Formation Of Hemidesmosomes Research Articles

Gingival keratinocyte adhesion on atomic layer-deposited hydroxyapatite coated titanium.

This study aimed to evaluate the effects of the atomic layer deposited hydroxyapatite (ALD-HA) coating of the titanium (Ti) surface on human gingival keratinocyte (HGK) cell adhesion, spreading, viability, and hemidesmosome (HD) formation. Grade 2 square-shaped Ti substrates were used (n = 62). Half of the substrates were ALD-HA coated, while the other half were used as non-coated controls (NC). The ALD-HA surface was characterized with scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDS) analysis. The initial cell adhesion and HD formation of HGKs were evaluated after a 24-h cultivation period. The cell proliferation was assessed by cultivating cells for 1, 3, and 7d. The expression levels of the integrin mediating cell adhesion were detected with the Western Blot method. In addition, cell spreading and expression of the proteins mediating cell adhesion were imaged using a confocal microscope. SEM-EDS analysis demonstrated the formation of HA on the ALD-HA surfaces. The relative cell attachment was significantly higher (p < .05) on the ALD-HA compared to the NC surface after 1 and 3 d of cell culture. No significant difference was found in integrin α6 or β4 expression. The microscope evaluation showed significantly increased cell spreading with peripheral HD expression on ALD-HA compared to the NC surfaces (p = .0001). Moreover, laminin γ2 expression was significantly higher on the ALD-HA than on the NC surfaces (p < .001). Compared to the NC Ti surface, the ALD-HA coating has favorable effects on HGK proliferation, growth, and cell spreading. This indicates that the ALD-HA coating has good potential for improving mucosal attachment on implant surfaces.

Endogenous chondroitin extends lifespan by inhibiting VHA-7-mediated tubular lysosome formation

Chondroitin extends lifespan and healthspan in C. elegans, but the relationship between extracellular chondroitin and intracellular anti-aging mechanisms is unknown. The basement membrane (BM) that contains chondroitin proteoglycans is anchored to cells via hemidesmosomes (HDs), and it accumulates damage with aging. In this study, we found that chondroitin regulates aging through the formation of HDs and inhibition of tubular lysosomes (TLs). Reduction of chondroitin due to a mutation in sqv-5/Chondroitin synthase (ChSy) causes the earlier and excessive formation of TLs and leakage of the lysosomal nuclease in a manner dependent on VHA-7, the a-subunit of V-type ATPase. VHA-7, whose mutation suppresses the short lifespan of the sqv-5 mutant, is initially localized to the basal side of the hypodermal cells and transported to lysosomes with aging. These results demonstrate that endogenous chondroitin suppresses aging by inhibiting the earlier excessive formation of TLs. This is a novel anti-aging mechanism that is controlled by the BM.

Radiofrequency Treatment Attenuates Age-Related Changes in Dermal-Epidermal Junctions of Animal Skin.

The dermal-epidermal junction (DEJ) is essential for maintaining skin structural integrity and regulating cell survival and proliferation. Thus, DEJ rejuvenation is key for skin revitalization, particularly in age-related DEJ deterioration. Radiofrequency (RF) treatment, known for its ability to enhance collagen fiber production through thermal mechanisms and increase heat shock protein (HSP) expression, has emerged as a promising method for skin rejuvenation. Additionally, RF activates Piezo1, an ion channel implicated in macrophage polarization toward an M2 phenotype and enhanced TGF-β production. This study investigated the impact of RF treatment on HSP47 and HSP90 expression, known stimulators of DEJ protein expression. Furthermore, using in vitro and aged animal skin models, we assessed whether RF-induced Piezo1 activation and the subsequent M2 polarization could counter age-related DEJ changes. The RF treatment of H2O2-induced senescent keratinocytes upregulated the expression of HSP47, HSP90, TGF-β, and DEJ proteins, including collagen XVII. Similarly, the RF treatment of senescent macrophages increased Piezo1 and CD206 (M2 marker) expression. Conditioned media from RF-treated senescent macrophages enhanced the expression of TGF-β and DEJ proteins, such as nidogen and collagen IV, in senescent fibroblasts. In aged animal skin, RF treatment increased the expression of HSP47, HSP90, Piezo1, markers associated with M2 polarization, IL-10, and TGF-β. Additionally, RF treatment enhanced DEJ protein expression. Moreover, RF reduced lamina densa replication, disrupted lesions, promoted hemidesmosome formation, and increased epidermal thickness. Overall, RF treatment effectively enhanced DEJ protein expression and mitigated age-related DEJ structural changes by increasing HSP levels and activating Piezo1.

