MEETING REPORT

8th MEETING OF THE ESPCR, PRAGUE, 23-26 SEPTEMBER 1998

by Dr Sheila Mac Neil

This meeting was held at the time of the 650th Anniversary of Charles University, Prague and was organised by the 1st Faculty of Medicine, Charles University. The spirit of the city of Prague and of Charles University was evident in every aspect of the meeting. A strong sense of history and of continuity graced the proceedings, the kindness and gentle humour of our Czech hosts made this an extremely pleasant meeting.

Who was there?

The ESPCR meeting was attended by 142 participants from 17 different countries. In 2˝ days there were 10 scientific sessions and 6 plenary lectures and 54 posters. A busy time but still with plenty of time for excellent music and food and wonderful beer and time to catch up with friends and colleagues.

Nature of report

Several colleagues have kindly contributed a summary of the highlights of sessions they chaired and have also identified particular talks or posters that they found raised their pulse rate. In this respect there was one particular plenary lecture which stood out above all others - John Pawelek and colleagues provided new experimental evidence that could change how we think about metastatic melanoma ... For details, see a fuller description later in this report (under "Our favourite things").

SESSION I Melanin-synthesis, properties and function

Report kindly provided by Professor Patrick Riley

Six papers were presented in this Session and 3 posters (P1, P3 and P52) were included in the discussion. The first talk was a brief and lucid exposition by Professor Christopher Ramsden of a series of chemical studies inspired by some new reactions found as a result of work on tyrosinase oxidation of analogue substrates. In this he outlined the production of the indoliumolate betanes and described work which led on to the study of the facile formation of quinomethanes from orthoquinones under conditions in which the alpha carbon has acidic character. This latter characteristic of these reactions enabled a reinterpretation of the famous Gates' synthesis of morphine. In the discussion of this paper Dr. Edward Land briefly mentioned the poster display P1 devoted to the generation of orthoquinones by pulse radiolysis, a technique that permits the measurement of the rate constants of subsequent reactions, such as cyclisation or isomerisation to form the corresponding quinomethanes.

Dr. Allesandra Napolitano then presented some new chemical evidence enabling a more detailed description to be given of the intermediates in the oxidation of cysteinyl dopa leading to the formation of benzothiazines. This talk was followed by a discourse by Professor Bruno Nicolaus concerning the fundamental structural features of melanins. In this he emphasised the importance of the chemical "backbone" of conjugated double bonds permitting the possibility of semiconductor properties of melanins. He pointed out that the melanisation of deep-sea vertebrates for example, where no light-absorbing function would be expected, have evolutionary value and leads to the expectation that there are other functions of melanins that are important. Whether melanin should be considered as jewel or garbage remained, however, an open question.

Dr. Jim Gallas then gave a spirited exposition of work on neutron diffraction analysis of experimental melanin solutions. He showed that there were specific aggregation patterns with a 3.4 Angström stacking characteristic, suggesting a layering phenomenon. These aggregates could be disrupted by hydrogen peroxide. Professor Milan Elleder then gave a brilliantly illustrated talk showing UV irradiation-induced fluorescence in pigmented tissue. This was also shown to be a feature of synthetic melanin and could be stimulated by prior treatment with hydrogen peroxide. The following talk by Dr. Luciana Mosca illustrated similar fluorescence in material generated by the oxidation of tyrosyl residues in opiate peptides. In discussion, the possibility that the fluorescence was dependent on degradation which involved Fenton chemistry, and thus bore similarities to the methodologies employed in posters P3 by Donato et al. and P52 by Wakamatsu, Ito and Koch was proposed. However, caution in the interpretation of the autofluorescence phenomena was urged by Professor Tadeusz Sarna who pointed out that, at least in vivo, there were a number of other compounds that could undergo autofluorescence and might be influenced by melanin in the micro-environment. Professor Karin Schallreuter strongly suggested that the fluorescence was related to other compounds that were known to be present in melanosomes such as the pterins, an interpretation that was not accepted by Professor Elleder who pointed that this could not be the case with synthetic melanins. The interesting differences between the case of induction of autofluorescence between eumelanins and phaeomelanins was also briefly mentioned, but lack of time prevented a full discussion of this interesting new data which may have diagnostic significance in view of the apparent increase in phaeomelanin synthesis in dysplastic naevi (a topic mentioned in a later session of the meeting).

