Downregulation of LAMB3 Altered the Carcinogenic Properties of Human Papillomavirus 16-Positive Cervical Cancer Cells.

Nearly all cervical cancer cases are caused by infection with high-risk human papillomavirus (HR-HPV) types. The mechanism of cervical cell transformation is related to the powerful action of viral oncoproteins and cellular gene alterations. Transcriptomic data from cervical cancer and normal cervical cells were utilized to identify upregulated genes and their associated pathways. The laminin subunit beta-3 (LAMB3) mRNAwas overexpressed in cervical cancer and was chosen for functional analysis. The LAMB3 was predominantly expressed in the extracellular region and the plasma membrane, which play a role in protein binding and cell adhesion molecule binding, leading to cell migration and tissue development. LAMB3 was found to be implicated in the pathway in cancer and the PI3K-AKT signaling pathway. LAMB3 knockdown decreased cell migration, invasion, anchorage-dependent and anchorage-independent cell growth and increased the number of apoptotic cells. These effects were linked to a decrease in protein levels involved in the PI3K-AKT signaling pathway and an increase in p53 protein. This study demonstrated that LAMB3 could promote cervical cancer cell migration, invasion and survival.

Differential expression of PEA3 in odontogenic cysts and tumors.

BACKGROUND: PEA3 transcription factor has been identified as a downstream target of the MAPK and PI3K pathways, and PEA3 overexpression has been observed in a variety of tumor types. We aimed to evaluate PEA3 expression in odontogenic cysts and tumors and compare the expression among odontogenic lesions. In addition, the correlations between PEA3 expression and clinicopathological characteristics of conventional ameloblastoma and unicystic ameloblastoma were investigated. METHODS: This study was performed on 165 samples of odontogenic cysts and tumors including 20 dentigerous cysts, 20 odontogenic keratocysts, 16 adenomatoid odontogenic tumors, 5 ameloblastic fibromas, 45 unicystic ameloblastomas, and 59 conventional ameloblastomas. The sections were immunohistochemically stained with mouse monoclonal anti-PEA3 antibody and PEA3 expression was evaluated as the immunoreactive score. RESULTS: PEA3 expression was absent in all dentigerous cysts (DCs) and odontogenic keratocysts, while all adenomatoid odontogenic tumors showed either no (75%) or low (25%) expression of PEA3. Most of the ameloblastic fibromas (60%) displayed no PEA3 expression. A high expression of PEA3 was observed in a substantial number of unicystic ameloblastomas (48.9%) and conventional ameloblastomas (49.2%) in our study. PEA3 expression in DCs, odontogenic keratocysts and adenomatoid odontogenic tumors were significantly different from that in conventional ameloblastomas and that in unicystic ameloblastomas (p < 0.05). The expression of PEA3 was significantly different in the age groups of unicystic ameloblastomas and histological subtypes of conventional ameloblastomas (p < 0.05). CONCLUSION: PEA3 overexpression is predominant in unicystic ameloblastomas and conventional ameloblastomas compared to other odontogenic lesions, indicating a pivotal role of PEA3 as a downstream effector of MAPK pathway in these two odontogenic lesions.

An integrative analysis of genome-wide methylation and expression in ameloblastoma: A pilot study.

OBJECTIVE: DNA methylation regulates the expression of various genes involved in tumorigenesis. Ameloblastoma is a benign odontogenic jaw tumor. It is locally aggressive with a high level of recurrence. A delay in treatment can lead to severe facial disfigurement. To the best of our knowledge, this is the first integrated analysis of DNA methylation and gene expression in ameloblastoma with the aim to identify genes that may be regulated by DNA methylation. MATERIALS AND METHODS: We used an Infinium MethylationEPIC array to measure genome-wide methylation and the Illumina HiSeq platform to obtain gene expression data in ameloblastoma tissues from five patients and dental follicles from three healthy subjects. An integration analysis was performed using City of Hope CpG Island Analysis Pipeline software. RESULTS: We identified 25,255 differentially methylated CpG sites and 17 differentially methylated CpG islands; six of the islands were negatively correlated with the expression of BAIAP2, DUSP6, FGFR2, FOXF2, NID2, and PAK6. Pyrosequencing and immunostaining techniques were further used to validate FGFR2, NID2, and PAK6. CONCLUSIONS: This analysis identifies a group of novel genes that may be regulated by DNA methylation and will possibly lead to new insights into the pathology and invasion mechanism of ameloblastoma.

