Clock gene Per2 modulates epidermal tissue repair in vivo.

Wound healing can be influenced by genes that control the circadian cycle, including Per2 and BMAL1, which coordinate the functions of several organs, including the skin. The aim of the study was to evaluate the role of PER2 during experimental skin wound healing. Two groups (control and Per2-KO), consisting of 14 male mice each, were anesthetized by inhalation, and two 6 mm wounds were created on their dorsal skin using a punch biopsy. A silicone ring was sutured around the wound perimeter to restrict contraction. The wound healing process was clinically measured daily (closure index) until complete wound repair. On Day 6, histomorphometric analysis was performed using the length and thickness of the epithelial migration tongue, in addition to counting vessels underlying the lesion by immunofluorescence assay and maturation of collagen fibers through picrosirius staining. Bromodeoxyuridine (BrdU) incorporation and quantification were performed using the subcutaneous injection technique 2 h before euthanasia and through immunohistochemical analysis of the proliferative index. In addition, the qualitative analysis of myofibroblasts and periostin distribution in connective tissue was performed by immunofluorescence. Statistically significant differences were observed in the healing time between the experimental groups (means: 15.5 days for control mice and 13.5 days for Per2-KO; p = 0.001). The accelerated healing observed in the Per2-KO group (p < 0.05) was accompanied by statistical differences in wound diameter and length of the migrating epithelial tongue (p = 0.01) compared to the control group. Regarding BrdU immunoreactivity, higher expression was observed in the intact epithelium of Per2-KO animals (p = 0.01), and this difference compared to control was also present, to a lesser extent, at the wound site (p = 0.03). Immunofluorescence in the connective tissue underlying the wound showed a higher angiogenic potential in the Per2-KO group in the intact tissue area and the wound region (p < 0.01), where increased expression of myofibroblasts was also observed. Qualitative analysis revealed the distribution of periostin protein and collagen fibers in the connective tissue underlying the wound, with greater organization and maturation during the analyzed period. Our research showed that the absence of the Per2 gene positively impacts the healing time of the skin in vivo. This acceleration depends on the increase of epithelial proliferative and angiogenic capacity of cells carrying the Per2 deletion.

Up-regulation of TNF-alpha/NFkB/SIRT1 axis drives aggressiveness and cancer stem cells accumulation in chemoresistant oral squamous cell carcinoma.

Tumor resistance remains an obstacle to successfully treating oral squamous cell carcinoma (OSCC). Cisplatin is widely used as a cytotoxic drug to treat solid tumors, including advanced OSCC, but with low efficacy due to chemoresistance. Therefore, identifying the pathways that contribute to chemoresistance may show new possibilities for improving the treatment. This work explored the role of the tumor necrosis factor-alpha (TNF-alpha)/NFkB signaling in driving the cisplatin resistance of OSCC and its potential as a pharmacological target to overcome chemoresistance. Differential accessibility analysis demonstrated the enrichment of opened chromatin regions in members of the TNF-alpha/NFkB signaling pathway, and RNA-Seq confirmed the upregulation of TNF-alpha/NFkB signaling in cisplatin-resistant cell lines. NFkB was accumulated in cisplatin-resistant cell lines and in cancer stem cells (CSC), and the administration of TNF-alpha increased the CSC, suggesting that TNF-alpha/NFkB signaling is involved in the accumulation of CSC. TNF-alpha stimulation also increased the histone deacetylases HDAC1 and SIRT1. Cisplatin-resistant cell lines were sensitive to the pharmacological inhibition of NFkB, and low doses of the NFkB inhibitors, CBL0137, and emetine, efficiently reduced the CSC and the levels of SIRT1, increasing histone acetylation. The NFkB inhibitors decreased stemness potential, clonogenicity, migration, and invasion of cisplatin-resistant cell lines. The administration of the emetine significantly reduced the tumor growth of cisplatin-resistant xenograft models, decreasing NFkB and SIRT1, increasing histone acetylation, and decreasing CSC. TNF-alpha/NFkB/SIRT1 signaling regulates the epigenetic machinery by modulating histone acetylation, CSC, and aggressiveness of cisplatin-resistant OSCC and the NFkB inhibition is a potential strategy to treat chemoresistant OSCC.

