Integrative Histologic and Bioinformatics Analysis of BIRC5/Survivin Expression in Oral Squamous Cell Carcinoma.

Survivin is a well-known protein involved in the inhibition of apoptosis in many different cancer types. The aim of this study was to perform an integrated bioinformatic and histologic analysis in order to study the expression and prognostic role of Survivin and its related gene BIRC5 in oral cancer. Publicly available databases were accessed via Gene Expression Omnibus and Oncomine, in addition raw data from The Cancer Genome Atlas (TCGA) were also obtained in order to analyze the rate of gene mutation, expression and methylation in patients with oral squamous cells carcinoma (OSCC). Immunohistochemistry (IHC) was also performed in order to evaluate the nuclear and cytoplasmic expression of Survivin and their correlation with cell proliferation in samples from OSCC patients. Results of this study revealed that Survivin is rarely mutated in OSCC samples and upregulated when compared to non-cancerous tissue. A negative correlation between the methylation of the island cg25986496 and BIRC5 mRNA expression was detected from TCGA data. IHC staining revealed that cytoplasmic (and not nuclear) expression of Survivin is associated with poor overall survival in OSCC patients, while the nuclear expression correlates with higher proliferation rate. In addition, data from TCGA database revealed that BIRC5 gene expression is an independent prognostic factor for OSCC patients.

Differentiating tumor heterogeneity in formalin-fixed paraffin-embedded (FFPE) prostate adenocarcinoma tissues using principal component analysis of matrix-assisted laser desorption/ionization imaging mass spectral data.

RATIONALE: Many patients with adenocarcinoma of the prostate present with advanced and metastatic cancer at the time of diagnosis. There is an urgent need to detect biomarkers that will improve the diagnosis and prognosis of this disease. Matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI-IMS) is playing a key role in cancer research and it can be useful to unravel the molecular profile of prostate cancer biopsies. METHODS: MALDI imaging data sets are highly complex and their interpretation requires the use of multivariate statistical methods. In this study, MALDI-IMS technology, sequential principal component analysis (PCA) and two-dimensional (2-D) peak distribution tests were employed to investigate tumor heterogeneity in formalin-fixed paraffin-embedded (FFPE) prostate cancer biopsies. RESULTS: Multivariate statistics revealed a number of mass ion peaks obtained from different tumor regions that were distinguishable from the adjacent normal regions within a given specimen. These ion peaks have been used to generate ion images and visualize the difference between tumor and normal regions. Mass peaks at m/z 3370, 3441, 3447 and 3707 exhibited stronger ion signals in the tumor regions. CONCLUSIONS: This study reports statistically significant mass ion peaks unique to tumor regions in adenocarcinoma of the prostate and adds to the clinical utility of MALDI-IMS for analysis of FFPE tissue at a molecular level that supersedes all other standard histopathologic techniques for diagnostic purposes used in the current clinical practice. Copyright © 2016 John Wiley & Sons, Ltd.

Orai1 expression pattern in tooth and craniofacial ectodermal tissues and potential functions during ameloblast differentiation.

BACKGROUND: Orai1 is a plasma membrane protein that forms the pore of the calcium release activated calcium channel. Humans with mutated Orai1 present with hereditary combined immunodeficiency, congenital myopathy and anhidrotic ectodermal dysplasia. Consistent with the ectodermal dysplasia phenotype, enamel formation and mineralization is also abnormal in Orai1 deficient patients. The expression pattern and potential functions of Orai1 in enamel formation remains unclear. To contribute toward understanding the role of Orai1 in amelogenesis we characterized ORAI1 protein developmental pattern in comparison with other ectodermal organs. We also examined the effects of Orai1 down-regulation in ameloblast cell proliferation and differentiation. RESULTS: Our data show strong expression of ORAI1 protein during the ameloblast secretory stage, which weans at the end of the maturation stage. In salivary glands, ORAI1 is expressed mainly in acini cells. ORAI1 expression is also found in hair follicle and oral epithelium. Knockdown of Orai1 expression decreases cell proliferation and results in RNA expression levels changes of key ameloblast genes regulating enamel thickness and mineralization. CONCLUSIONS: This study provides insights in the anhidrotic ectodermal dysplasia phenotype due to Orai1 mutation and highlights the importance of calcium signaling in controlling ameloblast differentiation and maturation during tooth development.

