Gingival Epithelium Cells Research Articles

Caspase-3 and gasdermin E detection in peri-implantitis

Peri-implantitis could lead to progressive bone loss and implant failure; however, the mechanism of peri-implantitis remains unclear. Based on emerging evidence, pyroptosis, a novel proinflammatory programmed death, contributes to different oral infectious diseases. In the present study, we investigated the involvement of cleaved caspase-3 and gasdermin E (GSDME) in peri-implantitis and established a pyroptosis model in vitro. By collecting and examining the inflamed biopsies around peri-implantitis, we found that the pyroptosis-related markers (caspase-3, GSDME, and IL-1β) were enhanced relative to levels in control individuals. Furthermore, human gingival epithelium cells (HGECs) induced by tumor necrosis factor-α (TNF-α) exhibited pyroptosis morphological changes (cell swelling and balloon-shaped bubbles) and upregulated expression of pyroptosis-related markers. Pretreated with Ac-DEVD-CHO (a caspase-3 inhibitor) or GSDME small interference RNA (siRNA) were found to attenuate pyroptosis in HGECs. In conclusion, our findings revealed a high expression of caspase-3 and GSDME in the inflamed biopsies of peri-implantitis and confirmed that the caspase-3/GSDME pathway mediates TNF-α-triggered pyroptosis in human gingival epithelium cells, which provides a new target for peri-implantitis treatment.

Influence of magnetized abutment on biological seal of the implant–soft tissue interface by reconstructed human gingiva (RHG) model

Background: osseointegration in implants have received a lot of attention. Recently, the focus has extended towards the development of abutment materials supporting gingival soft tissue attachment. The attachment of the soft tissue to the implant/abutment surface is necessary to form a biological seal that protects the pathogenic microbial colonization which can lead to periimplantitis and bone resorption culminating in dental implant failure. Aim/Hypothesis: To evaluate the Influence of magnetized abutment surface on biological seal of the implant–soft tissue interface that would enable junctional epithelium(JE) attachment to the abutment through an internal basal lamina and hemidesmosomes in order to prevent the JE from migrating too in depth. Materials and Methods: We developed novel magnetic nanoparticles (MNs) coated with chitosan(CS) (CS-MNs) for enhancing cellular migration using magnetic force. CS-MNs were characterized by XRD, UV, SEM. The CS-MNs where spherical shape with a diameter from 10 nm to 15 nm. The magnetized abutments surface was treated with CS-MNs. Influence of CS-MNs treated abutment on reconstructed human gingiva (RHG) model consisting of differentiated gingival epithelium cells was assessed in terms of Epithelial attachment, down-growth along the abutment surface, quantitative analysis by permeability and cell attachment tests. Phenotype via histomorphology, scanning electron microscopy, and immunohistochemistry 10 days after implantation. Results: Abutment values obtained from the intraexperiment replicates including measurement of the left and right side of the abutment (total of four measurements) were first averaged, and Differences were determined using one-way ANOVA followed by Dunnett's uncorrected multiple comparisons test. Differences were considered significant at P < 0.05. histomorphometric analyses showed epithelial growth near perpendicular to the surface of abutments. The magnification clearly showed individual keratinocytes spread and attached to abutment surfaces via filopodia extensions. Conclusions and Clinical Implications: The current RHG model shows the active migration of epithelial cells near perpendicularly along the magnetized abutment surface with higher gingival fibroblast attachment as compared with an untreated surface. This novel system is thought to be useful in the development of biological seal around the implant and cell-based strategies for the repair or replacement of tissue and other novel therapies. Keywords: Soft tissue-implant interactions, biological seal, tissue-engineered oral mucosa, three-dimensional in vitro model

Immunomodulation‐Based Strategy for Improving Soft Tissue and Metal Implant Integration and Its Implications in the Development of Metal Soft Tissue Materials

