Human Buccal Epithelial Cells Research Articles

Comprehensive in vitro and in vivo evaluation of therapeutic potential of Bacopa-derived asiatic acid against a human oral pathogen Streptococcus mutans.

Dental caries is a common human oral disease worldwide, caused by an acid-producing bacteria Streptococcus mutans. The use of synthetic drugs and antibiotics to prevent dental caries has been increasing, but this can lead to severe side effects. To solve this issue, developing and developed countries have resorted to herbal medicines as an alternative to synthetic drugs for the treatment and prevention of dental caries. Therefore, there is an urgent need for plant-derived products to treat such diseases. Bacopa monnieri, a well-documented medicinal plant, contains 52 phytocompounds, including the pentacyclic triterpenoid metabolite known as asiatic acid (ASTA). Hence, this study aimed to demonstrate, for the first time, the antibacterial activity of phytocompound ASTA against S. mutans. The findings revealed that ASTA significantly inhibited the growth of S. mutans and the production of virulence factors such as acidurity, acidogenicity, and eDNA synthesis. Molecular docking analysis evaluated the potential activity of ASTA against S. mutans virulence genes, including VicR and GtfC. Furthermore, toxicity assessment of ASTA in human buccal epithelial cells was performed, and no morphological changes were observed. An in vivo analysis using Danio rerio (zebrafish) confirmed that the ASTA treatment significantly increased the survival rates of infected fish by hindering the intestinal colonization of S. mutans. Furthermore, the disease protection potential of ASTA against the pathognomonic symptom of S. mutans infection was proven by the histopathological examination of the gills, gut, and kidney. Overall, these findings suggest that ASTA may be a promising therapeutic and alternative drug for the treatment and prevention of oral infection imposed by S. mutans.

Investigation of Hydrocolloid Plant Polysaccharides as Potential Candidates to Mimic the Functions of MUC5B in Saliva

The successful substitution of complex physiological fluids, such as human saliva, remains a major challenge in drug development. Although there are a large number of saliva substitutes on the market, their efficacy is often inadequate due to short residence time in the mouth, unpleasant mouthfeel, or insufficient protection of the teeth. Therefore, systems need to be identified that mimic the functions of saliva, in particular the salivary mucin MUC5B and the unique physiological properties of saliva. To this end, plant extracts known to contain hydrocolloid polysaccharides and to have mucus-forming properties were studied to evaluate their suitability as saliva substitutes. The aqueous plant extracts of Calendula officinalis, Fucus sp. thalli, and lichenan from Lichen islandicus were examined for composition using a range of techniques, including GC-MS, NMR, SEC, assessment of pH, osmolality, buffering capacity, viscoelasticity, viscoelastic interactions with human saliva, hydrocolloid network formation, and in vitro cell adhesion. For this purpose, a physiologically adapted adhesive test was developed using human buccal epithelial cells. The results show that lichenan is the most promising candidate to mimic the properties of MUC5B. By adjusting the pH, osmolality, and buffering capacity with K2HPO4, it was shown that lichenan exhibited high cell adhesion, with a maximum detachment force that was comparable to that of unstimulated whole mouth saliva.

Human buccal epithelial cells as a model system for molecular analysis of DNA, RNA and protein

We report use of human buccal epithelial cells as an easy model system for isolation and molecular analysis of genomic DNA, RNA, and protein to study any gene of interest by Polymerase Chain Reaction (PCR), RNA expression by Reverse Transcription-PCR (RT-PCR), protein-profiling, and expression by western blot as well as DNA-methylation by Msp I/Hpa II-restriction digestion. We used simple methods to isolate genomic DNA, RNA and protein from human buccal cells collected by oral swab and cultured them in-vitro. The microscopic observation of haematoxylin and eosin (EA-50) stained cells, genomic PCR of house-keeping genes (GAPDH and β-actin), RT-PCR of GAPDH and β-actin mRNAs and whole cell protein-profiling by Sodium Dodecyl Sulphate-Polyacrylamide Gel Electrophoresis (SDS-PAGE) were carried out. Expression of β-actin protein in supernatant and pellet fractions of the cells was determined by western blot analysis. MTT-assay was carried out to assess the cell viability and cell growth. Green Fluorescence Protein (GFP)-DNA was expressed in these cells by transient transfection. DNA-methylation in the genome was analyzed by Msp I/ Hpa II restriction digestion and agarose gel electrophoresis. Thus these methods can be used for molecular analysis of DNA, RNA and protein from the human buccal epithelial cells for studying and monitoring health, disease, population genetics/genomics and epidemiology under different conditions.

