Objective To investigate the effect of homograft of marrow mesenchymal stem cells (MSCs) seeded onto poly-L-lactic acid (PLLA)/gelatin on repair of articular cartilage defects. Methods The MSCs derived from36 Qingzilan rabbits, aging 4 to 6 months and weighed 2.5-3.5 kg were cultured in vitroand seeded onto PLLA/gelatin. The MSCs/ PLLA/gelatin composite was cultured and transplanted into full thickness defects on intercondylar fossa. Thirty-six healthy Qingzilan rabbits were made models of cartilage defects in the intercondylar fossa. These rabbits were divided into 3 groups according to the repair materials with 12 in each group: group A, MSCs and PLLA/gelatin complex(MSCs/ PLLA/gelatin); group B, only PLLA/gelatin; and group C, nothing. At 4,8 and 12 weeks after operation, the gross, histological and immunohistochemical observations were made, and grading scales were evaluated. Results At 12 weeks after transplantation, defect was repaired and the structures of the cartilage surface and normal cartilage was in integrity. The defects in group A were repaired by the hylinelike tissue and defects in groups B and C were repaired by the fibrous tissues. Immunohistochemical staining showed that cells in the zones of repaired tissues were larger in size, arranged columnedly, riched in collagen Ⅱ matrix and integrated satisfactorily with native adjacent cartilages and subchondral bones in group A at 12 weeks postoperatively. In gross score, group A(2.75±0.89) was significantly better than group B (4.88±1.25) and group C (7.38±1.18) 12 weeks afteroperation, showing significant differences (P＜0.05); in histological score, group A (3.88±1.36) was better than group B (8.38±1.06) and group C (13.13±1.96), and group B was better than group C, showing significant differences (P＜0.05). Conclusion Transplantation of mesenchymal stem cells seeded onto PLLA/gelatin is a promising way for the treatment of cartilage defects.
Objective To prepare a self-made compound, hemostatic jelly with polylactic acid(PLA), which has the hemostatic and absorbable effect on injured cancellous bone. Methods Two bone defects of 5 mm in diameter and 4 mm in depth were subjected on 20 health rabbits by drilling through their either outside plate of the iliac, and were filled with hemostatic jelly(group A), bone wax(group B) and blank(group C) respectively. Hemostasis were observed and recorded after 1 and 10 minutes. Five specimens were harvested at 2, 4, 8 and 12 weeks postoperatively for histological observation. Results ① Hemostatic effect: Bleeding of injured spongy bone stopped within 10 minutes after the treatment of hemostatic jelly and bone wax, but bleeding of balnk did not stop. Hemostatic jelly and bone wax adhered to bone defects firmly within 10 minutes was after the treatment. ② Absorbable effect: Hemostatic jelly and bone defects have not changed visibly in the first 2 weeks. With histological observation 4 to 8 weeks after the operation, hemastatic jelly was absorbed gradually and replaced by osteogenous tissue. It was absorbed completely after 8 to 12 weeks. Bone wax was not absorbed after 12 weeks, no new bone tissue was observed at bone wax area. The blank was replaced by connective tissue and osteogenous tissue partially after 12 weeks. Conclusion The compound hemostatic jelly manifests both hemostatic and absorbable effects on injured cancellous bone and may substitute for bone wax in clinical application.
Objective To study the result of using nerve conduit coated with chitin and filled with a guide-fiber to repair peripheral nerve defect. Methods Twenty-four female adult SD rats were made the model of 14 mm-gap on bilateral sciatic nerve under sterile condition. The rats were randomly divided into 4 groups(n=6),group A: polymer polyglycolic-lactic acid(PGLA) nerve conduit coated with chitin and filled with a guide-fiber as experimental group to repair 14 mm gap of rat sciatic nerve；group B: PGLA nerve conduit coated with chitin； group C: PGLA nerve conduit； group D: autograft (control group). The repair result was evaluated by normal observation, EMG testing and S-100 histological immunostaining analysis 4 and 12 weeks after operation.Results Four weeks after the operation，there were new regenerated immature fibers in groups A,B and C, 12 weeks after the operation, the regenerated nerve fibers were seen to have bridged the gap. There were myelinated fibers equably distributed and rarely newgenerated nerve fibers in distal parts of group D. The repair result of PGLA nerve conduit coated with a chitin and filled with guide-fiber was better than that of groups B and C（Plt;0.05）. There was significant difference of nerve fiber diameter，thickness of myelin sheath and fiber density in group D from those in groups A, B and C（Plt;0.05），but there were degenerative changes such as vacuoles insheaths and myelin separation in proximal and few new regenerated nerve fibers in distal parts of group D. Conclusion PGLA nerve conduit coated with chitin and filled with a guide-fiber offers a possible substitute for the repair of peripheral nerve defect.
