Focus on collagen: in vitro systems to study fibrogenesis and antifibrosis state of the art.
Focus on collagen: in vitro systems to study fibrogenesis and antifibrosis state of the art.
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DOI：
10.1186/1755-1536-2-7
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发表时间：
2009-12-15
期刊：
影响因子：
--
通讯作者：
Raghunath M
中科院分区：
文献类型：
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作者：
Chen CZ;Raghunath M
纤维化是一个主要的全球性疾病负担，但有效的抗纤维化化合物仍然没有出现。造成这种情况的部分原因是，制药行业的学术实验室和研发部门在动物试验之前重新制定体外纤维化过程以进行筛选程序方面一直面临困难。抗纤维化化合物的有效体外表征受到细胞培养环境的阻碍，这些环境缺乏关键的辅因子，或者不是导致不溶性细胞周胶原基质形成的生物合成和沉积途径的整体代表。为了理解抗纤维化药物的体外筛选所面临的任务，我们将首先通过将其分类为胶原生物合成上游的事件和胶原I的实际生物合成和沉积级联来回顾纤维化过程。我们指出的疏忽，如省略维生素C，一个重要的辅因子的生产稳定的前胶原分子，以及鲜为人知的在体外缓慢的前胶原处理胶原蛋白C-蛋白酶/BMP-1，另一个原因是最小的胶原蛋白沉积在细胞培养。我们回顾了当前的细胞培养和胶原定量方法，以及针对细胞数量进行标准化以进行有意义数据检索的高含量选项和要求。只有当胶原蛋白形成了一个纤维状基质，成为交联，投资配体，并可以重塑和吸收，纤维化的全貌可以反映在体外。我们在这里展示如何实现这一点。一个经过深思熟虑的体外纤维化系统代表了蛮力化学库筛选和合理的动物实验之间缺失的环节，从而为开发更好的抗纤维化药物提供了成本效益和简化的程序。
Fibrosis represents a major global disease burden, yet a potent antifibrotic compound is still not in sight. Part of the explanation for this situation is the difficulties that both academic laboratories and research and development departments in the pharmaceutical industry have been facing in re-enacting the fibrotic process in vitro for screening procedures prior to animal testing. Effective in vitro characterization of antifibrotic compounds has been hampered by cell culture settings that are lacking crucial cofactors or are not holistic representations of the biosynthetic and depositional pathway leading to the formation of an insoluble pericellular collagen matrix. In order to appreciate the task which in vitro screening of antifibrotics is up against, we will first review the fibrotic process by categorizing it into events that are upstream of collagen biosynthesis and the actual biosynthetic and depositional cascade of collagen I. We point out oversights such as the omission of vitamin C, a vital cofactor for the production of stable procollagen molecules, as well as the little known in vitro tardy procollagen processing by collagen C-proteinase/BMP-1, another reason for minimal collagen deposition in cell culture. We review current methods of cell culture and collagen quantitation vis-à-vis the high content options and requirements for normalization against cell number for meaningful data retrieval. Only when collagen has formed a fibrillar matrix that becomes cross-linked, invested with ligands, and can be remodelled and resorbed, the complete picture of fibrogenesis can be reflected in vitro. We show here how this can be achieved. A well thought-out in vitro fibrogenesis system represents the missing link between brute force chemical library screens and rational animal experimentation, thus providing both cost-effectiveness and streamlined procedures towards the development of better antifibrotic drugs.