| Research background:Bone and joint disease (osteoarthrosis, OA) is a frequently occurring non-inflammatorylesion that is characterized by progressive loss of movable joint. Since there’s no blood vessels,nerves nor lymphatic vessels in articular cartilage and the chondrocyte has very limitedproliferation potential, the articular cartilage can hardly repair itself. Its clinical pathologicalfeatures include articular cartilage degeneration and hyperplasia of bone under ligamentattachment or cartilage due to various reasons. The disease would eventually lead to structuraldamage of the joint and impairment of movements, accompanied by joint pain, stiffness,deformity that severely impact the quality of patients’ life. Due to its particular position andstructural, functional complexity, combined with the complex jaw-conversion process indevelopment, temporomandibular joint is more susceptible to joint disease.With thedevelopment of cartilage tissue engineering, how to rapid amplification of large number of stemcells and their more effectively to the cartilage cells to repair damaged cartilage has bocome oneof the research hotspots.Purpose:1、Based on the source of rabbit adipose-derived stem cells (ADSCs) were isolated,purified and cultured in vitro, observe the proliferation ability and biological characteristics andtheir multilineage differentiation potential.2、 Analysis the ability.of different concentration of ginseng saponins on humanRg1/ADSCs into chondrogenic differentiation.3、Establishment of temporomandibular joint osteoarthrosis model, application withhyaluronic acid as matrix composite ginsenoside Rg1/ADSCs on the rabbit temporomandibularjoint cartilage to repair and improve the damaged the feasibility of osteoarthrosis and thetemporomandibular joint function.Methods:1、March age healthy New Zealand rabbits and take its fat, and isolat and culture theADSCs cells were subcultured after identification and adipogenic, osteogenic and chondrogenicdifferentiation, observe multilineage potential ability. 2、ADSCs chondrocyte differentiation, and through MTT assay for the detection ofcartilage cell activity and immunohistochemical assay for the detection of cell type II collagenexpression of different concentration, determination of ginseng saponins on human Rg1/ADSCschondrogenic differentiation capacity of ginsenoside Rg1, and get the optimal effectiveconcentration values.3、32New Zealand white rabbits were randomly divided into four groups: blank group,model group, control group and experimental group,8in each group. Osteoarthritis model wasestablished by intra-articular injection of papain. Two weeks after successful model building,medication was given for the rabbits in control group and experimental group.0.6mLginsenoside Rg1/ADSCs suspension was injected into superior joint space for the rabbits incontrol group, once a week;0.6mL ginsenoside Rg1/ADSCs complex was injected for therabbits in experimental group, once a week. Animals in each group were sacrificed3and9weeks after administration, respectively (16for each time:n=4) Specimens were prepared forscanning electron microscopy (SEM), histological observation and real-time quantitative PCR(qPCR) of condylar cartilage. And the data were analyzed by analysis of variance and t testusing SPSS17.0statistical package.Result.1、Isolation of the ADSCs shape as the shuttle type, passage after rapid growth, most ofthem fibrous growth, induced, by oil red O staining showed cell lipid droplets in appearance;under osteogenic induction, cells showed the colony growth, alizarin red staining cells calciumnodule formation; into cartilage induction by toluidine blue staining, cartilage cells, cytokinesisis light blue, dark blue for the cell nucleus.2、Ginsenoside Rg1can enhance and promote ADSCs cartilage cell transformation ability,and in the40μmoL/L dilutions of cartilage cell formation have significant promoting effect.3、 Animal experiment, by gross observation, scanning electron microscopy andimmunohistochemical visible model of articular cartilage lesions in severe group, treatmentgroup and model group contrast, lesions of light and improvement in good condition, and theexperimental group in the hyaline cartilage formation and cartilage cell proliferation anddifferentiation are better than the control group. Real-time PCR and Western blot results show,model group MMP-13,the mRNA and protein expression were significantly higher than theother three groups, experimental group and blank group expression is relatively flat but lower than that of the control group, the difference was statistically significant; the model group(TIMP-1),the mRNA and protein expression amount lower than the other three groups, theexperimental group and the group of relatively flat blank expression but higher than that of thecontrol group, the difference was statistically significant.Conclusion:1、To culture ADSCs is convenient and passage of stable performance, good multilineagedifferentiation potential, and can be achieved through a variety of induced differentiationmedium success will ADSCs to adipogenic, osteogenic, chondrogenic transformation, is theideal seed cells.2、 Ginsenoside Rg1/ADSCs to promote chondrocyte transformation function,chondrogenesis is obvious, and the concentration of40μmoL/L is the best.3、 With hyaluronic acid as matrix composite ginsenoside Rg1/ADSCs on rabbittemporomandibular joint lesion area of cartilage repair and improvement of thetemporomandibular joint osteoarthritis in rabbit has obvious effect. |
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