Circulating microparticle effects on phenotypically distinct pulmonary endothelium

项目摘要 肺动脉和毛细血管内皮细胞在结构和功能上是异质的， 包括炎症反应。在肺动脉高压（PAH）中，CD 68阳性 单核细胞聚集在小动脉周围，而不是像急性炎症中那样迁移到肺泡腔中。 肺损伤此外，炎性浸润是晚期疾病的特征，局限于小动脉， 在PAH的早期阶段并不明显。炎症细胞浸润的贡献 包括释放生长因子和进一步炎症刺激。影响人类行为的精确机制 炎性细胞在选定大小血管中暂时聚集到血管周围空间以及 表型上不同的内皮仍然是未知的。表型不同的细胞内的微粒信号传导 内皮细胞尚未被考虑，可能对粘附分子有重要意义 表情我们发现晚期PAH大鼠的微粒被内吞并定位于 肺动脉内皮细胞（PAECs）核周间隙，诱导粘附分子表达， 促进白细胞粘附。然而，这些事件均未发生在肺微血管 内皮细胞（PMVEC），用相同的治疗。此外，从早期-晚期的循环中分离的微粒， 阶段PAH大鼠对两种细胞类型都没有这些影响。为了确定微粒的作用机理， 我们检查了PAEC中负责囊泡运输和细胞内信号传导的Rab蛋白 本地化PAEC和PMVEC的RNAseq分析和RT-PCR显示Rab 36仅在 PAEC。Rab 36是一个新发现的RabGT酶，负责囊泡向高尔基体的逆行转运 和核。基于这些数据，该提案测试了后期阶段的总体假设， 在PAH中循环的微颗粒，选择性地增加粘附分子在肺动脉中的表达。 肺动脉依赖Rab 36。特定目标测试的假设， 从晚期而非早期PAH获得的微粒：1.刺激表达和活动 粘附分子在肺动脉中的选择性。2.依赖于Rab 36的摄取， 肺动脉中的亚细胞定位。

PROJECT SUMMARY Pulmonary artery and capillary endothelial cells are structurally and functionally heterogeneous in multiple ways including their inflammatory response. In pulmonary arterial hypertension (PAH), CD68-positive monocytes accumulate around arterioles as opposed to migrating into the alveolar space as they do in acute lung injury. Further, inflammatory infiltrates are a feature of late-stage disease, are localized to small arterioles, and are not as observable in the early stages of PAH. The contributions of the inflammatory cell infiltrates include release of growth factors and further inflammatory stimuli. The precise mechanisms that influence the temporal recruitment of inflammatory cells to the perivascular space in select size vessels and the influence of the phenotypically distinct endothelium remain unknown. Microparticle signaling within phenotypically distinct endothelium has not been considered and could have significant implications toward adhesion molecule expression. We found that microparticles from late-stage PAH rats were endocytosed and localized to the perinuclear space in pulmonary artery endothelial cells (PAECs), induced adhesion molecule expression, and promoted leukocyte adhesion. However, none of these events occurred in pulmonary microvascular endothelium (PMVECs) with the same treatment. Further, microparticles isolated from the circulation of early- stage PAH rats had none of these effects on either cell type. To determine the mechanism of microparticle signaling in the PAECs we examined Rab proteins responsible for vesicle trafficking and intracellular localization. RNAseq analysis and RT-PCR of PAECs and PMVECs revealed Rab36 is expressed only in the PAECs. Rab36 is a newly identified RabGTPase responsible for retrograde transport of vesicles to the Golgi and nucleus. Based on these data this proposal tests the overall HYPOTHESIS that late-stage microparticles, circulating in PAH, increase adhesion molecule expression selectively in the pulmonary arteries dependent on Rab36. SPECIFIC AIMS test the hypotheses that circulating microparticles obtained from late-stage, but not early-stage PAH: 1. Stimulate expression and activity of adhesion molecules selectively in the pulmonary artery. 2. Are dependent on Rab36 for uptake and subcellular localization in the pulmonary artery.