Keratinocyte Adhesion Research Articles

The transient appearance of zipper-like actin superstructures during the fusion of osteoclasts

Multinucleated osteoclasts are responsible for bone resorption. Hypermultinucleated osteoclasts are often observed in some bone-related diseases such as Paget's disease and cherubism. The cellular mechanics controlling the size of osteoclasts is poorly understood. We introduced EGFP-actin into RAW 264.7 cells to monitor actin dynamics during osteoclast differentiation. Before their terminal differentiation into osteoclasts, syncytia displayed two main types of actin assembly, podosome clusters and clusters of zipper-like structures. The zipper-like structures morphologically resembled the adhesion zippers found at the initial stage of cell-cell adhesion in keratinocytes. In the zipper-like structure, Arp3 and cortactin overlapped with the distribution of dense F-actin, whereas integrin β3, paxillin and vinculin were localized to the periphery of the structure. The structure was negative for WGA-lectin staining and biotin labeling. The zipper-like structure broke down and transformed into a large actin ring, called a podosome belt. Syncytia containing clusters of zipper-like structures had more nuclei than those with podosome clusters. Differentiated osteoclasts with a podosome belt also formed the zipper-like structure at the cell contact site during cell fusion. The breakdown of the cell contact site resulted in the fusion of the podosome belts following plasma membrane fusion. Additionally, osteoclasts in mouse calvariae formed the zipper-like structure in the sealing zone. Therefore, we propose that the zipper-like actin superstructures might be involved in cell-cell interaction to achieve efficient multinucleation of osteoclasts. Understanding of the zipper-like structure might lead to selective therapeutics for bone diseases caused by hypermultinucleated osteoclasts.

The DSPII splice variant is critical for desmosome-mediated HaCaT keratinocyte adhesion

Desmosomes are intercellular junctions specialised for strong adhesion that are prominent in the epidermis and heart muscle. Defective desmosomal function due to inherited mutations in the constitutive desmosomal gene desmoplakin (DSP) causes skin or heart disorders and in some instances both. Different mutations have different disease-causing molecular mechanisms as evidenced by the varying phenotypes resulting from mutations affecting different domains of the same protein, but the majority of these mechanisms remain to be determined. Here, we studied two mutations in DSP that lead to different dosages of the two major DSP splice variants, DSPI and DSPII, and compared their molecular mechanisms. One of the mutations results in total DSP haploinsufficiency and is associated with autosomal dominant striate palmoplantar keratoderma (PPK). The other leads to complete absence of DSPI and the minor isoform DSPIa but normal levels of DSPII, and is associated with autosomal recessive epidermolytic PPK, woolly hair and severe arrhythmogenic dilated cardiomyopathy. Using siRNA treatments to mimic these two mutations and additionally a DSPII-specific siRNA, we found striking differences between DSP isoforms with respect to keratinocyte adhesion upon cellular stress with DSPII being the key component in intermediate filament (IF) stability and desmosome-mediated adhesion. In addition, reduction in DSP expression reduced the amount of plakophilin 1, desmocollin (DSC) 2 and DSC3 with DSPI having a greater influence than DSPII on the expression levels of DSC3. These results suggest that the two major DSP splice variants are not completely redundant in function and that DSPII dosage is particularly important for desmosomal adhesion in the skin.

Adherence and viability of primary human keratinocytes and primary human dermal fibroblasts on acrylonitrile-based copolymers with different concentrations of positively charged functional groups

