欢迎来深圳市科润达生物工程有限公司!我们将为您提供周到的服务!销售热线:86-0755-26814430 15711973608
联系我们Contact us
公司名称:深圳市科润达生物工程有限公司
联系人:科润达客服
地址:深圳市坪山区坑梓中兴路14号中城生命科学园A栋3层
首页 > 技术文章 > 过敏性湿疹病人体内金黄色葡萄球菌内毒素对组胺和白三烯释放的研究
过敏性湿疹病人体内金黄色葡萄球菌内毒素对组胺和白三烯释放的研究

过敏性湿疹病人体内金黄色葡萄球菌内毒素对组胺和白三烯释放的研究(国外学术刊物发表)

 

Staphylococcus aureus Enterotoxins

Induce Histamine and Leukotriene Release

in Patients with Atopic Eczema

K. Neuber, J. Wehner, K. Hakansson, and J. Ring

 

 

 

 

Abstract

Peripheral blood basophils from patients with atopic eczema stimulated with the

staphylococcal enterotoxins (SE) A, B, D, E and toxic shock syndrome toxin 1 (TSST-1

secreted significantly higher amounts of histamine and leukotriene C4 (LTC4 ) than

healthy controls. Furthermore, priming experiments with IL-3, IL-8 and GM -CSF followed

by stimulation with enterotoxins showed additional histamine and LTC4 release

in the group of atopic eczema patients. Histamine and leukotriene generation from

atopic basophils stimulated with staphylococcal enterotoxins may indicate a role of

these toxins as possible allergens in at least a subgroup of patients with atopic eczema.

 

Introduction

Normal as well as diseased skin of patients with atopic eczema (AE) is severely colonized

with Staphylococcus aureus [1]. Recently, it has been shown that the majority of

S. aureus strains isolated from the skin of patients with AE produce staphylococcal

enterotoxins (SEA, B, C, D, E) or toxic shock syndrome toxin-1 (TSST-1). About 50 %

of the patients have specific IgE directed to staphylococcal toxins [2]. The activation of

T-cells by staphylococcal superantigens results in the release of a TH2-like cytokine

pattern (IL-4, IL-5) in vitro [3] which, via induction of several effector cells, may be the

cause of increased IgE-synthesis and eosinophilia. In this study the influence of

staphylococcal enterotoxins (SEs) on histamine and leukotriene release of basophils was

studied.

 

Material and Methods

Patients and Controls. Basophils were obtained from healthy volunteers (n = 9) and

from patients with AE (n = 17). The AE diagnosis was performed according to the

criteria of Hanifin and Rajka [4]. All patients had a skin colonization with S. aureus as

was determined by skin smears. The donors did not receive systemic steroid or

antihistamine treatment.

 

New Trends in Allergy IV

(ed. by J. Ring, H. Behrendt, D. Vieluf)

© Springer-Verlag Berlin Heidelberg 1997

 

Blood Collection and Cell Preparation. Peripheral blood leukocytes (PBL) were obtained

by dextrane-sedimentation for 60-90 min. After three washing steps, the pellet

was resuspended in 5 ml HEPES-ACM buffer (pH 7.4).

Stimulation Conditions. As stimuli served S. aureus enterotoxins (SEA, SEB, SED, SEE

and TSST-1) obtained from Toxin Technology (Sarasota, USA) diluted in carbonate

buffer (pH 8.0). For priming experiments interleukins-3, 8 and granulocyte-macrophage

colony-stimulating factor (GM -CSF), all obtained from Biermann DPC (Bad Nauheim,

Germany), were used. As controls served spontaneous as well as anti-IgE (10-2 M

induced histamine and leukotriene release. Total leukocyte histamine content was

determined by sonication of cells for 10 s. Aliquots (500 μl) of leukocyte suspension

(1.0-1.5 x107/ml) were stimulated with SEs (100 ng/500 μl) for 30 min at 37 °C in

duplicates. For priming experiments leukocytes were prestimulated (10 min) with

interleukins in the following concentrations: IL-3 (500 ng/500 μl), IL-8 (40 ng/500 μl)

and GM-CSF (20 ng/500 μl). Afterwards, toxins were added for 30 min at 37 °C. After

incubation period reactions were stopped by adding 500 μl 0.01 N TFA (C2HF3O2).

Supernatants were stored at –20 °C.

Histamine and LTC4 ELISA. Histamine and LTC4 contents of supernatants were determined

by ELISA obtained from IBL (Hamburg, Germany) according to the prescription

of the manufacturer. The release of histamine, either spontaneously or as

induced by stimuli, was recorded as a percentage of the total net leukocyte histamine

release.

Analysis of Data. All calculations were performed using the SPSS Statistics package

(SPSS, Vers. 5.02). Results were analysed using the nonparametric Wilcoxon signedranks

test for paired data or the Wilcoxon non-paired rank-sum test for nonpaired data.

Only p-values below 0.05 were accepted as significant.

 

Results

Enterotoxin Induced Histamine and LTC4 Release. Basophils from healthy subjects did

not release more than 2 % histamine to any staphylococcal toxin. In contrast, 14 AE

patients (82 %) showed relevant histamine release upon stimulation with one or more

staphylococcal enterotoxins (Fig. 1).

