Immunoelectron microscopy study of superficial skin nerves in drug-induced acute urticaria / Estudo de microscopia imunoeletrônica dos nervos superficiais da pele na urticária aguda induzida por medicamentos

BACKGROUND: Few studies have evaluated the ultrastructure of the superficial skin nerves in urticaria. OBJECTIVE: The objective of this study was to describe findings in superficial skin nerves in cases of drug-induced acute urticaria. METHODS: Seven patients with drug-induced acute urticaria were included in the study. Skin biopsies were obtained from the urticarial lesion and from the apparently normal skin. The 14 fragments collected were processed for immunogold electron microscopy using single stains for antitryptase and anti-FXIIIa antibodies, as well as double immunogold labeling for both. RESULTS: Some sections showed mast cells in the process of degranulation. Following double immunogold staining, 10 nm (FXIIIa) and 15 nm (Tryptase) gold particles were found together throughout the granules in mast cells, indicating that tryptase and FXIIIa are located inside each one of the granules of these cells. Interestingly, we found strong evidence of the presence of tryptase and factor XIIIa in the superficial skin nerves of these patients, both in cases of urticarial lesions (wheals) and in the apparently normal skin. CONCLUSIONS: Tryptase and FXIIIa are present in the superficial nerves of the skin in drug-induced acute urticaria. This is the first report of tryptase and FXIIIa expression in the superficial skin nerves of patients with urticaria. Tryptase may be participating in neural activation in these patients, while FXIIIa may be present in the nerves to guarantee the functional integrity of structures.

FUNDAMENTOS: Poucos autores têm estudado a ultraestrutura dos nervos superficiais na urticária. OBJETIVO: Descrever os achados nos nervos cutâneos superficiais em casos de urticária aguda induzida por medicamentos. MÉTODOS: Sete pacientes com urticária aguda induzida por medicamentos foram incluídos no estudo. Foram obtidas biopsias da pele da lesão urticariforme e da pele aparentemente normal. Os 14 fragmentos coletados foram processados usando imunomarcação com ouro para anticorpos anti-triptase e anti-FXIIIa separadamente, além da dupla imunomarcação com ambos anticorpos. A seguir as amostras foram submetidas à análise por microscopia imunoeletrônica. RESULTADOS: Alguns cortes demonstraram mastócitos em processo de degranulação. Após a imunomarcação dupla, partículas de ouro de 10 nm (FXIIIa) e partículas de ouro de 15 nm (Triptase) apresentavam-se juntas em grânulos de mastócitos indicando que a triptase e o FXIIIa se localizam dentro de cada um dos grânulos dessas células. Curiosamente, foi encontrada uma forte evidência da presença da triptase e do fator XIIIa nos nervos superficiais dos pacientes avaliados, tanto em lesões urticadas, como na pele aparentemente normal. CONCLUSÕES: A triptase e o FXIIIa estão presentes nos nervos superficiais da pele na urticária aguda medicamentosa. Este é o primeiro relato da expressão de triptase e de FXIIIa nos nervos superficiais na urticária. A triptase poderia estar participando da ativação neural nos pacientes estudados. O FXIIIa poderia estar presente nos nervos, com a finalidade de manter a integridade funcional dessas estruturas.

Protease and Protease-Activated Receptor-2 Signaling in the Pathogenesis of Atopic Dermatitis