AbstractThe difficulties associated with metal implants and soft tissue integration have significantly affected the applications of metal implants in soft‐tissue‐related areas. Prompted by the close association between soft tissue integration and the immune response, an immunomodulation‐based strategy is proposed to manipulate the immune microenvironment and improve metal implant–soft tissue integration. Considering their vital roles in soft tissue responses to metal implants, macrophages are used and the cytokines fingerprints of M1 and M2 macrophage immune microenvironments are evaluated for their potential modulatory effects on metal implant–soft tissue integration. The modulatory effects of different immune microenvironments on model soft tissue cells (human gingival epithelium cells) cultured on model metal implants (titanium alloy disks) are then described, with the underlying possible mechanism FAK‐AKT‐mTOR signaling unveiled. As further proof of concept, IL‐4/PDA (polydopamine)‐coated titanium alloy implants, aiming at modulating M2 macrophage polarization, are prepared and found to improve the in vivo metal implant‐soft tissue integration. It is the authors' ambition that this immunomodulation‐based strategy will change the negative perception and encourage the active development of metal materials with favorable soft tissue integration properties, thus improving the success rates of perforating metal implants and broadening their application in soft‐tissue‐related areas.

Evaluation of a novel oral mucosa in vitro implantation model for analysis of molecular interactions with dental abutment surfaces.

BackgroundAbutment surfaces are being designed to promote gingival soft tissue attachment and integration. This forms a seal around prosthetics and consequently ensures long‐term implant survival. New scalable and reproducible models are necessary to evaluate and quantify the performance of these surfaces.PurposeTo evaluate a novel implantation model by histomorphometric and immunohistochemical characterization of the interactions between human oral gingival tissue and titanium abutments with either novel anodized or conventional machined surface.Materials and MethodsAbutments were inserted into an organotypic reconstructed human gingiva (RHG) model consisting of differentiated gingival epithelium cells on a fibroblast populated lamina propria hydrogel following a tissue punch. Epithelial attachment, down‐growth along the abutment surface, and phenotype were assessed via histomorphology, scanning electron microscopy, and immunohistochemistry 10 days after implantation.ResultsThe down‐growing epithelium transitioned from a gingival margin to a sulcular and junctional epithelium. The sulcus depth and junctional epithelial length were similar to previously reported pre‐clinical and clinical lengths. A collagen IV/laminin 5 basement membrane formed between the epithelium and the underlying connective tissue. The RHG expanded in thickness approximately 2‐fold at the abutment surface. The model allowed the evaluation of protein expression of adhering soft tissue cells for both tested abutments.ConclusionsThe RHG model is the first in vitro 3D model to enable the assessment of not only human epithelial tissue attachment to dental abutments but also the expression of protein markers involved in soft tissue attachment and integration. The two abutments showed no noticeable difference in epithelial attachment.

Immunohistochemical Analysis of S100-Positive Langerhans Cells in the Healthy Gingiva and Periodontal Disease

Langerhans cells are antigen presenting cells, located in the stratified squamous epithelium of the skin and oral epithelium. A high density of Langerhans cells in gingival epithelium associated with an inflammatory process has been demonstrated in the literature by numerous studies, although the relationship with inflammation was less investigated.The aim of our study was to evaluate the distribution, density, morphology of Langerhans cells in both gingival epithelium and the associated inflammatory infiltrate in the lamina propria of the gingiva. The relationship between Langerhans cells and inflamatory process was also assesed. The present study included a number of 51 gingival biopsies. 12 of these didn� t show significant changes on routine examination. In all of the other 39 cases the presence of inflammatory lesions was noticed. The inflamatory infiltrate was described by using hematoxylin eosin staining and quantitative score, with values between 0 and 3. Langerhans cells were immunohistochemicaly highlighted by using S100 antibody and quantified with hot spot method.We found an increase of Langerhans cells density in all cases of inflammation as compared with healthy gingival tissue. The highest density was found in the 13 cases with severe inflammation. At these last cases was noticed a migration tendency of Langerhans cells from infiltrate inflamatory area to basal layer of the epithelium. The results of this study indicated that Langerhans cells number is higher in inflamatory gingival tissue comparative with normal tissue, and the density of these cells is in relation with the severity of the inflammatory process.

