共查询到20条相似文献,搜索用时 0 毫秒
1.
Zhou M Liu Z Liu C Jiang X Wei Z Qiao W Ran F Wang W Qiao T Liu C 《Journal of biomedical materials research. Part B, Applied biomaterials》2012,100(1):111-120
Successful construction of a small-diameter bioartificial vascular graft remains a great challenge. This study reports on novel tissue engineering vascular grafts (TEVGs) constructed by endothelial progenitor cells and heparin-coated decellularized vessels (DV). The DVs were fabricated from canine carotid arteries with observable depletion of cellular components. After heparin coating, the scaffolds possessed excellent antithrombogeneity. Canine endothelial progenitor cells harvested from peripheral blood were expanded and seeded onto heparin-coated DVs and cocultured in a custom-made bioreactor to construct TEVGs. Thereafter, the TEVGs were implanted into the carotid arteries of cell-donor dogs. After 3 months of implantation, the luminal surfaces of TEVGs exhibited complete endothelium regeneration, however, only a few disorderly cells and thrombosis overlaid the luminal surfaces of control DVs grafts, and immunofluorescent staining showed that the seeded cells existed in the luminal sides and the medial parts of the explanted TEVGs and partially contributed to the endothelium formation. Specifically, TEVGs exhibited significantly smaller hyperplastic neointima area compared with the DVs, not only at midportion (0.64 ± 0.08 vs. 2.13 ± 0.12 mm(2) , p < 0.001), but also at anastomotic sites (proximal sites, 1.03 ± 0.09 vs. 3.02 ± 0.16 mm(2), p < 0.001; distal sites, 1.84 ± 0.15 vs. 3.35 ± 0.21 mm(2), p < 0.001). Moreover, TEVGs had a significantly higher patency rate than the DVs after 3 months of implantation (19/20 vs. 12/20, p < 0.01). Overall, this study provided a new strategy to develop small-diameter TEVGs with excellent biocompatibility and high patency rate. 相似文献
2.
The effect of dynamic culture conditions on endothelial cell seeding and retention on small diameter polyurethane vascular grafts 总被引:3,自引:0,他引:3
Due to the limited number of cells available in endothelial cell (EC) seeding of small diameter vascular grafts, high seeding rate and ideal proliferation are normally required and can be achieved by optimizing the EC seeding and culture procedures. In this study, by using rotational seeding at 0.16 rpm for 12 h in an incubator, 90% cells were successfully seeded on the polyurethane vascular grafts. Following a period of 72 h of static culture, the cell retention after 6 h of flushing could reach 90%. The retention was further enhanced after perfuse culture (9 cm/s). The optimal procedures to prepare a polyurethane vascular graft (4-mm i.d., 4 cm long) populated with firmly attached EC were therefore: (1) seeding the graft with 0.5 ml of cell suspension containing approximately 10(5) cells rotated at 0.16 rpm for 12 h; (2) culturing the seeded graft in static for 72 h; and (3) culturing the graft by perfusion (9 cm/s) for another 72 h to 7 days. These procedures consistently resulted in a graft covered with confluent vein EC that fully retained on the surface after 6 h of in vitro flushing. 相似文献
3.
Vascular tissue engineering represents a promising approach for the development of living small-diameter vascular grafts that can be used for replacement therapy. The culture of strong human tissue-engineered (TE) vascular grafts has required long culture times, up to several months, whether or not combined with gene therapy. This article describes the culture of strong, genetically unmodified, human TE vascular grafts in 4 weeks Small-diameter vascular grafts were engineered using a fast-degrading polyglycolic acid scaffold coated with poly-4-hydroxybutyrate combined with fibrin gel and seeded with myofibroblasts isolated from discarded saphenous veins from patients undergoing coronary bypass surgery. The TE grafts were subjected to dynamic strain conditions. After 28 d of in vitro culture, the grafts demonstrated burst pressures of 903 +/- 123 mmHg. Comparison with native vessels (intact human left internal mammary arteries (LIMAs) and saphenous veins) showed no significant differences in the amount of DNA, whereas the TE vessels contained approximately 50% of the native collagen content. In the physiological pressure range, up to 300 mmHg, the mechanical properties of the TE vessels were comparable to the LIMA. In this study, we showed that dynamic conditioning combined with fibrin gel cell seeding enhances the mechanical properties of small-diameter TE grafts. These grafts might provide a promising alternative to currently used vascular replacements. 相似文献
4.
