Objective To review research progress of corneal tissueengineering.Methods The recent articles on corneal tissue engineering focus on source and selection of corneal cells, the effects of growth factors on culture of corneal cells in vitro. The preparation and selection of three-dimensional biomaterial scaffolds and their b and weak points were discussed. Results The corneal tissue engineering cells come from normal human corneal cells. The embryo corneal cell was excellent. Several kinds of growth factors play important roles in culture, growth and proliferation of corneal cell, and incroporated into matrix.Growth factors including basic fibroblast growth factor, keratinocyte growth factor, transforming growth factor β1 and epidermal growth factor was favor to corneal cell. Collagen, chitosan and glycosaninoglycans were chosen as biomaterial scaffolds. Conclusion Human tissue engineering cornea can be reconstructed and transplanted. It has good tissue compatibility and can be used as human corneal equivalents.
Objective To establish a better method of isolating andculturing ofneural stem cells(NSCs) in neonatal rat brain. Methods Tissue of brain was isolated from neonatal rats. Different medium and culture concentration were used toculture NSCs of neonatal rat. The culture concentration used were 1×10 4, 1×105, 1×106and 1×107/ml respectively. Ingredient of medium was classified into group 1 to 8 respectively according to whether to add 2% B27, epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) as well as the difference in culture concentration. The cells were induced to differentiate asto be confirmed as NSCs, and then were checked by phase contrast microscopy and identified by immunocytochemistry. Results The cells isolated and cultured gathered into neurospheres. The cells were capable of proliferating and maintaining longterm survival in vitro. The cells could be differentiated into neurons and glia.It was to the benefit of the survival of NSCs to add 5% fetal bovine serum(FBS)into the medium at the beginning of the culturing. When 10% FBS was added intothe medium, the neurospheres differentiated quickly. When concentration 1×106/ ml was used, the growth rate of the cells was the highest of all the concentrations. Reasonably higher cell concentration promoted the proliferation of NSCs. It was necessary to add 2% B27, EGF, and bFGF into the medium. The cells had the best growth when 2% B27, 20 ng/ml bFGF and 20 ng/ml EGF were added into the culture medium. EGF and bFGF had cooperative effect. Conclusion A better method of isolating and culturing of NSCs in neonatal rat brain is established and the foundation for future research is laid.
OBJECTIVE: From the point of view of material science, the methods of tissue repair and defect reconstruct were discussed, including mesenchymal stem cells (MSCs), growth factors, gene therapy and tissue engineered tissue. METHODS: The advances in tissue engineering technologies were introduced based on the recent literature. RESULTS: Tissue engineering should solve the design and preparation of molecular scaffold, tissue vascularization and dynamic culture of cell on the scaffolds in vitro. CONCLUSION: Biomaterials play an important role in the tissue engineering. They can be used as the matrices of MSCs, the delivery carrier of growth factor, the culture scaffold of cell in bioreactors and delivery carrier of gene encoding growth factors.
Objective To review the recent progress of the researches in the field of cartilage tissue engineering, and to discuss the challenges in construction of tissue engineered cartilage. Methods Literature related with cartilage tissue engineering was reviewed and analyzed. Results Some techniques have been appl ied in cl inical. As far as the seeding cells, induced pluripotent stem cells have attracted much more attention. Current strategies of scaffold designing are trying to imitate both component and structure of natural extracellular matrix. Cartilage regeneration through the autologous cell homing technique el iminate the transplantation of exotic cells and has become the hot topic. Conclusion Successful treatment of the damaged cartilage using tissue engineering method will depend on the advances of stem cell technology development, biomimetic scaffolds fabrication and proper appl ication of growth factors.
Objective To summarize the research progress of controlled release of angiogenic factors and osteogenic factors in bone tissue engineering. Methods The domestic and abroad literature on the controlled release structure of growth factors during bone regeneration in recent years was extensively reviewed and summarized. Results The sustained-release structure includes direct binding, microsphere-three-dimensional scaffold structure, core-shell structure, layer self-assembly, hydrogel, and gene carrier. A sustained-release system composed of different sustained-release structures combined with different growth factors can promote bone regeneration and angiogenesis. Conclusion Due to its controllability and persistence, the growth factor sustained-release system has become a research hotspot in bone tissue engineering and has broad application prospects.