Antifungal effect of tissue conditioners containing poly(acryloyloxyethyltrimethyl ammonium chloride)-grafted chitosan on Candida albicans growth in vitro

Background/purposeDenture stomatitis is a pathological condition affecting the mucosa underneath ill-fitting dentures, and Candida albicans is considered its main etiologic factor. Tissue conditioners are temporary lining materials often applied to dentures to treat inflamed tissues. However, tissue conditioners do not exert antifungal activity, and the soft surface texture harbors C. albicans easily. The aim of this study was to examine the antifungal activity of tissue conditioners modified with chitosan (CS) or a quaternized chitosan (QCS), which was synthesized by grafting 2-[(acryloyloxy)ethyl] trimethyl ammonium chloride onto CS. Materials and methodsTissue conditioners containing varying weight percentages of CS or QCS were prepared as experimental discs 10 mm in diameter and 1 mm in thickness. Samples were co-cultured with C. albicans and the number of colony forming units was recorded. Other evaluations included cell toxicity and tensile bond strength to the resin denture base. ResultsIt was found significantly fewer fungal colonies in tissue conditioners modified with CS or QCS, and none when the weight percentage of QCS exceeded 5%. CS and QCS did not affect the viability of human gingival epithelium cells or fibroblasts, and tensile bond strength did not differ between control and modified tissue conditioners. ConclusionThis study provides a foundation for the development of QCS as a novel and safe antifungal agent applied to tissue conditioners in clinical practice.

Rapid HPLC of Cyanidin and Delphinidin of an Anthocyanin Complex Exposed to Human Gingival Epithelial Cells

A rapid isocratic HPLC was developed and validated for use in simultaneous analysis of cyanidin and delphinidin extracted from purple cobs of Zea mays L. ceritina Kulesh. (CC), blue petals of Clitoria ternatea L. (CT) and an anthocyanin complex (AC). The method was shown to be rapid, precise and accurate within 5 20 μg/ml (r &gt; 0.997) with limits of detection and quantitation of 0.45 and 1.52 μg/ml for cyanidin and 4.04 and 13.3 μg/ml for delphinidin, respectively. It could quantitatively detect and compare changes in cyanidin and delphinidin from the AC exposed to human gingival epithelium cells.

DNA methylation levels of SOCS1 and LINE-1 in oral epithelial cells from aggressive periodontitis patients

ObjectiveDNA methylation has been shown to be critical in the regulation of inflammatory genes. Infections are able to trigger susceptibility to disease and it can be considered as potential epimutagenic factors in reshaping the epigenome. Therefore, what would be the DNA methylation status in cells present in an infected and inflamed oral environment? The aim was to verify the DNA methylation pattern in oral epithelium cells from aggressive periodontitis (AgP) patients in a specific gene involved in the inflammation control, as suppressor of cytokine signalling (SOCS)1 and in a broader way through long interspersed nuclear element (LINE)-1. DesignGenomic DNA from oral cells of 30 generalized AgP patients and 30 healthy patients were purified and modified by sodium bisulfite. DNA methylation patterns were analyzed using combined bisulfite restriction analysis (COBRA) for SOCS1 and LINE-1. ResultsAn overall scenario of demethylation was seen for both groups, whereas the healthy group presented a higher percentage of demethylation (p<0.001), also presenting the majority of total demethylated samples (83.3% versus 70.8% in the AgP group). Total LINE-1 methylation or at each specific loci presented significant differences amongst groups. ConclusionEpithelial cells, present in an infected and inflamed oral environment, show different DNA methylation status from those present in a healthy oral environment, regarding the SOCS1 and LINE-1. In addition, the investigation allows detecting alterations in the DNA in a non-limited manner, since the results observed might reflect a generalized condition of the oral epithelial cells, besides reflecting the condition of the gingival epithelium cells.