Repurposing NFκB and HDAC inhibitors to individually target cancer stem cells and non-cancer stem cells from mucoepidermoid carcinomas.

Drug resistance remains a major obstacle in the treatment of mucoepidermoid carcinomas (MEC) leading to tumor recurrence, disease progression, and metastasis. Emerging evidence suggests that drug resistance is mediated by the presence of a highly adaptative subpopulation of cancer cells known as cancer stem cells (CSC). We have previously reported that solid tumors use NFkB signaling as a chemotherapy-resistant mechanism. We have also shown that interfering with the epigenome of solid tumors is an effective strategy to control the population of CSC. Here, we sought to investigate the effects of the NFkB inhibitor emetine and the HDAC inhibitor SAHA on the biology of MEC CSC and assessed whether this combination therapy would favor the standard of care therapy comprised of the administration of Cisplatin (CDDP). Our findings suggested that the administration of low concentrations of emetine and SAHA is more effective in disrupting CSC in MEC, while the administration of emetine in combination with CDDP constitutes an effective therapy to target non-CSC MEC tumor cells.

Proposal of a secure and efficient protocol for a murine oral carcinogenesis model induced by 4-nitroquinoline-1-oxide (4NQO).

An important rat model using the chemical carcinogen 4-nitroquinoline-1-oxide (4NQO) has been described for the study of the process of oral carcinogenesis. This model replicates the gradual progression seen in oral carcinoma patients. However, due to its high level of toxicity, its use in fundamental research is challenging. Here, we propose a secure and efficient modified protocol based on a lower dose of 4NQO concentration as well as an increased water supply and hypercaloric diet, in order to reduce the damage caused to the animals during the process of oral carcinogenesis. Twenty-two male Wistar rats were exposed to 4NQO, evaluated clinically once a week and euthanized at 12 and 20 weeks for histopathological analysis. The protocol involves a staggered dose of 4NQO up to a concentration of 25 ppm, associated with two days of pure water, a 5% glucose solution once a week and a hypercaloric diet. This modified protocol prevents the immediate consequences of the carcinogen. At week 7, all animals displayed clinically evident tongue lesions. From a histological perspective, after 12 weeks of 4NQO exposure, 72.7% of the animals developed epithelial dysplasia and 27.3% developed in situ carcinoma. In the group exposed for 20 weeks, epithelial dysplasia and in situ carcinoma were diagnosed in one case each, whereas invasive carcinoma was diagnosed in 81.8% of the cases. Nonsignificant modification of animal's behavior and weight was observed. This new proposed 4NQO protocol was secure and effective for studying oral carcinogenesis and can be used to conduct lengthy investigations.

Inflammatory Cells Can Alter the Levels of H3K9ac and γH2AX in Dysplastic Cells and Favor Tumor Phenotype.