SESSION II Melanogenesis

This contained 4 talks. Most controversial of these was that presented by Professor Karin Schallreuter. Professor Tony Thody comments on this session.

"One would have expected a little more on the MSH peptides in view of the current controversy concerning their significance as pigmentary hormones in humans (see Session IX for MSH and immunomodulation). The topic was at least touched on by Karin Schallreuter in her plenary lecture on pterins and pigmentation. It is now clear from her work that 6-tetrahydrobiopterin (6-GH4) has an important role in melanogenesis by regulating the availability of L-tyrosine and tyrosinase activity. She now has new data which suggests that a-MSH figures in this control through its ability to bind 6-BH4 and the implications are that a-MSH is able to regulate melanogenesis through activation of the MC-1 receptor and also through actions which are independent of the receptor. This is an extremely novel idea and hopefully we shall hear more on the subject at future ESPCR meetings".

Nico Smit presented work on the flavonenzyme DT-diaphorase suggesting that this enzyme may enhance oxidative stress by generating redox cycling catecholes and depletion of NAD(P)H unless other detoxifying enzymes are present.

Benathan and colleagues looked at the effects of thiol-modulating agents on melanogenic activity of normal and malignant pigment cells. Their results suggested that the balance between cysteine and glutathione may play an important role in regulating melanogenic activity of pigment cells. Ullrich Schraermeyer et al. presented evidence that melanosomes in retinal pigment epithelial cells are active lysosomes involved in the degradation pathway of rod outer segments of the eye.

SESSION III Microphthalmia encoded Transcription Factor and melanogenesis

Report kindly provided by Professor Vincent Hearing.

Vincent Hearing and Anthony Thody co-chaired this brief but interesting session which centred around the role of MITF (Microphthalmia encoded Transcription Factor) and other transcription factors that regulate mammalian melanogenesis. It has been known for some time that MITF, a basic helix-loop-helix transcription factor, regulates tyrosinase gene expression (as well as TRP1/TyrpI and/or TRP2/Dct, although these have been disputed) through binding to the E-box upstream regulatory region present in all 3 encoded genes. In this session, we heard that at least one, and probably many other, factors also play important roles in this regulation. Initially, M. Furumura reported his studies looking at transcriptional modulation of TRP genes by murine ITF2, another bHLH transcription factor originally identified by him as being upregulated during pheomelanogenesis. ITF2 was able to trans-activate the TRP1 gene as strongly as MITF, stimulated tyrosinase gene expression somewhat, and TRP2 expression not at all. Interestingly, ITF2 was able to inhibit MITF stimulation of TRP gene expression, probably by generating inactive heterodimers between MITF and ITF2.

J. Vachtenheim next reported on the role of MITF in human melanoma cells; they showed that expression of MITF is repressed in some cultured melanoma cells, and that this was associated with downregulation of tyrosinase, TRP1 and TRP2 genes in those cells. They directly demonstrated MITF regulation of those genes in human melanocytes by transfecting MITF back into those cells and demonstrating consequent upregulation of one or more of those TRP genes in the transfected cells. Their data support the critical nature of MITF expression in the pigmented phenotype of human melanocytes, and further suggest that repressors may be present in unpigmented cells which may also play important roles in regulating expression and catalytic function of those enzymes. Finally, S. Olaizola-Horn reported on the regulation of MITF and tyrosinase genes. Tyrosinase expression can be regulated via the cAMP and PKC pathways, but intracellular signalling pathways regulating MITF expression are not yet known. This study showed that treatment of melanocytes with forskolin to stimulate cAMP levels induced MITF gene expression within 4 hours and tyrosinase gene expression within 48 hours. Depletion of PKC by prolonged TPA treatment resulted in a decrease in pigmentation but was accompanied by increased MITF mRNA levels. These results suggest that tyrosinase and MITF gene expression are both regulated via cAMP and PKC pathways, but independently. This study further shows that factors other than MITF regulate tyrosinase gene expression and thus human pigmentation. The general conclusion of this session is that MITF is an important regulatory factor in controlling TRP gene expression, but that one or more other transcription factors are also important at this level of regulation.

