Segregation of two glutaminase isoforms in islets of Langerhans.

Despite the importance of glutamatergic signalling in the co-ordination of hormone secretion, the identity of the enzyme for the production of glutamate in beta-cells is still unresolved. We have found that the endocrine pancreas co-expresses two isoforms of GA (glutaminase), denoted as kidney-type (KGA) and liver-type (LGA), with a complementary cellular pattern of expression. Whereas KGA was mainly present in alpha-cells, LGA was very abundant in beta-cells. This spatial segregation may have important functional implications, facilitating a differential regulation of glutamate production in insulin- and glucagon-secreting cells.

Novel protein kinase C and matrix metalloproteinase inhibitors of vegetable origin as potential modulators of Langerhans cell migration following hapten-induced sensitization.

BACKGROUND: Migration and maturation of epidermal dendritic cells, the Langerhans cells (LC), are central events in the initiation of the cutaneous immune response. LC migration from skin to draining lymph nodes is regarded as an indispensable step for the early phase of antigen-specific sensitization. Among the several agents which influence the ability of LC to migrate, previous studies have revealed that matrix metalloproteinases (MMPs) and protein kinase C (PKC) contribute to promoting LC migration. In this work, we studied the effect of two recently developed PKC and MMPs inhibitors of vegetable origin on the migration of in vitro activated LC. METHODS: The migratory capacity of epidermal and in vitro generated LC was assessed using a reconstituted basement membrane assay (Matrigel), mimicking the prerequisite passage through the dermal-epidermal basement membrane on the way to the lymph nodes. RESULTS: Contact with chemical allergens, Bandrowski's base or 2,4-dinitrobenzenesulfonic acid (DNBS), triggered migration. In the presence of PKC inhibitors, D-erythro-sphingosine and OX100, or an inhibitor of MMPs, LU105, allergen-induced migration of LC was strongly decreased. The association between OX100 and LU105 was more efficient in modulating the migration of activated LC compared to each molecule tested separately. CONCLUSIONS: These results showed that PKC and MMPs inhibitors act in synergy to inhibit the migration of activated epidermal dendritic cells in vitro. They underscore the role of PKC and MMPs inhibitors and suggest they may be of relevance for therapeutically regulating epidermal dendritic cell migration in inflammatory dermatoses.

Imiquimod, a topical immune response modifier, induces migration of Langerhans cells.

Langerhans cells are bone marrow derived dendritic cells that represent the major antigen-presenting cells in the skin. Langerhans cells take up and process antigen within the epidermis and present processed antigen to T lymphocyte in the regional lymph nodes and thus form an integral part of the cutaneous immune response. The cutaneous immune response can be modified by a number of pharmacologic agents, including corticosteroids, cyclosporine, and retinoids as well as physical agents, such as ultraviolet light. For the most part these agents act by suppressing immune function. A topical immune response modifier, imiquimod has been shown to enhance the cutaneous immune response. Imiquimod has anti-viral and anti-tumor effects in animal models and has been approved for the topical treatment of external genital and perianal warts in humans. The biologic activity of imiquimod in part is due to its effect as a cytokine inducer. Preliminary data suggested that imiquimod could have an effect on Langerhans cells. In order to clarify this effect on Langerhans cells, we examined Langerhans cell morphology and migration in imiquimod-treated skin. The density of Ia + cells decreased 2 d after treatment, falling to approximately 43% by day 10. The Ia positive in cells remaining in the skin appeared larger and more dendritic suggesting an activated state. ATPase staining of epidermal sheet confirmed the decreased number of Langerhans cells. To clarify status of Langerhans cells, the activation of B7 was examined. Activation of B7-1 or B7-2 was not detected. Imiquimod, however, did enhance Langerhans cell migration from skin to draining lymph nodes. This enhanced Langerhans cell migration was also associated with an enhanced allergic contact hypersensitivity. These results suggest that the mechanism of modulation of immune response by imiquimod is in part due to effects on Langerhans cells.

Enhanced inflammation and immunosuppression by ultraviolet radiation in xeroderma pigmentosum group A (XPA) model mice.

