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Extracellular vesicles released from cells regulate many normal and pathological conditions. Little is known about the role of epidermal keratinocyte microvesicles (KC-MVs) in epithelial–stromal interaction that is essential for wound healing. We investigated, therefore, whether MV-like structures are present in human wounds and whether they affect wound healing–associated gene expression in dermal fibroblasts. In human wounds, MV-like vesicles were observed during active epithelial migration and early granulation tissue formation. When KC-MVs derived from keratinocyte-like cells (HaCaT) were added to fibroblast cultures, expression of 21 genes was significantly regulated (P<0.05) out of 80 genes investigated, including matrix metalloproteinase-1 and -3, interleukin-6 and -8, and genes associated with transforming growth factor-β signaling. Similar changes were observed at the protein level. MVs from normal epidermal keratinocytes showed similar response to HaCaT cells. KC-MVs activated ERK1/2, JNK, Smad, and p38 signaling pathways in fibroblasts with ERK1/2 signaling having the most prominent role in the MV-induced gene expression changes. KC-MVs stimulated fibroblast migration and induced fibroblast-mediated endothelial tube formation but did not affect collagen gel contraction by fibroblasts. The results demonstrate that keratinocyte microvesicles have a strong and a specific regulatory effect on fibroblasts that may modulate several aspects of wound healing.

Soft tissue calcification occurs in many parts of the body, including the gingival tissue. Epithelial cell-derived MVs can control many functions in fibroblasts but their role in regulating mineralization has not been explored. We hypothesized that microvesicles (MVs) derived from gingival epithelial cells could regulate calcification of gingival fibroblast cultures in osteogenic environment. Human gingival fibroblasts (HGFs) were cultured in osteogenic differentiation medium with or without human gingival epithelial cell-derived MV stimulation. Mineralization of the cultures, localization of the MVs and mineral deposits in the HGF cultures were assessed. Gene expression changes associated with MV exposure were analyzed using gene expression profiling and real-time qPCR. Within a week of exposure, epithelial MVs stimulated robust mineralization of HGF cultures that was further enhanced by four weeks. The MVs taken up by the HGF's did not calcify themselves but induced intracellular accumulation of minerals. HGF gene expression profiling after short exposure to MVs demonstrated relative dominance of inflammation-related genes that showed increases in gene expression. In later cultures, OSX , BSP and MMPs were significantly upregulated by the MVs. These results suggest for the first time that epithelial cells maybe associated with the ectopic mineralization process often observed in the soft tissues.

Periodontal diseases manifest by the formation of deep pockets between the gingiva and teeth where multispecies bacterial biofilms flourish, causing inflammation and bone loss. Epithelial cell receptor αvβ6 integrin that regulates inflammation by activating the anti-inflammatory cytokine transforming growth factor-β1, is highly expressed in healthy junctional epithelium that connects the gingiva to the tooth enamel. However, its expression is attenuated in human periodontal disease. Moreover, Itgb6 −/− mice display increased periodontal inflammation compared to wild-type mice. We hypothesized that bacterial biofilms present in the periodontal pockets suppress αvβ6 integrin levels in periodontal disease and that this change aggravates inflammation. To this end, we generated three-week-old multi-species oral biofilms in vitro and treated cultured gingival epithelial cells (GECs) with their extracts. The biofilm extracts caused suppression of β6 integrin expression and upregulation of pro-inflammatory cytokines, including interleukin-1β and -6. Furthermore, GECs with β6 integrin siRNA knockdown showed increased interleukin-1β expression, indicating that αvβ6 integrin-deficiency is associated with pro-inflammatory cytokine responsiveness. FSL-1, a synthetic bacterial lipopeptide, also suppressed β6 integrin expression in GECs. Therefore, biofilm components, including lipopeptides, may downregulate αvβ6 integrin expression in the pocket epithelium and thus promote epithelial cell-driven pro-inflammatory response in periodontal disease.

