The effect of epidermal levels of urocanic acid on 25-hydroxyvitamin D synthesis and inflammatory mediators upon narrowband UVB irradiation.

BACKGROUND/PURPOSE: Urocanic acid (UCA) absorbs ultraviolet (UV)B radiation in the epidermis which may interfere with phototherapy. Therefore, the influence of individual levels of UCA on immune reactivity and vitamin D synthesis induced by narrowband UVB radiation was assessed. METHODS: Twenty-eight subjects with irritant contact dermatitis of the hands were irradiated with suberythemal doses of narrowband UVB radiation on their unaffected lower forearms on three consecutive days. Stratum corneum tape strips and epidermal interstitial fluid (ISF) as well as blood samples were analyzed. RESULTS: Narrowband UVB irradiation led to the conversion of trans-UCA into its cis-isomer in the epidermis. The observed increase in 25-hydroxyvitamin D serum concentrations was inversely correlated with the baseline levels of trans-UCA. Furthermore, UVB irradiation induced significant changes in the levels of CXCL10/IP-10, CCL2/MCP-1, CCL4/MIP-1ß, and the IL-1RA/IL-1α ratio. The levels of IL-1α and CXCL9/MIG showed a trend toward increase. The changes in the levels of inflammatory and immunomodulatory mediators did not depend on baseline levels of trans-UCA. CONCLUSION: The results suggest that epidermal levels of trans-UCA affect vitamin D synthesis, but not cutaneous immune reactivity upon repeated exposure to suberythemal doses of narrowband UVB radiation. However, this requires further exploration.

Basophil-derived amphiregulin is essential for UVB irradiation-induced immune suppression.

UVB irradiation (290-320 nm) is used to treat skin diseases like psoriasis and atopic dermatitis, and is known to suppress contact hypersensitivity (CHS) reactions in mouse models. Regulatory T cells (Treg cells) have been shown to be responsible for this UVB-induced suppression of CHS. The epidermal growth factor (EGF)-like growth factor amphiregulin (AREG) engages EGFR on Treg cells and, in different disease models, it was shown that mast cell-derived AREG is essential for optimal Treg cell function in vivo. Here we determined whether AREG has a role in UVB-induced, Treg cell-mediated suppression of CHS reactions in the skin. Our data show that AREG is essential for UVB-induced CHS suppression. In contrast to the general assumption, however, mast cells were dispensable for UVB-induced immune suppression, whereas basophil-derived AREG was essential. These data reveal, to our knowledge, a previously unreported function for basophils in the homeostasis of immune responses in the skin. Basophils thus fulfill a dual function: they contribute to the initiation of effective type 2 immune responses and, by enhancing the suppressive capacity of local Treg cell populations, also to local immune regulation in the skin.

UVB immunosuppression: vitamin D or not vitamin D? That is the question.

UVB radiation stimulates the production of vitamin D and also has immunosuppressive effects. Vitamin D is also known to alter immunological function. Thus, a relevant question is whether vitamin D is a mediator of the immunological effects of UVB. In this issue, Schwarz et al. have addressed this issue and have concluded that although vitamin D has similar effects to UVB on the immune system, UVB-induced immunosuppression can be achieved without the requirement for vitamin D action.

1,25-dihydroxyvitamin D exerts similar immunosuppressive effects as UVR but is dispensable for local UVR-induced immunosuppression.

Low-dose UV radiation (UVR) inhibits the induction of contact hypersensitivity and induces regulatory T cells (Tregs), which because of their antigen specificity harbor therapeutic potential. Topical application of 1α,25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3)) is known to induce Tregs as well, which implies that 1,25(OH)(2)D(3) might be involved in UVR-induced immunosuppression. It was the aim of this study to clarify this issue, to further characterize 1,25(OH)(2)D(3)-induced Tregs and to determine whether they differ from UVR-induced Tregs. Our data demonstrate that 1,25(OH)(2)D(3)-induced Tregs act in an antigen-specific manner and belong to the Foxp3-expressing subtype of Tregs as demonstrated by diphtheria toxin (DT)-mediated depletion of Foxp3(+) Tregs in DEREG (depletion of Tregs) mice. Using Langerin-DTR (DT receptor) knock-in mice, it was shown that Langerhans cells (LCs) are required for the induction of Tregs by 1,25(OH)(2)D(3), as depletion of LCs but not Langerin(+) dermal dendritic cells abrogated the induction of Tregs. Taken together, 1,25(OH)(2)D(3) affects the immune system in a similar manner as UVR, probably using the same pathways. However, vitamin D receptor knockout mice were equally susceptible to UVR-induced immunosupppression as wild-type controls. This indicates that 1,25(OH)(2)D(3) exerts similar immunosuppressive effects as UVR but is dispensable for local UVR-induced immunosuppression.