Oral potentially malignant disorders (OPMD) are clinical presentations that carry an increased risk of cancer development. Currently, epithelial dysplasia grade is based on architectural and cytological epithelial changes and is used to predict the malignant transformation of these lesions. However, predicting which OPMD will progress to a malignant tumor is very challenging. Inflammatory infiltrates can favor cancer development, and recent studies suggest that this association with OPMD lesions may be related to the etiology and/or aggressive clinical behavior of these lesions. Epigenetic changes such as histone modifications may mediate chronic inflammation and also favor tumor cells in immune resistance and evasion. This study aimed to evaluate the relationship between histone acetylation (H3K9ac) and DNA damage in the context of dysplastic lesions with prominent chronic inflammation. Immunofluorescence of "low-risk" and "high-risk" OPMD lesions (n = 24) and inflammatory fibrous hyperplasia (n = 10) as the control group was performed to assess histone acetylation levels and DNA damage through the phosphorylation of H2AX (γH2AX). Cell co-culture assays with PBMCs and oral keratinocyte cell lines (NOK-SI, DOK, and SCC-25) were performed to assess proliferation, adhesion, migration, and epithelial-mesenchymal transition (EMT). Oral dysplastic lesions showed a hypoacetylation of H3K9 and low levels of γH2AX compared to control. The contact of dysplastic oral keratinocytes with PBMCs favored EMT and the loss of cell-cell adhesion. On the other hand, p27 levels increased and cyclin E decreased in DOK, indicating cell cycle arrest. We conclude that the presence of chronic inflammation associated to dysplastic lesions is capable of promoting epigenetic alterations, which in turn can favor the process of malignant transformation.

Effect of ZEB1 Associated with microRNAs on Tumor Stem Cells in Head and Neck Cancer.

Cancer biologists have focused on studying cancer stem cells (CSCs) because of their ability to self-renew and recapitulate tumor heterogeneity, which increases their resistance to chemotherapy and is associated with cancer relapse. Here, we used two approaches to isolate CSCs: the first involved the metabolic enzyme aldehyde dehydrogenase ALDH, and the second involved the three cell surface markers CD44, CD117, and CD133. ALDH cells showed a higher zinc finger E-box binding homeobox 1 (ZEB1) microRNA (miRNA) expression than CD44/CD117/133 triple-positive cells, which overexpressed miRNA 200c-3p: a well-known microRNA ZEB1 inhibitor. We found that ZEB1 inhibition was driven by miR-101-3p, miR-139-5p, miR-144-3p, miR-199b-5p, and miR-200c-3p and that the FaDu Cell Line inhibition occurred at the mRNA level, whereas HN13 did not affect mRNA expression but decreased protein levels. Furthermore, we demonstrated the ability of the ZEB1 inhibitor miRNAs to modulate CSC-related genes, such as TrkB, ALDH, NANOG, and HIF1A, using transfection technology. We showed that ALDH was upregulated upon ZEB1-suppressed miRNA transfection (Mann-Whitney ** p101 = 0.009, t-test ** p139 = 0.009, t-test ** p144 = 0.002, and t-test *** p199 = 0.0006). Overall, our study enabled an improved understanding of the role of ZEB1-suppressed miRNAs in CSC biology.

Antisclerostin Effect on Osseointegration and Bone Remodeling.

Objective: This study reviewed the literature on local or systemic administration of antisclerostin, presenting results associated with osseointegration of dental/orthopedic implants and stimulation of bone remodeling. Materials and Methods: An extensive electronic search was conducted through MED-LINE/PubMed, PubMed Central, Web of Science databases and specific peer-reviewed journals to identify case reports, case series, randomized controlled trials, clinical trials and animal studies comparing either the systemic or local administration of antisclerostin and its effect in osseointegration and bone remodeling. Articles in English and with no restriction on period were included. Results: Twenty articles were selected for a full-text, and one was excluded. Finally, 19 articles were included in the study (16 animal studies and 3 randomized control trials). These studies were divided into two groups, which evaluated (i) osseointegration and (ii) bone remodeling potential. Initially 4560 humans and 1191 animals were identified. At least 1017 were excluded from the studies (981 humans and 36 animals), totaling 4724 subjects who completed (3579 humans and 1145 animals). (a) Osseointegration: 7 studies described this phenomenon; 4 reported bone-implant contact, which increased in all included studies. Similar results were found for bone mineral density, bone area/volume and bone thickness. (b) Bone remodeling: 13 studies were used for description. The studies reported an increase in BMD with sclerostin antibody treatment. A similar effect was found for bone mineral density/area/volume, trabecular bone and bone formation. Three biomarkers of bone formation were identified: bone-specific alkaline phosphatase (BSAP), osteocalcin and procollagen type 1 N-terminal Pro-peptide (P1NP); and markers for bone resorption were: serum C-telopeptide (sCTX), C-terminal telopeptides of type I collagen (CTX-1), ß-isomer of C-terminal telopeptides of type I collagen (ß-CTX) and tartrate-resistant acid phosphatase 5b (TRACP-5b). There were limitations: low number of human studies identified; high divergence in the model used (animal or human); the variance in the type of Scl-Ab and doses of administration; and the lack of reference quantitative values in the parameters analyzed by authors' studies (many articles only reported qualitative information). Conclusion: Within the limitations of this review and carefully observing all data, due to the number of articles included and the heterogeneity existing, more studies must be carried out to better evaluate the action of the antisclerostin on the osseointegration of dental implants. Otherwise, these findings can accelerate and stimulate bone remodeling and neoformation.

