Sunlight suppressing rejection of 280- to 320-nm UV-radiation-induced skin tumors in mice.

Repeated exposure of female C3H/HeNCR- mice to sunlight prevented the normal immunologic rejection of a UV-induced tumor. This systemic immunologic alteration was transferred to syngeneic lethally X-irradiated animals with lymphoid cells from mice exposed to sunlight. The lymphoid cells also were able to suppress the capacity of lymphoid cells from normal animals to reject a UV-induced tumor. The 295- to 320-nm wave band appeared to be responsible for this immunosuppressive effect of sunlight because suppression was prevented by filtration of the radiation through Mylar and by application of a sunscreen containing para-aminobenzoic acid. These observations may have importance in understanding the pathogenesis of sunlight-induced skin cancer in humans.

Induction and transplantation of murine skin cancers induced by methoxsalen plus ultraviolet (320-400 nm) radiation.

Properties of skin tumors resulting from treatment of mice with ip methoxsalen plus UVA (320-400 nm) radiation (PUVA) were compared to those of skin tumors induced by UVB (280-320 nm) radiation. Repeated treatment of C3H/HeN murine mammary tumor virus-negative (C3H-) mice with PUVA produced both fibrosarcomas and squamous carcinomas that were indistinguishable morphologically from UVB-induced tumors. Fragments of 12 primary PUVA-induced tumors were transplanted into normal and immunosuppressed syngeneic recipients, and all grew progressively. In contrast, 19 primary tumors induced by repeated exposure of C3H- mice to UVB grew only in the immunosuppressed mice and not in the normal recipients. The growth of graded doses of cells from 5 PUVA-induced tumors was compared in normal and UVB-irradiated recipients. No preferential growth in UVB-irradiated mice was observed, even though this is characteristic of UVB-induced tumors. Thus PUVA-induced tumors do not appear to have the same antigenic properties as UVB-induced tumors.

UV radiation-induced tumors in haired mice: identification as squamous cell carcinomas.

The most common tumor induced by UV radiation in haired mice is considered to be a fibrosarcoma on the basis of its presentation as a nodule in the skin and on the basis of a spindled appearance upon light microscopic examination. A squamous cell carcinoma is thought to be a much less common tumor. In the present report this concept was reevaluated in mammary tumor virus-free C3H/HeNCr (C3H-) mice. From first appearance, almost all lesions upon gross morphologic examination have an epidermal component and initially are similar to solar keratoses in humans. The lesions then become nodular and eventually develop central ulceration, often with a rolled border characteristic of squamous cell carcinomas. The morphology upon light microscopic examination ranged from well-differentiated squamous cell carcinoma to a poorly differentiated spindle cell neoplasm. Occasionally, variable patterns of squamous differentiation were seen in the same lesion. Immunoperoxidase examination with a polyclonal antikeratin serum demonstrated the presence of keratin in 84 of 87 tumors. Frequent, poorly formed desmosomes were found on ultrastructural examination. These tumors usually had a regressor phenotype upon transplantation into recipients. In conclusion, almost all UV radiation-induced tumors in C3H- mice are squamous cell carcinomas, and these tumors are usually antigenic.

Impaired immune function in patients with xeroderma pigmentosum.

The development of contact allergy in sun-exposed skin is markedly impaired in patients with xeroderma pigmentosum as compared to the responses in healthy control subjects. The degree of this immunological impairment is directly related to the severity of the cutaneous disease. These findings raise the possibility that sunlight-induced alterations of immune function may be involved in the marked susceptibility of these patients to the development of nonmelanoma skin cancer.

The effect of a sunscreen containing para-aminobenzoic acid on the systemic immunologic alterations induced in mice by exposure to UVB radiation.

Application of a sunscreen containing para-aminobenzoic acid partially abrogated certain of the systemic immunologic alterations produced in mice by exposure to UVB (280-320 nm) radiation from sunlamp bulbs. The sunscreen reduced the degree of UVB-induced suppression of contact hypersensitivity to a chemical applied subsequently to nonirradiated skin. In addition, it reduced the frequency with which mice became susceptible to the growth of a highly antigenic, syngeneic, UVB-induced tumor following chronic treatment with UVB radiation, but this effect was not statistically significant. The tumor-susceptible state was transferred from animals treated with sunscreen and UVB radiation to lethally x-irradiated mice by injection of spleen cells. The gross morphology of the skin of mice treated with sunscreen and UVB radiation was normal but the histologic changes induced by UVB irradiation in skin were only partially abrogated by the sunscreen.

Studies on the mechanism of systemic suppression of contact hypersensitivity by UVB radiation. II. Differences in the suppression of delayed and contact hypersensitivity in mice.

