Opposing RNAi Molecule Gradient Constructs to Repair Osteochondral Defects

相反的 RNAi 分子梯度构建修复骨软骨缺损

基本信息

批准号：
9728716
负责人：
Eben Alsberg
金额：
$ 35.57万
依托单位：
UNIVERSITY OF ILLINOIS AT CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-04-01 至 2022-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9728716
关键词：
Address Affect Animal Model Biochemical Biocompatible Materials Biological Assay Biomechanics Biopolymers Bone Marrow Bone Morphogenetic Proteins Cartilage Cell Transplantation Cells Charge Chondrogenesis Clinical Defect Degenerative polyarthritis Dextrans Dimensions Disease Dose Encapsulated Engineering Gel Gene Expression Gene Silencing Genes Genetic Transcription Growth Factor Histologic Human Hydrogels Injury Joints Knee Laboratories Lead Mechanics Mesenchymal Stem Cells Messenger RNA MicroRNAs Microfluidic Microchips Modeling Molecular Morphology Natural regeneration Non-Viral Vector Oryctolagus cuniculus Osteogenesis Pain Pathway interactions Patients Polyethyleneimine Process Proteins Quality of life RNA RNA Interference RNA delivery Reporter Genes Small Interfering RNA Source Stains System Technology Testing Therapeutic Tissue Engineering Tissues Transfection Work articular cartilage bone clinical translation crosslink density design disability healing implantation improved in vivo knock-down microfluidic technology osteochondral repair osteochondral tissue osteogenic public health relevance recruit repaired scaffold spatiotemporal stem cell differentiation subchondral bone tissue regeneration tool

项目摘要

DESCRIPTION: The treatment of osteochondral defects (OCDs), which involve damage to both the subchondral bone and articular cartilage in the affected joint, is challenging. Such debilitating defects lead to mechanical instability, pain and worsening osteoarthritic degeneration. Current therapies fail to consistently repair and restore tissue function. Osteochondral tissue engineering technology utilizing biomaterials in combination with recruited and/or transplanted cells, and/or bioactive factors has emerged as a promising alternative approach. Human mesenchymal stem cells (hMSCs) are an attractive cell source as they can easily be isolated from bone marrow, expanded in culture without losing multipotency, and under appropriate conditions can differentiate into cells of the osteogenic and chondrogenic lineages. RNA interference (RNAi) is a powerful tool permitting inhibition of gene expression at the post-translational level by the targeted destruction of specific mRNA molecules, and has the potential to revolutionize the functional repair of damaged tissue by decreasing the expression of specific proteins that negatively impact healing processes or by altering stem cell differentiation pathways. Importantly, RNAi molecules have been identified that can promote the osteogenic and chondrogenic differentiation of hMSCs. However, effective delivery of RNAi molecules to target cells in vivo remains a significant challenge limiting its therapeutic potentia. We have engineered biopolymer hydrogels capable of locally delivering bioactive RNAi molecules with tailorable release profiles for delivery to surrounding and encapsulated cells, and these gels have been used to spatially and temporally control cell gene expression and fate. Therefore, the central hypothesis of this application is that the controlled spatial and temporal presentation of dual opposing RNAi molecule gradients in a biopolymer hydrogel will drive osteogenesis and chondrogenesis of encapsulated hMSCs in opposite directions to form osteochondral constructs that can promote the healing of OCDs. This will be addressed by the following specific aims: (1) Engineer biopolymer hydrogels with opposing concentration gradients of two different siRNAs for spatiotemporally controlled, sustained gene knockdown, (2) Deliver RNAi molecules that promote osteogenesis and chondrogenesis from biopolymer gradient hydrogels and investigate their capacity to spatially guide the osteogenic and chondrogenic differentiation of encapsulated hMSCs, (3) Develop opposing RNAi molecule gradient hydrogels with tailorable dimensions using microfluidic technology, and (4) Assess the ability of the hydrogel constructs containing hMSCs and opposing RNAi molecule gradients to drive osteogenesis and chondrogenesis in vivo upon implantation into a rabbit OCD model. This application aims to demonstrate the utility of a new tissue engineering approach for enhanced osteochondral tissue regeneration, which would have great clinical utility by improving the quality of life of patients suffering from OCDs.

描述：骨软骨缺损（OCD）的处理是挑战。这种衰弱的缺陷导致机械不稳定性，疼痛和令人担忧的骨关节炎变性。当前的疗法无法始终如一地修复和恢复组织功能。骨软骨组织工程技术利用生物材料与招募和/或移植细胞结合使用，并且/或生物活性因素已成为有望的替代方法。人间充质干细胞（HMSC）是一个有吸引力的细胞来源，因为它们可以轻松地从骨髓中分离出来，在培养中扩展而不会失去多重性，并且在适当的条件下可以区分成骨和软骨源的细胞。 RNA干扰（RNAI）是一种强大的工具，可以通过针对特定的mRNA分子的靶向破坏来抑制基因表达在翻译后水平上，并且有可能通过降低特定蛋白质的表达来革新受损组织的功能修复，从而通过更改愈合过程或通过改变干细胞分化的影响来降低特定蛋白的表达。重要的是，已经鉴定出可以促进HMSC的成骨和软骨分化的RNAi分子。但是，在体内有效递送RNAi分子向靶细胞递送仍然是限制其治疗潜力的重大挑战。我们已经设计了能够在局部传递具有可定制释放曲线的生物活性RNAi分子的生物聚合物水凝胶，以递送到周围和封装的细胞，并且这些凝胶已用于空间和临时控制细胞基因的表达和命运。因此，该应用的中心假设是，在生物聚合物水凝胶中对双重对立的RNAi分子梯度的受控空间和临时表示将驱动成骨和封装的HMSC的软骨形成，以相反的方向形成骨软骨构建体，以促进OCD的愈合。 This will be addressed by the following specific aims: (1) Engineer biopolymer hydrogels with opposing concentration gradients of two different siRNAs for spatially controlled, sustained gene knockdown, (2) Deliver RNAi molecules that promote osteogenesis and chondrogenesis from biopolymer gradient hydrogels and investigate their capacity to spatially guide the osteogenic and chondrogenic differentiation of encapsulated hMSCs, （3）使用微流体技术开发具有可定制尺寸的相对RNAi分子梯度水凝胶，并且（4）评估含有HMSC的水凝胶构建体的能力，以及在植入Rabbit OCD模型中植入Vivo中的成骨和软骨发生的RNAi分子梯度。该应用旨在证明一种新的组织工程方法的实用性，以增强骨软骨组织再生，这将通过改善患有OCD的患者的生活质量而具有很大的临床效用。