The impact of low-dose carcinogens and environmental disruptors on tissue invasion and metastasis.

The purpose of this review is to stimulate new ideas regarding low-dose environmental mixtures and carcinogens and their potential to promote invasion and metastasis. Whereas a number of chapters in this review are devoted to the role of low-dose environmental mixtures and carcinogens in the promotion of invasion and metastasis in specific tumors such as breast and prostate, the overarching theme is the role of low-dose carcinogens in the progression of cancer stem cells. It is becoming clearer that cancer stem cells in a tumor are the ones that assume invasive properties and colonize distant organs. Therefore, low-dose contaminants that trigger epithelial-mesenchymal transition, for example, in these cells are of particular interest in this review. This we hope will lead to the collaboration between scientists who have dedicated their professional life to the study of carcinogens and those whose interests are exclusively in the arena of tissue invasion and metastasis.

The tick tock of odontogenesis.

Although a big deal of dental research is being focused to the understanding of early stages of tooth development, a huge gap exist on our knowledge on how the dental hard tissues are formed and how this process is controlled daily in order to produce very complex and diverse tooth shapes adapted for specific functions. Emerging evidence suggests that clock genes, a family of genes that controls circadian functions within our bodies, regulate also dental mineralized tissues formation. Enamel formation, for example, is subjected to rhythmical molecular signals that occur on short (24h) periods and control the secretion and maturation of the enamel matrix. Accordingly, gene expression and ameloblast functions are also tightly modulated in regular daily intervals. This review summarizes the current knowledge on the circadian controls of dental mineralized tissues development with a special emphasis on amelogenesis.

Oral epithelial stem cells - implications in normal development and cancer metastasis.

Oral mucosa is continuously exposed to environmental forces and has to be constantly renewed. Accordingly, the oral mucosa epithelium contains a large reservoir of epithelial stem cells necessary for tissue homeostasis. Despite considerable scientific advances in stem cell behavior in a number of tissues, fewer studies have been devoted to the stem cells in the oral epithelium. Most of oral mucosa stem cells studies are focused on identifying cancer stem cells (CSC) in oral squamous cell carcinomas (OSCCs) among other head and neck cancers. OSCCs are the most prevalent epithelial tumors of the head and neck region, marked by their aggressiveness and invasiveness. Due to their highly tumorigenic properties, it has been suggested that CSC may be the critical population of cancer cells in the development of OSCC metastasis. This review presents a brief overview of epithelium stem cells with implications in oral health, and the clinical implications of the CSC concept in OSCC metastatic dissemination.

UM-SCC-103: a unique tongue cancer cell line that recapitulates the tumorigenic stem cell population of the primary tumor.

OBJECTIVE: A new head and neck cancer cell line was developed from a highly aggressive HNSCC of the oral cavity diagnosed in a 26-year-old pregnant woman. METHODS: Cells from the primary tumor were passaged in culture and genotyped as a unique cell line. The resultant cell line was assessed for its ability to replicate the primary tumor. RESULTS: The primary tumor and cell line contained 19.03% and 19.62% CD44(high) cells, respectively. CD44(high) cancer stem cells from UM-SCC-103 formed tumors after flank injections in mice that reconstituted the heterogeneity of the primary tumor. CD44 staining and histology in the primary tumor and tumors grown in vivo from the cell line were similar. CD44(high) cells from the primary tumor resulted in lung colony formation in 2 out of 2 tail vein injections in mice, whereas CD44(low) cells did not. Similarly, CD44(high) cells from UM-SCC-103 formed lung tumors in 2 out of 4 mice, whereas CD44(low) cells did not. CONCLUSION: The similarity in marker expression and tumorigenic behavior between the primary tumor and the resulting cell line strongly suggests that the cell line resembles the primary tumor that it was derived from and provides an important new research tool for the study of head and neck carcinomas in young patients.