Exposing mice to UV radiation in the UVB range (280-320 nm) causes a selective immune suppression that contributes to the development of UVB-induced skin cancers. Among the immune responses suppressed by UVB irradiation are contact and delayed hypersensitivity reactions to haptens administered at unexposed sites. In these studies we provide evidence that delayed and contact hypersensitivity to the same hapten are not equivalent reactions and that they are suppressed in UVB-irradiated mice by 2 different mechanisms. This conclusion is based on the findings that: suppression of contact hypersensitivity could not be overcome by immunizing UVB-irradiated mice with hapten-coupled antigen-presenting cells derived from normal donors; and treatment of UVB-irradiated mice with methylprednisolone before immunization prevented the suppression of delayed hypersensitivity but had no effect on the suppression of contact hypersensitivity. The decreased ability to induce contact hypersensitivity in UVB-irradiated mice could be transferred to x-irradiated mice by reconstituting them with spleen cells from UVB-irradiated donors. The induction of hapten-specific suppressor cells, however, required both UVB irradiation and priming with hapten. Based on these results, we postulate that UVB irradiation induces a population of suppressor-inducer cells with specificity for a modified skin antigen and that this antigen serves as a carrier molecule for haptens that induce contact hypersensitivity and for tumor-specific transplantation antigens on UVB-induced tumors.

Modulation of immune function by UV radiation.

In addition to its carcinogenic activity, ultraviolet (UV) radiation is capable of modifying certain immunologic reactions. Immunologic alterations induced in mice by UV radiation include both local and distant effects. Local alterations result from a direct effect of UV radiation on an immune reaction that takes place at the site of irradiation. Distant alterations are those in which exposure of skin to UV radiation at one site modifies an immune reaction occurring at a distant, unexposed site. Based on recent studies, we propose that there may be two types of distant alterations. One is nonspecific, may be due to accumulation of leukocytes at the site of UV-induced inflammation, and is exemplified by the suppression of delayed hypersensitivity and local graft-versus-host (GVH) reactions. The second may result from DNA damage, may involve a soluble mediator, and is manifested by the systemic suppression of contact hypersensitivity and the formation of antigen-specific suppressor T lymphocytes. These immunologic effects of exposure to UV radiation may be important in the pathogenesis of skin cancer and other cutaneous diseases.

Suppression of graft-versus-host reactivity in the mouse popliteal node by UVB radiation.

A single exposure of recipient (C57BL6 X C3H-) F1 (B6C3F1) mice to UVB radiation suppressed the graft-versus-host (GVH) reaction to injected C3H- lymphoid cells, as measured by the popliteal lymph node weight gain assay. Several observations provided evidence to suggest that this effect of UVB radiation is nonspecific and involves an alteration of the host lymphoid cell component of the reaction. First, the nonspecific trauma of mild thermal injury also suppressed the GVH reaction. Second, although treatment of mice with rose bengal and visible radiation suppresses contact hypersensitivity while treatment with eosin and visible radiation does not, both types of phototoxic treatment suppressed the GVH reaction. Third, implantation of spleens from normal B6C3F1 mice into UVB-treated or thermally injured recipient mice at the time of injection of graft cells overcame the suppression of the GVH reaction. Finally, treatment of donor B6C3F1 mice with UVB radiation did not suppress the host-versus-graft reaction in recipient C3H- mice, which suggests that radiation does not alter the stimulatory function of B6C3F1 cells. These findings are all consistent with a hypothesis that UVB radiation suppresses GVH reactivity by reducing the host component of this immune response through diversion of cells from the site of the reaction. Thus an alteration of cell trafficking appears to be an additional pathway by which UVB radiation can produce immunosuppression.

Differences in the immunologic reactivity of mice treated with UVB or methoxsalen plus UVA radiation.

Skin tumors induced in mice by chronic exposure to UVB radiation are often highly antigenic and regress when transplanted into normal syngeneic animals, but grow progressively in immunosuppressed mice. Exposure of mice to subtumorigenic doses of UVB radiation can abolish this immunologic rejection phenomenon. In this study, we have investigated the effects of treatment with 8-methoxypsoralen plus UVA radiation (PUVA) on the rejection of antigenic UVB-induced tumors. PUVA treatment, with either topical or systemic administration of the psoralen, did not alter the normal process of rejection of UVB-induced tumors. Mice treated with both minimally and markedly phototoxic doses of PUVA rejected tumors with a frequency similar to that seen in untreated animals, although these tumors grew progressively in UVB-irradiated mice. These results indicate that the effects of PUVA treatment differ from those of UVB irradiation in that PUVA treatment does not alter the immunologic rejection of UVB-induced tumors.

The effects of indomethacin on long-wave ultraviolet-induced delayed erythema.