【Abstract】 Objective To explore the interventional effect of platelet lysate (PL) on osteogenic differentiation ofBMSCs by induction in rats in vitro. Methods Twenty-four clean-grade adult Wistar rats, weighing from 250 g to 300 g, maleor female, were included in this study. PL was obtained through three times of centrifugation and repeated freeze-thaw for the blood aspirated from cardiac cavities in 16 Wistar rats. ELISA assay was conducted to detect the concentration of growth factors PDGF, TGF-β1, IGF-1 and VEGF in PL. The BMSCs harvested by flushing femurs of 8 adult Wistar rats were isolated, cultivated and expanded in vitro. The cells at the 4 passage were performed for osteogenic differentiation by induction in three groups of A (5% PL of final concentration in basic induction medium), B (1% PL of final concentration in basic induction medium), and C (no presence of PL in basic induction medium as a control). The morphological changes of the cells were dynamically observed with inverted phase contrast microscope during the whole period. At different time-points, ALP staining (7 days) and ALP/TP (2, 8, 12 days) of the cells were detected to evaluate ALP activity, and the mineral formation in extracellular martrix was examined with Al izarin red staining which provided quantitative analysis of mineral deposits. Results ELISA assay showed that the content of PDGF, TGF-β1, IGF-1 and VEGF in PL reached (300 ± 30), (140 ± 25), (80 ± 35), (70 ± 20) pg/mL, respectively. Morphological observation displayed BMSCs in group A or B gradually turned from spindle-shape to square- or polygon-shape as the morphorlogical type of osteoblast-l ike cells at 7 days. The cells in group A showed slower shape changesbut higher prol iferation than that in group B or C. Moreover, at the 20 days, the cells in group A still displayed dense gro wth and produced obviously decreased amount of mineral deposits in ECM when compared with group B or C. At the 7 days, the cells ofgroup A showed smaller amount of granules positive for ALP staining in cytoplasm when compared with groups B and C, and displayed marked reduction in ALP activity assay at the 2, 8, and 10 days compared with that of groups B and C (P lt; 0.05). At the 20 days, Al izarin red staining showed the number of mineral deposits in groups A, B and C were 7.67 ± 1.10, 12.87 ± 0.81 and 15.59 ± 0.25, respectively, while the area of mineral deposits were (161 778.70 ± 44 550.80), (337 349.70 ± 56 083.24), and (415 921.70 ± 71 725.39) pixels, respectively. The number of mineral deposits and the area of mineral deposits in group A were smaller than those in groups B and C (P lt;0.05). But there was no statistically significant difference between groups B and C (P gt; 0.05). Conclusion PL is a kind of system carrying various growth factors. Exposure of PL inhibits both ALP activity and mineral formation of BMCs in a dose-dependent way under the osteogenic induction environment.
Objective To investigate the effect of acid, basic fibroblast growth factor (aFGF, bFGF) and epidermal growth factor (EGF), andtheir combination on the proliferation of rabbit anterior cruciate ligament (ACL) and medial collateral ligament (MCL) in vitro. Methods Thecells of ACL and MCL were isolated and subcultured from the knee joints of tenweek-old New Zealand white rabbits. The cells were seeded into 96-well corning cluster plates. Three growth factors of different concentration alone or in combination were added into the culture medium respectively, which were 0, 1, 5, 10, 50 and 100 ng/ml for aFGF, bFGF and 0, 1.56, 3.13, 6.25, 12.5, 25 and 50 ng/ml for EGF. The proliferation of the fibroblasts was measured for 48 h with XTT method. Results All of the three growth factors alone promoted the cell proliferation of ACL and MCL fibroblasts. The concentration of aFGF hada significant effect on the proliferation of both ACL and MCL fibroblasts. The concentration of 1 ng/ml bFGF and 5 ng/ml EGF was most effective in promoting the proliferation of ACL, and both bFGF and EGF had a significant effect on MCL. 5ng/ml aFGF with 50 ng/ml EGF had effect on ACL. 1 ng/ml aFGF with 3.13 ng/ml EGF had effect on MCL. Conclusion The three growth factors may promote the cell proliferation of ACL and MCL. These findings suggest that topical application of aFGF, either alone or in combination with EGF may have the potential to promote the proliferation of rabbit ACL and MCL,and aFGF of low concentration in combination with EGF is more effective than single growth factor.
