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Biomedical subjects

M Haftek

Publications and source records attributed to M Haftek.

At least 19 recordsLinked to original sources
PubMed2005

[Netherton syndrome: a type of infantile erythroderma with failure to thrive, immune deficiency, rickets. Report of 3 cases].

We report the cases of 2 boys and 1 girl suffering from Netherton syndrome. Both boys presented with a non-bullous congenital erythroderma and were diagnosed early as Netherton syndrome with hair biopsies. Both had severe failure to thrive, signs of atopy, several episodes of bacterial infection, and rickets (with a high blood level of vitamin D in the first boy, and vitamin D deficiency in the second). In the third case, the pilar abnormality appeared at the age of 3 years. The girl had ichtyosis linearis circumflexa, failure to thrive and severe constipation. Netherton syndrome is a rare disorder characterized by severe ichtyosis, signs of atopy, immune deficiency and failure to thrive. The disease is severe and comprises many complications in early infancy. It is due to a genetic disorder of recessive autosomal transmission, and the gene, SPINK5, is located in the chromosome 5. Prenatal diagnosis is possible. Two of our patients had rickets, which has never been described in such patients population.

Carrier Proteins↗
PubMed2005

Hereditary mucoepithelial dysplasia: clinical, ultrastructural and genetic study of eight patients and literature review.

BACKGROUND: Hereditary mucoepithelial dysplasia is a dominantly inherited disease, mainly characterized by chronic mucosal lesions associated with keratitis, non-scarring alopecia, keratosis pilaris and perineal intertrigo. Since the original report by Witkop, this condition has been considered to be a disorder of desmosome/gap junction formation, but there has been no ex vivo investigation of these components using genetic and immunolabelling techniques. OBJECTIVES: To perform light and immunoelectron microscopic studies, and partial genetic analysis on five patients in a family and three sporadic cases and to point out similarities of this rare disorder with chronic mucocutaneous candidiasis and other follicular keratosis syndromes, i.e. ichthyosis follicularis-alopecia-photophobia (IFAP), keratitis-ichthyosis-deafness (KID) and Siemens syndromes. METHODS: Biopsies from the involved oral mucosa and armpit skin of patient 1 were prepared for standard histopathology, electron microscopy and immunocytochemistry. Microsatellite genotyping was performed in three affected family members. Direct sequencing after polymerase chain reaction amplification of the entire coding region was performed. RESULTS: A 14-year-old male had recurrent keratitis, widespread keratosis pilaris, perineal intertrigo, hypotrichosis and oral mucosal involvement. A similar phenotype was noted in four members of his family and in three sporadic cases. Histological examination of oral mucosa and skin samples showed a psoriasiform pattern, dyskeratotic features and cytoplasmic vacuoles. Expression of connexins (Cx), desmosomal, adherens junction and cytoskeleton proteins (Cx 26, 32 and 43, desmogleins 1 and 2, plakoglobin, desmoplakins I-II, plakophilin 1, beta-catenin, E-cadherin, keratins, beta-tubulin, vimentin and actin) was normal. Ultrastructural studies showed a reduced number of desmosomes. Dyskeratotic cells exhibited internalized gap junctions, long filamentous inclusions reactive with antikeratin antibodies, and bundles of perinuclear fibres resembling clear tonofilaments. Genetic analysis in the studied family excluded the desmosomal cadherins in chromosome 18q12 as candidate genes. CONCLUSIONS: A diagnosis of hereditary mucoepithelial dysplasia should be strongly suggested by the triad of non-scarring alopecia, well-demarcated erythema of oral mucosa and psoriasiform perineal rash, after exclusion of the clinically related follicular keratosis syndromes. Defective expression of cytoskeleton elements and/or a modification of mechanisms regulating junction-cytoskeleton assembly may be primarily responsible for impaired epithelial cohesion.

Adolescent↗
PubMed2004

Hereditary 'white nails': a genetic and structural study.

