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OBJECTIVES: In order to ensure improved and accelerated bone regeneration, nano-hydroxyapatite scaffolds are often enriched with different bioactive components to further accelerate and improve bone healing. In this review, we critically examined whether the enrichment of nHAp/polymer scaffolds with growth factors, hormones, polypeptides, microRNAs and exosomes improved new bone formation in vivo. MATERIALS AND METHODS: Out of 2989 articles obtained from the literature search, 106 papers were read in full, and only 12 articles met the inclusion criteria for this review. RESULTS: Several bioactive components were reported to stimulate accelerated bone regeneration in a variety of bone defect models, showing better results than bone grafting with nHAp scaffolds alone. CONCLUSIONS: The results indicated that composite materials based on nHAp are excellent candidates as bone substitutes, while nHAp scaffold enrichment further accelerates bone regeneration. The standardization of animal models should be provided in order to clearly define the most significant parameters of in vivo studies. Only in this way can the adequate comparison of findings from different in vivo studies be possible, further advancing our knowledge on bone regeneration and enabling its translation to clinical settings.
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This study aimed to compare the biological properties of newly synthesized cements based on calcium phosphate with a commercially used cement, mineral trioxide aggregate (MTA). Strontium (Sr)-, Copper (Cu)-, and Zinc (Zn)-doped hydroxyapatite (miHAp) powder was obtained through hydrothermal synthesis and characterized by scanning electron microscopy (SEM), X-ray diffraction (XRD), and energy dispersive X-ray spectrometry (EDX). Calcium phosphate cement (CPC) was produced by mixing miHAp powder with a 20 wt.% citric acid solution, followed by the assessment of its compressive strength, setting time, and in vitro bioactivity. Acetylsalicylic acid (ASA) was added to the CPC, resulting in CPCA. Biological tests were conducted on CPC, CPCA, and MTA. The biocompatibility of the cement extracts was evaluated in vitro using human dental pulp stem cells (hDPSCs) and in vivo using a zebrafish model. Antibiofilm and antimicrobial effect (quantified by CFUs/mL) were assessed against Streptococcus mutans and Lactobacillus rhamnosus. None of the tested materials showed toxicity, while CPCA even increased hDPSCs proliferation. CPCA showed a better safety profile than MTA and CPC, and no toxic or immunomodulatory effects on the zebrafish model. CPCA exhibited similar antibiofilm effects against S. mutans and L. rhamnosus to MTA.
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Aspirina , Fosfatos de Calcio , Cobre , Estroncio , Zinc , Estroncio/química , Estroncio/farmacología , Fosfatos de Calcio/química , Fosfatos de Calcio/farmacología , Humanos , Animales , Aspirina/farmacología , Aspirina/química , Cobre/química , Zinc/química , Zinc/farmacología , Cementos Dentales/química , Cementos Dentales/farmacología , Biopelículas/efectos de los fármacos , Ensayo de Materiales , Pez Cebra , Pulpa Dental/citología , Pulpa Dental/efectos de los fármacos , Streptococcus mutans/efectos de los fármacos , Células Madre/efectos de los fármacos , Difracción de Rayos X , Materiales Biocompatibles/química , Materiales Biocompatibles/farmacología , Proliferación Celular/efectos de los fármacosRESUMEN
Despite advancements in treatment, the squamous cell carcinoma (OSCC) patient survival rate remains stagnant. Conventional therapies have limited effectiveness, necessitating novel agents. Our study aims to synthesize and characterize amorphous calcium phosphate nanoparticles (nACPs), assess their potential cytotoxic effects on premalignant and malignant OSCC cells, and investigate possible mechanisms of action. The morphological features of nACP were investigated by field emission scanning coupled with energy dispersive spectroscopy (EDS), Fourier transform infrared spectroscopy (FTIR), and particle size distribution (PSD). Then, we examined the effect of nACPs on nanoparticle uptake, cell adhesion, viability, invasion ability, cell cycle, and gene expression. nACP uptake was dose-dependent, induced limited selectivity in cytotoxicity between healthy and malignant cells, and affected cellular adhesion and invasion. Early apoptosis was the predominant type of cell death. The nACP effect on viability was verified by alterations in the genes associated with apoptosis and proliferation. A high concentration of nACP was shown to arrest the cell cycle progression in the G0/G1 phase of both malignant and premalignant cells. This type of nACP justifies the development of a strategy for its potential use as an anti-cancer agent and/or anti-cancer active carrier for various drugs in oral cancer treatments.
