Exposure of plasminogen and a novel plasminogen receptor, Plg-RKT, on activated human and murine platelets.
- 作者列表："Whyte CS","Morrow GB","Baik N","Booth NA","Jalal MM","Parmer RJ","Miles LA","Mutch NJ
:Plasminogen activation rates are enhanced by cell surface binding. We previously demonstrated that exogenous plasminogen binds to phosphatidylserine-exposing and spread platelets. Platelets contain plasminogen in their α-granules, but secretion of plasminogen from platelets has not been studied. Recently, a novel transmembrane lysine-dependent plasminogen receptor, Plg-RKT, has been described on macrophages. Here, we analyzed the pool of plasminogen in platelets and examined whether platelets express Plg-RKT. Plasminogen content of the supernatant of resting and collagen/thrombin-stimulated platelets was similar. Pretreatment with the lysine analog, ε-aminocaproic acid, significantly increased platelet-derived plasminogen (0.33 vs 0.08 nmol/108 platelets) in the stimulated supernatant, indicating a lysine-dependent mechanism of membrane retention. Lysine-dependent, platelet-derived plasminogen retention on thrombin and convulxin activated human platelets was confirmed by flow cytometry. Platelets initiated fibrinolytic activity in fluorescently labeled plasminogen-deficient clots and in turbidimetric clot lysis assays. A 17-kDa band, consistent with Plg-RKT, was detected in the platelet membrane fraction by western blotting. Confocal microscopy of stimulated platelets revealed Plg-RKT colocalized with platelet-derived plasminogen on the activated platelet membrane. Plasminogen exposure was significantly attenuated in thrombin- and convulxin-stimulated platelets from Plg-RKT-/- mice compared with Plg-RKT+/+ littermates. Membrane exposure of Plg-RKT was not dependent on plasminogen, as similar levels of the receptor were detected in plasminogen-/- platelets. These data highlight Plg-RKT as a novel plasminogen receptor in human and murine platelets. We show for the first time that platelet-derived plasminogen is retained on the activated platelet membrane and drives local fibrinolysis by enhancing cell surface-mediated plasminogen activation.
: 纤溶酶原激活速率通过细胞表面结合而增强。我们以前证明外源性纤溶酶原与磷脂酰丝氨酸接触和扩散的血小板结合。血小板在其 α-颗粒中含有纤溶酶原，但尚未研究从血小板分泌纤溶酶原。最近，在巨噬细胞上描述了一种新的跨膜赖氨酸依赖性纤溶酶原受体plg-rkt。在这里，我们分析了血小板中的纤溶酶原池，并检查了血小板是否表达plg-rkt。静息和胶原/凝血酶刺激的血小板的上清液的纤溶酶原含量是相似的。用赖氨酸类似物 ε-氨基己酸预处理显著增加刺激上清液中的血小板衍生纤溶酶原 (0.33对0.08 nmol/108血小板)，表明赖氨酸依赖性膜保留机制。通过流式细胞术证实赖氨酸依赖性、血小板衍生的纤溶酶原在凝血酶和惊厥活化的人血小板上的保留。血小板在荧光标记的纤溶酶原缺乏凝块和浊度测定凝块溶解试验中引发纤维蛋白溶解活性。通过蛋白质印迹法在血小板膜组分中检测到与plg-rkt一致的17-kda条带。刺激的血小板共聚焦显微镜显示Plg-RKT与血小板衍生的纤溶酶原在活化的血小板膜上共定位。与plg-rkt +/+ 同窝小鼠相比，来自plg-rkt-/-小鼠的凝血酶和惊厥刺激的血小板中的纤溶酶原暴露显著减弱。Plg-rkt的膜暴露不依赖于纤溶酶原，因为在纤溶酶原-/-血小板中检测到类似水平的受体。这些数据强调Plg-RKT是人和鼠血小板中新的纤溶酶原受体。我们首次证明血小板衍生的纤溶酶原保留在活化的血小板膜上，并通过增强细胞表面介导的纤溶酶原激活来驱动局部纤维蛋白溶解。
METHODS::The ATP binding-cassette superfamily corresponds the mostly transmembrane transporters family found in humans. These proteins actively transport endogenous and exogenous substrates through biological membranes in body tissues, so they have an important role in the regulation of many physiological functions necessary for human homeostasis, as well as in response regulation to several pharmacological substrates. The development of multidrug resistance has become one of the main troubles in conventional chemotherapy in different illnesses including cancer, being the increased efflux of antineoplastic drugs the main reason for this multidrug resistance, with a key role of the ABC superfamily. Likely, the interindividual variability in the pharmacological response among patients is well known, and may be due to intrinsically factors of the disease, genetic and environmental ones. Thus, the understanding of