Defining Metaniches in the Oral Cavity According to Their Microbial Composition and Cytokine Profile.

The human oral microbiota consists of over 700 widespread taxa colonizing the oral cavity in several anatomically diverse oral niches. Lately, sequencing of the 16S rRNA genes has become an acknowledged, culture-independent method to characterize the oral microbiota. However, only a small amount of data are available concerning microbial differences between oral niches in periodontal health and disease. In the context of periodontitis, the cytokine expression in the gingival crevicular fluid has been studied in detail, whereas little is known about the cytokine profile in hard and soft tissue biofilms. In order to characterize oral niches in periodontal health, the oral microbiota and cytokine pattern were analyzed at seven different sites (plaque (P), gingival crevicular fluid (GCF), saliva (S), tongue (T), hard palate (HP), cheek (C) and sublingual area (U)) of 20 young adults using next-generation sequencing and multiplex immunoassays. Site-specific microbial compositions were detected, which clustered into three distinct metaniches ("P-GCF", "S-T-HP" and "C-U") and were associated with niche-/metaniche-specific cytokine profiles. Our findings allow the definition of distinct metaniches according to their microbial composition, partly reflected by their cytokine profile, and provide new insights into microenvironmental similarities between anatomical diverse oral niches.

Molecular community profiling of the bacterial microbiota associated with denture-related stomatitis.

Denture-associated stomatitis (DS) affects over two-thirds of denture-wearers. DS presents as erythema of the palatal mucosa in areas where denture-surface associated polymicrobial biofilms containing the fungus Candida albicans exist. The contribution of the oral bacterial microbiota toward the infection is unknown. Therefore, this study characterised the bacterial microbiota of sites within the oral cavity to identify potential associations with occurrence of DS. Denture-wearing patients were recruited (denture stomatitis (DS) n = 8; non-denture stomatitis (NoDS) n = 11) and the oral bacterial microbiota of the tongue, palate and denture-fitting surface was characterised using next-generation sequencing. Operational taxonomic units (OTUs) were identified to bacterial genera and species, and presence/absence and relative abundances were examined. A significant (P = 0.007) decrease in the number of OTUs and thus, diversity of the microbiota was observed in tongue samples of DS patients (vs non-DS). The microbiota of denture-fitting surfaces and palatal mucosae were similar. Large differences in the abundance of bacterial genera and species were observed at each sample site, and unique presence/absence of bacteria was noted. Presence/absence and relative abundance of specific bacteria associated with DS warrants further in vitro and in vivo evaluation, particularly as our previous work has shown C. albicans virulence factor modulation by oral bacteria.

Candida glabrata Has No Enhancing Role in the Pathogenesis of Candida-Associated Denture Stomatitis in a Rat Model.

Denture stomatitis (DS) is a condition characterized by inflammation of the oral mucosa in direct contact with dentures and affects a significant number of otherwise healthy denture wearers. Candida-associated DS is predominantly caused by Candida albicans, a dimorphic fungus that readily colonizes and forms biofilms on denture materials. Previous studies showed a requirement for Candida biofilm formation on both palate and dentures in infection and identified fungal morphogenic transcription factors, Efg1 and Bcr1, as key players in DS pathogenesis. While both C. albicans and Candida glabrata are frequently coisolated in mucosal candidiasis, a pathogenic role for C. glabrata in DS remains unknown. Using an established rat model of DS, we sought to determine whether C. glabrata alone or coinoculation with C. albicans establishes colonization and causes palatal tissue damage and inflammation. Rats fitted with custom dentures were inoculated with C. albicans and/or C. glabrata and monitored over a 4-week period for fungal burden (denture/palate), changes in body weight, and tissue damage via lactate dehydrogenase (LDH) release as well as palatal staining by hematoxylin and eosin (H&E) and immunohistochemistry for myeloperoxidase (MPO) as measures of inflammation. C. glabrata colonized the denture/palate similarly to C. albicans In contrast to C. albicans, colonization by C. glabrata resulted in minimal changes in body weight, palatal LDH release, and MPO expression. Coinoculation with both species had no obvious modulation of C. albicans-mediated pathogenic effects. These data suggest that C. glabrata readily establishes colonization on denture and palate but has no apparent role for inducing/enhancing C. albicans pathogenesis in DS.IMPORTANCE Many denture wearers suffer from Candida-associated denture stomatitis (DS), a fungal infection of the hard palate in contact with dentures. Biofilm formation by Candida albicans on denture/palate surfaces is considered a central process in the infection onset. Although Candida glabrata is frequently coisolated with C. albicans, its role in DS pathogenesis is unknown. We show here, using a contemporary rat model that employed a patented intraoral denture system, that C. glabrata established stable colonization on the denture/palate. However, in contrast to C. albicans inoculated rats, rats inoculated with C. glabrata exhibited minimal changes in weight gain or palatal tissue damage. Likewise, coinoculation with the two Candida species resulted in no exacerbation of C. albicans-induced DS pathology. Together, our findings indicate that C. glabrata has no inducing/enhancing role in DS pathogenesis.

