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J P Molès

Publications and source records attributed to J P Molès.

18 recordsLinked to original sources
PubMed2004

Differential expression of a human endogenous retrovirus E transmembrane envelope glycoprotein in normal, psoriatic and atopic dermatitis human skin.

BACKGROUND: Psoriasis is a common inflammatory skin disease characterized by uncontrolled proliferation of keratinocytes and recruitment of T lymphocytes into the skin. The possible role of human endogenous retroviruses (HERVs) in the induction of psoriasis has been suggested, based upon the previous observations of retrovirus-like particles in psoriasis from skin lesional plaques, urine and stimulated lymphocytes. OBJECTIVES: To investigate the expression of HERV-E transmembrane envelope glycoprotein (HERV-E env) in normal, psoriatic and atopic human skin, and to examine the influence of ultraviolet (UV) B irradiation on HERV-E env expression in normal human epidermal keratinocytes. METHODS: The analysis was performed on both skin biopsies and organotypic skin cultures using immunofluorescence and Western immunoblotting. UVB irradiation (312 nm) of cultured normal human keratinocytes was performed using a dose of 30 mJ cm(-2). RESULTS: Positive staining was observed in most of the psoriatic and atopic skin samples, whereas only 15% of the normal skin samples were faintly positive. In addition, the pattern of expression of HERV-E env differed markedly in psoriasis vs. atopy. By Western blotting analysis, two main proteins of 54 and 57 kDa were detected in extracts of normal skin, normal keratinocyte cultures and reconstructed epidermis from psoriatic and normal punch biopsies. An increased level of expression of these proteins was noted in extracts from psoriatic vs. normal reconstructed epidermis. The overexpression of the 57-kDa protein in normal human cultured keratinocytes was dramatically reduced by UVB irradiation. CONCLUSIONS: These data suggest for the first time that HERV-E env is expressed in normal and pathological human skin. Further studies are now required to elucidate the role of such viral proteins in the pathogenesis of psoriasis.

Adolescent↗
PubMed2000

A three-dimensional skin culture model for mouse keratinocytes: application to transgenic mouse keratinocytes.

The study of mouse epidermal biology has been hampered by the lack of a good in vitro model for the culture of mouse keratinocytes which allowed the reconstruction of a fully differentiated epidermis. We adapted the Pruniéras' model, also called the Dead de-Epidermized Dermis model (DED), to mouse keratinocytes and showed that a neo-epidermis can be reconstructed exhibiting a complete differentiation program. We also used this model to culture transgenic mouse keratinocytes. We observed that transgene expression occurred in the correct location and that the neo-epidermis mimed previous in vivo observations obtained with integrin skin-targeted transgenic mice. Therefore, this model will be a powerful tool to further investigate normal mouse and transgenic keratinocyte biology.

Animals↗
PubMed1999

P16 UV mutations in human skin epithelial tumors.

The p16 gene expresses two alternative transcripts (p16alpha and p16beta) involved in tumor suppression via the retinoblastoma (Rb) or p53 pathways. Disruption of these pathways can occur through inactivation of p16 or p53, or activating mutations of cyclin dependant kinase 4 gene (Cdk4). We searched for p16, Cdk4 and p53 gene mutations in 20 squamous cell carcinomas (SSCs), 1 actinic keratosis (AK), and 28 basal cell carcinomas (BCCs), using PCR-SSCP. A deletion and methylation analysis of p16 was also performed. Six different mutations (12%) were detected in exon 2 of p16 (common to p16alpha and p16beta), in five out of 21 squamous lesions (24%) (one AK and four SCCs) and one out of 28 BCCs (3.5%). These included four (66%) ultraviolet (UV)-type mutations (two tandems CC : GG to TT : AA transitions and two C : G to T : A transitions at dipyrimidic site) and two transversions. P53 mutations were present in 18 samples (37%), mostly of UV type. Of these, only two (one BCC and one AK) harboured simultaneously mutations of p16, but with no consequence on p16beta transcript. Our data demonstrate for the first time the presence of p16 UV induced mutations in non melanoma skin cancer, particularly in the most aggressive SCC type, and support that p16 and p53 are involved in two independent pathways in skin carcinogenesis.

Alternative Splicing↗
PubMed1999

[Beta-catenin mutations in a common skin cancer: pilomatricoma].

The normal hair development requires WNT signalling pathways. A constitutive activation of beta-catenin, one of the components of this cascade, induces an abnormal proliferation of hair matrix cells. This activation is observed in a human skin tumours called pilomatricoma. Mutations of the beta-catenin gene are detected in 75% of the tumours analysed.

