OBJECTIVE To explore the effect of basic fibroblast growth factor (bFGF) combined with autogenous vein graft conduit on peripheral nerve regeneration. METHODS Fifty four New Zealand rabbits were divided into three groups. The main trunk of sciatic nerve of rabbit in one side was severed and bridged by autogenous vein. 0.2 ml bFGF solution (4,000 U/ml) was intravenously injected to the vein graft conduit as group A, the same amount of saline solution as group B, and no solution injection as group C. Microscopic examination, axon video analysis and nerve conduct velocity were performed at the 10th, 30th, and 100th day after operation. RESULTS The nerve fibers were grown into vein graft conduit in all groups at 30th after operation, they were more and regular in group A than that of group B and C, and the axon regeneration rate in group A was more than that of group B and C. CONCLUSION bFGF combined with autogenous vein graft conduit can markedly promote nerve regeneration.
Objective To explore the effects of exogenous basic fibroblast growth factor (bFGF) on insheathed tendon healing and adhesion formation. Methods Ninety Leghorn chickens were randomly divided into 3 groups (groups A, B and C), 30 animals for each group, and the right third digitorum longus tendon of the chicken was transected to make defect models. In group A, the tendon was sutured in situ after transection. In group B, the tendon was sutured after 0.6 μl fibrin sealant (FS) was applied at repair site. In group C, the tendon was sutured after 0.6 μl FS mixed with 500 ng bFGF was appliedat repair site. At 1, 2, 4 and 8 weeks after operation, the tendons of 6 chickens in each group were harvested for morphological and histological evaluation. Six specimens of each group was obtained for biomechanical test at 8 weeks. Results The gross observation showed that the differences of grading of tendon adhesion were not significant between groups A, B, and C 8 weeks after operation(Pgt;0.05). Histological evaluation showedthat there were no significant differences in fibroblast counting and the content of collagen fibers between groups A and B(P＞0.05). The angiogenesis, fibroblast proliferation and collagen production in the sheath, epitendon and parenchyma at repair site in group C occurred earlier and were more than those in groups A and B, showing significant differences (Plt;0.05). The biomechanical tests showed that the gliding excursionof the tendon in group A, B and C were 3.44±0.43、3.51±0.56 and 2.84±0.42 mm respectively; the work of flexion were 14.87±1.72、14.08±1.85 and 20.62±3.52 Nmm respectively; the ultimate tensile strength of the tendon was10.34±1.45，11.26±1.83 and 15.02±2.20 N respectively; showing no significant differences between groups A and B(Pgt;0.05), but showing significant differences between group C and groups A, B(Plt;0.05). Conclusion The exogenous bFGF at tendon repair site can facilitate insheathed tendon healing, but also increase the tendon adhesion formation.
Objective To review the recent researches of basic fibroblast growth factor (bFGF) in tendon tissue engineering. Methods Recentoriginal related literature was extensively reviewed and analyzed. Results bFGF played an important role in establishing standard tendon tissue engineering cell lines, inducing the compound and analysis of extracellular matrix, enhancing interactions between cells and extracellular matrix and accelerating tissue engineering materials’ neovascularization. Conclusion The progresses in increasing endogenetic bFGF expression, controlling the release of exogenous bFGF and improving the bioutilization of bFGF has laid foundation for wider use of bFGF in tendon tissue engineering.
Objective To investigate the effects of the insulin-like growth factor 1 (IGF-1), the transforming growth factor β1(TGFβ1), and the basic fibroblast growth factor (bFGF) on proliferation and cell phenotype of the human fetal meniscal cells, and to find out the best combination and concentration of the growth factors for the meniscus tissue engineering. Methods The fetus came from the healthy woman accidental abortion and the procedure had got her approval.The human fetal meniscal fibrochondrocytes were cultured in vitro. The cell phenotype was identifiedby the collagen type Ⅱ immunohistochemistry and Aggrecan immunofluorescence. Inthe growth factor groups, the 3rd passage meniscal cells synchronized by the serum starvation method and were mixed with IGF-1 (1, 10, 50, 100 μg/L), TGF-β1 (0.1, 1.0, 5.0, 10.0, 50.0 μg/L), and bFGF (5, 10, 50, 100, 200 μg/L), respectively, and in the combination groups, the combinations of bFGF and TGF-β1, bFGF and IGF-1, TGF-β1 and IGF-1 were established at their optimal effect concentrations. The control group was also established for comparison. The dose-response relationship was studied at 48 h and 72 h bythe MTT colorimetric method. Results The 3rd passage meniscalcells could express collagen type Ⅱ and Aggrecan before and after the addition of the three growth factors. The proliferating effects of the growth factors (IGF-1 50 μg/L，TGF-β1 5 μg/L，bFGF 50 μg/L) on the 3rd passage cells at 48 h and 72 h were significantly better in the growth factor groups than in the control group (Plt;0.05)，and the combination groups of bFGF 50 μg/L and IGF-1 50 μg/L, IGF-1 50 μg/L and TGF-β1 5 μg/L showed a significantly higher proliferatingeffect than that in the single growth factor group (Plt;0.05). bFGF 50 μg/L and TGF-β1 5 μg/L had no synergetic effect (Pgt;0.05). Conclusion IGF-1, TGF-β1 and bFGF can promote the proliferation of the human fetal meniscal cells, respectively, and the combinations of bFGF and IGF-1, IGF-1 and TGF-β1 at their optimal concentrations can have better proliferating effects than the single growth factor. They can be used for the in vitro amplification of the meniscal seed cells.
