Ultraviolet radiation and the contact hypersensitivity reaction in mice

The article reviews the application of the contact hypersensitivity assay in mice to the science of photoimmunology. The contact hypersensitivity (CHS) reaction, which is suppressed by UV irradiation in mice similarly to their ability to respond immunologically to skin tumors, has been used very profitably to reveal many of the regulating factors that control photoimmunosuppression, such as the identity of the photoreceptors that initiate immunosuppression, the defects induced in the cutaneous antigen presenting pathway, the local cytokine imbalance, and the protective intervention by various molecules, drugs, or interacting UV wavebands. Technical hints to optimize the measurement of the CHS response are suggested, including information on UV radiation wavebands and dosages and sensitivities of different mouse strains.     

Polymorphisms in the promoter region of ESR2 gene and susceptibility to ovarian cancer

Susceptibility to ovarian cancer might be affected by genetic variations in genes involved in estrogen biosynthesis, metabolism or signal transduction. In this study we tested the hypothesis that single nucleotide polymorphisms (SNPs) in the promoter of human ESR2 gene, coding for estrogen receptor β, may be associated with increased risk for ovarian cancer. Three SNPs in the promoter region of human ESR2 gene were genotyped by means of allele-specific tetra-primer PCR. A total of 184 ovarian cancer cases and the same numbers of controls were included in the study. With regard to homozygous analysis, the AA genotype of SNP rs3020449 was found to be significantly more frequent in ovarian cancer cases staged as FIGO III + IV than in cases staged as I + II (OR 2.717, p = 0.027). With regard to allele frequency, the G allele of this SNP was less frequent in FIGO I + II cases than in cases with higher FIGO stages (OR 1.739, p = 0.018). With regard to genotype frequency, allele frequency, allele positivity or haplotype frequency of SNPs rs2987983, rs3020449 and rs3020450 we did not observe a significant difference between the cancer and the control group. Our data suggest that SNPs in the promoter region of ESR2 gene do not affect susceptibility to ovarian cancer, but SNP rs3020449 might affect progression of this disease.     

Conserved loci on the X chromosome confer phosphate homeostasis in mice and humans

Several genes expressed in kidney and other tissues determine phosphate homeostasis in extracellular fluid. The major form of inherited hypophosphatemia in humans involves an X-linked locus (HPDR, Xp22.31-p21.3). It has two murine homologues (Hyp and Gy) which map to closely-linked but separate loci (crossover value 0.4%-0.8%). Both murine mutations impair Na(+)-phosphate cotransport in renal brush border membrane; an associated renal disorder of 1,25-dihydroxyvitamin D3 (1,25(OH)2D) metabolism has been characterized in Hyp mice. Whereas experiments with cultured Hyp renal epithelium indicate that the gene is expressed in kidney, studies showing the development of the mutant renal phenotype in normal mice parabiosed to Hyp mice implicate a circulating factor; these findings can be reconciled if the humoral factor is of renal origin. The gene dose effect of HPDR, Hyp and Gy on serum phosphorus values is consistently deviant and heterozygotes resemble affected hemizygotes. The deviant effect is also seen on renal phosphate transport; all mutant females (Hyp/Hyp and Hyp/+) have similar phenotypes. On the other hand, there is a normal gene dose effect of HPDR in mineralized tissue; tooth PRATIO (pulp area/tooth area) values for heterozygotes are distributed between those for affected males and normals. The tooth data imply that the X chromosome locus is expressed in both renal and non-renal cells. The polypeptide product of the X chromosome gene(s) is still unknown.     

Measurement of ultraviolet radiation-induced suppression of recall contact and delayed-type hypersensitivity in humans

This article describes methodology used for assessment of ultraviolet radiation-induced suppression of recall responses in humans. Nickel allergy is common in the general population and patch testing of nickel-allergic volunteers provides a convenient model of contact hypersensitivity. Similarly, Mantoux-positive volunteers, recruited from within hospital staff, are used as a model for delayed-type hypersensitivity. Use of secondary rather than primary immune responses allows placement of multiple test sites on each volunteer. Further, each volunteer acts as his or her own unirradiated control. This enables UV immunosuppression to be studied with relatively few human volunteers, and makes determination of UV immunosuppression dose responses feasible in human subjects. The method can also be used for assessment of the level of immune protection afforded by agents such as sunscreens or biologically active substances.     

