[Anti- SSA/Ro and Anti-SSB/ La Antibodies in Patients with Yusho].

To investigate chronic immune effects of polychlorinated biphenyl (PCB) and polychlorinated dibenzofuran (PCDF), anti-SSA/Ro and anti-SSB/La antibodies were studied in serum of 213 patients with Yusho and 63 control subjects in 2013. Anti-SSA/Ro antibodies were found in 4.2% (9/213) of patients with Yusho and 1.6% (1/63) of control subjects. The prevalence rates of anti-SSA/Ro antibodies in Yusho patients were not associated with blood PCB concentration or blood 2,3,4,7,8-pentachlorodibenzofuran (PeCDF) concentration. Anti-SSB/La antibodies were found in 1.4% (3/213) of patients of Yusho, but none of controls. Furthermore, high concentrations of PCB and 2,3,4,7,8-PeCDF were present in Yusho patients with anti-SSB/La antibodies. We conclude that anti-SSB/La antibody is present in patients with Yusho, and it may be associated with PCB and 2,3,4,7,8-PeCDF in the blood.

[Serum Levels of Regulatory T Cell in the Yusho Patients].

Dioxin and dioxin-like compounds receptor (Ahr) mainly expressed on the surface of regulatory T (Treg) cell and Th17 cell could regulate immunological functions in the Yusho patients. We prospectively analyzed data obtained in a total of 56 cases of Yusho, which include patients identified ('Nintei' ) or non-identified ( 'Minintei') or identified as a family member, at the annual health check in 2014. The number of Treg cell showed lower among identified patients compared with non-identified group or family identified group (p = 0.4184 and p = 0.291, respectively). There was also a strong correlation between serum levels of neutral fat and the number of Treg cells (p = 0.0313). These results suggest that Treg cell plays a principal role in the immune response among Yusho patients.

[Antibodies to specific nuclear antigens in patients with Yusho].

To investigate chronic immune effects of polychlorinated biphenyl (PCB) and polychlorinated dibenzofuran (PCDF), autoantibodies including anti-Smith (Sm) antibody, anti-ribonucleoprotein (RNP) antibody, anti-centromere antibody and anti-double-stranded DNA (dsDNA) antibody, were studied in 168 patients with Yusho and 54 controls in 2012. Autoatibodies were present in some patients of Yusho; 1 case (0.6%) for anti-Sm antibody, 4 cases (2.4%) for anti-centromere antibody and 11 cases (6.5%) for anti-dsDNA antibody. However, these autoantibodies were not demonstrated in any controls. There was a significantly higher prevalence of elevated anti-centromere antibody in subjects with high PCB concentration than in those with low PCB concentration. We conclude that anti-centromere antibody is present in patients with Yusho, and it may be associated with blood PCB concentration.

Reduction of CC-chemokine ligand 5 by aryl hydrocarbon receptor ligands.

BACKGROUND: The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that recognizes a large number of xenobiotics, such as polycyclic aromatic hydrocarbons (PAHs), dioxins, and some endogenous ligands. Despite numerous investigations targeting AhR ligands, the precise physiological role of AhR remains unknown. OBJECTIVE: We explored novel AhR target genes, especially focused on inflammatory chemokine. METHODS: We treated (1) HaCaT, a human keratinocyte cell line, (2) normal human epidermal keratinocytes (NHEKs), and (3) mouse primary keratinocytes with AhR ligands, such as 6-formylindolo[3,2-b]carbazole (FICZ; endogenous ligand) and benzo[a]pyrene (BaP; exogenous ligand). Then, we detected mRNA and protein of chemokine using quantitative RT-PCR and ELISA. We next clarified the relationship between AhR and chemokine expression using AhR siRNA. In addition, we measured serum chemokine levels in patients with Yusho disease (oil disease), who were accidentally exposed to dioxins in the past. RESULTS: We identified CC-chemokine ligand 5 (CCL5), a key mediator in the development of inflammatory responses, as the AhR target gene. AhR ligands (FICZ and BaP) significantly reduced CCL5 mRNA and protein expression in HaCaT cells. These effects were observed in NHEKs and mouse primary keratinocytes. AhR knockdown with siRNA restored CCL5 inhibition by AhR ligands. In addition, AhR ligands exhibited a dose-dependent suppression of CCL5 production induced by Th1-derived cytokines. Finally, serum levels of CCL5 in patients with Yusho disease, were significantly lower than in controls. CONCLUSION: Our findings indicate that CCL5 is a target gene for AhR, and might be associated with the pathology of dioxin exposure.

