Department of Biosciences and Bioengineering, Indian Institute of Technology Guwahati, Guwahati 781039, Assam, India.
Centre for Nanotechnology, Indian Institute of Technology Guwahati, Guwahati 781039, Assam, India.
ACS Appl Mater Interfaces. 2022 Mar 2;14(8):10167-10186. doi: 10.1021/acsami.2c00312. Epub 2022 Feb 16.
Bioengineering an in vitro liver model recapitulating the native liver microarchitecture consisting of parenchymal and non-parenchymal cells is crucial in achieving cellular crosstalk and hepatic metabolic functions for accurate hepatotoxicity prediction. Bioprinting holds the promise of engineering constructs with precise control over the spatial distribution of multiple cells. Two distinct tissue-specific liver extracellular matrix (ECM)-based bioinks with excellent printability and rheological attributes are formulated for supporting parenchymal and non-parenchymal cells. A physiologically relevant human vascularized liver model is bioprinted with a novel liver ECM-based bioink laden with human adipose mesenchymal stem cell-derived hepatocyte-like cells (HLCs), human umbilical vein endothelial cells (HUVECs), and human hepatic stellate cells (HHSCs) using an extrusion-based bioprinting approach and validated for hepatotoxicity assessment. The HLC/HUVEC/HHSC-laden liver model resembles native alternate cords of hepatocytes with a functional sinusoidal lumen-like network in both horizontal and vertical directions, demonstrating enhanced albumin production, urea synthesis, and cytochrome P450 (CPR) activity. Furthermore, the liver model is evaluated for drug toxicity assessment following 24 h exposure to different concentrations of (i) non-hepatotoxicants aspirin and dexamethasone, (ii) idiosyncratic hepatotoxicant trovafloxacin mesylate, and (iii) clinical hepatotoxicant acetaminophen and troglitazone. A follow-up cell viability and metabolic competence evaluation by estimating DNA concentration, lactate dehydrogenase activity, and CPR activity revealed a dose-dependent clinically relevant hepatotoxic response. These results corroborated that the developed clinically relevant vascularized liver model is affordable and would aid pharmaceutical companies in speeding up the drug development and provide a robust platform for hepatotoxicity screening.
生物工程体外肝脏模型，重现由实质细胞和非实质细胞组成的固有肝脏微结构，对于实现细胞串扰和肝脏代谢功能以准确预测肝毒性至关重要。生物打印有望对多种细胞的空间分布进行精确控制，从而构建工程结构。两种具有出色打印性能和流变特性的独特组织特异性肝脏细胞外基质（ECM）生物墨水被配方用于支持实质细胞和非实质细胞。使用基于挤出的生物打印方法，用新型富含人脂肪间充质干细胞衍生的肝细胞样细胞（HLC）、人脐静脉内皮细胞（HUVEC）和人肝星状细胞（HHSC）的基于肝脏 ECM 的生物墨水打印出具有生理相关性的人血管化肝脏模型，并进行肝毒性评估验证。HLC/HUVEC/HHSC 负载的肝脏模型类似于固有交替的肝细胞索，在水平和垂直方向上均具有功能性窦状腔样网络，表现出增强的白蛋白产生、尿素合成和细胞色素 P450（CPR）活性。此外，在 24 小时内以不同浓度（i）非肝毒性药物阿司匹林和地塞米松、（ii）特发性肝毒性药物甲磺酸曲伐沙星和（iii）临床肝毒性药物对乙酰氨基酚和曲格列酮对肝脏模型进行药物毒性评估。通过估计 DNA 浓度、乳酸脱氢酶活性和 CPR 活性进行后续细胞活力和代谢能力评估，显示出剂量依赖性的临床相关肝毒性反应。这些结果证实，所开发的具有临床相关性的血管化肝脏模型具有成本效益，将有助于制药公司加快药物开发，并为肝毒性筛选提供强大的平台。
