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, α₁proteinase inhibitor (α₁PI, α₁-antitrypsin), has been reported to increase in concentration during inflammation. Quantitative determination of serum α₁PI has traditionally been performed nephelometrically; however, antigenically quantitated levels may not be representative of functional capacity. It has previously been observed that α₁PI in serum exhibits bimodal behavior as the result of various concentrations of proteinase inhibitors, specifically α₂macroglobulin (α₂M) and inter-α-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 α₁PI and α₂M has been empirically derived allowing, for the first time, the calculation of their specific activities in serum. As predicted, the serum concentration of α₁PI was found to be highly correlated with residual uninhibited PPE catalytic activity in healthy individuals, but not in individuals exhibiting fragmented or complexed α₁PI. Using these techniques, both the antigenic and functional levels of α₁PI 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 α₁PI compared with their orally healthy counterparts or normal controls. In contrast, functional analysis of serum α₁PI revealed no differences between the three subject populations. The elevated levels of antigenically determined serum α₁PI reflect the inflammatory status of periodontal disease. These results support the importance of and provide methodology for determining the functionally active levels of α₁PI allowing reexamination of changes detected during the acute phase of inflammation, replacement therapy, and longitudinal studies in relevant disease processes including malignancy and diabetes.