Proteases in the skin are essential to epidermal permeability barrier homeostasis. In addition to their direct proteolytic effects, certain proteases signal to cells by activating protease-activated receptors (PARs), the G-protein-coupled receptors. The expression of functional PAR-2 on human skin and its role in inflammation, pruritus, and skin barrier homeostasis have been demonstrated. Atopic dermatitis (AD) is a multifactorial inflammatory skin disease characterized by genetic barrier defects and allergic inflammation, which is sustained by gene-environmental interactions. Recent studies have revealed aberrant expression and activation of serine proteases and PAR-2 in the lesional skin of AD patients. The imbalance between proteases and protease inhibitors associated with genetic defects in the protease/protease inhibitor encoding genes, increase in skin surface pH, and exposure to proteolytically active allergens contribute to this aberrant protease/PAR-2 signaling in AD. The increased protease activity in AD leads to abnormal desquamation, degradation of lipid-processing enzymes and antimicrobial peptides, and activation of primary cytokines, thereby leading to permeability barrier dysfunction, inflammation, and defects in the antimicrobial barrier. Moreover, up-regulated proteases stimulate PAR-2 in lesional skin of AD and lead to the production of cytokines and chemokines involved in inflammation and immune responses, itching sensation, and sustained epidermal barrier perturbation with easier allergen penetration. In addition, PAR-2 is an important sensor for exogenous danger molecules, such as exogenous proteases from various allergens, and plays an important role in AD pathogenesis. Together, these findings suggest that protease activity or PAR-2 may be a future target for therapeutic intervention for the treatment of AD.

Methylthioadenosine phosphorylase expression in cutaneous squamous cell carcinoma.

The methylthioadenosine phosphorylase (MTAP) gene is a tumour suppressor gene, located on chromosome 9p21, 100 kb telomeric of the p15 and p16 genes, which are often deleted in tumor cells. The role of MTAP protein expression in the genesis of cutaneous squamous cell carcinoma (SCC) is currently not known. In a previous study we have shown the frequent occurrence of allelic imbalance/loss of heterozygosity (AI/LOH) in cutaneous SCCs using AI/LOH markers flanking the p15, p16, and MTAP genes and demonstrated reduction in p15 and p16 protein expression in comparison to normal human skin. The present study is a continuation to our previous studies, aimed at determining possible roles played by MTAP protein expression in the genesis of cutaneous SCC. The expression of MTAP protein was detected using immunohistochemical approach in 109 micro array cutaneous SCC and 20 normal human skin tissue samples. The expression of MTAP was not significantly different in the cutaneous SCC cases as compared with normal human skin. This may indicate that MTAP protein expression does not contribute to the genesis of cutaneous SCC.

Angiotensin converting enzyme from sheep mammary, lingual and other tissues.

Occurrence of angiotensin converting enzyme (ACE) in mammary gland and tongue taste epithelium was demonstrated for the first time. Six times higher ACE activity in lactating mammary gland, than non-lactating mammary gland, suggested pregnancy and lactation hormonal dependent expression of ACE in female mammals. ACE activity was highest in choroid plexus, less in spinal cord and moderate in cerebrum, medulla, cerebellum and pons. Distribution of ACE in different regions of skin, kidney and among other tissues was different. Presence of ACE in adrenal glands, pancreas, bone marrow and thyroid gland indicated functions other than blood pressure homeostasis for this enzyme.

Inhibition of cutaneous oxidative stress and two-stage skin carcinogenesis by Hemidesmus indicus (L.) in Swiss albino mice.

Chemopreventive potential of H. indicus on 7,12-dimethyl-benz[a]anthracene (DMBA)-initiated and 12-O-tetradecanoyl 13-phorbol acetate (TPA) promoted murine skin carcinogenesis has been assessed. Topical application of H. indicus resulted in significant protection against cutaneous tumorigenesis. Topical application of plant extract prior to that of TPA resulted in significant inhibition against TPA-caused induction of epidermal ornithine decarboxylase (ODC) activity and DNA synthesis. Application of H. indicus at a dose level of 1.5 and 3.0 mg/kg body weight in acetone prior to that of TPA treatment resulted in significant inhibition of oxidative stress. The level of lipid peroxidation was significantly reduced. In addition, depleted levels of glutathione and reduced activities of antioxidant enzymes were restored respectively). The results indicate that H. indicus is a potent chemopreventive agent in skin carcinogenesis.

Induction of inflammatory cell accumulation using human mast cell tryptases.

