles at the genus level showed that the relative abundances of the genera Bacteroides, Lactobacillus and Porphyromonas were enriched in the gingiva. The abundance of the genera Streptococcus, Faecalibaculum and Veillonella was increased in palatal samples, while the abundance of Neisseria and Bradyrhizobium was enriched in buccal samples. The genera Corynebacterium, Stenotrophomonas, Streptococcus and Fusobacterium were proportionally enriched in old samples, while Prevotella and Lacobacillus were enriched in adult samples. Network analysis showed that the genus Lactobacillus performed as a central node in the buccal module, while in the gingiva module, the central nodes were Nesterenkonia and Hydrogenophilus. FCM showed that the proportion of Th1 cells in the tongue samples (38.18 % [27.03-49.34 %]) (mean [range]) was the highest. The proportion of γδT cells in the buccal mucosa (25.82 % [22.1-29.54 %]) and gingiva (20.42 % [18.31-22.53 %]) samples was higher (P<0.01) than those in the palate (14.18 % [11.69-16.67 %]) and tongue (9.38 % [5.38-13.37 %] samples. The proportion of Th2 (31.3 % [16.16-46.44 %]), Th17 (27.06 % [15.76-38.36 %]) and Treg (29.74 % [15.71-43.77 %]) cells in the old samples was higher than that in the adult samples (P<0.01). Further analysis of the interplays between the microbiomes and immune cells indicated that Th1 cells in the adult group, nd Th2, Th17 and Treg cells in the old group were the main immune factors strongly associated with the oral microbiota. For example, Th2, Th17 and Treg cells showed a significantly positive correlation with age-related microorganisms such as Sphingomonas, Streptococcus and Acinetobacter, while Th1 cells showed a negative correlation. Another positive correlation occurred between Th1 cells and several commensal microbiomes such as Lactobacillus, Jeotgalicoccus and Sporosarcina. Th2, Th17 and Treg cells showed the opposite trend. Together, our findings identify the niche-specialist and age-related characteristics of the oral microbial ecosystem and the potential associations between the microbiomes and the mucosal immune cells, providing critical insights into mucosal microbiology.

Dynamic changes of exhaustion features in T cells during oral carcinogenesis.

OBJECTIVES: This study aimed to clarify the dynamic changes of exhaustion features in T cells during oral carcinogenesis. MATERIALS AND METHODS: Mice were randomly divided into 4NQO group and control group. The exhaustion features of CD4+ and CD8+ T cells of both groups were detected by flow cytometry. Furthermore, multiplex immunohistochemistry was used to evaluate the expression of inhibitory receptors in human normal, dysplastic, and carcinogenesis tissues. Finally, anti-PD-1 antibody treatment was performed at the early premalignant phase of oral carcinogenesis. RESULTS: The proportion of naive T cells in 4NQO group was lower than those in control group, while the proportion of effector memory T cells was higher in 4NQO group. The expression of inhibitory receptors on CD4+ and CD8+ T cells increased gradually during carcinogenesis. In contrast, the secretion of cytokines by CD4+ and CD8+ T cells decreased gradually with the progression stage. Strikingly, those changes occurred before the onset of oral carcinogenesis. The expression of inhibitory receptors on T cells increased gradually as the human tissues progressed from normal, dysplasia to carcinoma. Interestingly, PD-1 blockade at the early premalignant phase could reverse carcinogenesis progression by restoring T cell function. CONCLUSIONS: T-cell dysfunction was established at the early premalignant phase of oral carcinogenesis; PD-1 blockade at the early premalignant phase can effectively reverse T-cell exhaustion features and then prevent carcinogenesis progression.

TDO2+ myofibroblasts mediate immune suppression in malignant transformation of squamous cell carcinoma.

Characterization of the dynamic change in the immunological landscape during malignant transformation from precancerous lesions to cancerous lesions in squamous cell carcinoma (SCC) is critical for the application of immunotherapy. Here, we performed single-cell RNA-Seq (scRNA-Seq) of 131,702 cells from 13 cancerous tissues of oral squamous cell carcinoma (OSCC), 3 samples of precancerous oral leukoplakia, and 8 adjacent normal samples. We found that tumor-infiltrating CD4+ and CD8+ T cells were functionally inhibited by immunosuppressive ligands expressed on various types of myeloid cells or neutrophils in the process of oral carcinogenesis. Notably, we identified a subset of myofibroblasts that exclusively expressed tryptophan 2,3-dioxygenase (TDO2). These TDO2+ myofibroblasts were located distally from tumor nests, and both CD4+ and CD8+ T cells were enriched around them. Functional experiments revealed that TDO2+ myofibroblasts were more likely to possess the ability for chemotaxis toward T cells but induced the transformation of CD4+ T cells into Tregs and caused CD8+ T cell dysfunction. We further showed that use of the TDO2 inhibitor LM10 attenuated the inhibitory states of T cells, restored the T cell antitumor response, and prevented the progression of OSCC malignant transformation in murine models. Our study reveals a multistep transcriptomic landscape of OSCC and demonstrates that TDO2+ myofibroblasts are potential targets for immunotherapy.