Immune effects of hexachlorobenzene in the rat: role of metabolism in a 13-week feeding study.

The role of metabolism and porphyria in the immunomodulating effects of hexachlorobenzene (HCB) was investigated. To this end, female Wistar rats received a diet with two different doses of HCB and pentachlorobenzene (PCB), either in combination or not with the cytochrome P450IIIA1 inhibitor, triacetyloleandomycin (TAO). Urinary metabolites and urinary and liver porphyrins were measured at regular intervals and data have been published elsewhere. Skin lesions were scored weekly and after 13 weeks of exposure lymphoid organs were weighed, spleens were examined by morphometry, and serum.(auto)antibody levels were determined by ELISA. The probability of causal relationships between the different parameters was determined by analysis of correlation. HCB caused a dose-dependent increase of the incidence, but not the severity, of skin lesions, and dose-dependently increased weights of popliteal lymph nodes and spleen and serum levels of IgM, IgA, and autoantigen-specific IgM. IgG and IgG autoantibody levels were not changed. The splenomegaly could be attributed to an expansion of all splenic compartments. PCB caused no skin lesions and had only minor, predominantly immunosuppressive effects. TAO virtually lacked immunomodulating activity of its own, hardly affected the induction of skin lesions by HCB, did not change the immune effects of HCB, and suppressed IgG levels when combined with PCB. Comparison of the immunological data with those found in the same rats as to induction of porphyria and biotransformation of HCB and PCB, indicates that the HCB-induced porphyria, being markedly reduced by coadministration of TAO, is not involved in the immunomodulating effects of HCB. The same conclusion could be drawn for the oxidative HCB metabolites, since TAO inhibited their formation, while the same metabolites were formed upon administration of PCB that lacked the immunostimulatory effects of HCB. Therefore, HCB itself, its nonoxidative metabolites, or their precursors are likely candidates for inducing the immune effects. Further, an immune component in the HCB-induced skin lesions, usually associated with dermal porphyrin accumulation, is suggested by the observations that TAO profoundly reduced induction of porphyria, but not of skin lesions and immune effects, by HCB.

Porphyria cutanea tarda associated with autoimmune hypothyroidism, vitiligo and alopecia universalis.

The aetiology of porphyria cutanea tarda (PCT) has not been elucidated, but the possibility of an autoimmune mechanism has been proposed. We report a case of an unknown clinical combination of PCT with autoimmune hypothyroidism, alopecia universalis and vitiligo with thyroid and parietal cell circulating antibodies. This is highly suggestive of underlying autoimmune damage in this patient.

Studies of laminin and type IV collagen in blisters of porphyria cutanea tarda and drug-induced pseudoporphyria.

Blisters from five patients with porphyria cutanea tarda and two patients with drug-induced pseudoporphyria were examined by direct immunofluorescence and by immunofluorescence mapping with antibodies against laminin and type IV collagen to determine the level of subepidermal separation. Primary screening by direct immunofluorescence revealed the characteristic immune deposits in the vessel walls of the upper dermal plexus in all cases and at the dermoepidermal junction in five of seven cases. Type IV collagen and laminin were reactive in six and five cases, respectively, and appeared in the floor of the bulla. The findings were identical in porphyria and pseudoporphyria. In one case in which the bullous pemphigoid antigen could be detected, it appeared in the epidermal roof of the bulla. These findings indicate that the split in porphyria and pseudoporphyria occurs in the lamina lucida. We propose that a multistep mechanism involved in the induction of blisters may be similar in porphyria and pseudoporphyria.

Antinuclear antibodies and the diagnosis of systemic lupus erythematosus in patients with acute intermittent porphyria.