An in Vitro Study on Comparative Adherence of Oral Candida Albicans and Non–albicans Candida to Human Buccal Epithelial Cells and Its Antifungal Susceptibility Pattern

An in Vitro Study on Comparative Adherence of Oral Candida Albicans and Non–albicans Candida to Human Buccal Epithelial Cells and Its Antifungal Susceptibility Pattern

Diode Character of Local Conductivity of Human Buccal Epithelial Cell Membranes

AFM methods revealed unilateral (diode) local electrical conductivity of the membrane of human buccal epithelium cells at the nanoscale and its correlation with the topology of micromechanical properties.

AFM methods for studying the morphology and micromechanical properties of the membrane of human buccal epithelium cell

Using AFM methods in air under normal conditions in a wide range of local force effects ({F}_{const}< 40 μN) the relief, functional micromechanical properties (elasticity coefficient K, Young’s modulus E, elastic {Delta h}_{dfrm} and plastic {Delta h}_{stiff} deformations) and adhesive properties (work A of adhesive forces {F}_{adh}={F}_{adh}(x;y)) of the membranes of living adult cells of human buccal epithelium were studied in the presence of a protective layer < 100 nm of buffer solution that prevented the cells from drying. Almost all geometric and functional characteristics of the membrane in the local approximation at the micro- and nanolevels are affected by size effects and obey the laws of fractal geometry. The Brownian multifractal relief of the membrane is characterized by dimension {D}_{f} < 2.56 and irregularities < 500 nm vertically and < 2 μm horizontally. Its response to elastic (≤ 6 nN), active (6–21 nN), or passive (> 21 nN) stimulation ({F}_{const}) is a non-trivial selective process and exhibits a correspondingly elastic (K= 67.4 N/m), active (K= 80.2 N/m) and passive (K= 84.5 N/m) responses. K=K({F}_{const}) and E=E({F}_{const}) depend on {F}_{const}. Having undergone slight plastic deformations {Delta h}_{stiff} < 300 nm, the membrane is capable of restoring its shape. We mapped (E=E(x;y), {D}_{f} = 2.56; {Delta h}_{dfrm}={Delta h}_{dfrm}(x;y), {D}_{f} = 2.68; {Delta h}_{stiff}={Delta h}_{stiff}(x;y), {D}_{f } = 2.42, A=Aleft(x;yright) and {F}_{adh}={F}_{adh}(x;y)) indicating its complex cavernous structure.

Introducing a new type of alternative laryngeal mucosa model.

Research of human vocal fold (VF) biology is hampered by several factors. The sensitive microstructure of the VF mucosa is one of them and limits the in vivo research, as biopsies carry a very high risk of scarring. A laryngeal organotypic model consisting of VF epithelial cells and VF fibroblasts (VFF) may overcome some of these limitations. In contrast to human VFF, which are available in several forms, availability of VF epithelial cells is scarce. Buccal mucosa might be a good alternative source for epithelial cells, as it is easily accessible, and biopsies heal without scarring. For this project, we thus generated alternative constructs consisting of immortalized human VF fibroblasts and primary human buccal epithelial cells. The constructs (n = 3) were compared to native laryngeal mucosa at the histological and proteomic level. The engineered constructs reassembled into a mucosa-like structure after a cultivation period of 35 days. Immunohistochemical staining confirmed a multi-layered stratified epithelium, a collagen type IV positive barrier-like structure resembling the basement membrane, and an underlying layer containing VFF. Proteomic analysis resulted in a total number of 1961 identified and quantified proteins. Of these, 83.8% were detected in both native VF and constructs, with only 53 proteins having significantly different abundance. 15.3% of detected proteins were identified in native VF mucosa only, most likely due to endothelial, immune and muscle cells within the VF samples, while 0.9% were found only in the constructs. Based on easily available cell sources, we demonstrate that our laryngeal mucosa model shares many characteristics with native VF mucosa. It provides an alternative and reproducible in vitro model and offers many research opportunities ranging from the study of VF biology to the testing of interventions (e.g. drug testing).