Polymorphisms of the TNF gene and the TNF receptor superfamily member 1B gene are associated with susceptibility to ulcerative colitis and Crohn's disease, respectively

The importance of tumor necrosis factor (TNF)-alpha and the TNF receptor gene polymorphisms in the etipathogenesis of inflammatory bowel disease (IBD) has not been elucidated. DNA from peripheral blood samples was obtained from 124 patients with Crohn's disease (CD), 106 patients with ulcerative colitis (UC), and 111 unrelated healthy controls. We examined two single nucleotide polymorphisms (SNPs) of the TNF-alpha gene, TNF (-308 G/A and -238 G/A), an SNP of the TNF receptor superfamily member 1A gene, TNFRSF1A(also known as TNFR1), at codon 12 in exon 1 (CCA/CCG), and two SNPs of the 1B gene, TNFRSF1B (also known as TNFR2), (1466 A/G and 1493 C/T). There was a difference in the carrier frequency for haplotype AG (-308 A, -238 G) between UC patients and the controls (OR=4.76, 95% CI=1.53-14.74, P<0.01). We found a significant difference in carrier frequency for haplotype AT (1466 A, 1493 T) of the TNFRSF1B gene between CD patients and the controls (OR=2.13, 95% CI=1.08-4.21, P<0.05). The significance proved to be greater in CD patients with both internal and external fistula (OR=4.8, 95% CI=1.73-13.33, P<0.01), and in those who were poor responders ( n=22) to our treatments, which consisted of nutritional therapy, medical therapy and surgical therapy (OR=9.24, 95% CI=3.37-25.36, P<0.001). This study suggests that one of the genes responsible for UC may be the TNF gene, or an adjacent gene, and that TNFRSF1B gene polymorphisms contribute greatly to the increased onset risk of CD and to the disease behavior.     

Multiple Gln/Asn-rich prion domains confer susceptibility to induction of the yeast [PSI(+)] prion

The yeast prion [PSI(+)] results from self-propagating aggregates of Sup35p. De novo formation of [PSI(+)] requires an additional non-Mendelian trait, thought to result from a prion form of one or more unknown proteins. We find that the Gln/Asn-rich prion domains of two proteins, New1p and Rnq1p, can control susceptibility to [PSI(+)] induction as well as enhance aggregation of a human glutamine expansion disease protein. [PSI(+)] inducibility results from gain-of-function properties of New1p and Rnq1p aggregates rather than from inactivation of the normal proteins. These studies suggest a molecular basis for the epigenetic control of [PSI(+)] inducibility and may reveal a broader role for this phenomenon in the physiology of protein aggregation.     

IFN-gamma and TNF-alpha are involved in urushiol-induced contact hypersensitivity in mice

Contact hypersensitivity (CHS) is a cutaneous T-cell-mediated immunological reaction to applied haptens. Activated antigen-specific T cells release several cytokines and chemokines followed by the recruitment of inflammatory cells and skin damage. CD8+ T cells and CD4+ T cells have been involved in the establishment of previously described CHS. In this study, we investigated the induction of CHS by urushiol in mice. Maximum swelling in mouse ears was elicited 24 h after challenge with urushiol on day 9 of sensitization. IFN-gamma, TNF-alpha and IFN-gamma-inducible protein 10 (IP-10) mRNA were expressed after challenge of the antigen in urushiol-sensitized mice, but not in unsensitized mice. IFN-gamma knockout (KO) mice and TNF-alpha KO mice failed to elicit CHS with urushiol. Contact hypersensitivity and expressions of IFN-gamma, TNF-alpha and IP-10 mRNA were markedly suppressed in CD4+ and CD8+ cell-depleted mice. These results suggest that IFN-gamma, TNF-alpha, and possibly IP-10, play a critical role in CHS induced by urushiol, depending on both CD4+ T cells and CD8+ T cells.     

