Stawowy Philipp, Margeta Christian, Kallisch Heike, Seidah Nabil G, Chrétien Michel, Fleck Eckart, Graf Kristof
Department of Medicine/Cardiology, Deutsches Herzzentrum Berlin, D-13353 Berlin, Germany.
Cardiovasc Res. 2004 Jul 1;63(1):87-97. doi: 10.1016/j.cardiores.2004.03.010.
Heart failure is characterized by an imbalance of matrix synthesis/turnover, finally resulting in fibrosis. Cardiac myocytes and fibroblasts play a pivotal role in the remodeling process. Cardiac remodeling involves the expression of TGF-beta1 and matrix metalloproteinases (MMPs) in cardiac fibroblasts (CFBs). Furin, a subtilisin/kexin-like proprotein convertase (PC), activates TGF-beta1 and membrane-bound MT1-MMP, which facilitates pro-gelatinase A (MMP-2) activation. Even though several reports identified TGF-beta1 as a pro-fibrotic cytokine in the heart, it increases MMP-activity and cell migration/invasion in several cell types. The present study was done to investigate the contribution of TGF-beta1 and furin to CFBs MMP-activity and motility.
Stimulation of CFBs from adult Sprague-Dawley rats with TGF-beta1 (20 ng/ml) induced furin, but had no effect on the closely related PC5. Inhibition of furin inhibited angiotensin II-induced TGF-beta1 activation, indicating that TGF-beta1 amplifies its activating convertase in CFBs. Pretreatment of CFBs with TGF-beta1 (20 ng/ml, 24 h) increased their migration by about two-fold (p<0.05), which was accompanied by an enhanced expression and activity of MT1-MMP and MMP-2. Brefeldin A (BFA), a Golgi-disturbing agent, inhibited MT1-MMP activation, indicating that it occurs in the trans-Golgi network (TGN), where furin is concentrated and colocalized with MT1-MMP. Inhibition of furin significantly inhibited TGF-beta1-induced MT1-MMP/MMP-2 activation. Furthermore, inhibition of furin attenuated TGF-beta1-enhanced migration on gelatin-coated membranes (p<0.05). This was comparable to the effects of the MMP-inhibitor GM6001, pointing out that MMPs are major mediators of TGF-beta1-enhanced CFB motility.
We demonstrate that TGF-beta1 induces MMP-activity in CFBs, thereby facilitating CFBs motility. Furthermore, TGF-beta1 amplifies its activating convertase furin, which is also required for MT1-MMP/MMP-2 activation in CFBs. Thus, furin is central for TGF-beta1 and MT1-MMP activation and might be a novel target in cardiac remodeling.
心力衰竭的特征是基质合成/周转失衡，最终导致纤维化。心肌细胞和成纤维细胞在重塑过程中起关键作用。心脏重塑涉及心脏成纤维细胞（CFB）中转化生长因子-β1（TGF-β1）和基质金属蛋白酶（MMP）的表达。弗林蛋白酶是一种枯草杆菌蛋白酶/凯新样前蛋白转化酶（PC），可激活TGF-β1和膜结合的MT1-MMP，从而促进前明胶酶A（MMP-2）的激活。尽管有几份报告将TGF-β1确定为心脏中的促纤维化细胞因子，但它在几种细胞类型中会增加MMP活性以及细胞迁移/侵袭。本研究旨在探讨TGF-β1和弗林蛋白酶对CFB的MMP活性和运动性的作用。
用TGF-β1（20 ng/ml）刺激成年Sprague-Dawley大鼠的CFB可诱导弗林蛋白酶，但对密切相关的PC5没有影响。抑制弗林蛋白酶可抑制血管紧张素II诱导的TGF-β1激活，表明TGF-β1在CFB中放大了其激活转化酶。用TGF-β1（20 ng/ml，24小时）预处理CFB可使其迁移增加约两倍（p<0.05），同时伴有MT1-MMP和MMP-2的表达及活性增强。布雷菲德菌素A（BFA）是一种干扰高尔基体的药物，可抑制MT1-MMP的激活，表明其激活发生在反式高尔基体网络（TGN）中，弗林蛋白酶在该区域浓缩并与MT1-MMP共定位。抑制弗林蛋白酶可显著抑制TGF-β1诱导的MT1-MMP/MMP-2激活。此外，抑制弗林蛋白酶可减弱TGF-β1增强的在明胶包被膜上的迁移（p<0.05）。这与MMP抑制剂GM6001的作用相当，表明MMP是TGF-β1增强CFB运动性的主要介质。
我们证明TGF-β1可诱导CFB中的MMP活性，从而促进CFB的运动性。此外，TGF-β1放大了其激活转化酶弗林蛋白酶，而弗林蛋白酶也是CFB中MT1-MMP/MMP-2激活所必需的。因此，弗林蛋白酶是TGF-β1和MT1-MMP激活的核心，可能是心脏重塑中的一个新靶点。
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