Department of Clinical Chemistry, Central Diagnostic Laboratory, Maastricht University Medical Center;
Cardiovascular Research Institute Maastricht (CARIM), Maastricht University.
Clin Chem. 2017 Mar;63(3):683-690. doi: 10.1373/clinchem.2016.261644. Epub 2017 Jan 10.
We have found previously that in acute myocardial infarction (AMI), cardiac troponin T (cTnT) is degraded in a time-dependent pattern. We investigated whether cTnT forms differed in patients with chronic cTnT increases, as seen with renal dysfunction, from those in the acute phase of myocardial infarction.
We separated cTnT forms by gel filtration chromatography (GFC) in end-stage renal disease (ESRD) patients: prehemodialysis (pre-HD) and post-HD (n = 10) and 2 months follow-up (n = 6). Purified (cTnT) standards, quality control materials of the clinical cTnT immunoassay (Roche), and AMI patients' sera also were analyzed. Immunoprecipitation and Western blotting were performed with the original cTnT antibodies from the clinical assay and antibodies against the N- and C-terminal end of cTnT.
GFC analysis revealed the retention of purified cTnT at 27.5 mL, identical to that for cTnT in quality controls. For all ESRD patients, one cTnT peak was found at 45 mL, pre- and post-HD, and stable over time. Western blotting illustrated that this peak corresponded to cTnT fragments <18 kDa missing the N- and C-terminal ends. AMI patients' sera revealed cTnT peaks at 27.5 and 45 mL, respectively, corresponding to N-terminal truncated cTnT of 29 kDa and N- and C-terminal truncated fragments of <18 kDa, respectively.
We found that cTnT forms in ESRD patients are small (<18 kDa) and different from forms seen in AMI patients. These insights may prove useful for development of a more specific cTnT immunoassay, especially for the acute and diagnostic phase of myocardial infarction.
我们之前发现，在急性心肌梗死（AMI）中，心肌肌钙蛋白 T（cTnT）呈时间依赖性降解。我们研究了肾功能不全导致慢性 cTnT 升高的患者与 AMI 急性期患者的 cTnT 形式是否存在差异。
我们通过凝胶过滤色谱（GFC）在终末期肾病（ESRD）患者中分离 cTnT 形式：血液透析前（pre-HD）和血液透析后（n=10）以及 2 个月随访（n=6）。还分析了纯化的（cTnT）标准品、临床 cTnT 免疫分析的质控材料（罗氏）和 AMI 患者的血清。使用来自临床检测的原始 cTnT 抗体和针对 cTnT N 端和 C 端的抗体进行免疫沉淀和 Western 印迹。
GFC 分析显示，纯化的 cTnT 在 27.5ml 处保留，与质控品中的 cTnT 相同。对于所有 ESRD 患者，在血液透析前和后以及随时间推移，均在 45ml 处发现一个 cTnT 峰。Western 印迹表明，该峰对应于缺失 N 端和 C 端的<18kDa 的 cTnT 片段。AMI 患者的血清显示出分别在 27.5 和 45ml 处的 cTnT 峰，分别对应于 29kDa 的 N 端截断的 cTnT 和<18kDa 的 N 端和 C 端截断的片段。
我们发现 ESRD 患者的 cTnT 形式较小（<18kDa）且与 AMI 患者的形式不同。这些见解可能对开发更特异的 cTnT 免疫分析有用，特别是在 AMI 的急性和诊断阶段。
