Specific Activity of α1Proteinase Inhibitor and α2Macroglobulin in Human Serum: Application to Insulin-Dependent Diabetes Mellitus

Abstract

The shifting balance between proteinases and proteinase inhibitors in blood, a function of their relative affinities and concentrations, has long been hypothesized to influence immune competency. The identification of proteinase-activated receptor responses in cells of the mononuclear phagocyte system suggests a potential explanation. The major serum proteinase inhibitor, alpha1proteinase inhibitor (alpha1PI, alpha1-antitrypsin), has been reported to increase in concentration during inflammation. Quantitative determination of serum alpha1PI has traditionally been performed nephelometrically; however, antigenically quantitated levels may not be representative of functional capacity. It has previously been observed that alpha1PI in serum exhibits bimodal behavior as the result of various concentrations of proteinase inhibitors, specifically alpha2macroglobulin (alpha2M) and inter-alpha-trypsin inhibitor, which compete in binding to a panel of serine proteinases. Consequently, it has not previously been possible to assign a numerical value for the specific activity of these competing proteinase inhibitors in serum. By applying known constants representing the association of proteinase inhibitors with porcine pancreatic elastase (PPE), the theoretical relationship between the functional and antigenic values for alpha1PI and alpha2M has been empirically derived allowing, for the first time, the calculation of their specific activities in serum. As predicted, the serum concentration of alpha1PI was found to be highly correlated with residual uninhibited PPE catalytic activity in healthy individuals, but not in individuals exhibiting fragmented or complexed alpha1PI. Using these techniques, both the antigenic and functional levels of alpha1PI were determined in sera from subjects with insulin-dependent diabetes mellitus (IDDM) who had been clinically diagnosed as having either periodontal disease or gingival health. Determination of quantitative levels by antigen-capture suggests that the IDDM subjects with periodontitis manifest dramatically increased levels of fragmented serum alpha1PI compared with their orally healthy counterparts or normal controls. In contrast, functional analysis of serum alpha1PI revealed no differences between the three subject populations. The elevated levels of antigenically determined serum alpha1PI reflect the inflammatory status of periodontal disease. These results support the importance of and provide methodology for determining the functionally active levels of alpha1PI allowing reexamination of changes detected during the acute phase of inflammation, replacement therapy, and longitudinal studies in relevant disease processes including malignancy and diabetes.