Enzymes and sensitization via skin exposure: A critical analysis.

Some proteins, including enzymes, can induce allergic sensitization of various types, including allergic sensitization of the respiratory tract. There is now an increased understanding of the role that the skin plays in the development of IgE-mediated allergy and this prompts the question whether topical exposure to enzymes used widely in consumer cleaning products could result in allergic sensitization. Here, the evidence that proteins can interact with the skin immune system and the way they do so is reviewed, together with a consideration of the experience gained over decades of the use of enzymes in laundry and cleaning products. The conclusion drawn is that although transcutaneous sensitization to proteins can occur (typically through compromised skin) resulting in IgE antibody-mediated allergy, in practice such skin contact with enzymes used in laundry and cleaning products does not appear to pose a significant risk of allergic disease. Further, the evidence summarized in this publication support the view that proteins do not pose a risk of allergic contact dermatitis.

Proteomics analysis to compare the venom composition between Naja naja and Naja kaouthia from the same geographical location of eastern India: Correlation with pathophysiology of envenomation and immunological cross-reactivity towards commercial polyantivenom.

BACKGROUND: Cobra bite is frequently reported across the Indian subcontinent and is associated with a high rate of death and morbidity. In eastern India (EI) Naja naja and Naja kaouthia are reported to be the two most abundant species of cobra. RESEARCH DESIGN AND METHODS: The venom proteome composition of N. naja (NnV) and N. kaouthia (NkV) from Burdwan districts of EI were compared by separation of venom proteins by 1D-SDS-PAGE followed by LC-MS/MS analysis of protein bands. The potency of commercial polyantivenom (PAV) was assessed by neutralization, ELISA, immuno-blot and venom-PAV immunoaffinity chromatography studies. RESULTS: Proteomic analysis identified 52 and 55 proteins for NnV and NkV, respectively, when searched against the Elapidae database. A small quantitative difference in venom composition between these two species of cobra was observed. PAVs exhibited poor cross-reactivity against low molecular mass toxins (<20 kDa) of both cobra venoms, which was substantiated by a meager neutralization of their phospholipase A2 activity. Phospholipase A2 and 3FTx, the two major classes of nonenzymatic and enzymatic proteins, respectively, were partially recognized by PAVs. CONCLUSIONS: Efforts must be made to improve immunization protocols and supplement existing antivenoms with antibodies raised against the major toxins of these venoms.

Stress-Induced Mucus Secretion and Its Composition by a Combination of Proteomics and Metabolomics of the Jellyfish Aurelia coerulea.

BACKGROUND: Jellyfish respond quickly to external stress that stimulates mucus secretion as a defense. Neither the composition of secreted mucus nor the process of secretion are well understood. METHODS: Aurelia coerulea jellyfish were stimulated by removing them from environmental seawater. Secreted mucus and tissue samples were then collected within 60 min, and analyzed by a combination of proteomics and metabolomics using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) and ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS/MS), respectively. RESULTS: Two phases of sample collection displayed a quick decrease in volume, followed by a gradual increase. A total of 2421 and 1208 proteins were identified in tissue homogenate and secreted mucus, respectively. Gene Ontology (GO) analysis showed that the mucus-enriched proteins are mainly located in extracellular or membrane-associated regions, while the tissue-enriched proteins are distributed throughout intracellular compartments. Tryptamine, among 16 different metabolites, increased with the largest-fold change value of 7.8 in mucus, which is consistent with its involvement in the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway 'tryptophan metabolism'. We identified 11 metalloproteinases, four serpins, three superoxide dismutases and three complements, and their presence was speculated to be related to self-protective defense. CONCLUSIONS: Our results provide a composition profile of proteins and metabolites in stress-induced mucus and tissue homogenate of A. coerulea. This provides insight for the ongoing endeavors to discover novel bioactive compounds. The large increase of tryptamine in mucus may indicate a strong stress response when jellyfish were taken out of seawater and the active self-protective components such as enzymes, serpins and complements potentially play a key role in innate immunity of jellyfish.

