Janusz M J, Doherty N S
Marion Merrell Dow Research Institute, Cincinnati, OH 45054.
J Immunol. 1991 Jun 1;146(11):3922-8.
The granule proteases of human neutrophils are thought to be responsible for the connective tissue destruction associated with certain inflammatory diseases. Using a model system for the degradation of a macromolecular connective tissue substrate, purified neutrophil elastase and cathepsin G were both individually able to degrade cartilage matrix proteoglycan and this degradation was blocked by the appropriate specific inhibitors. Neutrophil granule lysate also produced cartilage matrix degradation but little inhibition of degradation occurred when either elastase or cathepsin G inhibitor was used alone. However, a combination of elastase and cathepsin G inhibitors each at 100 microM or each at 10 microM blocked cartilage matrix degradation by 89% +/- 1 and 65% +/- 9 (mean +/- SEM, n = 3), respectively. The magnitude of the cartilage degradation mediated by neutrophil lysate, and its sensitivity to specific inhibitors, was reproduced using purified elastase and cathepsin G at the concentrations at which they are present in neutrophil lysate. Human neutrophils stimulated with opsonized zymosan degraded cartilage matrix in a dose-dependent manner in the presence of serum antiproteases. Supernatants from stimulated neutrophils cultured in the presence of serum did not degrade cartilage matrix, indicating that neutrophil mediated degradation in the presence of serum was confined to the protected subjacent region between the inflammatory cell and the substratum. A combination of elastase and cathepsin G inhibitors each at 500 microM or each at 100 microM blocked subjacent cartilage matrix degradation by stimulated human neutrophils by 91% +/- 3 and 54% +/- 8 (mean +/- SEM, n = 5), respectively, whereas either the elastase or cathepsin G inhibitor alone was much less effective. These studies demonstrate that neutrophil-mediated cartilage matrix degradation is produced primarily by elastase and cathepsin G. Furthermore, these results support the hypothesis that inflammatory neutrophils form zones of close contact with substratum that exclude serum antiproteases and that this subjacent degradation of cartilage matrix by stimulated neutrophils can be blocked by a combination of synthetic elastase and cathepsin G inhibitors.
人们认为人类中性粒细胞的颗粒蛋白酶与某些炎症性疾病相关的结缔组织破坏有关。利用一种用于降解大分子结缔组织底物的模型系统，纯化的中性粒细胞弹性蛋白酶和组织蛋白酶G各自都能够降解软骨基质蛋白聚糖，并且这种降解被相应的特异性抑制剂所阻断。中性粒细胞颗粒裂解物也能导致软骨基质降解，但单独使用弹性蛋白酶或组织蛋白酶G抑制剂时，降解的抑制作用很小。然而，弹性蛋白酶和组织蛋白酶G抑制剂各以100微摩尔或各以10微摩尔的组合分别使软骨基质降解减少89%±1和65%±9（平均值±标准误，n = 3）。使用纯化的弹性蛋白酶和组织蛋白酶G，以它们在中性粒细胞裂解物中的浓度进行实验，重现了由中性粒细胞裂解物介导的软骨降解程度及其对特异性抑制剂的敏感性。用调理酵母聚糖刺激的人类中性粒细胞在血清抗蛋白酶存在的情况下以剂量依赖方式降解软骨基质。在血清存在下培养的受刺激中性粒细胞的上清液不能降解软骨基质，这表明在血清存在下中性粒细胞介导的降解局限于炎症细胞与基质之间受保护的相邻区域。弹性蛋白酶和组织蛋白酶G抑制剂各以500微摩尔或各以100微摩尔的组合分别使受刺激的人类中性粒细胞对相邻软骨基质的降解减少91%±3和54%±8（平均值±标准误，n = 5），而单独使用弹性蛋白酶或组织蛋白酶G抑制剂的效果则要差得多。这些研究表明，中性粒细胞介导的软骨基质降解主要由弹性蛋白酶和组织蛋白酶G产生。此外，这些结果支持这样一种假说，即炎症性中性粒细胞与基质形成紧密接触区域，排除血清抗蛋白酶，并且受刺激的中性粒细胞对软骨基质的这种相邻降解可被合成的弹性蛋白酶和组织蛋白酶G抑制剂的组合所阻断。