SESSION IV UV light, photoprotection, phototherapy

Report kindly provided by Professor Césarini.

This session was chaired by Professor Tad Sarna and Professor Césarini. The session was composed of 11 formal oral presentations to which 4 posters can be linked and were briefly discussed at the end (the programme of the session was organised in co-operation with the European Society for Photobiology).

Three major topics were exposed: photoprotection, progress in photodynamic therapy and some insights in melanoma cell biology.

J.P. Césarini and H.C. Wulf presented the natural photoprotection offered by melanins for different phototypes and the acquired photoprotection after serial exposures to UVA + B radiations. Topical sunscreens (Sun Protection Factor based on protection against erythema) are able to suppress the actinic erythema but other UV effects like immunosuppression or indirect evidence for genotoxicity, are still present in the absence of erythema. The importance of skin-vehicle interaction was emphasised by B. Gabard and the skin penetration of UV filters seems a critical point for the quality of sunscreens. R. Pedeux had shown p53 expression in melanoma cells in culture. In work presented by E. Wencz, the association of UVA sensitivity and phaeomelanogenesis was found phototoxic, the melanin content being strongly correlated with UVA-induced single strand breaks in melanocytes.

The second part of the session was devoted to photodynamic phototherapy. S.B. Brown, after a review of the literature, explained how light source dosimetry and treatment protocols can be improved, and pointed also the need for more basic studies. G. Jori found that the tumour response (pigmented melanoma in mice) was affected by a variety of parameters and that hyperthermal conditions (43-44oC) contributed to the damage to the tumour, the photo bleached tumour being more susceptible to the PDT treatment. M. Jiraskova emphasised the red fluorescence observed after intralesional injection of PDT, the fluorescence being of great help for the evaluation of tumour extent. G.M.J. Biejersbergen-van-Henegouwen suggested that photobinding of chemical with DNA or proteins is a pre-requisite to obtain specific suppression of hypersensitivity in extra-corporeal phototherapy with UVA. The induced singlet oxygen produces immune suppression.

The third part was more specifically devoted to some biological aspects of melanoma. H.Z. Hill found that serum-free conditioned medium of cultured melanoma cells, following irradiation with ionising radiations, contains some antigenic proteins that increases the survival of melanoma cells. This may explain some drawback of radiotherapy and chemotherapy of melanoma. E.M. Link demonstrated that UV and ionising radiations may trigger the melanoma metastasis. Some "physiological" factors, like eicosanoids, may trigger the metastatic cascade. The production of eicosanoids was accompanied by the activation of ACTH and a-MSH mediated immune system (adrenal axis feedback loop).

SESSION V Pigment cell cultivation (My comments on this session).

Ullrich Schraermeyer reported on experience with both explant and enzymically dispersed culture of porcine and human iridial melanocytes. Roger Bowers reported on premature death of avian melanocytes in Barred White Leghorn feathers. He was able to induce premature death in vitro by the addition of L-dopa plus a-MSH or by not changing medium. Both led to an increase in oxygen radical accumulation and development of apoptosis in melanocytes. Using this model, he was then able to look at strategies to "rescue" melanocytes from oxygen radical accumulation. Addition of superoxide dismutase was particularly effective and he suggested that this avian in vitro system could be used to study premature ovarian melanocyte cell death analogous to that found in vitiligo.