Fluoride releasing photothermal responsive TiO2 matrices for antibiosis, biosealing and bone regeneration

Peri-implantitis induced by infection leads to gingival recession, alveolar resorption and eventual dental implant failure. So, antibiosis and biosealing of abutments as well as osseointegration of roots need to be projected seriously during the whole service lifespan of dental implants. In this work, a multipurpose photothermal therapy strategy based on Si/P/F doped TiO2 matrix is proposed to address the above issues. This TiO2 matrix not only has outstanding photothermal response, but also triggers the release of F ions under near-infrared (NIR) light irradiation. Local hyperthermia assisted with the released F ions reduces adenosine triphosphate (ATP) synthesis of staphylococcus aureus (S. aureus), increases bacterial membrane permeability, and induces abundant of reactive oxygen species, resulting in the oxidation of cellular components and eventual death of bacteria. Furthermore, the synergic action of mild photothermal stimulation and Si/P/F ions of TiO2 matrix up-regulates gingival epithelial cells behavior (e.g., hemidesmosome formation) and osteoblasts response in vitro. In an infected model, this TiO2 matrix obviously eliminates bacteria, reduces inflammatory response, improves epithelial sealing and osseointegration, and reduces alveolar resorption by regulating NIR irradiation.

CREB3L2 Regulates Hemidesmosome Formation during Epithelial Sealing.

The long-term success rate of dental implants can be improved by establishing a favorable biological sealing with a high-quality epithelial attachment. The application of mesenchymal stem cells (MSCs) holds promise for facilitating the soft tissue integration around implants, but the molecular mechanism is still unclear and the general application of MSC sheet for soft tissue integration is also relatively unexplored. We found that gingival tissue-derived MSC (GMSC) sheet treatment significantly promoted the expression of hemidesmosome (HD)-related genes and proteins in gingival epithelial cells (GECs). The formation of HDs played a key role in strengthening peri-implant epithelium (PIE) sealing. Further, high-throughput transcriptome sequencing showed that GMSC sheet significantly upregulated the PI3K/AKT pathway, confirming that cell adhesion and HD expression in GECs were regulated by GMSC sheet. We observed that the expression of transcription factor CREB3L2 in GECs was downregulated. After treatment with PI3K pathway inhibitor LY294002, CREB3L2 messenger RNA and protein expression levels were upregulated. Further experiments showed that overexpression or knockdown of CREB3L2 could significantly inhibit or promote HD-related genes and proteins, respectively. We confirmed that CREB3L2 was a transcription factor downstream of the PI3K/AKT pathway and participated in the formation of HDs regulated by GMSC sheet. Finally, through the establishment of early implant placement model in rats, we clarified the molecular function of CREB3L2 in PIE sealing as a mechanical transmission molecule in GECs. The application of GMSC sheet-implant complex could enhance the formation of HDs at the implant-PIE interface and decrease the penetration distance of horseradish peroxidase between the implant and PIE. Meanwhile, GMSC sheet reduced the length of CREB3L2 protein expression on PIE. These findings elucidate the potential function and molecular mechanism of MSC sheet regulating the epithelial sealing around implants, providing new insights and ideas for the application of stem cell therapy in regenerative medicine.

Keratinocyte Proliferation and Hemidesmosome Formation on Surfaces for Dental Implants: In Vitro Study.

To demonstrate the likelihood of the polyetheretherketone (PEEK), zirconia (ZrO2), and titanium (Ti) disks to support proliferation and hemidesmosome formation of gingival cells. Water contact angle was performed on each material, and surface roughness (Ra) was measured. Scanning electron microscopy and x-ray photoelectron spectroscopy were used. Later, oral keratinocyte cells were cultured on disks, and metabolic activity and expression of hemidesmosome markers, integrin α6 and β4, in relation to the biomaterial disks at 1, 3, and 5 days of cell culture were quantified. Tissue culture polystyrene was used as the control. Statistical analysis was performed with analysis of variance (ANOVA) with Tukey post hoc comparison test. A P value of < .05 was considered statistically significant. The water contact angle ranged from 70.2 degrees (Ti) to a maximum of hydrophobicity of 93.3 degrees (PEEK). Ra was highest on ZrO2, followed by PEEK. Ti showed the most keratinocyte metabolic activity at 1, 3, and 5 culture periods. Contrarily, ZrO2 and PEEK disks had lower keratinocyte metabolic activity at all observation times, with no statistical differences between both groups. Integrin α6 and β4 expression was highest on TCPS and ZrO2 compared to Ti and PEEK. Keratinocytes proliferated faster on Ti than on ZrO2 and PEEK substrates, and expression of hemidesmosome formation markers, integrin α6 and β4, were higher on ZrO2 than either Ti or PEEK. Int J Oral Maxillofac Implants 2023;38:496-502. doi: 10.11607/jomi.9894.