An in vitro study on comparative adherence of oral Candida albicans and non–albicans Candida to human buccal epithelial cells and its antifungal susceptibility pattern

Background: Candida spp. are intra-oral commensals in 40-60% of human subjects and the precise factors involved in the development of Candidiasis are still not clear. The adherence of Candida to epithelial cells is one of the main pathogenic characteristics of the genus. Studies have shown that there are variations in the adherence capabilities of different Candida spp., which may explain why some species colonize mucosal surfaces more frequently than others. The aim of this study was to estimate the adherence properties of Candida spp. to buccal epithelial cells and to determine the antifungal susceptibility. Methods: 87 isolates of Candida spp. from known non-diabetic patients with oral Candidiasis were collected from diagnostic microbiology laboratories in central Kerala. The method of Kimura and Pearsall et al was followed with minor modifications to test the adherence. Results: All the three species exhibited adherence with varying capacity. C. albicans was significantly more adherent to buccal epithelial cells than C. tropicalis and C. krusei. Resistance was not observed among the isolates against fluconazole, voriconazole and clotrimazole. Conclusions: In conclusion Candida spp. isolated from oral candidiasis patients exhibited a higher adherence capacity to normal human buccal epithelial cells considered an essential virulence property. As anticipated C. albicans displayed higher virulence activity than non-Candida albicans species.

Syzygium aromaticum extracts debilitate Candida albicans by radically inhibiting its morphological plasticity and biofilm formation

ABSTRACT The effects of Syzygium aromaticum extracts on the growth, germ tube production, biofilm formation and adhesion of Candida albicans to human buccal epithelial cells (BEC) were investigated. We found larger zones of inhibition of growth of C. albicans with the essential oil (EO) and methanolic extract (ME) than amphotericin B. Similarly, the MIC values for the EO and ME were lower than that of amphotericin B. The EO and ME caused greater reductions in biofilm formation than amphotericin-B. Pre-treatment of C. albicans and BEC with the EO caused greater reductions in adhesion to BEC than amphotericin B.

Study of the influence of hops on the safety of brewing semi-products

In order to ensure quality control and food safety, the in vivo effect of hops on the frequency of cell aberrations was studied for the first time using a micronucleus test in human buccal epithelium. Results were obtained of in vivo studies of wort samples to identify possible negative effects of hops, which contribute to an increase in extractive components in the finished drink during dry hopping and contain a large number of biologically active compounds (bitter and phenolic substances, essential oils), on the human body. The frequency of nucleus abnormalities in human buccal epithelium cells of persons using hoped and unhopped must samples was analyzed for the frequency of cellular abnormalities in human buccal epithelium. The following types of disorders were found: cells with micronuclei, two nuclei, notches, perinuclear vacuoles, "tongue" and "broken egg" type protrusions, karyopyknosis, karyolisis and karyorexis. Based on the data obtained, indices of repair and accumulation of cytogenetic disorders were calculated. No effect of hops on the reparation index and the index of accumulation of cytogenetic disorders was revealed. Factor analysis methods show that the main contribution to the variance of the trait system will be made by the factor "specificity of the cytogenetic indices under study".

Study of tribological properties of human buccal epithelium cell membranes using probe microscopy

In this work demostrates a unique method for determining the absolute value of the friction force of a nanoobject on the surface of a cell membrane using atomic force microscopy. The tribological properties of membranes of adult human buccal epithelium cells in the presence of a protective adsorption buffer layer of ~ 100 nm on their surface were studied using atomic force microscopy in the contact scanning mode. Local mapping of the tribological characteristics of the surface was carried out, viz. friction FL = FL(x, y) and adhesion Fadh = Fadh(x, y) forces were measured. Studies of the friction force Ffr on the membrane surface at the nanolevel showed that its value varies discretely with an interval equal to lLF ≈ 100 nm. It was shown that such discreteness is determined by the interval lLF of the action of adhesive forces Fadh and indicates the fractal nature of the functional dependence of the friction force on the coordinate Ffr = Ffr(x). Thus, for nano-objects with dimensions ≤ lLF, the absolute value of Ffr decreases according to a power law with an increase in the size of the object, which contradicts the similar dependence of the friction force for macro-objects in the global approximation.