Equity-diversity-inclusion (EDI)-related strategies used by dental schools during the admission/selection process: a narrative review.

INTRODUCTION: Decades of evidence have demonstrated a lack of workforce diversity and sustaining disparities in academic dentistry and professional practice. Underrepresented minority students may face challenges and implicit bias during the dental schools' admission/selection process. This review collected papers from different countries to summarize the Equity-Diversity-Inclusion (EDI)-related strategies that dental schools worldwide have used in their admissions process to increase diversity. METHODS: A comprehensive search using MEDLINE (via PubMed), ERIC, Cochrane Reviews, Cochrane Trials, American Psychological Association Psyc Info (EBSCO) and Scopus was done between January and March-2023. All types of articles-designs were included, except comments and editorials, and all articles selected were in English. Two independent investigators screened the articles. Extracted data were general characteristics, study objectives, and EDI-related strategies. RESULTS: Sixteen publications were used to construct this manuscript. The year with the greatest number of publications was 2022. Type of studies were case studies/critical reviews (50%), cross-sectional (including survey and secondary data analysis) (n = 5, 31.25%), qualitative methods of analysis (n = 2, 12.5%), and retrospective/secondary data collection (n = 1, 6.25%). The strategies described in the articles were related to (1) considering the intersectionality of diversity, (2) using noncognitive indicators during the school admissions process to construct a holistic selection process, (3) diversifying, professionalizing, and providing training to admissions persons who had leadership roles with the support from the dental school and the university, and (4) allocating financial investments and analyzing current policies and procedures regarding EDI. CONCLUSIONS: This review aggregated interesting findings, such as: some schools are considering the intersectionality of diversity as a way to include underrepresented minorities and to diversify the students-body. The recent growth in publications on EDI during dental admission/selection process might indicate a positive movement in this field.

Colorimetric sensing assay based on aptamer-gold nanoparticles for rapid detection of salivary melatonin to monitor circadian rhythm sleep disorders.

Salivary melatonin is a clinically used biomarker for diagnosing circadian rhythm sleep disorders. Current melatonin detection assays are complex, expensive, and in many cases do not adequately measure low levels of salivary melatonin. Precisely measuring melatonin levels at multiple time points is crucial for determining dim light melatonin onset to evaluate its circadian fluctuation as well as the extent of circadian disruption and consequently adapt treatment regimens. Moreover, melatonin low levels in saliva challenges the reliability of routine clinical testing. This paper presents the development of a novel, highly sensitive, yet cost-effective, colorimetric assay for the rapid detection of salivary melatonin utilizing aptamer-AuNPs. Among several types of the aptamer tested, the 36-mer MLT-A-2 aptamer-AuNP probe showed the highest sensitivity with a melatonin limit of detection of 0.0011 nM along with a limit of quantification of 0.0021 nM in saliva. Moreover, our assay showed preferential interaction with melatonin when tested in presence of other structurally similar counter-targets. Taken together, this study provides new parameters for a melatonin assay that meets adequate levels of sensitivity and selectivity. The developed colorimetric assay could be adapted in a point-of-care system for profiling salivary melatonin levels at multiple time points during 24 h, crucial for accurately diagnosing and monitoring circadian rhythm sleep disorders and beyond.

Structural properties and binding mechanism of DNA aptamers sensing saliva melatonin for diagnosis and monitoring of circadian clock and sleep disorders.

