HECA-452 is a non-function blocking antibody for isolated sialyl Lewis x adhesion to endothelial expressed E-selectin under flow conditions
HECA-452 is a non-function blocking antibody for isolated sialyl Lewis x adhesion to endothelial expressed E-selectin under flow conditions
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DOI：
10.1016/j.jim.2012.07.003
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发表时间：
2012-10-31
影响因子：
2.2
通讯作者：
Goetz, Douglas J.
中科院分区：
文献类型：
--
作者：
Kummitha, China Malakondaiah;Shirure, Venktesh S.;Goetz, Douglas J.
炎症内皮细胞上表达的E-选择素和白细胞表面的唾液酸化Lewis x(SLE(X))在白细胞向炎症部位募集的过程中，在白细胞与内皮细胞的相互作用中起着关键作用。HECA-452是一种识别SLE(X)的单抗，经常被来自不同领域的研究人员使用，他们试图解开白细胞黏附的机制。关于HECA-452抑制碳水化合物介导的白细胞与E-选择素黏附的能力的数据仍然存在矛盾，部分原因是白细胞上存在各种潜在的E-选择素反应部分。认识到这一点，我们使用了一种补充的方法来深入了解HECA-452黏附试验。具体地说，我们使用SLE(X)微球研究了HECA-452是一种无功能的阻断单抗的假设，该单抗可用于分离的SLE(X)介导的与内皮表达的E-选择素的黏附。流式细胞仪分析表明，HECA-452识别并结合到SLE(X)微球上。以1.5dyn/cm(2)的速度在人脐静脉内皮细胞(HUVEC)上灌流SLE(X)微球，发现微球通过E-选择素与4h白介素1β(IL-1β)激活的HUVEC结合。用HECA-452处理SLE(X)微球不影响SLE(X)微球在IL-1β激活的人脐静脉内皮细胞上的初始连接和聚集。神经氨酸酶和岩藻糖苷酶对SLE(X)微球的处理表明，E-选择素结合需要唾液酸和岩藻糖，而HECA-452对SLE(X)的识别不依赖于E-选择素识别所需的岩藻糖部分。后一项发现表明，E-选择素和HECA-452的SLE(X)抗原之间存在潜在的细微差异。综上所述，这些数据表明，HECA-452是一种非抑制SLE(X)介导的与内皮表达的E-选择素的黏附。(C)2012爱思唯尔B.V.保留所有权利。
E-selectin, expressed on inflamed endothelium, and sialyl Lewis x (sLe(x)), present on the surface of leukocytes, play a key role in leukocyte-endothelial interactions during leukocyte recruitment to sites of inflammation. HECA-452 is a monoclonal antibody (mAb) that recognizes sLe(x) and is routinely used by investigators from diverse fields who seek to unravel the mechanisms of leukocyte adhesion. The data regarding the ability of HECA-452 to inhibit carbohydrate-mediated leukocyte adhesion to E-selectin remains conflicted, in part due to the presence of a variety of potential E-selectin reactive moieties on leukocytes. Recognizing this, we utilized a complementary approach to gain insight into HECA-452 adhesion assays. Specifically, we used sLe(x) microspheres to investigate the hypothesis that HECA-452 is a non-function blocking mAb for isolated sLe(x) mediated adhesion to endothelial expressed E-selectin. Flow cytometric analysis revealed that HECA-452 recognizes and binds to the sLe(x) microspheres. Perfusion of the sLe(x) microspheres over human umbilical vein endothelial cells (HUVEC) at 1.5 dyn/cm(2) revealed that the microspheres attach to 4 h interleukin (IL)-1 beta activated HUVEC specifically via E-selectin. Pretreatment of the sLe(x) microspheres with HECA-452 did not influence sLe(x) microsphere initial tethering and accumulation on IL-1 beta activated HUVEC. Neuraminidase and fucosidase treatments of sLe(x) microspheres revealed that sialic acid and fucose are required for E-selectin binding, whereas HECA-452 recognition of sLe(x) does not depend on the fucose moiety to the extent required for E-selectin recognition. This latter finding suggests there are potential subtle differences between the sLe(x) antigens for E-selectin and HECA-452. Combined, the data indicate that HECA-452 is a non-inhibitor of sLe(x)-mediated adhesion to endothelial expressed E-selectin. (C) 2012 Elsevier B.V. All rights reserved.