Anomaly of cornea and ocular adnexa in spinster homolog 2 (Spns2) knockout mice

Spinster 2 (Spns2) is a transporter that pumps sphingosine-1-phosphate (S1P), a bioactive lipid mediator synthesized in the cytoplasm, out of cells into the inter cellular space. S1P is a signal that modulates cellular behavior during embryonic development, inflammation and tissue repair, etc. A Spns2-null (KO) mouse is born with failure of eyelid closure (eyelid-open-at birth; EOB) and develop corneal fibrosis in adulthood. It remains elusive whether corneal lesion is caused by exposure to keratitis (lagophthalmos) of EOB phenotype or the loss of Spns2 directly perturbs the corneal tissue morphogenesis and intra-eyelid structures. Therefore, we investigated differences between the cornea and ocular adnexa morphogenesis in KO and wild-type (WT) embryos and adults as well.The loss of Spns2 perturbs cornea morphogenesis during embryonic development as early as E16.5 besides EOB phenotype. Histology showed that the corneal stroma was thinner with less extracellular matrix accumulation, e.g., collagen and keratocan in the KO mouse. Epithelial stratification, expression of keratin 12 and formation of desmosomes and hemidesmosomes were also perturbed in these KO corneas. Lacking Spns2 impaired morphogenesis of the Meibomian glands and of orbicularis oculi muscles. KO glands were labeled for ELOVL4 and PPARγ and were Oil-Red O-positive, suggesting KO acinar cells possessed functionality as the glands.This is the first report on the roles of Spns2 in corneal and Meibomian gland morphogenesis. Corneal tissue destruction in an adult KO mouse might be due to not only lagophthalmos but also to an impaired morphogenesis of cornea, Meibomian glands, and orbicularis oculi muscle.

Protein Ligand Nanopattern Size Selects for Cellular Adhesion via Hemidesmosomes over Focal Adhesions.

Hemidesmosomes (HDs) are multiprotein complexes that firmly anchor epidermal cells to the basement membrane of skin through the interconnection of the cytoplasmic intermediate filaments with extracellular laminin 332 (Ln332). Considerably less attention has been paid to HDs compared to focal complexes/focal adhesions (FC/FAs) in mechanistic single-cell structures due to the lack of suitable in vitro model systems. Here nanopatterns of Ln332 (100-1000nm) are created to direct and study the formation of HD in adherent HaCaT cells. It is observed that HaCaT cells at Ln 332 nanopatterns adhere via hemidesmosomes, in stark contrast to cells at homogeneous Ln332 surfaces that adhere via FC/FAs. Clustering of α6 integrin is observed at nanopatterned Ln332 of 300nm patches and larger. Cells at 500nm diameter patterns show strong colocalization of α6 integrin with ColXVII or pan-cytokeratin compared to 300nm/1000nm indicating a threshold for HD initiation >100nm but a pattern size selection for maturation of HDs. It is demonstrated that the pattern of Ln332 can determine the cellular selection of adhesion types with a size-dependent initiation and maturation of HDs. The protein nanopatterning approach that is presented provides a new in vitro route to study the role of HDs in cell signaling and function.