Objective To review the research progress of the current methods of inducing bone marrow mesenchymal stem cells (BMSCs) to chondrogenic differentiation in vitro so as to provide references for researches in cartilage tissue engineering. Methods Various methods of inducing BMSCs differentiation into the chondrogenic l ineage in vitro inrecent years were extensively reviewed and analyzed. Results Adding exogenous growth factors is still the mainly methodof inducing BMSCs differentiation into the chondrogenic l ineage; among the members, transforming growth factor β (TGF-β) family is recognized as the most important chondrogenic induction factor. Other important inducing factors include various chemical factors, physical factors, transgenic methods, and the microenvironmental induction. But the problems of low inducing efficiency and unstable inducing effects still exist. Conclusion The progress of chondrogenic induction of BMSCs promotes its util ization in cartilage tissue engineering. Further researches are needed for establ ishing more efficient, simpler, and safer inducing methods.
To compare the platelet enrichment ratio of platelet-rich plasma (PRP) prepared by different centrifuge methods and to compare the concentration of growth factors released from autologous platelet-rich gel (APG) with the whole blood. Methods Thirteen diabetic patients with refractory skin lesions were enrolled in APG treatment. ① Three kinds of centrifuge methods were selected for PRP by 11 diabetic patients: A(n=6): 529 × g for 4 minutes in the first centrifugeand 854 × g for 6 minutes in the second centrifuge; B (n=5): 313 × g for 4 minutes in the first centrifuge and 1 252 × g for 6 minutes in the second centrifuge; C (n=5): 176 × g for 5 minutes in the first centrifuge and 1 252 × g for 5 minutes in the second centrifuge. Platelet counted on the whole blood and PRP was determined. The APG, produced by combining the PRPwith thrombin and calcium gluconate (10 ∶ 1) was used by patients. ② PDGF-BB, TGF-β1, VEGF, EGF, and IGF-1 were measured in the APG and the whole blood using the enzyme-l inked immunosorbent assay method. Results ① The average platelet concentration was higher in group B [(1 363.80 ± 919.74) × 109/ L] than in groups A[(779.67 ± 352.39) × 109/ L)] and C[(765.00 ± 278.78) × 109/ L] and the platelet recovery rate was 75.2% ± 21.0% in group B. ② The concentration of growth factors all increased with the increasing platelet number. On average, for the whole blood as compared with APG, the PDGF-BB concentration increased from (145.94 ± 133.24) pg/mL to (503.81 ± 197.86) pg/mL (P lt; 0.05); TGF-β1 concentration increased from (3.31 ± 2.27) ng/mL to (5.67 ± 4.80) ng/mL (P lt; 0.05); IGF-1concentration increased from (14.54 ± 35.34) ng/mL to (110.56 ± 84.36) ng/mL (P lt; 0.05); and EGF concentration increased from (160.73 ± 71.10) pg/mL to (265.95 ± 138.43) pg/mL (P lt; 0.05). No increase was found for VEGF(P gt; 0.05). ③ There was positive correlation between the platelet concentration and PDGF-BB and TGF-β1 (r = 0.627, r = 0.437, P lt; 0.05). ④ Thirteen diabetic repractory dermal ulcers received APG treatment for 18 times, 9 ulcers (69.2%) and 10 sinuses (88.3%) were cured at the end of 12-week treatment. Conclusion The method ofgroup B is the best centrifuge method. A variety of growth factors are detected and released from the platelets at significant levels in APG. There is positive correlation between the platelet concentration and PDGF-BB and TGF-β1 .
Objective To introduce the basic research and cl inical appl ication of the injectable bone repair biomaterials. Methods The recent original articles about the injectable bone repair biomaterials were extensively reviewed. Results The injectable bone repair biomaterials could fill irregularly shaped defects and might allow bone augmentation, both with minimal surgical intervention, and the injectable bone repair material had a good prospect by the medical profession and attach great importance to the academic material, but there were some deficiencies and shortcomings. Conclusion The injectable bone repair biomaterials may be a future approach to repair bone defect．