Langerhans cells that migrate to skin after intravenous infusion regulate the induction of contact hypersensitivity

Intravenous infusion of hapten-derivatized epidermal cells (EC) in syngeneic mice leads to two competing signals for contact hypersensitivity (CH), a dominant effector signal attributable to Langerhans cells (LC) and a suppressor signal from Thy-1+ EC. In vitro exposure of LC to low dose ultraviolet B (UVB) radiation before hapten-derivatization and infusion not only results in the abrogation of their effector signal but also causes the down-regulation of subsequent CH responses. To delineate the relevance of i.v. immunization to the study of CH and of LC as the immunologic targets of low dose UVB radiation, we examined the migratory and immunogenic properties of EC after i.v. infusion. Unsorted EC migrated from blood to skin and lymphoid tissues, reaching steady state distributions at 16 h after infusion. No significant differences were observed between the trafficking of EC in syngeneic and allogeneic transfers. LC localized preferentially to skin, whereas Thy-1+ EC trafficked to skin, the thymus, mesenteric lymph nodes, and spleen. The pattern of trafficking of unirradiated and low dose UVB-irradiated LC were identical, suggesting that low dose UVB radiation had little effect on LC migration. Finally, skin graft experiments demonstrated i.v. infused, hapten-derivatized LC that migrate to skin to retain their capacity to induce CH, a property that was converted by in vitro pretreatment with low dose UVB radiation into down-regulation. These findings confirm the relevance and utility of the i.v. immunization model in the study of CH and the influence of low dose UVB on this immune response. Our data also provide a basis for investigating the role of disparate trafficking patterns in generating effector and suppressor signals when hapten-derivatized EC are employed for CH.     

IL-10 contributes to the inhibition of contact hypersensitivity in mice treated with photodynamic therapy

We have explored the effect of photodynamic therapy (PDT) with verteporfin on the induction and expression of contact hypersensitivity (CHS) to 2,4-dinitrofluorobenzene (DNFB) in normal mice and IL-10-deficient mice. Our results indicate that DNFB sensitized mice given PDT with verteporfin and whole body red light irradiation exhibited a significant reduction in CHS compared with control animals. Administration of rIL-12 reversed the effect(s) of PDT as did treatment of mice with anti-IL-10-neutralizing Ab. Knockout mice deficient in IL-10 were found to be resistant to the inhibitory effects of PDT. In vitro proliferative responses using spleen cells from DNFB-sensitized and PDT-treated mice showed a significantly lower response to DNBS as compared with cells from DNFB-sensitized mice or DNFB and PDT-treated IL-10-deficient mice. Finally, naive mice exposed to PDT exhibited an increase in skin IL-10 levels, which peaked between 72 and 120 h post-PDT. Together these data support the role of IL-10 as a key modulator in the inhibition of the CHS response by whole body PDT.     

The role of CXCR2 activity in the contact hypersensitivity response in mice

The recruitment of polymorphonuclear neutrophil leukocytes (PMN) into a challenge site, and their subsequent activation, are thought to play a role in the elicitation of the contact hypersensitivity (CHS) response. The present study investigated the role played by CXCR2 activity in tissue PMN infiltration and subsequent triggering of CHS. Our results show that the cutaneous infiltration by PMN, induced by hapten challenge was dramatically inhibited in sensitized, CXCR2-deficient (CXCR2(-/-)) mice. Inhibition of PMN recruitment into the hapten-challenged ears of CXCR2(-/-) mice was associated with a consistent reduction of the CHS response (ear swelling) in CXCR2(-/-) mice as compared with that observed in neutropenic, wild-type (CXCR2(+/+)) mice. Prevention of skin PMN infiltration and the ear swelling response by the absence of functional CXCR2 was observed regardless of the hapten used. These data clearly suggest that CXCR2 activity plays an essential role in mediating cutaneous recruitment and activation of PMN, and thus indirectly regulates recruitment of hapten-primed T cells into challenge sites, with the subsequent elicitation of the CHS response. The role played by CXCR2 activity in the CHS response provides the rationale for testing CXCR2 inhibitors as a new therapeutic approach to skin diseases.     

