Hyaluronan participates in the epidermal response to disruption of the permeability barrier in vivo

Hyaluronan (hyaluronic acid, HA) is a glycosaminoglycan in the extracellular matrix of tissues that plays a role in cellular migration, proliferation and differentiation. Injury to the stratum corneum elicits an epidermal hyperproliferative response, a pathogenic feature in many cutaneous diseases including eczema and psoriasis. Because HA is abundant in the matrix between keratinocytes, we asked whether the presence of HA is required for epidermal hyperplasia to occur in response to barrier injury. Disruption of the stratum corneum, by acetone application on the skin of hairless mice, led to a marked accumulation of HA in the matrix between epidermal basal and spinous keratinocytes, and also within keratinocytes of the upper epidermis. To test whether HA may have a functional role in epidermal hyperplasia, we used Streptomyces hyaluronidase (StrepH), delivered topically, to degrade epidermal HA and blunt the accumulation of epidermal HA after acetone. StrepH signficantly reduced epidermal HA levels, and also significantly inhibited the development of epidermal hyperplasia. This reduction in epidermal thickness was not attributable to any decrease in keratinocyte proliferation, but rather to an apparent acceleration in terminal differentiation (ie, increased keratin 10 and filaggrin expression). Overall, the data show that HA is a significant participant in the epidermal response to barrier injury.     

The combination of plasmid interleukin-12 with a single DNA vaccine is more effective than Mycobacterium bovis (bacille Calmette-Guèrin) in protecting against systemic Mycobacterim avium infection

Sub‐unit vaccines utilizing purified mycobacterial proteins or DNA vaccines induce partial protection against mycobacterial infections. For example, immunization with DNA vaccines expressing the gene for the immunodominant 35 000 MW protein, common to Mycobacterium avium and Mycobacterium leprae but absent from the Mycobacterium tuberculosis complex, conferred significant protection against infection with either virulent M. avium or M. leprae in mice. However, the level of protection was equivalent to that obtained with the viable, attenuated vaccine, Mycobacterium bovis, bacille Calmette–Guèrin (BCG). The cytokine, interleukin (IL)‐12, is essential for priming naïve CD4⁺ T lymphocytes to differentiate into interferon‐γ (IFN‐γ)‐secreting T cells. We have used a novel self‐splicing vector expressing both chains of murine IL‐12 to determine if plasmid IL‐12 would increase the efficacy of a vaccine expressing the M. avium 35 000 MW protein (DNA‐Av35). Co‐immunization with p2AIL‐12 and DNA‐Av35 led to a significant increase in the number of antigen‐specific IFN‐γ secreting cells and total amount of IFN‐γ released, but a concomitant fall in the antibody response to the 35 000 MW protein. This pattern of response was associated with enhanced clearance of M. avium from the liver and spleen of coimmunized mice, and was significantly more effective than BCG or DNA‐Av35. alone. Following M. avium challenge there was significant increase in the expansion of the 35 000 MW antigen‐reactive T cells in the coimmunized mice. Therefore, plasmid‐delivered IL‐12 acts as an effective adjuvant to increase the protective efficacy of a single DNA vaccine against M. avium infection above that achieved by BCG, and this strategy may improve the efficacy of subunit vaccines against M. leprae and M. tuberculosis.     

Peanut-induced anaphylaxis in children and adolescents: Data from the European Anaphylaxis Registry

Background

Peanut allergy has a rising prevalence in high-income countries, affecting 0.5%–1.4% of children. This study aimed to better understand peanut anaphylaxis in comparison to anaphylaxis to other food triggers in European children and adolescents.

Methods

Data was sourced from the European Anaphylaxis Registry via an online questionnaire, after in-depth review of food-induced anaphylaxis cases in a tertiary paediatric allergy centre.

Results

3514 cases of food anaphylaxis were reported between July 2007 - March 2018, 56% in patients younger than 18 years. Peanut anaphylaxis was recorded in 459 children and adolescents (85% of all peanut anaphylaxis cases). Previous reactions (42% vs. 38%; p = .001), asthma comorbidity (47% vs. 35%; p < .001), relevant cofactors (29% vs. 22%; p = .004) and biphasic reactions (10% vs. 4%; p = .001) were more commonly reported in peanut anaphylaxis. Most cases were labelled as severe anaphylaxis (Ring&Messmer grade III 65% vs. 56% and grade IV 1.1% vs. 0.9%; p = .001). Self-administration of intramuscular adrenaline was low (17% vs. 15%), professional adrenaline administration was higher in non-peanut food anaphylaxis (34% vs. 26%; p = .003). Hospitalization was higher for peanut anaphylaxis (67% vs. 54%; p = .004).