Pancreatic ductal adenocarcinoma cells employ integrin α6β4 to form hemidesmosomes and regulate cell proliferation

Pancreatic ductal adenocarcinoma (PDAC) has a dismal prognosis due to its aggressive progression, late detection and lack of druggable driver mutations, which often combine to result in unsuitability for surgical intervention. Together with activating mutations of the small GTPase KRas, which are found in over 90% of PDAC tumours, a contributory factor for PDAC tumour progression is formation of a rigid extracellular matrix (ECM) and associated desmoplasia. This response leads to aberrant integrin signalling, and accelerated proliferation and invasion. To identify the integrin adhesion systems that operate in PDAC, we analysed a range of pancreatic ductal epithelial cell models using 2D, 3D and organoid culture systems. Proteomic analysis of isolated integrin receptor complexes from human pancreatic ductal epithelial (HPDE) cells predominantly identified integrin α6β4 and hemidesmosome components, rather than classical focal adhesion components. Electron microscopy, together with immunofluorescence, confirmed the formation of hemidesmosomes by HPDE cells, both in 2D and 3D culture systems. Similar results were obtained for the human PDAC cell line, SUIT-2. Analysis of HPDE cell secreted proteins and cell-derived matrices (CDM) demonstrated that HPDE cells secrete a range of laminin subunits and form a hemidesmosome-specific, laminin 332-enriched ECM. Expression of mutant KRas (G12V) did not affect hemidesmosome composition or formation by HPDE cells. Cell-ECM contacts formed by mouse and human PDAC organoids were also assessed by electron microscopy. Organoids generated from both the PDAC KPC mouse model and human patient-derived PDAC tissue displayed features of acinar-ductal cell polarity, and hemidesmosomes were visible proximal to prominent basement membranes. Furthermore, electron microscopy identified hemidesmosomes in normal human pancreas. Depletion of integrin β4 reduced cell proliferation in both SUIT-2 and HPDE cells, reduced the number of SUIT-2 cells in S-phase, and induced G1 cell cycle arrest, suggesting a requirement for α6β4-mediated adhesion for cell cycle progression and growth. Taken together, these data suggest that laminin-binding adhesion mechanisms in general, and hemidesmosome-mediated adhesion in particular, may be under-appreciated in the context of PDAC.Proteomic data are available via ProteomeXchange with the identifiers PXD027803, PXD027823 and PXD027827.

Junctional epithelium and hemidesmosomes: Tape and rivets for solving the "percutaneous device dilemma" in dental and other permanent implants.

The percutaneous device dilemma describes etiological factors, centered around the disrupted epithelial tissue surrounding non-remodelable devices, that contribute to rampant percutaneous device infection. Natural percutaneous organs, in particular their extracellular matrix mediating the “device”/epithelium interface, serve as exquisite examples to inspire longer lasting long-term percutaneous device design. For example, the tooth's imperviousness to infection is mediated by the epithelium directly surrounding it, the junctional epithelium (JE). The hallmark feature of JE is formation of hemidesmosomes, cell/matrix adhesive structures that attach surrounding oral gingiva to the tooth's enamel through a basement membrane. Here, the authors survey the multifaceted functions of the JE, emphasizing the role of the matrix, with a particular focus on hemidesmosomes and their five main components. The authors highlight the known (and unknown) effects dental implant – as a model percutaneous device – placement has on JE regeneration and synthesize this information for application to other percutaneous devices. The authors conclude with a summary of bioengineering strategies aimed at solving the percutaneous device dilemma and invigorating greater collaboration between clinicians, bioengineers, and matrix biologists.

TNF-α regulates the composition of the basal lamina and cell-matrix adhesions in gingival epithelial cells

ABSTRACT Laminin 5, type 4 collagen, and α6β4 integrin contribute to the formation of hemidesmosomes in the epithelia of periodontal tissues, which is critical for the development and maintenance of the dentogingival junction. As it is not known whether TNF-α alters the composition of the epithelial pericellular matrix, human gingival epithelial cells were cultured in the presence or absence of TNF-α. Treatment with TNF-α accelerated epithelial cell migration and closure of in vitro wounds. These data indicate unexpectedly, that TNF-α promotes the formation of the pericellular matrix around epithelial cells and enhances adhesion of epithelial cells to the underlying matrix, properties which are important for cell migration and the integrity of the dentogingival junction.

Plectin ensures intestinal epithelial integrity and protects colon against colitis

Plectin, a highly versatile cytolinker protein, provides tissues with mechanical stability through the integration of intermediate filaments (IFs) with cell junctions. Here, we hypothesize that plectin-controlled cytoarchitecture is a critical determinant of the intestinal barrier function and homeostasis. Mice lacking plectin in an intestinal epithelial cell (IEC; PleΔIEC) spontaneously developed colitis characterized by extensive detachment of IECs from the basement membrane (BM), increased intestinal permeability, and inflammatory lesions. Moreover, plectin expression was reduced in the colons of ulcerative colitis (UC) patients and negatively correlated with the severity of colitis. Mechanistically, plectin deficiency in IECs led to aberrant keratin filament (KF) network organization and the formation of dysfunctional hemidesmosomes (HDs) and intercellular junctions. In addition, the hemidesmosomal α6β4 integrin (Itg) receptor showed attenuated association with KFs, and protein profiling revealed prominent downregulation of junctional constituents. Consistent with the effects of plectin loss in the intestinal epithelium, plectin-deficient IECs exhibited remarkably reduced mechanical stability and limited adhesion capacity in vitro. Feeding mice with a low-residue liquid diet that reduced mechanical stress and antibiotic treatment successfully mitigated epithelial damage in the PleΔIEC colon.

