| The first demonstration of the presence of leptin in 1994, Rockefeller University, Laboratory for the use of cloning sites were cloned mouse and human homologous sequences (ob gene) and confirmed, and has proven its genetic, molecular structure and its physiological role. Leptin produced mainly by white fat, but other organizations can produce, such as cartilage, bone, brown fat, and so on. In recent years, people involved in many areas of leptin research and found that many are associated with many diseases, these diseases, including obesity, diabetes, osteoporosis, etc. In particular, found that, in the process of endochondral bone, the leptin also play a role, possibly through the promotion of cartilage cell proliferation makes the growth plate increased, and ultimately makes the long bone growth. There experiments confirmed that above the existence of human articular chondrocytes leptin receptor, and has done a series of animal experiments confirmed that leptin can increase the thickness of articular cartilage in adult mice. Of leptin in the pathogenesis of osteoarthritis also have a certain role, it was found that patients with osteoarthritis synovial fluid contains leptin, and is proportional to the severity of arthritis, but the concrete impact on the relationship between the two is also the need for further studies confirmed.To cover the joint surface cartilage layer of bright, low friction, flexible, high permeability of the specificity of connective tissue, mainly by cartilage cells and extracellular matrix composition, conduction and distribution of its function is to load, maintain and bear the articular surface of the contact stress. Articular cartilage is no vascular, lymphatic and nervous organization, so self-repair capacity is limited, minor injuries may lead to progressive damage. It is the case, for many years has been the cartilage injury and repair the focus of clinical and basic research, difficult. Today, people are now worked out a number of ways to repair damaged cartilage, such as the scaffold to replace the damaged area, using a number of drugs to promote the reconstruction of cartilage, etc., has yielded some results, but due to various reasons, can not be universal access to treatment. Cartilage contains a variety of growth factors and secrete a number of matrix components, mainly including transforming growth factor-β(transforming growth factor-β, TGF-β), insulin-like growth factor (insulin-like growth factor, IGF), bone morphogenetic protein (bone morphogenetic protein, BMP), fibroblast growth factor (fibroblast growth factor, FGF), II collagen (type II collagen), proteoglycan (aggrecan) and so on, which by autocrine, paracrine, etc., together on cartilage cell division, growth, maturity and aging at all stages.Objective: This study was designed to study three-dimensional culture of leptin on the in vitro activity of rabbit articular cartilage cells, proliferation, cell phenotype and function, and to explore its matrix degradation of cartilage repair and key genes related to col-II, aggrecan, TGF -β1 expression, thus the results to better guide clinical on the cartilage tissue repair and cartilage tissue engineering development.Method1 take 3-4 weeks old rats were obtained under sterile conditions after execution of articular cartilage cells, with the mechanical method and enzyme eliminate the rule of law into the cell suspension, monolayer culture in vitro amplified three-dimensional culture;2 on cultured rat articular cartilage cells in morphology and identification;3 Experimental group (all with three generations of cartilage cells) were divided into two groups according to test methods under the conditions of a simple three-dimensional cultured chondrocytes, leptin (a concentration of 10ng/ml) under the conditions of the role of three-dimensional cultured chondrocytes;4 Using MTT method and flow cytometry cells were treated with leptin to stimulate cartilage cell proliferation at different times;5 using PCR to detect cartilage cells were treated with leptin to stimulate the expression of different times of TGF-β1, aggrecan situation;6 The SPSS13.0 software, the results were analyzed statistically.Results1 Using cell culture method has been successfully cultivated in vitro in rat articular cartilage cells, and, through morphological observation and special staining confirmed.2 Leptin cell proliferation was significantly higher than the control group, the best action time for the role of the 24h, to continue to leptin stimulated cell proliferation was not obvious, and has inhibitory trend.3 Leptin group aggrecan, TGF-β1 gene expression was significantly higher than the control group, the best time is when the role of 24h.Conclusion1 leptin can significantly improve the activity of cartilage cells;2 leptin can promote the in vitro three-dimensional culture of rabbit articular chondrocytes3 Leptin could enhance TGF-β1 mRNA expression in order to prevent degradation of extracellular matrix, there is potential for cartilage repair.4 Leptin can enhance aggrecan mRNA expression, can promote the generation of extracellular matrix, cartilage cells to provide a good living environment. |
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