Ultraviolet light phototherapy for allergic rhinitis.

Phototherapy has a profound immunosuppressive effect and is widely used for the treatment of immune mediated skin diseases. Phototherapy is able to inhibit immediate type hypersensitivity reaction in the skin. Intranasal phototherapy is a new approach for treatment of allergic rhinitis. In two open studies, 308 nm excimer laser and topical PUVA therapy efficiently inhibited clinical symptoms of allergic rhinitis. In a randomized, double-blind study combined low dose UVB, low dose UVA and visible light proved to be effective in reducing symptom scores for sneezing, rhinorrhea, nasal itching and the total nasal score in ragweed allergic patients. Mechanism of action of phototherapy is complex, it reduces the antigen presenting capacity of dendritic cells, induces apoptosis of immune cells and inhibits synthesis and release of pro-inflammatory mediator from several cell types. Therefore, intranasal phototherapy may represent an alternative treatment of allergic rhinitis and other inflammatory and immune mediated mucosal diseases.

Nerve growth factor, neuropeptides, and mast cells in ultraviolet-B-induced systemic suppression of contact hypersensitivity responses in mice.

The induction of systemic immunosuppression following ultraviolet B radiation exposure has been linked with the release of inflammatory and immunomodulatory mediators by cells of the epidermis and dermis. Nerve growth factor has not previously been linked with ultraviolet-B-induced immunosuppressive effects. Nerve growth factor antibodies abrogated ultraviolet-B-induced systemic suppression of contact hypersensitivity responses in BALB/C mice. Subcutaneous injection of nerve growth factor (20 microg per mouse) into dorsal skin 5 d before hapten sensitization on ventral skin suppressed contact hypersensitivity responses in mast-cell-replete but not Wf/Wf mast-cell-depleted mice. Nerve growth factor injected 24 h prior to challenge was not able to suppress the efferent phase of the contact hypersensitivity response. Subcutaneous injection of nerve growth factor (20 microg per mouse) did not suppress contact hypersensitivity responses in capsaicin-pretreated (neuropeptide-depleted) BALB/c mice, and thus sensory c-fibers are necessary for nerve-growth-factor-mediated systemic suppression of contact hypersensitivity responses. Increased concentrations of nerve growth factor within epidermal keratinocytes 8 h after ultraviolet B irradiation were confirmed immunohistochemically. These findings support a role for keratinocyte-derived nerve growth factor via its action on sensory c-fibers, and subsequent release of neuropeptides to mediate mast cell degranulation in systemic suppression of contact hypersensitivity responses in mice following ultraviolet B exposure.

Pefloxacin and ciprofloxacin increase UVA-induced edema and immune suppression.

BACKGROUND: Quinolone antibiotics are popularly prescribed antibiotics because of their wide antibacterial spectrum and lowered bacterial resistance. Quinolone antibiotics are one of the well-known photosensitizers that induce phototoxicity. Their role in photocarcinogenesis has been suggested in some studies. MATERIAL AND METHODS: Mice were treated with two quinolone antibiotics (ciprofloxacin, which is less phototoxic, and pefloxacin, which is more phototoxic) to study the effect of the antibiotics on sunburn and immune suppression by ultraviolet A (UVA) irradiation. The effects of a combined treatment with UVA and these quinolone antibiotics were measured on back skin swellings, sunburn cell formations, depletion of epidermal Langerhans cells, and local and systemic suppression of contact hypersensitivity. RESULTS: Mice treated with both UVA and quinolone showed significantly increased back skin swellings and decreased epidermal Langerhans cells than mice treated with UVA only. Sunburn cells were increased significantly in mice treated with pefloxacin and 50 J/cm2 of UVA. Combination of pefloxacin and UVA suppressed local contact hypersensitivity significantly, but not systemic contact hypersensitivity. CONCLUSION: Phototoxic quinolones augmented the effect of UVA by increasing sunburn and apoptosis, depleting Langerhans cells and suppressing local immune response. By affecting apoptosis and immune suppression, they may facilitate photocarcinogenesis caused by UVA.

