Modulation of airway hyperresponsiveness by thiols in a murine in vivo model of allergic asthma

OBJECTIVE AND DESIGN: Since oxidative stress contributes to the pathogenesis of asthma, this study addressed the question whether supplementing the endogenous antioxidant, glutathione (GSH), would alleviate features of allergic asthma in the mouse. MATERIAL AND METHODS: Ovalbumin-sensitized mice received aerosols of the GSH-donors, glutathione-ethyl ester (GSEt) or N-acetylcysteine, before or during respiratory allergen challenges, or during methacholine challenges given one day after the last allergen challenge. Lung GSH levels were measured shortly after allergen or methacholine challenge. In addition, the effect of GSH supplements on airway hyperresponsiveness and inflammatory cell numbers in the airway lumen was assessed. RESULTS: GSEt decreased allergen-induced airway hyperresponsiveness when given in combination with methacholine. However, when given before or during allergen challenge, both GSH-donors failed to decrease the methacholine-induced airway contractility, change cell numbers in the airway lumen, or increase lung GSH levels. In addition, allergen challenges of sensitized mice did not decrease lung GSH levels. CONCLUSION: In contrast to guinea pigs and humans, allergen challenges in mice does not lead to acute oxidative stress.     

Glutathione-conjugated toluene diisocyanate causes airway inflammation in sensitised mice

Toluene diisocyanate (TDI) is a highly volatile compound that reacts readily with nucleophilic compounds, sulfhydryl groups in particular. Since the epithelial lining fluid of the airways contains high levels of the sulfhydryl, glutathione (GSH), inhalation of TDI is likely to result in the formation of GS-TDI conjugates. We therefore investigated whether GS-TDI is capable of provoking irritant and/or allergic reactions. Irritant effects of GS-TDI were studied after intratracheal administration of a range of doses of GS-TDI in saline to naive BALB/c mice. GS-TDI caused a dose-dependent increase in neutrophils in the lungs 24 h after instillation. A dose equivalent to 150 g of TDI or lower had no effect. For provocation of allergic reactions, mice were sensitised by application of 1% TDI onto the skin on days 0 and 1, and challenged intratracheally with a sub-irritant dose of GS-TDI on day 8. GS-TDI did not induce non-specific tracheal hyperreactivity to carbachol 24 and 48 h after challenge in TDI-sensitised mice. However, it increased the numbers of neutrophils in the lungs as compared with the control mice. These findings suggest that GSH conjugation does not diminish the capacity of TDI to elicit irritant-induced inflammation in the lungs of mice at doses above 150 g of TDI in the conjugate. Moreover, the capacity to induce allergic-specific inflammation was retained at concentrations of GS-TDI being devoid of irritant activity. However, the GS-TDI conjugate failed to induce non-specific tracheal hyperreactivity. This may be the consequence of the deposition of excess of GSH upon local dissociation of the conjugate.     

Factors that define the susceptibility of endothelial cells to tumor necrosis factor and lipid A.

Factors defining the previously established differences in susceptibility of endothelial cells to the tumor-necrotizing agents, tumor necrosis factor (TNF) and lipid A, were investigated. Venous and arterial endothelial cells isolated from bovine and human umbilical cords showed large differences in proliferation rate and susceptibility to TNF and lipid A as measured by [3H]thymidine incorporation. The faster proliferating cells appeared more strongly inhibited by the agents. Also, using different isolates of human venous endothelial cells a similar correlation between proliferation rate and degree of inhibition was found. Growth stimulation of human venous cells with endothelial cell growth factor, but not the growth factor combined with heparin, increased the inhibitory action of the agents. Influence of the serum source was investigated by culturing human and bovine cells in the presence of human or bovine serum. Lipid A, but not TNF, was more inhibitory to cells of both species when cultured in bovine serum as compared to human serum. Supernatant of cultured tumor cells and serum from tumor-bearing mice increased the inhibitory effects of the agents as well. Data show that the action of the tumor-necrotizing agents on endothelial cells is defined by various factors, such as origin of the cells and culture conditions. In general, factors promoting cell proliferation tended to increase the susceptibility of the cells to the agents. The reported high vulnerability of tumors, wound tissue and placenta to induction of hemorrhage by these agents may be partially due to an enhanced sensitivity of the fast proliferating endothelium in these tissues.     

Chemical-induced autoimmune reactions and Spanish toxic oil syndrome. Focus on hydantoins and related compounds

Autoimmune diseases comprise a wide spectrum of overlapping, systemic and organ-specific disorders. Although, etiology and pathogenesis of such disorders are largely unknown, endogenous host factors and exogenous agents, such as viruses, bacteria, and small molecular weight chemicals, drugs and food components, are believed to be involved. The toxicological significance of low molecular weight compounds on induction of autoimmune disorders is illustrated by the toxic oil syndrome (TOS), a chemically induced epidemic, observed in Spain since 1981. The causative chemical(s) of TOS is still elusive, but an association between ingestion of refined aniline-adulterated rapeseed oil and the syndrome is well-documented. Epidemiological, clinical and immunopathological symptoms of TOS are briefly reviewed. The striking resemblance with immunological disorders, observed in man upon medication with hydantoins and related compounds, is demonstrated. The likeliness of formation of a hydantoin-related compound in the aniline-adulterated oil is evidenced and its role as possible toxic agent in TOS is proposed. Further, the presence of hydantoins and related compounds in food is briefly reviewed and it is suggested that these chemicals may account for a portion of idiopathic autoimmune diseases observed in man. The need for development of animal models to assess this kind of immunotoxicological effects is stressed.     