Next was a presentation from M. Regnier from L’Oreal on the use of keratinocyte-melanocyte co-cultures and pigmented reconstructed human epidermis to study modulation of melanogenesis. Dr. Regnier demonstrated that co-seeding melanocytes and keratinocytes on acellular human dermis gave a model in which UV induction of pigmentation could be observed and the efficacy of topical applications of pro or de-pigmenting agents could be followed. In question time, I asked whether a-MSH induced pigmentation in these models and whether the model had ever been constructed with fibroblasts present. Dr. Regnier replied that MSH was not melanogenic in this model in their experience and the contribution or otherwise of the fibroblast had not been examined in this model.

This related to a presentation by Paula Eves et al. (from my own laboratory) where we found that the addition of fibroblasts to such a reconstructed 3D skin composite (based on sterilised human de-epidermised acellular dermis to which melanocytes and keratinocytes and fibroblasts were added) actually reduced the spontaneous pigmentation of the skin composites. In agreement with Dr. Regnier, however, we also find MSH to be without effect on pigmentation in this model irrespective of the presence or absence of fibroblasts.

Using this model we demonstrated that, in collaboration with Professor Ghanem, the human melanoma cell line (HBL), which is poorly invasive on its own, could be demonstrated to traverse the basement membrane when keratinocytes were also present. Keratinocytes on their own did not invade the basement membrane. This suggests some interaction between the melanoma cell line and keratinocytes which may be relevant to initial escape of melanoma cells from the primary tumour.

Sviderskaya et al. then reported on the establishment of 3 unpigmented lines of a new cell type from neonatal murine skin. These cells appear to be neural crest-like.

Aranberger et al. then gave a brief video (in the absence of any of the authors) of grafting of melanocytes and keratinocytes for patients with vitiligo.

SESSION VI Pigment cells and oxidative stress (My comments on this session).

For several years now pigment cell melanoma cell biologists have been asking whether melanocytes and melanoma cells differ in their handling of oxidative stress and indeed whether a failure to handle oxidative stress might contribute to melanocyte failure and their ultimate removal (as in vitiligo) or even to melanocytic transformation.

Continued work on this theme from the group of Professor Frank Meyskens focused on differences in the basal levels of the superoxide anion and hydrogen peroxide in cutaneous metastatic melanoma cells and cultured melanocytes (I am indebted to Dr. John Haycock for this summary of this work). These authors found that metastatic melanoma cells have higher levels of both superoxide anion and hydrogen peroxide than normal human melanocytes as determined by FACS analysis etc.

Next, he reported that melanocytes did not respond to an oxidative stress with an increase in NF-kB DNA binding activity, whereas metastatic melanoma cells did. Also, the level of constitutive NF-kB activity in metastatic melanoma cells was higher than in melanocytes. This could be reduced by incubating the cells with two types of antioxidants: (i) pyrrolidine dithiocarbamate or (ii) 1, 10-ortho phenathroline (a metal ion chelator which acts as antioxidant by removing transition metal ions, thereby preventing metal catalysed oxidation proceeding by the Fenton reaction).

He speculated that an NF-kB response might be seen if a high enough oxidative stress were given. The absence of a response, he thought, was due to a high level of intracellular antioxidant protection.

The interest in NF-kB Rel family members was extended to include various heterodimer formations, identified in the two cell types by immunoprecipitation. These included various combinations of p50, p65, p75 and p52. Under basal (unstimulated) conditions some combinations of Rel dimers were either very low or undetectable in the metastatic melanoma cells in contrast to the melanocytes.