BACKGROUND: Hereditary subtotal leuconychia is a rare nail disease. The gene(s) underlying this phenotype is (are) not known. Immunohistochemical and ultrastructural studies of nails are performed infrequently. OBJECTIVES: To perform genetic linkage analysis and to assess ultrastructure and soft/hard keratin expression in hereditary white nails. METHODS: We have analysed microscopically and ultrastructurally the white nails of a patient from a family in which the trait is inherited in an autosomal dominant manner as an isolated symptom. No skin lesions or hair abnormalities could be detected. Genetic linkage studies were performed on DNA samples obtained from several members of the affected family. A longitudinal surgical biopsy of the nail from a great toe was split in two parts. One part was fixed in formalin and processed for histopathology. Another part was further subdivided and embedded either in Epon, following fixation in 2% glutaraldehyde, or in Lowicryl K4M, after fixation in 3% paraformaldehyde. Dewaxed nail sections and Lowicryl ultrathin sections were also stained with various antikeratin antibodies. RESULTS: Genetic linkage studies of the family pointed to the disease gene mapping to the chromosomal 12q13 region. Genes mapping within this chromosomal region include the genes coding for type II (basic) cytokeratins and hard keratins. The nail matrix presented an abnormal hypergranulosis. The upper part of the nail plate, originating from the proximal nail matrix, had a nonhomogeneous lamellar appearance, with numerous intracellular 'lipidic' vacuoles and 'empty' spaces separating keratin filament bundles. These cells were progressively shed at the nail surface. The cell loss was compensated by hyperproliferation of the distal matrix and of the nail bed keratinocytes, with persistent marked parakeratosis and loose arrangement of keratin bundles. The distal matrix and the nail bed contributed equally to formation of the lower plate. This presented the characteristics of a tissue composed of soft keratins. Accordingly, there was virtually no labelling with the Hb1 antibody to a basic hard keratin in the white nail, whereas the labelling with AE3 antibody to all type II keratins and with KL1 recognizing suprabasal soft keratins was normal or even enhanced. CONCLUSIONS: Genetic linkage indicates that the gene defect underlying the leuconychia in the family studied resides on chromosome 12q13. As the type II keratins map within this chromosomal interval, it is possible that a mutation in one of these keratin genes may be a cause of the hereditary leuconychia. The white appearance of nails in this disease seems to be due to an abnormal keratinization of cells originating from the proximal nail matrix, leading to the presence of abundant intracellular vacuoles and to a lesser compactness of keratins.

Chromosome Mapping↗
PubMed2003

[Structural and ultrastructural data on human cutaneous lipids].

The principal function of intercellular lipids of the epidermal horny layer is the constitution of an occlusive barrier and regulation of transepidermal water fluxes. Self-assembly of lipid bilayers and formation of multilayered structures can be observed in the intercellular spaces of the stratum corneum. Progressive organisation of lipids provokes the lateral displacements of hydrophilic compartments, squeezed aside along the lipid layers towards intercellular junctions (corneodesmosomes). Proteolytic enzymes contained in the hydrophilic material are thus delivered to the extracellular portions of cohesive junctional proteins. Consequently, modifications of the composition and structure of the stratum corneum lipids may influence epidermal desquamation through the regulation of corneodesmosome accessibility to extracellular enzymes.

Epidermis↗
PubMed2003

[Sub-total hereditary leukonychia, histopathological and electron microscopy study of "milky" nails].

INTRODUCTION: Leuconychia is the most common of ungueal discoloration or dyschromia. The hereditary form, sub-total or total, is very rare. We report the observation of a family in which thirteen members, distributed over four generations, presented a partial or subtotal leukonychia of all nails. On the basis of this observation and histopathological analysis of the propositus nail, we discuss the different hypotheses proposed to account for white nails. OBSERVATION: A 45 year-old man without previous medical history, showed partial or sub-total leukonychia of his twenty nails. Clinical examination revealed soft nails with slow growth. Discoloration or dyschromia was associated with koilonychia on the fingers without other cutaneous or visceral abnormalities. A longitudinal biopsy of the whole ungueal apparatus of the large toe was performed. Histopathological analysis showed parakeratosis and an abnormal granular layer thickened on the proximal and ventral womb. These abnormalities were responsible for heterogeneous HES coloration, lamellar and dissociated aspect of the nail plates. Electron microscopy revealed dissociated keratin bundles and the existence of intracytoplasmic clear vacuoles probably of lipid origin. This aspect was observed on the proximal part of the dorsal tablet and disappeared in the distal portion of the nail. DISCUSSION: According to Newton's theorem, a surface appears white when it reflects all the radiation of visible light. This mechanism can be proposed to explain leukonychia. On histological level, parakeratosis and dissociation of the keratin bundles may play a role in the modification of the solar light reflection by ungueal plates. In our case, parakeratosis and disorganization of keratin bundles were present in the white part of the nail, but also in the distal, pinkish crescent (distal dorsal plate). Electron microscopy analysis showed clear vacuoles located in the white part of the nail, whereas they were not seen in the distal part. These observations confirm the probable participation of parakeratosis and keratin abnormalities in this pathology but also suggest an important role of lipid vacuoles. Therefore, some white nails may in fact be "milky" nails. Finally, disorganization of the keratin bundles observed, was also reported in other genodermatosis e.g.: epidermolysis bullosa simplex. In this latter pathology, some hard keratin genes mutations (K5 and K14) have been demonstrated. According to these results, a genetic study is on going in this family in order to search for a mutation in one of the hard keratin genes.