Regulation of autoimmune disease progression by Pik3ip1 through metabolic reprogramming in T cells and therapeutic implications.

Metabolic alterations could profoundly affect immune functions and influence the progression and outcome of autoimmune diseases. However, the detailed mechanisms and their therapeutic potential remain to be defined. Here, we show that phosphatidylinositide 3-kinase interacting protein 1 (Pik3ip1), a newly identified negative immune regulator, is notably down-regulated in several major autoimmune diseases through a previously unidentified mechanism mediated by interleukin-21/p38 mitogen-activated protein kinase/a disintegrin and metalloprotease-17 (ADAM17) pathway. Down-regulation of Pik3ip1 in T cells causes a major metabolic shift from oxidative phosphorylation toward aerobic glycolysis, leading to their overactivation and aggressive disease progression in experimental autoimmune encephalomyelitis (EAE) mouse model. Suppression of hypoxia-inducible factor 1α (Hif1α) or pharmacologic inhibition of glycolysis could reverse these phenotypes and largely mitigate EAE severity. Our study reveals a previously unrecognized role of Pik3ip1 in metabolic regulation that substantially affects the inflammatory loop in the autoimmune setting and identifies the Pik3ip1/Hif1α/glycolysis axis as a potential therapeutic target for treatment of autoimmune diseases.

A comprehensive profile of TCF1+ progenitor and TCF1- terminally exhausted PD-1+CD8+ T cells in head and neck squamous cell carcinoma: implications for prognosis and immunotherapy.

The heterogeneity of exhausted T cells (Tex) is a critical determinant of immune checkpoint blockade therapy efficacy. However, few studies have explored exhausted T cell subpopulations in human cancers. In the present study, we examined samples from two cohorts of 175 patients with head and neck squamous cell cancer (HNSCC) by multiplex immunohistochemistry (mIHC) to investigate two subsets of Tex, CD8+PD1+TCF1+ progenitor exhausted T cells (TCF1+Texprog) and CD8+PD1+TCF1- terminally exhausted T cells (TCF1-Texterm). Moreover, fresh tumor samples from 34 patients with HNSCC were examined by flow cytometry and immunohistochemistry to further investigate their properties and cytotoxic capabilities and their correlation with regulatory T cells (Tregs) in the tumor immune microenvironment (TIME). mIHC and flow cytometry analysis showed that TCF1-Texterm represented a greater proportion of CD8+PD1+Tex than TCF1+Texprog in most patients. TCF1+Texprog produced abundant TNFα, while TCF1-Texterm expressed higher levels of CD103, TIM-3, CTLA-4, and TIGIT. TCF1-Texterm exhibited a polyfunctional TNFα+GZMB+IFNγ+ phenotype; and were associated with better overall survival and recurrence-free survival. The results also indicated that larger proportions of TCF1-Texterm were accompanied by an increase in the proportion of Tregs. Therefore, it was concluded that TCF1-Texterm was the major CD8+PD1+Tex subset in the HNSCC TIME and that these cells favor patient survival. A high proportion of TCF1-Texterm was associated with greater Treg abundance.

Crosstalk between the oral microbiota, mucosal immunity, and the epithelial barrier regulates oral mucosal disease pathogenesis.

Oral mucosal disease (OMD), which is also called soft tissue oral disease, is described as a series of disorders or conditions affecting the mucosa and soft tissue in the oral cavity. Its etiology is unclear, but emerging evidence has implicated the influence of the composition of the oral mucosa and saliva-resident microbiota. In turn, this dysbiosis effects the immune response balance and epithelial barrier function, followed by the occurrence and progression of OMD. In addition, oral microbial dysbiosis is diverse in different types of diseases and different disease progressions, suggesting that key causal pathogens may exist in various oral pathologies. This narrative literature review primarily discusses the most recent findings focusing on how microbial dysbiosis communicates with mucosal adaptive immune cells and the epithelial barrier in the context of five representative OMDs, including oral candidiasis (OC), oral lichen planus (OLP), recurrent aphthous ulcer (RAU), oral leukoplakia (OLK), and oral squamous cell carcinoma (OSCC), to provide new insight into the pathogenetic mechanisms of OMDs.

TCF-1 maintains CD8+ T cell stemness in tumor microenvironment.

