纤溶酶是负责处理增殖性玻璃体视网膜病变患者玻璃体中血小板衍生生长因子-C的主要蛋白酶。
Plasmin is the major protease responsible for processing PDGF-C in the vitreous of patients with proliferative vitreoretinopathy.
作者信息
Lei Hetian, Velez Gisela, Hovland Peter, Hirose Tatsuo, Kazlauskas Andrius
机构信息
Department of Ophthalmology, Schepens Eye Research Institute, Harvard Medical School, 20 Staniford Street, Boston, MA 02114, USA.
出版信息
Invest Ophthalmol Vis Sci. 2008 Jan;49(1):42-8. doi: 10.1167/iovs.07-0776.
PURPOSE
Proliferative vitreoretinopathy (PVR) is the primary cause of failure of retinal reattachment surgery. Growth factors such as platelet-derived growth factor (PDGF) are strongly associated with PVR. Of the five PDGF family members, PDGF-C predominates in the vitreous of experimental and clinical PVR. PDGF-C is secreted as a latent protein that requires proteolytic processing for activation. Although tissue plasminogen activator (tPA) is primarily responsible for processing PDGF-C in cultured cells, it constitutes a minority of the processing activity in the vitreous of experimental animals and in patients with PVR. Identifying the major PDGF-C processing protease was the purpose of this study.
METHODS
The presence of serum proteins in the vitreous was detected by Coomassie blue staining and Western blotting. PDGF-C processing activity was detected in an in vitro processing assay using either native or recombinant PDGF-C as the substrate. Plasmin activity was blocked using alpha(2)-plasmin inhibitor. Phosphorylation of the PDGF receptor (PDGFR) was monitored by antiphosphotyrosine Western blotting. Vitreous specimens were collected from experimental rabbits or from patients undergoing vitrectomy to repair retinal detachment or for other reasons.
RESULTS
A number of prominent serum proteins (albumin and IgG) were detected in the vitreous of all patients undergoing retinal surgery. The level of these proteins markedly increased in the vitreous of rabbits as they developed PVR. These observations suggested that serum-borne proteases are also likely to be present in the vitreous. Indeed, plasmin (a protease capable of processing PDGF-C) was present in the vitreous from PVR rabbits and retinal surgery patients. Plasmin was dramatically more effective than tPA in processing PDGF-C in an in vitro assay. Blocking plasmin activity eliminated most of the processing activity in the vitreous of patients and rabbits with PVR.
CONCLUSIONS
Plasmin was the major PDGF-C processing protease in the vitreous of PVR rabbits and patients undergoing retinal surgery. Blocking plasmin prevented the generation of active PDGF-C, which is the major PDGF isoform relevant for PVR. These observations are the first report of an in vivo protease responsible for processing PDGF-C. In addition, plasmin was identified as a novel therapeutic target for patients with PVR.
目的
增殖性玻璃体视网膜病变（PVR）是视网膜复位手术失败的主要原因。血小板衍生生长因子（PDGF）等生长因子与PVR密切相关。在PDGF的五个家族成员中，PDGF-C在实验性和临床性PVR的玻璃体中占主导地位。PDGF-C以潜伏蛋白的形式分泌，需要蛋白水解加工才能激活。虽然组织型纤溶酶原激活剂（tPA）主要负责在培养细胞中加工PDGF-C，但在实验动物和PVR患者的玻璃体中，它仅占加工活性的一小部分。本研究的目的是确定主要的PDGF-C加工蛋白酶。
方法
通过考马斯亮蓝染色和蛋白质印迹法检测玻璃体中血清蛋白的存在。使用天然或重组PDGF-C作为底物，通过体外加工试验检测PDGF-C加工活性。使用α(2)-纤溶酶抑制剂阻断纤溶酶活性。通过抗磷酸酪氨酸蛋白质印迹法监测PDGF受体（PDGFR）的磷酸化。从实验兔或接受玻璃体切除术以修复视网膜脱离或因其他原因接受手术的患者中收集玻璃体标本。
结果
在所有接受视网膜手术的患者的玻璃体中均检测到多种显著的血清蛋白（白蛋白和IgG）。随着兔发生PVR，这些蛋白在玻璃体中的水平显著升高。这些观察结果表明，血清来源的蛋白酶也可能存在于玻璃体中。事实上，纤溶酶（一种能够加工PDGF-C的蛋白酶）存在于PVR兔和视网膜手术患者的玻璃体中。在体外试验中，纤溶酶加工PDGF-C的效果明显优于tPA。阻断纤溶酶活性消除了PVR患者和兔玻璃体中的大部分加工活性。
结论
纤溶酶是PVR兔和接受视网膜手术患者玻璃体中主要的PDGF-C加工蛋白酶。阻断纤溶酶可防止活性PDGF-C的产生，而活性PDGF-C是与PVR相关的主要PDGF亚型。这些观察结果是关于体内负责加工PDGF-C的蛋白酶的首次报道。此外，纤溶酶被确定为PVR患者的一个新的治疗靶点。
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