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Undiagnosed and untreated oral precancerous lesions often progress into malignancies. Photodynamic therapy (PDT) might be a minimally invasive alternative to conventional treatments. 5-aminolevulinic acid (5-ALA) is one of the most commonly used photosensitizers in PDT, and it is effective on many cancer types. However, its hydrophilic characteristic limits cell membrane crossing. In the present study, the effect of a newly formulated gel containing 5% 5-ALA in combination with red light (ALAD-PDT) on a premalignant oral mucosa cell line was investigated. The dysplastic oral keratinocyte (DOK) cells were incubated with ALAD at different concentrations (0.1, 0.5, 1, and 2 mM) at two different times, 45 min or 4 h, and then irradiated for 7 min with a 630 nm LED (25 J/cm2). MTT assay, flow cytometry, wound healing assay, and quantitative PCR (qPCR) were performed. ALAD-PDT exerted inhibitory effects on the proliferation and migration of DOK cells by inducing ROS and necrosis. mRNA analysis showed modulation of apoptosis-related genes' expression (TP53, Bcl-2, survivin, caspase-3, and caspase-9). Furthermore, there was no difference between the shorter and longer incubation times. In conclusion, the inhibitory effect of the ALAD-PDT protocol observed in this study suggests that ALAD-PDT could be a promising novel treatment for oral precancerous lesions.
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Collagen membranes are routinely used in oral surgery for bone regeneration. Despite their numerous advantages, such as stimulating bone growth, bacterial contamination still remains one of the disadvantages of membrane use. Thus, we assessed the biocompatibility and osteogenic and antibacterial properties of a collagen membrane (OsteoBiol) modified with chitosan (CHI) and hydroxyapatite nanoparticles (HApNPs). Attenuated total reflectance-Fourier transform infrared spectroscopy (ATR FT-IR), X-ray powder diffraction (XRD), and field emission scanning electron microscopy (FE-SEM) were performed for membrane characterization. Biocompatibility was assessed on dental pulp stem cells (DPSCs) by an MTT assay, while the osteogenic effect was assessed by an ALP activity assay and qPCR analysis of osteogenic markers (BMP4, ALP, RUNX2, and OCN). Antimicrobial properties were investigated by counting colony-forming units (CFUs) of Streptococcus mitis, Porphyromonas gingivalis, and Fusobaterium nucleatum on membranes and in the surrounding medium. Membranes showed no cytotoxicity. ALP activity was higher and ALP, BMP4, and OCN genes were up-regulated in DPSCs on modified membranes compared to unmodified membranes. The CFUs were reduced on modified membranes and in the medium. Modified membranes showed great biocompatibility and a high osteoinductive effect. Additionally, they showed antimicrobial and antibiofilm effects against periopathogens. It can be concluded that the incorporation of CHI and hydroxyapatite nanoparticles in collagen membranes may be advantageous to promote osteogenesis and reduce bacterial adhesion.
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Antiinfecciosos , Quitosano , Osteogénesis , Quitosano/farmacología , Quitosano/química , Durapatita/farmacología , Durapatita/química , Diferenciación Celular , Espectroscopía Infrarroja por Transformada de Fourier , Colágeno/farmacología , Antiinfecciosos/farmacología , Células Cultivadas , Proliferación CelularRESUMEN
(1) Treatment failure of oral squamous cell carcinoma (OSCC) is generally due to the development of therapeutic resistance caused by the existence of cancer stem cells (CSCs), a small cell subpopulation with marked self-renewal and differentiation capacity. Micro RNAs, notably miRNA-21, appear to play an important role in OSCC carcinogenesis. Our objectives were to explore the multipotency of oral CSCs by estimating their differentiation capacity and assessing the effects of differentiation on stemness, apoptosis, and several miRNAs' expression. (2) A commercially available OSCC cell line (SCC25) and five primary OSCC cultures generated from tumor tissues obtained from five OSCC patients were used in the experiments. Cells harboring CD44, a CSC marker, were magnetically separated from the heterogeneous tumor cell populations. The CD44+ cells were then subjected to osteogenic and adipogenic induction, and the specific staining was used for differentiation confirmation. The kinetics of the differentiation process was evaluated by qPCR analysis of osteogenic (Bone Morphogenetic Protein-BMP4, Runt-related Transcription Factor 2-RUNX2, Alkaline Phosphatase-ALP) and adipogenic (Fibroblast Activation Protein Alpha-FAP, LIPIN, Peroxisome Proliferator-activated Receptor Gamma-PPARG) markers on days 0, 7, 14, and 21. Embryonic markers (Octamer-binding Transcription Factor 4-OCT4, Sex Determining Region Y Box 2-SOX2, and NANOG) and micro RNAs (miRNA-21, miRNA-133, and miRNA-491) were also correspondingly evaluated by qPCR. An Annexin V assay was used to assess the potential cytotoxic effects of the differentiation process. (3) Following differentiation, the levels of markers for the osteo/adipo lineages showed a gradual increase from day 0 to day 21 in the CD44+ cultures, while stemness markers and cell viability decreased. The oncogenic miRNA-21 also followed the same pattern of gradual decrease along the differentiation process, while tumor suppressor miRNA-133 and miRNA-491 levels increased. (4) Following induction, the CSCs acquired the characteristics of the differentiated cells. This was accompanied by loss of stemness properties, a decrease of the oncogenic and concomitant, and an increase of tumor suppressor micro RNAs.
