Legendre P, Salsmann A, Rayes J, Trassard O, Kieffer N, Baruch D
INSERM U143, Le Kremlin-Bicetre, France.
J Thromb Haemost. 2006 Jan;4(1):236-46. doi: 10.1111/j.1538-7836.2005.01653.x.
Alpha(IIb)beta3-mediated platelet adhesive interactions in the vasculature, which are dependent on the functional state of this receptor, may be sensitive to shear forces.
To evaluate the influence of the alpha(IIb)beta3 affinity state on cell attachment under flow, we compared Chinese hamster ovary cells expressing the low affinity alpha(IIb)beta3 wild-type (wt) receptor to those expressing the high affinity alpha(IIb)beta3 T562N receptor.
We designed a real-time videomicroscopy adhesion assay for von Willebrand factor (VWF) or fibrinogen under flow conditions.
At 50 s(-1), alpha(IIb)beta3 T562N supported higher cell adhesion to fibrinogen (63.3 +/- 2.9 cells/field) than alpha(IIb)beta3 wt (38.7 +/- 2.4 cells/field, P < 0.0001). At 100 s(-1), alpha(IIb)beta3 T562N mediated cell adhesion (40.5 +/- 3.8 cells/field), while alpha(IIb)beta3 wt did not (5.3 +/- 1.4 cells/field, P < 0.001), allowing to discriminate the efficiency of each receptor. Similar findings were observed for adhesion to VWF. Complete inhibition of cell adhesion to fibrinogen was achieved with 800 microM fibrinogen gamma-chain dodecapeptide [HHLGGAKQAGDV (H12)], while Arg-Gly-Asp-Ser (RGDS) peptide (10-1000 microM) induced a dose-dependent cell detachment. These results suggest that the H12 motif allows initial attachment, in contrast to the RGDS site, which strengthens the stability of adhesion. Interestingly, compared with wt, a 10-fold lower concentration of RGDS was required to reach a similar reduction of cell adhesion mediated by alpha(IIb)beta3 T562N.
Our data show that alpha(IIb)beta3 activation is associated with a stabilization of integrin binding to fibrinogen or VWF under shear.
血管系统中α（IIb）β3介导的血小板黏附相互作用取决于该受体的功能状态，可能对剪切力敏感。
为了评估α（IIb）β3亲和力状态对流动条件下细胞黏附的影响，我们将表达低亲和力α（IIb）β3野生型（wt）受体的中国仓鼠卵巢细胞与表达高亲和力α（IIb）β3 T562N受体的细胞进行了比较。
我们设计了一种实时视频显微镜黏附试验，用于在流动条件下检测血管性血友病因子（VWF）或纤维蛋白原。
在50秒^（-1）时，α（IIb）β3 T562N支持的细胞对纤维蛋白原的黏附（63.3±2.9个细胞/视野）高于α（IIb）β3 wt（38.7±2.4个细胞/视野，P<0.0001）。在100秒^（-1）时，α（IIb）β3 T562N介导细胞黏附（40.5±3.8个细胞/视野），而α（IIb）β3 wt则不介导（5.3±1.4个细胞/视野，P<0.001），从而能够区分每种受体的效率。对于与VWF的黏附也观察到了类似的结果。用800微摩尔纤维蛋白原γ链十二肽[HHLGGAKQAGDV（H12）]可完全抑制细胞对纤维蛋白原的黏附，而精氨酸-甘氨酸-天冬氨酸-丝氨酸（RGDS）肽（10-1000微摩尔）诱导剂量依赖性的细胞脱离。这些结果表明，与RGDS位点相反，H12基序允许初始黏附，而RGDS位点增强了黏附的稳定性。有趣的是，与wt相比，达到类似程度的α（IIb）β3 T562N介导的细胞黏附减少所需的RGDS浓度低10倍。
我们的数据表明，α（IIb）β3激活与剪切力作用下整合素与纤维蛋白原或VWF结合的稳定有关。