As shown in several studies, various properties of biomaterials such as stiffness, surface roughness, chemical composition or the amount of functional groups at the surface can influence adhesion, viability, proliferation and functionalities of cells. The aim of this work was to explore whether a cell-selective effect could be achieved for acrylonitrile-based copolymers containing different contents of positively charged functional groups, which were introduced by incorporation of methacrylic acid-2-aminoethylester hydrochloride (AEMA) units. The p(AN-co-AEMA) copolymers were synthesized by suspension polymerization in water and processed into disk shaped test specimen via a sintering process to ensure the absence of organic solvents in the copolymers. Copolymers with an AEMA content of 1.4, 1.6, and 4.4 mol-% were investigated according to their cell-selective capacity, which should support the adhesion, viability and proliferation of keratinocytes, while the adherence of fibroblasts should rather be disabled. The test samples were seeded with primary human keratinocytes and primary human dermal fibroblasts in mono- as well as in co-cultures. Tissue culture plate polystyrene (TCP) was used to control the physiologic growth of the cells. Density and viability of attached and non-adherent cells were analyzed by live/dead staining, lactate dehydrogenase (LDH) assay and flow cytometry with DAPI staining. For the assured discrimination of adherent cell types in coculture a keratin/vimentin-staining was performed. On copolymers with 4.4 mol-% AEMA adherent keratinocytes in monoculture and cocultured keratinocytes and fibroblasts showed a higher viability, a lower impairment of cell membranes and higher densities of viable cells compared to both other copolymers. For adherent fibroblasts these parameters did not differ between the copolymers and an increasing ratio of keratinocytes to fibroblasts in cocultures were found with increasing AEMA content. The results showed that keratinocytes and fibroblasts can be influenced by copolymers with different contents of positively charged functional groups. Since the tendency of a better adherence and viability of keratinocytes with increasing amounts of positively charged functional groups was shown, the potential enhancement by further increase of the amount of positively charged functional groups shall be tested in a future study.

Semi-interpenetrated Hydrogels Composed of PVA and Hyaluronan or Chondroitin Sulphate: Chemico-Physical and Biological Characterization

Physical hydrogels based on poly (vinyl alcohol) (PVA) were synthesized in the presence of natural polysaccharides, either hyaluronan or Chondroitin sulphate, in order to obtain versatile biomaterials with enhanced performances. The physical network is obtained by the freeze thawing technique, a simple method widely used for structuring PVA blends. The chemico-physical characterization of resulting materials consisted of swelling studies and mechanical analysis. Furthermore the release of embedded polysaccharides from the network was evaluated to improve understanding of the strength of hydrogen bonding between the different polymeric chains, and the effect of the sulphate groups on the interaction promoting network formation and stability. Biological response in terms of cytotoxicity, adhesion and cell vitality of murine fibroblast and human keratinocytes showed that the addition of glycosaminoglycans to a PVA polymer leads to a biomaterial with potential applications in biomedical fields.

Silica Nanoparticles Suppress Fibronectin-Mediated Adhesion and Migration in Normal Human Keratinocytes

Nanotechnology has been increasingly applied to various fields, such as biology, chemistry, physics, medicine and engineering. However, a major concern that has been the topic in nanoscience is whether exposure of humans to engineered nanoparticles might cause toxic effects. In the present in vitro study, the influence of silica nanoparticles on fibronectin-mediated cellular response was assessed in normal human keratinocytes. Our results demonstrated that silica nanoparticles but not silica microparticles significantly suppressed cell adhesion and migration to fibronectin. This phenomenon was not observed in cell response to Poly-L-Lysine, which mediates cell adhesion and migration in a way different from that of fibronectin. Moreover, it seemed that this suppression was not due to cytotoxic effects induced by silica nanoparticles. Subsequently, we also showed that silica nanoparticles impaired the fibronectin-induced activation of FAK and its downstream PI3K, AKT and Src. Taken together, our data suggests that silica nanoparticles may negatively modulate cell response to fibronectin.