Circadian desynchrony with the external light-dark cycle influences the rhythmic secretion of melatonin which is among the first signs of circadian rhythm sleep disorders. An accurate dim light melatonin onset (established indicator of circadian rhythm sleep disorders) measurement requires lengthy assays, and antibody affinities alterations, especially in patients with circadian rhythm disorders whose melatonin salivary levels vary significantly, making antibodies detection mostly inadequate. In contrast, aptamers with their numerous advantages (e.g., target selectivity, structural flexibility in tuning binding affinities, small size, etc.) can become preferable biorecognition molecules for salivary melatonin detection with high sensitivity and specificity. This study thoroughly characterizes the structural property and binding mechanism of a single-stranded DNA aptamer full sequence (MLT-C-1) and its truncated versions (MLT-A-2, MLT-A-4) to decipher its optimal characteristics for saliva melatonin detection. We use circular dichroism spectroscopy to determine aptamers' conformational changes under different ionic strengths and showed that aptamers display a hairpin loop structure where few base pairs in the stem play a significant role in melatonin binding and formation of aptamer stabilized structure. Through microscale thermophoresis, aptamers demonstrated a high binding affinity in saliva samples (MLT-C-1F Kd = 12.5 ± 1.7 nM; MLT-A-4F Kd = 11.2 ± 1.6 nM; MLT-A-2F Kd = 2.4 ± 2.8 nM; limit-of-detection achieved in pM, highest sensitivity attained for MLT-A-2F aptamer with the lowest detection limit of 1.35 pM). Our data suggest that aptamers are promising as biorecognition molecules and provide the baseline parameters for the development of an aptamer-based point-of-care diagnostic system for melatonin detection and accurate profiling of its fluctuations in saliva.

Deciphering the functions of Stromal Interaction Molecule-1 in amelogenesis using AmelX-iCre mice.

Introduction: The intracellular Ca2+ sensor stromal interaction molecule 1 (STIM1) is thought to play a critical role in enamel development, as its mutations cause Amelogenesis Imperfecta (AI). We recently established an ameloblast-specific (AmelX-iCre) Stim1 conditional deletion mouse model to investigate the role of STIM1 in controlling ameloblast function and differentiation in vivo (Stim1 cKO). Our pilot data (Said et al., J. Dent. Res., 2019, 98, 1002-1010) support our hypothesis for a broad role of Stim1 in amelogenesis. This paper aims to provide an in-depth characterization of the enamel phenotype observed in our Stim1 cKO model. Methods: We crossed AmelX-iCre mice with Stim1-floxed animals to develop ameloblast-specific Stim1 cKO mice. Scanning electron microscopy, energy dispersive spectroscopy, and micro- CT were used to study the enamel phenotype. RNAseq and RT-qPCR were utilized to evaluate changes in the gene expression of several key ameloblast genes. Immunohistochemistry was used to detect the amelogenin, matrix metalloprotease 20 and kallikrein 4 proteins in ameloblasts. Results: Stim1 cKO animals exhibited a hypomineralized AI phenotype, with reduced enamel volume, diminished mineral density, and lower calcium content. The mutant enamel phenotype was more severe in older Stim1 cKO mice compared to younger ones and changes in enamel volume and mineral content were more pronounced in incisors compared to molars. Exploratory RNAseq analysis of incisors' ameloblasts suggested that ablation of Stim1 altered the expression levels of several genes encoding enamel matrix proteins which were confirmed by subsequent RT-qPCR. On the other hand, RT-qPCR analysis of molars' ameloblasts showed non-significant differences in the expression levels of enamel matrix genes between control and Stim1-deficient cells. Moreover, gene expression analysis of incisors' and molars' ameloblasts showed that Stim1 ablation caused changes in the expression levels of several genes associated with calcium transport and mitochondrial kinetics. Conclusions: Collectively, these findings suggest that the loss of Stim1 in ameloblasts may impact enamel mineralization and ameloblast gene expression.

On-demand chlorine dioxide solution enhances odontoblast differentiation through desulfation of cell surface heparan sulfate proteoglycan and subsequent activation of canonical Wnt signaling.

