Coby Out-Luiting

Increased Risk of Cancer Other Than Melanoma in CDKN2A Founder Mutation (p16-Leiden)-Positive Melanoma Families

Purpose: We report the largest study to date analyzing the risk of cancers other than melanoma in melanoma families positive for the same CDKN2A mutation. Experimental Design: We studied family members of 22 families positive for the p16-Leiden founder mutation who had attended a surveillance clinic or were their close relatives. Within this cohort, observed and expected rates of cancer were computed by mutation status consisting of 221 (proven plus obligate) carriers, 639 (proven plus obligate) noncarriers, and 668 first-degree relatives whose carrier risk was estimated from the relationship to known carriers and the age and melanoma status of that person and their relatives. Results: Our analysis shows a relative risk (RR) of cancer other than melanoma and nonmelanoma skin cancer of 4.4 [95% confidence interval (95% CI), 3.3-5.6], predominantly attributable to the increased risk for pancreatic cancer (RR, 46.6; 95% CI, 24.7-76.4), but also for other cancers. We provide substantial proof for pancreatic cancer being a key component of the p16-Leiden phenotype. Inclusion of this cancer in a penetrance analysis leads to an estimated RR of pancreatic cancer for mutation carriers of 47.8 (95% CI, 28.4-74.7). Conclusions: This study shows clear evidence of increased risk of cancers other than melanoma in CDKN2A families carrying the p16-Leiden mutation. Further research is necessary to determine if similar risks apply to families with CDKN2A mutations other than p16-Leiden.

Gene expression profiling identifies tumour markers potentially playing a role in uveal melanoma development.

Microarray is a powerful tool to compare the gene expression of different tumour specimens and cell lines simultaneously and quantitatively. To get a better insight into genes that are involved in uveal melanoma tumorigenesis, we compared the gene expression profiles of 12 different uveal melanoma cell lines with three melanocyte cell cultures obtained from healthy donor eyes. Gene expression profiles were obtained by nylon filter arrays, containing 1176 gene spots related to cancer development. The expression levels of selected genes were validated on cell lines and primary uveal melanomas by real time RT–PCR, and were subsequently included in cluster analysis. Four candidate tumour markers, Laminin Receptor 1, Endothelin 2, Von Hippel Lindau Binding protein 1 and Cullin 2, have been selected from genes that were differentially expressed in the uveal melanoma cell lines compared to the normal uveal melanocytes. In primary uveal melanomas, these four markers could discriminate between two classes of uveal melanoma, which may be indicative of a differential disease process.

Efficacy of micronized titanium dioxide-containing compounds in protection against UVB-induced immunosuppression in humans in vivo

Micronized pigment-containing sunscreens may provide a good alternative to chemical sunscreens in protection against ultraviolet (UV) B-induced immunosuppression. The metal particles in these products are likely to remain on the skin surface where they can offer broadband protection for both the UVA and UVB regions. We have tested the protective capacity of three titanium dioxide (TiO2)-containing compounds in humans in vivo. The effect on sunburn cell formation has been investigated using transmission electron microscopy, while the mixed epidermal cell lymphocyte reaction (MECLR) has been used as a model for immunosuppression. Furthermore, the influence of titanium on the integrity of the stratum corneum barrier (intercellular lipids and desmosomes) has been examined using freeze fracture electron microscopy. We find that all three compounds protect against sunburn cell formation. The immunoprotection studies show that one of the three compounds does not prevent UVB-induced changes of the MECLR responses. Application of this compound without subsequent UVB irradiation also induces a significant decrease of the MECLR responses. Moreover, the same compound affects the intercellular lipid layers, and desmosomes cannot be detected. The deleterious effect of this compound is probably caused by an incomplete hydrolysis during the TiO2 synthesis. Our findings indicate that micronized pigment-containing compounds can offer good protection against short-term UVB-induced immunomodulation in humans in vivo. However, accurate screening of the synthesis of these compounds is a prerequisite for their safe use as sunscreening agents in human subjects.

