Chong Tae W, Smith Robert L, Hughes Michael G, Camden Jeremy, Rudy Christine K, Evans Heather L, Sawyer Robert G, Pruett Timothy L
Department of Surgery, Surgical Infectious Disease Laboratory, University of Virginia, Charlottesville, Virginia 22908, USA.
J Surg Res. 2006 Jan;130(1):52-7. doi: 10.1016/j.jss.2005.04.043. Epub 2005 Sep 8.
Hepatitis C (HCV) is a worldwide health problem, affecting nearly 170 million people. Current models for studying Hepatitis C have focused primarily on the use of poorly permissive cell lines and viral constructs, because of the lack of a suitable animal model or an in vitro system for studying functional infection. As hepatocytes are the primary reservoir for the virus in vivo, we report on a model using primary human hepatocytes cultured in spheroid formation.
The hepatocytes were harvested from uninfected liver resections and cultured as spheroids (that promotes a differentiated phenotype) or monolayers. Spheroids expressed the putative receptors CD81 and human scavenger receptor B1 in a variable pattern throughout the culture period. Samples were inoculated with infectious HCV serum, and HCV RNA was detected using RT-PCR. RNA was detected in the cells and culture medium by 3 days and 5 days after inoculation, respectively. Selection of HVR1 variants occurred in a differential pattern based on culture technique, suggesting that viral selection was dependent on host phenotype. Detection of NS5A by Western blot analysis of infected samples and immunofluorescence for HCV core protein was seen only in infected spheroids.
The use of spheroid formation to study Hepatitis C is associated with the establishment of HVR1 selection and functional infection. This represents a promising alternative model to study Hepatitis C.
丙型肝炎（HCV）是一个全球性的健康问题，影响着近1.7亿人。由于缺乏合适的动物模型或用于研究功能性感染的体外系统，目前用于研究丙型肝炎的模型主要集中在使用低允许性细胞系和病毒构建体上。由于肝细胞是体内病毒的主要储存库，我们报道了一种使用培养成球体的原代人肝细胞的模型。
从未感染的肝脏切除术中获取肝细胞，并培养成球体（促进分化表型）或单层细胞。在整个培养期间，球体以可变模式表达推定受体CD81和人清道夫受体B1。用感染性HCV血清接种样品，并使用RT-PCR检测HCV RNA。接种后3天和5天分别在细胞和培养基中检测到RNA。基于培养技术，HVR1变体的选择以不同模式发生，这表明病毒选择取决于宿主表型。仅在感染的球体中通过对感染样品的蛋白质印迹分析检测到NS5A，并通过免疫荧光检测到HCV核心蛋白。
使用球体形成来研究丙型肝炎与HVR1选择的建立和功能性感染有关。这代表了一种有前途的研究丙型肝炎的替代模型。
