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1.
Deng C  Tian H  Zhang P  Sun J  Chen X  Jing X 《Biomacromolecules》2006,7(2):590-596
Advances in tissue engineering require biofunctional scaffolds that can provide not only physical support for cells but also chemical and biological cues needed in forming functional tissues. To achieve this goal, a novel RGD peptide grafted poly(ethylene glycol)-b-poly(L-lactide)-b-poly(L-glutamic acid) (PEG-PLA-PGL/RGD) was synthesized in four steps (1) to prepare diblock copolymer PEG-PLA-OH and to convert its -OH end group into -NH(2) (to obtain PEG-PLA-NH(2)), (2) to prepare triblock copolymer PEG-PLA-PBGL by ring-opening polymerization of NCA (N-carboxyanhydride) derived from benzyl glutamate with diblock copolymer PEG-PLA-NH(2) as macroinitiator, (3) to remove the protective benzyl groups by catalytic hydrogenation of PEG-PLA-PBGL to obtain PEG-PLA-PGL, and (4) to react RGD (arginine-glycine-(aspartic amide)) with the carboxyl groups of the PEG-PLA-PGL. The structures of PEG-PLA-PGL/RGD and its precursors were confirmed by (1)H NMR, FT-IR, amino acid analysis, and XPS analysis. Addition of 5 wt % PEG-PLA-PGL/RGD into a PLGA matrix significantly improved the surface wettability of the blend films and the adhesion and proliferation behavior of human chondrocytes and 3T3 cells on the blend films. Therefore, the novel RGD-grafted triblock copolymer is expected to find application in cell or tissue engineering.  相似文献   

2.
Cell therapy combined with biomaterial scaffolds is used to treat cartilage defects. We hypothesized that chondrogenic differentiation bone marrow-derived mesenchymal stem cells (BM-MSCs) in three-dimensional biomaterial scaffolds would initiate cartilaginous matrix deposition and prepare the construct for cartilage regeneration in situ. The chondrogenic capability of human BM-MSCs was first verified in a pellet culture. The BM-MSCs were then either seeded onto a composite scaffold rhCo-PLA combining polylactide and collagen type II (C2) or type III (C3), or commercial collagen type I/III membrane (CG). The BM-MSCs were either cultured in a proliferation medium or chondrogenic culture medium. Adult human chondrocytes (ACs) served as controls. After 3, 14, and 28 days, the constructs were analyzed with quantitative polymerase chain reaction and confocal microscopy and sulfated glycosaminoglycans (GAGs) were measured. The differentiated BM-MSCs entered a hypertrophic state by Day 14 of culture. The ACs showed dedifferentiation with no expression of chondrogenic genes and low amount of GAG. The CG membrane induced the highest expression levels of hypertrophic genes. The two different collagen types in composite scaffolds yielded similar results. Regardless of the biomaterial scaffold, culturing BM-MSCs in chondrogenic differentiation medium resulted in chondrocyte hypertrophy. Thus, caution for cell fate is required when designing cell-biomaterial constructs for cartilage regeneration.  相似文献   

3.
Cartilage-derived growth factor purified to homogeneity by affinity chromatography on columns of heparin-Sepharose was mitogenic for early passage bovine fetal chondrocytes. Hyaluronate and sulfated glycosaminoglycan synthesis in these cells was analyzed by differential enzymatic digestion of the glycosaminoglycans labeled with [14C] glucosamine or [35S]. It was found that chondrocyte proliferation was accompanied by about a four-fold increase in hyaluronate synthesis over a two-day period, while the synthesis of sulfated glycosaminoglycans decreased by about 2-fold. Chromatographic analysis of the sulfated glycosaminoglycans showed decreases in chondroitin 4 and 6 sulfates. It was concluded from these results that cartilage-derived growth factor was a proliferative factor for chondrocytes and differed from the somatomedins.  相似文献   