High density of CD8 T cell and immune imbalance of T lymphocytes subsets are associated with proliferative verrucous leukoplakia.

Oral leukoplakia (OL) and proliferative verrucous leukoplakia (PVL) are oral potentially malignant disorders (OPMDs) that microscopically show no or varying degrees of dysplasia. Even sharing clinical and microscopic aspects, PVL shows a more aggressive clinical behaviour, with a malignant transformation rate greater than 40%. Inflammatory infiltrate associated with dysplastic lesions may favour malignant transformation of OPMDs. This study aimed to evaluate the density of T cells and cytokines in dysplastic lesions from OL and PVL patients. Additionally, we evaluated whether soluble products produced in vitro by dysplastic keratinocytes are capable of modulating apoptosis rates and Th phenotype (Th1, Th2, Th17 and Treg) of peripheral blood mononuclear cells. The density of CD3, CD4 and CD8 T cells was assessed by immunohistochemistry. Cytokines and chemokines profile from frozen tissue samples were analysed using the LUMINEX system. Apoptosis rates and Th phenotype modulation were evaluated by flow cytometry. Our results showed an increase in the number of CD8 T cell in the subepithelial region from PVL dysplastic lesions in relation to OL samples. PVL showed increased levels of IL-5 and a decrease in IL-1ß and IFN-γ levels compared to OL. Soluble products of PVL and oral carcinoma cell cultures were able to reduce apoptosis rate and promote an imbalance of Th1/Th2 and Th17/Treg. The high-subepithelial density of CD8 T cells and immune imbalance of T lymphocytes subsets probably play an important role in the pathogenesis of PVL and may explain its more aggressive behaviour in relation to OL.

Oral squamous cell carcinoma cancer stem cells have different drug sensitive to pharmacological NFκB and histone deacetylation inhibition.

Despite many progresses in the development of new systemic therapies for oral squamous cell carcinoma (OSCC), the five-year survival rate of OSCC is low. The traditional chemotherapies approach (cisplatin - CDDP) shows some limitations like drug toxicity, limited efficacy, and drug resistance. Promising studies suggested OSCC cancer stem cells (CSC) presented resistance to CDDP. We have previously studied many targets, and we extensively showed the efficacy of the NFκB signaling and the role of histones acetylation, on different malignant tumors, including adenoid cystic carcinoma and mucoepidermoid carcinoma, but until then the effects of the NFkB inhibitor and histone deacetylase (HDAC) inhibitor on the biology of OSCC were not evaluated. Here we assessed the pharmacological inhibitor of NFκB emetine and HDAC inhibitor SAHA on the behavior of CSC derived from OSCC. Our data suggested that CDDP administration resulted in reduced viability of bulk OSCC cells and increased CSC. A single and isolated shot of emetine and SAHA were able to disrupt CSC by inhibiting the NFκB pathway and increasing the histone acetylation levels, respectively. Further, the combined administration of emetine and SAHA presented the same CSC disruption as seen in emetine alone.

Clinical Performance Comparing Titanium and Titanium-Zirconium or Zirconia Dental Implants: A Systematic Review of Randomized Controlled Trials.