T cell factor 1 (TCF-1) is a transcription factor (TF) of the canonical Wnt signaling pathway that encoded by the Tcf7. The crucial role of TCF-1 in T cell development and memory formation has been widely recognized. Recent studies have demonstrated that exhausted CD8+ T cell with the expression of TCF-1 may have inspiring function to amplify immunoreaction and improve the response to immunotherapy in chronic viral infection and cancer. In this short review, we summarized recent progress in intratumoral exhausted CD8+ T cells expressing TCF-1 that represent a fantastic subset with stem cell-like properties that associated with improved antitumor immunity and response to immune checkpoint blockade (ICB).

Prognostic value of VISTA in solid tumours: a systematic review and meta-analysis.

In the last few years, V-domain Ig-containing suppressor of T cell activation(VISTA) has been reported as a prognostic biomarker in articles including various solid tumours. However, their conclusions have been controversial. For this reason, we performed this meta-analysis to further verify the prognostic value of VISTA in solid tumours. All relevant literature was identified from PubMed, Embase, the Cochrane Library and Web of Science. Ten studies, including 2, 440 patients, were eligible for the analysis. The pooled results showed that high expression of VISTA was associated with favourable overall survival (OS) than that seen with low expression of VISTA (7 studies, hazard ratio (HR) = 0.75, 95% confidence interval (CI): 0.66-0.86, P < 0.001). In addition, high expression of VISTA significantly correlated with high numbers of CD8 (+) tumour infiltrating lymphocytes (TILs) (3 studies, risk ratio (RR) = 1.80, 95% CI: 1.41-2.31, P < 0.001). In conclusion, these results indicate that VISTA is a potential prognostic biomarker in solid tumours.

Prognostic value of tertiary lymphoid structure and tumour infiltrating lymphocytes in oral squamous cell carcinoma.

Tertiary lymphoid structures (TLS) are ectopic lymphoid structures in cancers that are largely associated with favourable prognosis. However, the prognostic value of TLSs in oral squamous cell carcinoma (OSCC) is largely unknown, and the association between tumour infiltrating lymphocytes (TILs) and TLSs has been rarely explored in OSCC. In this study, associated markers of TLS, including peripheral node address (PNAd) in high endothelial venules, CD20 in B cells and CD3 in T cells, were examined in 168 OSCC patients, and survival analysis was performed between TLS-positive and TLS-negative cohorts. We detected the presence of TILs by staining CD8+ cytotoxic T cells and CD57+ NK cells as well. TLSs appeared as highly organized structures in 45 (26.8%) cases. TLS-positive patients had a better 5-year overall survival (OS) rate (88.9% vs. 56.1%, P < 0.001) and relapse-free survival (RFS) rate (88.9% vs. 63.4%, P = 0.002). Moreover, the presence of TLS was an independent prognostic factor for both the 5-year OS rate (hazard ratio [HR] = 3.784; 95% confidence interval [CI], 1.498-9.562) and RFS rate (HR = 3.296; 95% CI, 1.279-8.490) in multivariate analysis. Furthermore, a higher density of CD8+ T cells and CD57+ NK cells was found in TLS-positive sections than in TLS-negative counterparts (P < 0.001), and their combination provided a higher predictive accuracy (AUC = 0.730; 95% CI, 0.654-0.805). In conclusion, our results suggest that TLS is an independent positive prognostic factor for OSCC patients. These findings provide a theoretical basis for the future diagnostic and therapeutic value of TLSs in OSCC treatment.

High Abundance of Intratumoral γδ T Cells Favors a Better Prognosis in Head and Neck Squamous Cell Carcinoma: A Bioinformatic Analysis.

γδ T cells are a small subset of unconventional T cells that are enriched in the mucosal areas, and are responsible for pathogen clearance and maintaining integrity. However, the role of γδ T cells in head and neck squamous cell carcinoma (HNSCC) is largely unknown. Here, by using RNA-seq data from The Cancer Genome Atlas (TCGA), we discovered that HNSCC patients with higher levels of γδ T cells were positively associated with lower clinical stages and better overall survival, and high abundance of γδ T cells was positively correlated with CD8+/CD4+ T cell infiltration. Gene ontology and pathway analyses showed that genes associated with T cell activation, proliferation, effector functions, cytotoxicity, and chemokine production were enriched in the group with a higher γδ T cell abundance. Furthermore, we found that the abundance of γδ T cells was positively associated with the expression of the butyrophilin (BTN) family proteins BTN3A1/BTN3A2/BTN3A3 and BTN2A1, but only MICB, one of the ligands of NKG2D, was involved in the activation of γδ T cells, indicating that the BTN family proteins might be involved in the activation and proliferation of γδ T cells in the tumor microenvironment of HNSCC. Our results indicated that γδ T cells, along with their ligands, are promising targets in HNSCC with great prognostic values and treatment potentials.