Objective To observe the proliferation and migration of endothelial cells after 30% total burn surface area (TBSA) of deep partial thickness scald, and the effect of basic fibroblast growth factor (bFGF) on angiogenesis during wound healing.Methods A total of 133 male Wistar ratswere divided randomly into normal control (n=7), injured control group (n=42), bFGF group (n=42) andanti-c-fos group (n=42). The apoptosis expression of fibroblasts was determinedwith in situ hybridization and the changes of proliferation cell nuclear antigen(PCNA), focal adhesion rinase(FAK), c-fos and extracellular signalregulated kinase(ERK) proteins expression were detected with immunohistochemistry staining technique after 3 hours, 6 hours, 1 day, 3 days, 7 days, 14 days and 21 days of scald.Results In injured control group and bFGF group, theproliferation rate of the vascular endothelial had evident changes 7 days and14 days after scald； the expression of FAK was increased 14 days after scald. ERK proteins expression was different between injury control group and bFGF group at initial stage after scald. Stimulation of ERKs by bFGF led to up-regulation of c-fos and b expression of FAK. Conclusion Exogenous bFGF extended the influence on wound healing process by ERK signaling pathway, affecting migration cascade of vascular endothelial cell. The oncogene proteins play an important role on accelerating angiogenesis duringwound healing.
Objective To evaluate the effect of composite (bFGF/PDPB) of basic fibroblast growth factor（bFGF） and partially deproteinized bone (PDPB) on the repair of femoral head defect. Methods Forty-eight femoral heads with defect derived from 24 New Zealand rabbits were divided into 3 groups at random, which were implanted with bFGF/PDPB(group A), PDPB(group B) and nothing(group C) respectively.The rabbits were sacrificed at 2,4,and8 weeks after operation, and then the femoral heads were obtained. The specimens injected with Chinese ink were created. Then X-ray examination, histopathological and morphological examination of blood vessel, and image analysis were made. Results The bone defects healed completely 8 weeks after operation in group A. The implants in the repaired tissue were not substituted completely in group B. The bone defects did not heal completely in group C. Two weeks after operation, affluent newly formed vessels were seen in repaired areas in groupA. No significant difference between group A and group B was observed 8 weeks after operation. In group C, newly formed vessels were scarce 2, 4, and 8 weeks after operation. There were 3 sides rated excellent, 2 good and 1 fair in group A; 1 excellent, 2 good, 2 fair and 1 poor in group B; and 1 fair and 5 poor in group C according to the X-ray evaluation 8 weeks after operation. Eight weeks after operation, the volume fraction of bone trabecula in repaired tissue was higher in group A than that in group B (Plt;0.05), and the fraction in group C was thelowest among the 3 groups (Plt;0.05). Conclusion The composite ofbFGF and PDPB can effectively promote the repair of femoral head defect of rabbit.