Transcriptomic analysis of peripheral blood mononuclear cells in head and neck squamous cell carcinoma patients.

OBJECTIVES: To investigate the gene expression profile of peripheral blood mononuclear cells (PBMCs) from head and neck squamous cell carcinoma (HNSCC), including oral cancer (OC) and oropharyngeal cancer (OPC) patients, and compare them with healthy controls (HC). MATERIALS AND METHODS: Transcriptomic analysis of PBMCs was performed by RNA-sequencing. The upregulated candidate genes were selected for validation by quantitative real-time polymerase chain reaction (qPCR). In addition, related plasma protein levels were determined by enzyme-linked immunosorbent assay (ELISA). RESULTS: Three significantly upregulated genes, including high mobility group nucleosomal binding domain 2 (HMGN2), folate receptor gamma (FOLR3), and amphiregulin (AREG), were selected. In the first cohort, the results showed that only HMGN2 expression was significantly increased in OC patients. In the larger sample size, the overall results demonstrated that HMGN2 expression had a tendency to increase in both OC and OPC patients compared with HC. Interestingly, the plasma HMGN2 (HMG-17) protein level exhibited the same trend as that observed at the transcriptional level. CONCLUSION: HMGN2 expression and plasma HMG-17 (HMGN2 protein) were increased in both cancer patients compared with HC. This gene may be important for further functional studies in the PBMCs of HNSCC patients.

Alteration of Extracellular Matrix Components in the Anterior Pituitary Gland of Neonatal Rats Induced by a Maternal Bisphenol A Diet during Pregnancy.

The extracellular matrix (ECM) plays crucial roles in the anterior pituitary gland via the mechanism of cell-ECM interaction. Since bisphenol A (BPA), a well-known endocrine disruptor, can cross through the placenta from mother to fetus and bind with estrogen receptors, cell populations in the neonatal anterior pituitary gland could be the target cells affected by this chemical. The present study treated maternal rats with 5000 µg/kg body weight of BPA daily throughout the pregnancy period and then investigated the changes in ECM-producing cells, i.e., pericytes and folliculostellate (FS) cells, including their ECM production in the neonatal anterior pituitary at Day 1. We found that pericytes and their collagen synthesis reduced, consistent with the increase in the number of FS cells that expressed several ECM regulators-matrix metalloproteinase (MMP) 9 and the tissue inhibitors of metalloproteinase (TIMP) family. The relative MMP9/TIMP1 ratio was extremely high, indicating that the control of ECM homeostasis was unbalanced. Moreover, transmission electron microscopy showed the unorganized cell cluster in the BPA-treated group. This study revealed that although the mother received BPA at the "no observed adverse effect" level, alterations in ECM-producing cells as well as collagen and the related ECM balancing genes occurred in the neonatal anterior pituitary gland.

Evaluation of NID2 promoter methylation for screening of Oral squamous cell carcinoma.

BACKGROUND: Oral squamous cell carcinoma (OSCC) is an aggressive human malignancy. Because of late diagnosis and recurrence of OSCC, the treatment of patients with OSCC is often ineffective. Thus, finding novel biomarkers of OSCC are essential. Here we derived a methylation marker by utilizing methylation microarray data and testing its capacity in cross-sectional study designed for OSCC detection and screening. METHODS: According to bioinformatics analysis of total of 27,578 cg sites, cg22881914 of Nidogen 2 (NID2) methylation was selected for evaluation. Next, we confirmed the methylation status by bisulfite sequencing from the microdissected OSCC cells in comparison with the microdissected oral epithelia. Subsequently, we developed a simple technique using real-time PCR with the specific probe to examine the ability for the detection of OSCC in the oral epithelial samples, which included 103 oral rinse and 82 oral swab samples. RESULTS: Based on the comparison of microdissected tissue, cg22881914 of NID2 was proved to be methylated in most OSCC cells but unmethylated in the normal oral epithelia. Furthermore, the methylated NID2-relied quantitative PCR approach has demonstrated that this marker assists in distinguishing among patients with OSCC from normal oral epithelia, smokers, and patients with oral lichen planus using the non-invasive oral rinse and swab samples. CONCLUSIONS: Specific methylation at cg22881914 of NID2 of OSCC could be used as an important potential marker for detecting OSCC. Thus, to certify the utility of this marker, further studies with a larger sample size are needed.

Distribution of the Epstein-Barr virus in the normal stomach and gastric lesions in Thai population.