Heparan sulfate proteoglycans (HSPGs) surround the surface of odontoblasts, and their modification affects their affinity for Wnt ligands. This study proposes applying Matching Transformation System® (MA-T), a novel chlorinated oxidant, to enhance dentinogenesis. MA-T treatment in odontoblasts decreased sulfation of HSPG and upregulated the expression of dentin sialophosphoprotein (Dspp) and Dentin Matrix Protein 1 (Dmp1) via activation of canonical Wnt signaling in vitro. Ex vivo application of MA-T also enhanced dentin matrix formation in developing tooth explants. Reanalysis of a public single-cell RNA-seq dataset revealed significant Wnt activity in the odontoblast population, with enrichment for Wnt10a and Wnt6. Silencing assays showed that Wnt10a and Wnt6 were redundant in inducing Dspp and Dmp1 mRNA expression. These Wnt ligands' expression was upregulated by MA-T treatment, and TCF/LEF binding sites are present in their promoters. Furthermore, the Wnt inhibitors Notum and Dkk1 were enriched in odontoblasts, and their expression was also upregulated by MA-T treatment, together suggesting autonomous maintenance of Wnt signaling in odontoblasts. This study provides evidence that MA-T activates dentinogenesis by modifying HSPG and through subsequent activation of Wnt signaling.

Novel trends, challenges and new perspectives for enamel repair and regeneration to treat dental defects.

Dental enamel is the hardest tissue in the human body, providing external protection for the tooth against masticatory forces, temperature changes and chemical stimuli. Once enamel is damaged/altered by genetic defects, dental caries, trauma, and/or dental wear, it cannot repair itself due to the loss of enamel producing cells following the tooth eruption. The current restorative dental materials are unable to replicate physico-mechanical, esthetic features and crystal structures of the native enamel. Thus, development of alternative approaches to repair and regenerate enamel defects is much needed but remains challenging due to the structural and functional complexities involved. This review paper summarizes the clinical aspects to be taken into consideration for the development of optimal therapeutic approaches to tackle dental enamel defects. It also provides a comprehensive overview of the emerging acellular and cellular approaches proposed for enamel remineralization and regeneration. Acellular approaches aim to artificially synthesize or re-mineralize enamel, whereas cell-based strategies aim to mimic the natural process of enamel development given that epithelial cells can be stimulated to produce enamel postnatally during the adult life. The key issues and current challenges are also discussed here, along with new perspectives for future research to advance the field of regenerative dentistry.

Self-Crosslinkable Oxidized Alginate-Carboxymethyl Chitosan Hydrogels as an Injectable Cell Carrier for In Vitro Dental Enamel Regeneration.

Injectable hydrogels, as carriers, offer great potential to incorporate cells or growth factors for dental tissue regeneration. Notably, the development of injectable hydrogels with appropriate structures and properties has been a challenging task, leaving much to be desired in terms of cytocompatibility, antibacterial and self-healing properties, as well as the ability to support dental stem cell functions. This paper presents our study on the development of a novel self-cross-linkable hydrogel composed of oxidized alginate and carboxymethyl chitosan and its characterization as a cell carrier for dental enamel regeneration in vitro. Oxidized alginate was synthesized with 60% theoretical oxidation degree using periodate oxidation and characterized by Fourier Transform Infrared spectroscopy, proton nuclear magnetic resonance spectroscopy, and Ultraviolet-visible absorption spectroscopy. Then, hydrogels were prepared at three varying weight ratios of oxidized alginate to carboxymethyl chitosan (4:1, 3:1, and 2:1) through Schiff base reactions, which was confirmed by Fourier Transform Infrared spectroscopy. The hydrogels were characterized in terms of gelation time, swelling ratio, structure, injectability, self-healing, antibacterial properties, and in vitro characterization for enamel regeneration. The results demonstrated that, among the three hydrogels examined, the one with the highest ratio of oxidized alginate (i.e., 4:1) had the fastest gelation time and the lowest swelling ability, and that all hydrogels were formed with highly porous structures and were able to be injected through a 20-gauge needle without clogging. The injected hydrogels could be rapidly reformed with the self-healing property. The hydrogels also showed antibacterial properties against two cariogenic bacteria: Streptococcus mutans and Streptococcus sobrinus. For in vitro enamel regeneration, a dental epithelial cell line, HAT-7, was examined, demonstrating a high cell viability in the hydrogels during injection. Furthermore, HAT-7 cells encapsulated in the hydrogels showed alkaline phosphatase production and mineral deposition, as well as maintaining their round morphology, after 14 days of in vitro culture. Taken together, this study has provided evidence that the oxidized alginate-carboxymethyl chitosan hydrogels could be used as an injectable cell carrier for dental enamel tissue engineering applications.