Objective To study the mechanism of ectopic osteogenesis of nacre/Polylactic acid (N/P) artificial bone combined with allogenic osteoblasts, and to explore the possibility as a scaffold material of bone tissue engineering. Methods The allogenic- osteoblasts seeded onto N/P artificial bone were co-cultured in vivo 1 week.The N/P artificial bone with allogenic osteoblasts were implanted subcutaneously into the left back sites of the New Zealand white rabbits in the experimental group and the simple N/P artificial bone into the right ones in the control group. The complexes were harvested and examined by gross observation, histologic analysis and immunohistochemical investigation 2, 4 and 8 weeks after implantation respectively.Results In experimental group, the osteoid formed after 4 weeks, and the mature bone tissue withbone medullary cavities formed after 8 weeks; but in control group there was nonew bone formation instead of abundant fibrous tissue after 4 weeks, and more fibrous tissue after 8 weeks.Conclusion N/P artificial bone can be used as an optical scaffold material of bone tissue engineering.
摘要:目的:研究生物降解聚DL乳酸(PDLLA)自锁式捆绑带固定骨折的生物力学性能。方法:80只新西兰大白兔随机分为两组,建立股骨干非负重骨折动物模型,应用生物降解自锁式捆绑带固定骨折为实验组,钢丝固定骨折为对照组,分别于术后1、4、8、12周行生物力学检查进行比较。结果:捆绑带组在术后4、8、12周均比钢丝组的弯曲强度高,但4周、12周时Pgt;005,无统计学差异,8周时Plt;005,提示有统计学差异。离体同种固定物不同时间段抗拉强度自身比较:钢丝固定术后4阶段抗拉强度比较Pgt;005,任何两两比较都没有统计学差异,抗拉强度未随术后时间延长发生明显下降。捆绑带固定术后4周与术后1周比较Pgt;005,抗拉强度无明显降低,但术后8周和术后12周时Plt;005,抗拉强度明显下降。结论:生物降解自锁式捆绑带在非负重骨折治疗中可发挥良好的固定作用。生物降解自锁式捆绑带降解时,应力传导促进了骨折的愈合。Abstract: Objective: To study the biomechanics function of selflocking cerclage band made of biodegradable material polyDLlactic acid (PDLLA) in the fixation of fractures. Methods: Eighty rabbits were divided into two groups. Femur fracture models were made. Fractures were fixed using biodegradable selflocking cerclage band in experimental group and metal fixation material in control group. The biomechanics was analyzed and compared after 1, 4, 8 and 12 weeks respectively． Results: The bending strength of experimental group is more ber than that of control group after 4, 8 and 12 weeks, but it was not statistically significant at 4 and 12 weeks (Pgt;005). It was statistically significant at 8 weeks (Plt;005). The tensile strength of the same cerclage instrument was compared at different stage in vitro, and the result of the control group was not statistically significant at the four stage (〖WTBX〗P〖WTBZ〗gt;005). Regarding the changes of tensile strength of the cerclage instrument at different stage, the result of the experimental group was not statistically significant after 1 and 4 weeks (Pgt;005). However, the decrease of tensile strength was statistically significant after 8 and 12 weeks (Plt;005). Conculsion: Biodegradable selflocking cerclage band could be used in thetreatment of nonweightbearing fractures. The stress force conducting promotes healing of fracture when the selflocking biodegradable cerclage band degrades.