Biophysical studies of the gingival epithelium

Objectives: As the gingival epithelial cells undergo transition between the surface, crevicular and junctional areas, it is reported that their resting potential could also get altered due to change/variations in the membrane transport protein composition. Hence, the purpose of the present study is (i) to study the biophysical properties of the surface, the crevicular, and the junctional epithelial gingival cells and to assess their implications to the cell to cell and the cell to tooth attachment mechanisms; (ii) to study the effect of certain delipidifying agents such as Sodium Deoxycholate (DOC) and Sodium Dodecyl Sulfate (SDS) on the transmembrane electrical activity of the gingival epithelial cells; and, (iii) to study the histological changes due to these treatments and assess their bearings on the biophysical observations. Materials and Methods: The biophysical and histological investigations on the healthy human gingival epithelium were carried out on the fresh biopsy material obtained from a homogeneous group of willing donors between 11-15 years of age, who were advised for extraction of first premolar due to orthodontic reasons. The biophysical measurement involved recording of the transmembrane potentials using glass ultra-microelectrodes. For histologic studies, the hematoxylin and eosin (H&E) staining of the tissue sections was performed. Results: The mean values of the membrane potential in the three types of the gingival epithelium cells vary appreciably. While it was lowest in the junctional cells (2.83 ± 0.98 mV), it was highest in the surface epithelial cells (22.96 ± 5.19 mV). The crevicular cells showed a value greater than the junctional cells but lesser than the surface cells (9.3 ± 1.73 mV). Conclusion: The membrane transport protein density appears to decrease in the following order: surface > crevicular > junctional cells. The crosslinking force of calcium (Ca 2+ ) ions with their increased magnitude from the junctional epithelial cells to the cementum or enamel of the tooth appears to be the most convincing model of the epithelial attachment at the Dentoenamel (DE) junction.

Higher density of label-retaining cells in gingival epithelium

Epithelial sheets made of cultured oral mucosal epithelial cells have been used to treat various epithelial defects [1]. These epithelial sheets are derived from oral epithelial stem cells exhibiting high proliferation potential. In the skin, research into the keratinocyte stem cell niche has advanced remarkably. However, the oral mucosal stem cell niche is less well understood. Thus, we investigated the density of label-retaining cells (LRC) in different areas of oral mucosa to elucidate the localization of oral mucosal stem cells.

Expression of Integrins in Human Gingiva

The distribution of the alpha 1-alpha 6 subunits of beta 1 integrins was studied by using a panel of monoclonal antibodies in indirect immunofluorescence microscopy. The results showed that the beta 1 subunit was expressed at the cell membrane of basal cells of gingival epithelium, throughout the cells of junctional epithelium (JE), and in cells of connective tissue, including endothelial cells and, more faintly, in inflammatory cells in gingival connective tissue. The alpha 4 subunit was expressed selectively in inflammatory cells, and the alpha 5 subunit was expressed in cells throughout gingival connective tissue. An overall cell membrane immunoreactivity for the alpha 2 and alpha 3 subunits was shown in basal cells of gingival epithelium and in cells of JE, corresponding to the epithelial localization of the beta 1 subunit. The alpha 6 subunit was polarized to the basal aspects of basal epithelial cells, but was also present in an overall cell surface distribution in basal cells and in cells of JE. The beta 4 integrin subunit was mainly expressed at the basal aspects of basal cells in gingival epithelium and JE. The results indicate that the alpha 2/beta 1, alpha 3/beta 1, alpha 6/beta 1, and alpha 6/beta 4 integrins are all expressed in human gingival epithelium. Of these, the alpha 6/beta 4 integrin complex is the major candidate for mediation of the attachment of epithelial cells to the basement membrane facing the connective tissue and probably also the tooth.

Interleukin-1 modulates T6 expression on a putative intra-epithelial Langerhans cell precursor population.

T6 is an antigen which is a highly specific marker for Langerhans cells. Previous studies have demonstrated that Interleukin-1 (IL-1) and an IL-1 inhibitor (ILS) modulate T6 expression (T6E) in explant culture. The present study examined the effects of IL-1 and ILS on T6E in dispersed gingival epithelial cell (EC) cultures. EC were obtained by trypsinization of gingival fragments obtained during periodontal surgery. T6E by EC was demonstrated using OKT6 monoclonal antibody in an immuno-peroxidase technique. In both unseparated and T6-depleted EC, IL-1 (0.5 U/mL) stimulated T6E, and this effect was abrogated by ILS (1:30). ILS alone depressed T6E in unseparated EC cultures. All effects were consistent between four- and 24-hour culture periods, and no treatment affected EC viability, thus excluding cell proliferation or necrosis as a vector for the action of IL-1 and ILS. These results indicate that a population of epithelial cells exists which is induced to express T6 under the influence of IL-1. IL-1 and ILS act in combination to regulate T6E on these precursor cells and on DR(-) Langerhans cells in gingival epithelium.