Changes in immunoreactivity of allergen-reduced peanuts due to post-enzyme treatment roasting.

The objective of this study was to evaluate the effect of enzyme treatment and post-enzyme treatment roasting on allergenicity of raw peanut kernels. Raw peanuts were treated by single- and two-enzyme treatments, respectively. Enzyme-treated raw peanuts were dry roasted. Reductions of four major allergens (Ara h 1, 2, 3, and 6) and in vitro allergenicity of peanuts were evaluated. Quantitative measurements show that enzyme treatment of raw peanuts reduced Ara h 1, 2, 3 and 6 in raw peanuts by 99-100%, 95-99%, 35-46% and 85-88%, respectively. Roasting of enzyme-treated peanuts significantly reduced the total soluble protein (P < 0.05), Ara h 3 and 6 (P < 0.0001), slightly increased Ara h 1 in the extracts (P < 0.05), but did not significantly affect Ara h 2. Immunoblot shows that the IgE-bindings of both soluble and insoluble proteins of enzyme-treated peanuts were slightly enhanced by roasting but still tremendously lower than that of untreated peanuts.

An overview of the immune modulating effects of enzymatic toxins from snake venoms.

Snake venoms are complex mixtures of organic and inorganic compounds, including proteins belonging to the protease (serine and metalloproteinases), oxidase (L-amino acid oxidases), and phospholipase (especially phospholipases A2) enzyme classes. These toxins account for the serious deleterious effects of snake envenomations, such as tissue necrosis, neurotoxicity, and hemorrhage. In addition to their toxic effects, snake venom toxins have served as important tools for investigating the mechanisms underlying envenomation and discovering new pharmacologically active compounds with immunotherapeutic potential. In this sense, the present review discusses the new findings and therapeutic perspectives in the immune modulating potential of enzymatic toxins from snake venoms belonging to the classes metalloproteinase, serine protease, L-amino acid oxidase, and phospholipase A2.

Profiling of proteins secreted in the bovine oviduct reveals diverse functions of this luminal microenvironment.

The oviductal microenvironment is a site for key events that involve gamete maturation, fertilization and early embryo development. Secretions into the oviductal lumen by either the lining epithelium or by transudation of plasma constituents are known to contain elements conducive for reproductive success. Although previous studies have identified some of these factors involved in reproduction, knowledge of secreted proteins in the oviductal fluid remains rudimentary with limited definition of function even in extensively studied species like cattle. In this study, we used a shotgun proteomics approach followed by bioinformatics sequence prediction to identify secreted proteins present in the bovine oviductal fluid (ex vivo) and secretions from the bovine oviductal epithelial cells (in vitro). From a total of 2087 proteins identified, 266 proteins could be classified as secreted, 109 (41%) of which were common for both in vivo and in vitro conditions. Pathway analysis indicated different classes of proteins that included growth factors, metabolic regulators, immune modulators, enzymes, and extracellular matrix components. Functional analysis revealed mechanisms in the oviductal lumen linked to immune homeostasis, gamete maturation, fertilization and early embryo development. These results point to several novel components that work together with known elements mediating functional homeostasis, and highlight the diversity of machinery associated with oviductal physiology and early events in cattle fertility.

Induction of defense-related enzymes in patchouli inoculated with virulent Ralstonia solanacearum

Background: Defense-related anti-oxidative response is a vital defense mechanism of plants against pathogen invasion. Ralstonia solanacearum is an important phytopathogen. Bacterial wilt caused by R. solanacearum is the most destructive disease and causes severe losses in patchouli, an important aromatic and medicinal plant in Southeast Asia. The present study evaluated the defense response of patchouli inoculated with virulent R. solanacearum. Results: Results showed that the basic enzymatic activities differed not only between the leaves and stems but also between the upper and lower parts of the same organ of patchouli. POD, SOD, PPO, and PAL enzymatic activities were significantly elevated in leaves and stems from patchouli inoculated with R. solanacearum compared to those in control. The variation magnitude and rate of POD, PPO, and PAL activities were more obvious than those of SOD in patchouli inoculated with R. solanacearum. PAGE isoenzymatic analysis showed that there were one new POD band and two new SOD bands elicited, and at least two isoformic POD bands and two SOD bands were observably intensified compared to the corresponding control. Conclusion: Our results suggest that not only defense-related enzymatic activities were elevated but also the new isoenzymatic isoforms were induced in patchouli inoculated with R. solanacearum.