Indomethacin, a known inhibitor of prostaglandin synthesis, administered topically as a 2.5% solution, intradermally in 10 mug doses, and orally in a dose of 150 mg/day for 2 days did not diminish the delayed erythema produced a long-wave ultraviolet light (320-400 nm). The delayed phototoxic erythema produced by 8-methoxypsoralen and subsequent exposure the long-wave ultraviolet light was similarly unaffected. By comparison, topical and intradermal indomethacin treatment produced sustained decrease in the erythemal response to ultraviolet radiation in the UVB ranges (290-320 nm).

Systemic suppression of contact hypersensitivity in mice by psoralen plus UVA radiation (PUVA).

Treatment of mice with 8-methoxypsoralen plus longwave UV radiation (UVA, 320-400 nm) decreased their response to contact sensitizers applied subsequently to unirradiated skin. This decreased reactivity exhibited a delayed time course, it affected the afferent but not the efferent phase of the reaction, and it was associated with the development of splenic suppressor cells. These suppressor cells were antigen-specific T lymphocytes, and they prevented the induction, but not the elicitation, of contact hypersensitivity in recipient mice. In all of these characteristics, the decreased reactivity induced by treatment with psoralen plus UVA radiation (PUVA) resembled that produced by UV radiation of shorter wavelengths (less than 320 nm). These studies suggest that PUVA treatment may initiate the same sequence of cellular events as does exposure to sunlamp (UVB, 280-320 nm) radiation, leading to preferential activation of the suppressor cell pathway.

The role of UVB radiation in the induction and elicitation of photocontact hypersensitivity to TCSA in the mouse.

Photocontact hypersensitivity (PHS) to 3,3',4',5 tetrachlorosalicylanilide (TCSA) can be induced in mice by using cyclophosphamide as an immunopotentiator. Only UVA (320-400 nm) radiation was required for both sensitization and elicitation of PHS. The reaction was successfully transferred to syngeneic mice by injecting them with lymph node cells from sensitized donors, a finding that demonstrates the immunologic nature of PHS. The presence of UVB (280-320 nm) radiation was not necessary for sensitization and did not increase PHS beyond the levels observed with UVA radiation alone. Ultraviolet radiation in the UVB range (plus a small amount of UVA radiation) from FS40 sunlamps in the dose employed did not induce statistically significant PHS to TCSA, nor did it elicit a significant response in mice sensitized with TCSA plus UVA radiation. However, treatment of mice with UVB radiation at a distant site 6 days before sensitization suppressed the induction of PHS. This suppression appeared to be analogous to the systemic suppression of ordinary contact hypersensitivity in mice by UVB radiation.

Therapy of psoriasis by tar photosensitization.

For the first time since Goeckerman suggested the therapeutic potential of tar photosensitization of psoriatic skin, this mechanism has been shown to the effective in treating generalized psoriasis vulgaris. Long treatment times, need for special high-intensity sources, the presence of skin pain, and absence of information about long-term effects makes this treatment impractical at present.

Influence of PUVA and UVB radiation on delayed hypersensitivity in the guinea pig.

Exposure of guinea pigs to UVA (320--400 nm) radiation following administration of 8-methoxypsoralen by gavage (referred to by the acronym, PUVA) or exposure to UVB (290--320 nm) radiation, produced suppression of the cutaneous delayed hypersensitivity reaction at the site of exposure to radiation and at distant nonexposed sites. In these experiments, the animals were immunized by injection of dinitrophenyl-bovine gamma-globulin (DNP-BGG) in complete Freund's adjuvant and delayed hypersensitivity responses were provoked by intradermal injections of DNP-BGG, DNP and BGG on the flanks. Exposure to erythemogenic doses of either PUVA or UVB radiation for 7 days prior to immunization and for the 7 days between immunization and challenge (total period of radiation: 14 days) produced inhibiton of responses to each of the test substances. In addition, treatment with erythemogenic doses of PUVA either for 7 days prior to immunization or during the interval between immunization and challenge with DNP-BGG, inhibited the delayed hypersensitivity responses at the site of irradiation and at a nonexposed site. These findings suggest that in vivo exposure to nonionizing radiation leads to both local and systemic alteration of certain immune responses.

The influence of PUVA and UVB radiation on skin-graft survival in rabbits.

The survival time of full-thickness skin grafts in rabbits was prolonged by administration of methoxsalen and subsequent exposure of the donor and recipient graft sites to longwave ultraviolet radiation (UVA). Erythemogenic doses of radiation were required to prolong graft survival. Similar exposure to mid-ultraviolet radiation (UVB) did not significantly prolong the survival time to grafts.

Reduction of the fraction of circulating helper-inducer T cells identified by monoclonal antibodies in psoriatic patients treated with long-term psoralen/ultraviolet-A radiation (PUVA).