Humans↗
PubMed2002

[The stratum corneum].

Epidermal differentiation is a continuous process, constituting and renewing a protective layer at the skin surface: the stratum corneum, composed of cornified keratinocytes that is a barrier to water diffusion. Massive loss of physiologic liquids is one of the severest consequences of extensive burns. The stratum corneum also protects the subjacent tissues from xenobiotic aggression, ultraviolet radiation (70 p. cent of UVB absorption) and from mechanic aggression. The integrity of the stratum corneum depends on three elements: 1) the physico-chemical quality of the cornified cells, 2) the persistence of mechanical junctions uniting these cells and 3) the organization and composition of the lipid "mortar" in the intercellular spaces. Since all these components are issued from the keratinocyte differentiation process, any perturbation may, in time, induce modifications in the "barrier" function of the epidermis. The barrier quality varies, depending on its localization (soles of the feet, palms of the hands and transitional epidermis of the lips), during skin healing (priority to keratinocyte proliferation to the detriment of maturation) under the influence of treatment (retinoids, vitamin D derivatives), in the course of aging and diseases: ichtyosis and other keratinization genodermatoses, benign (including psoriasis) and malignant hyperproliferative diseases. Furthermore, the relative impermeability of the stratum corneum is an important factor limiting the penetration and diffusion of allergens, but also local drugs delivered with local and systemic trans-epidermal treatments. Further knowledge is required on the function of the epidermal barrier and the mechanism regulating cohesion/desquamation of the stratum corneum to understand certain hereditary diseases, improve efficacy of topical therapeutic products and optimize cosmetic formulations.

Body Water↗
PubMed2001

Refined characterization of corneodesmosin proteolysis during terminal differentiation of human epidermis and its relationship to desquamation.

Corneodesmosin is a putative adhesion glycoprotein located in the extracellular part of the desmosomes in the upper layers of the epidermis. Synthesized by granular keratinocytes as a 52-56-kDa protein, corneodesmosin is progressively proteolysed during corneocyte maturation. This processing is a prerequisite for desquamation. Two glycine- and serine-rich domains of the protein might take on the conformation of adhesive secondary structures similar to glycine loops. Corneodesmosin proteolysis was further characterized. Deglycosylation experiments and reactivity with lectins demonstrated that the corneodesmosin carbohydrate moiety does not prevent the proteolysis. Immunoblotting, immunohistochemistry, and immunoelectron microscopy experiments using affinity-purified anti-peptide antibodies raised to four of the five structural domains of corneodesmosin and a monoclonal antibody against its fifth central domain showed that the first step in corneodesmosin processing is the cleavage of its extremities and probably occurs before its incorporation into desmosomes. Then the glycine loop-related domains are cleaved, first the N-terminal and then part of the C-terminal domain. At the epidermis surface, the multistep proteolytic cleavage leaves intact only the central domain, which was detected on exfoliated corneocytes and probably lacks adhesive properties. Importantly, corneodesmosin was demonstrated to be a preferred substrate of two serine proteases involved in desquamation, the stratum corneum tryptic and chymotryptic enzymes.

Amino Acid Sequence↗
PubMed2001

Topically applied vitamin C enhances the mRNA level of collagens I and III, their processing enzymes and tissue inhibitor of matrix metalloproteinase 1 in the human dermis.

