Bone Tissue Engineering Using Mineralized Collagen-GAG Scaffolds

使用矿化胶原蛋白-GAG 支架的骨组织工程

• 批准号: 8440695
• 负责人: Timothy A Miller
• 金额: --
• 依托单位: VA GREATER LOS ANGELES HEALTHCARE SYSTEM
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-10-01 至 2015-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8440695
• 关键词: 3-Dimensional; Adhesions; Affect; Afghanistan; Aging; Architecture; Autologous; Back; Binding; Biocompatible; Biological Sciences; Bone Marrow; Bone Tissue; Bone Transplantation; Caring; Cells; Cephalic; Collagen; Defect; Development; Diffusion; Dose; Excision; Exhibits; Extracellular Matrix; GAG Gene; Glycolic-Lactic Acid Polyester; Glycosaminoglycans; Goals; Harvest; Healed; Hemostatic Agents; Human; Implant; In Vitro; Infiltration; Inflammation; Injury; Integrins; Iraq; Laboratories; Limb structure; Malignant Neoplasms; Mechanics; Medical center; Mesenchymal Stem Cells; Modeling; Nutrient; Oryctolagus cuniculus; Osteogenesis; Osteomyelitis; Pain; Patients; Population; Property; Quality of Care; Recombinants; Role; Services; Signal Transduction; Signaling Molecule; Site; Solid; Stromal Cells; Structure; System; Techniques; Technology; Test Result; Testing; Tissue Engineering; Trauma; Vascularization; Veterans; Wound Healing; base; biomaterial development; bone; bone morphogenetic protein 2; calcium phosphate; combat; comparative efficacy; craniofacial; healing; improved; in vivo; mineralization; nanoparticulate; operation; osteogenic; prevent; public health relevance; reconstruction; repaired; scaffold; tissue culture

DESCRIPTION (provided by applicant): Our laboratory has been focused on the development of a synthetic bone graft substitute (BGS) for the past 15 years. Our experimental bone graft substitutes have included PLGA- (Poly-lactide-co-glycolide) and collagen-based scaffolds seeded with autologous bone marrow stromal cells (BMSC), and stimulated by osteoinductive agents including BMP-2. With these BGS models, we have generated new bone both in vitro, using a 3-dimensional tissue culture system, and in vivo, resulting in the healing of critical-sized cranial defects in the rabbit. While our studies have been promising, a clinically useful BGS remains elusive. The carrier is of critical importance in the development of a BGS. PLGA has shown much promise, being both biocompatible and biodegradable. However, there are limitations to the use of PLGA in tissue engineering. PLGA induces the formation of acidic degradation products that can affect local PH, possibly inducing inflammation. Collagen based scaffolds, including collagen-GAG, have been widely used in bone tissue engineering because collagen is a naturally existing component of extracellular matrix that promotes cell binding, exhibits low antigenicity and has excellent haemostatic property. We demonstrated that human mesenchymal stem cells (MSC) seeded in collagen scaffolds exhibit accelerated and robust mineralization and bone formation in comparison to their counterparts seeded in PLGA scaffolds. However, we also discovered in our in vitro studies that cell-seeded collagen scaffolds undergo significant contraction compared to PLGA scaffolds. The contraction can greatly hamper application of collagen implants in repairing bony defects. In order to limit volume loss and destruction of collagen implants, we propose to stabilize the structure of collagen scaffolds by incorporating nanoparticulate calcium phosphate (CaP) on the collagen-GAG fibrils. Our initial test results indicate using MC-GAG scaffolds can greatly prevent contraction caused by adhesion and differentiation of hMSCs. It is also superior to collagen-GAG in supporting new bone formation. Our current proposal intends to further investigate the feasibility of MC-GAG scaffolds in bone tissue engineering. In this proposal, we will compare collagen-GAG and MC-GAG scaffolds for their ability to support osteogenic differentiation in rabbit BMSCs cultured in vitro. We will also compare the efficacy of cellular scaffolds made of collagen-GAG and MC-GAG in healing a critical-sized rabbit cranial defect by performing histological and mechanical analysis of newly formed bone. Finally, we will study integrins and their downstream signaling molecules responsible for the differences between collagen-GAG and MC-GAG in their ability to control contraction and support osteogenic differentiation in rBMSCs. We expect that MC-GAG scaffolds will (1) enhance osteogenic differentiation and reduce contraction in rabbit BMSCs compared to collagen alone, and (2) accelerate new bone formation in a rabbit cranial defect model compared to collagen alone. We anticipate that changes in integrin signaling are responsible for enhanced differentiation and reduced contraction of rBMSCs in MC-GAG scaffolds.

描述（由申请人提供）： 在过去的15年中，我们的实验室一直关注合成骨移植替代品（BGS）的发展。我们的实验性骨移植替代物包括Plga-（聚乳酸 - 糖糖苷）和基于胶原蛋白的支架，这些支架与自体骨髓基质细胞（BMSC）种子，并由包括BMP-2在内的骨诱导剂刺激。使用这些BGS模型，我们使用3维组织培养系统和体内产生了新的骨骼，从而导致兔子中关键尺寸的颅骨缺陷的愈合。尽管我们的研究很有希望，但临床上有用的BG仍然难以捉摸。载体在BGS的发展中至关重要。 PLGA表现出了很多希望，既具有生物相容性又可生物降解。但是，在组织工程中使用PLGA存在局限性。 PLGA诱导可能影响局部pH的酸性降解产物的形成，可能引起炎症。基于胶原蛋白的支架，包括胶原蛋白 - GAG，已广泛用于骨组织工程中，因为胶原蛋白是促进细胞结合的细胞外基质的自然存在的成分，它表现出低抗原性，并且具有极好的止血特性。我们证明，与在PLGA支架中播种的对应物相比，在胶原支架中播种的人间充质干细胞（MSC）表现出加速而强大的矿化和骨形成。但是，我们在体外研究中还发现，与PLGA支架相比，细胞种子的胶原蛋白支架经历了明显的收缩。收缩可能会大大阻碍胶原蛋白植入物在修复骨缺陷中的应用。为了限制胶原蛋白植入物的体积损失和破坏，我们建议通过在胶原-GAG原纤维上掺入纳米磷酸钙（CAP）来稳定胶原蛋白支架的结构。我们的初始测试结果表明，使用MC-GAG支架可以极大地阻止HMSC的粘附和分化引起的收缩。它在支撑新的骨形成方面也优于胶原蛋白-GAG。我们目前的建议旨在进一步研究MC-GAG支架在骨组织工程中的可行性。在此提案中，我们将比较胶原蛋白-GAG和MC-GAG支架的能力，以支持在体外培养的兔BMSC中的成骨分化的能力。我们还将通过对新形成的骨骼进行组织学和机械分析来比较胶原蛋白-GAG和MC-GAG制成的细胞支架在愈合临界大小的兔颅缺损方面的功效。最后，我们将研究整联蛋白及其下游信号传导分子，以控制胶原蛋白-GAG和MC-GAG之间在控制收缩和支持RBMSC中成骨分化的能力方面的差异。我们预计，与单独的胶原蛋白相比，MC-GAG支架将（1）增强成骨分化并减少兔BMSC的收缩，并且（2）与单独的胶原蛋白相比，在兔颅缺陷模型中加速了新的骨形成。我们预计整联蛋白信号传导的变化将导致MC-GAG支架中RBMSC的分化增强和减少收缩。