ABSTRACT
The objective of this study was to assess the feasibility of the Arteriograph 24 device to measure 24-hour PWV and central systolic blood pressure (cSBP) in patients with type 2 diabetes (T2DM) and non-diabetic controls and compare daytime and nighttime characteristics in the two groups. Twenty-four-hour PWV and cSBP was measured in 58 patients with T2DM (mean age: 66â ±â 9 years, 50% women, mean duration of T2DM: 7.8â ±â 1.5 years) and 62 age- and sex-matched controls. Seventy percent of participants (71% T2DM patients and 69% controls) had sufficient readings to generate an acceptable 24-hour report (≥14 day and ≥7 night readings). Lower nocturnal than daytime PWV and cSBP were observed in both groups. Nocturnal PWV and cSBP dipping were attenuated in T2DM patients compared to controls (PWV: -0.3â ±â 0.9 vs. -0.7â ±â 0.9â m/s, P â =â 0.04, cSBP: -8â ±â 14 vs. -18â ±â 18â mmHg, P â <â 0.01). No group differences in PWV or cSBP were observed during daytime (T2D vs. controls, PWV: 9.2â ±â 1.1 vs. 9.2â ±â 1.3â m/s, P â =â 0.99, cSBP: 133â ±â 19 vs. 137â ±â 25â mmHg, P â =â 0.42) or nighttime (PWV: 8.9â ±â 1.3 vs. 8.4â ±â 1.3â m/s, P â =â 0.14, cSBP 124â ±â 20 vs. 118â ±â 27â mmHg, P â =â 0.26). The study findings indicate that the nocturnal dipping of PWV and cSBP is attenuated in T2DM patients. The significant number of missing measurements raises concerns regarding the clinical utility of the Arteriograph 24 device.
Subject(s)
Diabetes Mellitus, Type 2 , Vascular Stiffness , Humans , Female , Middle Aged , Aged , Male , Blood Pressure/physiology , Pulse Wave Analysis , Diabetes Mellitus, Type 2/complications , Feasibility Studies , Blood Pressure DeterminationABSTRACT
In human autoimmune diseases, low plasma levels of complement factors C3 and C4 are commonly used as a proxy for complement activation. The measurements of C3 and C4 concentrations (the result of synthesis and consumption) however, show low sensitivity in patient follow-up. We find that the estimation of the C3dg fragment released during complement activation is a better parameter for complement activation. Available techniques for measuring the activation fragment C3dg, e.g. immune-electrophoresis or involving PEG-precipitation, are time-consuming and difficult to standardize. Here we examine the specificity and use of an antibody with mono-specificity for a neoepitope at the N-terminus of C3dg, which is only exposed after cleavage of C3. We present a stable, reproducible, and easy-to-use, time-resolved immunoassay with specificity for C3dg that can be used to directly evaluate ongoing complement activation. We demonstrate that the assay can be applied to clinical samples with a high specificity (95%) and a positive likelihood ratio of 10. It can also differentiate the complement related disease Systemic Lupus Erythematosus from controls and other immune-mediatedimmune mediated diseases like Rheumatoid Arthritis (86% specificity) and Spondyloarthritis (91% specificity). Further, we establish how the assay may also be used for experimental research in in vivo mouse models.
Subject(s)
Antibodies/immunology , Arthritis, Rheumatoid/diagnosis , Complement Activation , Fluoroimmunoassay , Lupus Erythematosus, Systemic/diagnosis , Peptide Fragments/blood , Spondylarthritis/diagnosis , Animals , Arthritis, Rheumatoid/blood , Arthritis, Rheumatoid/immunology , Biomarkers/blood , Case-Control Studies , Complement C3b , Cross-Sectional Studies , Disease Models, Animal , Epitopes , Female , Humans , Lupus Erythematosus, Systemic/blood , Lupus Erythematosus, Systemic/immunology , Male , Mice, Inbred C57BL , Predictive Value of Tests , Reproducibility of Results , Spondylarthritis/blood , Spondylarthritis/immunologyABSTRACT
We aimed at establishing a sensitive and robust assay for estimation of systemic complement activation at complement component C3 level in mouse and human plasma samples. In order to capture the activation products iC3b and C3dg in a specific and physiological relevant manner we utilized a construct consisting of the iC3b/C3dg-binding site of human complement receptor 2 (CR2) attached to an Fc-part of mouse IgG. This construct binds C3dg and iC3b from both mice and humans. We purified the CR2-IgG construct from mouse B myeloma cell line supernatants, J558L-CR2-IgG, by protein G affinity chromatography. The CR2-IgG construct was used for capturing C3 fragments in microtiter wells and an anti-mouse or an anti-human-C3 antibody was used for detection of bound C3 fragments. Initially we tested the specificity of the assays with the use of purified C3 fragments. Further, with the use of the CR2-based assay, we measured an up to three-fold higher signal in activated mouse serum as compared to non-activated mouse serum, whereas activated serum from a C3 knock-out mouse gave no signal. We tested in vivo generated samples from a mouse experiment; complement activation was induced by injecting cobra venom factor or heat aggregated IgG into C57bl6 mice, followed by withdrawal of EDTA blood samples at different time points and measurement of iC3b/C3dg. We observed a clear time-dependent distinction in signals between samples with expected high and low complement activation. Furthermore, with the use of the assay for human C3 fragments, we observed that patients with systemic lupus erythematosus (SLE) (n = 144) had significantly higher iC3b/C3dg levels as compared to healthy individuals (n = 144) (p < 0.0001). We present two functional immunoassays, that are able to measure systemic levels of the C3-activation products iC3b and C3dg in mice and humans. To our knowledge, these are the first assays for complement activation that use a physiological relevant capture construct such as CR2. These assays will be a relevant tool when investigating mouse models and human diseases involving the complement system.