4.
5.
In vitro cultivation of primary bovine knee chondrocytes (BKCs), using bovine pituitary extract (BPE) and porous scaffolds composed of polyglycolide (PGA) and 85/15 poly(lactide-co-glycolide) (PLGA), was investigated. Here, BPE was prepared from fresh bovine pituitaries, and cylindrical PGA/PLGA scaffolds with various chemical compositions were fabricated by solvent merging/particulate leaching method. Experimental results showed that in microcarrier systems, the rate of BKC growth on PGA surfaces is faster than that on PLGA surfaces, and the decrease in the medium pH value of BKCs-adsorbed PGA particles is faster than that of BKCs-adsorbed PLGA particles. After 28-day construct cultivation, the BKC amount and the content of glycosaminoglycans and collagen per construct increased with BPE protein concentration. For a constant BPE protein concentration, a higher PGA percentage in scaffold leads to a better biological environment for the growth of BKCs and the synthesis of extracellar matrices.  相似文献   

6.
Cultivation of bovine knee chondrocytes (BKCs) in various cationic additives was studied using chitosan-gelatin scaffolds, whose surfaces were modified by human fibronectin (HFN). Here, the genipin-crosslinked scaffolds were fabricated by the freezing/lyophilization method with various concentrations of the precursory gels. The experimental results indicated that a lower freezing temperature led to higher moisture content, porosity, and specific surface area of a scaffold. The higher the precursor concentration, the larger the moisture content of a scaffold. A fast biodegradation of scaffold matrix was generated by a high porosity with BKCs. A higher concentration of HFN coated on scaffold surfaces yielded a faster rate of BKC attachment from the culture medium. The amounts of BKCs, glycosaminoglycans, and collagen over 28-day cultivation increased with the scaffold porosity, the coating concentration of HFN, the seeding density of BKCs, and the calcium concentration in medium.  相似文献   

7.
Bone tissue engineering offers promising alternatives to repair and restore tissues. Our laboratory has employed poly(lactide-co-glycolide) PLAGA microspheres to develop a three dimensional (3-D) porous bioresorbable scaffold with a biomimetic pore structure. Osseous healing and integration with the surrounding tissue depends in part on new blood vessel formation within the porous structure. Since endothelial cells play a key role in angiogenesis (formation of new blood vessels from pre-existing vasculature), the purpose of this study was to better understand human endothelial cell attachment, viability, growth, and phenotypic expression on sintered PLAGA microsphere scaffold. Scanning electron microscopy (SEM) examination showed cells attaching to the surface of microspheres and bridging the pores between the microspheres. Cell proliferation studies indicated that cell number increased during early stages and reached a plateau between days 10 and 14. Immunofluorescent staining for actin showed that cells were proliferating three dimensionally through the scaffolds while staining for PECAM-1 (platelet endothelial cell adhesion molecule) displayed typical localization at cell-cell contacts. Gene expression analysis showed that endothelial cells grown on PLAGA scaffolds maintained their normal characteristic phenotype. The cell proliferation and phenotypic expression were independent of scaffold pore architecture. These results demonstrate that PLAGA sintered microsphere scaffolds can support the growth and biological functions of human endothelial cells. The insights from this study should aid future studies aimed at enhancing angiogenesis in three dimensional tissue engineered scaffolds.  相似文献   

8.
A biocompatible and elastomeric nanofibrous scaffold is electrospun from a blend of poly(1,8-octanediol-co-citrate) [POC] and poly(L-lactic acid) -co-poly-(3-caprolactone) [PLCL] for application as a bioengineered patch for cardiac tissue engineering. The characterization of the scaffolds was carried out by Fourier transform infra red spectroscopy, scanning electron microscopy (SEM), and tensile measurement. The mechanical properties of the scaffolds are studied with regard to the percentage of POC incorporated with PLCL and the results of the study showed that the mechanical property and degradation behavior of the composites can be tuned with respect to the concentration of POC blended with PLCL. The composite scaffolds with POC: PLCL weight ratio of 40:60 [POC/PLCL4060] was found to have a tensile strength of 1.04 ± 0.11 MPa and Young's Modulus of 0.51 ± 0.10 MPa, comparable to the native cardiac tissue. The proliferation of cardiac myoblast cells on the electrospun POC/PLCL scaffolds was found to increase from Days 2 to 8, with the increasing concentration of POC in the composite. The morphology and cytoskeletal observation of the cells also demonstrated the biocompatibility of the POC containing scaffolds. Electrospun POC/PLCL4060 nanofibers are promising elastomeric substrates that might provide the necessary mechanical cues to cardiac muscle cells for regeneration of the heart.  相似文献   