Objective To explore the impact of basic fibroblast growth factor (bFGF) and parathyroid hormone-related protein (PTHrP) on early and late chondrogenic differentiation of rabbit bone marrow mesenchymal stem cells (BMSCs) induced by transforming growth factor β1 (TGF-β1). Methods BMSCs were isolated from 3 healthy Japanese rabbits (2-month-old, weighing 1.6-2.1 kg, male or female), and were clutured to passage 3. The cells were put into pellet culture system and were divided into 5 groups according to different induce conditions: TGF-β1 group (group A), TGF-β1/bFGF group (group B), TGF-β1/21 days bFGF group (group C), TGF-β1/PTHrP group (group D), and TGF-β1/21 days PTHrP group (group E). At the beginning, TGF-β1 (10 ng/mL) was added to all groups, then bFGF and PTHrP (10 ng/mL) were added to groups B and D respectively; bFGF and PTHrP (10 ng/mL) were added to groups C and E at 21 days respectively. The gene expressions of collagen type I (Col I), Col II, Col X, matrix metalloproteinases (MMP)-13, and alkaline phosphatase (ALP) activity were detected once every week for 6 weeks. The 1, 9-dimethylmethylene blue (DMMB) staining was used to observe the extracellular matrix secretion at 6 weeks. Results The expression of Col I in groups C and E showed a significant downward trend after 3 weeks; the expression in group A was significantly higher than that in groups C and E at 4 and 5 weeks (P lt; 0.05), and than that in groups B and D at 3-6 weeks (P lt; 0.05); and significant differences were found between groups B and C at 3 and 4 weeks, and between groups D and E at 3 weeks (P lt; 0.05). After 3 weeks, the expressions of Col II and Col X in groups C and E gradually decreased, and were significantly lower than those in group A at 4-6 weeks (P lt; 0.05). Groups B and D showed no significant difference in the expressions of Col II and Col X at all time points, but there was significant difference when compared with group A (P lt; 0.05). MMP-13 had no obvious expression at all time points in group A; significant differences were found between group B and groups A, C at 3 weeks (P lt; 0.05); and the expression was significantly higher in group D than in groups A and E (P lt; 0.05). ALP activity gradually increased with time in group A; after 4 weeks, ALP activity in groups C and E obviously decreased, and was significantly lower than that in group A (P lt; 0.05); there were significant differences between groups B and C, and between groups D and E at 2 and 3 weeks (P lt; 0.05). DMMB staining showed more cartilage lacuna in group A than in the other groups at 6 weeks. Conclusion bFGF and PTHrP can inhibit early and late chondrogenic differentiation of BMSCs by changing synthesis and decomposition of the cartilage extracellular matrix. The inhibition is not only by suppressing Col X expression, but also possibly by suppressing other chondrogenic protein.
Objective To study the effect of two cytokines, basic fibroblast growth factor(bFGF) and insulin-like growth factor-I(IGF-I), on cell proliferation in chondrocytes of adult rabbits. Methods The primary chondrocytes of adult rabbits were harvested and cultured with bFGF and IGF-I at different concentrations,respectively, as well as with the mixture of the two cytokines; the quantity of cultured chondrocytes was detected by MTT assay at the 24th, 48th and 72th hours; and the final fold increase of different groups was measured by cell count for the 3rd passage; and the proliferation index of the groups was recorded by flowing cytometer on the 14th day. Results ① The cultured chondrocytes with either bFGF, IGF-I or their mixture were significantly more than that of control group at the 24th, 48th and 72th hours (P＜0.01). ② After the 3rd passage, the final folds of proliferation were significantly higher in the groups with cytokinesthan in the control group (P＜0.01); and the final fold with the mixture ofcytokines was significantly higher than that of both IGF-I and bFGF (P＜0.01). ③ Theproliferation index was significantly higher in the groups with cytokines than in the control group (P＜0.01); the proliferation index with the mixture of cytokines was significantly higher than that of both IGF-I and bFGF (P＜0.05); besides, proliferation index was higher when cytokine was applied twice than once (P＜0.05). Conclusion bFGF and IGF-I could promote chondrocytes proliferation of adult rabbits obviously and they are synergistic in cell proliferation.
