Detection and characterization of cell type-specific extracellular vesicles in obesity-driven hepatocellular carcinoma

项目摘要 肥胖症在世界范围内的流行已达到大流行的程度。脂肪与 积累或肝脂肪变性和肝细胞癌（HCC）的发展早已为人所知。 肥胖引起的脂肪肝及其更严重的并发症非酒精性脂肪性肝炎 NASH（NASH），被归类为非酒精性脂肪性肝病（NAFLD），影响高达40%的美国人口。 基于相关研究和实验室研究，已经提出了几种机制来解释肥胖如何 增加癌症风险。一个重要的发现，说明了肥胖的肿瘤促进作用是低- 分级，异常炎症反应，导致细胞因子，如TNF α的产生增加 和IL-6小鼠模型的研究表明，肥胖促进的HCC发展依赖于 增加这些炎症细胞因子的产生。然而，关键问题是， 肥胖症中的异常炎症的起始尚未被明确定义。 我们假设，在肥胖症，细胞外囊泡（EV）来源于肝细胞成为致病 并驱使受体免疫细胞，如嗜中性粒细胞和单核细胞/巨噬细胞，向异常 与HCC发展相关的炎症。为了证明这一假设，我们将建立两个 新系统; A）新的小鼠系，其中特异性细胞/组织来源的EV被选择性标记为 绿色荧光蛋白（GFP），和B）一种简单但功能强大的基于电动的微型装置， 基于EV的介电性质，从生物流体中以高纯度和高收率快速提取EV。与这些 系统，我们的目标是 1)确定H-EV是否成为致病性并诱导导致HCC的异常炎症，以及 2)通过新型介电泳（DEP）技术分离H-EV，并评估其亲 炎症特征 这项研究将使我们证明肝细胞来源的EV的致病性，这可能提供 HCC发展的新机制和该疾病的EV生物标志物。

PROJECT SUMMARY The worldwide prevalence of obesity has reached pandemic proportions. An association between fat accumulation or hepatosteatosis and hepatocellular carcinoma (HCC) development has been long known. Obesity-induced hepatosteatosis, together with its more severe complication nonalcoholic steatohepatitis (NASH), classified as the nonalcoholic fatty liver disease (NAFLD) affects up to 40% of the US population. Based on correlative and bench studies, several mechanisms have been proposed to explain how obesity increase cancer risk. An important finding that accounts for the tumor-promoting effect of obesity is the low- grade, aberrant inflammatory response, which results in the elevated production of cytokines, such as TNF and IL-6. Studies with mouse models demonstrated that obesity-promoted HCC development is dependent on the enhanced production of these inflammatory cytokines. However, critical questions concerning how the aberrant inflammation in obesity is initiated have been yet to be clearly defined. We hypothesize that in obesity, extracellular vesicles (EVs) derived from hepatocytes become pathogenic and drive recipient immune cells, such as neutrophils and monocytes/macrophages, towards abnormal inflammation associated with the development of HCC. To demonstrate this hypothesis, we will establish two novel systems; A) a new mouse line in which specific cell/tissue-derived EVs are selectively labeled with green fluorescent protein (GFP), and B) a simple, yet powerful electrokinetic-based micro-device that can rapidly extract EVs based on their dielectric properties from biofluids with high purity and yield. With these systems, we aim to 1) determine if H-EVs become pathogenic and induce aberrant inflammation leading to HCC, and 2) isolate H-EVs through the novel dielectrophoretic (DEP) technology and assess their pro- inflammatory trait. This study will lead us to demonstrate the pathogenicity of hepatocyte-derived EVs, which potentially provides a novel mechanism for the development of HCC and the EV biomarker for the disease.