Objective To investigate the feasibility of fetal liver cells for liver tissue engineering, the supporting function of poly L lactic acid (PLLA) scaffold for fetal liver cells and the effects of oncostatin M (OSM), nicotinamide (NA) and dimethyl sulfoxide(DMSO) on growth and hepatic differentiation. Methods After three dimensional PLLA scaffolds having a porous structure were prepared by using NH 4HCO 3 particle, fetal liver cells obtained from E14.5 C57BL/6CrSlc murine embryos were inoculated in the scaffolds. Cells were cultured in Williams’E medium with or without OSM, NA and DMSO for 30 days. Changes in cell number, liver-specific function, and cellular morphology were observed. Results When compared with in monolayer culture, cell number and albumin secretion increased obviously in three-dimensional PLLA. Alburmin secretion increased slightly in OSM group of monolayer culture, but increased obviously in OSM groupo of PLLA culture and in OSM/NA/DMSO group of both monlayer and PLLA cultures. Conclusion The three-dimensional PLLA scaffold is a good supporting material for the cultivation of tetal liver cells. OSM, NA and DMSO remarkaly stimulated maturation of hepatic parenchymal cells in vitro in terms of morphology and liver-specific function.
Objective To explore the biocompatibility of poly(lacticacid/glycolic acid/asparagic acid-co-polyethylene glycol) biomaterials (PLGA-ASP-PEG) and biological behaviors of cultured marrow stroml stem cells (MSCs) combined with this new type of scaffold in tissue engineering. Methods The PLGA-ASP-PEG tri-block copolymers were obtained through bulk ringopening copolymerization method.MSCs were isolated from the bone marrow of 4 week old New Zealand rabbits. The 3rdgeneration MSCs were cultured combining with PLGA-ASP-PEG in vitro, while cells cultured in PLGA as control group. The cell adhesion rate and the adhesivepower were examined by conventional precipitation method and micropipette aspiration technique respectively. The morphological features were studied by scanning electron microscope. The proliferation behavior of the cells was analyzed by MTT assay. The cell cycle, proliferation index, DNA index and apoptosis of the cells were detected by flow cytometry. The synthesis of protein and collagen were examined by Coomassie Brilliant Blue dyes and 3H-Proline incorporation test. Results The MSCs adhered and grew well on the surface of the biomaterial PLGA-ASP-PEG. The powers of cell adhesion, proliferation and protein and collagen synthesis of the cells were all significantly higher than those of PLGA group (P＜0.05), but the apoptosis rate was significantly lower than that of PLGA group (P＜0.05). The DNA indexes showed the cells of both PLGA-ASP-PEG group and PLGAgroup were normal diploid cells. Conclusion PLGA-ASP-PEG showedgood biocompatibilityand the biological properties improved greatly compared with the PLGA scaffold materials. These results demonstrated that the promise of PLGAASPPEG canbe used as an ideal scaffold material for construction of tissue engineered bone to restore bone defects in bone tissue engineering．
ObjectiveTo investigate the effect of silk fibroin-poly-L-lactic acid (SF-PLLA) microcarriers on the expansion and differentiation of adipose-derived stem cells (ADSCs).MethodsADSCs were extracted from adipose tissue donated voluntarily by patients undergoing liposuction by enzymatic digestion. The 3rd generation ADSCs were inoculated on CultiSpher G and SF-PLLA microcarriers (set up as groups A and B, respectively), and cultured in the rotary cell culture system. ADSCs cultured in normal two-dimensional plane were used as the control group (group C). Scanning electron microscope was used to observe the microcarriers structure and cell growth. Live/Dead staining and confocal fluorescence microscope was used to observe the distribution and survival condition of cells on two microcarriers. DNA quantification was used to assess cell proliferation on two microcarriers. Real-time fluorescence quantitative PCR (qRT-PCR) was used to detect chondrogenesis, osteogenesis, and adipogenesis related gene expression of ADSCs in 3 groups cultured for 18 days. Flow cytometry was used to identify the MSCs surface markers of ADSCs in 3 groups cultured for 18 days, and differential experiments were made to identify differentiation ability of the harvested cells.ResultsADSCs could be adhered to and efficiently amplified on the two microcarriers. After 18 days of cultivation, the total increment of ADSCs of the two microcarriers were similar (P>0.05). qRT-PCR results showed that chondrogenesis related genes (aggrecan, cartilage oligomeric matrix protein, SOX9) were significantly up-regulated for ADSCs on SF-PLLA microcarriers and adipogenesis related genes (peroxisome proliferator-activated receptor γ, lipoprotein lipase, ADIPOQ) were significantly up-regulated for ADSCs on CultiSpher G microcarriers, all showing significant differences (P<0.05). Flow cytometry and differentiation identification proved that the harvested cells of the two groups were still ADSCs.ConclusionThe ADSCs can be amplified by SF-PLLA microcarriers, and the chondrogenic differential ability of harvested cells was up-regulated while the adipogenic differential was down-regulated.