Antifungal Activity of Green and Red Brazilian Propolis Extracts

Background: Diseases associated to Candida spp. are recurrent and can be difficult to treat, mainly due to the new strains resistant to the limited number of available antifungals. Objective: The objective of this study is to evaluate the in vitro and in vivo activity of Brazilian green propolis (GrProp) and red propolis (RdProp) ethanolic extracts against standard strains of Candida albicans, C. tropicalis, C. krusei, C. glabrata, C. parapsilosis and 10 clinical isolates of C. albicans. Methods: Antifungal activity in vitro was tested using the M-27-A3/CLSI protocol. The in vivo antifungal activity was evaluated using Tenebrio molitor model and, the effect of extracts on adhesion of C. albicans in human buccal epithelial cells (BECs) was also studied. Results: GrProp and RdProp exhibited antifungal activity against at least one of the Candida strains tested. The adhesion inhibition of C. albicans in BECs was 45% (GrProp28), 60% (Gr- Prop50) and 82% (RdProp), in comparison to amphotericin B (82%). All propolis extracts showed synergistic activity with fluconazole and amphotericin B. GrProp50 (10 mg/kg) showed better protection of T. molitor, blocking the progression of C. albicans infection, increasing survival and delayed the larvae death. Conclusion: Brazilian GrProp and RdProp extracts inhibit the in vitro C. albicans growth and protect T. molitor against infection by this yeast. The physiochemical parameters found for the analyzed samples were in accordance to the standards established by the Brazilian Legislation for propolis and derivatives. GrProp and RdProp have potential to be used against Candida spp. infections, mainly in association with fluconazole or amphotericin B.

Human buccal epithelial cell envelope reactions to external micromechanical stimuli

The reaction of the shell of an adult living cell of human buccal epithelium to an external micromechanical stimulus was studied. The dependence of the magnitude of the reaction on the level of passive or active stimulation, namely, the level of force exposure, was established. It is shown that the cell can exhibit both active and passive reactions to an external micromechanical stimulus, as well as control the level of exposure to adhesive forces. Keywords: human buccal epithelium cell, shell, micromechanical stimulus, shell elasticity, AFM force spectroscopy.

Investigation of the relief geometry of the membrane of human buccal epithelium cells using atomic force microscopy methods

The analysis of the shape of the Brownian relief of the surface of human buccal epithelium cells was carried out using the method of fractal geometry. The scaling coefficients eta, zeta and the fractal dimension Df were used to estimate their actual areas Sc(fact) and linear characteristics lf. Keywords: atomic force microscopy, buccal epithelium, fractal number.

Исследование геометрии рельефа оболочки клеток буккального эпителия человека методами атомно-силовой микроскопии

The analysis of the shape of the Brownian relief of the surface of human buccal epithelium cells was carried out using the method of fractal geometry. The scaling coefficients ,  and the fractal dimension Df were used to estimate their actual areas Scfact and linear characteristics lf.

Peran Hemaglutinin dan Hemolisin pada Escherichia coli Sorbitol-negatif Isolat Burung Puyuh pada Proses Infeksi Secara in Vitro

Abstract Escherichia coli sorbitol-negative in quails cause economic loss due to the death, growth rate inhibition, decreased egg production, and increased medical treatment. Escherichia coli sorbitol-negative has many virulence factors, including hemagglutinin and hemolysin. The aim of this study is to determine the role of hemagglutinin and hemolysin in the infection process of Escherichia coli sorbitol-negative in vitro. This study was performed using 23 isolates of Escherichia coli sorbitol-negative from quails, 52.2% (12 of 23 isolates) had hemagglutinin, while 34.8% (8 out of 23 isolates) had hemolysin. Isolates with hemagglutinin were more attached to human buccal epithelial cells than isolates without hemagglutinin (P &lt;0.05). Isolates with hemolysin were less phagocyted by macrophages compared to isolates which without hemolysin (P &lt;0.05). Escherichia coli sorbitol-negative isolates from quails that have hemagglutinin and hemolysin are pathogenic isolates that possess the potential to cause colibasilosis and transmission between quails and other birds.