Confluence switch signaling regulates ECM composition and plasmin proteolytic cascade in keratinocytes

In culture, cell confluence generates signals that commit actively growing keratinocytes to exit the cell cycle and differentiate to form a stratified epithelium. Using a comparative proteomic approach, we studied this 'confluence switch' and identified a new pathway triggered by cell confluence that regulates basement membrane (BM) protein composition by suppressing the uPA-uPAR-plasmin pathway. Indeed, confluence triggers adherens junction maturation and enhances TGF-β and activin A activity, resulting in increased deposition of PAI-1 and perlecan in the BM. Extracellular matrix (ECM)-accumulated PAI-1 suppresses the uPA-uPAR-plasmin pathway and further enhances perlecan deposition by inhibiting its plasmin-dependent proteolysis. We show that perlecan deposition in the ECM strengthens cell adhesion, inhibits keratinocyte motility and promotes additional accumulation of PAI-1 in the ECM at confluence. In agreement, during wound-healing, perlecan concentrates at the wound-margin, where BM matures to stabilize keratinocyte adhesion. Our results demonstrate that confluence-dependent signaling orchestrates not only growth inhibition and differentiation, but also controls ECM proteolysis and BM formation. These data suggest that uncontrolled integration of confluence-dependent signaling, might favor skin disorders, including tumorigenesis, not only by promoting cell hyperproliferation, but also by altering protease activity and deposition of ECM components.

Novel cholinergic peptides SLURP‐1 and ‐2 regulate epithelialization of cutaneous and oral wounds

It is well established that auto/paracrine acetylcholine (ACh) is essential for wound epithelialization, and that the mechanisms include regulation of keratinocyte motility and adhesion via nicotinic ACh receptors (nAChRs). Keratinocyte nAChRs can be also activated by non-canonical ligands, such as secreted mammalian Ly-6/urokinase-type plasminogen activator receptor-related protein (SLURP)-1 and -2. In this study, we determined effects of recombinant (r)SLURP-1 and-2 on migration of human epidermal and oral keratinocytes under agarose and epithelialization of cutaneous and oral mucosal excisional wounds in mice, and also identified nAChRs mediating SLURP signals. Both in vitro and in vivo, rSLURP-1 decreased and SLURP-2 increased epithelialization rate. The mixture of both peptides accelerated epithelialization even further, indicating that their simultaneous signaling renders an additive physiologic response. The specificity of rSLURP actions was illustrated by similar effects on cutaneous and oral wounds, which feature distinct responses to injury, and also by abrogation of rSLURP effects with neutralizing antibodies. rSLURP-1 acted predominantly via the α7 nAChR-coupled up-regulation of the sedentary integrins α2 and α3 , whereas SLURP-2--through α3, and α9 nAChRs up-regulating migratory integrins α5 and αV . The biologic effects of rSLURPs required the presence of endogenous ACh, indicating that auto/paracrine SLURPs provide for a fine tuning of the physiologic regulation of crawling locomotion via the keratinocyte ACh axis. Since nAChRs have been shown to regulate SLURP production, cholinergic regulation of keratinocyte migration appears to be mediated by a reciprocally arranged network. The cholinergic peptides, therefore, may become prototype drugs for the treatment of wounds that fail to heal.

Proteomic Identification of Betaig-h3 as a Lysophosphatidic Acid-Induced Secreted Protein of Human Mesenchymal Stem Cells: Paracrine Activation of A549 Lung Adenocarcinoma Cells by Betaig-h3

Lysophosphatidic acid (LPA) is enriched in the serum and malignant effusion of cancer patients and plays a key role in tumorigenesis and metastasis. LPA-activated mesenchymal stem cells promote tumorigenic potentials of cancer cells through a paracrine mechanism. LPA-conditioned medium (LPA CM) from human adipose tissue-derived mesenchymal stem cells (hASCs) elicited adhesion and proliferation of A549 human lung adenocarcinoma cells. To identify proteins involved in the LPA-stimulated paracrine functions of hASCs, we analyzed the LPA CM using liquid-chromatography tandem mass spectrometry-based shotgun proteomics. We identified βig-h3, an extracellular matrix protein that is implicated in tumorigenesis and metastasis, as an LPA-induced secreted protein in hASCs. LPA-induced βig-h3 expression was abrogated by pretreating hASCs with the LPA receptor(1/3) inhibitor Ki16425 or small interfering RNA-mediated silencing of endogenous LPA(1). LPA-induced βig-h3 expression was blocked by treating the cells with the Rho kinase inhibitor Y27632, implying that LPA-induced βig-h3 expression is mediated by the LPA(1)- Rho kinase pathway. Immunodepletion or siRNA-mediated silencing of βig-h3 abrogated LPA CM-stimulated adhesion and proliferation of A549 cells, whereas retroviral overexpression of βig-h3 in hASCs potentiated it. Furthermore, recombinant βig-h3 protein stimulated the proliferation and adhesion of A549 human lung adenocarcinoma cells. These results suggest that hASC-derived βig-h3 plays a key role in tumorigenesis by stimulating the adhesion and proliferation of cancer cells and it can be applicable as a biomarker and therapeutic target for lung cancer.