9.
Electrospun composite scaffolds show high ability to be used in regenerative medicine and drug delivery, due to the nanofibrous structure and high surface area to volume ratio. In this study, we used nanofibrous scaffolds fabricated by chitosan (CS), poly(vinyl alcohol) (PVA), carbopol, and polycaprolactone using a dual electrospinning technique while curcumin (Cur) incorporated inside of the CS/PVA fibers. Scaffolds were fully characterized via scanning electron microscopy, water contact angle, tensile measurement, hydration, protein adsorption, and wrinkled tests. Furthermore, viability of the buccal fat pad-derived mesenchymal stem cells (BFP-MSCs) was also investigated using MTT assay for up to 14 days while cultured on these scaffolds. Cell cycle assay was also performed to more detailed evaluation of the stem cells growth when grown on scaffolds (with and without Cur) compared with the culture plate. Results demonstrated that Cur loaded nanofibrous scaffold had more suitable capability for water absorption and mechanical properties compared with the scaffold without Cur and it could also support the stem cells viability and proliferation. Cur release profile showed a decreasing effect on BFP-MSCs viability in the initial stage, but it showed a positive effect on stem cell viability in a long-term manner. In general, the results indicated that this nanofibrous scaffold has great potential as a delivery of the Cur and BFP-MSCs simultaneously, and so holds the promising potential for use in various regenerative medicine applications.  相似文献   

10.
Zhang P  Wu H  Wu H  Lù Z  Deng C  Hong Z  Jing X  Chen X 《Biomacromolecules》2011,12(7):2667-2680
Various surface modification methods of RGD (Arg-Gly-Asp) peptides on biomaterials have been developed to improve cell adhesion. This study aimed to examine a RGD-conjugated copolymer RGD/MPEG-PLA-PBLG (RGD-copolymer) for its ability to promote bone regeneration by mixing it with the composite of poly(lactide-co-glycotide) (PLGA) and hydroxyapatite nanoparticles surface-grafted with poly(L-lactide) (g-HAP). The porous scaffolds were prepared using solvent casting/particulate leaching method and grafted to repair the rabbit radius defects after seeding with autologous bone marrow mesenchymal cells (MSCs) of rabbits. After incorporation of RGD-copolymer, there were no significant influences on scaffold's porosity and pore size. Nitrogen of RGD peptide, and calcium and phosphor of g-HAP could be exposed on the surface of the scaffold simultaneously. Although the cell viability of its leaching liquid was 92% that was lower than g-HAP/PLGA, its cell adhesion and growth of 3T3 and osteoblasts were promoted significantly. The greatest increment in cell adhesion ratios (131.2-157.1% higher than g-HAP/PLGA) was observed when its contents were 0.1-1 wt % but only at 0.5 h after cell seeding. All the defects repaired with the implants were bridged after 24 weeks postsurgery, but the RGD-copolymer contained composite had larger new bone formation and better fusion interface. The composites containing RGD-copolymer enhanced bone ingrowth but presented more woven bones than others. The combined application of RGD-copolymer and bone morphological protein 2 (BMP-2) exhibited the best bone healing quality and was recommended as an optimal strategy for the use of RGD peptides.  相似文献   

11.
This study was performed to determine the microscopic biological response of human nasal septum chondrocytes and human knee articular chondrocytes placed on a demineralized bovine bone scaffold. Both chondrocytes were cultured and seeded onto the bovine bone scaffold with seeding density of 1 × 105 cells per 100 μl/scaffold and incubated for 1, 2, 5 and 7 days. Proliferation and viability of the cells were measured by mitochondrial dehydrogenase activity (MTT assay), adhesion study was analyzed by scanning electron microscopy and differentiation study was analyzed by immunofluorescence staining and confocal laser scanning electron microscopy. The results showed good proliferation and viability of both chondrocytes on the scaffolds from day 1 to day 7. Both chondrocytes increased in number with time and readily grew on the surface and into the open pores of the scaffold. Immunofluorescence staining demonstrated collagen type II on the scaffolds for both chondrocytes. The results showed good cells proliferation, attachment and maturity of the chondrocytes on the demineralized bovine bone scaffold. The bovine bone being easily resourced, relatively inexpensive and non toxic has good potential for use as a three dimensional construct in cartilage tissue engineering.  相似文献   