The spectrum of intraoral bacteria seen in patients with cleft palates in an African setting.

Dehiscence or palatal fistula formation following palatoplasty is a complication that has grave consequences for the patient that include tissue loss, emotional distress to the parents and patient, and further medical costs. Palatal dehiscence or fistula formation is multifactorial following surgery-tension of wound closure, poor patient adherence to postoperative orders and wound infection are the most common causes for this. Oral colonization with pathogenic organisms could play a role in wound healing complications. Identification of intraoral bacteria among patients with cleft palates has thus far not been performed. To identify the spectrum of intraoral bacteria in cleft palate patients in an African setting; a retrospective, chart review was performed at Inkosi Albert Luthuli Central Hospital-a quaternary hospital in Durban, South Africa. All patients with unrepaired cleft palates who underwent palatoplasty in 2015 were included. Fifty-two patients were included. Preoperative throat/palatal swabs were taken prior to palatoplasty. The various bacteria cultured from the aforementioned swabs were cataloged. Various bacteria were cultured. In total, 23 patients (44.2%) had positive swab cultures. Eighteen cultures (34.6%) had gram-positive growth only, four cultures (7.7%) had gram-negative growth only, while one patient (1.9%) cultured both a gram-positive and a gram-negative organism. Streptococcus viridans was the most commonly cultured organism (19.2%) while beta-hemolytic streptococci were cultured from only 4 swabs (7.7%). Our study cataloged the commonly occurring bacteria found in unrepaired cleft palate patients in Africa. Further research into the clinical significance of each bacteria is advised.

A Case Report of Mucormycosis with Palatal Ulcer: A Rare Clinical Dilemma.

Mucormycosis represents a group of life-threatening infections caused by fungi of the order mucorales of subphylum mucormycotina (formely known as the class zygomycetes). It is highly invasive and progressive disease resulting in higher rates of morbidity and mortality. Newer therapies have declined the rates of mortality. Early initiation of therapy often before confirmation of the diagnosis is necessary to optimize outcomes. Here, we report a case of mucormycosis with palatal ulcer in a 63-year male patient, which is a rare anatomic site for the occurrence of mucormycosis or is a rare clinical presentation. The patient was successfully treated with surgical debridement and amphotericin B.

Actinomycotic osteomyelitis of palate masquerading periapical pathology: A rare case report.

Osteomyelitis is an infection that is challenging to manage due to the poor vascularization of bone that favors the proliferation of microorganisms. We report a case of osteomyelitis occurring in endodontically treated teeth in the maxillary palatal region. Clinically and radiographically, it was initially diagnosed as osteomyelitis and was treated accordingly with antibiotics for 1 year with no reported healing. Later, biopsy was done and the findings were consistent with that of chronic osteomyelitis in association with infection by Actinomyces organisms. Thus, the case highlights the rare occurrence of actinomycotic osteomyelitis in maxilla and the importance of biopsy and histopathology which will help in correct diagnosis and rapid resolution through appropriate antibiotic therapy.