Autoimmune bullous diseases in childhood

Autoimmune blistering disorders are a heterogeneous group of diseases that result from autoantibodies generated against target antigens found in the skin and mucous membranes. This process leads to a variety of disruptions in keratinocyte adhesion and cellular integrity, resulting in fluid accumulation and development of blisters. Physicians should have an appreciation and understanding of autoimmune blistering disorders in the pediatric population when formulating a differential diagnosis of a patient who presents with skin blistering. Early detection and discrimination between the varied autoimmune blistering disorders can change the course of treatment and outcome. Due to the similarity in clinical presentation among different diseases within this category, histopathologic evaluation and, especially, immunofluorescence studies are necessary to establish the definitive diagnosis.

Bioactive support for cell cultivation and potential grafting. Part 1: Surface modification of 2-hydroxyethyl methacrylate hydrogels for avidin immobilization

AbstractSynthetic hydrogels are often used in biomedical applications as many of them are compatible with living tissue and moreover they can meet most criteria for artificial tissue properties. For applications in tissue engineering modification of polymer surface using some bioactive compounds (e.g. saccharides, proteins) for promoting the process of cell adhesion and proliferation is widely used. In this work, a series of modified hydrogels was prepared by three different methods: by copolymerization of 2-hydroxyethyl methacrylate with methacrylic acid, by hydrolysis and oxidation of poly(2-hydroxyethyl methacrylate) to obtain carboxyl-rich supports. The influence of the reaction conditions of the hydrogel surface treatment on the total carboxylic group content and the swelling degree was studied. The modified hydrogels were characterized by attenuated total reflectance FT-IR spectroscopy and refractive index measurements. Obtained carboxylic groups on the hydrogel surface have allowed the immobilization of avidin in two ways: 1) electrostatically through dissociated carboxylic groups and 2) covalently bonded through activated carboxylic groups by Nhydroxysuccinimide. The capacity of hydrogels for avidin immobilization was determined by Bradford spectrophotometric method. The results so far obtained from the preliminary biological tests showed that immobilized avidin on the hydrogel surface provides better adhesion and proliferation of keratinocytes compared to supports without avidin.

N-glycosylation of ß4 integrin controls the adhesion and motility of keratinocytes.

α6ß4 integrin is an essential component of hemidesmosomes and modulates cell migration in wound healing and cancer invasion. To elucidate the role of N-glycosylation on ß4 integrin, we investigated keratinocyte adhesion and migration through the re-expression of wild-type or N-glycosylation-defective ß4 integrin (ΔNß4) in ß4 integrin null keratinocytes. N-glycosylation of ß4 integrin was not essential for the heterodimer formation of ß4 integrin with α6 integrin and its expression on a cell surface, but N-glycosylation was required for integrin-mediated cell adhesion and migration. Concomitantly with the reduction of ß4 integrin in the membrane microdomain, the intracellular signals of Akt and ERK activation were decreased in cells expressing ΔNß4 integrin. Forced cross-linking of ß4 integrin rescued the decreased ERK activation in ΔNß4 integrin-expressing cells to a similar extent in wild-type ß4 integrin-expressing cells. Surprisingly, compared with cells expressing wild-type ß4 integrin, an alternation in N-glycan structures expressed on epidermal growth factor receptor (EGFR), and the induction of a stronger association between EGFR and ß4 integrin were observed in ΔNß4 integrin-expressing cells. These results clearly demonstrated that N-glycosylation on ß4 integrin plays an essential role in keratinocyte cellular function by allowing the appropriate complex formation on cell surfaces.