12.
A novel fibrous membrane of carboxymethyl chitin (CMC)/poly(vinyl alcohol) (PVA) blend was successfully prepared by electrospinning technique. The concentration of CMC (7%) with PVA (8%) was optimized, blended in different ratios (0–100%) and electrospun to get nanofibers. Fibers were made water insoluble by chemical followed by thermal cross-linking. In vitro mineralization studies identified the ability of formation of hydroxyapatite deposits on the nanofibrous surfaces. Cytotoxicity of the nanofibrous scaffold was evaluated using human mesenchymal stem cells (hMSCs) by the MTT assays. The cell viability was not altered when these nanofibrous scaffolds were pre-washed with phosphate buffer containing saline (PBS) before seeding the cells. The SEM images also revealed that cells were able to attach and spread in the nanofibrous scaffolds. Thus our results indicate that the nanofibrous CMC/PVA scaffold supports cell adhesion/attachment and proliferation and hence this scaffold will be a promising candidate for tissue engineering applications.  相似文献   

13.
In the present contribution, the potential for use of the ultrafine electrospun fiber mats of poly(3-hydroxybutyrate) (PHB) and poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) as scaffolding materials for skin and nerve regeneration was evaluated in vitro using mouse fibroblasts (L929) and Schwann cells (RT4-D6P2T) as reference cell lines. Comparison was made with PHB and PHBV films that were prepared by solution-casting technique. Indirect cytotoxicity assessment of the as-spun PHB and PHBV fiber mats with mouse fibroblasts (L929) and Schwann cells (RT4-D6P2T) indicated that the materials were acceptable to both types of cells. The attachment of L929 on all of the fibrous scaffolds was significantly better than that on both the film scaffolds and tissue-culture polystyrene plate (TCPS), while RT4-D6P2T appeared to attach on the flat surfaces of TCPS and the film scaffolds much better than on the rough surfaces of the fibrous scaffolds. For L929, all of the fibrous scaffolds were superior in supporting the cell proliferation to the film counterparts, but inferior to TCPS at days 3 and 5, while, for RT4-D6P2T, the rough surfaces of the fibrous scaffolds appeared to be very poor in supporting the cell proliferation when comparing with the smooth surfaces of TCPS and the film scaffolds. Scanning electron microscopy was also used to observe the behavior of both types of cells that were cultured on both the fibrous and the film scaffolds and glass substrate for 24 h.  相似文献   

14.
Polyglycolide (PGA)/poly(lactide-co-glycolide) (PLGA) scaffolds were fabricated by a solvent casting/particulate leaching method using hexafluoroisopropanol (HFIP) or acetone for material dissolution and NaCl particles as porogen. The results revealed that the mechanical strength increased as the PGA percentage in a HFIP-processed scaffold increased. Chemical ingredients did not substantially affect the mechanical strength of acetone-processed scaffolds. Large NaCl particles led to weak mechanical strength, low porosity, and small specific surface area. For a fixed composition, PGA crystals in a HFIP-processed scaffold were smaller than those in an acetone-processed scaffold. High PGA fractions yielded partly fused PGA/PLGA scaffolds. A faster degradation rate of a scaffold could result from a higher PGA percentage, smaller NaCl particles, or the existence of chondrocytes. The combination of PGA and PLGA, which compensated each other for bioactivity, would be beneficial to cartilage regeneration.  相似文献   

