近日，英国科研人员开发出一种独特的实验室细胞和组织培养技术，其培养条件与人体内环境很相似，相关论文发表在近日出版的《解剖学杂志》上。该技术利用一种塑料支架，使细胞在一种更接近体内细胞生长环境的三维环境中生长。而此前，细胞培养均在培养皿上进行。科研人员还发现，利用这个系统，能够更有效地进行药物研发，并减少一些不必要的动物实验。          

    这项技术的专利权由英国杜伦大学的研究人员及其附属的ReInnervate有限公司持有。该研究小组获得的大量证据显示，这是一种成本低、操作简单的三维细胞培养方式。          

    在药物研发中，很大一部分药物往往在检测阶段宣告失败，从而造成数百万英镑的成本损失。因为大多数药物首先在二维培养的细胞中进行检测，但是人体形成组织和细胞生长则是以一种更为复杂的三维方式进行的。          

    在药物研发领域，肝脏细胞常常用于检测药物的毒性。这项新的研究检测了一种名为MTX的癌症药物对三维和二维培养的肝脏细胞的毒性作用，结果表明，MTX在高剂量时能导致肝脏损伤。          

    检测显示，使用三维支架培养的细胞结构和特征与人体中自然生长的肝脏细胞最相似。当接触到MTX时，生长在二维培养条件下的细胞在极低剂量下就会死亡，而生长在三维支架上的细胞则抵抗力更强，并能更准确地反映出人体内细胞在接触相似药物剂量时的行为表现。此外，研究人员还检测了10种在由多孔聚苯乙烯材料构成支架上培养的不同组织，包括骨骼、肝脏、脂肪和骨髓干细胞。          

    研究人员发现，这项技术能培养用于药物研发的人类干细胞。它的使用可以减少动物实验。此前，美国一所大学的研究人员证实，他们设计的一种新的成体干细胞培养方法能使成体干细胞高效长成角膜干细胞，研究证实，这种培养技术已成功用于治疗70只兔子的角膜疾病。该研究的目的是希望能够修复受损的上皮细胞并因此恢复角膜功能。这项研究同时证实，利用健康眼睛的角膜干细胞治疗人的角膜病变是可行的。          

    据了解，该项技术目前正被用于人类患者，并获得满意结果。相关论文描述了一种新的细胞培养方法，并证实了这个过程的临床应用价值。该项研究还显示，源于活组织切片样本的新培养技术能使干细胞的数量增加，从而获得足够用于有效治疗的细胞，细胞样本可以从健康眼睛结构中获得。          

    将塑料芯片与一种羊膜芯片结合起来后，这种培养方法能使获得的细胞保持其特征。其新颖之处集中在使用塑料芯片的第一阶段：将从健康眼睛获得的组织片段分割成更小的片段，以便在芯片上培养。          

    组织片段的数量越多就能获得更多数量的干细胞。然后，将获得的细胞样本送到解剖病理学实验室进行细胞的可用性和治疗鉴定。之后，将这些细胞转移到羊膜上继续培养，这种膜很适合培养用于眼睛再生治疗的干细胞移植。          

    一旦放在羊膜上，这些干细胞能以均一的方式扩散，从而确保更好的细胞等同性，以便选择最适合用于治疗的单元。这种方法能精确找到用于眼睛细胞移植的细胞群，保证了被移植细胞的质量和数量。（科学时报）

原始出处:

Journal of Anatomy 211 (4), 567–576.
Volume 211 Issue 4 Page 567-576, October 2007

doi:10.1111/j.1469-7580.2007.00778.x

Culture of HepG2 liver cells on three dimensional polystyrene scaffolds enhances cell structure and function during toxicological challenge

• Maria Bokhari 1,31School of Biological and Biomedical Science, Durham University, South Road, Durham, DH1 3LE, UK3ReInnervate Limited, School of Biological and Biomedical Science, Durham University, South Road, Durham DH1 3LE, UK,
• Ross J. Carnachan 1,21School of Biological and Biomedical Science, Durham University, South Road, Durham, DH1 3LE, UK2IRC in Polymer Science and Technology, Department of Chemistry, Durham University, South Road, Durham DH1 3LE, UK,
• Neil R. Cameron 2,32IRC in Polymer Science and Technology, Department of Chemistry, Durham University, South Road, Durham DH1 3LE, UK3ReInnervate Limited, School of Biological and Biomedical Science, Durham University, South Road, Durham DH1 3LE, UK and
• Stefan A. Przyborski 1,31School of Biological and Biomedical Science, Durham University, South Road, Durham, DH1 3LE, UK3ReInnervate Limited, School of Biological and Biomedical Science, Durham University, South Road, Durham DH1 3LE, UK

• 1School of Biological and Biomedical Science, Durham University, South Road, Durham, DH1 3LE, UK2IRC in Polymer Science and Technology, Department of Chemistry, Durham University, South Road, Durham DH1 3LE, UK3ReInnervate Limited, School of Biological and Biomedical Science, Durham University, South Road, Durham DH1 3LE, UK
  CorrespondenceStefan A. Przyborski, Reader in Stem Cell Biology, Director and Chief Scientific Officer of Relnnervate Limited, School of Biological and Biomedical Science, University of Durham, South Road, Durham DH1 3LE, UK. T: +44 (0)191 3341341; F: +44 (0)191 3341201; E: stefan.przyborski@durham.ac.uk
  Key words 3-D cell growth; cell culture; drug screening; function; HepG2 cells; liver; polymer scaffold; toxicity; ultra-structure.

Cultured cells are dramatically affected by the micro-environment in which they are grown. In this study, we have investigated whether HepG2 liver cells grown in three dimensional (3-D) cultures cope more effectively with the known cytotoxic agent, methotrexate, than their counterparts grown on traditional two dimensional (2-D) flat plastic surfaces. To enable 3-D growth of HepG2 cells in vitro, we cultured cells on 3-D porous polystyrene scaffolds previously developed in our laboratories. HepG2 cells grown in 3-D displayed excellent morphological characteristics and formed numerous bile canaliculi that were seldom seen in cultures grown on 2-D surfaces. The function of liver cells grown on 3-D supports was significantly enhanced compared to activity of cells grown on 2-D standard plasticware. Unlike their 2-D counterparts, 3-D cultures were less susceptible to lower concentrations of methotrexate. Cells grown in 3-D maintained their structural integrity, possessed greater viability, were less susceptible to cell death at higher levels of the cytotoxin compared to 2-D cultures, and appeared to respond to the drug in a manner more comparable to its known activity in vivo. Our results suggest that hepatotoxicity testing using 3-D cultures might be more likely to reflect true physiological responses to cytotoxic compounds than existing models that rely on 2-D culture systems. This technology has potential applications for toxicity testing and drug screening.