Maxillary Osteomyelitis by Actinomyces in a Child / Osteomielitis Maxilar por Actinomyces en un Niño

ABSTRACT: Actinomycosis is a bacterial infection caused by Actinomyces species, which usually affect the soft tissues of the cervicofacial region of adult males. Clinically, it's characterized by a slow-growing indurated mass, especially in the submandibular area. However, in a few cases, the jaws bones can be affected developing osteomyelitis characteristics. The aim of this paper is to report a rare clinical case of Actinomycotic Osteomyelitis affecting the maxilla of a child, involving the maxillary sinus, orbital and zygomatic areas that was treated by the association of antibiotic therapy and surgical debridement. The patient's 2 years follow-up was uneventful and no signs of the lesion recurrence.

RESUMEN: La actinomicosis es una infección bacteriana causada por la especie Actinomyces, que generalmente afecta los tejidos blandos de la región cervicofacial de los hombres adultos. Clínicamente, se caracteriza por una masa endurecida de crecimiento lento, especialmente en la zona submandibular. Sin embargo, en algunos casos, los huesos de las mandíbulas pueden ser afectados desarrollando características de osteomielitis. El objetivo de este trabajo es reportar un caso clínico poco frecuente de osteomielitis actinomicótica que afecta el maxilar de un niño, envolviendo el área del seno maxilar, y zonas orbitales y cigomáticas que fueron tratadas con la asociación de terapia con antibióticos y desbridamiento quirúrgico. El seguimiento del paciente por 2 años ocurrió sin incidentes y sin signos de recidiva de las lesiones.

Palatal Petechiae in the Absence of Group A Streptococcus in Pediatric Patients with Acute-Onset Neuropsychiatric Deterioration: A Cohort Study.

BACKGROUND: Palatal petechiae are 95% specific for streptococcal pharyngitis. Despite this, and despite prior research demonstrating that Group A Streptococcus (GAS) is a common antecedent to pediatric acute-onset neuropsychiatric syndrome (PANS) episodes, we anecdotally observed a low rate of documented GAS in patients with PANS and palatal petechiae. This retrospective chart review was conducted to formally report the rate of palatal petechiae and concurrent GAS in a cohort of patients with PANS and investigate other etiologic factors. METHODS: The clinical notes of 112 patients seen at the Stanford PANS Clinic who met PANS research criteria were reviewed for mention of palatal petechiae. The medical records of patients who demonstrated palatal petechiae on physical examination were reviewed for signs of infection, a clinical history of trauma, and laboratory results that could indicate other causes of petechiae. RESULTS: Twenty-three patients had documented palatal petechiae on physical examination (ages 5-16, 13/23 [57%] male). Fifteen patients had a rapid GAS test and GAS culture in the Stanford PANS clinic, all with negative results. Evidence of recent GAS infection was found in 8/23 (32%) patients (elevated GAS titers [n = 6] or documentation of a positive rapid GAS test at another facility [n = 2]), one of whom also had potential herpes simplex virus (HSV) infection. One patient had potential HSV infection and recent palatal trauma. No patients had thrombocytopenia. 14/23 (61%) of patients with palatal petechiae had no discernable cause of petechiae. 10/19 (53%) of patients had antihistone antibodies. CONCLUSIONS: Despite the established relationship between palatal petechiae and GAS, no patient with palatal petechiae in our clinic tested positive for GAS and only 32% had evidence of recent GAS. Most did not have an identifiable cause for the palatal lesions. This finding suggests the potential for alternative causes of palatal petechiae or undetectable GAS in our patient population. The high prevalence of palatal petechiae without GAS infection suggests that the pathogenesis of PANS is multifactorial and may involve disruption or inflammation of the microvasculature. Additional research is needed to further elucidate these findings.

Invasive fungal rhinosinusitis with palatal erosion in an elderly edentulous patient with uncontrolled diabetes: report of a rare case.

OBJECTIVES: To report a rare case of chronic invasive fungal rhinosinusitis with palatal erosion. BACKGROUND: Restoring and maintaining oral health of diabetic elderly patients with increased risk of infections is a challenge to the dentist. Patients suffering from uncontrolled diabetes are susceptible to fungal infections. Palatal erosion due to fungal rhinosinusitis is rare. MATERIALS AND METHODS: Case report of a 65 years old illiterate female patient from low socio-economic strata, suffering from uncontrolled diabetes and poor systemic health presenting with chronic invasive fungal rhinosinusitis leading to palatal erosion. CONCLUSION: Such a case is a diagnostic challenge to a dentist. Therefore understanding the disease process and its possible outcomes is desirable. The treatment warrants a multidisciplinary approach.