Cancer-testis (CT) antigens are predominantly expressed in testis or placenta, but absent in most adult tissues. During malignant transformation CT genes are often activated. CT antigen 16 (CT16, PAGE5) is frequently expressed in advanced melanoma but its biological function has been unknown. To examine the role of CT16 in cell survival we knocked it down in A2058 melanoma cells using specific siRNAs and exposed the cells to cancer drug cisplatin known to induce apoptosis. As a result, cell survival was markedly decreased. To study the effects of CT16 on cell survival in more detail, the cellular gene expression profiles were investigated after CT16 silencing in CT16 positive A2058 melanoma cells, as well as after CT16 overexpression in CT16 negative WM-266-4 melanoma cells. Among the 11 genes both upregulated by CT16 silencing and downregulated by CT16 overexpression or vice versa, 4 genes were potentially apoptotic or antiapoptotic genes. CT16 was recognized as a positive regulator of antiapoptotic metallothionein 2A and interleukin 8 genes, whereas it inhibited the expression of apoptosis inducing dickkopf 1 (DKK1) gene. In addition CT16 enhanced the expression of fatty acid binding protein 7, a known promoter of melanoma progression. The effect of CT16 on DKK1 expression was p53 independent. Furthermore, CT16 did not regulate apoptotic genes via DNA methylation. In twenty melanoma metastasis tissue samples average DKK1 mRNA level was shown to be significantly (p<0.05) lower in high CT16 expressing tumors (n = 3) when compared to the tumors with low CT16 expression (n = 17). Thus, our results indicate that CT16 promotes the survival of melanoma cells and is therefore a potential target for future drug development.

Objectives Decontamination of biofilm‐colonized rough implant surfaces remains challenging. We investigated the effect of airflow with glycine powder (AFG) on decontamination of mature oral multispecies biofilm from a sandblasted and acid etched (SLA) titanium surface. Materials and Methods Subgingival dental plaque was cultured on SLA disks anaerobically for 21 days. AFG with various settings and distances was applied directly on the disks with or without previous rinse of 0.9% NaCl. The specimens were then analyzed through scanning electron microscope and remaining bacteria on the implant surface were quantified and statistically compared. Results Mature oral biofilm with cocci and rods as major morphotypes, as well as spiral‐ and filamentous‐shaped organisms, was formed on the untreated disks. Saline rinsing removed the thick biofilm layer but left numerous of coccoid bacteria in rough surface pits. AFG effectively removed most of the bacteria from the pits. Both 25% and 50% power settings were equally effective at 3‐mm distance. With 50% power, AFG successfully removed bacteria at both 3‐ and 6‐mm distance. When AFG was applied on native biofilm without prior rinsing with saline, it effectively removed the biofilm including bacteria in the pits. Conclusion Application of AFG appears effective in removing bacteria from rough implant surfaces.

Objectives Peri‐implantitis (PI) is caused by bacteria in the peri‐implant space but the consensus on microbial profile is still lacking. Current microbial sampling of PI lesions has largely focused on analyzing bacterial species that have been shed from the implant surface and captured in the pocket fluid. The purpose of the present study was to investigate the morphotypes of bacteria in biofilm covering the implant threads and explore whether certain morphotypes were associated with PI. Methods Fourteen failed implants were removed and instantly processed for scanning electron microscope analysis. The implants were imaged at three equally divided sub‐crestal levels of the exposed area. Bacterial morphotypes were identified and quantified by three examiners. Mobility and years in function were correlated to the presence of different morphotypes. Results The implants demonstrated the presence of variable bacterial morphotypes that did not correlate to disease progression in our study. Some implants were dominated by filaments and others showed the presence of combinations of cocci/rods or spirilles/spirochetes. In general, all implants showed variable morphologic biofilm composition. However, individual implants tended to have similar composition throughout the entire implant. Rods and filaments were dominant morphotypes throughout the surfaces and cocci showed increased presence toward the apical third. There were some differences in the biofilm morphology with mobility and time in function. Conclusions The profiles of bacterial biofilm morphotypes in failing implants with similar clinical presentations were highly variable. While there were significant differences between implants, similar morphotypes in individual implants were often found throughout the entire surface.

