Increased Neutrophil Elastase and Proteinase 3 and Augmented NETosis Are Closely Associated With β-Cell Autoimmunity in Patients With Type 1 Diabetes

Abstract

Type 1 diabetes (T1D) is an autoimmune disease resulting from the self-destruction of insulin-producing β-cells. Reduced neutrophil counts have been observed in patients with T1D. However, the pathological roles of neutrophils in the development of T1D remain unknown. Here we show that circulating protein levels and enzymatic activities of neutrophil elastase (NE) and proteinase 3 (PR3), both of which are neutrophil serine proteases stored in neutrophil primary granules, were markedly elevated in patients with T1D, especially those with disease duration of less than 1 year. Furthermore, circulating NE and PR3 levels increased progressively with the increase of the positive numbers and titers of the autoantibodies against β-cell antigens. An obvious elevation of NE and PR3 was detected even in those autoantibody-negative patients. Increased NE and PR3 in T1D patients are closely associated with elevated formation of neutrophil extracellular traps. By contrast, the circulating levels of α1-antitrypsin, an endogenous inhibitor of neutrophil serine proteases, are decreased in T1D patients. These findings support an early role of neutrophil activation and augmented neutrophil serine proteases activities in the pathogenesis of β-cell autoimmunity and also suggest that circulating NE and PR3 may serve as sensitive biomarkers for the diagnosis of T1D.

NETosis-associated serum biomarkers are reduced in type 1 diabetes in association with neutrophil count

“…These results are in contrast to a recent study of paediatric subjects with T1D in China [9], and although the reasons for the discrepancy are not clear, they could be related to the populations studied (onset age, genetics, environment, standard of care) or to the analytical methods employed. We did not see any indication that ethnicity was a clear explanation, although the number of subjects of Chinese descent in our study was limited and a relevant difference in genetics between the populations cannot be ruled out.…”

“…In this study, we sought to confirm the recent report of elevated NETosis-associated factors in the serum of subjects with T1D [9] in order to explore further the utility of this biomarker for predicting clinical outcomes in T1D.…”

“…In addition, multiple laboratories have reported a reduction in the number of circulating neutrophils near the time of T1D diagnosis as well as prior to onset [7,8]. Somewhat paradoxically considering these findings, a recent study has raised a significant amount of interest in the field, showing extremely high levels of neutrophil-derived enzymes in the serum of newly diagnosed T1D patients [9]. Specifically, these products were associated with NETosis, a neutrophil-specific form of anti-microbial cell death.…”

“…To address whether this reduction could be observed with other NETosis markers, we proceeded to analyse NE, a second marker that was reported to be elevated in T1D along with PR3 [9]. As with PR3, NE was also found at lower average levels in T1D, with the lowest levels being found in the subgroup closer to diagnosis (Fig.…”

“…The most marked elevation in a serum NETosis-associated biomarker reported previously was in PR3 [9], so we first evaluated levels of this enzyme in our cohorts using a commercial ELISA. In contrast to previous findings, average serum PR3 levels were actually modestly decreased, by 26Á2%, in subjects with T1D compared with matched controls (Fig.…”