Streptococcus mutans counts in patients wearing removable retainers with silver nanoparticles vs those wearing conventional retainers: A randomized clinical trial.

INTRODUCTION: The rough surface of Hawley removable appliances provides an environment for plaque accumulation, leading to enamel demineralization. The aim of this study was to evaluate the effect of silver nanoparticles incorporated into acrylic baseplates of orthodontic retainers on Streptococcus mutans colony-forming units. METHODS: Sixty-six orthodontic patients at the debonding stage were randomly assigned to 2 sex-matched groups with stratified block randomization: group 1 received conventional removable retainers; group 2 received removable retainers containing silver nanoparticles (about 40 nm in size and 500 ppm in concentration). After comprehensive orthodontic treatment, patients who revealed no clinical evidence of dental caries, periodontal pockets, or systemic disease were considered eligible for this study. Swab samples were taken from the maxillary palatal side from the patient in the dental chair at retainer placement (T1, 1 week after debonding the fixed orthodontic appliance) and T2, 7 weeks later. The main outcome was to compare the number of S mutans colony-forming units between the 2 groups 7 weeks after retainer delivery. The results were analyzed by using analysis of covariance. The participants and the assessors were blinded to the allocation groups. RESULTS: Twenty-nine patients in the control group and 32 in the intervention group were analyzed. At T1, the intervention group had higher S mutans colony counts relative to the control group. The analysis of covariance test showed a significant reduction of colonies in the intervention group after 7 weeks. The mean difference of colony counts between the 2 groups was 40.31 (95% confidence interval, 24.83-55.79; P < 0.001). CONCLUSIONS: Adding silver nanoparticles to the acrylic plate of retainers had a strong antimicrobial effect against S mutans under clinical conditions. REGISTRATION: This study was registered as a clinical trial at the Iranian Clinical Trial Center under the code number IRCT201309239086N2. FUNDING: This trial was supported by Hamadan Dental Research Centre, Hamadan University of Medical Sciences, Hamadan, Iran.

A multispecies probiotic reduces oral Candida colonization in denture wearers.

PURPOSE: The prevalence of Candida infections has been rising with an increasingly aging population and a larger population of immunocompromised individuals. The use of probiotics may be an alternative approach to antifungal agents in the prevention and treatment of oral candidiasis. This study aimed to evaluate the short-term effect of probiotics in reducing the infection level of oral Candida in candidiasis-asymptomatic elderly denture wearers. MATERIALS AND METHODS: In a double-blind randomized study, 59 denture wearers harboring Candida spp. in the oral cavity with no clinical symptoms were allocated into two groups: probiotic and placebo. All patients were instructed to clean the denture daily. The probiotic group poured a capsule containing lyophilized Lactobacillus rhamnosus HS111, Lactobacillus acidophillus HS101, and Bifidobacterium bifidum daily on the palatal surface of the maxillary denture, whereas the placebo group was submitted to the same regimen using placebo capsules. Candida spp. infection levels were evaluated in palate mucosa samples obtained before and after a 5-week experimental period. RESULTS: All patients harbored Candida in the palate mucosa at baseline. Fifty-five individuals completed the experimental period. The detection rate of Candida spp. was 92.0% in the placebo group after the experimental period, whereas it was reduced to 16.7% in the probiotic group. The reduction promoted by the probiotic regimen was independent of baseline characteristics such as Candida infection level and colonizing species, age of denture, and other variables. CONCLUSION: The probiotic product was effective in reducing the colonization of the oral cavity with Candida in candidiasis-asymptomatic elderly denture wearers, suggesting that this multispecies probiotic could be used to prevent oral candidiasis. CLINICAL IMPLICATIONS: Colonization of oral surfaces by Candida is considered a risk factor for invasive fungal infections. The use of a product with L. rhamnosus, L. acidophilus, and B. bifidum may represent an alternative treatment for reduction of Candida infections in elderly denture wearers.

Assessment of Candida species colonization and denture-related stomatitis in bar- and locator-retained overdentures.