The Clinical, Microbiological, and Immunological Effects of Probiotic Supplementation on Prevention and Treatment of Periodontal Diseases: A Systematic Review and Meta-Analysis.

(1) Background: Periodontal diseases are a global health concern. They are multi-stage, progressive inflammatory diseases triggered by the inflammation of the gums in response to periodontopathogens and may lead to the destruction of tooth-supporting structures, tooth loss, and systemic health problems. This systematic review and meta-analysis evaluated the effects of probiotic supplementation on the prevention and treatment of periodontal disease based on the assessment of clinical, microbiological, and immunological outcomes. (2) Methods: This study was registered under PROSPERO (CRD42021249120). Six databases were searched: PubMed, MEDLINE, EMBASE, CINAHL, Web of Science, and Dentistry and Oral Science Source. The meta-analysis assessed the effects of probiotic supplementation on the prevention and treatment of periodontal diseases and reported them using Hedge's g standardized mean difference (SMD). (3) Results: Of the 1883 articles initially identified, 64 randomized clinical trials were included in this study. The results of this meta-analysis indicated statistically significant improvements after probiotic supplementation in the majority of the clinical outcomes in periodontal disease patients, including the plaque index (SMD = 0.557, 95% CI: 0.228, 0.885), gingival index, SMD = 0.920, 95% CI: 0.426, 1.414), probing pocket depth (SMD = 0.578, 95% CI: 0.365, 0.790), clinical attachment level (SMD = 0.413, 95% CI: 0.262, 0.563), bleeding on probing (SMD = 0.841, 95% CI: 0.479, 1.20), gingival crevicular fluid volume (SMD = 0.568, 95% CI: 0.235, 0.902), reduction in the subgingival periodontopathogen count of P. gingivalis (SMD = 0.402, 95% CI: 0.120, 0.685), F. nucleatum (SMD = 0.392, 95% CI: 0.127, 0.658), and T. forsythia (SMD = 0.341, 95% CI: 0.050, 0.633), and immunological markers MMP-8 (SMD = 0.819, 95% CI: 0.417, 1.221) and IL-6 (SMD = 0.361, 95% CI: 0.079, 0.644). (4) Conclusions: The results of this study suggest that probiotic supplementation improves clinical parameters, and reduces the periodontopathogen load and pro-inflammatory markers in periodontal disease patients. However, we were unable to assess the preventive role of probiotic supplementation due to the paucity of studies. Further clinical studies are needed to determine the efficacy of probiotic supplementation in the prevention of periodontal diseases.

Gemini surfactant-based nanoparticles T-box1 gene delivery as a novel approach to promote epithelial stem cells differentiation and dental enamel formation.