Polymorphisms in autophagy genes and susceptibility to tuberculosis

Recent data suggest that autophagy is important for intracellular killing of Mycobacterium tuberculosis, and polymorphisms in the autophagy gene IRGM have been linked with susceptibility to tuberculosis (TB) among African-Americans, and with TB caused by particular M. tuberculosis genotypes in Ghana. We compared 22 polymorphisms of 14 autophagy genes between 1022 Indonesian TB patients and 952 matched controls, and between patients infected with different M. tuberculosis genotypes, as determined by spoligotyping. The same autophagy polymorphisms were studied in correlation with ex-vivo production of TNF, IL-1β, IL-6, IL-8, IFN-γ and IL-17 in healthy volunteers. No association was found between TB and polymorphisms in the genes ATG10, ATG16L2, ATG2B, ATG5, ATG9B, IRGM, LAMP1, LAMP3, P2RX7, WIPI1, MTOR and ATG4C. Associations were found between polymorphisms in LAMP1 (p = 0.02) and MTOR (p = 0.02) and infection with the successful M. tuberculosis Beijing genotype. The polymorphisms examined were not associated with M. tuberculosis induced cytokines, except for a polymorphism in ATG10, which was linked with IL-8 production (p = 0.04). All associations found lost statistical significance after correction for multiple testing. This first examination of a broad set of polymorphisms in autophagy genes fails to show a clear association with TB, with M. tuberculosis Beijing genotype infection or with ex-vivo pro-inflammatory cytokine production.     

Inhibitory effect of dietary carotenoids on dinitrofluorobenzene-induced contact hypersensitivity in mice

We evaluated the effect of carotenoids on the dinitrofluorobenzene (DNFB)-induced contact hypersensitivity in mice. Dietary carotenoids significantly inhibited ear swelling and reduced the contents of TNF-α and histamine in the DNFB-treated mice. Our results suggest that dietary carotenoids exerted an anti-inflammatory effect by suppressing mast cell degranulation in vivo.     

Estrogen receptor-beta signaling protects epidermal cytokine expression and immune function from UVB-induced impairment in mice.

A previous study in the hairless mouse, in which the photoimmune protective properties of a topical phytoestrogen or 17-beta-estradiol were abrogated by the estrogen receptor antagonist ICI 182,780, revealed that estrogen receptor (Er) signaling is involved in the regulation of the suppression of immune function by UVB (290-320 nm) radiation. Here we identify the expression of Er-beta but not Er-alpha mRNA in hairless mouse skin, whereas Er-alpha and Er-beta mRNA were present in normal haired mouse skin. This suggests that the non-classical estrogen target Er-beta is involved in the photoimmune modulation, and is consistent with Er-alpha being more closely associated with hair growth control, as indicated by other studies. In mice with a null mutation for Er-beta, there was a significant exacerbation of the solar simulated UV (290-400 nm)-induced suppression of contact hypersensitivity. Immunohistochemical analysis revealed that the Er-beta deficiency inhibited the normally immunoprotective upregulation by the UVA (320-400 nm) waveband of the epidermal expression of the cytokines IFN-gamma and IL-12. Er-beta deficiency also significantly increased the UVB-induced expression of the immunosuppressive cytokine IL-10. Thus Er signalling via the Er-beta is evidently a major regulator of the UVA and UVB waveband interactions that determine the skin's immune functional status, and achieves this by normalization of the cutaneous cytokine array in the UV-irradiated skin.     