The aim of this study was to assess the prevalence of denture-related stomatitis (DRS) in different attachment-retained overdenture wearers and its association with particular colonizing Candida species. Thirty-seven edentulous patients with implant-supported maxillary or mandibular overdentures were enrolled. A full clinical history was obtained, including details of patients' oral hygiene practices and the levels of erythema based on Newton's classification scale. Swabs were taken from the palate and investigated mycologically to identify the yeast colonies. Quantitative and qualitative microbiological assessments were performed, which included recording the total numbers of colonies (cfu), their color, and their morphological characteristics. Significant differences were found in cfu values between the attachment and inner surfaces of locator- and bar-retained overdentures (P < .05). Candida albicans was the most common species in both evaluations, being isolated from 81.3% of bar-retained overdentures and 38.1% of locator-retained overdentures. DRS developed in all patients using bar-retained overdentures but in only 71.4% of those using locator-retained overdentures. No statistically significant relationship was found between bar and locator attachments according to smoking habit, overnight removal, or plaque and gingival indices (P > .05).

Phase 1 clinical trial on the effect of palatal brushing on denture stomatitis.

PURPOSE: This study aimed to assess the efficacy of palatal brushing in the treatment of denture stomatitis. MATERIALS AND METHODS: After screening 143 individuals with a potential diagnosis of denture stomatitis, 48 patients (mean age: 66.0 ± 11.2 years) were enrolled in a two-center phase 1 clinical trial with a one-group pretest/posttest design. The intervention of interest was manual palatal brushing after each meal and before bedtime. Clinical and microbiologic examinations were performed at baseline and 1 and 3 months after treatment. Additional data were obtained using a validated questionnaire. The primary and secondary outcomes were the remission of denture stomatitis and diminution of Candida colony-forming units (CFUs), respectively. Descriptive and nonparametric statistical tests were conducted to analyze the data. RESULTS: At the 3-month follow-up, denture stomatitis was completely cured in 10.4% of the participants, and 70.8% of denture wearers showed improvement in the clinical signs of denture stomatitis. There was a significant reduction in the area and severity of the palatal inflammation (P < .0001). The effect size ranged from medium to large (0.34 to 0.54) depending on the classification used for the diagnosis of denture stomatitis. A significant reduction in the number of Candida CFUs isolated from the palatal mucosa and dentures (P ≤ .05) was observed. CONCLUSIONS: The results of this study suggest that palatal brushing is an effective treatment of denture stomatitis.

Crossover clinical trial of the influence of the use of adhesive on biofilm formation.

STATEMENT OF PROBLEM: Contrasting results have been reported regarding the influence of the use of adhesive on biofilm formation. PURPOSE: The purpose of this study was to evaluate the influence of the use of adhesive on the formation of biofilm on the internal surface of complete dentures and the palatal mucosa of denture wearers. MATERIAL AND METHODS: Thirty participants with well-fitting complete dentures were randomly divided according to the experimental design: protocol 1, adhesive use during the first 15 days, followed by no use of adhesive over the next 15 days; protocol 2, no use of adhesives during the first 15 days, followed by adhesive use over the next 15 days. After each period, material from the mucosa and intaglio of the maxillary dentures was collected. Replicate aliquots were plated onto Petri dishes containing selective media for Candida spp, Streptococcus mutans, and a nonselective culture medium. Colony-forming units were expressed as log (CFU+1)/mL. In addition, the internal surfaces of the maxillary and mandibular complete dentures were stained and photographed. From the photographs, the total internal surface and the surface stained with biofilm were quantified (software ImageTool 3.00), and the percentage of the biofilm-covered area (%) on the maxillary and mandibular dentures was calculated and compared with 2-way ANOVA. For the nonselective culture medium, data were compared with the paired-sample t test, and the Wilcoxon signed rank test was performed to compare the colony counts of Candida spp and Streptococcus mutans (α=.05). RESULTS: Similar colony counts were found with or without the use of adhesive for the mucosa and internal surfaces of maxillary dentures, irrespective of the culture medium (P>.05). The area of dentures covered with biofilm was influenced by the use of adhesive (P=.025), regardless of the type of denture (P=.121). CONCLUSIONS: The use of adhesive did not alter the colony counts of microorganisms from the palatal mucosa and maxillary dentures of complete denture wearers during the 15-day period, but it did influence the area covered with biofilm on the internal surfaces of the complete dentures.