Conclusions

The European Anaphylaxis Registry data confirmed peanut as one of the major causes of severe, potentially life-threatening allergic reactions in European children, with some characteristic features e.g., presence of asthma comorbidity and increased rate of biphasic reactions. Usage of intramuscular adrenaline as first-line treatment is low and needs to be improved. The Registry, designed as the largest database on anaphylaxis, allows continuous assessment of this condition.

     

Systemic oxidative and antioxidative status in Chinese patients with asthma

BACKGROUND: Patients with asthma generate an increased amount of reactive oxygen species from peripheral blood cells. Reactive oxygen species produce many of the pathophysiologic changes associated with asthma and may contribute to its pathogenesis. OBJECTIVE: We investigated changes in antioxidant enzyme activities and oxidized glutathione (glutathione disulfide; GSSG) levels in erythrocytes from a group of healthy control Chinese subjects (n=135) and patients with asthma (n=106). METHODS: Baseline pulmonary function was measured for all subjects. Antioxidant status was evaluated by measuring erythrocyte superoxide dismutase, catalase, and glutathione peroxidase activities. Oxidative stress was also measured in terms of GSSG in erythrocytes with a kinetic microassay. RESULTS: Patients with asthma had significantly increased erythrocyte superoxide dismutase and catalase activities compared with controls (61.10 +/- 1.30 U/g hemoglobin [Hb] vs 55.51 +/- 1.82 U/g Hb [P=.018] and 0.0637 +/- 0.0021 U/g Hb vs 0.0257 +/- 0.0120 U/g Hb [P <.001] for the asthma and control groups, respectively). Conversely, erythrocyte glutathione peroxidase activity decreased (44.21 +/- 1.33 mU/g Hb vs 50.07 +/- 1.39 mU/g Hb for the asthma and control groups, respectively; P=.003). Patients with asthma also had significantly higher GSSG levels in erythrocyte hemolysates compared with controls (167.40 +/- 2.93 micromol/L vs 44.98 +/- 0.44 micromol/L for the asthma and control groups, respectively; P <.001), indicating increased oxidative stress. CONCLUSIONS: Asthma is accompanied by an alteration in systemic antioxidant status due to possible oxidative stress in this disease.     

Stability of lyophilised specimens for the molecular detection of viral DNA/RNA.

BACKGROUND: The range of nucleic acid-based technologies for the molecular detection of pathogens has grown rapidly in recent years. The influx of new testing methods into the clinical laboratory, demands for evaluation and standardisation of methods, interpretation of results and evaluation of laboratory performance have highlighted the need for internal and External Quality Assessment (EQA) systems more than ever before. External Quality Assessment panels demand reproducible, stable specimens of consistent form, suitable for transportation. OBJECTIVES: To determine the stability of freeze-dried viral specimens in terms of molecular detection. STUDY DESIGN: When EQA specimens are prepared, they undergo long-term storage and testing as part of the quality control (QC) process. The frequency and nature of testing is dependent on the resources and methodologies available at the time. A range of virus preparations used for EQA was monitored over a period of months to years in a retrospective study; the available quality monitoring data for the five viruses, including storage temperature and method of detection were analysed. RESULTS: The nucleic acid (DNA or RNA) of the freeze-dried viruses included in the study was readily detectable over a long period of time. Quantitative analysis indicated that detectable concentrations of nucleic acid post-freeze drying were similarly maintained. Storage temperature was an important factor in the stability of HCV, but other viruses were unaffected by storage at different temperatures. CONCLUSIONS: In summary, the molecular detection of nucleic acid (DNA or RNA) in freeze-dried specimens of HSV1, HSV2, HBV, HCV and HIV is possible even after prolonged storage, in some cases at a range of temperatures. Freeze drying allows large-scale production of viral specimens of high quality for EQA, which are stable in varying storage and shipment conditions. Furthermore, detection of each virus was possible with a range of commonly used molecular diagnostic methods.     

Chromosome abnormalities in chronic lymphocytic leukemia revealed by TPA as a mitogen

Bone marrow and peripheral blood cultures of chronic lymphocytic leukemia patients were mitogenically stimulated with TPA (12-0-tetradecanylphorbol-13-acetate). Clonal cytogenetic abnormalities were detected in frequencies varying from 15% to 100%, in five of the six patients studied. Parallel studies with pokeweek mitogen showed a much lower level of stimulation and only two abnormal clones were detected. The chromosome abnormalities described in this study are similar to those reported in CLL by other authors, particularly with respect to trisomy 12 and deletion 11q. A significant frequency of hypodiploidy and chromosome deletion was also detected in this study, and further studies are underway to determine the significance of these findings.     