Animals↗
PubMed1998

[The gene of Gorlin's syndrome (basocellular nevomatosis) and the revival of the developmental genes in human carcinogenesis].

Interaction between developmental biology and human pathology has brought new insight on basal cell carcinoma oncogenesis. Alterations of the Patched gene appear to be responsible for the Gorlin syndrome and to be detected in 50% of the sporadic basal cell carcinomas. This relation has recently been extended to all the components of the Hedgehog signalling pathway.

Basal Cell Nevus Syndrome↗
PubMed1997

Correlation between hyperproliferation and suprabasal integrin expression in human epidermis reconstituted in culture.

In normal epidermis integrin expression is largely confined to the basal layer. However, during wound healing and in psoriatic lesions suprabasal expression is observed. Although the potential importance of suprabasal integrin expression in the pathogenesis of psoriasis has been established, the cause of suprabasal expression is unknown. We now describe changes in integrin expression that occur with time when normal human keratinocytes are grown on two types of dermal equivalent, de-epidermized dermis and collagen gels containing fibroblasts. We show that suprabasal integrin expression is correlated with suprabasal expression of the EGF receptor, but not with expression of keratin 10 or keratin 16. By quantitating the proportion of basal keratinocytes expressing the proliferation marker Ki-67 we could show that suprabasal integrin expression is correlated with high proliferative activity within the basal layer. Taken together with our earlier work, these results suggest that suprabasal integrin expression is linked to hyperproliferation and not to abnormal terminal differentiation or to inflammation; they also establish dermal equivalent cultures as useful experimental models with which to manipulate keratinocyte integrin expression.

Antibodies, Monoclonal↗
PubMed1997

The epidermal stem cell compartment: variation in expression levels of E-cadherin and catenins within the basal layer of human epidermis.

The basal layer of the epidermis contains two types of proliferating keratinocyte: stem cells, with high proliferative potential, and transit amplifying cells, which are destined to undergo terminal differentiation after a few rounds of division. It has been shown previously that two- to three-fold differences in the average staining intensity of fluorescein-conjugated antibodies to beta 1 integrin subunits reflect profound differences in the proliferative potential of keratinocytes, with integrin-bright populations being enriched for stem cells. In the search for additional stem cell markers, we have stained sections of normal human epidermis with antibodies to proteins involved in intercellular adhesion and quantitated the fluorescence of individual cell-cell borders. In the basal layer, patches of brightly labeled cells were detected with antibodies to E-cadherin, beta-catenin, and gamma-catenin, but not with antibodies to P-cadherin, alpha-catenin, or with pan-desmocollin and pan-desmoglein antibodies. In the body sites examined, palm and foreskin, integrin-bright regions were strongly labeled for gamma-catenin and weakly labeled for E-cadherin and beta-catenin. Our data suggest that there are gradients of both cell-cell and cell-extracellular matrix adhesiveness within the epidermal basal layer and that the levels of E-cadherin and of beta- and gamma-catenin may provide markers for the stem cell compartment, stem cells expressing relatively higher levels of gamma-catenin and lower levels of E-cadherin and beta-catenin than other basal keratinocytes.

Cadherins↗
PubMed1996

1,25-Dihydroxyvitamin D3 and its synthetic derivatives MC903 and EB1089 induce a partial tumoral phenotype reversal in a skin-equivalent system.

The antitumoral potency of 1,25-dihydroxyvitamin D3 and its synthetic derivatives MC903, EB1089, and KH1060 was investigated on a tumoral Bowen-like epidermis reconstructed from an immortalized human keratinocyte cell line transfected by expression vectors coding for E6 and E7 of human papillomavirus 16. Treatment of skin equivalents by vitamin D derivatives (10(-9) M or 10(-12) M) was performed during (from day 1 to day 15 of culture) or after tissue reconstruction (from day 15 to day 30). Pharmacologic effects were evaluated by morphologic and immunohistologic analysis and compared with those of controls (vehicle alone) and with treatment of skin equivalents derived from normal keratinocytes. When performed during epidermal reconstruction, treatment of tumoral skin equivalents induced only minor morphologic and immunohistologic changes. Conversely, when performed after epidermal reconstruction, treatment with 1,25-dihydroxyvitamin D3, MC903, and EB1089 clearly improved the phenotype of treated tissues. Morphologic analysis showed reorganization of epidermal layers with the appearance of a distinct basal layer and of stratified orthokeratotic stratum corneum. Immunohistochemical analysis demonstrated that the terminal differentiation markers profilaggrin and cytokeratin 10 were re-expressed in the treated tissues while absent in controls. Overall, the results indicate that 1,25-dihydroxyvitamin D3, MC903, and EB1089 can induce a partial reversion of the tumoral phenotype in this in vitro model.