Interaction between Interleukin-1&lt;i&gt;α&lt;/i&gt; and Protease-activated Receptor-2 in Human Buccal Mucosa Epithelial Cells

Interaction between Interleukin-1&lt;i&gt;α&lt;/i&gt; and Protease-activated Receptor-2 in Human Buccal Mucosa Epithelial Cells

Quantitative assessment of micronuclei in oral exfoliative buccal squamous cells in smokers, chewers and healthy individuals using Giemsa Stain and Papainicolous Stain

Context: The main reason for oral cancer is tobacco consumption in various forms, such as chewing and smoking. Micronuclei in oral exfoliated cells are markers of chromosomal damage caused by toxic genetic agents from tobacco-related substances, alcohol, etc. The number of micronuclei indicates the cell's genotoxic effect and can be used as a biomarker to assess the changes in the cellular changes. Recently research is focused on determining early genotoxicity changes using exfoliative cytology in human buccal epithelial cells. Aim: Quantitative analysis of micronuclei in oral exfoliated oral, buccal mucosal cells using Papainicolous stain with Giemsa stain in tobacco users in the form of chewing and smoking in comparison with healthy individuals. Materials and methods: A total of seventy-five cases were selected for this study. Among seventy-five participants divided into three groups based on habits tobacco smokers (n=25), tobacco chewers (n=25), and healthy individuals (n=25). Results: The mean number of micronuclei in Giemsa stain was 255.2 ± 78 compared with PAP stain155.12 ± 78.5. The increased mean number of micronuclei in chewers 273.24 ± 80.7, and different stain comparison Giemsa stain showed highest in chewers. A mean number of micronuclei showed statistically highly significant (p≤0.05). Pair-wise comparison of micronuclei inter and intra groups in all three groups, smokers and chewers showed highly statistically substantial (p≤ 0.0001) compared with healthy individuals. Conclusion: Micronuclei provide evidence that consumption of smokeless tobacco may be developing oral cancer, and micronuclei assay can be used as biomarkers of genotoxicity and carcinogenesis progression. Key word: Cytological smear, Genotoxicity, Micronuclei, Oral cancer, Tobacco chewers

Cysteamine derived N/S co-doped carbon dots for fluorescence imaging of pathogenic bacteria and human buccal epithelial cells

Here, we report cysteamine derived nitrogen and sulfur co-doped carbon dots (Cy@CD) for imaging pathogenic bacteria and human buccal epithelial cells. The Cy@CD was synthesized using one-step simple microwave-assisted hydrothermal synthesis with precursors, namely, cysteamine and 50X TAE buffer. The spectroscopic characterization of Cy@CD showed (λex/λem) at 350/434 nm with a decent quantum yield of 13.2%. Cy@CD was deployed in tagging bacterial cells such as E. coli, S. aureus, P. aeruginosa and B. subtilis and human buccal epithelial cells. The multicolour emissive Cy@CD tagged microbial cells were also cross-validated by fluorometry analysis. The fluorescent Cy@CD could be used as an ideal substitute for the harmful synthetic fluorescent stain in cell imaging applications.

Regulation of Jun and Fos AP-1 transcription factors by JNK MAPKs signaling cascade in areca nut extract treated KB cells

The edible endosperm of Areca catechu is recognized as a potent carcinogenic agent either consumed alone or in combination with tobacco. Habitual chewing of areca nut leads to orally potential malignant disorders which are highly effective in malignant transformation and thereby lead to oral carcinogenesis. Human buccal epithelial KB carcinoma cells were used as an experimental cell system to inspect the mechanistic act of aqueous extract of areca nut on biochemical status and their implications on transcriptional activation of cancer signaling cascade that could possibly trigger numerous oncogenic players and finally decides the cells fate. Extract treated cells showed reduced viability with altered balance between oxidants and antioxidants which lead to redox status and which is known to distort various biological processes within the cell system. Results of RT-PCR demonstrated decreased expression of BCl2, cell cycle regulators along with Activator Protein −1 (AP-1) components. While Bax, p16 and p21 mRNAs showed increased expression in extract treated KB cells. Likewise, the translational levels of proliferation cell nuclear antigen (PCNA), tumor suppressor p53, retinoblastoma (Rb) and cyclin dependent kinase 4 (CDK4) were decreased along with AP-1 subunits (c-Jun/c-Fos) with increased protein levels of p21 in extract treated KB cells. Further, the downstream activation and regulation of AP-1 transcription factors could be through stress activated c-Jun – N terminal Kinase (JNK) Mitogen Activated Protein Kinases (MAPKs) which downregulated both Jun and Fos mRNA transcripts in areca nut extract exposed KB cells. Thus, outcome of the study provides insights into mechanistic path of pathogenesis of areca related disorders. Further, it could aid in designing new therapeutic modalities that specific targets these oncogenic players and help in disease management.