Giraudi Pablo J, Becerra Varenka J Barbero, Marin Veronica, Chavez-Tapia Norberto C, Tiribelli Claudio, Rosso Natalia
Fondazione Italiana Fegato, Centro Studi Fegato, Area Science Park Basovizza Bldg. Q SS14 Km 163.5, 34149 Trieste, Italy.
Fondazione Italiana Fegato, Centro Studi Fegato, Area Science Park Basovizza Bldg. Q SS14 Km 163.5, 34149 Trieste, Italy; Liver Research Unit, Médica Sur Clinic & Foundation, Puente de Piedra 150, Col. Toriello Guerra, Tlalpan, C.P. 14050 Mexico City, Mexico.
Exp Mol Pathol. 2015 Feb;98(1):85-92. doi: 10.1016/j.yexmp.2014.12.006. Epub 2014 Dec 20.
BACKGROUND & AIMS: Non-alcoholic fatty liver disease is characterized by an initial accumulation of triglycerides that can progress to non-alcoholic steatohepatitis, which can ultimately evolve to cirrhosis and hepatocellular carcinoma. Hepatic stellate cells play a key role in liver fibrogenesis by an increased activation and an altered profile of genes involved in the turnover of extracellular matrix components. To reproduce in-vitro the functional cell connections observed in vivo it is essential to consider cell-to-cell proximity and interaction. The aim of this study was to determine the response to free fatty acids in a simultaneous co-culture model of hepatocytes and hepatic stellate cells.
Simultaneous co-culture model and monoculture of each cell type (control) were exposed to FFA for 24 up to 144 h. Quantification of steatosis; stellate cell activation; assessment of fibrogenic response; expression and activity of metalloproteinases as well as collagen biosynthesis were evaluated.
Free fatty acids induced comparable steatosis in simultaneous co-culture and monoculture. However, the activation of the stellate cells assessed by alpha-smooth muscle actin expression is greater when cells were in close contact. Furthermore, a time-dependent increment of tissue inhibitor metalloproteinase-2 protein was observed, which was inversely correlated with protein expression and activity of matrix-metalloproteinases, suggesting enhanced collagen biosynthesis. This behavior was absent in cell monoculture.
These data indicate that cell-to-cell proximity between hepatocytes and stellate cells is necessary for the initiation of the fibrotic process.
非酒精性脂肪性肝病的特征是甘油三酯最初蓄积，可进展为非酒精性脂肪性肝炎，最终可演变为肝硬化和肝细胞癌。肝星状细胞通过增加激活以及参与细胞外基质成分周转的基因谱改变，在肝纤维化形成中起关键作用。为了在体外重现体内观察到的功能性细胞连接，考虑细胞间的接近度和相互作用至关重要。本研究的目的是确定在肝细胞和肝星状细胞同步共培养模型中对游离脂肪酸的反应。
将同步共培养模型和每种细胞类型的单培养物（对照）暴露于游离脂肪酸24至144小时。评估脂肪变性的量化；星状细胞激活；纤维化反应评估；金属蛋白酶的表达和活性以及胶原蛋白生物合成。
游离脂肪酸在同步共培养和单培养中诱导了相当的脂肪变性。然而，当细胞紧密接触时，通过α-平滑肌肌动蛋白表达评估的星状细胞激活程度更高。此外，观察到组织金属蛋白酶抑制剂-2蛋白呈时间依赖性增加，这与基质金属蛋白酶的蛋白表达和活性呈负相关，提示胶原蛋白生物合成增强。这种现象在细胞单培养中不存在。
这些数据表明，肝细胞和星状细胞之间的细胞间接近度是纤维化过程起始所必需的。