Enamel is the highest mineralized tissue in the body protecting teeth from external stimuli, infections, and injuries. Enamel lacks the ability to self-repair due to the absence of enamel-producing cells in the erupted teeth. Here, we reported a novel approach to promote enamel-like tissue formation via the delivery of a key ameloblast inducer, T-box1 gene, into a rat dental epithelial stem cell line, HAT-7, using non-viral gene delivery systems based on cationic lipids. We comparatively assessed the lipoplexes prepared from glycyl-lysine-modified gemini surfactants and commercially available 1,2-dioleoyl-3-trimethylammonium-propane lipids at three nitrogen-to phosphate (N/P) ratios of 2.5, 5 and 10. Our findings revealed that physico-chemical characteristics and biological activities of the gemini surfactant-based lipoplexes with a N/P ratio of 5 provide the most optimal outcomes among those examined. HAT-7 cells were transfected with T-box1 gene using the optimal formulation then cultured in conventional 2D cell culture systems. Ameloblast differentiation, mineralization, bio-enamel interface and structure were assessed at different time points over 28 days. Our results showed that our gemini transfection system provides superior gene expression compared to the benchmark agent, while keeping low cytotoxicity levels. T-box1-transfected HAT-7 cells strongly expressed markers of secretory and maturation stages of the ameloblasts, deposited minerals, and produced enamel-like crystals when compared to control cells. Taken together, our gemini surfactant-based T-box1 gene delivery system is effective to accelerate and guide ameloblastic differentiation of dental epithelial stem cells and promote enamel-like tissue formation. This study would represent a significant advance towards the tissue engineering and regeneration of dental enamel.

Bioprinting of alginate-carboxymethyl chitosan scaffolds for enamel tissue engineeringin vitro.

Tissue engineering offers a great potential in regenerative dentistry and to this end, three dimensional (3D) bioprinting has been emerging nowadays to enable the incorporation of living cells into the biomaterials (such a mixture is referred as a bioink in the literature) to create scaffolds. However, the bioinks available for scaffold bioprinting are limited, particularly for dental tissue engineering, due to the complicated, yet compromised, printability, mechanical and biological properties simultaneously imposed on the bioinks. This paper presents our study on the development of a novel bioink from carboxymethyl chitosan (CMC) and alginate (Alg) for bioprinting scaffolds for enamel tissue regeneration. CMC was used due to its antibacterial ability and superior cell interaction properties, while Alg was added to enhance the printability and mechanical properties as well as to regulate the degradation rate. The bioinks with three mixture ratios of Alg and CMC (2-4, 3-3 and 4-2) were prepared, and then printed into the calcium chloride crosslinker solution (100 mM) to form a 3D structure of scaffolds. The printed scaffolds were characterized in terms of structural, swelling, degradation, and mechanical properties, followed by theirin vitrocharacterization for enamel tissue regeneration. The results showed that the bioinks with higher concentrations of Alg were more viscous and needed higher pressure for printing; while the printed scaffolds were highly porous and showed a high degree of printability and structural integrity. The hydrogels with higher CMC ratios had higher swelling ratios, faster degradation rates, and lower compressive modulus. Dental epithelial cell line, HAT-7, could maintain high viability in the printed constructs after 1, 7 and 14 d of culture. HAT-7 cells were also able to maintain their morphology and secrete alkaline phosphatase after 14 d of culture in the 3D printed scaffolds, suggesting the capacity of these cells for mineral deposition and enamel-like tissue formation. Among all combinations Alg4%-CMC2% and in a less degree 2%Alg-4%CMC showed the higher potential to promote ameloblast differentiation, Ca and P deposition and matrix mineralizationin vitro. Taken together, Alg-CMC has been illustrated to be suitable to print scaffolds with dental epithelial cells for enamel tissue regeneration.

Expression of Beta-Catenin, Cadherins and P-Runx2 in Fibro-Osseous Lesions of the Jaw: Tissue Microarray Study.

Fibrous dysplasia (FD) and hyperparathyroidism-jaw tumor syndrome (HPT-JT) are well-characterized benign bone fibro-osseous lesions. The intracellular mechanism leading to excessive deposition of fibrous tissue and alteration of differentiation processes leading to osteomalacia have not yet been fully clarified. Tissue Microarray (TMA)-based immunohistochemical expression of ß-catenin, CK-AE1/AE3, Ki-67, cadherins and P-Runx2 were analyzed in archival samples from nine patients affected by FD and HPT-JT and in seven controls, with the aim of elucidating the contribution of these molecules (ß-catenin, cadherins and P-Runx2) in the osteoblast differentiation pathway. ß-catenin was strongly upregulated in FD, showing a hyper-cellulated pattern, while it was faintly expressed in bone tumors associated with HPT-JT. Furthermore, the loss of expression of OB-cadherin in osteoblast lineage in FD was accompanied by N-cadherin and P-cadherin upregulation (p < 0.05), while E-cadherin showed a minor role in these pathological processes. P-Runx2 showed over-expression in six out of eight cases of FD and stained moderately positive in the rimming lining osteoblasts in HPT-JT syndrome. ß-catenin plays a central role in fibrous tissue proliferation and accompanies the lack of differentiation of osteoblast precursors in mature osteoblasts in FD. The study showed that the combined evaluation of the histological characteristics and the histochemical and immunohistochemical profile of key molecules involved in osteoblast differentiation are useful in the diagnosis, classification and therapeutic management of fibrous-osseous lesions.