Mauro Picardo and colleagues gave a presentation continuing their work into investigating the anti-oxidative status of patients with melanoma. They presented evidence that whereas in normal subjects, there was a correlation between epidermal and peripheral blood mononuclear cell levels of superoxide dismutase, catalase and vitamin E, this correlation was not seen in melanoma patients suggesting that some patients with melanoma may have a constitutive metabolic alteration. This, in turn, may contribute to their susceptibility to external oxidative stress.

In a second presentation from this group, Dr. V. Maresca presented evidence that the potent inflammatory cytokine, TNF- a, can itself generate pro-oxidative stress in melanoma cells. The response of cells to TNF- a may differ depending on the levels of intracellular antioxidants and peroxidisable compounds in the cells.

The final talk from Roger Bowers and colleagues continued Roger's work in trying to (a) deliberately drive to the point of destruction his chicken melanocytes and then (b) rescue them from impending death by paying attention to their ability to cope with oxidative stress. In this study, he showed that the addition of iron to the failing melanocytes increased their mortality rate significantly and that, as expected, drugs which compromised the ability of the cell to cope with oxidative stress (via buthionine sulphoximine addition, a glutathione inhibitor) and a superoxide dismutase inhibitor (diethyldithiocarbamate) both increased mortality rate in the feather melanocytes of the Barred White Leghorn.

SESSION VII Neuromelanogenesis

This was chaired by Professor M.G. Peter and Professor Bengt Larsson. (Comments by myself)

The first presentation by Vincent Hearing concerned macrophage migration inhibitory factor (MIF). This was originally identified as a lymphocyte-derived protein that inhibited monocyte migration. More recently it has been found to catalyse the conversion of dopaminechrome and norapenaphrinechrome, toxic quinone products of the neurotransmitters dopamine and norapenaphrine to indole quinone derivatives that may serve as precursors in neuromelanin. He demonstrated that MIF rescue cells from dopaminechrome-induced death in vitro and speculated that as MIF was highly expressed in human brain, it may participate in a detoxification pathway for catacholamine products and could, therefore, have an important protective role for neural tissues.

M. Miranda et al. then presented data of possible relevance to Parkinson's disease. In Parkinson's disease there is degeneration of the dopaminergic cells in the nigro-striatum system. A low level of tyrosine hydroxylase prevents transformation of L-tyrosine to L-dopa. A common therapy has been the administration of the dopamine precursor (L-dopa) but it does have severe side-effects. An alternative approach of stereotactic injection of liposome-entrapped tyrosinase was used to significantly increase the levels of dopamine in the rat brain.

SESSION VIII Gene expression in pigment cells

Chaired by Friedrich Beermann and myself. (Comments by myself).

This contained 5 talks.

In the first from Richard King and colleagues, transcripts of the Hermanski Pudlak Syndrome (HPS) chain were mapped. They identified several mutations and polymorphisms in this gene in individuals with HPS.

G. Kraehn and colleagues then presented work on differential expression of receptor tyrosine kinases in melanocytic skin lesions.

Staying with differentially expressed genes, the next presentation from R. Hipfel et al. searched for differentially expressed genes in malignant melanoma and congenital nevi biopsies. Using the technique of differential display more than 120 melanoma-specific and about 100 nevi-specific products were cloned and screened. Of these, 10 melanoma-specific transcripts were confirmed and sequenced. Three of these genes were singled out for particular interest as they concerned a cysteine protease, a protein proteinase inhibitor and a gene product that is mainly expressed in the brain and may function as a transcription factor.

The next presentation from J. Utikal et al. focused on c-myc oncogene expression and reported over-expression of c-myc with late stage melanoma which the authors speculated might be due to an increased number of c-myc - chromosome-8 copy number.

The last presentation from U. Leiter et al. concerned the apoptotic pathway in melanoma. These authors concluded that bcl2 gene expression increases in malignant melanoma which might reflect an increased malignant potential caused by an inhibition of apoptosis conferring a growth advantage in melanoma metastasis.

SESSION IX Melanoma - experimental aspects

This was chaired by Professor Doré and Professor Garbe. (Comments by myself).