Dietary butter protects against ultraviolet radiation-induced suppression of contact hypersensitivity in Skh:HR-1 hairless mice.

Dietary fats modulate a wide variety of T cell functions in mice and humans. This study examined the effects of four different dietary fats, predominantly polyunsaturated sunflower oil, margarine, and predominantly saturated butter, clarified butter, on the T cell-mediated, systemic suppression of contact hypersensitivity by ultraviolet radiation in the Skh:HR-1 hairless mouse. Diets containing either 200 g/kg or 50 g/kg butter or clarified butter as the sole fat source protected against systemic photoimmunosuppression, whether the radiation source was unfiltered ultraviolet B (280-320 nm) or filtered solar simulated ultraviolet radiation (290-400 nm), in comparison with diets containing either 200 or 50 g/kg margarine or sunflower oil. There was a linear relationship (r > 0.9) between protection against photoimmunosuppression and the proportion of clarified butter in mice fed a series of 200 g/kg mixed fat diets that provided varying proportions of clarified butter and sunflower oil. The dietary fats did not modulate the contact hypersensitivity reaction in unirradiated animals. The observed phenomena were not primary due to the carotene, tocopherol, cholecalciferol, retinol, lipid hydroperoxide or the nonfat solid content of the dietary fats used and appeared to be a result of the different fatty acid composition of the fats.

Effect of a helium-neon laser on cutaneous inflammation.

A Helium-Neon (He-Ne) laser with a wavelength of 632.8 nm is known to have photobiological effects and is widely used for reducing the pain of herpes zoster and accelerating wound healing, however the cellular mechanism and effect of the He-Ne laser are poorly understood. The present study was designed to examine the influence of He-Ne laser irradiation on irritant and allergic contact dermatitis of the mouse ear and on histamine release from rat peritoneal mast cells. Irradiation was applied with a He-Ne laser (12.2 J/cm2) at 1 h, 10 min, 5 min and 0 min before, and 5 min, 6 hs and 24 hs after a challenge of an irritated contact dermatitis (ICD) or allergic contact dermatitis (ACD) was made on the right ears of ICR-mice. Twenty-four hours after the challenge, the swelling of the ear was measured with a dial thickness gauge, and the anti-inflammatory effect of He-Ne laser irradiation was expressed as an ear thickness ratio (ETR). Although the laser did not decelerate the ETR from ICD, the allergic response was decelerated. Irradiation at 5 min after the challenge of contact dermatitis increased the thickness ratio. Next, the influence of the He-Ne laser on histamine release from Wistar-rat peritoneal mast cells was observed. The spontaneous histamine release was inhibited by laser irradiation, while substance P and compound 48/80-induced histamine release were not inhibited. From these results, it can be suggested that He-Ne laser irradiation has an anti-inflammatory effect on cutaneous inflammation.

[Improved management of allergic contact eczema]. / Förbättrad handläggning av allergiska kontakteksem.

Allergic contact dermatitis (ACD) is common. Its diagnosis requires patch testing to establish ACD, followed by repeat open application testing (ROAT) to confirm that the putative allergen is in fact responsible for the ACD. The aim of primary management is to eliminate or minimise exposure to the allergen and its cross reactants. To promote healing and ease discomfort, local treatment is given, which in principle always consists of a short course of corticosteroids and an emollient. Sometimes antibiotics may be necessary, as well as systemic corticosteroid treatment for a brief period. In cases of persistent ACD, and sometimes in cases with a multifactorial background, further treatment may be required, first and foremost phototherapy. In isolated cases, treatment with low-energy roentgen (grenz rays), retinoids or cyclosporine may be called for. Desensitisation is not possible, and at present treatment with chelating agents and diet has no place in the management of ACD.

Suppressive and enhancing effects of ultraviolet B radiation on expression of contact hypersensitivity in man.