OBJECTIVES: This study aimed to compare clinical results between titanium (Ti), zirconia (Zr), or titanium-zirconium (TZ) dental implants and to analyze survival rate (SR), bleeding on probing (BoP), marginal bone loss (MBL), and/or probing depth (PD). DATA SOURCE: Manual and electronic searches were conducted (PubMed and Web of Science) to identify randomized controlled trials that compared the outcomes of at least two implant types (control and test group) within the same study. The focused question was determined according to the PICOT strategy. Seven studies were included out of 202 research studies initially found. The follow-up periods ranged from 12 to 80 months, and the mean age was from 43.3 to 65.8 years old. The SR for Ti, TZ, and Zr implants ranged from 92.6% to 100%, 95.8% to 100%, and 87.5% to 91.25%, respectively; MBL for Ti, TZ, and Zr implants varied from -1.17 mm to -0.125 mm for Ti, -0.6 mm to -0.32 mm for TZ, and -0.25 mm to -1.38 mm for Zr. Studies showed a low incidence of mucositis and peri-implantitis; however, BoP for Zr was 16.43%, Ti ranged between 10% and 20%, and TZ from 10% to 13.8%. PD for Ti ranged from 1.6 mm to 3.05 mm, TZ was 3.12 mm (only one study), and Zr ranged from 2.21 mm to 2.6 mm. CONCLUSION: All three types of implants showed similar tissue behavior. However, the TZ group had better results when compared with Ti and Zr for SR, MBL, and BoP, except for PD. Furthermore, the worst SR was found in the Zr implants group.

Distinct global DNA methylation and NF-κB expression profile of preimplantation biopsies from ideal and non-ideal kidneys.

BACKGROUND: Epigenetic mechanisms may affect the ideal and non-ideal kidneys selected for transplantation and their inflammatory gene expression profile differently and may contribute to poor clinical outcomes. OBJECTIVE: Study the Global DNA methylation and the expression profiles of the DNA methyltransferases (DNMTs) and nuclear factor kappa B (NF-κB) in preimplantation kidney biopsies from ideal and non-ideal kidneys (expanded criteria donor (ECD) and with KDPI > 85%). METHODS: In a sample consisting of 45 consecutive pre-implantation biopsies, global DNA methylation levels were detected by LINE-1 repeated elements using bisulfite pyrosequencing. DNMT gene expression was assessed by real-time quantitative polymerase chain reaction, and NF-κB protein expression by immunofluorescence. RESULTS: ECD kidneys displayed increased methylation levels in LINE-1, and DNMT1 and DNMT3B expression was upregulated when comparing ECD to standard criteria donor kidneys. Similarly, kidneys with KDPI > 85% exhibited increased LINE-1 methylation and DNMT1 upregulation when compared to a KDPI ≤ 85%. NF-κB protein expression levels were greatly increased in both types of non-ideal kidneys compared to ideal kidneys. Moreover, hypermethylation of LINE-1 was associated with cold ischemia time > 20 h and ECD kidney classification. CONCLUSIONS: This study shows that global DNA hypermethylation and high expression of NF-κB occurred in both types of non-ideal kidneys and were associated with prolonged cold ischemia time. Global DNA methylation can be a useful tool to assess non-ideal kidneys and hence, could be used to expand the pool of kidneys donors.

Tensile Strength Essay Comparing Three Different Platelet-Rich Fibrin Membranes (L-PRF, A-PRF, and A-PRF+): A Mechanical and Structural In Vitro Evaluation.