It contained the most exciting presentation of the meeting (not just my opinion but resoundingly confirmed by Tony Thody, Frank Meyskens, Jan Borovansky and no doubt many others). For details of this presentation by John Pawelek, please see under "Our favourite things".

There were 6 other talks in this very lively session.

Friedrich Beermann and colleagues have used transgenic mice methodology to develop animals with tumours of the retinal pigment epithelium. This has been achieved using SV40 transforming sequences directed to the developing RPE using the promoter of tyrosinase-related protein-1 (TRP-1) in transgenic embryos. Work has advanced to the stage that primary tumour cell lines and metastasis derived from these have now been established and characterised.

Ruth Halaban and colleagues presented work in which they have examined to what extent the retinoblastoma tumour suppressor protein (RB) contributes to tumourigenicity in melanocytes. They were able to neutralise RB function and this led to releasing melanocytes from their normal cell cycle constraints but these melanocytes remained phorbol ester dependent and underwent accelerated cell death in the absence of phorbol ester. Thus, it appears that loss of this protein is insufficient for melanocytic transformation but clearly contributes to regulation of melanocytes by external growth inhibitory signals.

The next talk was from my own laboratory (B. Richardson) on the influence which sex steroids can exert on melanoma cell invasion (at least in vitro). Epidemiological studies show female survival benefit in advanced metastatic melanoma which is largely unexplained. Using a very simple model of a melanoma cell line (A375-SM cells) invading through a layer of human fibronectin over 20 hours, we were able to show that the female steroid 17ß-oestradiol and, to an even greater extent, oestrone, significantly reduce invasion of cells. Other androgenic and adrenal steroids were ineffective. This in vitro data begins to offer an explanation to the apparent female survival advantage in metastatic melanoma.

The next talk was from Dr. J. Haycock from my laboratory working in collaboration with Professor Ghanem. Jointly, the two laboratories have previously shown that a-MSH is able to oppose the actions of the pro-inflammatory cytokine TNF-a in both melanocytes and melanoma cells. The current study progresses this work to show that a-MSH can be demonstrated to oppose the actions of TNF-a at the level of activation of the transcription factor NF-kB. Thus, TNF-a would normally activate NF-kB to a maximal degree within 1-2 hours in cutaneous and ocular melanocytes and melanoma cells. We have demonstrated that a-MSH can reduce this activation by 50% on average, demonstrating for the first time in melanocytes and melanoma cells that the immunomodulatory action of a-MSH may rise by inhibiting the normal cytokine induced activation of NF-kB in both melanocytes and melanoma cells.

The next talk from P. Parsons et al. concerned the anti-tumour activity of agents which affect acetylation of histone. Cell killing was accompanied by hyperacetylation of histone H4.

The last talk in this session was from Stan Pavel and colleagues. They presented evidence that the composition of melanin in melanosomes of dysplastic naevi is very abnormal with increased concentrations of sulphur and, hence, phaeomelanin. According to Frank Meyskens - "they are building a compelling case that the naevi are under chronic oxidative conditions and, hence, internal genotoxic stress".

SESSION X Melanoma - clinical aspects

Report kindly provided by Professor Frank Meyskens.

Progress in the clinical area lags behind that of the tremendous advances occurring in our basic understanding of melanoma. There were, however, a couple of novel observations reported. Blum and his colleagues (Tuebingen) reported on a large comparative study of clinical examination to ultrasound evaluation of regional lymph nodes. Comparison to histopathological analysis indicated that ultrasound was more effective than palpation. Also, from the same group (Blaheta) was presented the results of another large study showing that sentinel node evaluation and biopsy combined with RT-PCR for tyrosinase in apparently negative cases increased the diagnostic accuracy. If the results of these two studies can be confirmed, their usage will be important in determining who might and might not benefit from adjuvant interferon, a difficult and expensive intervention. The Ulm group (Kaskel) also reported a large series in which S-100 was evaluated as a marker of disease. Its elevation in serum was found to correlate with high frequency with the appearance of metastatic disease; however, more extensive studies will need to be done to determine its usefulness as a prognostic or response (to therapy) marker. The remainder of the presentations of the papers in this session dealt with the epidemiology of melanoma in Germany and, although of interest to workers and the public in Germany, did not provide new or unexpected information about the disease.