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Adipogénesis , Carcinoma de Células Escamosas , MicroARNs , Neoplasias de la Boca , Células Madre Neoplásicas , Osteogénesis , Humanos , Carcinoma de Células Escamosas/patología , MicroARNs/metabolismo , Neoplasias de la Boca/patología , Células Madre Neoplásicas/citología , Células Madre Neoplásicas/metabolismoRESUMEN
High elution and diffusion of 2-hydroxylethyl methacrylate (HEMA) and camphorquinone (CQ) through dentinal tubules may induce pulp injury and postoperative sensitivity. We aimed to investigate the melatonin protective effect in HEMA- and CQ-treated human dental pulp cells (hDPCs) as well as its relevance in a mechanism for postoperative sensitivity in diabetic patients. hDPCs were exposed to HEMA (5 mM) and/or CQ (1 mM) in the absence and presence of melatonin (MEL) (0.1 mM and 1 mM). Heme oxygenase-1 (HMOX1), NADPH oxidase-4 (NOX4), BCL-2-associated X-protein (BAX), B-cell lymphoma-2 (BCL-2) and caspase-3 (CASP3) gene expression levels, and superoxide dismutase (SOD) activity were measured in hDPCs while inducible nitric oxide synthase (iNOS) and melatonin protein expression were measured in human dental pulp as well, by RT-PCR, by ELISA, and spectrophotometrically. Bioinformatic analyses were performed by using the ShinyGO (v.0.75) application. Type 2 diabetic patients showed a higher incidence of postoperative sensitivity and lower melatonin and higher iNOS content in dental pulp tissue compared with non-diabetic patients. Melatonin, when co-added in hDPC culture, reverses HEMA and CQ cytotoxic effects via anti-apoptotic and anti-inflammatory/antioxidant iNOS-related effects. Enrichment analyses showed that genes/proteins, altered by HEMA and CQ and normalized by melatonin, are the most prominently overrepresented in type 2 diabetes mellitus pathways and that they share subcellular localization in different oligomeric protein complexes consisting of anti- and pro-apoptotic regulators. This is the first evidence of the ability of melatonin to counteract iNOS-mediated inflammatory and stress effects in HEMA- and CQ-treated hDPCs, which could be of significance for the modulation of presently observed immediate postoperative sensitivity after composite restoration in type 2 diabetic patients.
