Laser capture microdissection and genetic analysis of carbon-labeled Kupffer cells
Laser capture microdissection and genetic analysis of carbon-labeled Kupffer cells作者机构：Department of Medicine Rhode Island Hospital and The Warren Alpert Medical School of Brown University Providence RI 02903 United States Department of Pathology Rhode Island Hospital and The Warren Alpert Medical School of Brown University Providence RI 02903 United States zentrum fur Kinder- und Jugendmedizin der Universitaitsmedizin der Johannes Gutenberg-Universitait 55101 Mainz Germany
出 版 物：《World Journal of Gastroenterology》 (世界胃肠病学杂志（英文版）)
年 卷 期：2009年第15卷第14期
页 面：1708-1718页
核心收录：
学科分类：1002[医学-临床医学] 100201[医学-内科学(含：心血管病、血液病、呼吸系病、消化系病、内分泌与代谢病、肾病、风湿病、传染病)] 10[医学]
基 金：Supported by NIH Grant DK068097 funds provided by Rhode Island Hospital the Deutsche Forschungsgemeinschaft grant (DFG) grant GE1193/1-1 NIH COBRE Award (RR-P20 RR17695)
主 题：Kupffer cells India ink Laser capturemicrodissection Bile duct ligation DNA microarray
摘 要：AIM： To develop a method of labeling and microdissecting mouse Kupffer cells within an extraordinarily short period of time using laser capture microdissection （LCM）. METHODS： Tissues are complex structures comprised of a heterogeneous population of interconnected cells. LCM offers a method of isolating a single cell type from specific regions of a tissue section. LCM is an essential approach used in conjunction with molecular analysis to study the functional interaction of cells in their native tissue environment. The process of labeling and acquiring cells by LCM prior to mRNA isolation can be elaborate, thereby subjecting the RNA to considerable degradation. Kupffer cell labeling is achieved by injecting India ink intravenously, thus circumventing the need for in vitro staining. The significance of this novel approach was validated using a cholestatic liver injury model. RESULTS： mRNA extracted from the microdissected cell population displayed marked increases in colonystimulating factor-1 receptor and Kupffer cell receptor message expression, which demonstrated Kupffer cell enrichment. Gene expression by Kupffer ceils derived from bile-duct-ligated, versus sham-operated, mice was compared. Microarray analysis revealed a significant （2.5-fold, q value 〈 10） change in 493 genes. Based on this fold-change and a standardized PubMed search, 10 genes were identified that were relevant to the ability of Kupffer cells to suppress liver injury. CONCLUSION; The methodology outlined herein provides an approach to isolating high quality RNA from Kupffer cells, without altering the tissue integrity.
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