The Epstein-Barr virus (EBV) is one of the infectious agents found in stomach tissue. Recently, EBV-associated gastric carcinoma (EBVaGC) was classified as a new subtype of gastric carcinoma. To date, there is a lack of knowledge about the distribution and prevalence of EBV infection in both the normal stomach and various gastric lesions, including EBVaGC, in the Thai population. In this study, we detected EBV in the normal stomach (NS; n = 19), chronic gastritis (CG; n = 36), intestinal metaplasia (IM; n = 40), gastric dysplasia (GD; n = 15), and gastric adenocarcinoma (GC; n = 33) by polymerase chain reaction (PCR) amplification of the latent membrane protein (LMP1) gene of EBV. EBV-PCR amplification was positive in 42.1%, 36.1%, 22.5%, 13.3%, and 33.3% of NS, CG, IM, GD, and GC, respectively. For further clarification in EBVaGC, we performed EBV-encoded small RNA in situ hybridization (EBER-ISH) in PCR-positive cases of GD and GC. Four GC cases were EBER-ISH positive (12.1%), while both GD cases were EBER-ISH negative. In addition, we determined the distribution of the EBV strain (type A or B) based on EBNA3C sequence and EBV variants based on LMP1 variation (wild-type and 30-bp deletion variants; wt-LMP1 or del-LMP1). The results showed that type A and wt-LMP1 were the most prevalent in all lesions. In conclusion, EBV is common in both the NS and gastric lesions, and the frequency of EBVaGC was 12.1% in Thai patients.

Head and neck squamous cell carcinoma drives long interspersed element-1 hypomethylation in the peripheral blood mononuclear cells.

OBJECTIVE: Alteration of long interspersed element-1 (LINE-1) methylation in peripheral blood mononuclear cells (PBMCs) has been simultaneously activated to breast carcinogenesis due to its secretion. To evaluate the effect in head and neck squamous cell carcinoma (HNSCC), LINE-1 methylation levels and patterns have been measured both in vitro and in vivo. METHODS: Analysis of LINE-1 methylation in cocultured models between HNSCC cell lines and normal PBMCs was performed. The observation of PBMCs of HNSCC patients compared to PBMCs of normal controls was performed using the semiquantitative combined bisulfite restriction analysis technique. RESULTS: Downregulation of LINE-1 methylation was significantly found in the PBMCs cocultured with all HNSCC cell lines compared to normal PBMCs. Likewise, a reduction in LINE-1 methylation levels was observed in PBMCs of HNSCC compared to normal PBMCs (p < 0.0001). Receiver operating characteristic analysis demonstrated the potential of the unmethylated alleles (u Cu C) of LINE-1 for distinguishing the PBMCs of HNSCC patients from normal controls with 100% sensitivity and specificity. CONCLUSION: Our data supported that the alteration of LINE-1 methylation levels in PBMCs was influenced by HNSCC secretions. Moreover, the unmethylated LINE-1 allele of PBMCs was proved to be an effective tumor marker and possesses a potential as HNSCC diagnostic tool.

The Occurrence of Vibrionaceae, Staphylococcaceae, and Enterobacteriaceae in Green Turtle Chelonia mydas Rearing Seawater.

In this study, levels of Vibrionaceae, Staphylococcaceae, and Enterobacteriaceae were observed in seawater from juvenile green turtle Chelonia mydas rearing tanks and in the incoming coastal seawater (the water supply). Bacterial loads were compared between the incoming coastal seawater and two different rearing conditions: in cement tanks at a low stocking density and in fiberglass tanks at a high stocking density. The total bacterial counts in seawater from fiberglass tanks were statistically greater than those in cement tanks. The nonlactose and lactose fermenting enterobacteria, tellurite-reducing bacteria, and total plate counts in water from all rearing containers were greater than those in coastal seaweater by a logarithmic fold change of 2--3. Differences in bacterial population structure of the incoming coastal seawater and rearing water were also addressed. The results from biochemical identification of 344 isolates revealed that the bacteria that were commonly found in water samples were Citrobacter spp., Enterobacteria spp., Edwardsiella spp., Staphylococcus spp., Staphylococcus aureus, Photobacterium spp., Vibrio alginolyticus, and Vibrio spp. Conclusively, the microbiological monitoring of rearing water provides important and essential information on the management of aquatic animal health and husbandry.

