Development of Robust Culture Systems for the Human Splenic Littoral Cell

描述(申请人提供)：脾是人类最大的次级免疫系统器官。脾不全的人死于严重的脓毒症的几率会增加。他们患自身免疫性疾病、某些癌症和缺血性心脏病的风险也更大。死亡率的提高归因于缺乏隔离在脾中的吞噬细胞，从而有效地消除了适当的靶点，尽管相关细胞可以在其他部位找到。虽然脾白髓的整体组织类似于淋巴结，但脾并不与淋巴系统相连。相反，病原体、红细胞、凋亡/坏死、改变和肿瘤细胞都在血液中传递到脾和离开脾。与白髓不同的是，脾红髓既是一个蓄水池，又是一个过滤器，它决定了流经其中的细胞是被保留、破坏还是回到循环中。人静脉窦衬里细胞，称为海岸细胞(LC)，约占红髓的30%。越来越多的证据表明，这些高度特化的脾细胞是红细胞和其他细胞是否被破坏、保留或重新进入循环的主要决定因素，尽管调节它们功能的机制完全不清楚。我们发现LCS不是一种经典的内皮细胞，因为它具有多个巨噬细胞和平滑肌相关抗原，是CD8+，缺乏传统的内皮标志物和造血祖细胞系。那么，为什么人们对LC函数知之甚少呢？LCs的获取、纯化和培养历来被证明是非常困难的。基于我们最近在获取新鲜脾以及识别区别于其他脾细胞的标记方面的成功，我们现在准备进一步分离和分析LC基因/蛋白表达(AIM I)。尽管像LC这样在复杂器官内形成特化结构的细胞通常很难在体外繁殖，但克服这些限制将代表着对LC功能理解的重大进步(S)。我们聚集了一组专家内皮细胞生物学家、免疫学家和病理学家，他们将为我们提供深入的指导，因为我们开发了多种LC培养系统(AIM II)，这些系统既具有生理相关性，又易于处理。这些方法将有助于发现LC功能的基础机制，并导致能够在不同疾病状态(如败血症、疟疾、血红蛋白病等)下调节脾滤过的治疗方法的开发。

DESCRIPTION (provided by applicant): The spleen is the largest secondary immune system organ in man. Individuals who are asplenic die at an increased rate from overwhelming sepsis. They are also at greater risk of autoimmune disease, some cancers and ischemic cardiac disease. Enhanced mortality is attributed to lack of phagocytes sequestered in spleen that efficiently eliminate appropriate targets, though related cells can be found at other sites. Although the overall organization of splenic white pulp is similar to that of lymph nodes, spleen is not connected to the lymphatic system. Instead, pathogens, RBCs, apoptotic/necrotic, altered and tumor cells all are delivered to and leave the spleen in blood. In contrast to white pulp, splenic red pulp serves as both a reservoir and a filter that determines whether cells coursing therein are retained, destroyed or returned to the circulation. The human venous sinus-lining cell, known as the littoral cell (LC), comprises ~30% of red pulp. Increasing evidence indicates these highly specialized splenocytes are the major determinant of whether RBCs and others are destroyed, retained or returned to the circulation, though the mechanisms that regulate their function are entirely unknown. Others and we showed that LCs are not a classical endothelial cells as they bear multiple macrophage and smooth muscle-associated antigens, are CD8+ and lack traditional markers of endothelial and also hematopoietic lineage. So, why is so little known about LC function? The acquisition, purification and culture of LCs have historically proved very difficult. Based on our recent success in obtaining fresh spleen as well as identifying markers that distinguish LCs from other splenocytes, we are now prepared to further isolate and profile LC gene/protein expression (AIM I). Although cells such as LCs that form specialized structures within complex organs are often hard to propagate ex vivo, overcoming these limitations would represent a significant advance in understanding LC function(s). We have assembled a group of expert endothelial cell biologist-immunologists, and pathologists who will provide us with in depth guidance as we develop diverse LC culture systems (AIM II) that are both physiologically relevant and tractable. These methods will enable discovery of the mechanisms that underlie LC function and lead to development of therapeutics able to modulate splenic filtration in different disease states (e.g. sepsis, malaria, hemoglobinopathies and others).