Ascorbic acid (vitamin C) is a cofactor required for the function of several hydroxylases and monooxygenases. It is not synthesized in humans and some other animal species and has to be provided by diet or pharmacologic means. Its absence is responsible for scurvy, a condition related in its initial phases to a defective synthesis of collagen by the reduced function of prolylhydroxylase and production of collagen polypeptides lacking hydroxyproline, therefore, they are unable to assemble into stable triple-helical collagen molecules. In fibroblast cultures, vitamin C also stimulates collagen production by increasing the steady-state level of mRNA of collagen types I and III through enhanced transcription and prolonged half-life of the transcripts. The aim of the experimental work has been to evaluate the effect on dermal cells of a preparation of vitamin C topically applied on one side vs placebo on the other side of the dorsal face of the upper forearm of postmenopausal women. Biopsies were collected on both sides and the level of mRNA measured by non competitive reverse transcription-polymerase chain reaction made quantitative by the simultaneous transcription and amplification of synthetic RNA used as internal standards. The mRNA of collagen type I and type III were increased to a similar extent by vitamin C and that of three post-translational enzymes, the carboxy- and amino-procollagen proteinases and lysyloxidase similarly increased. The mRNA of decorin was also stimulated, but elastin, and fibrillin 1 and 2 were not modified by the vitamin. The expression of matrix metalloproteinases 1, 2, and 9 was not significantly changed, but an increased level of tissue inhibitor of matrix metalloproteinase 1 mRNA was observed without modification of tissue inhibitor of matrix metalloproteinase 2 mRNA. The stimulating activity of topical vitamin C was most conspicuous in the women with the lowest dietary intake of the vitamin and unrelated to the level of actinic damage. The results indicate that the functional activity of the dermal cells is not maximal in postmenopausal women and can be increased.

Administration, Topical↗
PubMed2000

Inherited palmoplantar keratoderma and sensorineural deafness associated with A7445G point mutation in the mitochondrial genome.

We report a French pedigree with members having an inherited combination of non-epidermolytic palmoplantar keratoderma (NEPPK) and sensorineural deafness. The penetrance of both features was incomplete. Additional ectodermal defects were absent. The expression of numerous epidermal proteins (keratins, fillagrin, cornified envelope proteins, intercellular junction proteins including connexin 26, and loricrin) defined with immunolabelling was normal in the proband. The combination was shown to be associated with the A7445G point mutation in the mitochondrial genome (mtDNA). This mutation is responsible for a subtype of NEPPK which is so far the only mtDNA mutation-associated keratoderma.

Adolescent↗
PubMed2000

Genomic amplification of the human plakophilin 1 gene and detection of a new mutation in ectodermal dysplasia/skin fragility syndrome.

Ectodermal dysplasia/skin fragility syndrome is a recently described autosomal recessive disease affecting skin, nails, and hair (MIM 604536), that results from mutations in plakophilin 1, a structural component of desmosomes. We report a new plakophilin 1 mutation in an affected patient as well as detailing the intron-exon organization of the gene to facilitate future polymerase chain reaction-based mutation screening. Using polymerase chain reaction amplification of genomic DNA, we identified 15 exons spanning approximately 50 kb. Direct sequencing disclosed several nonpathogenic intragenic polymorphisms, as well as a homozygous splice site mutation (1233-2 A-->T; GenBank Z73678) in a 17 y old affected male. The clinical features comprised skin erosions, dystrophic nails, sparse hair, and painful thickening and cracking of palms and soles. Skin biopsy showed negative immunolabeling with an anti-plakophilin 1 antibody and small desmosomes. These results expand the database of plakophilin 1 mutations and demonstrate the importance of this protein in the stabilization of desmosomal adhesion in terminally differentiating keratinocytes.

Adolescent↗
PubMed2000

Penetration of human metastatic melanoma cells through an authentic dermal-epidermal junction is associated with dissolution of native collagen types IV and VII.

The events occurring during the penetration of melanoma cells through the dermal-epidermal junction, which is the first crucial step in the process of metastasis, are poorly understood, partly because no suitable tissue models exist. In the in vitro model reported here, two melanoma clones (T1C3, which generates lung metastases in experimental animals, and IC8, which does not) derived from a single parental cell line were co-seeded with normal allogenic keratinocytes onto acellular human de-epidermized dermis with preserved intact basement membrane and cultured for up to 1 month at an air-liquid interface. Histological, immunohistochemical and ultrastructural studies showed that melanoma cells from the metastatic clone (T1C3), but not from the non-metastatic clone (IC8), penetrated the dermal-epidermal junction to invade the dermis after 3 weeks of culture. Local invasion was associated with the dissolution of the native epidermal basement membrane collagens type IV and VII. Confocal laser scanning microscopy analysis demonstrated that numerous T1C3 cells were able to colonize the interstitial dermis and to rapidly penetrate empty dermal cavities. Our model represents a significant technical advance over others currently available since: (i) the organized three-dimensional architecture of the native dermal-epidermal junction is preserved; (ii) the active invasion process coincides with the dissolution of native components of the epidermal basement membrane, i.e. collagen types IV and VII; and (iii) the ability of melanoma cells to cross the dermal-epidermal junction correlates with their metastatic potential. This model provides a valuable tool for the study of the time-course of the cellular and molecular events that occur during the earliest steps of invasion in cutaneous melanoma. It also offers new opportunities to study the possible role of the keratinocyte environment in melanoma invasion.

