OBJECTIVE This paper aims to investigate the suitable cell density and the best formation time of tissue engineered autologous cartilage and to provide theoretical basis and parameters for clinical application. METHODS The chondrocytes isolated from mini swines’ ears were mixed with injectable biocompatible matrix (Pluronic), and the density of cell suspensions were 10, 20, 30, 40, 50, 60, 70 x 10(4)/ml. The chondrocyte-polymer constructs were subcutaneously injected into the abdomen of autologous swine. The specimens were observed grossly and histologically after 6 weeks, and investigated the suitable cell density. Then the chondrocyte-polymer constructs with suitable cell density were transplanted into the abdomen of autologous swine and evaluated grossly and histologically in 1, 3, 6, 9, 15 weeks after transplantation to investigate the best formation time of tissue engineered cartilage. RESULTS The experiments demonstrated that the tissue engineered autologous cartilage was similar to the natural cartilage on animals with normal immune system in histological characteristics. The optimal chondrocyte density is 50 x 10(6)/ml, and the proper harvest time is the sixth week. CONCLUSION With tissue engineering skills, we have identified the optimal chondrocyte density and the proper harvest time.
OBJECTIVE To review the recent research progress of bone-marrow stromal stem cells (BMSCs) in the conditions of culture in vitro, chondrogenic differentiation, and the application in cartilage tissue engineering. METHODS: Recent original articles related to such aspects of BMSCs were reviewed extensively. RESULTS: BMSCs are easy to be isolated and cultivated. In the process of chondrogenesis of BMSCs, the special factors and interaction between cells are investigated extensively. BMSCs have been identified to form cartilage in vivo. One theory is the committed chondrocyte from BMSCs is only a transient stage. CONCLUSION: BMSCs are the alternative seeding cells for cartilage tissue engineering. The conditions promoting mature chondrocyte should be further investigated.
Objective To evaluate the effect of WO-1 on repair of the bone defect in the New Zealand rabbit radius by an oral or local administration. Methods Bone defects were surgically created in the bilateral radii of 36 Zealand rabbits (1.6-2.0 kg), which were randomly divided into3 groups. In Group A, the defective areas were given WO-1 0.1 ml (50 mg/ml) by the local injections; in Group B, the rabbits were given WO-1 5 mg each day by the oral administration. Group C was used as a control group. Among each of the 3 groups, 4 rabbits were randomly selected and were sacrificed at 20, 30 and 60 days after operation, respectively. Then, the serological, X-ray and histological examinations were performed. Results The serum alkaline phosphatase and bone glaprotein levels were significantly higher at 20 and 30 days after operation in Groups A and B than in Group C, but significantly lower at 60 days after operation in Groups A and B than in Group C（Plt;0.01）. The X-ray and histological examinations at 20, 30 and 60 days after operation revealed that the callus formation and remodeling were earlier in Groups A and B thanin Group C, and the remodeling was earlier and better in Group A than in Group B. Conclusion WO-1 can promote the repair of the radial defect in a rabbit; however, further studies on the doseeffect relationship, administration time, and administration route are still needed.
Objective To study the grafting effect of tissue engineered artificial rat skin equivalent on full thickness wounds. Methods Full thickness wounds(Φ20mm) were made on the backs of twenty four nude mice which be divided in artificial skin(AS) group, chitosan membrane(CH) group and control group. All wounds were covered with AS, CH and petrolatum gauze , respectively. The wounds were observed daily by infrared ray scanning and histological examination on the 3rd , 7th, 14th, and 21st days. Results The wounds in AS group healed better than those in CH group and control group. The artificial skin achieved a good adherence to wound and there were some crescent regenerative blood vessel appeared in the AS group on the 3rd day of grafting. Then, the epidermal cells in artificial skin proliferated and differentiated to form a new epidermis consisting of stratum basal, stratum spinosum, stratum granulosum, stratum corneum almost like the natural skin. Dermis of the sd extracellular matrix secreted by fibroblasts; the chitosan lattice was degraded and replaced by the extracellular matrix. On the 14th day of grafting, the wounds healed. The color of artificial skin grafted was very similar to the natrual skin and the formed scar was very smaal. Conclusion A kind of new reconstructive tissue engineering artificial skin has good histocompatibility and can be transplanted into the full-thickness wounds.
OBJECTIVE: To study the expression of type I collagen and its receptor system-integrin alpha 2 beta 1 in different passages of osteoblasts. METHODS: The expression of type I collagen and integrin alpha 2 beta 1 in the primary, sixth and fifteenth passage of osteoblasts were detected by S-P immunohistological staining technique, and their mRNA expression by quantity RT-PCR technique. RESULTS: Type I collagen and integrin alpha 2 beta 1 were expressed in different passages of osteoblasts and there was no significant difference among three passages by immunohistological technique. Their mRNA expression was gradually decreased with subculture. CONCLUSION: Type I collagen promotes the adhesion and phenotype expression of osteoblasts through its receptor-integrin alpha 2 beta 1. The reductive expression of type I collagen-receptor system will decline the phenotype of osteoblasts.