Mucormycosis in a diabetic ketoacidosis patient.

Oral cavity is considered to be a kaleidoscope for body's general health. Many systemic conditions do present with diverse oral manifestations. Mucormycosis involving the oral cavity is one such entity that presents as necrosis of bone in immunocompromised patients. Mucormycosis is an opportunistic fungal infection that mainly affects the patients with uncontrolled diabetes mellitus. Hereby, we report a case of mucormycosis involving the palate in a patient with diabetic ketoacidosis.

Microbial geography of the oral cavity.

We aimed to determine the bacterial diversity of different oral micro-niches and to assess whether saliva and plaque samples are representative of oral microbial composition. We took minute samples from each surface of the individual teeth and gingival crevices of two healthy volunteers (112 samples per donor), as well as samples from the tongue dorsum and non-stimulated and stimulated saliva. DNA was extracted from 67 selected samples of each donor, and the 16S rRNA gene was amplified by PCR and pyrosequenced to obtain, on average, over 2,700 reads per sample, which were taxonomically assigned to obtain a geographic map of bacterial diversity at each tooth and sulcus location. Analysis of the data shows considerable differences in bacterial composition between teeth at different intra-oral locations and between surfaces of the same tooth. The most pronounced differences were observed in incisors and canines, where genera like Streptococcus were found at 40% to 70% on the vestibular surfaces but were almost absent on the lingual sides. Saliva samples, especially non-stimulated saliva, were not representative of supra-and subgingival plaque in the two individuals tested. We suggest that more precise sampling is required for the proper determination of oral microbial composition and to relate that diversity to epidemiological, clinical, and etiological parameters.

Effects of local simvastatin-alendronate conjugate in preventing periodontitis bone loss.

BACKGROUND AND OBJECTIVE: Local host-modulation therapy is an emerging approach to prevent disease progression in sites with moderate periodontitis. The combination of simvastatin and alendronate would be an intriguing host-modulatory strategy because of the bone-anabolic properties of simvastatin and the antiresorptive/bone-targeting characteristics of alendronate. The objective of this study was to evaluate the effects of local administration of a simvastatin-alendronate-ß-cyclodextrin (SIM-ALN-CD) conjugate for preventing experimental periodontitis bone loss. MATERIAL AND METHODS: Twenty-four mature female Sprague-Dawley rats were treated with three, 12 µL injections, administered one week apart, bilaterally into the palatal/interproximal gingiva. The injections contained: (i) a conjugate of 0.5 mg of SIM and 3.75 mg of ALN-CD in H2 O; (ii) H2 O alone; or (iii) no treatment. One week later, the same sites were subjected to induction of experimental periodontitis by three injections (i.e. one injection administered every other day for five d) of 0.01 mg of Escherichia coli endotoxin [lipopolysaccharide (LPS)] in phosphate-buffered saline (PBS) or PBS alone. After an additional week, the rats were killed, the palates were harvested and interproximal bone volume and adjacent thickness were calculated using microcomputed tomography. Subsequently, specimens were decalcified, and interproximal histologic sections were stained with hematoxylin and eosin for evaluation of alveolar crest osteoclasts and surrounding inflammation. Values were compared among treatment groups using analysis of variance and the Kruskal-Wallis test. RESULTS: Interproximal bone volume was reduced by LPS injections (p ≤ 0.04), yet when experimental periodontitis was preceded by treatment with SIM-ALN-CD, more bone was preserved than after treatment with carrier alone (p = 0.007). While LPS caused a significant loss in bone thickness over the palatal roots (p ≤ 0.04), the injection protocol (PBS) also caused a significant loss of palatal bone thickness (p ≤ 0.03). However, prophylactic SIM-ALN-CD injections resulted in no further loss of bone thickness during experimental periodontitis. LPS injections gave histologic evidence of increased osteoclasts and subsulcular inflammation, both of which were reduced when preceded by treatment with SIM-ALN-CD (p ≤ 0.0002). CONCLUSION: The primary conclusion of this study was that locally applied SIM-ALN-CD has the potential to prevent episodes of periodontitis bone loss.