Protein aggregation of SERCA2 mutants associated with Darier disease elicits ER stress and apoptosis in keratinocytes

Mutations in sarcoplasmic/endoplasmic reticulum calcium ATPase 2 (SERCA2) underlie Darier disease (DD), a dominantly inherited skin disorder characterized by loss of keratinocyte adhesion (acantholysis) and abnormal keratinization (dyskeratosis) resulting in characteristic mucocutaneous abnormalities. However, the molecular pathogenic mechanism by which these changes influence keratinocyte adhesion and viability remains unknown. We show here that SERCA2 protein is extremely sensitive to endoplasmic reticulum (ER) stress, which typically results in aggregation and insolubility of the protein. Depletion of ER calcium stores is not necessary for the aggregation but accelerates the progression. Systematic analysis of diverse mutants identical to those found in DD patients demonstrated that the ER stress initiator is the SERCA2 mutant protein itself. These SERCA2 proteins were found to be less soluble, to aggregate and to be more polyubiquitinylated. After transduction into primary human epidermal keratinocytes, mutant SERCA2 aggregates elicited ER stress, caused increased numbers of cells to round up and detach from the culture plate, and induced apoptosis. These mutant induced events were exaggerated by increased ER stress. Furthermore, knockdown SERCA2 in keratinocytes rendered the cells resistant to apoptosis induction. These features of SERCA2 and its mutants establish a mechanistic base to further elucidate the molecular pathogenesis underlying acantholysis and dyskeratosis in DD.

Pemphigus Treatment in Japan

Pemphigus is a group of autoimmune blistering diseases characterized by blisters and erosive lesions on the skin and mucous membranes due to loss of cell and cell adhesion of keratinocytes. In 2010, new Japanese guidelines for the management of pemphigus were published for dermatologists. Systemic corticosteroids are the gold standard and the first choice of treatment of pemphigus.

Collagen XXIII, Novel Ligand for Integrin α2β1 in the Epidermis

Cellular receptors for collagens belong to the family of β(1) integrins. In the epidermis, integrin α(2)β(1) is the only collagen-binding integrin present. Its expression is restricted to basal keratinocytes with uniform distribution on the cell surface of those cells. Although α(2)β(1) receptors localized at the basal surface interact with basement membrane proteins collagen IV and laminin 111 and 332, no interaction partners have been reported for these integrin molecules at the lateral and apical membranes of basal keratinocytes. Solid phase binding and surface plasmon resonance spectroscopy demonstrate that collagen XXIII, a member of the transmembrane collagens, directly interacts with integrin α(2)β(1) in an ion- and conformation-dependent manner. The two proteins co-localize on the surface of basal keratinocytes. Furthermore, collagen XXIII is sufficient to induce adhesion and spreading of keratinocytes, a process that is significantly reduced in the absence of functional integrin α(2)β(1).

Upregulation of CD26 expression in epithelial cells and stromal cells during wound-induced skin tumour formation