Predictable outcomes intended by the application of PRF (platelet-rich fibrin) derivative membranes have created a lack of consideration for their consistency and functional integrity. This study aimed to compare the mechanical properties through tensile strength and analyze the structural organization among the membranes produced by L-PRF (leukocyte platelet-rich fibrin), A-PRF (advanced platelet-rich fibrin), and A-PRF+ (advanced platelet-rich fibrin plus) (original protocols) that varied in centrifugation speed and time. L-PRF (n = 12), A-PRF (n = 19), and A-PRF+ (n = 13) membranes were submitted to a traction test, evaluating the maximum and average traction. For maximum traction, 0.0020, 0.0022, and 0.0010 N·mm−2 were obtained for A-PRF, A-PRF+, and L-PRF, respectively; regarding the average resistance to traction, 0.0012, 0.0015, and 0.006 N·mm−2 were obtained, respectively (A-PRF+ > A-PRF > L-PRF). For all groups studied, significant results were found. In the surface morphology observations through SEM, the L-PRF matrix showed a highly compact surface with thick fibers present within interfibrous areas with the apparent destruction of red blood cells and leukocytes. The A-PRF protocol showed a dense matrix composed of thin and elongated fibers that seemed to follow a preferential and orientated direction in which the platelets were well-adhered. Porosity was also evident with a large diameter of the interfibrous spaces whereas A-PRF+ was the most porous platelet concentrate with the greatest fiber abundance and cell preservation. Thus, this study concluded that A-PRF+ produced membranes with significant and higher maximum traction results, indicating a better viscoelastic strength when stretched by two opposing forces.

MutSα expression predicts a lower disease-free survival in malignant salivary gland tumors: an immunohistochemical study

Ten cases of normal salivary gland (NSG) and 92 of SGT (54 benign and 38 malignant) were retrieved. Immunohistochemistry was performed for hMSH2, hMSH3, hMSH6. Scanned slides were digitally analyzed based on the percentage of positive cells with nuclear staining. Cases were further classified in MutSαhigh and MutSβhigh based on hMSH2-hMSH6 and hMSH3-hMSH6 expression, respectively. Results: hMSH3 expression was lower in malignant SGT compared to NSG and benign cases. Adenoid cystic carcinoma (ACC) cases with perineural invasion presented a lower percentage of hMSH3 positive cells. hMSH6 was downregulated in both benign and malignant SGT compared to NSG. Malignant SGT cases with MutSαhigh expression had lower disease-free survival compared to MutSαlow cases. A 10.26-fold increased risk of presenting local recurrence was observed. Conclusions: Our findings suggest that a lack of hMSH3 protein function is associated with a more aggressive phenotype (malignancy and perineural invasion) and that MutSα overexpression predicts a poor clinical outcome in malignant SGT.(AU)

Expression profile of the PI3K-AKT-mTOR pathway in head and neck squamous cell carcinoma: Data from Brazilian population.

OBJECTIVE: The phosphatidylinositol 3-kinase/protein kinase-B/mammalian target of the rapamycin (PI3K-AKT-mTOR) signaling pathway is an important regulator of cell proliferation, survival, and motility. The gain or loss of function of proteins related to this pathway results in the neoplastic transformation in several types of cancers. This study aimed to evaluate the expression profile of the PI3K-AKT-mTOR pathway in patients with head and neck squamous cell carcinoma (HNSCC) and HNSCC cell lines. STUDY DESIGN: The study involved 26 formalin-fixed paraffin-embedded tissue samples from patients with HNSCC. The analysis of immunohistochemical expression of PI3K, AKT, p-mTOR, and phosphatase and tensin homolog (PTEN) proteins was performed by a quantitative assessment. The in vitro gene and protein expression evaluation was performed by real-time polymerase chain reaction and Western blot assay, respectively, in the human cell lines SCC-9 and FaDu. RESULTS: High levels of PI3K, AKT, and p-mTOR were found in most HNSCC tumors. Following this result, we observed low amounts or absence of PTEN in most samples. Additionally, the FaDu cells (pharynx) showed higher AKT expression but lower expression of p-mTOR compared with SCC-9 cells (oral cavity), which hints at a loco-anatomical relevance. CONCLUSION: Overall, this study found increased expression of the PI3K-AKT-mTOR pathway along with evident PTEN reduction in head and neck cancer.