15.
AIM: To evaluate adhesion, proliferation and differentiation of human dental pulp stem cells (hDPSCs) on four commercially available scaffold biomaterials.METHODS: hDPSCs were isolated from human dental pulp tissues of extracted wisdom teeth and established in stem cell growth medium. hDPSCs at passage 3-5 were seeded on four commercially available scaffold biomaterials, SureOss (Allograft), Cerabone (Xenograft), PLLA (Synthetic), and OSTEON II Collagen (Composite), for 7 and 14 d in osteogenic medium. Cell adhesion and morphology to the scaffolds were evaluated by scanning electron microscopy (SEM). Cell proliferation and differentiation into osteogenic lineage were evaluated using DNA counting and alkaline phosphatase (ALP) activity assay, respectively.RESULTS: All scaffold biomaterials except SureOss (Allograft) supported hDPSC adhesion, proliferation and differentiation. hDPSCs seeded on PLLA (Synthetic) scaffold showed the highest cell proliferation and attachment as indicated with both SEM and DNA counting assay. Evaluating the osteogenic differentiation capability of hDPSCs on different scaffold biomaterials with ALP activity assay showed high level of ALP activity on cells cultured on PLLA (Synthetic) and OSTEON II Collagen (Composite) scaffolds. SEM micrographs also showed that in the presence of Cerabone (Xenograft) and OSTEON II Collagen (Composite) scaffolds, the hDPSCs demonstrated the fibroblastic phenotype with several cytoplasmic extension, while the cells on PLLA scaffold showed the osteoblastic-like morphology, round-like shape.CONCLUSION: PLLA scaffold supports adhesion, proliferation and osteogenic differentiation of hDPSCs. Hence, it may be useful in combination with hDPSCs for cell-based reconstructive therapy.  相似文献   

16.
Porous scaffolds for tissue engineering have been prepared from poly(3-hydroxybutyrate) (PHB) and a copolymer of poly(3-hydroxybutyrate) and polyethylene glycol (PHB-PEG) produced by bioPEGylation. The morphology of the scaffolds and their capacity for adsorption of the model protein bovine serum albumin (BSA) have been studied. Scaffolds produced from bioPEGylated PHB adsorbed more BSA, whereas the share of protein irreversibly adsorbed on these scaffolds was significantly lower (33%) than in the case of PHB homopolymer-based scaffolds (47%). The effect of protein adsorption on scaffold biocompatibility in vitro was tested in an experiment that involved the cultivation of fibroblasts (line COS-1) on the scaffolds. PHB-PEG scaffolds had a higher capacity for supporting cell growth than PHB-based scaffolds. Thus, the bioPEGylated PHB-based polymer scaffolds developed in the present study have considerable potential for use in soft tissue engineering.  相似文献   

17.
We investigated the performance of small-caliber polyurethane (PU) small-diameter vascular prosthesis generated using the electrospinning technique. PU was electrospun into small-diameter, small-caliber tubular scaffolds for potential application as vascular grafts. We investigated the effects of electrospinning conditions (solution concentration, mandrel rotation speed) on the microstructure and porosity of the scaffolds for the purpose of preparing scaffolds with optimum microstructures and properties. We evaluated the mechanical properties of the scaffolds by tensile tests and the cytotoxicity of the PU small-diameter, small-caliber PU synthetic vascular graft by the MTT assay. The adhesion of endothelial cells to the PU scaffold was characterized by Hoechst staining and fluorescence microscopy, and we measured endothelial cell proliferation on the PU scaffold by the CCK-8 assay. We analyzed the prosthesis microstructure and endothelial cell morphology using scanning electron microscopy. With increasing PU concentration in the electrospinning solution, the fiber diameter of the vascular graft increased and the porosity decreased. In addition, with increasing electrospinning time, the wall thickness increased and the porosity decreased. We found that regular fiber orientation can be obtained by adjusting the rotation speed of the mandrel. Cell proliferation was not inhibited as the small-caliber PU synthetic vascular grafts showed little cytotoxicity. The endothelial cells had faster adherence to the PU scaffolds than to the PTFE surface during the initial contact. After prolonged cell culture, significantly higher endothelial cell proliferation rate was observed in the PU scaffold groups than the PTFE group. We obtained small-caliber PU vascular grafts with optimal fiber arrangement, excellent mechanical properties, and optimal biocompatibility by optimizing the electrospinning conditions. This study provides in vitro biocompatibility data that is helpful for the clinical application of the PU small-diameter, small-caliber PU vascular grafts.  相似文献   