OUR FAVOURITE THINGS

The presentation which attracted most attention and probably has caused many of us to go away and re-evaluate some of our ideas was that from John Pawelek in which he showed that experimentally mouse melanoma cells can fuse with macrophages to form hybrids, many of which are more aggressive metastatically in vivo than the parent cell line. In vitro, these cells were found to be more heavily pigmented and responsive to MSH than the parent cell line. The hybrids also responded to MSH with increased chemotaxis - a property not noted in the parent cell line.

John took great pains to point out that this is not a new idea - fusion of cancer cells with macrophages has been previously noted for other cancers including melanoma (Munzarova and colleagues, Lancet 1987 and Melanoma Research 1992) and it offers another explanation for how melanoma cells acquire metastatic success. Quoting from Munzarova et al. "We are of the opinion that the rapid assimilation of multiple properties from various populations of cells (and especially those of macrophages) obtained by fusion and processes after it are a better explanation of many of these qualities than the requirement of a single cell lineage to undergo sequentially so numerous mutagenic alterations" (Munzarova et al. Neoplasma 1992; 39: 70-86). This fusion behaviour which John and colleagues demonstrated can happen reproducibly in vitro (Rachkovsky et al. Clinical Experimental Metastasis, 1998, 16: 299-312) offers a new paradigm for the study of melanoma cells and their host interactions. It has implications for detection of metastatic melanoma, for drug resistance and, down the line, it may offer new avenues for therapeutic and preventive intervention.

Does this occur clinically? Is this what is going on with advanced metastatic melanoma? It was unfortunate that John’s talk was delivered near the end of the meeting so that there was less opportunity for discussion of the import of this work which is potentially enormous.

Other plenary lectures

In addition to John Pawelek and Karin Schallreuter (already mentioned in this report), we had excellent contributions from Professor Pat Riley, Professor Tadeusz Sarna, Dr. Nico Smit and Professor M. Elleder.

Professor Riley gave a plenary lecture explaining that the unusual kinetic behaviour of tyrosinase is due to its activation by dihydric phenol substrates which are formed indirectly (including dopa which is not a direct product of the tyrosinase reaction) as previously proposed by Raper and his co-workers (my thanks to Jan Borovansky for this succinct summary of Pat Riley's talk).

Professor Sarna looked into the complex question of whether melanins act as antioxidants and how phototherapy of pigmented tissues must take into account the ability of melanin to bind photosensitisers. Also, oxidative degradation of melanin can significantly reduce its antioxidant efficiency. Professor Sarna's talk underlined once more that the exact role of melanin in photoprotection remains unclear.

Dr. Nico Smit, in a plenary lecture emphasising how much the culture conditions can influence the pigmentary biology of the melanocyte, made a plea for some standardisation of culture conditions in order to be able to compare results obtained in different laboratories.

Finally, Professor Elleder gave an excellent review of lipopigments. Frustratingly, the chemical nature of lipopigments is still unknown despite serious analytical effort in this area. He pointed out that lipopigments may contain a number of associated compounds, one of which may be melanin although this is the subject of some debate. The best understood lipopigment is that occurring in a fatal neurodegenerative disease where the origin of the pigment is an agregate of extremely hydrophobic proteins (enzyme subunits of a mitochondrial enzyme).

We were also delighted at this meeting to, as a Society, offer honorary life membership to Professors Riley and Duchon in recognition of their considerable achievements to research in pigment biology and their unflagging support of the European Society for Pigment Cell Research.