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Diabetes Mellitus Tipo 2 , Melatonina , Humanos , Melatonina/farmacología , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Metacrilatos/farmacología , Óxido Nítrico Sintasa de Tipo II/genética , Óxido Nítrico Sintasa de Tipo II/metabolismo , Pulpa Dental/metabolismo , Antioxidantes , Proteínas Proto-Oncogénicas c-bcl-2/metabolismoRESUMEN
Students are particularly vulnerable from the mental health aspect, which was especially recognized during the COVID-19 pandemic. This study aimed to reveal the impact of COVID-19 on quality of life (QoL) and mental health among dental students. The study was conducted on a sample of 797 students (207 male and 592 female) with an average age of 21.7 ± 2.4, from the School of Dental Medicine, University of Belgrade. The measurements used in the study were the Demographic and Academic Questionnaire, Questionnaire about exposure to COVID-19, COVID-19-Impact on QoL Questionnaire (COV19-QoL), Generalized Anxiety Disorder 7-item (GAD-7) scale, and Patient Health Questionnaire (PHQ-9). The mean total score for COV19-QoL was 2.9 ± 0.9, while the diagnostic criteria of GAD-7 and depression met 19.9% and 31.4% of students, respectively. There was a positive and strong correlation between QoL, anxiety, and depression. During COVID-19, predictors for lower perceptions of QoL were female gender and death of close relatives (p = 0.049, p = 0.005, respectively). At the same time, predictors for GAD were female gender, living in dormitories, and death of close relatives (p = 0.019, p = 0.011, p = 0.028, respectively), while for depression they were year of study, living with parents, and death of close relatives due to COVID-19 (p = 0.012, p = 0.008, p = 0.029, respectively). The study showed that students' QoL and mental health during the pandemic were at high risk.
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COVID-19 , Masculino , Femenino , Humanos , Adulto Joven , Adulto , COVID-19/epidemiología , Pandemias , Calidad de Vida , Salud Mental , Estudios Transversales , Depresión/epidemiología , Estudiantes de Odontología , Ansiedad/epidemiologíaRESUMEN
OBJECTIVE: The study aimed to investigate acetylsalicylic acid (ASA) effects on osteo/odontogenic differentiation and proliferation of dental pulp stem cells (DPSCs) in vitro and the potential involvement of adenosine monophosphate-activated protein kinase (AMPK) pathway in these processes. DESIGN: DPSCs were isolated from third molars pulp tissues of five patients and grown in osteogenic medium alone or supplemented with ASA. Expression of DPSCs markers was tested by flow-cytometry. Cytotoxicity of ASA at concentrations of 10, 50 and 100 µg/ml was tested by MTT and NR assays. Osteo/odontogenic differentiation was analyzed via alizarin red staining and ALP activity. Quantitative PCR (qPCR) was used for osteo/odontogenic markers' (DSPP, BMP2, BMP4, BSP, OCN and RUNX2) and c-Myc expression analysis. AMPK inhibition of ASA-induced osteo/odontogenesis was tested by qPCR of selected markers (DSPP, OCN and RUNX2). RESULTS: Cytotoxicity assays showed that only the highest ASA dose decreased cell viability (89.1 %). The smallest concentration of ASA applied on DPSCs resulted in a remarkable enhancement of osteo/odontogenic differentiation, as judged by increased mineralized nodules' formation, ALP activity and gene expression of analyzed markers (increase between 2 and 30 folds), compared to untreated cells. ASA also increased DPSCs proliferation. Interestingly, AMPK inhibition per se upregulated DSPP, OCN and RUNX2; the gene upregulation was higher when ASA treatment was also included. c-Myc expression level decreased in cultures treated with ASA, indicating undergoing differentiation processes. CONCLUSIONS: Low concentrations of ASA (corresponding to the standard use in cardiovascular patients), were shown to stimulate osteo/odontogenic differentiation of dental pulp stem cells.
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Subunidad alfa 1 del Factor de Unión al Sitio Principal , Pulpa Dental , Humanos , Aspirina/farmacología , Proteínas Quinasas Activadas por AMP , Células Madre , Odontogénesis/fisiología , Diferenciación Celular , Osteogénesis/fisiología , Proliferación Celular , Células CultivadasRESUMEN
BACKGROUND: Dental stem cells, which originate from the neural crest, due to their easy accessibility might be good candidates in neuro-regenerative procedures, along with graphene-based nanomaterials shown to promote neurogenesis in vitro. We aimed to explore the potential of liquid-phase exfoliated graphene (LPEG) film to stimulate the neuro-differentiation of stem cells from apical papilla (SCAP). METHODS: The experimental procedure was structured as follows: (1) fabrication of graphene film; (2) isolation, cultivation and SCAP stemness characterization by flowcytometry, multilineage differentiation (osteo, chondro and adipo) and quantitative PCR (qPCR); (3) SCAP neuro-induction by cultivation on polyethylene terephthalate (PET) coated with graphene film; (4) evaluation of neural differentiation by means of several microscopy techniques (light, confocal, atomic force and scanning electron microscopy), followed by neural marker gene expression analysis using qPCR. RESULTS: SCAP demonstrated exceptional stemness, as judged by mesenchymal markers' expression (CD73, CD90 and CD105), and by multilineage differentiation capacity (osteo, chondro and adipo-differentiation). Neuro-induction of SCAP grown on PET coated with graphene film resulted in neuron-like cellular phenotype observed under different microscopes. This was corroborated by the high gene expression of all examined key neuronal markers (Ngn2, NF-M, Nestin, MAP2, MASH1). CONCLUSIONS: The ability of SCAPs to differentiate toward neural lineages was markedly enhanced by graphene film.