Cephaeline is an inductor of histone H3 acetylation and inhibitor of mucoepidermoid carcinoma cancer stem cells.

AIM: To evaluate the potential use of Cephaeline as a therapeutic strategy to manage mucoepidermoid carcinomas (MEC) of the salivary glands. MATERIAL AND METHODS: UM-HMC-1, UM-HMC-2, and UM-HMC-3A MEC cell lines were used to establish the effects of Cephaeline over tumor viability determined by MTT assay. In vitro wound healing scratch assays were performed to address cellular migration while immunofluorescence staining for histone H3 lysine 9 (H3k9ac) was used to identify the acetylation status of tumor cells upon Cephaeline administration. The presence of cancer stem cells was evaluated by the identification of ALDH enzymatic activity by flow cytometry and through functional assays using in vitro tumorsphere formation. RESULTS: A single administration of Cephaeline resulted in reduced viability of MEC cells along with the halt on tumor growth and cellular migration potential. Administration of Cephaeline resulted in chromatin histone acetylation as judged by the increased levels of H3K9ac and disruption of tumorspheres formation. Interestingly, ALDH levels were increased in UM-HMC-1 and UM-HMC-3A cell lines, while UM-HMC-2 showed a reduced enzymatic activity. CONCLUSION: Cephaeline has shown anti-cancer properties in all MEC cell lines tested by regulating tumor cells' viability, migration, proliferation, and disrupting the ability of cancer cells to generate tumorspheres.

Understanding the role of endotoxin tolerance in chronic inflammatory conditions and periodontal disease.

OBJECTIVE: This review aims to present the current understanding of endotoxin tolerance (ET) in chronic inflammatory diseases and explores the potential connection with periodontitis. SUMMARY: Subsequent exposure to lipopolysaccharides (LPS) triggers ET, a phenomenon regulated by different mechanisms and pathways, including toll-like receptors (TLRs), nuclear factor kappa-light-chain enhancer of activated B-cells (NFκB), apoptosis of immune cells, epigenetics, and microRNAs (miRNAs). These mechanisms interconnect ET with chronic inflammatory diseases including periodontitis. While the direct correlation between ET and periodontal destruction has not been fully elucidated, emerging reports point towards the potential tolerization of human periodontal ligament cells (hPDLCs) and gingival tissues with a significant reduction of TLR levels. CONCLUSIONS: There is a potential link between ET and periodontal diseases. Future studies should explore the crucial role of ET in the pathogenesis of periodontal diseases, as evidence of a tolerized oral mucosa may represent an intrinsic mechanism capable of regulating the oral immune response. A clear understanding of this host immune regulatory mechanism might lead to effective and more predictable therapeutic strategies to treat chronic inflammatory diseases and periodontitis.

New tendencies in non-surgical periodontal therapy.

The aim of this review was to update the evidence of new approaches to non-surgical therapy (NSPT) in the treatment of periodontitis. Preclinical and clinical studies addressing the benefits of adjunctive antimicrobial photodynamic therapy, probiotics, prebiotics/synbiotics, statins, pro-resolving mediators, omega-6 and -3, ozone, and epigenetic therapy were scrutinized and discussed. Currently, the outcomes of these nine new approaches, when compared with subgingival debridement alone, did not demonstrate a significant added clinical benefit. However, some of these new alternative interventions may have the potential to improve the outcomes of NSPT alone. Future evidence based on randomized controlled clinical trials would help clinicians and patients in the selection of different adjunctive therapies.

Epigenetic modulation of the tumor microenvironment in head and neck cancer: Challenges and opportunities.