【Abstract】 Objective Sonic hedgehog (Shh) signaling pathway is involved in an important part of regulating angiogenesis. To investigate the effects of recombinant Shh N-terminant (rShh-N) on the expression and secretion of angiogenesis-related factor—vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF). Methods Bone marrow mesenchymal stem scells (BMSCs) were isolated from 3-day-old healthy Sprague Dawley rats and cultured to passage 3 in vitro. rShh-N at the concentrations of 0, 10, 100, and 200 ng/mL were applied to culture BMSCs in groups A, B, C, and D, respectively. At 12, 24, 48, and 72 hours of culture, the expressions of VEGF and bFGF mRNA and the levels of VEGF and bFGF in supernatant were measured with real-time quantitative PCR and ELISA, respectively. Results At the gene level, compared with group A, the expressions of VEGF and bFGF mRNA were enhanced in group D (P lt; 0.05) and the upregulation was more significant at 12 and 48 hours than 24 and 72 hours (P lt; 0.01). In group C, bFGF mRNA expression was substantially promoted at 12-72 hours (P lt; 0.05) and VEGF mRNA level was upregulated at 24-72 hours (P lt; 0.05), and both reached peak at 72 hours (P lt; 0.01). In group B, VEGF mRNA expression was inhibited at 12 hours (P lt; 0.05), but the level increased at 48 and 72 hours (P lt; 0.05); bFGF mRNA expression was obviously promoted at 12-48 hours (P lt; 0.05) and the maximum appeared at 48 hours (P lt; 0.01). At the protein level, the secretion of VEGF and bFGF in group D was significantly increased at 12-72 hours, as compared with group A (P lt; 0.05). In group C, VEGF and bFGF secretion was increased at 24-72 hours (P lt; 0.05). The secretion of VEGF in group B was inhibited at 12 and 48 hours (P lt; 0.05) and was promoted at 24 hours (P lt; 0.05); bFGF secretion was up-regulated at 24 and 48 hours (P lt; 0.05). The secretion of VEGF and bFGF in supernatant at 【Abstract】 Objective Sonic hedgehog (Shh) signaling pathway is involved in an important part of regulating angiogenesis. To investigate the effects of recombinant Shh N-terminant (rShh-N) on the expression and secretion of angiogenesis-related factor—vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF). Methods Bone marrow mesenchymal stem scells (BMSCs) were isolated from 3-day-old healthy Sprague Dawley rats and cultured to passage 3 in vitro. rShh-N at the concentrations of 0, 10, 100, and 200 ng/mL were applied to culture BMSCs in groups A, B, C, and D, respectively. At 12, 24, 48, and 72 hours of culture, the expressions of VEGF and bFGF mRNA and the levels of VEGF and bFGF in supernatant were measured with real-time quantitative PCR and ELISA, respectively. Results At the gene level, compared with group A, the expressions of VEGF and bFGF mRNA were enhanced in group D (P lt; 0.05) and the upregulation was more significant at 12 and 48 hours than 24 and 72 hours (P lt; 0.01). In group C, bFGF mRNA expression was substantially promoted at 12-72 hours (P lt; 0.05) and VEGF mRNA level was upregulated at 24-72 hours (P lt; 0.05), and both reached peak at 72 hours (P lt; 0.01). In group B, VEGF mRNA expression was inhibited at 12 hours (P lt; 0.05), but the level increased at 48 and 72 hours (P lt; 0.05); bFGF mRNA expression was obviously promoted at 12-48 hours (P lt; 0.05) and the maximum appeared at 48 hours (P lt; 0.01). At the protein level, the secretion of VEGF and bFGF in group D was significantly increased at 12-72 hours, as compared with group A (P lt; 0.05). In group C, VEGF and bFGF secretion was increased at 24-72 hours (P lt; 0.05). The secretion of VEGF in group B was inhibited at 12 and 48 hours (P lt; 0.05) and was promoted at 24 hours (P lt; 0.05); bFGF secretion was up-regulated at 24 and 48 hours (P lt; 0.05). The secretion of VEGF and bFGF in supernatant at
OBJECTIVE To investigate the effects of basic fibroblast growth factor(bFGF) on repairing transected sciatic nerves in rats. METHODS The animal models of the transected sciatic nerve of 40 SD rats were established, which divided into 4 groups: normal saline (NS) group, nerve growth factor (NGF) group, bFGF group and normal control group. The epineurium of the transected sciatic nerve was sutured under microscope, then bFGF or NGF was dropped into local sites and injected intramuscularly once a day for 30 days after operation. Functional repair for the transected sciatic nerves was studied by nerve conductive velocity (NCV) and sciatic nerve function index (SFI). RESULTS As a criterion, the level of the normal control group was regarded as zero, SFI of NS group, NGF group and bFGF group were -114.30 +/- 10.34, -70.50 +/- 11.01, -50.45 +/- 7.82 respectively at 1 month after operation, and they were -54.96 +/- 16.46, -35.21 +/- 10.80, -27.53 +/- 11.23 respectively in 3 months after operation. NCV of bFGF group was significantly faster than NS group and NGF group. CONCLUSION bFGF can significantly promote the functional repair of injured peripheral nerve, and its effects are better than NGF.