Antineoplastic Agents↗
PubMed1994

p53 immunohistochemical analysis in breast cancer with four monoclonal antibodies: comparison of staining and PCR-SSCP results.

The expression of p53 protein was examined in a series of 136 primary breast carcinomas, 106 of which were analysed with a panel of four monoclonal antibodies (MAbs 1801, 240, DO7 and DO1). p53 expression was detected with at least one antibody in 40 tumours (38%), whereas only 15 tumours (14%) were positive with all four antibodies. Some variability in the immunostaining could be observed depending on the antibody used. This was noticeable both for the number of positive cells within a section and for the intensity of staining. We therefore selected a panel of 17 tumour sections (nine were highly positive, three with medium to low staining and five with low to negative staining), which we analysed by polymerase chain reaction-single-strand conformation polymorphism analysis (PCR-SSCP) for the presence of a p53 mutation at the molecular level. Mutations were identified in 15 cases. Therefore the proportion of p53-stained cells does not seem to be an exact representation of the number of cancer cells bearing a mutation within a tumour. A statistically significant correlation was observed between p53 expression, regardless of the number of positive antibodies, and grade III disease (P < 0.0001), oestrogen (P < 0.0001) or progesterone receptor negativity (P = 0.0061), increased Ki 67 index (P = 0.0018), epidermal growth factor receptor (EGFR) positivity (P = 0.0076) and aneuploidy (P = 0.037). No correlation was observed with tumour size or lymph node involvement. In univariate analysis p53 expression was not correlated with disease-free survival, in contrast to the classical prognostic parameters, which were statistically correlated. In this series p53 expression was not a marker of early recurrence.

Antibodies, Monoclonal↗
PubMed1994

Comparative analysis of normal and psoriatic skin both in vivo and in vitro.

The morphological and biochemical characteristics of psoriasis are well documented, but the pathogenesis of this disease is not clearly understood. A variety of in vitro models of psoriasis have been developed in attempts to identify the trigger factors, but no model so far reproduces the stable psoriatic phenotype accurately. In the present work, we initially checked the immunohistochemical distribution of proliferation/differentiation markers in psoriatic skin in vivo, and our results largely confirm previously reported data. However the study was performed using a new series of monoclonal antibodies to keratin. Subsequently we took normal or psoriatic skin biopsies, reconstructed skin equivalents using a recently described model and analysed the proliferation/differentiation status of the resulting epidermis. Dramatic morphological and antigenic differences were found between normal and psoriatic skin in vivo, but whatever the source of the initial biopsy, a unique in vitro phenotype was obtained in the reconstructed epidermis. This phenotype was marked by mild hyperproliferation and an altered distribution of differentiation-associated antigens suggesting a need for extracutaneous stimuli to maintain the psoriatic phenotype in vitro.

Antibodies, Monoclonal↗
PubMed1994

TP53 tumor suppressor gene and skin carcinogenesis.

The tumor suppressor gene TP53 encodes for a nuclear phosphoprotein involved in the control of cell proliferation, particularly in stressed cells. TP53 gene mutations are the most frequent genetic event found in human cancers. Most mutations locate in the highly conserved domains of the gene. Their localizations vary according to the tissue and tumor type, but define some hot spot regions that may have a certain degree of tissue specificity. In certain cases, the type of nucleotide substitutions observed can help to find the carcinogenic agent. In recent years, TP53 gene mutations have been frequently observed in human skin tumors. In epithelial carcinomas, they involve mainly exons 5, 7, and 8. Interestingly, many are C to T transitions at dipyrimidine sites; particularly, one can find CC to TT double-base changes that are known to be specific to ultraviolet radiation. These data confirm at the molecular level the role of ultraviolet radiation as an important etiologic factor in the genesis of these lesions. The high incidence of TP53 mutations suggest that they play a role in keratinocyte transformation. Nevertheless, this event has not yet been defined as an early or late event. In melanomas, most studies have shown the detection of the p53 protein by immunohistochemistry, suggestive of the presence of a mutation in the gene prolonging the protein half-life. Anti-p53 reactivity is frequent in these tumors and seems to correlate with tumor aggressiveness. Confirmation and characterization of TP53 gene mutation at the DNA level would help to precisely define the role of this gene in the development of these tumors.