Leukocytes from patients with AE generated significantly higher amounts of LTC4

(Fig. 2) after stimulation with SEA (p<0.02), SEB (p<0.02), TSST-1 p<0.02) and SEE

p<0.04).

Histamine and LTC4 Release After Priming with IL-3, IL-8 and GM-CSF. Priming of

atopic basophils with IL-3, IL-8 or GM -CSF induced a significantly enhanced histamine

release compared to stimulation with enterotoxins alone (Table 1). In normal donors no

histamine release over 2 % was observed after cytokine priming and incubation with

enterotoxins. In healthy volunteers priming with interleukins (3, 8) and GM -CSF did not

enhance enterotoxin induced LTC4 generation. However, in patients with AE priming

of basophils led to an increase of LTC4 secretion by 875 % for IL-3, 187 % for IL-8 and

412 % for GM -CSF compared to cells stimulated without interleukins (Table 1).



 

Discussion

These data clearly show that SE stimulated basophils from patients with AE secrete

significantly higher amounts of histamine and LTC4 than healthy donors. Furthermore,

priming of basophils with IL-3, IL-8 and GM-CSF increased mediator release in AE.

Recently, an increasing number of data support the suggestion that parts of the

cutaneous microflora (e. g.) Staphylococcus aureus, Pityrosporum ovale) acting as permanent

stimuli for allergic skin reactions can be an important trigger for the exacerbation

of AE [3,5]. SEs have been demonstrated to be stimuli of TH2 cells in AE

inducing IL-4 and IL-5 secretion and IgE synthesis [3].

Histamine and LTC4 are very important mediators of inflammation in allergic

diseases [6]. Therefore, induction of mediator release by SEs in AE supports the

assumption that these toxins may trigger allergic inflammation. Since AE is associated

with superficial S. aureus skin colonization, secreted enterotoxins would be expected to

penetrate the skin. Binding of SEs to cell surface bound specific IgE antibodies can

mediate histamine release from basophils and mast cells. This may lead to local release

of mediators and cytokines resulting in late-phase reactions and itch.

In control experiments with buffer not containing Ca2+ SEs as well as cytokines

failed to induce histamine secretion even in patients with AE (data not shown). These

results are similar IgE mediated histamine and LTC4 formation.

One of the most important modulating effects of cytokines on inflammatory cells is

their capacity to prepare the cells for more efficient mediator release, a phenomenon

called “priming”. Blood basophils markedly increase their capacity to release preformed

histamine or to form de novo LTC4 following short incubation in vitro with IL-3, IL-8

and GM-CSF [7]. The fact that SE induced mediator release in AE is significantly

increased by cytokines supports the assumption that hypersensitivity reaction to

these toxins are pathophysiologically relevant in at least a subgroup of patients with AE.

Acknowledgements. The authors wish to thank Ingrid Köhler and Johanna Grosch for

excellent technical assistance.

 

References

1. Ring J, Abeck D, Neuber K. Atopic eczema: role of microorganisms on the skin surface. Allergy

1992; 47:265-269

2. Leung DYM, Harbeck R, Bina P, Reiser RF, Yang E, Yang DA, Norris DA, Hanifin JK, Sampson

HA. Presence of IgE antibodies to staphylococcal exotoxins on the skin of patients with atopic dermatitis.

J Clin Invest 1993; 92:1374-1380

3. Neuber K, Steinrücke K, Ring J. Staphylococcal enterotoxin B affects in vitro IgE synthesis, interferon-

g, interleukin-4 and interleukin-5 production in atopic eczema. Int Arch Allergy Immunol

1995; 107:179-182

4. Hanifin J, Rajka G. Diagnostic features of atopic dermatitis. Acta Derm Venereol (Stockh) 1980;

92:44-47

5. Kröger S, Neuber K, Gruseck E, Ring J, Abeck D. Pityrosporum ovale extracts increase

interleukin-4, interleukin-10 and IgE synthesis in patients with atopic eczema. Acta Derm Venereol

(Stockh) 1995; 75:357-360

6. Ruzicka T, Ring J. Role of inflammatory mediators in atopic eczema. In: Handbook of atopic

eczema. Eds.: Ruzicka T, Ring J, Przybilla B. Berlin: Springer, 1991; p 245-255

7. de Weck AL, Dahinden CA, Bischoff S. Cytokines in the regulation of allergic inflammation. In:

Advances in allergology and clinical immunology: the proceedings of the XVth congress of the

European Academy of Allergology and Clinical Immunology. Eds.: Godard P, Bousquet J, Michel

FB. Park Ridge: The Parthenon Publishing Group. 1992; p 67-74

下一篇:组胺ELISA Kit
地址:深圳市坪山区坑梓中兴路14号中城生命科学园A栋3层
联系人:科润达客服

深圳市科润达生物工程有限公司 版权所有  主营:TNF-α检测试剂盒|HMGB1多克隆抗体|糖代谢相关检测试剂|美国Invitrogen细胞因子

ICP备案号:粤ICP备18097826号 管理登陆 技术支持:化工仪器网   总流量:277245  网站地图

在线客服
在线客服
扫码关注我们
用心服务 成就你我