Basement Membrane↗
PubMed2000

[Acquired peeling skin syndrome in an adult].

BACKGROUND: Peeling skin syndrome is a rare form of congenital ichthyosis. The term was coined in 1982 by Levy and Goldsmith and the syndrome is clinically characterized by generalized scaling. Histologically, there is an epidermal separation in the stratum corneum. CASE REPORT: We report the case of a 73-year-old woman who had ichthyosis without cicatricial progressive alopecia since her first pregnancy. An ultrastructural study was performed confirming the clinical diagnosis of peeling skin syndrome. DISCUSSION: The peeling skin syndrome designates several different clinical entities classed by Traupe in type A and type B. Mevorah and al. expanded this classification with a type C. This classification has remained valid after additional information provided by ultrastructural studies and may suggest different pathogenic mechanisms underlying the dermatosis. A critical review of the literature shows that the case reported here is exceptional and had a late clinical onset.

Adult↗
PubMed1999

Modulation of a fibrotic process induced by transforming growth factor beta-1 in dermal equivalents.

Why initially normal wound healing sometimes shifts toward an impaired cicatrization is poorly understood. Collagen gels with incorporated fibroblasts constitute valuable in vitro models to study mechanisms of connective tissue reorganization. Such 1-week-old, partially contracted normal dermal equivalents were treated with concentrations of TGF-beta1 ranging from 1 to 10 ng/ml. The cytokine was applied in a single dose or four times at regular intervals, over a 2-week period. Dose-dependent activation of fibroblasts was observed after treatment. The cytokine induced a myofibroblastic transformation of dermal cells, significantly enhanced the process of dermal contraction, and stimulated synthesis of such proteins as cellular fibronectin, tenascin and smooth-muscle actin. Our approach is more informative than models using pathological or pretreated dermal cells, since it demonstrates newly induced modulation of fibrotic transformation in an initially normal dermal equivalent. This in vitro assay will enable us to study mechanisms involved in the shift between normal and impaired fibrotic transformation during wound healing.

Adult↗
PubMed1999

Internalization of gap junctions in benign familial pemphigus (Hailey-Hailey disease) and keratosis follicularis (Darier's disease).

Hereditary skin disorders involving acantholysis, such as Hailey-Hailey disease and Darier's disease, have been genetically linked to distinct chromosomal parts which do not code for known structural proteins. Such evidence suggests that the genomic abnormalities underlying these dermatoses may concern functional/regulatory mechanisms of keratinocyte cohesion. Epidermal communication junctions (gap junctions) are responsible for direct coupling of cells and, thus, co-ordinate the behaviour of keratinocytes within the tissue. Consequently, they remain one of the potential, and poorly studied, elements in the pathogenesis of hereditary acantholytic diseases. We have investigated the distribution and fate of gap junctions during non-immune acantholysis, using fine immunolocalization methods at the light and electron microscopic levels. Our results demonstrate normal expression of epidermal gap junction proteins, connexins 26 and 43, in non-lesional skin of Hailey-Hailey and Darier's diseases. The gap junctions were not primarily dismantled during acantholysis, typical of both of the studied dermatoses, but underwent internalization and subsequent cytoplasmic dispersion in the portions of cells which were no longer attached to the rest of the tissue. In Darier's disease, perifollicular acantholysis did not specifically concern epithelium of appendages coexpressing connexin 26 in addition to connexin 43, further indicating that the observed changes in gap junction localization were secondary to the loss of cell-cell contact. We demonstrated that the sequence of changes was identical in both diseases and that the previously described putative differences were apparently related to the degree of acantholysis present in the studied biopsies. The fate of the junctional structures and proteins, documented in the present study, is most probably a form of recycling process also used by normal keratinocytes during organogenesis and tissue differentiation.

Acantholysis↗
PubMed1998

Stratum corneum, corneodesmosomes and ex vivo percutaneous penetration.