Xeroderma pigmentosum group A (XPA) gene-deficient mice were developed by gene targeting in mouse embryonic stem cells. To examine whether these XPA-model mice display photodermatologic abnormalities similar to those in human xeroderma pigmentosum, we investigated the effects of acute ultraviolet radiation on the homozygous (-/-) mice compared to the wild type (+/+) and heterozygous (+/-) mice. A single irradiation with ultraviolet B or topical psoralen plus ultraviolet A treatment induced stronger and longer lasting ear swelling in the (-/-) mice than in the (+/+) and (+/-) mice. Histologic changes including epidermal necrosis, cell infiltration, and sunburn cell formation after ultraviolet B radiation were more prominent in the (-/-) model mice than in the control mice. The (-/-) model mice showed damage of ADPase(+)Langerhans cells at a lower ultraviolet B dose than did the control mice. Moreover, the reappearance of ADPase(+)Langerhans cells after ultraviolet B radiation was delayed in the (-/-) mice compared to the control mice. Although contact hypersensitivity was induced equally in all mice, ultraviolet B-induced local and systemic immunosuppression were greatly enhanced in the (-/-) model mice. The data suggest that the XPA gene-deficient mice may be a useful model of human XPA, because the responses to UV radiation in the mice were very similar to those in the patients with XPA. Moreover, it is possible that enhanced ultraviolet immunosuppression is involved in the development of skin cancers in xeroderma pigmentosum.

[The effect of Tomesa therapy on epidermal Langerhans cells in experimental animals]. / Die Wirkung der Tomesa-Therapie auf epidermale Langerhanszellen im Experiment.

In the last years a new therapy of psoriasis was developed, which consists in a treatment with salt solutions, resembling the water of the Dead Sea, and ultraviolet light (Tomesa-therapy). We studied the influence of the used salt on ATPase positive epidermal Langerhans cells in murine ear skin. An irreversible partial reduction of the Langerhans cell ATPase was found after salt treatment of separated epidermis or of full skin preparations. These results may have implications for the optimization and broader application of this therapy.

Alterations in Langerhans cells and Thy-1+ dendritic epidermal cells in murine epidermis during the evolution of ultraviolet radiation-induced skin cancers.

To understand the role of cutaneous immune cells in host resistance to the induction and growth of skin cancer, we investigated the number and morphology of murine dendritic epidermal cells (dEC) during the evolution of ultraviolet (UVA) UV-induced skin cancers. Female C3H/HeN mice were treated topically with 8-methoxypsoralen followed by ultraviolet A (UVA) radiation 3 times/week or irradiated with UVB radiation 3 times/week. In both psoralen plus UVA- and UVB-treated mice, ATPase+ and Ia+ Langerhans cells almost completely disappeared from the treated skin during the early latency period of tumor development (4 weeks) but reappeared in the epidermis late in the latency period (between 15 and 22 weeks). The ATPase+ cells that reappeared in the epidermis had a rounder, less dendritic morphology than normal Langerhans cells. Thy-1+ dEC were totally depleted from the epidermis in both treatment groups at the end of first week of treatment and were nearly absent from the skin during the entire latency period. After tumors appeared (29 weeks), Thy-1+ dEC were still absent or detected only in small numbers in skin surrounding the tumors. ATPase+ and Ia+ cells present in skin around the tumors constituted 60 to 80% of the number in nonirradiated skin. Mice that received UVA radiation alone developed no tumors. ATPase+ and Ia+ Langerhans cells and Thy-1+ dEC were detected in UVA-treated epidermis after 22 weeks and 43 weeks, although the numbers were lower than those in unirradiated mice. Most psoralen plus UVA-induced tumors (81%) were squamous cell carcinomas, whereas only 24% of UVB-induced tumors were of this histological type. Our results demonstrate that UV-induced skin cancers developed in the presence of ATPase+ and Ia+ cells in the epidermis and in the absence of Thy-1+ dEC.

Evaluation of ATPase-positive Langerhans' cells in skin lesions of lupus erythematosus and experimentally induced inflammations.

We examined the time-dependent dynamics of epidermal Langerhans' cells (LC) in human systemic lupus erythematosus (SLE), in MRL/Mp-lpr/lpr(MRL/lpr) mice, and in various experimental cutaneous inflammations, such as the Arthus reaction, dinitrochlorobenzene allergic dermatitis, and croton oil primary irritant dermatitis, in order to clarify the pathomechanisms of lupus skin lesions. The numbers of LC in untreated SLE patients with newly developed skin lesions decreased in the central lesional sites and increased significantly in the peripheral lesional sites. In MRL/lpr mice, the number of LC increased significantly in the central lesional sites during the initial stage and increased in the peripheral lesional sites and decreased in the central lesional sites 2-4 weeks after the onset of skin lesions. In contrast, with experimental cutaneous inflammations of guinea pigs, the increase in numbers of LC in the peripheral lesional sites was not significant during the time course of the reaction. These results suggest that the increased numbers of LC during the active and early stages of skin lesions in human SLE and MRL/lpr mice are closely related to the specific spontaneous development of skin lesions, unlike the dynamics of LC in experimental cutaneous inflammations.