Participation of adhesion molecules in changing cell interactions during metastasis development

The review summarizes current information about the role and significance of adhesion molecules in the process of tumor metastasis. For example, different tumors is shown that changes in the expression of adhesion molecules leads to disruption of the regulatory interactions in the processes of proliferation, cytodifferentiation and migration of tumor cells, enables the cascade of pathological processes leading to the colonization of other tumor cells remote from the tumor bodies. The phenomenology of tumor development and metastasis is shown. The participation of the main groups of adhesion molecules (cadherins, integrins, selectins, immunoglobulins and white blood cell homing receptors) at the stages of metastasis is considered. A violation of cadherin expression is accompanied by a decrease in the expression of integrins involved in the formation of hemidesmosomes, while a cascade of reactions develops that contributes to the development of an aggressive malignant phenotype, cells acquire an increased ability to migrate and invade, resulting in the appearance of a tumor in secondary foci. The role of immunoglobulins in the development of metastatic process has been studied in less detail than in cadherins and integrins. It was found that immunoglobulins are involved in the formation of resistance of tumor cells to proapoptotic signals. Immunoglobulins increase the expression of metalloproteinase genes involved in the degradation of the extracellular matrix, which is a factor in the initiation of metastasis. A correlation between increased selectin expression and increased metastasis is shown. An increase in the expression level of white blood cell homing receptors leads to an increase in the invasive potential of tumors. The results of the study of cell adhesion proteins serve as a fundamental basis for the development of methods of antitumor therapy. Over the past decades, a number of immunohistochemical protein detection reactions have become one of the diagnostic methods of cancer clinics. However, the role of cell adhesion molecules in ensuring the metastasis process, as well as their significance in the prognosis of the development of the tumor process and antitumor therapy, needs to be further studied.

Keratinocyte-Specific Peptide-Based Surfaces for Hemidesmosome Upregulation and Prevention of Bacterial Colonization.

Percutaneous devices like orthopedic prosthetic implants for amputees, catheters, and dental implants suffer from high infection rates. A critical aspect mediating peri-implant infection of dental implants is the lack of a structural barrier between the soft tissue and the implant surface which could impede bacteria access and colonization of exposed implant surfaces. Parafunctional soft tissue regeneration around dental implants is marked by a lack of hemidesmosome formation and thereby weakened mechanical attachment. In response to this healthcare burden, a simultaneously hemidesmosome-inducing, antimicrobial, multifunctional implant surface was engineered. A designer antimicrobial peptide, GL13K, and a laminin-derived peptide, LamLG3, were coimmobilized with two different surface fractional areas. The coimmobilized peptide surfaces showed antibiofilm activity against Streptococcus gordonii while enhancing proliferation, hemidesmosome formation, and mechanical attachment of orally derived keratinocytes. Notably, the coatings demonstrated specific activation of keratinocytes: the coatings showed no effects on gingival fibroblasts which are known to impede the quality of soft tissue attachment to dental implants. These coatings demonstrated stability and retained activity against mechanical and thermochemical challenges, suggesting their intraoral durability. Overall, these multifunctional surfaces may be able to reduce peri-implantitis rates and enhance the success rates of all percutaneous devices via strong antimicrobial activity and enhanced soft tissue attachment to implants.

Surfce Functionalized via AdLAMA3 Multilayer Coating for Re-epithelization Around Titanium Implants.