To investigate the previously postulated association of systemic lupus erythematosus (SLE) and porphyria 38 patients with various types of porphyria were investigated for clinical and laboratory evidence of a connective tissue disease. Antinuclear antibodies (ANAs) were found in 8/15 (53%) patients with acute intermittent porphyria. These patients were more likely to have had a recent acute attack of porphyria, but only one patient had clinical evidence of SLE. Antinuclear antibodies were not found in any patients with latent acute intermittent porphyria or appreciably in any of the other types of porphyria studied. This finding of ANAs in patients with acute intermittent porphyria may explain the previously described association with SLE. Strict diagnostic criteria need to be used in any one patient as these two disorders have many similar clinical manifestations.

Mechanisms of phototoxicity in porphyria cutanea tarda and erythropoietic protoporphyria.

The porphyrias are the only group of diseases caused by endogenous phototoxic agents. While patients with erythropoietic protoporphyria and those with porphyria cutanea tarda both have skin lesions on sun-exposed areas, there are differences in their cutaneous manifestations. Based on information discussed in this chapter, the following pathophysiologic mechanisms can be proposed. In porphyria cutanea tarda, photoactivation of the complement system in the presence of uroporphyrin results in activation of dermal mast cells, which release their proteases. This results in dermal-epidermal separation, reflected clinically as skin fragility and vesicles. The interaction between activated mast cells with fibroblasts, the nature of which is still unclear, may contribute to fibrosis and sclerodermoid skin changes. The stimulatory effect of uroporphyrin on collagen biosynthesis by fibroblasts, which occurs independent of irradiation, may be responsible for the sclerodermoid lesions seen at sun-exposed as well as sun-protected areas. In erythropoietic protoporphyria, mast cell activation can occur as the result of complement activation induced by protoporphyrin and irradiation. Protoporphyrin and irradiation may also directly induce the release of preformed and generated mediators from mast cells, a process mediated at least in part by peroxidation. The release of mast cell mediators may account for the erythema, edema, and urticaria observed in patients with erythropoietic protoporphyria upon exposure to sunlight. Interaction of mast cells with fibroblasts, and the direct membrane-damaging effect of protoporphyrin and irradiation on the latter, may contribute to the waxy thickening of skin seen in chronically sun-exposed areas of these patients. There are, however, many unanswered questions. What accounts for the different biological effects of mast cell-derived mediators: dermal-epidermal separation in one, erythema and urticaria in the other? The fragmentation of dermal collagen bundles associated with cleavage beneath the lamina densa, and the hyperpigmentation and hypertrichosis observed in some patients with porphyria cutanea tarda remain unexplained. What is the mechanism of the reduplication of blood vessel basal lamina in the non-sun-exposed areas of both types of patient? Are there any roles for cytokines and epidermal cell-derived eicosanoids? While it is clear that the pathogenesis of cutaneous lesions in porphyria cutanea tarda and erythropoietic protoporphyria involves interactions among inflammatory mediators and various cells in skin, much still needs to be done to further our understanding of their pathophysiology.

Antibodies to the Borrelia burgdorferi flagellum in patients with scleroderma, granuloma annulare and porphyria cutanea tarda.

It is generally accepted that cutaneous Lyme borreliosis comprises erythema chronicum migrans, lymphadenosis benigna cutis, and acrodermatitis chronica atrophicans. In recent years the tick-borne spirochete Borrelia burgdorferi has been associated with a number of other cutaneous disorders. We therefore investigated sera from 175 patients with localized scleroderma (morphea) (n = 64), systemic sclerosis (n = 74), granuloma annulare (n = 16) and porphyria cutanea tarda (n = 21) with the new, highly sensitive and specific Borrelia burgdorferi flagellum ELISA assay. As controls (n = 297) served normal healthy volunteers and patients with other skin diseases. It was found that the distribution of individual antibody values and the median antibody levels were identical in controls and in patients with scleroderma, granuloma annulare and porphyria cutanea tarda. These data do not support the hypothesis of an etiological association between Borrelia burgdorferi infection and scleroderma, granuloma annulare or porphyria cutanea tarda.

Increased frequency of HLA-A3 in subjects with sporadic porphyria cutanea tarda.

