Regeneration of hyaline cartilage in situ with bone marrow mesenchymal stem cells

骨髓间充质干细胞原位再生透明软骨

• 批准号: 9206892
• 负责人: CONG-QIU CHU
• 金额: --
• 依托单位: PORTLAND VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-01-01 至 2019-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9206892
• 关键词: Adipose tissue; Adult; Affect; Alpha Cell; Animal Model; Animals; Antibodies; Autologous; Biological Assay; Biomechanics; Bone Marrow; Cartilage; Cartilage injury; Cell Differentiation process; Cell Proliferation; Cells; Characteristics; Charge; Chimeric Proteins; Chondrocytes; Chondrogenesis; Chronic; Clinical Trials; Collagen; Collagen Type I; Collagen Type II; Contralateral; Data; Defect; Degenerative polyarthritis; Delayed Hypersensitivity; Development; Ensure; Failure; Family; Fibrocartilages; Fluorescence; Future; Genes; HMG Domain; Histologic; Histology; Human; Hyaline Cartilage; Hybrids; Immune response; Immunohistochemistry; Impairment; In Situ; In Vitro; Knee; Knowledge; Lead; Lesion; Life; Measures; Mediating; Membrane; Mesenchymal Differentiation; Mesenchymal Stem Cells; Methods; Modeling; Molecular; Natural regeneration; Oryctolagus cuniculus; Outcome; Pathology; Pathway interactions; Phenotype; Prevention; Procedures; Process; Production; Property; Proteins; Reaction; Recruitment Activity; Regulation; Repression; Research; SOX9 protein; Serological; Serum; Signaling Protein; Site; Source; Synovial Membrane; Synovial joint; T cell response; T-Lymphocyte; Techniques; Tertiary Protein Structure; Testing; Therapeutic Effect; Time; Tissues; Transplantation; Treatment Efficacy; United States; Variant; aggrecan; articular cartilage; cartilage development; cartilage regeneration; cartilage repair; clinical development; cost; design; experimental study; functional improvement; immunogenicity; improved; improved outcome; in vivo; in vivo regeneration; lymphocyte proliferation; migration; minimally invasive; novel therapeutics; prevent; public health relevance; repaired; self-renewal; subchondral bone; success; therapy design; tissue repair; transcription factor

 DESCRIPTION (provided by applicant): Cartilage injury is common which can lead to progression of osteoarthritis (OA) which is characterized by progressive breakdown of articular cartilage, and ultimately leads to functional failure of synovial joints. Regeneration of cartilage has been an attractive approach to cartilage repair and treatment of OA. However, autologous chondrocyte transplantation suffers from insufficient cell supply, new damage to the donor sites and chondrocytes dedifferentiation during in vitro expansion. Mesenchymal stem cells (MSC) which reside in the bone marrow and many adult tissues are capable of self-renewal and differentiation into chondrocytes and are a potential source for cartilage in situ regeneration. Microfracture induces migration of bone marrow MSC to the site of cartilage defect and promotes fibrocartilage production. However, fibrocartilage is non-durable and functionally inadequate in the long-term. SOX9 is a transcription factor belonging to the SOX (Sry-type HMG box) family and has been identified as a "master regulator" of the chondrocyte phenotype. We have generated a cell penetrating, super positively charged SOX9 fusion protein (scSOX9). In vitro studies have demonstrated that scSOX9 protein induced bone marrow derived MSC proliferation and differentiation into chondrocytes. This was evident by decreased production of collagen type I and type X but increased collagen type II and aggrecan, which are characteristics of articular chondrocytes. In a 21 day culture, the initial one time addition of scSOX9 was sufficient to differentiate MSC into chondrocytes and maintain the chondrocyte phenotype. Preliminary in vivo data demonstrated scSOX9 significantly improved the outcome of microfracture in cartilage repair in short-term. In this proposal, we aim to promote cartilage in situ regeneration by stimulating bone marrow MSC in a rabbit model of cartilage defect. Experiments are designed to administer scSOX9 protein at the site of microfracture of the defective cartilage. Administration of scSOX9 will be carried out by either direct delivery with a bioscaffold, collagen membrane at the site of microfracture. Regeneration of cartilage will be assessed at 12 and 24 weeks using histology and immunohistochemistry for characters of hyaline cartilage by examination of content of aggrecan and collagen type I, II and X and osteocalcine. The biomechanical properties of the repaired cartilage will also be evaluated. Immune response to scSOX9 will be assessed by measuring serum antibodies and T cells reaction to scSOX9. Efficacy of re-administration of scSOX9 with microfracture will be assessed in animals who received scSOX9 previously in contralateral knee. Molecular mechanism for which scSOX9 mediated hyaline cartilage repair will be investigated. The research plan takes advantage of the minimal invasive, simple and low cost microfracture to attract bone marrow MSC and pro-chondrogenic property of scSOX9 to regenerate articular cartilage in situ for therapy of cartilage defect. The success of this project will provide invaluable information for design of a clinical trial using the same techniques for therapy of cartilage lesions and prevention of progression to OA.

 描述（由申请人提供）： 软骨损伤很常见，可能导致骨关节炎（OA）的进展，其特征是关节软骨的进行性分解，最终导致滑膜关节的功能失效。软骨的再生一直是OA的软骨修复和处理的吸引人方法。然而，自体软骨细胞移植不足，在体外膨胀过程中，细胞供应不足，对供体部位的新损害和软骨细胞的去分化。位于骨髓和许多成年组织中的间充质干细胞（MSC）能够自我再生和分化为软骨细胞，并且是原位再生的潜在来源。微裂纹诱导骨髓MSC迁移到软骨缺陷部位，并促进纤维球杆菌的产生。然而，纤维球杆菌是不可固定的，并且在功能上长期不足。 Sox9是属于Sox（Sry-type HMG Box）家族的转录因子，已被确定为软骨细胞表型的“主调节器”。我们已经产生了一个细胞渗透，超级带电的SOX9融合蛋白（SCSOX9）。体外研究表明，SCSOX9蛋白会诱导骨髓衍生的MSC增殖，并分化为软骨细胞。 I型胶原蛋白和X型的产生降低，但增加了胶原蛋白II型和Aggrecan，这是关节软骨细胞的特征，这证明了这一点。在21天的培养物中，最初的SCSOX9添加足以将MSC区分为软骨细胞并保持软骨细胞表型。初步体内数据表明，SCSOX9在短期内显着改善了软骨修复中微骨折的结果。在此提案中，我们旨在通过在软骨缺陷的兔子模型中刺激骨髓MSC来促进软骨原位再生。实验旨在在有缺陷软骨的微裂缝部位施用SCSOX9蛋白。 SCSOX9的给药将通过在微裂缝部位的生物苏斯科纸，胶原蛋白膜进行直接递送来进行。通过组织学和免疫组织化学对软骨的再生将在12和24周中评估透明软骨的特征，通过检查Aggrecan和I型，II型，II和X型和骨分泌蛋白的含量。还将评估修复后的软骨的生物力学特性。通过测量血清抗体和T细胞对SCSOX9的反应来评估对SCSOX9的免疫反应。 SCSOX9用微骨折的重新服用的功效将在先前接受SCSOX9的动物中评估。将研究SCSOX9介导的透明软骨修复的分子机制。该研究计划利用了最小的侵入性，简单和低成本的微裂纹来吸引SCSOX9的骨髓MSC和促造成性质的特性，以将关节软骨再生用于治疗软骨缺陷的原位。这个项目的成功 将使用相同的技术治疗软骨病变和预防向OA进行的相同技术来设计临床试验的宝贵信息。