The stratum corneum is composed of cornified keratinocytes, joined together with corneodesmosomes and embedded in the relatively hydrophobic intercellular substance. Formation of this horny layer and its constant desquamation are fundamental processes leading to the establishment of an efficient epidermal barrier. We examined structural changes occurring in the stratum corneum ex vivo, in order to better understand the mechanisms regulating corneocyte desquamation at the epidermal surface and influencing percutaneous penetration of exogenous substances. Morphologic modifications were induced by occlusion, increasing the tissue hydration, or by application of propylene glycol, a hydrophilic solvent of small molecular size. Distribution of the major epidermal antigens, markers of terminal differentiation, was studied immunohistochemically and showed no modification related to the tissue alteration. Skin samples were fixed in paraformaldehyde and either postfixed in OsO4 and embedded in Epon or postfixed in RuO4 and embedded in epoxy and acrylic resins. Structural composition and spatial organization of the elements present in the intercorneocyte spaces were evaluated ultrastructurally, with special attention paid to the fate of corneodesmosomes. Our results suggest that the spontaneous organization of lipids excreted into the intercellular spaces and constitution of the lamellar multilayers in the lower stratum corneum are at the origin of partition of the intercellular compartment to the hydrophobic and hydrophilic domains. The latter, compatible with the proteinic contents, seem to be displaced by the hydrophobic components undergoing reorganization, towards the invariable hydrophilic foci represented by corneodesmosomes. This mechanism may be involved in the delivery of proteolytic enzymes, thus contributing to the progressive degradation of corneodesmosomal proteins.

Desmosomes↗
PubMed1998

Expression cloning of human corneodesmosin proves its identity with the product of the S gene and allows improved characterization of its processing during keratinocyte differentiation.

In human epidermis and other cornified squamous epithelia, corneodesmosin is located in the desmosomes of the upper living layers, and in related structures of the cornified layers, the corneodesmosomes. During maturation of the cornified layers, the protein undergoes a series of cleavages, thought to be a prerequisite of desquamation. Partial amino acid sequencing of corneodesmosin fragments suggested that it is related to the product of the S gene, previously identified in the human major histocompatibility complex. We report the expression cloning of corneodesmosin cDNA from a human epidermis library screened with monoclonal antibodies. Sequencing demonstrated that corneodesmosin is really the product of the S gene. However, analysis of 20 alleles of the gene revealed that its product is 27 amino acids longer than initially reported. Two additional polymorphic sites were described, and the position of the unique intron was ascertained. Corneodesmosin cDNA expression in COS-7 cells led to secretion of the protein. Precise epitope mapping allowed further characterization of the molecular forms of corneodesmosin present in the most superficial cornified layers, where fragments corresponding to the central region of the protein were detected. This indicated a cleavage of the N- and C-terminal domains of corneodesmosin before desquamation. These serine- and glycine-rich domains are proposed to mediate an adhesive function.

Amino Acid Sequence↗
PubMed1997

Hyperphosphorylation of beta-catenin on serine-threonine residues and loss of cell-cell contacts induced by calyculin A and okadaic acid in human epidermal cells.

Phosphorylation and dephosphorylation events may critically control junction assembly and stability, as well as regulate the formation of the cadherin-cytoskeleton complex, thus influencing the adhesive function of cells. In the present study, we have used specific activators and inhibitors of protein kinases and phosphatases to analyze the role of protein phosphorylation in the maintenance of epithelial architecture. Okadaic acid and calyculin A cell treatments induced two major effects: a dramatic alteration of the keratin network of epidermal cells and a complete disruption of cell-cell contacts. This loss in cell-cell contacts was not tissue and species restricted and the interactions of keratinocytes with the matrix were not involved. The observed changes were highly specific for these drugs and were obtained in the range of concentrations corresponding to the inhibition of protein phosphatase 1 (PP1). They were time- and dose-dependent, and reversible, excluding a cytotoxic effect of the drugs. A decrease in electrophoretic mobility of beta-catenin, a major protein involved in the regulation of intercellular adherens junctions, was observed in keratinocytes and fibroblasts treated with okadaic acid and calyculin A, suggesting a change in the protein phosphorylation level and/or protein conformation. Data from beta-catenin immunocomplex autoradiography performed after 32P in vivo incorporation in untreated and okadaic acid or calyculin A-treated HaCaT cells, demonstrated a higher level of phosphorylation of beta-catenin in treated cells compared to untreated ones. Analysis of 32P-labeled phosphoaminoacids demonstrated that beta-catenin was exclusively phosphorylated on serine-threonine residues but not on tyrosine residues. Immunoprecipitations and Western blotting using anti-phosphoserine and anti-phosphotyrosine antibodies confirmed these data. The change in beta-catenin phosphorylation on serine-threonine residues may play a role in the control of the cohesion between epithelial cells and may be involved in the regulation of the transduction signal.

Apoptosis↗