The HLA-A3 alloantigen, usually associated with the hemochromatosis gene, was detected in 12 of 18 patients (67%) with sporadic porphyria cutanea tarda but in only 23 per cent of 328 normal subjects (p = 0.0006). This difference remained significant after correcting for the number of HLA-A locus antigens tested (p = 0.025). These results suggest that an HLA-linked hemochromatosis allele is present and may account for the iron abnormalities in many patients with sporadic porphyria cutanea tarda.

Immunohistochemical studies on vitronectin in elastic tissue disorders, cutaneous amyloidosis, lichen ruber planus and porphyria.

Vitronectin, identical with serum-spreading factor and S-protein of complement, is a glycoprotein present in both plasma and tissue. It stimulates cell adhesion and spreading and affects the complement and coagulation pathways. Vitronectin immunoreactivity was recently found in conjunction with dermal and renal elastic fibres, in renal amyloid deposits in cases of AL- and AA-amyloidosis, and in sclerotic glomerular lesions. Skin specimens from lesions of patients with selected skin diseases were investigated with an avidin-biotin peroxidase technique using both monoclonal and polyclonal anti-vitronectin antibodies and an alkaline phosphatase anti-alkaline phosphatase technique using monoclonal anti-vitronectin antibodies. Vitronectin immunoreactivity was found in association with the abnormal elastic tissue in solar elastosis and pseudoxanthoma elasticum. It was also found in conjunction with dermal amyloid deposits in primary localized cutaneous amyloidosis and in Civatte bodies in cases of lichen ruber planus. In cases of erythropoietic protoporphyria and porphyria cutanea tarda, hyaline perivascular deposits also demonstrated positive vitronectin immunoreactivity. The presence of vitronectin immunoreactivity not only in normal and degenerated elastic fibres but also in various pathological tissue deposits suggests that vitronectin occurs both in elastic fibres and in different types of abnormal protein deposits.

Porphyria cutanea tarda and HLA-linked hemochromatosis. Evidence against a systematic association.

This study was designed to test the hypothesis that a hemochromatosis allele is implicated in the expression of porphyria cutanea tarda. HLA phenotypes were determined in 69 porphyria cutanea tarda patients, 42 of which had the sporadic type (normal erythrocyte uroporphyrinogen decarboxylase activity) and 27 unrelated patients who had the familial type (diminished erythrocyte uroporphyrinogen decarboxylase activity). The incidence of HLA antigen A3, a marker of the hemochromatosis allele, was identical in the sporadic patients (23.8%), in the familial patients (22.2%), and in the controls (24.5%). Furthermore, no clinical difference could be found between A3 and non-A3 patients. These results demonstrate no systematic association between hemochromatosis and porphyria cutanea tarda in the population studied. Another HLA-linked gene, however, could be implicated in the expression of the disease as HLA antigen DR7 presented an incidence statistically different (p less than 0.05) between sporadic (16.6%) and familial patients (43%).

Porphyria cutanea tarda associated with the acquired immune deficiency syndrome.

Three male subjects with cutaneous symptoms and biochemical signs typical of porphyria cutanea tarda (PCT) developed acquired immune deficiency syndrome (AIDS). All three were in a classic high risk group for the latter disease and developed a typical progressive illness. Two patients succumbed to opportunistic infections; the third is alive but critically ill. The symptomatic prodrome of AIDS developed concurrently with or followed the onset of symptoms of PCT in all three individuals. PCT and AIDS are both uncommon disorders; their association in three patients is thus of inherent clinical interest. If this association is not coincidental, it raises the possibility that the occurrence of photosensitivity, skin lesions, and evidence of biochemical changes characteristic of PCT may, in certain patients at risk for AIDS, presage the subsequent full clinical expression of the latter disease.

Complement cleavage products in the phototoxic reaction of porphyria cutanea tarda.

We have measured C3, C4, CH50 and complement cleavage products C3a and C5a in in sera and plasma from PCT patients and normal controls 10 min and 1, 4 and 24 h after UVA irradiation. We found elevated C3a concentrations in PCT patients immediately after UVA irradiation and 24 h later. The same was true for CH50, whereas C3, C4 and C5a did not change significantly. No such changes occurred in normal controls. Our data suggest that activation of the complement cleavage product C3a by porphyrin and UV light triggers a series of events that cause tissue damage.