Objective To investigate the current situation, problems of medicinal biotechnology in China, and to provide the relevant countermeasures for its development. Methods We surveyed the units which could carry out medicinal biotechnology projects in 30 provinces except Tibet, and compared the results with that in America.Results The questionnaire were returned from 25 provinces (83.4%), and there were 1 477 medicinal biotechnology projects carried out by 149 units in the past 10 years. These projects ranged from basic biotechnology to regenerative medicine and stem cell researches. The basic research projects constituted quite large percentage among all the projects. But the development levels in different areas were imbalanced, cross correlation with the development levels of economy. An echelon team of talents has been developed, most of them were trained in China. The invested capital differed considerably among units, in general the amounts were insufficient. Most invested capital came from the government. The number of patent application for projects based on independent-developed technology was small. This showed that project principals had a poor understanding of patents. More than half of units did not have a Bioethics Committee. From the search result for documents, the number of articles on stem research of China was close to that in America; and the number of articles on gene treatment and tissue engineering has already exceeded that of America. However, research on gene diagnosis of China was lagging far behind America. Conclusions An echelon team of talents has been developed, most of them are trained in China.We should give full play to the advantage of the distribution of qualified personal resources in developed economical areas so as to promote the applicability and popularity of medicinal biotechnology in less developed areas.Regarding to applicability and development, we should first develop applied technology to form the core competetiveness of basic research, technology development and application; we should also strengthen the training in ethics and regulation to establish a set of scientific assessment of medicinal biotechnology and management system.
Objective To sum up the research advances of the seed cell and the culture system using in tissue engineering cartilage. Methods The recent original articles about the seed cell and the culture system in tissue engineering cartilage were extensively reviewed. Results At present, autologous or homologous cells is still major seed cell and the three dimensional culture system is also major system for tissue engineering cartilage. Conclusion The source of seed cell for tissue engineering cartilage. Conclusion The source of seed cell for tissue engineering cartilage should be further explored, and the culture system need to be improved and developed.
OBJECTIVE To study the biocompatibility of skin reproductive membrane. METHODS According to ISO’s standards, the extractions of the skin reproductive membrane were prepared, and the acute systematic toxicity test, primary skin irritant test, cytotoxicity test, gene expression of type I collagen and fibronectin were detected to evaluate the biocompatibility of skin reproductive membrane. RESULTS All of those tests showed negative results. CONCLUSION The skin reproductive membrane has excellent biocompatibility in the level of the systematic, cellular and molecular biology.
Objective To study the difference of repairing segmental bone defect with bio-derived bone preserved by various methods.Methods Freeze-dried biomaterials had been stored in two different preservation solutions for three months,while the biomaterials stored for same period were observed as control group. The experimental model of 15 mm radial segmentaldefect was made in 60 New Zealand white rabbits, which were divided into groups A,B and C according to transplant materials preserved by various methods. Groups A and B were deeply divided into A1 and A2 subgroups, B1 and B2 subgroups according to whether materials were cocultured with osteoblasts. Tissue engineered bone was used to repair bone defects of left limbs in A1 and B1 subgroups, while simple material to repair defects of right limbs in A2 and B2 subgroups. Group C was divided into C1 and C2 subgroups. Freeze-dried material was used to repairbone defects of the left limbs, while defects of the right limbs as blank control group. The samples were harvested and observed by the roentgenographical, histomorphological, biomechanical and computerized graphical analysis at 4,8 and 16 weeks. Results All of the defects treated with implants exhibited new bone formation 4, 8 and 16 weeks postoperatively, increasing with time. The radiological, histomorphological and biomechanical evaluation showed that the ability of new bone formation was arranged in 6 subgroups as follows:A1gt;A2gt;C1gt;B1gt;B2gt;C2, the difference was significant between them (P＜0.001, P＜0.05).The ability of new bone formation was best and at 16 weeks the defect was bridged with the appearance of marrow cavities in A1 subgroup, the biomechanicalproperties in implants approached to those of normal bone. Conclusion The choice of proper preservation solution can improve the ability of repairing bone defect.
Objective To study the integration of rat marrow stromal stem cells (MSCs) after transplantation into acellular extracellular matrix (AECM). Methods We got 16 femurs from 8 Kunming rats, the femurs were treated by Triton X100 toget AECM, MSCs were collected from femoral marrow of 20 Kunming rats about a mouth old by PBS 4ml, centrifugalized and primary cultured in bottles,then therat MSCs were transplanted into AECM at a concentration of 5×106/ml and culturedfor 7 days. The integration of the donor cells was observed using one phase contrast microscope, a light microscope and a scanning electron microscope (SEM).Results In AECM bone lacunas there were MSCs nucleuses stained blue. The nucleuses were unevenly distributed in AECM with more in the peripheral AECM than in the central AECM and with more in the layer anear culture medium than in the layer far away from culture medium.AECM possessed a good spatial scaffold structure, the marrow stromal stem cells were well integrated into AECM.Conclusion AECM can be usedas a good scaffold material for tissue engineered bone construction.