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Dentures and epitheses are mostly made from poly(methyl methacrylate) (PMMA), which does not show antimicrobial properties. They present reservoirs of microorganisms grown in biofilms. The aim of this study is to prepare a PMMA enriched with gold nanoparticles (AuNPs)-PMMA/AuNPs and the examination of its physical, mechanical and antimicrobial properties. The AuNPS were synthetized from HAuCl4 using the ultrasonic spray pyrolysis method with lyophilization. The PMMA/AuNP samples were compared to PMMA samples. Density was measured by pycnometer. Microhardness was evaluated using the Vickers hardness test. Monomicrobial biofilm formation (Streptococcus mitis, Candida albicans, Staphylococcus aureus and Escherichia coli) was measured by colony-forming units (CFUs) and MTT test and visualized by SEM. AuNP release was measured indirectly (the CFUs of the medium around the sample). The density and microhardness of the PMMA/AuNPs were similar to those of the PMMA. CFU and MTT values for the biofilms formed on the PMMA for each of the tested species were higher than those of the biofilms formed on the PMMA/AuNPs. The CFUs of the medium around the sample were similar for both materials. PMMA/AuNPs showed a significant reduction in the monomicrobial biofilms of all tested species. AuNPs are not released from PMMA/AuNPs. Density, indirect measurement of residual monomer and dentures weight were similar between PMMA and PMMA/AuNPs. Microhardness, as a measure of the wear resistance, was also similar between tested discs.
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Oral carcinoma is the sixth most common malignancy worldwide, with survival rates of approximately 50%. The major type of oral cancer, present in 90% of the cases, is oral squamous cell carcinoma (OSCC). The genetic background predisposing an individual to OSCC is complex and largely unknown. Studies have suggested that endothelial nitric oxide synthase (eNOS) gene polymorphisms modulate the cancer risk, prompting us to assess the impact of three functional eNOS gene polymorphisms on OSCC risk. The present study included 50 patients with OSCC and 110 controls. Polymerase chain reaction and restriction fragment length polymorphism analysis were used for genotyping of single-nucleotide polymorphisms -786 T/C (rs2070744) and 894 G/T (rs1799983) and variable number of tandem repeats (VNTR) intron 4b/a polymorphism. Homozygous carriers of -786 T/C and intron 4b/a VNTR variant alleles paired with a significant increase of oral cancer risk [odds ratio (OR): 3.63, 95% confidence interval (CI): 1.08-12.21; P = 0.045 and OR: 11.29, 95% CI: 2.71-47.11; P < 0.001, respectively]. When combined, CC and 4b4a genotypes together led to a 21-fold OSCC risk increase (OR: 21, 95% CI: 2.07-213.29; P = 0.006). Haplotype analysis showed that the C-G-4b haplotype conferred an 11-fold increase in OSCC risk. In conclusion, eNOS polymorphisms considerably influence levels of OSCC risk in the Serbian population.
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Carcinoma de Células Escamosas , Neoplasias de la Boca , Estudios de Casos y Controles , Frecuencia de los Genes , Predisposición Genética a la Enfermedad , Haplotipos , Humanos , Óxido Nítrico Sintasa de Tipo III/genética , Polimorfismo de Nucleótido SimpleRESUMEN
Epithelial to mesenchymal transition (EMT) is a feature of several types of human cancer, including oral squamous cell carcinoma (OSCC). In the present study, tumor and margin cell cultures obtained from patients with OSCC were used to determine the expression patterns of certain EMT-associated markers, including vimentin, α-smooth muscle actin, SLUG and SNAIL. In addition, other EMT-associated features, including clonal, proliferative and migratory potential were compared between the two cell types. Cell cultures were generated from tumor and margin tissue samples from 6 patients and cultured up to the fifth passage. EMT marker expression was assessed by reverse transcription-quantitative PCR. Cell proliferation, colony formation and scratch wound healing assays were conducted to characterize the two cell types in terms of proliferation rates, clonality and motility. All of the studied markers were expressed in tumor and margin cells. Although no significant differences were noted with regard to the aforementioned markers, their expression tended to be higher in margin cultures than in tumor cultures. The expressions of the EMT markers were also higher in the fifth passage compared with those noted at the first with a few exceptions. The rates of proliferation and cell migration were decreased during passages, while the number of colonies was increased in both types of cell culture. Tumor and margin cells indicated certain similarities with regard to EMT transition characteristics.
