ENDOTHELIAL DYSFUNCTION AND VASOOCCLUSION IN SICKLE CELL ANEMIA

镰状细胞性贫血中的内皮功能障碍和血管闭塞

基本信息

批准号：
6667525
负责人：
VIJAY K. KALRA
金额：
$ 49.81万
依托单位：
UNIVERSITY OF SOUTHERN CALIFORNIA
依托单位国家：
美国
项目类别：
财政年份：
2002
资助国家：
美国
起止时间：
2002-04-15 至 2003-03-31
项目状态：
已结题

项目摘要

Vasoocclusive crisis is the major cause of morbidity and mortality in sickle cell disease (SCD). The molecular cascade giving rise to slowed blood-vessel flow remains elusive. Slowing of blood flow and hence oxygen delivery to tissues very likely relates to the adherence of sickle cell (SS) RBC to vascular endothelium, localized tissue hypoxia secondary to blood stasis and/or inflammation common in SCD. SS RBC adherence may affect blood flow by promoting, via surface expression of counter- receptors on the endothelium, the additional binding of SS RBC and leukocytes. We hypothesize that SS RBC adherence to endothelium generates cellular oxidant stress (evidenced by lipid peroxide formation and activation of transcription factor NF-kb), leading to a surface expression (SE) of a subset of cell adhesion molecules (CAMs), ICAM-1, E- selectin and BCAM-1. Similarly, infection/inflammation causes an increase in CAM expression. The induced SE of VCAM-1 leads to a greater adherence of less dense SS RBC through their alpha4beta1 ligand. The adherence and subsequent diapedesis of PMN and monocytes occurs via the increased SE of CAMs, which act as counter-receptors for integrins on these cells. Because the activation of NF-kB occurs partly through lipid peroxides generated by oxidant stresses such as vasoocclusion, e plan to determine whether overexpression of bcl-2 (a lipid peroxide-suppressing protooncogene) in cultured human umbilical vein endothelial cells (HUVEC) will prevent lipid peroxidation, NF-Kb activation and a CAM induction when transfected HUVEC are incubated with SS RBC. Dense deoxygenated SS RBC display phosphatidylserine (PS) on the external leaflet of the RBC bilayer and adhere to HUVEC through the PS receptor, so we will determine whether LPS and cytokines, which induce the expression of PS receptor, potentiate dense deoxygenated SS RBC adherence. Additionally, hypoxia increases monocyte trans endothelial migration, which may contribute to vasoocclusion; thus we propose investigating the mechanism of hypoxia-induced diapedesis of PMN and monocytes. We will also examine the mechanism(s) by which SS RBC, LPS and cytokine interaction with HUVEC alters CAM expression, the concomitant adherence of PMN and monocytes, and the augmented SS RBC adherence. Moreover, we will investigate CAM induction in endothelial cells derived from distinct vascular beds (pulmonary microvessel, pulmonary artery and brain), and potential differences in such induction effected by various shear stresses, mimicking conditions prevalent in various vascular beds. These completed studies will provide insight into new pharmacological and/or molecular approaches to ameliorate the clinical manifestations of vasoocclusion in sickle cell disease.

血管闭塞危象是导致患者发病和死亡的主要原因镰状细胞病（SCD）。分子级联反应减慢血管流动仍然难以捉摸。血流减慢，氧气随之减慢向组织的递送很可能与镰状细胞的粘附有关 (SS)红细胞至血管内皮，继发于局部组织缺氧 SCD 中常见的血瘀和/或炎症。 SS RBC 依从性可能通过促进反作用的表面表达来影响血流内皮细胞上的受体，SS RBC 的额外结合和白细胞。我们假设 SS RBC 粘附于内皮细胞产生细胞氧化应激（通过脂质过氧化物形成和转录因子 NF-kb 的激活），导致表面表达 (SE) 细胞粘附分子 (CAM)、ICAM-1、E-选择素子集和BCAM-1。同样，感染/炎症会导致 CAM 增加表达。 VCAM-1 诱导的 SE 导致更少的更大的粘附通过其 alpha4beta1 配体形成致密 SS RBC。的坚持和随后 PMN 和单核细胞的血细胞渗出是通过 SE 的增加而发生的 CAM，充当这些细胞上整合素的反受体。因为 NF-kB 的激活部分是通过脂质过氧化物发生的氧化应激，例如血管闭塞，我们计划确定是否 bcl-2（一种抑制脂质过氧化物的原癌基因）在培养的人脐静脉内皮细胞（HUVEC）将防止脂质转染 HUVEC 时的过氧化、NF-Kb 激活和 CAM 诱导与 SS RBC 一起孵育。致密脱氧 SS 红细胞在表面显示磷脂酰丝氨酸 (PS) RBC 双层的外部小叶并通过 PS 粘附到 HUVEC 受体，因此我们将确定 LPS 和细胞因子是否会诱导 PS 受体的表达，增强致密脱氧 SS 红细胞的粘附。此外，缺氧会增加单核细胞跨内皮迁移，这可能会导致血管闭塞；因此我们建议调查缺氧诱导中性粒细胞和单核细胞血细胞渗出的机制。我们也会检查 SS RBC、LPS 和细胞因子相互作用的机制 HUVEC 改变 CAM 表达，伴随 PMN 和单核细胞和增强的 SS 红细胞粘附。此外，我们将研究源自不同来源的内皮细胞的 CAM 诱导血管床（肺微血管、肺动脉和脑），以及受各种剪切力影响的这种感应的电位差应力，模仿各种血管床中普遍存在的条件。这些已完成的研究将提供对新药理学和/或改善临床表现的分子方法镰状细胞病中的血管闭塞。