Respiratory Allergy to Trimellitic Anhydride in Rats: Concentration-Response Relationships During Elicitation

The present study investigated whether airway responses of sensitized rats to trimellitic anhydride (TMA) were concentration dependent and whether these were related to irritation by TMA. Groups of BN and Wistar rats were sensitized by two dermal applications of TMA (50% w/v, followed by 25% w/v in vehicle). Controls received vehicle (acetone–olive oil 4:1, v/v). All animals were challenged 3 wk after the first sensitization by inhalation of one of a range of concentrations of TMA (0.2–61 mg/m³ for BN rats, 15–250 mg/m³ for Wistar rats). Breathing pattern, breathing frequency, and tidal volume were measured before, during, and after challenge to assess allergic and irritative airway responses. One day after challenge, nonspecific airway responsiveness to a range of concentrations of methacholine was measured. At necropsy on the same day, blood was withdrawn for measuring total serum immunoglobulin E (IgE) and organs were weighed. Larynx, trachea and lungs were examined histopathologically. In BN rats, TMA sensitization elevated total IgE levels; subsequent inhalation challenge with 2 mg/m³ of TMA and higher caused laryngeal inflammation with squamous epithelial metaplasia, and pulmonary hemorrhages. Concentration-related decreases in breathing frequency and alterations in breathing pattern, which differed from the irritation-induced pattern, were also observed at these levels. Inhalation challenge with TMA concentrations of 12 mg/m³ and higher increased lung weight. Increased nonspecific airway responsiveness was observed at the 2 next higher tested concentrations of 46 and 61 mg/m³. In unsensitized BN rats, only laryngeal squamous metaplasia was observed, albeit at higher challenge concentrations of TMA, and decreased breathing frequency, a typical breathing pattern characteristic of irritation. Identically sensitized Wistar rats showed airway inflammation and pulmonary hemorrhages upon challenge with TMA, but no functional changes, even at distinctly irritating concentrations of TMA up to 250 mg/m³. In conclusion, TMA challenge of sensitized BN rats caused challenge concentration-related allergic airway inflammation, asthmalike changes in breathing pattern, and increased nonspecific airway responsiveness. The lowest no-observed-effect level (NOEL) based on the most sensitive endpoint investigated was 0.2 mg/m³, a value that is well below the irritation concentration. The presence of a NOEL in the sensitized BN rat suggests that assessment of safe human exposure levels is feasible.     

Urinary biopterin levels in mice during graft-versus-host reactions and during exposure to 5,5-diphenylhydantoin.

Based on evidence that urinary neopterin levels are useful markers of disordered cellular immunity in man, we investigated murine urinary biopterin excretion during acute and chronic graft-versus-host (GvH)-reactions as well as after oral exposure to drugs with documented immune disregulating potential in man. Biopterin levels were determined in urine spot samples by reversed-phase high performance liquid chromatography and expressed in relation to the urinary creatinine content. Similarly increased and decreased biopterin levels were observed during acute and chronic GvH-disease in (C57BL/6J x DBA/2J)F1 (B6D2F1) mice. Increased and/or decreased levels of urinary biopterin were observed during treatment with 5,5-diphenylhydantoin (DPH), methimazole, propylthiouracil and nitrofurantoin, but no consistent pattern could be distinguished. The DPH-induced alterations were similar in B6 and B6D2F1 mice, were dose-dependent, reversible and independent of mature T-cells, as judged by the pronounced biopterin excretion of B6-nu/nu mice in comparison with their T-cell competent litter mates. The results indicate that monitoring of urinary biopterin excretion in mice does not represent a useful biochemical marker for T-cell activation.     

Regulatory T-lymphocytes in asthma.

T-helper cell type (Th)2 lymphocytes play an important role in the initiation, progression and persistence of allergic diseases, including asthma. However, little is known about immunoregulatory mechanisms that determine susceptibility to, severity of, or persistence of asthma. The concept of a disturbed Th1/Th2 balance, although having furthered the present understanding of immunoregulation in asthma, has recently been named a "procrustean paradigm", because of its failure to adequately explain many (pre)clinical observations. In recent years, the general knowledge regarding the regulation of infectious, autoimmune diseases, asthma and allergen immunotherapy by T-regulatory (Treg) cells, has rapidly increased. Many different Treg subsets have been described, including CD8+ Treg cells, natural killer (NK) cells and several different CD4+ Treg cell subsets. In this review, the authors will focus on two major and well-described CD4+ Treg cell subsets. These consist of naturally occurring CD25+ Treg cells and adaptive Treg cells that are postulated to prevent immune responses against self-antigens and adaptive immune responses, respectively. The adaptive T-regulatory cells are further subdivided into T-regulatory cells type 1 and T-helper cell type 3 that mediate suppression exclusively via the cytokines interleukin-10 and transforming growth factor-beta, respectively.