Establishment Of Cultured Adult Rabbit Articular Chondrocyte And Effects Of BFGF

Articular cartilage is a kind of special non-vessel tissue with weak self-repair or regeneration ability. Therefore, up till now, the repair of articular cartilage injury or defect has remained a puzzle. Tissue engineering is a burgeoning subject that came forth in 1980s. It is the result of the common development and integration of material science, engineering and life science as well as one of the main approaches contributing to the advancement of future medical science from replacement to regeneration. One of its branches is cartilage tissue engineering which includes three aspects:â‘ seed cells of certain quantity and qualityâ‘¡brackets for suitable materialâ‘¢cytokine.Whether chondrocyte repair or cartilage tissue engineering, seed cells of certain quantity and quality is the first and foremost precondition.The successsful isolation and cultivation in vitro of chondroncytes in 1960s brought to us fresh inspiration; as a result, at present, the cultivation of chondrocytes is not only important, but mature as well. There are certain limits, however, on cultivation of chondrocytes:scarce sources of chondrocytes or tissue;failure to obtain cartilage from adult body sufficient to make chondrocytes;impossibility to acquire young chondrocytes with high growing potential as seed cells for tissue engineering; great effects of the age of chondrocytes upon survival rate. Technological difficulties such as immune rejection to allogenic cells, low transformation rate and even reverse transformation of mesenchymal stem cell to chondrocytes, genetic mutation , difficulty in controlling the tranformation,and disadvantages such as aging and hard to stop in gene amplification still exist though more soures of seed cells of cartilage, such as mesenchymal stem cells, embryonic stem cells, muscles, adipose tissue-derived stem cells, etc. are reported and genetic method to get seed cells in vitro has been developed. Cultivation of chondrocytes remains the most significant approach of obtaining chondrocytes. Despite the success in cultivation of chondrocytes,the reports are always limited to baby rabbits while few reports on growing pattern of chondrocytes of adult rabbits have been published. As the cartilages of baby animals have not fully grown, the chondrocytes have enormous potential in differentiation and are easy to be cultured in vitro, great survival rate in cultivation in vitro, high proliferation rate, and late appearance of dedifferentiation phenomenon;whereas in clinical practice, patients are most adults whose cartilages are mature or pathological and whose chondrocytes are scanty with no active differentiation ability or are degenerated,thus there low comparability between basic experiment and clinical practice. Hence, in this study, adult rabbits are applied as subjects, in vitro monolayer cultivation method is adopted, and bFGF is employed on the cultivation of adult rabbit chondrocytes, to understand the effects of basic fibroblast growth factor ( bFGF) on the proliferation and metabolism of articular chondrocytes and provide experimental and theoretical evidence for the obtain of seed cells for cartilage tissue project and repair of articular cartilage injury.Since 1970s, along with the advancement of cytochemistry and cytobiology, people has begun exploring the repair of articular cartilage from cellular level, and thereby discovered that great number of hormones and growth factors such as insulin-like growth factor(IGF), fibroblast growth factor(FGF), transforming growth factor(TGF), and interleukin(IL) exert effects on biological behavior of chondrocytes. The study on basic fibroblast growth facto(rbFGF)is the most comprehensive, thus its mechanism has been clearly understood. bFGF is a heparin-binding polypeptide. It has strong mitogenic activity upon cells developed from mesoderm, neural ectoderm and other ectoderm and endoderm derivatives, and can increase the synthesis of chondrocyte collagen. bFGF has relatively strong effects on stimulating the synthesis of collagen in chondrocytes, inducing the proliferation of chondrocytes,induing and influencing the differentiation of chondrocytes,inhibiting the synthesis of mucopolysaccharides and fibronectin in collagen I, stimulating the production of collagen type II and chondroitin sulfate mucopolysaccharides. promoting mitosis and synthesis of chondrocyte DNA , quickly transforming into fibroblast when shortage of bFGF appears in chondrocytes. It is considered to be one of the most effective stimulating sources to the mitosis of articular chondrocytes and the strongest growth factor. The function of bFGF has been acknowledged and affirmed for it has mitogenic activity, and can regulate in vitro the differentiation of cells and elongation and growth of axon.On the basis of following facts:(1)possibility to obtain seed cells of sufficient quantity and quality is the most foundamental and vital precondition to the success of tissue engineering (2)at present, cultivation in vitro of chondrocytes is still the main approache of getting seed cells for tissue engineering, none other methods can rival with it yet as far as the current research situation is concerned(3)in most of the past cultivation in vitro of chondrocytes, young animals are the sources of seed cells, few researches on growing pattern of chondrocytes of adult rabbits have been conducted(4)though the chondrocytes of baby animals have enormous potential in differentiation and are easy to be cultured in vitro, great survival rate in cultivation in vitro, whereas in clinical practice, patients are most adults whose cartilages are mature or pathological and whose chondrocytes are scanty with no active differentiation ability or are degenerated, thus there low comparability between basic experiment and clinical practice, the results obtained from the experiment might be inaccurate(5)the mechanism of bFGF's function on adult rabbit chondrocytes has not been reported, neither has the possibility of adult rabbit chondrocytes influenced by bFGF being seed cells. Hence this experiment organically combine bFGF and the cultivation in vitro of adult rabbit chondrocytes to explore the function mechanism of bFGF on chondrocytes of adult rabbit as well as whether chondrocytes of adult rabbit are suitable for seed cells, providing experimental data for clinical practice and searching methods of obtaining seed cells.It has been found through the study that:(1) bFGF has the functions of promoting mitosis and synthesis of chondrocyte DNA , increase the synthesis of proteoglycan and collegen type II on adult rabbit chondrocytes cultured in vitro. Statistics shows distinct difference comparing with control group, which indicates that bFGF has the functions of promoting growth and proliferation on chondrocytes of adult rabbits.(2) It has also been proved that chondrocytes of adult rabbits also own growth activity, ability of surviving cultivation in vitro and being used as seed cells. However the fact that the dedifferentiation phenomenon appears early than that in baby rabbits indicates cells from three generation ago should be used. This study is innovative, data and results will definitely be useful as reference to further foundamental or clinical experiment.