Antipsoriatic, erythematogenic, and Langerhans cell marker depleting effect of bath-psoralens plus ultraviolet A treatment. Comparison of 8-methoxypsoralen and trimethylpsoralen photosensitization.

Successful results have been reported for bath-psoralens plus ultraviolet A (PUVA) treatment of psoriasis with the use of either a low-sensitizing psoralen (methoxsalen) or a high-sensitizing psoralen (trioxsalen), but no evidence has been presented that phototoxicity would be a prerequisite for antipsoriatic activity of these treatments. To study therapeutic-to-phototoxic ratios, bath-PUVA was applied to psoriasis patients with the use of either a 0.2 mg/L solution of trioxsalen or a 0.4 mg/L solution of methoxsalen. The average minimal phototoxic ultraviolet A (UVA) dose (MPD) obtained after 15 minutes' bathing was 0.86 joule/cm2 for trioxsalen and 9.76 joules/cm2 for methoxsalen. In the first part of the study phototoxically equipotent treatment schedules were used, compensating the much lower phototoxic potential of methoxsalen by using about 10 times larger UVA doses compared to the doses used with trioxsalen. A healing rate of 71% +/- 25% was recorded for trioxsalen and 63% +/- 34% for methoxsalen treatment (mean +/- SD). Both PUVA treatments decreased epidermal Langerhans cell counts to 10% to 20% of the control value. In the second part of the study, the much lower phototoxic potential of methoxsalen was not compensated for. Instead, methoxsalen bath-PUVA was carried out with the use of UVA doses physically similar to those used in the trioxsalen therapy series. Surprisingly enough, even this very suberythemal methoxsalen therapy caused a significant healing effect (51% +/- 10%). Langerhans cell depletion in methoxsalen-bathed areas (reduction to 53% of the control value) was much less than that caused in trioxsalen-bathed areas (reduction to 11% of the control value).(ABSTRACT TRUNCATED AT 250 WORDS)

Reappearance of epidermal Langerhans cells after PUVA therapy.

Epidermal Langerhans cells (LC) disappear during photochemotherapy (PUVA) with 8-methoxypsoralen (8-MOP) and long wavelength ultraviolet (UV-A) radiation. The time course of their disappearance during treatment and their reappearance after its completion was followed. Langerhans cells lost ATPase activity before they disappeared by ultrastructural criteria: thus 90% of ATPase-stained cells had disappeared after seven treatments (2 weeks) whereas it was only after fifteen treatments (5 weeks) that they were seen to be reduced on electron microscopy. Their numbers remained low throughout the course of treatment but they had returned to normal by 3 weeks after cessation of therapy. Since PUVA lamps also emit traces of medium wavelength UV (UV-B) the separate effects of UV-A and UV-B in the presence and absence of 8-MOP were examined. Within the dose range normally used for therapy, the LC disappeared only with the combination of UV-A and 8-MOP.

Effect of oral methoxsalen photochemotherapy on human Langerhans cell number. Dose-response and time-sequence studies.

In human adult volunteers, oral 8-methoxypsoralen and UVA (PUVA) caused an almost linear dose-response effect in depleting adenosine 1--5 J/cm2 were used. A higher dose did not appreciably augment the LC depleting effect although the intensity of the PUVA-induced skin inflammation increased. After a single PUVA dose of 5 J/cm2, a nadir in LC density was achieved on day 8 after irradiation, with a decrease from the starting mean count of 704 +/- 58 cells/mm2 to 195 +/- 173 cells/mm2. On day 15 after irradiation, the LC count was still low (261 +/- 249 cells/mm2). In comparison, a single erythematogenic irradiation with a medium-pressure mercury lamp emitting mainly UVB caused an LC depletion which was less intensive, peaked earlier and was almost completely restored by day 15. With both modalities morphological changes were induced in the LC, manifested initially as a shortening of the dendritic processes and later as cell enlargement and dendrite elongation.

Relationship between epidermal Langerhans cell density ATPase activity and the induction of contact hypersensitivity.

Langerhans cells have been implicated as playing a crucial role as antigen-processing cells in the induction of positive immune responses to antigens presented through the skin. We have investigated the effects of carcinogenic doses of UVB-irradiation, psoralen plus UVA light (PUVA), PABA containing sunscreen preparations, and topically applied corticosteroids on both Langerhans cell densities and immunologic responsiveness to contact sensitizers applied to the treated site. The data presented demonstrate that UV-irradiation of skin or topical application of corticosteroids can create a local milieu in which DNFB cannot function as an effective stimulator of contact hypersensitivity. Further, we have shown that the effect induced by UV light is reversible, does not appear to be related to the numerous tissue destructive effects of UVB-irradiation, and that the correlation with the density of ATPase-positive cells is not absolute.