Differences in LINE-1 methylation between endometriotic ovarian cyst and endometriosis-associated ovarian cancer.

BACKGROUND: Endometriosis in endometriosis-associated ovarian cancer (EAOC) refers to lesions that can derive from endometriotic ovarian cysts (ECs) that form in the ovarian endometrium with the potential to transform into full-blown ovarian cancer. Hypomethylation of long interspersed element-1 (LINE-1 or L1) is a common epigenomic event in several cancers and is strongly associated with ovarian cancer progression. OBJECTIVES: To evaluated alterations in LINE-1 methylation between EC, ovarian endometrioid adenocarcinoma (OEA), EAOC, and ovarian clear cell carcinoma (OCC). METHODS/ MATERIALS: First, LINE-1 methylation status in 19 normal endometrium, 29 EC, 35 OCC, and 22 OEA tissues from unrelated samples were compared. Then, specific areas of eutopic endometrium, contiguous endometriosis, and cancer arising from 16 EAOCs were collected by microdissection and analyzed for LINE-1 methylation status. RESULTS: The total LINE-1 methylation levels were significantly different among the endometrium, endometriosis, and ovarian cancer (P < 0.001). A stepwise decrease in LINE-1 methylation was observed in the following order: normal endometrium, EC, OEA, and OCC. Interestingly, endometriosis in EAOC of both OEA (P = 0.016) and OCC (P = 0.003) possessed a higher percentage of LINE-1 unmethylated loci than EC. CONCLUSION: Our data implicate that LINE-1 hypomethylation is an early molecular event involved in OEA and OCC malignant transformation. Precise measurements of LINE-1 methylation may help to distinguish EC and endometriosis in EAOC.

Identification of Molecular Mechanisms of Ameloblastoma and Drug Repositioning by Integration of Bioinformatics Analysis and Molecular Docking Simulation.

Background: Ameloblastoma (AM) is a benign tumor locally originated from odontogenic epithelium that is commonly found in the jaw. This tumor makes aggressive invasions and has a high recurrence rate. This study aimed to investigate the differentially expressed genes (DEGs), biological function alterations, disease targets, and existing drugs for AM using bioinformatics analysis. Methods: The data set of AM was retrieved from the GEO database (GSE132474) and identified the DEGs using bioinformatics analysis. The biological alteration analysis was applied to Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. Protein-protein interaction (PPI) network analysis and hub gene identification were screened through NetworkAnalyst. The transcription factor-protein network was constructed via OmicsNet. We also identified candidate compounds from L1000CDS2 database. The target of AM and candidate compounds were verified using docking simulation. Results: Totally, 611 DEGs were identified. The biological function enrichment analysis revealed glycosaminoglycan and GABA (γ-aminobutyric acid) signaling were most significantly up-regulated and down-regulated in AM, respectively. Subsequently, hub genes and transcription factors were screened via the network and showed FOS protein was found in both networks. Furthermore, we evaluated FOS protein to be a therapeutic target in AMs. Candidate compounds were screened and verified using docking simulation. Tanespimycin showed the greatest affinity binding value to bind FOS protein. Conclusions: This study presented the underlying molecular mechanisms of disease pathogenesis, biological alteration, and important pathways of AMs and provided a candidate compound, Tanespimycin, targeting FOS protein for the treatment of AMs.

Protein expression analysis for predicting recurrent laryngeal squamous cell carcinoma.

The expression of a number of proteins plays a major role in predicting recurrent laryngeal squamous cell carcinoma (LSCC). Thus, the aim of the present study was to investigate the expression of 16 selected proteins as prognostic indicators for recurrent and non-recurrent LSCC. Samples from a total of 41 patients with LSCC were investigated by immunohistochemistry. Digital image analysis was performed, and various associated factors were calculated. Histoscore (H-score) and receiver operating characteristic curves were used to divide protein expression in high and low for predicting disease recurrence. Disease-free survival (DFS) curves, crude hazard ratios (HRs) and adjusted HRs were analyzed and compared. Significantly different H-scores were found between the recurrent and non-recurrent groups in terms of pRb and c-Met expression. pRb was expressed at high levels in recurrent LSCC, while c-Met was expressed at low levels. Patients with low pRb expression had a longer DFS than those with high pRb expression (log-rank χ2, 5.161; P=0.023). Patients with high c-Met expression had a longer DFS than those with low c-Met expression (log-rank χ2, 6.441; P=0.011). Moreover, patients with high pRb expression and low c-Met expression had the shortest DFS (log-rank χ2, 11.827; P=0.008). Differentiated histological factors had an impact on the risk of recurrence (Cox regression test; crude HR, 9.53; 95% confidence interval, 1.214-74.819; P=0.032). The present study demonstrated that the grading of differentiated squamous cell carcinoma, pRb and c-Met expression are the most useful prognostic factors for the prediction of recurrent LSCC. These might be further applied as potential markers for clinical use.