THE ACTION SPECTRA FOR UV‐INDUCED SUPPRESSION OF MLR and MECLR SHOW THAT IMMUNOSUPPRESSION IS MEDIATED BY DNA DAMAGE

‐Ultraviolet‐B (UVB,280–320 nm) radiation can promote the induction of skin cancer by two mechanisms: damage of epidermal DNA and suppression of the immune system, allowing the developing tumor to escape immune surveillance. The mixed lymphocyte reaction (MLR) and the mixed epidermal cell lymphocyte reaction (MECLR) are commonly used methods to study the immunosuppressive effects of UVB radiation. To obtain a better understanding of the mechanism by which UVB radiation decreases the alloactivating capacity of in vitro‐irradiated cells, action spectra for the MLR and MECLR were determined. Suspensions of peripheral blood mononuclear cells or epidermal cells were irradiated with monochromatic light of 254, 297, 302 or 312 nm and used as stimulator cells in the MLR or MECLR. Using dose‐response curves for each wavelength, the action spectra were calculated. Both MLR and MECLR action spectra had a maximum at 254 nm and a relative sensitivity at 312 nm that was a thousand times lower than at 254 nm. Strikingly, the action spectra corresponded very closely to the action spectra that were found by Matsunaga et al. (Photochem. Photobiol. 54,403–410, 1991) for the induction of thymine dimers and (6‐4)photoproducts in irradiated calf thymus DNA solutions, strongly suggesting that the UV‐induced abrogation of the MLR and MECLR responses is mediated by UV‐induced DNA damage. Furthermore, the action spectra for the MLR and MECLR were similar, suggesting that they share a common mechanism for UV‐induced suppression.

UVB-induced leucocyte trafficking in the epidermis of photosensitive lupus erythematosus patients: Normal depletion of langerhans cells

Background: The pathogenic mechanisms of UV‐induced skin lesions of lupus erythematosus (LE) are unknown. In a recent study of pathogenic mechanisms of polymorphic light eruption (PLE), significantly more Langerhans cells (LCs) persisted in the epidermis after UVB overexposure than in healthy individuals. Interestingly, the same phenomenon was observed in one subacute cutaneous lupus erythematosus (SCLE) patient. It could therefore be hypothesized that both photodermatoses share a common pathogenic mechanism of photosensitivity. In the present study, we tested this hypothesis by investigating leucocyte trafficking in the initial phase of cutaneous LE after intense UVB exposure. Methods: In 22 photosensitive LE patients (12 chronic discoid lupus erythematosus, seven systemic lupus erythematosus and three SCLE) and nine age/sex‐matched controls, uninvolved buttock skin was exposed to six minimal erythemal dose (MED) UVB radiation. Subsequently, biopsies were taken after 24, 48 and 72 h, and one control biopsy was taken from unirradiated skin. Skin sections were stained for the presence of LCs, neutrophils and macrophages. Areal percentages of positively stained cells within the epidermis were quantified and compared between the patients and controls. Results: A gradual decrease of epidermal LCs and a gradual increase of epidermal neutrophils and macrophages at several timepoints after six MED irradiation was observed equally in both LE patients and controls. Conclusion: Immunohistopathology of irradiated uninvolved skin of photosensitive LE patients did not reveal the same pathologic trafficking of LCs and neutrophils as described for PLE patients. We conclude that different mechanisms are operative in the pathogenesis of PLE and photosensitive LE.

Ultraviolet-B radiation induces modulation of antigen presentation of herpes simplex virus by human epidermal cells

Although ultraviolet (UV) B radiation is known to be immunosuppressive, there is little information regarding a relevant immunological endpoint to assess human subjects in vivo. Therefore, we have examined the effect of in vivo UV radiation on the ability of human epidermal cells (EC) to present herpes simplex virus (HSV) antigens to memory T cells. Human volunteers, who were seropositive for HSV, were exposed to one minimal erythemal dose (MED) for four consecutive days. EC, prepared from suction blister roofs, were co-cultured with autologous T cells in the presence of HSV. HSV antigen presentation by UV-exposed EC was increased compared with control, nonexposed EC. This up-regulation correlated with an influx of macrophages into the epidermis, which are considered to be associated with UV-induced tolerance. Altering the UV protocol to a sub-erythemal UV dose for four consecutive days or to a single high dose of 2 MED, resulted in suppressed HSV antigen presentation, without the influx of the UV-macrophages. One of the goals of the present study was to eventually use this HSV system to investigate sunscreen immunoprotection. A pilot study with a TiO2-containing sunscreen suggested that the endpoint for UV-induced immunosuppression presented here is promising to be used for human in vivo sunscreen immunoprotection studies.