Autoinflammatory Diseases in Pediatric Dermatology-Part 1: Urticaria-like Syndromes, Pustular Syndromes, and Mucocutaneous Ulceration Syndromes. / Enfermedades autoinflamatorias en dermatología pediátrica. Parte 1: síndromes urticariformes, síndromes pustulosos y síndromes con ulceraciones cutáneo-mucosas.

Monogenic autoinflammatory diseases are a heterogeneous emergent group of conditions that are currently under intensive study. We review the etiopathogenesis of these syndromes and their principal manifestations. Our aim is to propose a classification system based on the clinicopathologic features of typical skin lesions for routine clinical use in dermatology. Our focus is on diagnosis in pediatric practice given that this is the period when the signs and symptoms of these syndromes first appear. In Part 1 we discuss the course of urticaria-like syndromes, which include cryopyrin-associated periodic conditions and hereditary periodic fever syndromes. Pustular syndromes are also covered in this part. Finally, we review the range of therapies available as well as the genetic mutations associated with these autoinflammatory diseases.

Sensitising effects of genetically modified enzymes used in flavour, fragrance, detergence and pharmaceutical production: cross-sectional study.

OBJECTIVES: The use of genetically engineered enzymes in the synthesis of flavourings, fragrances and other applications has increased tremendously. There is, however, a paucity of data on sensitisation and/or allergy to the finished products. We aimed to review the use of genetically modified enzymes and the enormous challenges in human biomonitoring studies with suitable assays of specific IgE to a variety of modified enzyme proteins in occupational settings and measure specific IgE to modified enzymes in exposed workers. METHODS: Specific IgE antibodies against workplace-specific individual enzymes were measured by the specific fluorescence enzyme-labelled immunoassay in 813 exposed workers seen in cross-sectional surveys. RESULTS: Twenty-three per cent of all exposed workers showed type I sensitisation with IgE antibodies directed against respective workplace-specific enzymes. The highest sensitisation frequencies observed were for workers exposed enzymes derived from α-amylase (44%), followed by stainzyme (41%), pancreatinin (35%), savinase (31%), papain (31%), ovozyme (28%), phytase (16%), trypsin (15%) and lipase (4%). The highest individual antibody levels (up to 110 kU/L) were detected in workers exposed to phytase, xylanase and glucanase. In a subgroup comprising 134 workers, detailed clinical diagnostics confirmed work-related symptoms. There was a strong correlation (r=0.75, p<0.0001) between the symptoms and antibody levels. Workers with work-related respiratory symptoms showed a higher prevalence for the presence of specific IgE antibodies against workplace-specific enzymes than asymptomatic exposed workers (likelihood ratio 2.32, sensitivity 0.92, specificity 0.6). CONCLUSIONS: Our data confirm the previous findings showing that genetically engineered enzymes are potent allergens eliciting immediate-type sensitisation. Owing to lack of commercial diagnostic tests, few of those exposed receive regular surveillance including biomonitoring with relevant specific IgE assays.

¿Se asocia la obesidad en la artritis psoriásica a una menor respuesta terapéutica y más efectos adversos con el tratamiento de fondo? / Is obesity in psoriatic arthritis associated with a poorer therapeutic response and more adverse effects of treatment with an anchor drug?