5-Methylcytosine immunohistochemistry for predicting cutaneous melanoma prognosis.

There is a correlation between DNA methylation and the diseased stage and poor survival. 5-methylcytosine (5-mC) is one of the epigenetic modifications of bases that researchers focus on. Staining with 5-mC immunohistochemistry was used to examine pathological samples taken from individuals diagnosed with cutaneous melanoma. Between Breslow levels 2 and 4, there was a significant difference in the H-score of 5-mC expression (p = 0.046). A significant reduction in 5-mC expression H-scores was seen in patients who were diagnosed with ulcers (p = 0.039). It was shown that patients with low 5-mC had a significantly worse overall survival rate (p = 0.027).

Exploring protein profiles and hub genes in ameloblastoma.

Ameloblastoma (AM) is a prominent benign odontogenic tumor characterized by aggressiveness, likely originating from tooth-generating tissue or the dental follicle (DF). However, proteomic distinctions between AM and DF remain unclear. In the present study, the aim was to identify the distinction between AM and DF in terms of their proteome and to determine the associated hub genes. Shotgun proteomics was used to compare the proteomes of seven fresh-frozen AM tissues and five DF tissues. Differentially expressed proteins (DEPs) were quantified and subsequently analyzed through Gene Ontology-based functional analysis, protein-protein interaction (PPI) analysis and hub gene identification. Among 7,550 DEPs, 520 and 216 were exclusive to AM and DF, respectively. Significant biological pathways included histone H2A monoubiquitination and actin filament-based movement in AM, as well as pro-B cell differentiation in DF. According to PPI analysis, the top-ranked upregulated hub genes were ubiquitin C (UBC), breast cancer gene 1 (BRCA1), lymphocyte cell-specific protein-tyrosine kinase (LCK), Janus kinase 1 and ATR serine/threonine kinase, whereas the top-ranked downregulated hub genes were UBC, protein kinase, DNA-activated, catalytic subunit (PRKDC), V-Myc avian myelocytomatosis viral oncogene homolog (MYC), tumor protein P53 and P21 (RAC1) activated kinase 1. When combining upregulated and downregulated genes, UBC exhibited the highest degree and betweenness values, followed by MYC, BRCA1, PRKDC, embryonic lethal, abnormal vision, Drosophila, homolog-like 1, myosin heavy chain 9, amyloid beta precursor protein, telomeric repeat binding factor 2, LCK and filamin A. In summary, these findings contributed to the knowledge on AM protein profiles, potentially aiding future research regarding AM etiopathogenesis and leading to AM prevention and treatment.

CpG methylation changes in Alu repetitive sequences in normal aging due to diastolic hypertension in human dermal fibroblasts from the facial area.

Aging fibroblasts, an important factor contributing to skin aging, are affected by numerous mechanisms, including alterations in DNA methylation and age-related diseases. The current study aimed to investigate the role of Alu methylation in aging fibroblasts and hypertension. The Alu methylation levels in dermal fibroblasts obtained from patients of different ages and blood pressure status were analyzed using the combined bisulfite restriction analysis technique. An inverse correlation was observed between Alu methylation in dermal fibroblasts and patient age. Dermal fibroblasts from the high-normal diastolic blood pressure group had higher Alu methylation levels compared with those from the normal group. The findings of the present study suggest that Alu methylation alterations can be observed with chronological aging and hypertension, and are a potential aging marker or therapeutic target.

Evaluation of P53 immunostaining in patients with cutaneous melanoma.

