Laboratory of Immunobiology, Department of Microbiology and Immunology, Rega Institute for Medical Research, University of Leuven, KU Leuven, Leuven, Belgium.
Department of General Internal Medicine, University Hospital Gasthuisberg, KU Leuven, Leuven, Belgium.
J Cell Mol Med. 2019 Jan;23(1):576-585. doi: 10.1111/jcmm.13962. Epub 2018 Oct 24.
Gelatin zymography analysis is a sensitive method and commonly used to characterize and quantify the presence of the gelatinases (MMP-2 and MMP-9) in biological samples. In human plasma samples from healthy controls and systemic lupus erythematosus (SLE) patients, we observed a gelatinolytic molecule at 80 kDa, suggestive for activated human MMP-9. However, by developing and using the EDTA/gelatin zymography method and after purification of the 80 kDa entity, we proved that this molecule was the C1s subunit of the complement system. The zymolytic capacity of C1s was validated and found to be enhanced, in the absence of calcium and in the presence of EDTA. Our findings indicate that for correct identification of gelatinolytic proteins in complex biological samples the use of EDTA/gelatin zymography for enzyme development is advised. In addition, by quantification of EDTA/gelatin zymography analysis and ELISA, we observed that the levels of C1s were higher in plasma and immune complexes of SLE patients than of healthy individuals. Therefore, our data imply that C1s may become a marker for the diagnosis of SLE.
明胶酶谱分析是一种敏感的方法，常用于对生物样本中明胶酶（MMP-2 和 MMP-9）的存在进行特征分析和定量。在来自健康对照者和系统性红斑狼疮（SLE）患者的人血浆样本中，我们观察到一种 80 kDa 的明胶酶解分子，提示为激活的人 MMP-9。然而，通过开发和使用 EDTA/明胶酶谱分析方法，并对 80 kDa 实体进行纯化后，我们证明该分子是补体系统的 C1s 亚基。C1s 的酶解能力得到验证，并发现其在没有钙和存在 EDTA 的情况下增强。我们的研究结果表明，为了正确鉴定复杂生物样本中的明胶酶解蛋白，建议使用 EDTA/明胶酶谱分析来进行酶开发。此外，通过 EDTA/明胶酶谱分析和 ELISA 的定量，我们观察到 SLE 患者的血浆和免疫复合物中 C1s 的水平高于健康个体。因此，我们的数据表明 C1s 可能成为 SLE 诊断的标志物。