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OBJECTIVES: Sutures are the most frequently used medical device for wound closure. They support tissue during the early phase of healing until it regains enough tensile strength. The aim of this study was to compare four different suture materials in terms of the influence on wound healing, microbial adherence, tissue reaction, and relevant clinical parameters which determine their clinical value. MATERIALS AND METHODS: Total number of 32 patients undergoing surgical extraction of four impacted third molars were involved in the study. Clinical parameters were estimated intraoperatively and during the control check-ups. Soft tissue healing around sutures were evaluated on the 3rd and 7th day postoperatively. Microbial colonization was assessed by means of qPCR. Also, histological analysis was done to assess inflammatory reaction. RESULTS: Significantly better soft tissue healing was found around monofilament and synthetic sutures compared to multifilament and natural ones respectively. Soft tissue healing was significantly better around all sutures on the 7th day than on the 3rd day postoperatively. CONCLUSIONS: Non-resorbable polypropylene suture showed superior clinical characteristics among all sutures. Moreover, the best healing of soft tissue and the least inflammatory reaction was found around this thread. The poorest soft tissue healing was found around non-resorbable silk suture. This suture elicited strongest inflammatory reaction and showed the greatest microbial adherence affinity compared to alternative sutures. CLINICAL RELEVANCE: Monofilament synthetic suture should be used in order to obtain the best soft tissue healing, reduce the risk of postoperative infection, and alleviate the suturing after oral surgery procedures.
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Procedimientos Quirúrgicos Orales , Polipropilenos , Seda , Infección de la Herida Quirúrgica/prevención & control , Suturas , Cicatrización de Heridas , Humanos , Técnicas de Sutura/instrumentación , Resistencia a la TracciónRESUMEN
Taking advantage of the flexibility of the apatite structure, nano- and micro-particles of hydroxyapatite (HAp) were doped with different combinations of rare earth ions (RE3+ = Gd, Eu, Yb, Tm) to achieve a synergy among their magnetic and optical properties and to enable their application in preventive medicine, particularly diagnostics based on multimodal imaging. All powders were synthesized through hydrothermal processing at T ≤ 200 °C. An X-ray powder diffraction analysis showed that all powders crystallized in P63/m space group of the hexagonal crystal structure. The refined unit-cell parameters reflected a decrease in the unit cell volume as a result of the partial substitution of Ca2+ with smaller RE3+ ions at both cation positions. The FTIR analysis additionally suggested that a synergy may exist solely in the triply doped system, where the lattice symmetry and vibration modes become more coherent than in the singly or doubly doped systems. HAp:RE3+ optical characterization revealed a change in the energy band gap and the appearance of a weak blue luminescence (λex = 370 nm) due to an increased concentration of defects. The "up"- and the "down"-conversion spectra of HAp:Gd/Yb/Tm and HAp:Gd/Eu powders showed characteristic transitions of Tm3+ and Eu3+, respectively. Furthermore, in contrast to diamagnetic HAp, all HAp:RE3+ powders exhibited paramagnetic behavior. Cell viability tests of HAp:Gd/Yb/Tm and HAp:Gd/Eu powders in human dental pulp stem cell cultures indicated their good biocompatibility.
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AIM: To characterize stem cells originating from different dental tissues (apical papilla [SCAP], dental follicle [DFSC], and pulp [DPSC]) and test the capacity of Raman microspectroscopy to distinguish between the three dental stem cell types. METHODS: SCAP, DFSC, and DPSC cultures were generated from three immature wisdom teeth originating from three patients. Cell stemness was confirmed by inducing neuro-, osteo-, chondro-, and adipo-differentiaton and by mesenchymal marker expression analysis by flow-cytometry and real-time polymerase chain reaction. Cellular components were then evaluated by Raman microspectroscopy. RESULTS: We found differences between SCAP, DFSC, and DPSC Raman spectra. The ratio between proteins and nucleic acids (748/770), a parameter for discriminating more differentiated from less differentiated cells, showed significant differences between the three cell types. All cells also displayed a fingerprint region in the 600-700 cm-1 range, and characteristic lipid peaks at positions 1440 cm-1 and 1650 cm-1. CONCLUSION: Although different dental stem cells exhibited similar Raman spectra, the method enabled us to make subtle distinction between them.