The aim of this study was to investigate the effects of human tryptases on inflammatory cell accumulation in vivo. Various concentrations of purified lung or skin tryptases were injected into the peritoneum of BALB/c mouse. At 6 hours or 16 hours following injection, cells from the peritoneal lavage were collected and stained with modified Wright's stain. Differential cell counts were performed and results were expressed as absolute numbers of each cell type per mouse peritoneum. The results showed a dose-dependent infiltration of neutrophils with a maximal increase of up to 32 fold or 43 fold in numbers at 16 hours following an injection of skin and lung tryptases, respectively. Skin tryptase was able to attract more eosinophils than lung tryptase. Significant increases in lymphocyte and macrophage numbers were also observed. In conclusion, both skin and lung tryptases are able to induce nucleated cell accumulation in the peritoneum of mice with similar specificity and potency.

Detection of Telomerase Activity in Psoriasis Lesional Skin and Correlaton with Ki-67 Expression and Suppression by Retinoic Acid

Telomerase activity is usually detected in most tumor tissues but not in normal tissues. Recently, there is increasing evidence that telomerase activity is associated with cell proliferation without malignancy, whereas there is little information about telomerase activity and its relationship with cell proliferation in chronic hyperproliferative skin diseases. Thus, we studied telomerase activity in skins from 10 patients with psoriasis and compared telomerase activity with the expression of Ki-67, a proliferation marker, using immunohistochemical staining. The effect of retinoic acid on the telomerase activity in HaCaT cells was also evaluated. Telomerase activity was detected in 7 (70%) of 10 lesional skins of psoriasis and none of the nonlesional skin. Telomerase activity in lesional skin was significantly associated with Ki-67 labelling index. Retinoic acid treatment on HaCaT cells inhibited telomerase activity, which correlated with inhibition of cell proliferation by the agent. The results of our study represent another example that shows telomerase activity correlates with cellular proliferation. Further studies on the regulation of the telomerase are needed to understand the cellular factors involved in controlling telomerase activity.

Isolation and properties of a multicatalytic proteinase complex from Xenopus laevis skin secretion

A multicatalytic proteinase complex present in the skin secretion of Xenopus laevis was purified and its enzymatic activity towards natural and synthetic peptides was investigated. We identified three activities: i) a C-terminal deamidation enzyme activity which exhibited selectivity for the Asp-Phe-NH2 and Phe-Leu-NH2 motifs of cerulein, minigastrin Leu-enkephalinamide, (des-Tyr1)Leu-enkephalinamide and diaminobenzylthiocyanate-DVDERDVRGFASFL NH2 (DABTCDR8kermit); ii) an endopeptidase activity that cleaves peptide bonds on the carboxyl side of hydrophobic amino acid residues such as Tyr-Gly of LHRH, Ile-Ala of PGLa and Leu-Ala of buccalin; iii) an enzyme activity that cleaves peptide bonds at the dibasic sites of peptides of the dynorphin family. The molecular weight determined by Sephacryl S-400 molecular sieve filtration indicated an Mr about 600 kDa. The activities characterized here exhibit and optimal pH of about 7.4. The activities of the multicatalytic complex were differentially inhibited by the classical inhibitors of proteases

Toxic effects of different concentrations of dimethoate on lysosomal enzymes of female albino rats.

The effect of three different concentrations of dimethoate on the activity of certain lysosomal enzymes, viz. beta-glucuronidase, beta-N-acetylglucosaminidase, cathepsin B and cathepsin D in serum, skin, liver, kidney and spleen and the stability of liver and kidney lysosomes was studied in female albino rats. The activity of beta-glucuronidase, beta-N-acetylglucosaminidase, cathepsin D was found to increase in serum and tissues in higher concentration (2.25 mg/100 g body weight) of dimethoate treated rats. A significant increase in the rate of release of beta-glucuronidase was found in the liver and kidney of higher concentration of dimethoate treated rats compared to controls. The results demonstrate that the activity of lysosomal enzymes increased in higher concentration of dimethoate treated rats than the lower concentration (0.56 mg/100 g body weight) of dimethoate treated rats.

Niemann-Pick disease in association with homozygous hemoglobin E: a case report.

The first case of Niemann-Pick disease Type A in a Thai infant was reported. The patient also had abnormal hemoglobin E. The diagnosis was based on the clinical features, bone marrow findings and sphingomyelinase levels in the culture of skin fibroblasts. The autosomal recessive mode of inheritance was confirmed in this case.