Genes, Tumor Suppressor↗
PubMed1994

Analysis of HPV16 E6 and mutant p53-transfected keratinocytes in reconstituted epidermis suggests that wild-type p53 inhibits cytokeratin 19 expression.

Using a reconstituted skin culture model we have analysed the effects of oncogenic human papillomavirus (HPV) and mutant TP53 genes on the proliferation and differentiation of human keratinocytes. Immortal cell lines generated by transfection of early passage normal human keratinocytes with HPV16 E7 plus mutant human TP53 (KN #1), HPV16 E7/E6 (KN #2), or HPV16 E7 plus murine p53 (KN #3) were examined. KN #1 and KN #2 behaved identically, reconstructing a tumor-like epidermis characterized by the lack of differentiation and the presence of an aberrant epidermal architecture. In contrast, KN #3 reconstructed an epidermis that was more similar to that obtained with normal keratinocytes. KN #1 and KN #2 were further characterized by the inversion of the proliferative compartment and the abnormal expression of cytokeratin 19 (CK19). Because p53 function is reduced in these cells, either by heterocomplex formation between endogenous wild-type p53 and transfected mutant p53 or by E6-induced degradation of wild-type p53, we hypothesized that CK19 expression may be normally repressed by wild-type p53. This hypothesis was supported by the strict correlation observed between TP53 mutation and CK19 expression in a set of human skin tumors. CK19 was detected in all eight carcinomas containing a mutated TP53 gene but in none of the 16 carcinomas containing only wild-type TP53. These results illustrate the utility of the in vitro reconstituted skin model for investigating the consequences of genetic alterations in human keratinocytes.

Cell Differentiation↗
PubMed1993

Mutation of the tumor suppressor gene TP53 is not detected in psoriatic skin.

We investigated the tumor-suppressor gene, TP53, in psoriatic skin at gene (GP53) and protein (p53) levels. No mutation was detected in the seven exons analyzed using a combination of polymerase chain reaction and single-strand conformation polymorphism techniques. In addition, by immunohistochemistry using three different anti-p53 antibodies, no positive staining, reported to be associated with mutations and/or abnormal expression of the TP53 gene, was observed in psoriatic skin biopsies, in contrast to recent reports. These results suggest that the proliferative status of psoriatic keratinocytes does not implicate the TP53 gene.

Base Sequence↗
PubMed1993

TP53 tumor-suppressor gene and human carcinogenesis.

The wild-type tumor-suppressor TP53 gene encodes for a nuclear protein which has been shown to act as a transcriptional modulator. The cellular role of the p53 protein is the control of cell proliferation, particularly important in stressed cells. The TP53 gene is frequently mutated in sporadic and familial human cancers. Most transforming mutations localize in highly conserved domains of the gene and define hot-spot regions that have a certain degree of tissue specificity. Moreover, most mutations are point mutations and the type and localization of the nucleotide substitution may sometimes help in recognizing the carcinogenic agent. This is the case for C to T transitions at dipyrimidine sites induced by UV radiation in cutaneous epitheliomas. Inactivation of p53 protein can also occur through mechanisms other than genetic alteration, such as binding to viral or cellular proteins. Loss of wild-type TP53 function seems therefore to play a crucial role in cell transformation in human cancers, either during carcinogenesis or later in tumor progression.

Cell Division↗
PubMed1993

p53 gene mutations in human epithelial skin cancers.

In the present study we analysed 38 epithelial skin cancers, 19 basal cell carcinomas (BCCs), 13 squamous cell carcinomas (SCCs) and six Bowen diseases (BwDs), using a combination of polymerase chain reaction (PCR) and single-stranded conformation polymorphism (SSCP) techniques for the presence of p53 and RAS gene mutations. Whereas 48% (9/19) of the BCCs tested presented a mutated p53 gene, the frequency was lower (15%, 2/13) in our series of SCCs and negative in the BwDs. Nine of the 11 characterized mutations were single-nucleotide substitutions and, interestingly, seven of these involved CC dimers, where a C was changed into a T or a G (three C-->T transitions and four C-->G transversions). This mutational pattern, added to the fact that all the mutated tumors occurred at sun-exposed body sites, implicates UV light in their genesis. Furthermore, we observed two internal deletions of 6 and 24 bp whose flanking sequences contained two or three Cs on either strand. In addition to molecular detection, we searched for p53 protein accumulation, by immunocytochemical staining, in a subset of 23 epithelial skin tumors (nine bearing a mutation, 14 which scored negative in our assay). Three commercially available anti-p53 antibodies (PAb CM1, mAbs DO7 and 1801) were used, and 3/23 (all showing a mutated p53 gene) presented specific nuclear staining. In contrast to other reported data we could not detect any activating RAS gene mutation in our series of human skin cancers.