Objetivos. Estudiar si en la artritis psoriásica (APs) hay asociación entre la obesidad, el control de la actividad inflamatoria y el aumento de efectos adversos con los fármacos modificadores de la enfermedad (FAME). Métodos. Revisión sistemática de la literatura utilizando las bases de datos Medline y Embase según las guías del consenso MOOSE. Se incluyeron estudios en pacientes con APs, en los que la obesidad fuera factor predictor de efectos adversos y el desenlace fuera toxicidad, incluido fallo de eficacia. La calidad se evaluó mediante una escala de riesgo de sesgos ad hoc. Se realizó un análisis cualitativo por tipo de estudio y población estudiada, calidad y resultados específicos. Resultados. Se encontraron 1.043 artículos, la mayoría se descartaron por título y abstract. Se estudiaron en detalle 10, excluyéndose finalmente 3. La mayoría concluye con resultados estadísticamente significativos que la obesidad en pacientes con APs e inhibidores del TNF-α (iTNF-α) se asocia a una probabilidad menor de alcanzar y mantener la mínima actividad inflamatoria, con mayor tasa de interrupción del tratamiento y menor tasa de respuesta cutánea. En relación con los FAME sintéticos convencionales, se observó en obesos una tendencia a un aumento moderado de las transaminasas con metotrexato (MTX). Conclusiones. La obesidad es un factor predictivo negativo de la respuesta clínica en pacientes con APs e iTNF-α. Exceptuando la hepatotoxicidad por el MTX, no se encontraron otros efectos adversos ni por otros fármacos en relación con la obesidad (AU)

Objectives. To assess the association between obesity, control of inflammatory activity and increased adverse effects in psoriatic arthritis (PsA) with disease-modifying anti-inflammatory drugs (DMARD). Methods. A systematic literature review was performed using MEDLINE and EMBASE databases following the guidelines of the Meta-analysis Of Observational Studies in Epidemiology (MOOSE) consensus statement. Studies were selected if they included patients with PsA, obesity was studied as a predictive factor, and the outcome was adverse effects, including efficacy failure. Quality was assessed using an ad hoc risk of bias tool. A qualitative analysis was carried out by type of study and study population, quality and specific results. Results. We found 1043 articles, discarding most of them on the basis of title and abstract. Ten articles were studied in detail and finally excluded three. The majority concluded, with statistically significant results, that in patients with PsA and treated with TNFα inhibitors (TNFαi), obesity is associated with poorer chances of achieving and maintaining a minimal disease activity, higher treatment discontinuation rates, and lower skin response. Regarding conventional synthetic DMARD, a trend toward a moderate increase in transaminases with methotrexate (MTX) was observed in obese patients with PsA. Conclusions. Obesity is a negative predictor of clinical response in patients with PsA being treated with TNFαi. Except MTX hepatotoxicity, no other adverse effects, either with TNFαi or other drugs, were found in relation to obesity in PsA (AU)

The impact of the immune system on the safety and efficiency of enzyme replacement therapy in lysosomal storage disorders.

In the light of clinical experience in infantile onset Pompe patients, the immunological impact on the tolerability and long-term efficacy of enzyme replacement therapy (ERT) for lysosomal storage disorders has come under renewed scrutiny. This article details the currently proposed immunological mechanisms involved in the development of anti-drug antibodies and the current therapies used in their treatment. Given the current understanding of the adaptive immune response, it focuses particularly on T cell dependent mechanisms and the paradigm of using lymphocytic negative selection as a predictor of antibody formation. This concept originally postulated in the 1970s, stipulated that the genotypically determined lack of production or production of a variant protein determines an individual's lymphocytic repertoire. This in turn is the key factor in determining the potential severity of an individual's immunological response to ERT. It also highlights the need for immunological assay standardization particularly those looking at describing the degree of functional impact, robust biochemical or clinical endpoints and detailed patient subgroup identification if the true evaluations of impact are to be realised.

The role of antibodies in enzyme treatments and therapeutic strategies.