Integrin αvβ6 is an epithelial-specific receptor that binds and activates latent transforming growth factor-β1 (TGF-β1). TGF-β1 has been implicated as an endogenous inducer of hair follicle (HF) regression during hair cycling. We hypothesized that αvβ6 integrin–mediated TGF-β1 signaling regulates hair regeneration and HF involution. In wild-type (WT) mice, the expression of integrin αvβ6 was strongly upregulated in the outer root sheath (ORS) during early hair regeneration, and was specifically enhanced in the HF bulge region. Expression gradually decreased in late anagen and remained restricted to the bulge region in the catagen and telogen stage HFs. The first spontaneous hair cycle was not altered in β6 integrin knockout (β6−/−) mice. However, after depilation, β6−/− mice exhibited retarded HF regression compared with WT controls. β6−/− follicles contained significantly higher numbers of proliferating Ki67-positive keratinocytes than WT follicles at an identical cycle stage. The β6−/− follicles also demonstrated significantly reduced levels of TGF-β1 expression and Smad2 phosphorylation during early anagen and anagen–catagen transition. Our study indicates that αvβ6 integrin has an important inhibitory role in keratinocyte proliferation in both HFs and interfollicular epidermis. Thus, downregulated TGF-β1 signaling in β6−/− mice may affect bulge niche stem cell behavior.

Desmosomes are a complex assembly of protein molecules that mediate adhesion between adjacent cells. Desmosome composition is well established and spatial relationships between components have been identified. Intercellular cell-cell adhesion is created by the interaction of extracellular domains of desmosomal cadherins, namely, desmocollins and desmogleins. High-resolution methods have provided insight into the structural interactions between cadherins. However, there is a lack of understanding about the architecture of the intact desmosomes and the physical principles behind their adhesive strength are unclear. Electron Tomography (ET) studies have offered three-dimensional visual data of desmosomal cadherin associations at molecular resolution. This review discusses the merits of two cadherin association models represented using ET. We discuss the possible role of sample preparation on the structural differences seen between models and the possibility of adaptive changes in the structure as a direct consequence of mechanical stress and stratification.

Exposure to airborne allergens from mice and rats can lead to laboratory animal asthma or allergy. Several biological methods can measure allergens contained in aerosols; however, they are time and cost intensive. An innovative methodology is proposed to warn laboratory animal facility workers of a possible rise in mouse and rat allergens by measuring the relationships between airborne allergens, particulate matter (PM), and volatile organic compounds (VOCs). By using a low-cost sensor (average difference respect to reference methods of 3 and 9% for PM 2.5 and VOC, respectively), Spearman’s rank correlation between allergens and time-averaged PM and VOCs was found to be 0.3 and − 0.07, respectively. These numbers indicate a poor correlation between allergens and PM 2.5 and VOC; however, by considering only the spikes in PM minute-by-minute data, the relation between time-average PM and allergens increases up to 0.71. This high value indicates the applicability of PM low-cost sensors in laboratory animal centers as a warning sign of raising values of allergens. Mouse and rat allergens are present in the animals’ urine, which can become aerosolized during animal activity, or task activities carried out in the laboratory facility. While previous references established a correlation between activities and mouse and rat allergens, the results are outdated and refer to a limited number of activities. For example, washing lab coats, changing uniforms, sitting in an office space, having lunch, or walking in any corridor are shown in this study to contain on average 1.77, 0.96, 0.65, 0.88, and 1.62 ng/m 3 of rat allergens, respectively. Thus, locations that do not contain any direct source of allergens are positive to the presence of mouse and rat allergens.

A reconsideration of Sir Arthur Wilson's plan for war with Germany is presented. OA