Braun M A, Gerlach D, Hartwig U F, Ozegowski J H, Romagné F, Carrel S, Köhler W, Fleischer B
First Department of Medicine, University of Mainz, Germany.
J Immunol. 1993 Mar 15;150(6):2457-66.
The pyrogenic (erythrogenic) exotoxins A and C (SPEA and SPEC) of Streptococcus pyogenes belong to the family of mitogenic toxins of which the staphylococcal enterotoxins are the prototypes. The erythrogenic toxin B (SPEB) is a proteinase precursor. All SPE have been reported to be superantigens. Here we have analyzed the human T cell response to these toxins. We used highly purified preparations of SPEA, SPEB, and SPEC from different S. pyogenes strains. These toxins were apparently homogenous in SDS-PAGE, IEF, and HPLC. In addition, recombinant SPEA and SPEC were produced in Escherichia coli. In cultures of PBMC, all three toxins expanded preferentially a fraction of T cells. Using mAb against V beta 2, -5, -6, -8, and -12, we investigated the phenotype of the stimulated cells. Natural SPEA, SPEB, and SPEC strongly stimulated V beta 8+ T cells, whereas recombinant SPEA and SPEC did not. Both natural and recombinant SPEA stimulated V beta 12+ cells and both natural and recombinant SPEC stimulated V beta 2+ cells. In accordance with these findings, a human V beta 8+ line responded to all three toxins derived from S. pyogenes but not to the recombinant proteins. An antiserum against natural SPEC neutralized specifically the V beta 2-stimulating activity of SPEC and the V beta 8-stimulating activity of all three toxins, but had no effect on the response to other superantigens. This shows that trace amounts of a potent novel V beta 8-stimulating activity not identical to SPEA and SPEC are responsible for the stimulation of V beta 8+ T cells by natural SPEA and SPEC reported previously. In a preliminary screening of S. pyogenes strains from patients, we found that this novel superantigen appears to be more widely distributed than SPEA and SPEC. Furthermore, we present evidence that also the superantigenic properties of SPEB are due to contaminations with this V beta 8 stimulator. The response to SPEB usually required 1000 times higher concentrations than to SPEA or SPEC. Antisera to SPEC but not to SPEB inhibited the response of PBMC and V beta 8+ Jurkat cells to SPEB. Furthermore, more stringent purification of SPEB yielded SPEB preparations devoid of mitogenic activity. These results indicate that the mitogenicity that is commonly attributed to SPEB is due to minute contaminations of the V beta 8 stimulator. These results raise two important caveats for the work with these highly potent T cell mitogens.(ABSTRACT TRUNCATED AT 400 WORDS)
化脓性链球菌的致热（产红细胞毒素）外毒素A和C（SPEA和SPEC）属于促细胞分裂毒素家族，其中葡萄球菌肠毒素是其原型。产红细胞毒素B（SPEB）是一种蛋白酶前体。据报道，所有的SPE都是超抗原。在此，我们分析了人类T细胞对这些毒素的反应。我们使用了来自不同化脓性链球菌菌株的高度纯化的SPEA、SPEB和SPEC制剂。这些毒素在SDS - PAGE、IEF和HPLC分析中显然是均一的。此外，重组SPEA和SPEC在大肠杆菌中产生。在PBMC培养物中，所有三种毒素都优先扩增了一部分T细胞。使用针对Vβ2、-5、-6、-8和-12的单克隆抗体，我们研究了受刺激细胞的表型。天然的SPEA、SPEB和SPEC强烈刺激Vβ8 + T细胞，而重组的SPEA和SPEC则不然。天然和重组的SPEA都刺激Vβ12 +细胞，天然和重组的SPEC都刺激Vβ2 +细胞。根据这些发现，一条人类Vβ8 +细胞系对来自化脓性链球菌的所有三种毒素有反应，但对重组蛋白无反应。一种针对天然SPEC的抗血清特异性中和了SPEC的Vβ2刺激活性以及所有三种毒素的Vβ8刺激活性，但对其他超抗原的反应没有影响。这表明，先前报道的天然SPEA和SPEC对Vβ8 + T细胞的刺激是由痕量的一种不同于SPEA和SPEC的强效新型Vβ8刺激活性引起的。在对患者来源的化脓性链球菌菌株进行的初步筛选中，我们发现这种新型超抗原的分布似乎比SPEA和SPEC更广泛。此外，我们提供证据表明，SPEB的超抗原特性也是由于被这种Vβ8刺激物污染所致。对SPEB的反应通常需要比SPEA或SPEC高1000倍的浓度。针对SPEC而非SPEB的抗血清抑制了PBMC和Vβ8 + Jurkat细胞对SPEB的反应。此外，对SPEB进行更严格的纯化得到了无促有丝分裂活性的SPEB制剂。这些结果表明，通常归因于SPEB的促有丝分裂活性是由于Vβ8刺激物的微量污染。这些结果为使用这些高效T细胞促有丝分裂原的研究提出了两个重要的注意事项。（摘要截短至400字）
