Laboratory of Medical Biochemistry, Department of Pharmaceutical Sciences, University of Antwerp, 2610 Wilrijk, Belgium.
Laboratory of Cell Biology and Histology, Faculty of Pharmaceutical, Biomedical and Veterinary Sciences, University of Antwerp, 2610 Wilrijk, Belgium.
Int J Mol Sci. 2023 Feb 13;24(4):3725. doi: 10.3390/ijms24043725.
Carboxypeptidase U (CPU, TAFIa, CPB2) is a potent attenuator of fibrinolysis that is mainly synthesized by the liver as its inactive precursor proCPU. Aside from its antifibrinolytic properties, evidence exists that CPU can modulate inflammation, thereby regulating communication between coagulation and inflammation. Monocytes and macrophages play a central role in inflammation and interact with coagulation mechanisms resulting in thrombus formation. The involvement of CPU and monocytes/macrophages in inflammation and thrombus formation, and a recent hypothesis that proCPU is expressed in monocytes/macrophages, prompted us to investigate human monocytes and macrophages as a potential source of proCPU. mRNA expression and the presence of proCPU/CPU protein were studied in THP-1, PMA-stimulated THP-1 cells and primary human monocytes, M-CSF-, IFN-γ/LPS-, and IL-4-stimulated-macrophages by RT-qPCR, Western blotting, enzyme activity measurements, and immunocytochemistry. mRNA and proCPU protein were detected in THP-1 and PMA-stimulated THP-1 cells as well as in primary monocytes and macrophages. Moreover, CPU was detected in the cell medium of all investigated cell types and it was demonstrated that proCPU can be activated into functionally active CPU in the in vitro cell culture environment. Comparison of mRNA expression and proCPU concentrations in the cell medium between the different cell types provided evidence that mRNA expression and proCPU secretion in monocytes and macrophages is related to the degree to which these cells are differentiated. Our results indicate that primary monocytes and macrophages express proCPU. This sheds new light on monocytes and macrophages as local proCPU sources.
羧肽酶 U（CPU、TAFiA、CPB2）是一种有效的纤维蛋白溶解抑制剂，主要由肝脏合成其无活性前体 proCPU。除了抗纤维蛋白溶解特性外，有证据表明 CPU 可以调节炎症，从而调节凝血和炎症之间的通讯。单核细胞和巨噬细胞在炎症和相互作用中起着核心作用凝血机制导致血栓形成。CPU 和单核细胞/巨噬细胞在炎症和血栓形成中的参与，以及最近关于 proCPU 在单核细胞/巨噬细胞中表达的假设，促使我们研究人类单核细胞和巨噬细胞作为 proCPU 的潜在来源。通过 RT-qPCR、Western blotting、酶活性测定和免疫细胞化学研究了 THP-1、PMA 刺激的 THP-1 细胞和原代人单核细胞、M-CSF、IFN-γ/LPS 和 IL-4 刺激的巨噬细胞中 proCPU/CPU 蛋白的 mRNA 表达和存在。在 THP-1 和 PMA 刺激的 THP-1 细胞以及原代单核细胞和巨噬细胞中检测到 proCPU 蛋白。此外，在所有研究的细胞类型的细胞培养基中均检测到 CPU，并且证明 proCPU 可以在体外细胞培养环境中被激活为功能活性的 CPU。不同细胞类型之间细胞培养基中 proCPU 浓度和 proCPU 浓度的 mRNA 表达比较提供了证据表明单核细胞和巨噬细胞中的 proCPU 表达和分泌与这些细胞分化的程度有关。我们的研究结果表明，原代单核细胞和巨噬细胞表达 proCPU。这为单核细胞和巨噬细胞作为局部 proCPU 来源提供了新的视角。