Ultraviolet B (UVB) radiation has multiple effects on the immune system, and these effects contribute to the development of UVB-induced skin cancers in mice, and probably man. Depending upon dose and duration of UVB exposure, the resultant immune aberrations may be strictly local (at the irradiated skin site) or systemic. One important local effect of acute, low-dose UVB regimens is impaired induction of contact hypersensitivity (CH). Because a significant proportion of humans who develop CH when hapten is painted on UVB-exposed skin fall to display a primary allergic reaction at that site, we inquired into the effects of UVB radiation on the expression of CH in man. A high proportion of individuals who were first exposed to a sensitizing dose of hapten via UVB-exposed skin displayed CH when challenged on unirradiated (normal) skin 11 d later. However, only 50% of these subjects developed CH when challenged simultaneously on skin that had been exposed to UVB radiation 11 d previously. Because the density of epidermal antigen-presenting cells was comparable in both responders and non-responders, we interpret these findings to mean that UVB radiation can create a sustained immunosuppressive microenvironment that inhibits the expression of CH. In separate experiments, when normal volunteers were sensitized with hapten via unirradiated (normal) skin, expression of CH at UVB-exposed challenge sites 11 d later was found to be enhanced, at least in some individuals, compared to expression of CH at unirradiated challenge sites. Thus, the local effects of UVB radiation on expression of CH in man may be enhancing or inhibitory, depending upon whether initial sensitization occurred through normal or through UVB-exposed skin.

Ultraviolet B light-induced suppression of contact sensitivity is not abolished by cyclophosphamide.

Dose responses for ultraviolet B light (UVB)-induced suppression of contact sensitivity were studied in mice, with and without cyclophosphamide (Cy) pretreatment, to investigate the role of Cy-sensitive suppression. Mice were irradiated on the back, sensitized on the abdomen, and challenged on the ears. Half of the mice were injected intraperitoneally with 200 mg/kg of Cy 3 days before being sensitized. Ultraviolet B light radiation reduced the ear swelling reactions in a linear relation to the log10 of the dose. Fifty percent suppression was shown by the computer-generated regression line at approximately 4.8 kJ/m2 of UVB radiation, with complete suppression at approximately 65 kJ/m2. In mice pretreated with Cy, ear swelling was increased, showing inhibition of a Cy-sensitive suppressive component of the contact sensitivity reaction. This Cy-sensitive component also was seen in mice treated with UVB, but with higher doses of UVB, there still was a UVB-dose-dependent decline in ear swelling in Cy pretreated mice, and there was complete suppression of reactions with the highest doses of UVB in the Cy-treated mice. Therefore, there is a second mechanism, not sensitive to Cy, that causes UVB-induced immune suppression.

Neutrophils, differentiated macrophages, and monocyte/macrophage antigen presenting cells infiltrate murine epidermis after UV injury.

We asked whether, as in humans, a population of antigen-presenting macrophages infiltrates the epidermis of ultraviolet (UV)-exposed BALB/c mice. Using three-color flow cytometry on cell suspensions plus in situ immunofluorescence microscopy, the phenotype of normal Langerhans cells was class II major histocompatibility complex (MHC+), CD11b+, NLDC-145+, BM8+ CD45+ and homogeneous. By contrast, in epidermal cells harvested 3 d following UV (UV-EC), there were two subsets of class II MHC+ cells: 1) class II MHChi CD11b+, and 2) class II MHClo CD11b-. Neither expressed the Langerhans cell markers BM8 and NLDC-145. In addition, there were two major populations of class II MHC- CD11b+ cells; half of these expressed the GR-1 neutrophil marker. Langerhans and dendritic epidermal T cells were markedly reduced after UV injury. By electron microscopy, immunomagnetic bead-purified CD11b+ cells in UV-EC were comprised of neutrophils, differentiated macrophages, and mononuclear cells with prominent lysosomes, but no Birbeck granules; the class II MHC+ subset resembled a monocytic cell in between differentiated macrophages and indeterminate dendritic cells. Functionally, immediately following in vivo UV exposure, the allogeneic antigen-presenting cell capacity of UV-EC was reduced to 21 +/- 6% of control epidermal cells (C-EC); by 3 d, antigen-presenting cell activity of UV-EC had recovered to 59 +/- 11% of C-EC, although at this time NLDC-145+ Langerhans cells had reached their lowest number. The recovered antigen-presenting cell activity was critically dependent upon the class II MHChiCD11b+ cells. Sensitization of BALB/c mice through skin that contained these antigen-presenting cells (3 d after UV) resulted in tolerance to dinitrofluorobenzene. By contrast, sensitization through UV-exposed skin immediately after the exposure resulted in unresponsiveness without tolerance, demonstrating temporal association of tolerance with leukocytic infiltration. In summary, murine epidermis responds to an acute UV injury in vivo with an initial abrogation of antigen-presenting activity followed by epidermal infiltration with neutrophils, differentiated macrophages, and monocytic antigen-presenting cells that are distinct from Langerhans cells with regard to expression of Langerhans cell markers and ultrastructure.