Emerging biotechnologies for evaluating disruption of stress, sleep, and circadian rhythm mechanism using aptamer-based detection of salivary biomarkers.

The internally driven 24-h cycle in humans, called circadian rhythm, controls physiological, metabolic, and hormonal processes, and is tied to the circadian clocks ticking in most of the cells and tissues. The central clock, located in suprachiasmatic nuclei of the hypothalamus, is directly influenced by external cues, particularly light, and entrains the peripheral clocks through neural and hormonal pathways to the external light-dark cycle. However, peripheral clocks also have self-sustained circadian rhythmicity and feeding is the potent synchronizer. The internal clock system regulates the sleep-wake cycle and maintains stress responses through the hypothalamus-pituitary-adrenal axis and autonomic pathways. Any misalignment in this complex network could lead to circadian clock disruption and endocrine and metabolic dysfunction that may induce inflammatory responses. The detrimental consequences of such dysfunction are broad and can lead to serious health problems; however, the extent of the circadian disruption is difficult to assess. New promising techniques based on biosensors and point-of-care devices using aptamers - single-stranded DNA or RNA biorecognition molecules that can measure biomarkers of stress, sleep, and circadian rhythms in bodily fluids such as saliva with high sensitivity and specificity - can provide timely and accurate diagnosis and allow for effective implementation of behavioral and therapeutic interventions. This review provides detailed insight into the complex crosstalk between stress, sleep, and circadian rhythm, their relationship with the body's homeostasis, and the consequences of circadian dysregulation. The review also summarizes the mechanisms of aptamer-based biosensors and/or point-of-care devices developed to date for the detection of salivary biomarkers linked to stress, sleep, and circadian rhythm. Lastly, the review outlines the knowledge gaps in the literature related to the detection of lower concentrations of biomarkers in saliva and discusses the prospects of aptamer-based detection of salivary biomarkers from a high-precision perspective that is crucial for clinical diagnosis, at a time when circadian disruption is evident in unprecedented proportions across the globe.

When the clock ticks wrong with COVID-19.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a member of the coronavirus family that causes the novel coronavirus disease first diagnosed in 2019 (COVID-19). Although many studies have been carried out in recent months to determine why the disease clinical presentations and outcomes can vary significantly from asymptomatic to severe or lethal, the underlying mechanisms are not fully understood. It is likely that unique individual characteristics can strongly influence the broad disease variability; thus, tailored diagnostic and therapeutic approaches are needed to improve clinical outcomes. The circadian clock is a critical regulatory mechanism orchestrating major physiological and pathological processes. It is generally accepted that more than half of the cell-specific genes in any given organ are under circadian control. Although it is known that a specific role of the circadian clock is to coordinate the immune system's steady-state function and response to infectious threats, the links between the circadian clock and SARS-CoV-2 infection are only now emerging. How inter-individual variability of the circadian profile and its dysregulation may play a role in the differences noted in the COVID-19-related disease presentations, and outcome remains largely underinvestigated. This review summarizes the current evidence on the potential links between circadian clock dysregulation and SARS-CoV-2 infection susceptibility, disease presentation and progression, and clinical outcomes. Further research in this area may contribute towards novel circadian-centred prognostic, diagnostic and therapeutic approaches for COVID-19 in the era of precision health.