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Pulpa Dental/citología , Saco Dental/citología , Células Madre Mesenquimatosas/química , Tercer Molar/citología , Espectrometría Raman , Adolescente , Diferenciación Celular , Citometría de Flujo , Humanos , Reacción en Cadena en Tiempo Real de la Polimerasa , Células Madre , DienteRESUMEN
BACKGROUND: Despite improvements in oral squamous cell carcinoma (OSCC) management, survival rates remain relatively low and novel anti-neoplastic agents are needed. Bromodomain and extra-terminal (BET) inhibitors proved to be promising agents for cancer treatment. We investigated the effects of three BET inhibitors (JQ1, IBET-151, IBET-762) on SCC-25 cell line and primary oral cancer cell culture. METHODS: Cell viability was evaluated by MTT. Protein levels of MCM5 and cleaved-PARP were estimated by Western blot. Clonogenic and migratory abilities were determined by colony forming and scratch assays. BET inhibitors effects on mRNA levels of E-Cadherin, Vimentin, SNAI1, SNAI2, CLU, SERPINI1, MCM5, c-Myc, E2F, IL7R, and PPARg were analyzed by qPCR. RESULTS: BET inhibitors significantly reduced oral cancer cell viability. JQ1 showed the greatest effect reducing cell viability to 10%, both in SCC-25 and primary OSCC cultures (P < 0.001), compared to control cells. Cells treated with BET inhibitors displayed a reduction to 50% in colony forming capacity compared to control cells (P < 0.0001) and the colonies were smaller; they also had a 50%-60% reduction in migratory capacity (P < 0.05) compared to untreated cells. BET inhibitors had a significant impact on genes related to epithelial to mesenchymal transition and other cancer cell markers, notably on MCM5, a gene related to cell cycle control. CONCLUSIONS: BET inhibitors induce both OSCC cell death and reduction of tumor aggressiveness. Molecular mechanisms of BET inhibition involve among others, MCM5 downregulation. Importantly, this study demonstrates for the first time the anti-tumoral effect of IBET-151 and IBET-762 in oral cancer.
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Antineoplásicos/farmacología , Azepinas/farmacología , Benzodiazepinas/farmacología , Carcinoma de Células Escamosas/patología , Supervivencia Celular/efectos de los fármacos , Neoplasias de la Boca/patología , Triazoles/farmacología , Carcinoma de Células Escamosas/genética , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Regulación hacia Abajo/efectos de los fármacos , Transición Epitelial-Mesenquimal/efectos de los fármacos , Humanos , Neoplasias de la Boca/genética , Células Madre Neoplásicas/efectos de los fármacosRESUMEN
BACKGROUND: Understanding the pathogenesis of basal cell carcinoma (BCC) and identifying the cells responsible for propagation and recurrence are crucial for the development of new treatment strategies. The aim of this study was to characterize the cells isolated from BCC and its margin. METHODS: Primary cultures were established from 10 BCCs, their respective close resection margins (3 mm) and 10 control tissues. Stem cell markers analysis was carried out by real-time PCR and/or flow cytometry. Spheroid formation and MTT assays were also performed. RESULTS: Real-time PCR showed a higher expression of embryonic (Oct4, Sox2 and Nanog) and mesenchymal (CD44 and CD73) stem cell markers in tumors compared to margins and controls (P < 0.05). Bmi-1 and GPR49 were also upregulated in tumors in comparison with margins. Both tumor and margin cells, but not normal, had the capacity to form spheroids. During passages, the number of spheres increased, while the diameter decreased. Tumor cells showed higher chemo-resistance compared to margin and control cells. CONCLUSIONS: Basal cell carcinomas expressed stem cell markers, pointing to the existence of a cancer cell side population with stemness characteristics. Margin also appeared to harbour a small number of cancer-initiating cells.