IL-3 production as an in vitro marker of the genetically determined traits of UVB susceptibility and UVB resistance.

The capacity of low-dose ultraviolet B (UVB) radiation to disrupt epidermal Langerhans cell function and to prevent the induction of contact hypersensitivity (CH) is genetically determined in mice and men. In mice, Tnf alpha and Lps are the genetic loci at which reside alleles that dictate susceptibility and resistance to the deleterious effect of UVB radiation. Detection of the UVB-susceptibility (UVB-S) and UVB-resistance (UVB-R) traits relies upon the in vivo end point of contact hypersensitivity, and is cumbersome, labor intensive, and time consuming. It has recently been reported that hapten-immune murine T cells can secrete interleukin-3 (IL-3) in vitro when exposed to hapten-derivatized syngeneic stimulator cells. To determine whether this assay might be useful in distinguishing UVB-R from UVB-S mice, panels of UVB-susceptible (C57BL/10, C3H/HeN) and UVB-resistant (A/J, BALB/c, C3H/HeJ) mice were sensitized epicutaneously with dinitrofluorobenzene (DNFB). When challenged in vitro 6 d later with dinitrophenyl-derivatized stimulator cells, T cells from all strains proliferated and secreted IL-3. Moreover, T cells from UVB-R mice that were sensitized through UVB-treated skin also made copious amounts of IL-3. However, T cells from UVB-S mice whose abdominal skin had been UVB irradiated prior to epicutaneous application of DNFB failed to secrete IL-3 in vitro, although the cells did proliferate. We conclude that following application of a sensitizing dose of hapten to UVB-exposed skin of UVB-S mice a) hapten-specific T cells are selectively unable to secrete IL-3 in vitro in response to hapten stimulation, and b) this inability is a reliable marker of the UVB-S trait. The IL-3 assay may prove useful in elucidating the mechanism by which UVB-exposed Langerhans cells activate regulatory T cells, and in detecting the UVB-S and UVB-R traits in humans.

Effect of local ultraviolet irradiation on infections of mice with Candida albicans, Mycobacterium bovis BCG, and Schistosoma mansoni.

In this study, we investigated whether mice given ultraviolet (UV)-B (280-320 nm) radiation in doses sufficient to alter cutaneous immune cells and impair the induction of contact hypersensitivity would also have impaired resistance to infectious agents administered at the site of UV irradiation. C3H mice were exposed to 400 J/m2 UVR from FS40 sunlamps on four consecutive days. Immediately after the last UV treatment, groups of mice were injected subcutaneously with Candida albicans, injected intradermally (ID) with Mycobacterium bovis bacillus Calmette-Guerin (BCG), or infected percutaneously with Schistosoma mansoni in UV-irradiated skin. The induction of the delayed hypersensitivity response to C. albicans and BCG, as assessed by footpad swelling, was unaffected by UV irradiation. However, the number of viable mycobacteria recovered from the lymphoid organs of BCG-infected mice was increased significantly in the UV-irradiated animals for a period of more than 2 months. Low-dose UV irradiation of the skin at the site of infection did not influence the number of S. mansoni parasites recoverable from the internal organs of mice that had been infected with cercariae percutaneously 6 weeks earlier. We conclude that the ability of UV radiation to impair the development of cell-mediated immunity to antigens introduced in a UV-irradiated site is not universal and depends on the particular antigen administered. We hypothesize that the involvement of epidermal Langerhans cells as the primary antigen-presenting cells in the induction of cell-mediated immunity may be the critical factor in determining whether a particular immune response will be affected by local UV irradiation.

Identification of the molecular target for the suppression of contact hypersensitivity by ultraviolet radiation.