this variability, especially the genetic variability associated with the efficacy and toxicity of drugs, can provide a safer and more effective pharmacological treatment, so ABC genes are considered as important regulators due to their relationship with the reduction in pharmacological response. In this review, updated information about transporters belonging to this superfamily was collected, the possible role of these transporters in cancer, the role of genetic variability in their genes, as well as some therapeutic tools that have been tried to raise against main transporters associated with chemoresistance in cancer.
METHODS:BACKGROUND:Cholinergic neurotransmission regulates neuroinflammation in Parkinson disease (PD). RESEARCH DESIGN AND METHODS:The authors conducted a delayed-start study of donepezil for cognitive decline in non-demented PD patients. The study consisted of a 96-week randomized placebo-controlled double-blind phase 1, followed by a 24-week donepezil extension phase 2. The primary outcome measure was a change in the Mini-Mental State Examination (MMSE) at week 120. RESULTS:A total of 98 patients were randomly allocated to the early-start (donepezil-to-donepezil) and delayed-start (placebo-to-donepezil) groups. Mean (SD) of the baseline MMSE was 27.6 (2.0) and 28.0 (2.1), respectively. MMSE change at week 120 was better in the early-start group than in the delayed-start group, but the difference was not significant. The MMSE declined in apolipoprotein ε4 carriers, but not in non-carriers, and the factor interaction (intervention × ε4 genotype) was highly significant (P < 0.001). Analyzed with the interaction, the difference was significant (group difference 1.95 [0.33 to 3.57], P = 0.018). The MMSE decline slope in phase 1 was significantly better in the early-start group than in the delayed-start group (P = 0.048). CONCLUSIONS:Cognitive function deteriorated in ε4 carriers, but not in non-carriers, and early-start donepezil may postpone cognitive decline in the former.
METHODS::Since the discovery of dental pulp stem cells, a lot of teams have expressed an interest in dental pulp regeneration. Many approaches, experimental models and biological explorations have been developed, each including the use of stem cells and scaffolds with the final goal being clinical application in humans. In this review, the authors' objective was to compare the experimental models and strategies used for the development of biomaterials for tissue engineering of dental pulp with stem cells. Electronic queries were conducted on PubMed using the following terms: pulp regeneration, scaffold, stem cells, tissue engineering and biomaterial. The extracted data included the following information: the strategy envisaged, the type of stem cells, the experimental models, the exploration or analysis methods, the cytotoxicity or viability or proliferation cellular tests, the tests of scaffold antibacterial properties and take into account the vascularization of the regenerated dental pulp. From the 71 selected articles, 59% focused on the "cell-transplantation" strategy, 82% used in vitro experimentation, 58% in vivo animal models and only one described an in vivo in situ human clinical study. 87% used dental pulp stem cells. A majority of the studies reported histology (75%) and immunohistochemistry explorations (66%). 73% mentioned the use of cytotoxicity, proliferation or viability tests. 48% took vascularization into account but only 6% studied the antibacterial properties of the scaffolds. This article gives an overview of the methods used to regenerate dental pulp from stem cells and should help researchers create the best development strategies for research in this field.