Head and neck cancer is globally challenging due to the resistance to therapy and aggressive behavior leading to high rates of mortality. Recent findings show that the tumor microenvironment plays a role in the maintenance and progression of many solid tumors, including head and neck cancer. The mechanisms involved in the modulation and regulation of the tumor microenvironment remain poorly understood. Increasing evidence suggests that epigenetic events can modulate the crosstalk between neoplastic and non-neoplastic cells during tumor progression. In this review, we explore the current understanding of the involvement of epigenetic events in the modulation of the tumor microenvironment and its impact on head and neck cancer behavior. We also explore the latest therapeutic strategies that use epigenetic-modulating drugs to manage tumor growth and progression.

Worldwide prevalence of PI3K-AKT-mTOR pathway mutations in head and neck cancer: A systematic review and meta-analysis.

A systematic review (SR) and meta-analysis were conducted to determine the prevalence of PI3K-AKT-mTOR signaling pathway mutations in patients with head and neck cancer (HNC). Overall, 105 studies comprising 8630 patients and 1306 mutations were selected. The estimated mutations prevalence was 13 % for PIK3CA (95 % confidence interval [CI] = 11-14; I2 = 82 %; p < 0.0001), 4% for PTEN (95 % CI = 3-5; I2 = 55 %; p < 0.0001), 3% for MTOR (95 % CI = 2-4; I2 = 5%; p = 0.40), and 2% for AKT (95 % CI = 1-2; I2 = 50 %; p = 0.0001). We further stratified the available data of the participants according to risk factors and tumor characteristics, including HPV infection, tobacco use, alcohol exposure, TNM stage, and histological tumor differentiation, and performed subgroup analysis. We identified significant associations between PI3K-AKT-mTOR pathway-associated mutations and advanced TNM stage (odds ratio [OR] = 0.20; 95 % CI = 0.09-0.44; I² = 71 %; p = 0.0001) and oropharyngeal HPV-positive tumors and PIK3CA mutations (OR = 17.48; 95 % CI = 4.20-72.76; I² = 69 %; p < 0.0002). No associations were found between alcohol and tobacco exposure, and tumor differentiation grade. This SR demonstrated that the PI3K-AKT-mTOR pathway emerges as a potential prognostic factor and could offer a molecular basis for future studies on therapeutic targeting in HNC patients.

miR-22 and miR-205 Drive Tumor Aggressiveness of Mucoepidermoid Carcinomas of Salivary Glands.

OBJECTIVES: To integrate mRNA and miRNA expression profiles of mucoepidermoid carcinomas (MECs) and normal salivary gland (NSGs) tissue samples and identify potential drivers. MATERIAL AND METHODS: Gene and miRNA expression arrays were performed in 35 MECs and six NSGs. RESULTS: We found 46 differentially expressed (DE) miRNAs and 3,162 DE mRNAs. Supervised hierarchical clustering analysis of the DE transcripts revealed two clusters in both miRNA and mRNA profiles, which distinguished MEC from NSG samples. The integrative miRNA-mRNA analysis revealed a network comprising 696 negatively correlated interactions (44 miRNAs and 444 mRNAs) involving cell signaling, cell cycle, and cancer-related pathways. Increased expression levels of miR-205-5p and miR-224-5p and decreased expression levels of miR-139-3p, miR-145-3p, miR-148a-3p, miR-186-5p, miR-338-3p, miR-363-3p, and miR-4324 were significantly related to worse overall survival in MEC patients. Two overexpressed miRNAs in MEC (miR-22 and miR-205) were selected for inhibition by the CRISPR-Cas9 method. Cell viability, migration, and invasion assays were performed using an intermediate grade MEC cell line. Knockout of miR-205 reduced cell viability and enhanced ZEB2 expression, while miR-22 knockout reduced cell migration and invasion and enhanced ESR1 expression. Our results indicate a distinct transcriptomic profile of MEC compared to NSG, and the integrative analysis highlighted miRNA-mRNA interactions involving cancer-related pathways, including PTEN and PI3K/AKT. CONCLUSION: The in vitro functional studies revealed that miR-22 and miR-205 deficiencies reduced the viability, migration, and invasion of the MEC cells suggesting they are potential oncogenic drivers in MEC.