Base Sequence↗
PubMed1992

p53 and RAS gene mutations in multiple myeloma.

We analysed genomic DNA from 30 patients with multiple myeloma (MM), searching for alterations in the p53 and RAS genes by a combination of polymerase chain reaction and single-strand conformation polymorphism techniques. Mutations in the p53 gene were observed in 20% (6 out of 30) of the patients, and were located in conserved sequence blocks within exons 5 and 7. These were single-nucleotide substitutions and consisted predominantly (4/6) of G:C to A:T transitions. Of the six patients with a mutated p53 gene, four were in the terminal phase of the disease. RAS gene mutations were found more frequently since they occurred in 47% (14 out of 30) of the patients. Mutations consisted of single-nucleotide substitutions, located in codons 12, 13 and 61 of either K- or N-RAS, to the exclusion of H-RAS. Moreover, one patient bore two simultaneous mutations, affecting simultaneously the K- and the N-RAS genes. RAS gene mutations were more frequently observed in patients with fulminating disease (10/15, 67%) than in patients with less aggressive forms of the disease (4/15, 26%). We also analysed genomic DNAs from 10 human myeloma cell lines, of which two bore mutations affecting codon 12 of the K-RAS gene, and one codon 12 of the N-RAS gene. The first two cell lines were obtained from freshly explanted tumor cells in which we observed identical mutations. Results presented here show that activating mutations in the RAS genes are, in MM, more frequent than those affecting the p53 gene and suggest that both events are related to terminal phases of the disease.

Amino Acid Sequence↗
PubMed1991

C-fos and c-jun proto-oncogene expression is decreased in psoriasis: an in situ quantitative analysis.

Psoriasis is a common, sometimes severe, non-malignant skin disease characterized by hyperproliferation and abnormal differentiation of keratinocytes. Because proto-oncogenes are implicated in both cell proliferation and differentiation, their expression could be modified in skin diseases such as psoriasis. The c-fos and c-jun proto-oncogenes, whose products associate to form a heterodimeric transcription factor, are among the first genes to be expressed when certain cells are stimulated to either proliferate or differentiate. Recent studies in our laboratory have shown that the c-fos proto-oncogene is highly expressed in normal human adult skin. In the present study, we used in situ hybridization with RNA to compare the expression and localization of c-fos and c-jun transcripts in 15 lesional and non-lesional psoriatic skin samples. Two clinical variants of psoriasis were studied: the most severe and chronic form or plaque-type psoriasis (N = 10) and rapidly resolutive guttate-type psoriasis (N = 5). Quantitative analysis was performed using a semi-automatic image analyzer and the "Starwise grain" software program. Our control samples included 10 normal skins and eight specimens from other benign hyperproliferative non-psoriatic skin diseases, consisting of three with inflammation (seborrheic dermatitis and atopic dermatitis), and 5 without inflammation (seborrheic keratoses). Control genes we used for in situ hybridization and RNA integrity were keratin 14, which is expressed in the epidermis and was normally expressed in all tissue analyzed, and ribosomal RNA. Our data showed that c-fos and c-jun were expressed to an equivalent extent, both spatially and quantitatively, in all specimens tested. Expression was significantly decreased in plaque-type but not in guttate-type psoriasis. It was also decreased in the three other benign inflammatory cutaneous hyperproliferative disorders, but not in the five non-inflammatory cases. These results were surprising because hyperproliferation was here associated with a decrease in proto-oncogene expression, thus suggesting that c-fos and c-jun do not play a crucial role in the control of keratinocyte proliferation in vivo. However, their reduced expression in some abnormally differentiated skins indicates that both c-fos and c-jun proto-oncogenes may play a key role in keratinocyte differentiation. Their altered expression correlated with severity of the disease and the presence of an inflammatory infiltrate. These data offer a new insight into the role and regulation of these proto-oncogenes in vivo in humans.

Chronic Disease↗