The role of antigen-presenting cells and interleukin-12 in the priming of antigen-specific CD4+ T cells by immune stimulating complexes

Immune stimulating complexes (ISCOMs) containing the saponin adjuvant Quil A are vaccine adjuvants that promote a wide range of immune responses in vivo, including delayed-type hypersensitivity (DTH) and the secretion of both T helper 1 (Th1) and Th2 cytokines. However, the antigen-presenting cell (APC) responsible for the induction of these responses has not been characterized. Here we have investigated the role of dendritic cells (DC), macrophages (Mφ) and B cells in the priming of antigen-specific CD4⁺ T cells in vitro by ISCOMs containing ovalbumin (OVA). OVA ISCOMs pulsed bone marrow (BM)-derived DC but not BM Mφ, nor naïve B cells prime resting antigen-specific CD4⁺ T cells, and this response is greatly enhanced if DC are activated with lipopolysaccharide (LPS). Of the APC found in the spleen, only DC had the capacity to prime resting antigen specific CD4⁺ T cells following exposure to OVA ISCOMs in vitro, while Mφ and B cells were ineffective. DC, but not B cells purified from the draining lymph nodes of mice immunized with OVA ISCOMs also primed resting antigen-specific CD4⁺ T cells in vitro, suggesting that DC are also critical in vivo. Using DC and T cells from interleukin (IL)-12 p40^−/− mice, we also identified a crucial role for IL-12 in the priming of optimal CD4⁺ T cell responses by OVA ISCOMs. We suggest that DC are the principal APC responsible for the priming of CD4⁺ T cells by ISCOMs in vivo and that directed targeting of these vectors to DC may enhance their efficancy as vaccine adjuvants.     

Identical allelic losses in mature teratoma and other histologic components of malignant mixed germ cell tumors of the testis

Teratomas of the testis in post-pubertal patients are histologically diverse tumors that often coexist with other types of germ cell tumors. Using laser capture microdissection and loss of heterozygosity analysis, we investigated the clonality of mature teratoma and its relationship to other components of malignant mixed germ cell tumors to gain potential insight into the histogenetic relationship of teratoma with other germ cell tumor components. All 16 patients had mature teratoma as one component of their mixed germ cell tumors. The other histological subtypes included immature teratoma, seminoma, embryonal carcinoma, yolk sac tumor, and choriocarcinoma. Laser-assisted microdissection was performed on the formalin-fixed, paraffin-embedded tissue. Polymerase chain reaction was used to amplify genomic DNA at specific loci on chromosome 1p36.2 (D1S508), 2q22-32 (D2S156), 9p21-22 (D9S162), 11p13 (D11S903), 12q22-23 (D12S1051), and 18q21 (D18S46). Fourteen of 16 (88%) cases showed allelic loss in one or more components of the mixed germ cell tumors. Fourteen of 16 mature teratomas showed allelic loss in at least one of six microsatellite polymorphic markers analyzed. The frequency of allelic loss in mature teratoma was 50% (7 of 14) with D1S508, 33% (5 of 15) with D2S156, 58% (7 of 12) with D9S162, 43% (6 of 14) with D11S903, 20% (3 of 15) with D12S1051, and 33% (5 of 15) with D18S46. Completely concordant allelic loss patterns between mature teratoma and all of the other germ cell tumor components were seen in 10 of 14 tumors in which mature teratoma showed loss of heterozygosity. Our data support the common clonal origin of mature teratoma with other components of malignant mixed germ cell tumors of the testis.     

Lymphopenia in acute nitrofurantoin pleuropulmonary reactions

Each of 5 patients with acute nitrofurantoin pleuropulmonary reactions had profound lymphopenia and 4 had eosinophilia developing early in the clinical course after the drug was withdrawn. The 2 patients tested had only one third of the normal numbers of E rosettes (T lymphocytes) in the peripheral blood during recovery. Lymphoblastic transformation tests with purified nitrofurantoin were done in 3 patients and all of them were negative; responses to phytohemagglutinin, concanavalin A, and pokeweed were decreased but still normal. The diagnosis of various nitrofurantoin hypersensitivity reactions relies on clinical data. The mechanisms of these reactions presently remain unclear.