P53 is a tumor suppressor gene that is mutated in numerous types of cancer. The aim of the present study was to determine the frequency of this mutation in cutaneous melanomas and to conduct clinicopathological characteristics and clinical outcome association analyses with the P53 mutation. P53 immunohistochemical staining was used as a surrogate marker for P53 mutation analysis to assess P53 status. In the present study, 50 pathological samples of cutaneous melanoma from 2012 to 2018 at Chulalongkorn University (Bangkok, Thailand), were subjected to anti-P53 immunohistochemistry, followed by an examination of the association between P53 statuses and clinical and pathological characteristics, along with clinical outcomes. A positive staining for anti-P53 antibody was detected in 30% of patients (15/50) with cutaneous melanomas. Positivity was significantly associated with female sex, nodular histological subtype and Breslow level 4. Cox regression analysis revealed that an age >65.5 years and Breslow grade 4 disease were associated with mortality. The Kaplan-Meier curve revealed a shorter duration of recurrence time in the P53 mutation than P53 wild type. In the present study, P53 mutations in specific cases of cutaneous melanoma were identified. Notably, patients who were older and/or had a Breslow score of 4 exhibited an increased risk of mortality. These findings suggested the potential involvement of P53 mutations in cutaneous melanoma, highlighting the necessity for further investigations to improve understanding of their roles.

Distribution of Epstein-Barr virus in the oral cavity of Thais with various oral mucosal conditions.

Objectives: We aimed to examine the presence of EBV, EBV strains, and variants among 3 oral conditions including normal oral mucosa (NOM), oral potentially malignant disorders/oral cancer (OPMDs/OC) and non-OPMDs/OC in a group of Thais. Material and methods: Oral exfoliated cells were obtained from 315 participants living in the northeastern and central regions of Thailand. The participants were divided into 3 groups encompassing the NOM, the OPMDs/OC and the non-OPMDs/OC groups. The presence of EBV was first determined by PCR using primers for LMP1 gene. Subsequently, EBV strains of EBNA3c and variants based on LMP1 sequences were determined by real-time PCR. Results: The prevalence of EBV in OPMDs/OC, non-OPMDs/OC and NOM were 72.0 %, 56.2 %, and 27.2 % respectively. EBV type A, B and AB were found in 52.1 %, 32.1 % and 15.8 % of all positive samples, respectively. The percentage of participants with EBV type A was more prominent in the NOM group (72.0 %) compared to the non-OPMDs/OC (54.8 %) and the OPMDs/OC group (41.8 %) whereas EBV type B was higher in the OPMDs/OC group (35.8 %) compared to the non-OPMDs/OC (31.5 %) and the NOM (24.0 %) groups. Regarding EBV variants, 30-bp deletion LMP1 variant (del-LMP1) which is more associated with malignant transformation was predominately found in the OPMDs/OC (32.8 %) and the non-OPMDs/OC (38.4 %) groups compared to the NOM group (20.0 %). Conclusions: High frequency of EBV was demonstrated in the OPMDs/OC group. EBV type A was more predominant in the NOM group whereas EBV type B was more prevalent in the OPMDs/OC group. The del-LMP1 variant was more common in the OPMDs/OC and the non-OPMDs/OC groups.

High 4-1BB Expression in PBMCs and Tumor Infiltrating Lymphocytes (TILs) in Patients with Head and Neck Squamous Cell Carcinoma.

OBJECTIVE: 4-1BB is a costimulatory immune-activating molecule. Increased amounts of this protein have previously been found in the plasma of patients with oropharyngeal and oral cancer. Here, we focused on this molecule that functions as part of the immune system. We investigated 4-1BB in the peripheral blood mononuclear cells (PBMCs) and tumor infiltrating lymphocytes (TILs) of patients with head and neck squamous cell cancer (HNSCC). MATERIALS AND METHODS: The expression level of 4-1BB in the PBMCs was determined using real-time polymerase chain reaction (PCR). The TIMER (Tumor Immune Estimation Resource) web server was utilized to approximate the 4-1BB level in HNSCC TILs. Moreover, 4-1BB immunohistochemistry (IHC) was used to validate TILs in four organs of HNSCC, including oral cancer (OC), oropharyngeal cancer (OPC), sinonasal cancer (SNC), and laryngeal cancer (LC), in both the tumor area and adjacent normal epithelium. The difference in 4-1BB expression levels in various groups was assessed using a Kruskal-Wallis test and an independent sample t-test. RESULTS: The level of 4-1BB expression in PBMCs was highest in OPC, followed by OC and healthy controls (HC). Significant differences were discovered between HC and OPC and between OC and OPC. Bioinformatics revealed a substantial correlation between 4-1BB expression level and lymphocyte infiltration in HNSCC, including B cells, CD8+ T cells, and CD4+ T cells. IHC validation in HNSCC tissue revealed that the average number of 4-1BB positive TILs in all four HNSCC subtypes was considerably greater than the number of lymphocytes seen in adjacent normal tissue. Interestingly, the number of lymphocytes that were 4-1BB positive increased in relation to the TIL level. CONCLUSION: A higher number of 4-1BB expression levels were found in the PBMCs and TILs of HNSCC patients, implying that 4-1BB may be a promising approach for HNSCC patients to improve their immune function. It is important to study and create a treatment that uses 4-1BB medicine as well as existing drugs.

