Serum anti-SERPINE1 antibody as a potential biomarker of acute cerebral infarction

The presence of disease-specific antigens and autoantibodies in the sera of patients with atherosclerosis-related diseases has been widely reported and is considered to result from inflammation of the arterial wall and the involvement of immune factors. The aim of this study was to identify a novel antibody in patients with ischemic stroke by serological identification of antigens using recombinant cDNA expression cloning from patients who had a transient ischemic attack (TIA). We identified the serpin peptidase inhibitor, clade E member 1 (SERPINE1), as a candidate antigen. The serum anti-SERPINE1 antibody levels quantified using amplified luminescent proximity homogeneous assay-linked immunosorbent assay were significantly higher in patients with ischemic stroke, including those with acute cerebral infarction (aCI), TIA, and chronic cerebral infarction, than in healthy donors. The antibody levels were strongly associated with old age, female sex, and presence of hypertension, diabetes mellitus, and cardiovascular disease. Age and intima-media thickness of the carotid artery were positively correlated with antibody levels, which suggests that SERPINE1 may reflect the progression of atherosclerosis. In a multivariate analysis, SERPINE1 antibody level was an independent predictor of aCI. Thus, the serum levels of anti-SERPINE1 antibody could potentially serve as a biomarker of atherothrombotic infarction.

Serum anti-AP3D1 antibodies are risk factors for acute ischemic stroke related with atherosclerosis

Atherosclerosis has been considered as the main cause of morbidity, mortality, and disability worldwide. The first screening for antigen markers was conducted using the serological identification of antigens by recombinant cDNA expression cloning, which has identified adaptor-related protein complex 3 subunit delta 1 (AP3D1) as an antigen recognized by serum IgG antibodies of patients with atherosclerosis. Serum antibody levels were examined using the amplified luminescent proximity homogeneous assay-linked immunosorbent assay (AlphaLISA) using a recombinant protein as an antigen. It was determined that the serum antibody levels against AP3D1 were higher in patients with acute ischemic stroke (AIS), transient ischemic attack, diabetes mellitus (DM), cardiovascular disease, chronic kidney disease (CKD), esophageal squamous cell carcinoma (ESCC), and colorectal carcinoma than those in the healthy donors. The area under the curve values of DM, nephrosclerosis type of CKD, and ESCC calculated using receiver operating characteristic curve analysis were higher than those of other diseases. Correlation analysis showed that the anti-AP3D1 antibody levels were highly associated with maximum intima-media thickness, which indicates that this marker reflected the development of atherosclerosis. The results of the Japan Public Health Center-based Prospective Study indicated that this antibody marker is deemed useful as risk factors for AIS.

Serum Anti-AP3D1 Antibodies Are Risk Factors for Acute Ischemic Stroke Related With Atherosclerosis

Atherosclerosis has been considered as the main cause of morbidity, mortality, and disability worldwide. The first screening for antigen markers was conducted using the serological identification of antigens by recombinant cDNA expression cloning, which has identified adaptor-related protein complex 3 subunit delta 1 (AP3D1) as an antigen recognized by serum IgG antibodies of patients with atherosclerosis. Serum antibody levels were examined using the amplified luminescent proximity homogeneous assay-linked immunosorbent assay (AlphaLISA) using a recombinant protein as an antigen. It was determined that the serum antibody levels against AP3D1 were higher in patients with acute ischemic stroke, transient ischemic attack , diabetes mellitus (DM), cardiovascular disease , chronic kidney disease (CKD), esophageal squamous cell carcinoma (ESCC), and colorectal carcinoma than those in the healthy donors. The area under the curve values of DM, nephrosclerosis type of CKD, and ESCC calculated using receiver operating characteristic curve analysis were higher than that of other diseases. Correlation analysis showed that the anti-AP3D1 antibody levels were highly associated with maximum intima-media thickness, which indicates that this marker reflected the development of atherosclerosis. The results of the Japan Public Health Center-based Prospective Study indicated that this antibody marker is deemed useful as risk factors for AIS.

Structural and functional characterization of a glycoside hydrolase family 3 β-N-acetylglucosaminidase from Paenibacillus sp. str. FPU-7

Chitin, a β-1,4-linked homopolysaccharide of N-acetyl-d-glucosamine (GlcNAc), is one of the most abundant biopolymers on Earth. Paenibacillus sp. str. FPU-7 produces several different chitinases and converts chitin into N,N′-diacetylchitobiose ((GlcNAc)2) in the culture medium. However, the mechanism by which the Paenibacillus species imports (GlcNAc)2 into the cytoplasm and divides it into the monomer GlcNAc remains unclear. The gene encoding Paenibacillus β-N-acetyl-d-glucosaminidase (PsNagA) was identified in the Paenibacillus sp. str. FPU-7 genome using an expression cloning system. The deduced amino acid sequence of PsNagA suggests that the enzyme is a part of the glycoside hydrolase family 3 (GH3). Recombinant PsNagA was successfully overexpressed in Escherichia coli and purified to homogeneity. As assessed by gel permeation chromatography, the enzyme exists as a 57-kDa monomer. PsNagA specifically hydrolyses chitin oligosaccharides, (GlcNAc)2–4, 4-nitrophenyl N-acetyl β-d-glucosamine (pNP-GlcNAc) and pNP-(GlcNAc)2–6, but has no detectable activity against 4-nitrophenyl β-d-glucose, 4-nitrophenyl β-d-galactosamine and colloidal chitin. In this study, we present a 1.9 Å crystal structure of PsNagA bound to GlcNAc. The crystal structure reveals structural features related to substrate recognition and the catalytic mechanism of PsNagA. This is the first study on the structural and functional characterization of a GH3 β-N-acetyl-d-glucosaminidase from Paenibacillus sp.