Substitution of the defective lysosomal enzyme in lysosomal storage disorders (LSDs) often elicits antibody formation towards the infused protein. Aside from Gaucher disease, antibodies often lead to infusion associated reactions and a reduced biochemical response. In Pompe disease, antibody titer is predictive of clinical outcome, but this is less apparent in other LSDs and warrants further study. Few laboratories are capable of enzyme-antibody determination: often physicians need to rely on the enzyme manufacturer for analysis. Currently, laboratories employ different antibody assays which hamper comparisons between cohorts or treatment regimens. Assay standardisation, including measurement of antibody-related enzyme inhibition, is therefore urgently needed. Successful immunomodulation has been reported in Pompe and in Gaucher disease, with variable success. Immunomodulation regimens that contain temporary depletion of B-cells (anti-CD20) are most used. Bone marrow transplantation in MPS-I results in disappearance of antibodies. No other clinical studies have been conducted in humans with immunomodulation in other LSDs.

Self-reactive T cells: suppressing the suppressors.

The immune system is a tightly regulated and complex system. An important part of this immune regulation is the assurance of tolerance toward self-antigens to maintain immune homeostasis. However, in recent years, antigen-specific cellular immune responses toward several normal self-proteins expressed in regulatory immune cells have been reported, especially in patients with cancer. The seemingly lack of tolerance toward such proteins is interesting, as it suggests a regulatory function of self-reactive T (srT) cells, which may be important for the fine tuning of the immune system. In particular, surprising has been the description of cytotoxic srT cells that are able to eliminate normal regulatory immune cells. Such srT cells may be important as effector cells that suppress regulatory suppressor cells. The current knowledge of the nature and function of srT cells is still limited. Still, the therapeutic targeting of srT cells offers a novel approach to harness immune-regulatory networks in cancer.

Proteomic approach for identification of immunogenic proteins of Mycoplasma mycoides subsp. capri.

In this study, an immunoproteomic approach was used to identify immunodominant proteins from Mycoplasma mycoides subsp. capri isolates. Membrane proteins, extracted through TX-114 phase partitioning, were separated using mono- and two-dimensional electrophoresis and detected by Western blotting with pooled sera from naturally infected goats. A total of 27 immunoreactive spots, corresponding to 13 different proteins, were identified using nanoLC-ESI-MSMS. Function annotation revealed that most of these proteins were metabolic enzymes involved in carbohydrate and energy metabolism. The immunogenic proteins identified in this study: pyruvate dehydrogenase, dihydrolipoamide acetyltransferase, dihydrolipoyl dehydrogenase, phosphate acetyltransferase, phosphopyruvate hydratase, adenine phopshoribosyltransferase, transketolase, translation elongation factor G, translation elongation factor Ts, FMN-dependent NADH-azoreductase, peptide methionine sulfoxide reductase, inorganic diphosphatase and trigger factor may be used as biomarkers for the serological diagnosis of contagious agalactia caused by M. mycoides subsp. capri.

The toxicology and immunology of detergent enzymes.

Detergent enzymes have a very good safety profile, with almost no capacity to generate adverse acute or chronic responses in humans. The exceptions are the limited ability of some proteases to produce irritating effects at high concentrations, and the intrinsic potential of these bacterial and fungal proteins to act as respiratory sensitizers, demonstrated in humans during the early phase of the industrial use of enzymes during the 1960s and 1970s. How enzymes generate these responses are beginning to become a little clearer, with a developing appreciation of the cell surface mechanism(s) by which the enzymatic activity promotes the T-helper (T(H))-2 cell responses, leading to the generation of IgE. It is a reasonable assumption that the majority of enzyme proteins possess this intrinsic hazard. However, toxicological methods for characterizing further the respiratory sensitization hazard of individual enzymes remains a problematic area, with the consequence that the information feeding into risk assessment/management, although sufficient, is limited. Most of this information was in the past generated in animal models and in vitro immunoassays that assess immunological cross-reactivity. Ultimately, by understanding more fully the mechanisms which drive the IgE response to enzymes, it will be possible to develop better methods for hazard characterization and consequently for risk assessment and management.

Relevance of sensitization to occupational allergy and asthma in the detergent industry.