Ultraviolet radiation has been found to alter the distribution and function of human lymphocytes. To determine whether photochemotherapy (PUVA) alters circulating levels of T cell subset marker-bearing lymphocytes, cells from 9 patients with psoriasis undergoing PUVA therapy for several years (mean 4.6 +/- 1.4 yr), 17 patients with active untreated psoriasis, and 20 healthy volunteers were reacted with monoclonal antibodies to T cell surface markers, including OKT3 (all peripheral blood T cells), OKT4 (helper/inducer T cells), OKT6 (common thymocytes), and OKT8 (suppressor/cytotoxic T cells), and analyzed by flow cytometry. There were no differences in the distribution of T cell subsets between healthy volunteers and patients with active psoriasis. In contrast, the percentages of lymphocytes reacting with OKT3 and OKT4 were lower (by 16% and 12% percent respectively, p less than 0.0025) in the PUVA-treated patients compared to healthy volunteers or patients with active psoriasis that had not received PUVA therapy. There was no difference in the percentage of OKT8 and OKT6 bearing cells. Squamous cell carcinoma of the skin subsequently developed in 2 of 3 PUVA-treated patients with the lowest percentages of T4-bearing cells. These findings indicate that long-term PUVA therapy is associated with a reduction in circulating helper/inducer T cells. This reduction may have a role in the altered immune function reported in PUVA-treated patients.

In vitro assay for phototoxic chemicals.

The photosensitizing potential of chemicals known to produce photosensitivity in humans was compared to chemicals not considered to be photosensitizers in an in vitro assay. The assay involved exposure of human lymphoid cells to UVA (320-400 nm), and in some cases UVB (280-320 nm) radiation, in the presence of the chemicals and the assessement of phototoxicity as measured by the incorporation of 3[H]-thymidine into nuclear DNA. All known photosensitizers tested were found to be phototoxic, while the nonphotosensitizing agents, with the exception of retinoic acid, were not phototoxic. Peripheral blood mononuclear cells were compared to a T lymphoblastoid cell line as target cells; the latter were superior in terms of convenience, cost and reproducibility of results. This test system has potential as a predictive assay for detecting additional phototoxic chemicals.

Dermatological and ocular examinations in rabbits chronically photosensitized with methoxsalen.

Four groups of female Dutch-belted rabbits (Oryctulagus cuniculus) were given methoxsalen (12 mg/kg) or placebo by oral intubation and 1 hr later were exposed to UVA for either 2 or 8 hr. This procedure was repeated 5 days each week for 18 mo. A fifth group received no drug and no UVA exposure. The skin of the animals given methoxsalen and UVA showed signs of acute and chronic phototoxicity. Multiple peripheral blood parameters of hepatic, renal and hematologic function were normal and were not different between groups. Complete ophthalmoscopic examinations were performed periodically. No cataracts were seen in any of the animals. This data provides the perspective that in one species the daily dose of methoxsalen and UVA required to induce chronic cutaneous photosensitization is lower than the daily dose required to induce cataracts. It is inadvisable to interpret this data as suggesting that no risk exists for patients being treated with oral methoxsalen photochemotherapy. The experimental evidence supporting photosensitization as a cause of cataracts and implicating a role of lens DNA in this cataractogenesis is reviewed. Because methoxsalen-UVA alterations of lens DNA or protein could lead to delayed onset of cataracts, and because of the serious nature and potential preventability of phototoxic lens opacification, appropriate protective eye wear is recommended for all patients receiving oral psoralen photochemotherapy.

Ultraviolet carcinogenesis in athymic nude mice.

We have investigated the development of skin cancer from exposure to ultraviolet (UV) radiation in C3H- nu/nu nude mice. Nude mice, nude mice reconstituted with thymuses, and nude mouse skin grafted onto normal haired mice had similar tumor incidences and rates of tumor development. All tumors were squamous cell carcinomas and both well-differentiated and poorly differentiated lesions occurred in each of the groups. Transplants of the tumors that developed in nude skin grew preferentially in immunosuppressed mice as compared with normal mice, indicating that tumors from each treatment group were antigenic. These results indicate that the presence or absence of a functioning thymus does not seem to influence UV carcinogenesis.

Recent advances in phototherapy and photochemotherapy of skin disease.

The therapeutic spectrum for ultraviolet radiation treatment of skin disease has continued to be broadened. Psoralen photochemotherapy is beneficial in chronic lichenoid graft-versus-host disease and disseminated granuloma annulare. This treatment is now being found more useful in atopic eczema and chronic photosensitivity with some modifications of the therapy. UV phototherapy has also been found useful in mild to moderate atopic eczema. The nature of these treatments is also changing with greater use of selective UV phototherapy and definition of the required schedule for maintenance treatment with UVB phototherapy. The mechanism of therapeutic benefit remains unknown although one possibility is selective phototoxicity for inflammatory cells in the dermis. Nonmelanoma skin cancer, premature aging of the skin and freckling are the main long-term adverse effects of these treatments.