Objective To fabricate a novel porous bioactivecomposite biomaterial consisting of poly lactic acid (PLA)bone matrix gelatin(BMG) by using the supercritical carbon dioxide fluid technique (SC-CO2) and to evaluate its osteoinductive activity. Methods The cortical bones selected from healthy adult donors were processed into BMG by the defatting, demineralizing, and deproteinizing processes. PLA and BMG were mixed at a volume radio of 3∶1; then, the PLA-BMG mixed material and the pure PLA material were respectively placed in the supercritical carbon dioxide reaction kettles, and were respectively added by the NaCl particles 100200 μm in diameter for theporosity of the materials so that the porous PLA-BMG composite material and the porous PLA composite material could be formed. The mouse osteoblastlike MC3T3-E1 cells were cultured in the dulbecco’s modified eagle medium (DMEM) supplemented with 10% fetal bovine serum. Then, 20 μl of the MC3T3E1 cell suspensions containing 2 ×106 cells /ml were delivered into the culturing plate (24 wells/plate) made of the different materials, which were co-cultured for 2 weeks. In the PLA-BMG group, 100 μg of the crushed PLA-BMG material was contained in each well; in the PLA group, 100 μg of the crushed PLA material was containedin each well; and in the DMEM group, only DMEM was contained, which served as the control group. There were 6 wells in each group. The quantitative analysis onthe calcification area was performed by the staining of the alizarin red S. Theco-cultured cells were harvested and lysated in 1 ml of 0.2% Nonidet P-40 by the ultrasonic lysating technique. Then, the ALP activity and the Ca content were measured according to the illuminations of the reagent kits. Results The porous PLABMG composite material showed a good homological porosity with a pore diameter of 50-150 μm and a good connectivity between the pores. The ALP activity, the Ca content, and the calcification area were significantly greater in the PLABMG group than in the PLA group and the control group (325.59±70.40 U/gprot, 3.51±1.64 mmol/gprot, 42.98±4.44% vs. 63.62±30.01 U/gprot, 1.04±0.21 mmol/gprot, 9.55±1.94%, and 2.40±1.47 U/gprot, 0.70±0.24 mmol/gprot, 0.86±0.41%; Plt;0.05). Meanwhile, there was a statistically significant difference between the PLA group and the control group in the ALP activity and the calcification area (Plt;0.05). Conclusion The porous PLABMG composite material prepared by the use of SC-CO2 has a good steoinductive activity and can be used as a promising bone biomaterial and a bone tissue engineered scaffold.
The poly-lactic acid membrane is characterized by its non-toxicity, non-carcinogenic and the property of degradation in the body in animal experiment, this membrane was wrapped around the tenorrhaphy site. The results showed that 3 to 9 weeks after operation there was an intact space surrounding the tendon, and the membrane began to degradate at 24 weeks, might achieve the purpose to prevent the formation of adhesion. From Nov. 1985 to Sept. 1989, this membrane was applied to 20 tendons of 14 patients. The follow-up results showed that the satisfactory rate was 81.8% .the indications of using poly-lacic acid membrane were primary, delayed primary and secondary suture of flexor tendon injuries in areas of the fingers.