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Carcinoma Basocelular/patología , Márgenes de Escisión , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patología , ARN/metabolismo , Neoplasias Cutáneas/patología , 5'-Nucleotidasa/genética , Antineoplásicos/farmacología , Biomarcadores/metabolismo , Carcinoma Basocelular/cirugía , Supervivencia Celular/efectos de los fármacos , Cisplatino/farmacología , Resistencia a Antineoplásicos , Fluorouracilo/farmacología , Proteínas Ligadas a GPI/genética , Expresión Génica , Humanos , Receptores de Hialuranos/genética , Receptores de Hialuranos/metabolismo , Proteína Homeótica Nanog/genética , Células Madre Neoplásicas/fisiología , Factor 3 de Transcripción de Unión a Octámeros/genética , Complejo Represivo Polycomb 1/genética , Cultivo Primario de Células , Receptores Acoplados a Proteínas G/genética , Factores de Transcripción SOXB1/genética , Neoplasias Cutáneas/cirugía , Esferoides Celulares , Células Tumorales CultivadasRESUMEN
Lanthanide-doped fluoride up-converting nanoparticles (UCNPs) represent the new class of imaging contrast agents which hold great potential for overcoming existing problems associated with traditionally used dyes, proteins and quantum dots. In this study, a new kind of hybrid NaYF4:Yb,Er/PLGA nanoparticles for efficient biolabeling were prepared through one-pot solvothermal synthesis route. Morphological and structural characteristics of the as-designed particles were obtained using X-ray powder diffraction (XRPD), scanning and transmission electron microscopy (SEM/TEM), energy dispersive spectroscopy (EDS), Fourier transform infrared (FTIR) and photoluminescence (PL) spectroscopy, while their cytotoxicity as well as up-conversion (UC) labeling capability were tested in vitro toward human gingival cells (HGC) and oral squamous cell carcinoma (OSCC). The results revealed coexistence of the cubic (Fm-3m) and hexagonal (P63/m) phase in spherical and irregularly shaped nanoparticles, respectively. PLGA [Poly(lactic-co-glycolic acid)] ligands attached at the surface of UCNPs particles provide their enhanced cellular uptake and enable high-quality cells imaging through a near-infrared (NIR) laser scanning microscopy (λexâ¯=â¯980â¯nm). Moreover, the fact that NaYF4:Yb,Er/PLGA UCNPs show low cytotoxicity against HGC over the whole concentration range (10-50⯵g/mL) while a dose dependent viability of OSCC is obtained indicates that these might be a promising candidates for targeted cancer cell therapy.
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Diagnóstico por Imagen , Erbio/química , Ácido Láctico/química , Neoplasias de la Boca/diagnóstico por imagen , Neoplasias de Células Escamosas/diagnóstico por imagen , Ácido Poliglicólico/química , Espectroscopía Infrarroja Corta , Iterbio/química , Adulto , Muerte Celular , Línea Celular Tumoral , Encía/patología , Humanos , Neoplasias de la Boca/patología , Nanopartículas/química , Nanopartículas/ultraestructura , Neoplasias de Células Escamosas/patología , Copolímero de Ácido Poliláctico-Ácido Poliglicólico , Espectroscopía Infrarroja por Transformada de Fourier , Difracción de Rayos X , Adulto JovenRESUMEN
PURPOSE: Recent evidence suggests that small subpopulations of stem-like cells are accountable for tumour initiation, progression and metastasis. Until now, studies were focused exclusively on the characterization of these cell populations within the tumour itself, while tumour margins were neglected, although it is known that the histological and molecular status of tumour margins may play a significant role in the course of the disease. Therefore, the aims of the study were to isolate cells from oral squamous cell carcinomas and their respective margins, to characterize these cells using specific markers, to assess their self-renewal potential and determine their chemoresistance. METHODS: Cell cultures were obtained from 12 tissue specimens (6 tumours and 6 margins). Total RNA was extracted and gene expression analysis was done by real-time PCR (RT-PCR). Flow cytometry, immunocytometry, sphere formation and MTT assays were also applied. RESULTS: With minor differences, cells originating from both tumours and tumour margins showed the presence of stem cell markers CD133, Nanog, Sox2, CD44, and Oct4, had the capacity to form spheroids and showed chemoresistance. CONCLUSIONS: Subpopulations of margin cells appeared to have stemness properties which might raise the question of re-evaluation of optimal surgical management.