Abstract

Alcohol injury induces hepatic fibrosis which gradually progresses to cirrhosis, sometimes may lead to liver cancer. Animal models are less efficient in mimicking responses of human liver cells, whereas in vitro models discussed so far are majorly based on rodent cells. In this work, a coculture of primary human hepatocytes (PHHs) with LX-2 cells was established on the unmodified (C:F_0:0), collagen-I modified (C:F_1:0), fibronectin modified (C:F_0:1) and 3:1 collagen-I to fibronectin modified (C:F_3:1) 3D electrospun fibrous scaffolds. The effect of alcohol injury was evaluated on this cell-scaffold model at 0–40 μl/ml alcohol concentrations over 14 days of culture period by using the gold standard sandwich culture as the control. Among all the culture groups, C:F_3:1 scaffold was able to maintain translational and transcriptional properties of human liver cells at all concentrations of alcohol treatment. The study reveals that, PHHs on C:F_3:1 were able to maintain ~4-fold and ~1.6-fold higher secretion of albumin than the gold standard sandwich culture on Day 3 and Day 7, respectively. When treated with alcohol, at concentrations of 20 and 40 μl/ml, albumin secretion was also observed to be higher (~2-fold) when compared to the gold standard sandwich culture. Again as expected, in C:F_3:1 culture group on 40 μl/ml alcohol treatment, albumin gene expression decreased by ~2-fold due to alcohol toxicity, whereas CYP2C9, CYP3A4, CYP2E1 and CYP1A2 gene expressions upregulated by ~3.5, ~~4, ~5 and ~15-fold, respectively in response to the alcohol injury. LX-2 cells also acquire more quiescent phenotype on C:F_3:1 scaffolds when compared to the gold standard sandwich culture upon alcohol treatment. Thus, C:F_3:1 scaffold with human liver cells was established as the potential platform to scan alcohol toxicity at varied alcohol concentrations. Thus, it can pave a promising path not only to support functional healthy human liver cells for liver tissue engineering but also to examine potential drugs to study the progression or inhibition of alcoholic liver fibrosis in vitro.

中文翻译：

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生物工程 3D 电纺纳米纤维支架与人类肝细胞在体外研究酒精性肝病

摘要

酒精损伤导致肝纤维化，逐渐发展为肝硬化，有时可能导致肝癌。动物模型在模拟人类肝细胞的反应方面效率较低，而在体外到目前为止讨论的模型主要基于啮齿动物细胞。在这项工作中，原代人肝细胞 (PHHs) 与 LX-2 细胞的共培养建立在未修饰 (C:F_0:0)、胶原蛋白-I 修饰 (C:F_1:0)、纤连蛋白修饰 (C:F_0: 1) 和 3:1 胶原蛋白-I 到纤连蛋白修饰 (C:F_3:1) 3D 电纺纤维支架。通过使用金标准夹心培养物作为对照，在 0-40 μl/ml 酒精浓度下，在 0-40 μl/ml 酒精浓度下评估酒精损伤的影响。在所有培养组中，C:F_3:1 支架能够在所有浓度的酒精处理下维持人肝细胞的翻译和转录特性。该研究表明，C:F_3:1 上的 PHHs 能够维持 ~4 倍和 ~1。在第 3 天和第 7 天，白蛋白的分泌量分别是金标准夹心培养物的 6 倍。当用酒精处理时，在 20 和 40 μl/ml 的浓度下，与金标准夹心培养物相比，还观察到白蛋白分泌更高（~2 倍）。正如预期的那样，在 40 μl/ml 酒精处理的 C:F_3:1 培养组中，由于酒精毒性，白蛋白基因表达下降了 ~2 倍，而 CYP2C9、CYP3A4、CYP2E1 和 CYP1A2 基因表达上调了 ~3.5，~对酒精损伤的反应分别为~4、~5 和~15 倍。与酒精处理后的金标准夹心培养相比，LX-2 细胞还在 C:F_3:1 支架上获得更多的静止表型。因此，C:F_3：1 具有人类肝细胞的支架被建立为在不同酒精浓度下扫描酒精毒性的潜在平台。因此，它不仅可以为肝组织工程支持功能健康的人类肝细胞，还可以为研究酒精性肝纤维化的进展或抑制的潜在药物铺平道路。体外。