The peri-implant epithelium (PIE) forms a crucial seal between the oral environment and the implant surface. Compared with the junctional epithelium (JE), the biological sealing of PIE is fragile, which lacks hemidesmosomes (HDs) and internal basal lamina (extracellular matrix containing laminin332, IBL) on the upper part of the interface. In the study, we aim to prepare a coating with good biocompatibility and ability to immobilize the recombinant adenovirus vector of LAMA3 (AdLAMA3) for promoting the re-epithelization of PIE. The titanium surface functionalized with AdLAMA3 was established via layer-by-layer assembly technique and antibody-antigen specific binding. The biological evaluations including cell adhesion and the re-epithelization of PIE were investigated. The results in vitro demonstrated that the AdLAMA3 coating could improve epithelial cell attachment and cell spreading in the early stage. In vivo experiments indicated that the AdLAMA3 coating on the implant surface has the potential to accelerate the healing of the PIE, and could promote the expression of laminin α3 and the formation of hemidesmosomes. This study might provide a novel approach and experimental evidence for the precise attachment of LAMA3 to titanium surfaces. The process could improve the re-epithelization of PIE.

Adenovirus-Mediated LAMA3 Transduction Enhances Hemidesmosome Formation and Periodontal Reattachment during Wound Healing

A robust dento-epithelial junction prevents external pathogenic factors from entering connective tissue and could be crucial for periodontal reattachment after periodontal surgery. The junctional epithelium (JE) is attached to the tooth surface through the hemidesmosome (HD) and internal basal lamina, where the primary component is laminin-332. Destruction of the JE leads to the loss of periodontal attachment. Traditional treatments are effective in eliminating local inflammation of the gingiva; however, few directly promote periodontal reattachment and HD formation. Here, we designed a gene-therapy strategy using the adenovirus-mediated human laminin-332 α3 chain (LAMA3) gene (Ad-LAMA3) transduced into a human-immortalized epidermal cell line (HaCaT) to study the formation of HD in vitro. Ad-LAMA3 promoted early adhesion and fast migration of HaCaT cells and increased expression of LAMA3 and type XVII collagen (BP180) significantly. Furthermore, HaCaT cells could facilitate formation of mature HDs after LAMA3 overexpression. In vivo experiments demonstrated that the JE transduced with Ad-LAMA3 could increase expression of LAMA3 and BP180 and “biological sealing” between the tooth and gingival epithelium. These results suggested that adenovirus-mediated LAMA3 transduction is a novel therapeutic strategy that promotes the stability and integration of the JE around the tooth during wound healing.

Surface Immobilization Chemistry of a Laminin-Derived Peptide Affects Keratinocyte Activity.

Many chemical routes have been proposed to immobilize peptides on biomedical device surfaces, and in particular, on dental implants to prevent peri-implantitis. While a number of factors affect peptide immobilization quality, an easily controllable factor is the chemistry used to immobilize peptides. These factors affect peptide chemoselectivity, orientation, etc., and ultimately control biological activity. Using many different physical and chemical routes for peptide coatings, previous research has intensely focused on immobilizing antimicrobial elements on dental implants to reduce infection rates. Alternatively, our strategy here is different and focused on promoting formation of a long-lasting biological seal between the soft tissue and the implant surface through transmembrane, cell adhesion structures called hemidesmosomes. For that purpose, we used a laminin-derived call adhesion peptide. However, the effect of different immobilization chemistries on cell adhesion peptide activity is vastly unexplored but likely critical. Here, we compared the physiochemical properties and biological responses of a hemidesmosome promoting peptide immobilized using silanization and copper-free click chemistry as a model system for cell adhesion peptides. Successful immobilization was confirmed with water contact angle and X-ray photoelectron spectroscopy. Peptide coatings were retained through 73 days of incubation in artificial saliva. Interestingly, the non-chemoselective immobilization route, silanization, resulted in significantly higher proliferation and hemidesmosome formation in oral keratinocytes compared to chemoselective click chemistry. Our results highlight that the most effective immobilization chemistry for optimal peptide activity is dependent on the specific system (substrate/peptide/cell/biological activity) under study. Overall, a better understanding of the effects immobilization chemistries have on cell adhesion peptide activity may lead to more efficacious coatings for biomedical devices.

Biology of soft tissue repair: gingival epithelium in wound healing and attachment to the tooth and abutment surface.