In Situ Action Spectra Suggest that DNA Damage is Involved in Ultraviolet Radiation-induced Immunosuppression in Humans

Abstract— The mixed epidermal cell lymphocyte reaction (MECLR) is a commonly used method to study the immunomodulatory effects of UV radiation. The in vitro action spectrum for the MECLR showed that the UV‐induced suppression of the MECLR responses is associated with UV‐induced DNA damage. To investigate whether in vivo DNA damage also leads to the abrogation of the MECLR, in situ action spectra were made for the MECLR and the induction of thymine dimers (T<>T). Human skin, obtained from plastic surgery, was exposed to monochromatic light of 254, 297, 302 and 312 nm. After irradiation, epidermal cells were isolated and used as stimulator cells in the MECLR or processed for flow cytometric detection of T<>T. On the basis of dose‐response curves for each wavelength, the action spectra for suppression of the MECLR and the induction of T<>T were calculated. These spectra showed close similarities, suggesting that, also in situ, UV‐induced DNA damage is involved in the UV‐induced suppression of the MECLR. Both action spectra showed a small decline from 254 nm to 302 nm, followed by a steep decline to 312 nm. These data show that, in situ, UVC can efficiently induce DNA damage and modulate cutaneous immune responses.

Differential Suppression of the Human Mixed Epidermal Cell Lymphocyte Reaction (MECLR) and Mixed Lymphocyte Reaction (MLR) by Cis‐Urocanic Acid

Abstract— Cis‐urocanic acid (UCA), formed in the stratum corneum by UV irradiation of trans‐UCA has been proposed as a mediator of UV‐induced immunosuppression in the skin. In this study, we examined the in vitro effect of cis‐UCA (6‐100 μg/mL) on the human mixed lymphocyte reaction (MLR) and the mixed epidermal cell lymphocyte reaction (MECLR). Addition of cis‐UCA (purified or in a mixture with trans‐UCA) did not affect the MLR but was able to induce a 20% suppression of the MECLR responses. Because this effect of cis‐UCA on the MECLR was not as strong as could be expected from previous in vivo results, we designed a set of experiments in order to enhance the in vitro immunosuppressive capacity of cis‐UCA. Firstly, we preincubated epidermal cells with UCA (50 u.g/mL) for 3 or 6 days before culture in the MECLR because in vivo repeated UV exposure can lead to a photostationary state, where cis‐UCA may be present for several weeks. This pretreatment with cis‐UCA resulted in a maximal decrease of the MECLR responses of 27%, whereas trans‐UCA had no effect. Secondly, we investigated whether UVB irradiation of epidermal cells could make cells more sensitive to cis‐UCA. However, addition of trans‐ or cis‐UCA did not potentiate the reduced alloac‐tivating capacity of UVB‐irradiated cells. Finally, we examined the possibility of a synergistic effect of cis‐UCA with histamine. Addition of histamine suppressed the MLR and MECLR responses, but neither cis‐ nor trans‐UCA were able to modulate this decrease. We conclude that cis‐UCA can partly downregulate the human MECLR but not the MLR. The mechanism involved in this differential downregulation is not known. In this respect it is striking that cis‐UCA does not potentiate the UVB‐ or histamine‐induced suppression of the MECLR.

UVB-induced suppression of the mixed epidermal cell lymphocyte reaction is critically dependent on irradiance.

The mixed epidermal cell lymphocyte reaction (MECLR) is a commonly used method to study the effects of ultraviolet B (UVB) radiation on the skin immune system. In UVB experiments dosimetry is very important. The influence of irradiance on the MECLR was studied in vitro using Philips FS40 lamps with variable UV intensities. Irradiation of isolated epidermal cells with high irradiance impaired the alloactivating capacity more than irradiation with low irradiance. In vivo, the influence of long‐term UVB exposure on the MECLR was studied by treating normal healthy volunteers with suberythemagenic doses of UVB thrice weekly during 4 weeks. The first set of experiments, using low irradiance Sylvania UV‐21 F75/85 W lamps, resulted in a decrease of MECLR responses of 83.1%. In the second set of experiments performed a year later, employing an identical protocol except for the use of high irradiance Waldmann UV‐21 F85/100 W lamps, an increase of MECLR responses of 99.7% was observed. Volunteers of both sets of experiments received equal doses of UVB. In conclusion, this study shows that in vitro UVB‐induced suppression of the MECLR is critically dependent on irradiance and therefore might explain contradictory results described in the literature. The in vivo data suggest that, comparable to the in vitro experiments, irradiance may influence the effects of UVB irradiation in vivo. Further experiments should prove whether this is indeed the case.