Modulation of limbal niche stiffness to regulate stem cell differentiation

调节角膜缘生态位硬度以调节干细胞分化

• 批准号: MR/K017217/1
• 负责人: Che Connon
• 金额: $ 70.82万
• 依托单位: University of Reading
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2013
• 资助国家: 英国
• 起止时间: 2013 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=MR%2FK017217%2F1
• 关键词: Modulation; limbal; niche; stiffness; regulate

Tissue engineering is a newly emerging biomedical methodology to assist and accelerate the regeneration of defective and damaged tissues based largely on the natural healing potentials of stem cells. For this new therapeutic strategy, it is indispensable to provide cells with a local environment (niche) that enhances and regulates their proliferation and differentiation. In this regard, biomaterial technology currently leads the way in trying to fulfil these requirements by recreating artificial three-dimensional environments. However, understanding how, why and in what environment these cells differentiate in a lineage-specific manner are essential for tissue engineering and stem cell therapy. We believe that this underlying biology (i.e. specific signals or stimuli which make the cells respond) has not yet been properly investigated and that this is likely to undermine current attempts to recreate the stem cell niche using biomaterials. Therefore, we plan to understand the role substrate stiffness plays in maintaining the corneal stem cell niche (the home for cells required to cover the surface of the cornea). Our expertise in ocular surface models means we are well placed to investigate the changes in gene and protein expression associated with substrate stiffness dependent (mechanosensitive) differentiation. Together these investigations will feed directly into the field of tissue engineering and our own on-going MRC, BBSRC and EPSRC research into ocular surface biomaterials and corneal stem cell transplantation. Acoustically induced inelastic light scattering (Brillouin microscopy) allows non-contact, direct readout of the mechanical properties of a material and has been used successfully to measure, at microscopic resolution, the biomechanical properties of the cornea and lens of the human eye. We plan to employ this sophisticated method to measure stiffness across the surface of normal and disease/wounded corneas producing high resolution maps of the tissues' mechanical properties. We will then seek to affect localised stiffness using an enzyme (collagenase) to break up the corneas main structural protein (collagen) in a highly controlled manner. The enzyme will be delivered by a gel that sticks to the cornea to allow precise positioning of the enzyme and a corresponding localised effect on stiffness. We have shown previously that corneal stem cells differentiate when grown on stiff collagen gels and that collagenase can reduce this stiffness with a proportional reduction in differentiated cells. Therefore we plan to investigate a new pre-corneal stem cell transplantation technique. In short, having validated that the corneal stem cell niche is sensitive to changes in its stiffness we will develop a method to control its stiffness prior to corneal stem cell transplantation with the belief that this will result in an improved residency time for the transplanted cells in an undifferentiated form. The predicted outcome being a significantly improved success rate for corneal stem cell transplantation. Recently the overall clinical success rate of corneal stem cell transplantation has remained static at approx. 75% with half of those able to discern two lines or more of symbols on an eye chart. Whilst this represents a remarkable surgical achievement there remains considerable room for improvement. We believe restoring the corneal tissues mechanical properties prior to stem cell transplantation represents a significant surgical enhancement.

组织工程是一种新出现的生物医学方法，可在很大程度上基于干细胞的自然愈合潜力来协助和加速有缺陷和受损的组织的再生。对于这种新的治疗策略，必须为细胞提供局部环境（小裂），从而增强和调节其增殖和分化是必不可少的。在这方面，生物材料技术目前通过重新创建人工的三维环境来实现这些要求。但是，了解这些细胞以谱系特异性方式分化的环境如何，为什么以及在什么环境中对于组织工程和干细胞疗法至关重要。我们认为，这种潜在的生物学（即使细胞响应的特定信号或刺激）尚未得到适当研究，并且这很可能破坏了使用生物材料来重现干细胞生态位的尝试。因此，我们计划了解底物刚度在维持角膜干细胞生态位的作用（覆盖角膜表面所需的细胞的家园）。我们在眼部表面模型中的专业知识意味着我们可以很好地研究与底物刚度依赖性（机械敏感）分化相关的基因和蛋白质表达的变化。这些研究将共同​​进行组织工程领域，以及我们自己正在进行的MRC，BBSRC和EPSRC对眼表面生物材料和角膜干细胞移植的研究。声学诱导的非弹性散射（布里渊显微镜）允许直接读出材料的机械性能，并已成功地用于测量微观分辨率，即角膜的生物力学特性和人眼的生物力学特性。我们计划采用这种复杂的方法来测量正常和疾病/受伤角膜表面的刚度，从而产生组织机械性能的高分辨率图。然后，我们将寻求使用酶（胶原酶）以高度控制的方式破坏角膜主结构蛋白（胶原蛋白）。酶将通过粘附在角膜上的凝胶递送，以允许酶的精确定位以及对刚度的相应局部作用。我们先前已经证明，在刚性胶原蛋白凝胶上生长时，角膜干细胞会分化，并且胶原酶可以通过分化细胞比例减少而降低这种刚度。因此，我们计划研究一种新的培养基干细胞移植技术。简而言之，在验证了角膜干细胞生态位对其刚度的变化敏感后，我们将开发一种在角膜干细胞移植之前控制其刚度的方法，并认为这会导致改善的居住时间，使得在居住时间改善了将细胞移植的时间一种未分化的形式。预测的结果是角膜干细胞移植的成功率显着提高。最近，角膜干细胞移植的总体临床成功率在大约上保持静态。 75％的人中有一半能够在眼图上辨别两条或更多符号的人。尽管这代表了一个了不起的手术成就，但仍有很大的改进空间。我们认为，在干细胞移植之前恢复角膜组织机械性能代表了显着的手术增强。

项目成果

Controlling the 3D architecture of Self-Lifting Auto-generated Tissue Equivalents (SLATEs) for optimized corneal graft composition and stability.

• DOI: 10.1016/j.biomaterials.2016.12.023
• 发表时间: 2017-03
• 期刊: Biomaterials
• 影响因子: 14
• 作者: Gouveia RM;González-Andrades E;Cardona JC;González-Gallardo C;Ionescu AM;Garzon I;Alaminos M;González-Andrades M;Connon CJ
• 通讯作者: Connon CJ

Low-glucose enhances keratocyte-characteristic phenotype from corneal stromal cells in serum-free conditions.

• DOI: 10.1038/srep10839
• 发表时间: 2015-06-03
• 期刊: Scientific reports
• 影响因子: 4.6
• 作者: Foster JW;Gouveia RM;Connon CJ
• 通讯作者: Connon CJ

Oxidized alginate hydrogels as niche environments for corneal epithelial cells.

• DOI: 10.1002/jbm.a.35011
• 发表时间: 2014-10
• 期刊: JOURNAL OF BIOMEDICAL MATERIALS RESEARCH PART A
• 影响因子: 4.9
• 作者: Wright, Bernice;De Bank, Paul A.;Luetchford, Kim A.;Acosta, Fernando R.;Connon, Che J.
• 通讯作者: Connon, Che J.

Application of retinoic acid improves form and function of tissue engineered corneal construct.

• DOI: 10.1080/15476278.2015.1093267
• 发表时间: 2015
• 期刊: Organogenesis
• 影响因子: 2.3
• 作者: Abidin FZ;Gouveia RM;Connon CJ
• 通讯作者: Connon CJ