Resposta imune inata na estomatite protética: interação in vitro entre células epiteliais de palato humano e Candida albicans / Innate imune response in denture stomatitis: in vitro interaction between human palatal epithelial cells and Candida albicans

A presença de Candida albicans nos biofilmes microbianos da superfície interna das próteses totais superiores está relacionada com uma doença inflamatória no palato, a estomatite protética. Constituinte da defesa inata do hospedeiro, o epitélio bucal, por sua vez, tem a capacidade de reconhecer e reagir aos fatores fúngicos a fim de evitar a invasão pelo microrganismo. O objetivo deste trabalho foi avaliar in vitro o efeito direto e indireto de C. albicans viável sobre as células epiteliais de palato humano (CEPH) ao longo do tempo. Objetivamos correlacionar os eventos de agressão, apoptose e invasão das CEPH provocados pelo fungo, com as respostas de defesa epitelial mediante produção de óxido nítrico (NO) e expressão gênica do peptídeo antimicrobiano β-defensina 2 (hBD-2). Material e Métodos: As CEPH foram obtidas, parte pelo método explante e parte pelo método enzimático, e mantidas em co-cultivo sobre uma camada de sustentação feederlayer (fibroblastos gengivais humanos mitoticamente inativados). Após desafios das CEPH com C. albicans ATCC 90028 por contato direto fungo-epitélio (D.D.) e indireto pelo sobrenadante da cultura do fungo hifal (D.I.), proporções de desafio de 0,01/1; 0,025/1 e 0,1/1 levedura/queratinócito (FUN/EPI) e tempos experimentais de 3, 6 e 10 h foram determinados; via ensaios de viabilidade celular por imunofluorescência (LIVE/DEAD), e análise qualitativa da invasão celular pelo fungo por meio do método colorimétrico com laranja de acridina. A apoptose epitelial foi determinada pela marcação nuclear fluorescente com Hoechtst 33258. A produção de óxido nítrico (NO) e a expressão de RNAm de hBD-2 foram avaliados por reação colorimétrica de Griess e RT-qPCR, respectivamente. Os resultados foram expressos como média ± desvio padrão e submetidos aos testes estatísticos ANOVA Fatorial, Teste de Contraste; ou Teste de Mann-Whitney (p<0,05). Resultados: Em 3 h, foi detectado aumento da apoptose das células epiteliais em relação ao...

The presence of the fungus Candida albicans in the microbial biofilm underlying maxillary prosthesis is related to an inflammatory reaction of the palatal mucosa, the denture stomatitis. As a component of the host innate defense, the oral epithelium has the ability to recognize and react to fungal factors in order to prevent the microrganism invasion. The aim of this study was to in vitro evaluate the direct and indirect effect of viable C. albicans on the human palatal epithelial cells (HPEC) over time. The aggressive events, such as apoptosis and HPEC invasion by the fungus, were correlated with epithelial defense responses through the nitric oxide (NO) production and antimicrobial peptides β-defensin (hBD-2) mRNA expression. Methods: The HPEC were obtained by explant and enzymatic methods, and were maintained in co-culture on a feeder-layer support (mitotically inactivated human gingival fibroblasts). After the HPEC challenges with C. albicans ATCC 90028 by direct contact fungus-epithelium (D.D.) and indirect contact by supernatant from hyphal fungus (D.I.), defiance ratios of 0.01/1, 0.025/1 and 0.1/1 yeast/keratinocyte (FUN/EPI) and experimental times of 3, 6 and 10 h were determined. These conditions were standardized by cell viability immunofluorescence assay (LIVE/DEAD), and cell invasion qualitative analysis (colorimetric method with acridine orange). The apoptotic cells were determined by fluorescent nuclear staining with Hoechtst 33258. The nitric oxide (NO) production and hBD-2 gene expression were evaluated by Griess colorimetric reaction and RT-qPCR, respectively. The results were expressed as mean ± standard deviation and were analyzed using the factorial ANOVA, Contrast Test; or Mann-Whitney Test (p<0,05). Results: At 3 h, the apoptotic epithelial cells under 0.1/1 FUN/EPI increased compared to epithelium unchallenged (p<0,05) that remained over time with increasing concentration and independent of D.D. and D.I. The onset...