We have previously described InvEE transgenic mice in which non-dividing, differentiating epidermal cells express oncogenically activated MAPK kinase 1 (MEK1). Skin wounding triggers tumour formation in InvEE mice via a mechanism that involves epidermal release of IL-1α and attraction of a pro-tumorigenic inflammatory infiltrate. To look for potential effects on the underlying connective tissue, we screened InvEE and wild-type epidermis for differential expression of cytokines and immune modulators. We identified a single protein, CD26 (dipeptidyl peptidase-4). CD26 serum levels were not increased in InvEE mice. In contrast, CD26 was upregulated in keratinocytes expressing mutant MEK1 and in the epithelial compartment of InvEE tumours, where it accumulated at cell-cell borders. CD26 expression was increased in dermal fibroblasts following skin wounding but was downregulated in tumour stroma. CD26 activity was stimulated by calcium-induced intercellular adhesion in keratinocytes, suggesting that the upregulation of CD26 in InvEE epidermis is due to expansion of the differentiated cell layers. IL-1α treatment of dermal fibroblasts stimulated CD26 activity, and therefore epidermal IL-1α release may contribute to the upregulation of CD26 expression in wounded dermis. Pharmacological blockade of CD26, via Sitagliptin, reduced growth of InvEE tumours, while combined inhibition of IL-1α and CD26 delayed tumour onset and reduced tumour incidence. Our results demonstrate that inappropriate activation of MEK1 in the epidermis leads to changes in dermal fibroblasts that, like the skin inflammatory infiltrate, contribute to tumour formation.

Protein 4.1R regulates cell adhesion, spreading, migration and motility of mouse keratinocytes by modulating surface expression of β1 integrin

Protein 4.1R is a membrane-cytoskeleton adaptor protein that has diverse roles in controlling the cell surface expression and/or function of transmembrane proteins, and in organizing F-actin. 4.1R is expressed in keratinocytes, but its role in these cells has not been explored. Here, we have investigated the role of 4.1R in skin using 4.1R(-/-) mice. Cell adhesion, spreading, migration and motility were significantly impaired in 4.1R(-/-) keratinocytes, and 4.1R(-/-) mice exhibited defective epidermal wound healing. Cultured 4.1R(-/-) keratinocytes on fibronectin failed to form actin stress fibres and focal adhesions. Furthermore, in the absence of 4.1R, the surface expression, and consequently the activity of β1 integrin were reduced. These data enabled the identification of a functional role for protein 4.1R in keratinocytes by modulating the surface expression of β1 integrin, possibly through a direct association between 4.1R and β1 integrin.

The Calcium-Sensing Receptor-Dependent Regulation of Cell–Cell Adhesion and Keratinocyte Differentiation Requires Rho and Filamin A

Extracellular Ca(2+) (Ca(2+)(o)) functioning through the calcium-sensing receptor (CaR) induces E-cadherin-mediated cell-cell adhesion and cellular signals mediating cell differentiation in epidermal keratinocytes. Previous studies indicate that CaR regulates cell-cell adhesion through Fyn/Src tyrosine kinases. In this study, we investigate whether Rho GTPase is a part of the CaR-mediated signaling cascade regulating cell adhesion and differentiation. Suppressing endogenous Rho A expression by small interfering RNA (siRNA)-mediated gene silencing blocked the Ca(2+)(o)-induced association of Fyn with E-cadherin and suppressed the Ca(2+)(o)-induced tyrosine phosphorylation of β-, γ-, and p120-catenin and formation of intercellular adherens junctions. Rho A silencing also decreased the Ca(2+)(o)-stimulated expression of terminal differentiation markers. Elevating the Ca(2+)(o) level induced interactions among CaR, Rho A, E-cadherin, and the scaffolding protein filamin A at the cell membrane. Inactivation of CaR expression by adenoviral expression of a CaR antisense complementary DNA inhibited Ca(2+)(o)-induced activation of endogenous Rho. Ca(2+)(o) activation of Rho required a direct interaction between CaR and filamin A. Interference of CaR-filamin interaction inhibited Ca(2+)(o)-induced Rho activation and the formation of cell-cell junctions. These results indicate that Rho is a downstream mediator of CaR in the regulation of Ca(2+)(o)-induced E-cadherin-mediated cell-cell adhesion and keratinocyte differentiation.

