In mammalian melanocytes, melanin synthesis is controlled by tyrosinase, the critical enzyme in the melanogenic pathway. We and others showed that the stimulation of melanogenesis by cAMP is due to an increased tyrosinase expression at protein and mRNA levels. However, the molecular events connecting the rise of intracellular cAMP and the increase in tyrosinase activity remain to be elucidated. In this study, using B16 melanoma cells, we showed that cAMPelevating agents stimulated mitogen-activated protein (MAP) kinase, p44mapk . This effect was mediated by the activation of MAP kinase kinase. cAMP-elevating agents induced a translocation of p44 mapk to the nucleus and an activation of the transcription factor AP-1. cAMP-induced AP-1 contained FOS-related antigen-2 in association with JunD, while after phorbol ester stimulation AP-1 complexes consist mainly of JunD/c-Fos heterodimers. In an attempt to connect these molecular events to the control of tyrosinase expression that appears to be the pivotal point of melanogenesis regulation, we hypothesized that following its activation by cAMP, p44 mapk activates AP-1. Then AP-1 could stimulate tyrosinase expression through the interaction with specific DNA sequences present in the mouse tyrosinase promoter.In melanocytes and melanoma cells, melanin synthesis is controlled by a cascade of enzymatic reactions regulated at the level of tyrosinase. This enzyme synthesizes dopaquinone from tyrosine and appears to control the rate-limiting step of melanogenesis. Melanin synthesis is stimulated by a large array of effectors including 1-oleyl-2-acetyl-glycerol (1), ultraviolet B radiations (2), and cAMP-elevating agents (forskolin, IBMX, 1 ␣-MSH) (3-5). Few data are available concerning the molecular mechanisms triggered by these melanogenic agents. Protein kinase C was thought to be involved in the induction of melanogenesis by 1-oleyl-2-acetyl-glycerol and ultraviolet B radiations (6, 7). However, a recent report of Carsberg et al. (8) has shown that the stimulation of melanogenesis by these agents was not affected by RO485, a potent inhibitor of protein kinase C. While the role of protein kinase C in the induction of melanogenesis remains controversial, compelling data have shown that cAMP-elevating agents stimulate melanogenesis in both melanocytes and melanoma cells, indicating that the cAMP pathway plays a key role in the regulation of melanogenesis (3-5). The effect of cAMP on melanogenesis is due to a stimulation of tyrosinase activity. This appears to be the consequence of an augmentation of enzymatic activity of preexisting tyrosinase (4, 9) following post-translational modifications such as (i) phosphorylation or glycosylation (10), (ii) association with an activator (11, 12), and (iii) dissociation from an inhibitor (13). Alternatively, cAMP was shown to increase tyrosinase mRNA (14, 15), resulting in an augmentation of tyrosinase amount, suggesting that cAMP stimulates tyrosinase transcription (16). However, the molecular events connecting the stimulation of t...

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Herlitz junctional epidermolysis bullosa (H-JEB

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Gambierdiscus toxicus has been described as the chief producer of ciguateric toxins: ciguatoxin (CTX) and maitotoxin (MTX). Dr. R. Bagnis (Malarde Institute, Tahiti) provided us with a strain of this benthic dinoflagellate in 198 1 to study its cytological, physiological, and toxicological characteristics.The growth of G. toxicus has been studied under various chemical and physical conditions enabling us to define optimal culture conditions. Since then, we have improved the cultivation procedure and have obtained large scale cultures of some clones and strains in different culture media.All the experiments showed that G. toxicus has complex nutritive requirements and a large inertia to response at some non-drastic environmental variations. Increased growth rates were observed when cultures were treated with antibiotics.The classical extraction procedure of the toxins has been adapted to our algal material. The method was simplified and resulted in enhanced toxinic yield.Our principal results have demonstrated that G. toxicus in culture remained very toxic; 600 to 2000 cells were sufficient to kill a 20 g female mouse within 24 hours (MLD = 1-3 mg/kg). No variation of the degree of toxicity has been observed for three years.A linear relationship between the number of G. toxicus cells (Coulter counted) and the weight of corresponding algal pellet has been found. This leads to an easier evaluation of the quantity of toxic extract required to calculate the minimum lethal dose (MLD).The extraction procedure results in two toxic fractions: a water-soluble (MTXlike) and a lipid-soluble (CTX-like). The latter corresponds to 10 to 25% of the total toxicity.A dose-time to death curve has been established with our CTX-like extracts.

Fertilization includes sperm-oocyte recognition, adhesion, binding, fusion and egg activation. Integrin receptors, which are adhesion molecules, are expressed on sea urchin, mouse, hamster and human unfertilized oocytes. Potential sperm ligands have been identified. A role for integrins during fertilization is supported by inhibition of sperm-egg adhesion and/or fusion by means of anti-integrin monoclonal antibodies, Arg-Gly-Asp (RGD) or disintegrin-like peptides. In several cell-cell interactions, such as lymphocyte activation, viral fusion, bacterial infection or macrophage phagocytosis, integrins act as co-receptors after activation and, by clustering in a multimolecular complex, are able to transduce signals through cytoskeletal proteins and adaptor kinases. Experimental data suggest that they may act in a similar way during fertilization and may participate to initiation and/or propagation of the calcium signal via stimulation of phospholipase C gamma and inositol trisphosphate production.

The extensive growth of Caulerpa taxifolia in the Mediterranean sea produces important quantities of bioactive secondary metabolites unable to enter the food chain. The cytotoxic effects of caulerpenyne, the major secondary metabolite from C. taxifolia, was studied in different in vitro models: skin cells, primary cultures of melanocytes and keratinocytes, immortalized keratinocytes (HaCaT and HESV), and bone marrow cells (hematopoietic progenitors CFU-GM). Typical dose-response curves from neutral red uptake and MTT assays were recorded in all models with IC50 ranging from 6 to 24 microM. Hematopoietic progenitors were more sensitive to caulerpenyne than melanocyte and keratinocyte cell lines, which could be due to their higher proliferative rate. The distribution of aggregates in colonies, macroclusters, and microclusters of hematopoietic progenitors was also altered in the presence of caulerpenyne. From our evaluation of the caulerpenyne concentrations required to result in cellular toxicity, the risks of cutaneous and/or food intoxication to humans may be considered minimal.