There exists considerable historic experience of the relationship between exposure and both the induction of sensitization and the elicitation of respiratory symptoms from industrial enzymes of bacterial and fungal origin used in a wide variety of detergent products. The detergent industry in particular has substantial experience of how the control of exposure leads to limitation of sensitization with low risk of symptoms. However, the experience also shows that there are substantial gaps in knowledge, even when the potential occupational allergy problem is firmly under control, and also that the relationship between exposure and sensitization can be hard to establish. The latter aspect includes a poor appreciation of how peak exposures and low levels of exposure over time contribute to sensitization. Furthermore, while a minority of workers develop specific IgE, essentially none appear to have symptoms, a situation which appears to contradict the allergy dogma that, once sensitized, an individual will react to much lower levels of exposure. For enzymes, the expression of symptoms occurs at similar or higher levels than those that cause induction. In spite of some knowledge gaps, medical surveillance programs and constant air monitoring provide the tools for successful management of enzymes in the occupational setting. Ultimately, the knowledge gained from the occupational setting facilitates the completion of safety assessments for consumer exposure to detergent enzymes. Such assessments have been proven to be correct by the decades of safe use both occupationally and in consumer products.

Neutralization of Pseudomonas aeruginosa enzymatic activity by antibodies elicited with proteins of Larrea divaricata Cav.

Larrea divaricata Cav. (Jarilla) is a bush widely used in folk therapy for the treatment of several pathologies. Partially purified proteins of crude extract (JPCE) cross-react with proteins of Gram-negative bacteria, including Pseudomonas aeruginosa, which is an opportunistic pathogen that causes several intrahospitalary infections. This bacterium produces many proteins with enzymatic activity, including hemolysins and proteases that play a major role in acute infection caused by this bacterium. The aim of our work was to investigate if antibodies against with L. divaricata neutralize the hemolytic and proteolytic activity of P. aeruginosa. The hemolytic activity of soluble cellular proteins was inhibited 100% and extracellular proteins (EP) showed an inhibition between 44 and 95% when both bacterial fractions were treated with anti-JPCE serum. Also, in EP the neutralization was directed towards the active site of the hemolysin. When protease activity of extracellular products was tested, bands of 217, 155, 121, 47 and 27 kDa were observed in native zymograms. Neutralization between 55 and 70% of the bands of 217, 155 and 121 kDa was observed when EP were treated with anti-JPCE serum. In conclusion, our data clearly demonstrate that antibodies elicited with L. divaricata' proteins are able to neutralize the hemolytic and proteolytic activity of P. aeruginosa cellular and extracellular proteins. Our study constitutes the first report that associates the immunogenicity of plant proteins and bacterial proteins with enzymatic activity. These findings could be relevant in the development of alternatives therapies for patients suffering intrahospitalary opportunistic infections with P. aeruginosa.

Influence of diet enriched with green tea on innate humoral and cellular immune response of kelp grouper (Epinephelus bruneus) to Vibrio carchariae infection.

Effect of diet enriched with green tea at 0, 0.01, 0.1 or 1.0% levels on immune responses such as non-specific humoral (lysozyme, antiprotease and complement) and cellular (myeloperoxidase content, production of reactive oxygen, and nitrogen species) and disease resistance on week 1, 2 or 4 in kelp grouper Epinephelus bruneus challenged with Vibrio carchariae (2.47 × 10(8) CFU ml(-1)) was quantified. At all doses green tea supplementation significantly enhanced the serum lysozyme activity from weeks 1 to 4. On the other hand, after week 2 the serum hemolytic complement activity, leucocyte myeloperoxidase content and reactive nitrogen species protection significantly increased in groups fed with 0.01 and 0.1% green tea supplementation diets. The serum antiprotease activity significantly increased in group fed with at 1.0% green tea from week 1 to 4. However, all diets except at 0.01% level resulted in a significant decrease in reactive oxygen species protection during the experimental period. Challenged groups fed with green tea enriched diet at 0.01 and 0.1% level had a higher relative percent survival than with 1.0% diet on week 1, 2 or 4. The results suggest that dietary administration of green tea supplementation at a concentration of 0.01 and 0.1% level positively enhances the non-specific humoral and cellular immune responses and disease resistance of kelp grouper E. bruneus to V. carchariae.