88 Absrrwrs JSlDiCSID Joint Merritzg

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CONTRBUTION OF XMRK, A NOVEL RECEPTOR TYROSINE KIN&E GENE, IN PlGMENT CELL AND MELANOMA FORMATION IN THE XPHOPHORVS MELANOMA MODEI. BW WooIc~ck. BM Schmidt, JR Vlelkind BC Cancer Research Centre and Dept. of Pathology, Universly oi EC, Vancouver. EC. Canada.

The Xrphophoms fish melanoma mode, is welt recognized as a mode, for the study of human melanoma Melanomas in thwe.fash are mediated by two major gene% IOCI. I) a” oncagenic sex-linked locus contains information ior the development of pwnent cells, localization of these) cells and melanoma susceptibd~ty, and ii) an autosoma, locus pmmoles pigment cell differentiation and behaves as a tumour suppressor Iocus, ie in wderthat fully makgna”, melanomas can form both wp,es of tha gene must be lost. We and others have cloned a novel tyrosine klnase receptor gene which belongs to the famdy of epaemal growth factor receptor genes and maps lo the ancqenic sex-llnked locus. The gene. X&-Z. is a duplicate version of another wtually Identical gene which IS also located on the sex chromosome, the fwo genes d~iier wth regard to their pmm”ter regmns We present evidence that Xmrk-2 is “ecessaly for melanoma formah”“: I) Xmrk gene expression 1s deregulated I” melanomas. ii) is pwbvely correlated with degree of sevwty of melanoma, and IS) IU expression decreases once melanoma growth ceases and 1s ~ImoSt non-endent in regressive melanomas TO further elwdate the iuncho” of this gene we have studied Xmrk-2 from three pigment cell mutants. I” two mutants pylment cells are not io”“ed. I” one the Xmrk.2 is deleted wh,le I” the other mutant 8 pm”, mutation msuits m a” ammo aad subsbtution I” the k1”ase doma!” which might abrogate signal transduction In the third mutant pigment cell formation is ectopic and premalure Here we found a t5bp insed msuking in a 5 leucine residue I” the signal peptlde of the Xmh protein. These data suggest that the gene is required for pigment cell davelopment Funher functional analysis by transiechng the latter Mu mutant XmrX genes into a fish cell ,h”e should help to dariiy the effects “‘these “Utah”“* 0” the function of Xmrk prote,”

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A NEW MODEL OF IN VIVO CELL PROLIFERATION OF HUMAN CUTANEOUS T-CELL LYMPHOMA IN SEVERE COMBINED IMMUNODEFKIENT MICE. hnura. A..‘# Takaon-Kondo.A..* lmada, K..*

Previous attempts to propagate the human cutaneous T-cell lymphoma (CTCL) cells in animals hove been unsuccessful except for a report by skin grafting. We have ma& an in wvo cell proliferation model of CTCL cells in severe combined immunodeficient (SCID) mice. SC[D mice were intraperitoneally injected with CTCL cells from ski” tumors or lymph nodes of 6 CTCL patients (4 mycosis fungoides, I cutaneous Kt- I lvnmhoma and I S&w svndmtre). lnlected CTCL cells from 3 of 6 patients proliferated _,.._ I. -.. .~ , I _ and infilrrated into a variety of murine organs including skin, 4 to 7 weeks after the inoculation of the cells. The cells proliferated and recovered from the mice expressed the sa,,c cell surface phenotype and had the same realTa”geme”t patterns of T-XII receptor ,?

and y chain genes as those of the original CTCL cells when examined by FACScan analysis and Southern blot hybridization. indicating that the cells proliferating in SCID mice were derived fnxn the original CTCL clone. Furtlwnwre. the histologic examination showed the localized intiltmtion of CTCL cells in murine epidermis which was quite similar to Pauttier’s microabscess. The animal model of in viw cell proliferation of human CTCL cells we hove developed will ix very useful for studying the mechanism of neoplastic cell growth m viva and also provide us new therapeutic approaches to ove~onx the disease

MOLECULAR GENETIC APPROACHES TO THE STUDY OF WOUND HEALING MECHANISMS IN THE SKIN OF TRANSGENIC MICE m Dept. Of BiologIcal Chemistry and Dermatology. The Johns Hopkins Universuy School of Medicine, Baltimore, MD 21205, U3.A

Reepithelialization of fuU-thiclmess skin wounds(wound closure) occurs via two major mechanisms : wound contraction, mediated by demml tibroblasu, and inward migration of epithelia, cells orgmating from the edges of the wound. The relalive contribution of these two mechanisms depend on parameters such as site, nature, and extent of injury, and a host of physiological factors. The signals responsible for the recruitment of keratinccyta irom wound edge tissue as well as the mechanisms of epithelial sheet migration into the wound site after skin injury remain largely unknown. We identilied a group OC genes whose products. keratio 6.16 and 17(K6.K16 and K17). are induced in keratinocytes at the wound edge within hours after skin injury. well before the onset of migration into the wound site. We are using these keratin genes as a paradigm 10 study the mechanisms regulating gene expression at rhe wound edge, the reprogramming of keratinwyte fate from a terminally differentwing to a migratory mode, and the cellular mechanisms of epitbelial sheet migration. Our studies take advantage of tie transgenic mouse technology, which offers the possibility of manipulating genes and assess the consequences in a genuine skin environment in viva This research has profound implications for our understanding various a.spats of skin biology. when normal, injured, or dii. as well as for the development of gene therapy strategies. Supported by grants from the National Institutes of Health, American Cancer Society, and Dermatology Foundation.

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DECREASE IN CELL MEMBRANE DAMAGE AND IN INTERLEUKIN-la MESSAGE IN CULTURED KERATINOCYTES TREATED WITH GREEN TEA POLYPHENOLS. GsbuL E 2 ‘v” ielinska. S Kondo. Division of Dermatology, Sunnybrook Health Science Cent% Univenily of Toronto. Toronto. Canada and Case Western Reserve University, Cleveland, Ohio

The use of naturally occurring botanic& in the treatment of a wide variety of human diseases has recently achieved 8 significant amoum of interest. Anecdotal ewdence suggests that one such botanical, green tea. may be a useful anti- inflammatory agent and may have antineoplaslic effects. The polypknolic fraction w3ated from green tea (CiTPl has been shown lo afford protection agsinst UVB induced cercinogenesis and against local and systemic suppression of contact hypersensitivity in mouse skin. To examine the potential mechanisms involved in this complex phenomena. we mvestigated the effect of GTP on UVB induced changes in cultured human keratinocytes. In particular, the effect .of membrane changes in keratinocytes was assessed by labelling keratinwytes with ‘H arachidonic acid.. GTP added to UVB irradiated keratinocytes reduced UVB induced release of labelled arachidonic acid in a dose-dependent manner. The effects of GTP on cytokine regulation was assessed by semi-quantitative PCR. The consritutive expression of interleukin-la (IL-la) was down-regulated by GTP in a dose-dependent manner at 24 hr. while TNFa message remained unchanged. The concenlration of GTF (0.01 -20 @g/ml) employed was not toxic to keratinofytes as assessed by tetrazalium Mm reduction. Our results suggest that GTP may modify UVB induced events by stabilizing cell membrane and down-regulating proinflammatory cytolrine IL-l@..

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MOOULATORY I:,-FECTS OF CELLPERMEABLE SPHlNGOSlNE AND CERAMIDES ON CELLIILAK KINETICS OF T GILLS TH2 SUPPRESSOR CELLS ARE MORE susc*:PrI”t.t: 10 SP”,N00S,N6 THAN THI CELLS IN MtmlNE DELAYED-TYPE