LINE-1 and Alu hypomethylation in mucoepidermoid carcinoma.

BACKGROUND: Mucoepidermoid carcinoma (MEC) can be classified into low-, intermediate-, and high-grade tumors based on its histological features. MEC is mainly composed of three cell types (squamous or epidermoid, mucous and intermediate cells), which correlates with the histological grade and reflects its clinical behavior. Most cancers exhibit reduced methylation of repetitive sequences such as Long INterspersed Element-1 (LINE-1) and Alu elements. However, to date very little information is available on the LINE-1 and Alu methylation status in MEC. The aim of this study was to investigate LINE-1 and Alu element methylation in MEC and compare if key differences in the methylation status exist between the three different cell types, and adjacent normal salivary gland cells, to see if this may reflect the histological grade. METHODS: LINE-1 and Alu element methylation of 24 MEC, and 14 normal salivary gland tissues were compared using Combine Bisulfite Restriction Analysis (COBRA). Furthermore, the three different cell types from MEC samples were isolated for enrichment by laser capture microdissection (LCM), essentially to see if COBRA was likely to increase the predictive value of LINE-1 and Alu element methylation. RESULTS: LINE-1 and Alu element methylation levels were significantly different (p<0.001) between the cell types, and showed a stepwise decrease from the adjacent normal salivary gland to the intermediate, mucous and squamous cells. The reduced methylation levels of LINE-1 were correlated with a poorer histological grade. In addition, MEC tissue showed a significantly lower level of LINE-1 and Alu element methylation overall compared to normal salivary gland tissue (p<0.001). CONCLUSIONS: Our findings suggest that LINE-1 methylation differed among histological grade mucoepidermoid carcinoma. Hence, this epigenetic event may hold value for MEC diagnosis and prognostic prediction.

Identification of Potential Molecular Mechanisms and Prognostic Markers for Oral Squamous Cell Carcinoma: A Bioinformatics Analysis.

Aims and Objectives: The goal of this study was to uncover crucial biochemical pathways, prognostic indicators, and therapeutic targets in patients with oral cancer in order to enhance therapy strategies. Materials and Methods: Five gene expression omnibus datasets were analyzed by using bioinformatics approaches to identify differentially expressed genes (DEGs). To determine biological alterations, gene ontology (GO) and KEGG pathway analyses were implied using the identified DEGs. Hub genes were determined using protein-protein interaction (PPI) network analysis and an interactome was constructed using NetworkAnalyst. Furthermore, five hub genes were evaluated for use as prognostic markers by using the human protein atlas (HPA) and the GEPIA2.0 database. In addition, the correlations between hub-gene expression and immune cell infiltration of oral squamous cell carcinoma (OSCC) tumors were analyzed using the tWumor immune estimation resource (TIMER) database. Results: A total of 2071 upregulated genes and 1893 downregulated genes were identified. GO and pathway analysis showed DEGs were enriched in multiple immune response terms and interaction of inflammatory cytokines. From the PPI network, five hub genes were identified that have a crucial role in OSCC. These included interferon regulatory factor 4 (IRF4), chemokine receptor 7 (CCR7), TNF receptor superfamily member 17 (TNFRSF17), CD27, and sphingosine-1-phosphate receptor 4 (S1PR4), which were predicted to be favorable prognostic markers for OSCC using HPA. Overall survival analysis revealed that low expression of the five hub genes was significantly associated with worse overall survival. Our analysis of tumor-associated immune infiltration revealed that increased IRF4 expression was positively correlated with the gene expression profiles suggestive of infiltration of all immune cell types, whereas increased CCR7 expression was negatively correlated with neutrophil infiltration. Increased expression of CD27, S1PR4, and TNFRSF17 was found to be negatively correlated with dendritic cell, M0 macrophage, and neutrophil infiltration. Conclusion: In summary, inflammation, and the immune response play an important role in OSCC. All five hub genes were good predictors of OSCC prognosis, suggesting that they could be used as potential therapeutic targets and tumor markers.