Dermal fibroblast and epidermal keratinocyte functionality on titania nanotube arrays

Transcutaneous implants that penetrate through the depth of the skin are used in numerous clinical applications, including prosthetics and dental implants. Favorable interactions between the implant surface and the respective skin layers are critical for the long-term success of transcutaneous implantable devices, hence, it is essential to understand the physiologic response elicited by skin–biomaterial interactions. Recent studies have shown that material surfaces that provide topographic cues at the nanoscale level may provide one possible solution to enhanced biomaterial integration, thus preventing biomaterial rejection. In this study titania nanotube arrays were fabricated using a simple anodization technique as potential interfaces for transcutaneous implantable devices. The in vitro functionality of human dermal fibroblasts and epidermal keratinocytes were evaluated on these nanotube arrays (diameter 70–90 nm, length 1–1.5 μm). Cellular functionality in terms of adhesion, proliferation, orientation, viability, cytoskeletal organization, differentiation and morphology were investigated for up to 4 days in culture using fluorescence microscope imaging, a cell viability assay, indirect immunofluorescence and scanning electron microscopy. The results reported in this study indicate increased dermal fibroblast and decreased epidermal keratinocyte adhesion, proliferation and differentiation on titania nanotube arrays.

IgG Autoantibodies Against Desmocollin 3 in Pemphigus Sera Induce Loss of Keratinocyte Adhesion

Pemphigus is considered an autoimmune bullous skin disorder associated with IgG against the desmosomal components, desmoglein 3 (Dsg3) and desmoglein 1 (Dsg1). This concept is supported by the in vitro and in vivo pathogenicity of anti-Dsg3/Dsg1 IgG and the mucosal blistering phenotype of mice with a genetic deficiency of Dsg3. Mice deficient for another desmosomal adhesion molecule, desmocollin 3 (Dsc3), show a similar pemphigus phenotype, and we investigated the pathogenicity of Dsc3-reactive IgG autoantibodies that were identified previously in a subset of patients with atypical pemphigus. We here demonstrate that IgG against Dsc3 causes loss of adhesion of epidermal keratinocytes. Specifically, IgG against Dsc3 was purified from Dsc3-reactive pemphigus sera by affinity column chromatography using recombinant human Dsc3. Affinity purified IgG was functionally active and did not only react with recombinant Dsc3 but also with epidermis and cultured human keratinocytes. Moreover, Dsc3-reactive IgG induced loss of adhesion of epidermal keratinocytes in a dispase-based keratinocyte dissociation assay that was reversed on pre-adsorption with human Dsc3 but not Dsg3. These findings demonstrate that IgG autoantibodies against an additional component of the desmosomes, Dsc3, induce loss of keratinocyte adhesion and thus may contribute to blister formation in pemphigus.

Hemidesmosomes and focal contact proteins: Functions and cross-talk in keratinocytes, bullous diseases and wound healing

The outer most layer of the skin, the epidermis, is attached to the dermis via a sheet of extracellular matrix proteins termed the basement membrane zone (BMZ). In the intact skin, adhesion of the keratinocytes in the basal layer of the epidermis to the BMZ is facilitated primarily by hemidesmosomes which associate with the keratin cytoskeleton. Cultured keratinocytes do not assemble bona fide hemidesmosomes although hemidesmosome protein clusters (stable anchoring contacts) are found along the substrate-attached surface of the cells and towards the leading edge of keratinocytes repopulating scratch wounds. Actin cytoskeleton-associated matrix adhesion devices termed focal contacts are not thought to play an important role in the adhesion of keratinocytes to the BMZ in intact skin but are prominent in cultured keratinocytes where they are believed to regulate cell migration. We review the molecular components, functions, dynamics and cross-talk of hemidesmosomes and focal contacts in keratinocytes. In addition, we briefly describe what is known about their role in autoimmune and genetic blistering diseases of the skin. We also discuss recent publications which indicate, contrary to expectation, that certain focal contact proteins retard keratinocyte migration while hemidesmosomal proteins regulate directed keratinocyte motility during wound healing.