Diets containing pomegranate polyphenol and soy isoflavone attenuate contact hypersensitivity in mice

Contact hypersensitivity (CHS) is frequently used as an animal model for human allergic contact dermatitis (ACD). Diets of pomegranate polyphenols (PPs) or soy isoflavones (SIs) each alleviated CHS symptoms; however, the effect of diets containing a mixture of PPs and SIs on CHS is unclear. We investigated the CHS-inhibitory effects of diets supplemented with a mixture of PPs and SIs at human physiologically relevant doses. Consuming the mixture of PPs and SIs attenuated ear swelling and reduced infiltration of Gr-1-positive cells. Ear swelling decreased in the PP and SI-treated mice compared to the SI-treated mice. The auricle tissues of the PP and SI-fed mice exhibited decreased production of CXCL2 and MCP-5 compared to the SI- and PP-treated mice, respectively. These results suggest that dietary supplementation with a mixture of PPs and SIs may have ACD-preventive effects and may prove more beneficial than supplementation with PPs or SIs alone.     

Polymorphisms in the promoter region of ESR2 gene and breast cancer susceptibility

Genetic variations like single nucleotide polymorphisms (SNPs) in genes involved in estrogen biosynthesis, metabolism and signal transduction have been suggested to affect breast cancer susceptibility. In this study we tested the hypothesis that polymorphisms in the promoter of ESR2 gene may be associated with increased risk for breast cancer. We analyzed three SNPs in the promoter region of human ESR2 gene by means of allele-specific tetra-primer PCR. A total of 318 sporadic breast cancer cases and 318 age-matched controls were included in the study. With regard to homozygous genotypes, women with sporadic breast cancer more frequently carried the CC genotype of ESR2 promoter SNP rs2987983 (OR 1.99, p = 0.005). Calculation of allele positivity demonstrated that presence of T allele of this SNP was more frequent in healthy women. Our data suggest that a SNP in the promoter region of ESR2 gene might be able to affect breast cancer risk. These results further support the emerging hypothesis that ERβ is an important factor in breast cancer development.     

Suppression of contact hypersensitivity after repeated exposures of humans to low doses of solar simulated radiation.

Although it is generally recognised that UV radiation (UVR) can induce suppression of contact hypersensitivity (CHS) in human subjects, most protocols to date have not tested the effect of low daily doses of solar simulated radiation (SSR). In the present study, healthy individuals, divided into four groups each consisting of approximately 34 subjects, were whole-body irradiated with 1.2 standard erythema doses of SSR for 2, 10 or 30 consecutive days, or were unirradiated. They were sensitised with diphenylocyclopropenone (DPCP) on one exposed body site 24 h after the final UVR. The occurrence and severity of the primary allergic response were noted, and both parameters were shown to be significantly lowered in the group irradiated for 30 days compared with the unirradiated group. Elicitation of CHS was undertaken 3 weeks after the sensitisation, using a range of concentrations of DPCP on a UV-protected body site. The extent of the CHS at 48 h was assessed by the clinical score, by an erythema meter and by histological examination of a biopsy taken from the site challenged with one selected concentration of DPCP. Although erythema and pigmentation did not differ between the groups, a significant negative correlation was found between the clinical CHS score and the number of days of UV exposure, at the lowest challenge dose of DPCP. In addition a significant negative correlation was revealed between the intensity of spongiosis (intraepidermal oedema and vesicles, as evaluated by histology) and the number of days of UV exposure. Thus small daily doses of SSR induce suppression of CHS in human subjects and the effect is cumulative, indicating that there is no adaptation to the immunomodulating effects of UVR, at least over the test period of 30 days.     

Consanguinity and susceptibility to infectious diseases in humans

Studies of animal populations suggest that low genetic heterozygosity is an important risk factor for infection by a diverse range of pathogens, but relatively little research has looked to see whether similar patterns exist in humans. We have used microsatellite genome screen data for tuberculosis (TB), hepatitis and leprosy to test the hypothesis that inbreeding depression increases risk of infection. Our results indicate that inbred individuals are more common among our infected cases for TB and hepatitis, but only in populations where consanguineous marriages are common. No effect was found either for leprosy, which is thought to be oligogenic, or for hepatitis in Italy where consanguineous marriages are rare. Our results suggest that consanguinity is an important risk factor in susceptibility to infectious diseases in humans.