18.
Nanocomposites have recently been identified as a useful scaffolding material in tissue engineering applications. Poly (3-hydroxybutyrate)/hydroxyapatite nanoparticles (P3HB)/(nHA) porous scaffolds were successfully fabricated through a solvent casting and particulate leaching technique. P3HB/nHA and P3HB scaffolds were prepared by the same technique for comparison. The structure of the nanocomposite and P3HB scaffolds was observed by SEM. The Energy Disperssive X-ray Analysis (EDXA, map of Ca) results indicated that HA nanoparticles were homogeneously dispersed in the P3HB matrix. X-ray diffraction (XRD) analysis showed that P3HB and HA coexist in the nanocomposite. Transmission electron microscopy (TEM) images also showed that the particle size of HA was 30 ~ 40 nm. The porosity of the scaffolds was 84%, and macropores and micropores coexisted and interconnected throughout the scaffolds. Acellular bioactivity experiments showed that more HA crystals formed on the surface of the nanocomposite scaffold than on the P3HB scaffold after 4 weeks immersion in Simulated Body Fluid (SBF). Cell culture experiments demonstrated that the P3HB/nHA nanocomposite scaffold had a better tendency of proliferation and Alkaline Phosphatase (ALP) activity to MG 63 cells than the pure P3HB scaffold. It was found that nHA addition can improve acellular and cellular bioactivity of the P3HB scaffold.  相似文献   

19.
Cell stimulation by bioactive molecules has become an important tool in tissue engineering. The homogeneous incorporation of such molecules within the bulk of a polymer-based scaffold compared to surface coating is considered advantageous for most applications and minimizes a burst effect. An efficient way of bulk loading is the incorporation of these molecules during the scaffold formation process. In this paper, two different integrated processes for the preparation of scaffolds from poly(epsilon-caprolactone) (PCL) loaded with a small molecule are investigated. Both formation and loading of the scaffold is carried out in a single-step process. Sudan Red G was selected as a model compound for lipophilic small molecules. A freeze drying and pressure quench (PQ) formation process was selected, and the influence of the small molecule on the formation processes and on the morphology of the obtained scaffold was evaluated and compared. It could be shown for both processes that the formation of loaded scaffolds is possible, and that the small molecule has a very high impact on the foam morphology. In case of the freeze-drying (FD) method, only a load of 1 wt% Sudan Red G was incorporated within the bulk and showed no influence on the foam morphology. In the case of PQ foaming, an incorporation of 43 wt% Sudan Red G was achieved (although tiny crystal needles of the small molecule were found on the surface) and a strong effect on the foam morphology was found. This paper presents an efficient method of incorporating small molecules by integrated processes.  相似文献   

20.
PLGA的不同组成对支架材料性能的影响研究   总被引:5,自引:0,他引:5  
研究PLGA的不同组成对支架材料的力学性能、降解性能和生物学性能的影响。采用溶液浇注/颗粒沥取法制备出不同组成的PLGA多孔支架,对支架的力学性能和降解速率进行考察,同时将人真皮成纤维细胞接种于不同组成的PLGA支架材料上,培养不同时间后,检测细胞的粘附率和增殖率,以及细胞产生的总胶原含量,并通过扫描电镜观察支架上的细胞形态。结果显示,随PLA比例的增加,支架的力学强度增加,降解速率降低,但都不是线性变化。70:30比例的支架,拉伸强度最高,而70:30和80:20两种比例的支架,其降解速率没有显著性差异。PLGA不同组成的支架,均具有良好的细胞相容性,成纤维细胞粘附率和增殖率在三种比例的支架上没有显著性差异,细胞在支架表面生长良好,分泌大量的细胞外基质,细胞基本铺满整个支架。本文研究发现,PLGA的组成对支架力学性能、降解性能和生物学性能有细小但显著的影响,这将对组织构建选用PLGA支架材料提供有益的帮助。  相似文献   

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