Epithelium attachment to the tooth or abutment surface is necessary to form a biological seal preventing pathogens and irritants from penetrating the body and reaching the underlying soft tissues and bone, which in turn can lead to inflammation and subsequent bone resorption. The present review investigated oral wound closure and the role of micro-environment, saliva, crevicular fluid and microbiota in wound healing. The importance of the junctional epithelium (peri-implant epithelium) attachment to the abutment surface was investigated. Current research focuses on macro-design, surface-topography, surface-chemistry, materials, coatings and wettability to enhance attachment, since these optimised surface properties are expected to promote keratinocyte attachment and spreading through hemi-desmosome formation. Detailed studies describing the extent of junctional epithelium attachment - e.g. barrier function, hemi-desmosomes, epithelium quality, composition of the external basement membrane or ability of the epithelium to resist microbial penetration and colonisation - are not yet reported in animals due to ethical considerations, scalability, expense, technical challenges and limited availability of antibodies. In vitro studies generally include relatively simple 2D culture models, which lack the complexity required to draw relevant conclusions. Additionally, human organotypic 3D mucosa models are being developed. The present review concluded that more research using these organotypic mucosa models may identify relevant parameters involved in soft-tissue-abutment interactions, which could be used to study different macro-shapes and surface modifications. Such studies would bridge the gap between clinical, animal and traditional in vitro cell culture studies supporting development of abutments aiming at improved clinical performance.

631 Laminin-511 is a key component of epidermal basement membrane to maintain epidermal homeostasis

Daily exposure to sunlight damages the epidermal basement membrane (BM) and disrupts epidermal homeostasis. However, the role of epidermal BM components in maintaining epidermal homeostasis is unclear. Here, we investigated the effect of sun exposure on BM components and epidermal homeostasis. We found that laminin-511 in epidermal BM is less in sun-exposed skin compared to sun-protected skin. b1 integrin was also decreased. Since laminin-511 interacts with b1 integrin expressed on basal keratinocytes, we hypothesized that laminin-511 plays a key role in epidermal homeostasis by maintaining epidermal proliferation and differentiation. Through a screening study, we identified 1-(2-hydroxyethyl)-2-imidazolidinone (HEI) as an inhibitor of laminin-511 degradation. HEI potently inhibited the activity of heparanase and matrix metalloproteinase 9 (MMP9), which degrade epidermal BM and are activated in sun-exposed skin. In the skin equivalent model of human skin, HEI promoted laminin-511 deposition at the epidermal BM, increased filaggrin and loricrin expression in granular layer in epidermis, and maintained Ki67 positive basal keratinocytes. HEI also increased laminin-511 deposition and promoted formation of continuous lamina densa and hemidesmosomes at the epidermal BM, maintained Ki67 positive basal keratinocytes, promoted filaggrin and loricrin expression, decreased transepidermal water loss (TEWL), and increased water content of stratum corneum in UVB exposed organ culture model. In a pilot study, it improved the barrier function of human facial skin. These results suggest laminin-511 is a key regulator of epidermal homeostasis by regulating proliferation and differentiation of basal keratinocytes in epidermis.

Solo, a RhoA-targeting guanine nucleotide exchange factor, is critical for hemidesmosome formation and acinar development in epithelial cells.

Cell-substrate adhesions are essential for various physiological processes, including embryonic development and maintenance of organ functions. Hemidesmosomes (HDs) are multiprotein complexes that attach epithelial cells to the basement membrane. Formation and remodeling of HDs are dependent on the surrounding mechanical environment; however, the upstream signaling mechanisms are not well understood. We recently reported that Solo (also known as ARHGEF40), a guanine nucleotide exchange factor targeting RhoA, binds to keratin8/18 (K8/K18) intermediate filaments, and that their interaction is important for force-induced actin and keratin cytoskeletal reorganization. In this study, we show that Solo co-precipitates with an HD protein, β4-integrin. Co-precipitation assays revealed that the central region (amino acids 330–1057) of Solo binds to the C-terminal region (1451–1752) of β4-integrin. Knockdown of Solo significantly suppressed HD formation in MCF10A mammary epithelial cells. Similarly, knockdown of K18 or treatment with Y-27632, a specific inhibitor of Rho-associated kinase (ROCK), suppressed HD formation. As Solo knockdown or Y-27632 treatment is known to disorganize K8/K18 filaments, these results suggest that Solo is involved in HD formation by regulating K8/K18 filament organization via the RhoA-ROCK signaling pathway. We also showed that knockdown of Solo impairs acinar formation in MCF10A cells cultured in 3D Matrigel. In addition, Solo accumulated at the site of traction force generation in 2D-cultured MCF10A cells. Taken together, these results suggest that Solo plays a crucial role in HD formation and acinar development in epithelial cells by regulating mechanical force-induced RhoA activation and keratin filament organization.