Rapid Prototyping PLGA Scaffold-collagen Gel For Cartilage Tissue Engineering

Cartilage defects are a major problem in orthopedic surgery.Because the cartilage tissue itself lacks a blood supply to support repair and remodeling,once damaged,the cartilage has little capacity for spontaneous healing.Most of the current therapies to repair damaged cartilage have limititions.Progress in tissue engineering provided a new concept for tissue regeneration and transplantation.The scaffold can provide suitable spatial structure for cell proliferation and matrix secreting,thus facilitate tissue forming.Since the scaffold surface properties play an important role in determining how cells respond to the biomaterials,the development of tissue engineering has recently focoused on the surface design of biodegradable scaffolds.Several approaches,such as alkali hydrolysis treatment,plasma treatment,ion irradiation and surface coating have been developed to improve the bioactivity of the surface.The complex scaffolds were composed of two or more biomaterials to consist a 3-D scaffold with suitable structure and function. It makes up the disadvantages of using artificial synthetical material or natural materials alone. The purpose of this study was to develop a new complex scaffold for cartilage tissue engineering,which owns the advantages of high cell seeding efficiency,good ability to support chondrogenesis as well as adequate physical properities.rapid prototyping PLGA porous scaffold was chosed as framework, then chondrocytes suspended in collagen solution was seed in framework,and gelling process was performed.We believe rapid prototyping PLGA-collagen gel complex could act as a new scaffold for cartilage regeneration.This study includes the following parts:1 Fabricating PLGA scaffold with LDM techonology The structure of scaffold, porosity and mechanical property can meet with the requirement of scaffold for tissue-engineered cartilage.2 Isolation of chondrocytes and identification of cell phenotype. Articular chondrocytes were isolated from the cartilages with the methods of enzymatic digestion.The morphological changes and growth feature of primary cultured and subcultured chondrocytes were observed under the inverted microscope each day. Toluidine blue staining and immunocytochemist ry were used to indentify the chondrocytes. The primarily cultured chondrocytes were in olygonal shape, and became dedifferentiation after 3 passages. The chondrocytes maintained the morphology and immunochemical staining pattern within the first 3 passages. The method used in this work for isolation and culture of chondrocytes is simple and feasible. The chondrocytes cultured in viro maintained the specific chondrocytes phenotype in the first 3 passages. The growth of the second passage was robust and may be suitable for most experiment. 3 The compatibility of original scaffold and hybrid scaffold.After examing water absorption capability of the scaffold, chondrocytes proliferation and differentiation on the scaffold were calculated and the results showed that rapid prototyping PLGA coating with collagen had a good biocompatibility for the attatchment and proliferation of the chondrocyte. It might be ideal biomaterials in tissue-engineered cartilage field.4 Seeding cell methods of seeding chondrocytes into scaffoldsMixture chondrocytes with collagen solution was incorporated into a PLGA 3D scaffold, and made into gel in the scaffold, then cultured invitro.Attachment of chondrocytes were evaluated by cell count.Distribution of chondrocytes into 3D scaffolds was observed by phase microscope.The results showed More than 90 percent of seeding chondrocytes were able to incorporated into PLGA scaffolds efficiently and uniformly, and prevent cell loss.5 cartilage tissue formation in nude miceThe complexes of seeding of chondrocytes encapsulated in collagen gel into RP PLGA scaffolds were transplanted into the back of nude mice. The specimens harvested at the 4th week,8th week and 12th week were analyzed by observation, histology and tested the quantification of glycosaminoglycan in neo-cartilage.The allograft of chondrocytes proliferated successfully in the back of the nude mice, new cartilage could be seen by the end of 12 weeks and the scaffolds have suitable degradation。The cartilage formed with the scaffold degrading.Our research has demonstrated that the attachment , proliferation ,differentiation of chondrocytes on rapid prototyping PLGA coating with collagen.The methods of gel encapsulated cells incorporated to rapid prototyping PLGA can fix cells in scaffold uniformly and efficiently. Meanwhile the complex of gel encapsulated cells incorporated to rapid prototyping PLGA has the capability of cartilaginous tissue forming.Rapid prototyping PLGA-collagen gel was suitable scaffold to carry cells in the research of tissue-engineered cartilage.