This study was conducted to explore the involvement of DNA damage in the suppression of contact hypersensitivity (CHS) by UV irradiation. The opossum, Monodelphis domestica, was used because cells of these marsupials have an enzyme that is activated by visible light (photoreactivating enzyme) and repairs ultraviolet radiation (UVR)-induced pyrimidine dimers in DNA. A single dose of 1,500 J/m2 of UVB (280-320 nm) radiation, representing 2 minimal erythema doses, was administered to the dorsal skin of opossums. This treatment prevented the opossums from developing a CHS response to dinitrofluorobenze (DNFB) applied either at the site of irradiation or an unirradiated site. In addition, this dose of UVR decreased the number of ATPase+ epidermal Langerhans cells in the dorsal epidermis to approximately 3% of that in unirradiated skin at the time of DNFB application. Treatment of the animals with wavelengths that activate the repair enzyme (320-500 nm, photoreactivating light, PRL) for 120 min immediately after UV irradiation inhibited the UVR-induced suppression of CHS almost completely. Exposure to PRL before UVR did not prevent UVR-induced suppression of CHS. PRL treatment after UV irradiation also prevented the decrease in the number of ATPase+ Langerhans cells. Measurements of lesions in DNA indicated that PRL treatment removed around 85% of the UVR-induced pyrimidine dimers. These data provide direct evidence that DNA, and most likely, the pyrimidine dimer, is the primary molecular target for the UVB-induced suppression of contact hypersensitivity to haptens applied to irradiated or unexposed skin.

The duration of Grenz ray-induced suppression of allergic contact dermatitis and its correlation with the density of Langerhans cells in human epidermis.

Recent investigations have shown that Grenz rays can suppress the allergic contact dermatitis reaction completely and that Langerhans cells, identified by OKT6 antibodies and electron microscopy, disappear from the epidermis at the same time. It is not known for how long this suppression lasts. This has been investigated in 28 nickel-sensitive patients who were given Grenz rays (3 Gy) on the back, once a week for 3 weeks. The patients were then divided into four groups and tested with patch tests for nickel at 1, 7, 14 and 21 days after the last Grenz ray treatment. Biopsies were taken from positive patch test sites, and from the corresponding opposite control. They were labelled with OKT6 antibodies to detect Langerhans cells. The patch test reactions were suppressed and the Langerhans cell density was decreased initially. These changes were restored after 3 and 6 weeks, respectively. The results show that the effect of Grenz rays on eczematous reactions extends to a maximum of 3 weeks and imply that Langerhans cells are necessary for the elicitation of the efferent phase of allergic contact dermatitis.

Epidermal Langerhans' cells in chronic eczematous dermatitis of the palms treated with PUVA and UVB.

Epidermal Langerhans' cells (LC) were studied in patients with chronic eczematous dermatitis of the palms. The monoclonal antibodies anti-Leu 6 and anti-HLA-DR were used, and the cells visualized with an immunoperoxidase technique. Increased numbers of LC were found in allergic contact dermatitis as well as in irritant contact dermatitis and hyperkeratotic dermatitis of the palms. The increased number of epidermal LC in lesional skin may facilitate presentation of exogenous or endogenous antigens to activated T-cells. One hand was treated with PUVA or UVB and the other hand served as a non-exposed control. PUVA treatment cleared the dermatitis and the LC number decreased markedly. With UVB treatment clinical improvement was achieved, and a less pronounced decrease in epidermal LC was noticed. There seems to be a crude relationship between the extent of clinical improvement and the reduction in epidermal LC numbers. The change in LC might be a primary event or secondary to a reduction of the inflammatory process.

Desensitization to DNFB in hairless mice.

Naive hairless mice may be rendered partly tolerant to dinitrofluorobenzene (DNFB) by painting DNFB on skin irradiated with 30 mJ.cm-2 ultraviolet B light (UVB) over 4 days. However, DNFB-sensitized hairless mice show no decrease in sensitivity when repainted with DNFB on skin irradiated with the same dose of UVB. Hence, established hypersensitivity appears not to be reduced by this method of inducing tolerance in naive mice.

A double-blind study of Grenz ray therapy in chronic eczema of the hands.

The effect of Grenz ray therapy as an adjunct to topical therapy in chronic symmetrical eczema of the hands was assessed in 24 patients by randomly allocating active treatment to one hand while the other, which received simulated therapy, served as a control. Three Gy of Grenz rays were applied on six occasions at intervals of 1 week. There was a significantly better response to active treatment 5 and 10 weeks after the start of treatment compared with the untreated control.