Scaffolds produced by rapid prototyping (RP) techniques have proved their value for tissue engineering applications, due to their ability to produce predetermined forms and structures featuring fully interconnected pore architectures. Nevertheless, low cell seeding efficiency and non-uniform distribution of cells remain major limitations when using such types of scaffold. This can be mainly attributed to the inadequate pore architecture of scaffolds produced by RP and the limited efficiency of cell seeding techniques normally adopted. In this study we aimed at producing scaffolds with pore size gradients to enhance cell seeding efficiency and control the spatial organization of cells within the scaffold. Scaffolds based on blends of starch with poly(ε-caprolactone) featuring both homogeneously spaced pores (based on pore sizes of 0.75 and 0.1 mm) and pore size gradients (based on pore sizes of 0.1-0.75-0.1 and 0.75-0.1-0.75 mm) were designed and produced by three-dimensional plotting. The mechanical performance of the scaffolds was characterized using dynamic mechanical analysis (DMA) and conventional compression testing under wet conditions and subsequently characterized using scanning electron microscopy and micro-computed tomography. Osteoblast-like cells were seeded onto such scaffolds to investigate cell seeding efficiency and the ability to control the zonal distribution of cells upon seeding. Scaffolds featuring continuous pore size gradients were originally produced. These scaffolds were shown to have intermediate mechanical and morphological properties compared with homogenous pore size scaffolds. The pore size gradient scaffolds improved seeding efficiency from ~35% in homogeneous scaffolds to ~70% under static culture conditions. Fluorescence images of cross-sections of the scaffolds revealed that scaffolds with pore size gradients induce a more homogeneous distribution of cells within the scaffold. 相似文献
5.
Endothelialization of PTFE vascular grafts under flow induces significant cell changes 总被引:9,自引:0,他引:9
Rademacher A Paulitschke M Meyer R Hetzer R 《The International journal of artificial organs》2001,24(4):235-242
BACKGROUND: To minimize thrombogeneity of small diameter PTFE grafts they are usually coated in vitro with endothelial cells under static culture conditions. The disadvantage of this technique is that a cell layer is formed that fails to withstand shear stress typical in normal blood flow. METHOD: Since the in vivo functional and structural status of endothelial cells correlates with the applied shear stress, we developed a computer-controlled perfusion system to seed and culture cells on PTFE-grafts up to a confluent monolayer under the influence of increasing shear stress. The confluence of endothelial coating was defined by immunohistological staining of cross sections, and by upper light microscopy of flattened graft samples. In addition, the expression of fibronectin as an important adhesion molecule was estimated. RESULTS AND DISCUSSION: The application of pulsatile shear stress (6.6 dyn/cm2, 5 min) to grafts endothelialized under perfusion (n = 7) did not lead to a disruption of the confluent cell layer. In contrast, a 5 min long shear stress of 3 dyn/cm2 was sufficient to wash more than 50% of cells off the PTFE-graft cultured under static conditions (n = 6). The perfusion cultures showed a significantly higher proliferation rate in comparison with static cultures. This effect was reproducibile in both serum-containing and serum-free culture media. The expression of fibronectin by endothelial cells was significantly higher in the perfused graft compared to the static one. These results suggest the practicability of endothelialized PTFE vascular grafts, preconditioned to shear rates similar to the in vivo situation, as an alternative bypass material in cardiac surgery. 相似文献
6.
The endothelization of polymers using surface modification has received great attention. In particular, creation of physical surface features such as craters or pores has been an active area of research. However, there have been no reported studies of the effects of pore sizes (wide range) and interpore distance on endothelial cell growth. This report details the study done on endothelial cell attachment on the surfaces of polymers modified by porogen leaching. The polymeric system studied includes PLLA and PLGA (80/20). Factors such as porogen type, pore size, and interpore distance were varied, and the surface was evaluated for its influence on endothelial cell growth. Three groups of pore sizes were evaluated: small (5-20 mum), medium (20-45 mum), and large pores (45-90 mum). Two porogens were evaluated: sugar and gelatin. In addition to counting the attached endothelial cells, their proliferation was also quantified. Pore size and interpore distances were evaluated using scanning electron microscopy (SEM), and cell morphology was studied by staining with crystal violet. Analysis of variance demonstrated that the main parameters, pore size and interpore distance were significant in endothelial cell growth. In PLGA (80/20), it was found that endothelial cell growth was enhanced by smaller pore size and lower interpore distance, whereas the growth was poor on PLLA regardless of pore features. 相似文献
7.
J P Mazzucotelli C Klein-Soyer A Beretz C Brisson G Archipoff J P Cazenave 《The International journal of artificial organs》1991,14(8):482-490
The establishment of an endothelial lining on vascular grafts to obtain a highly thromboresistant surface in a clinical situation requires optimization of cell collection, quality, adhesion and growth. We have studied the conditions for collection, seeding and growth of human saphenous vein endothelial cells (HSVEC), on Dacron or Gore-Tex expanded polytetrafluoroethylene (PTFE) vascular grafts. Carefully handled veins, as opposed to veins obtained using the usual procedures for coronary bypass graft preparation, yielded a higher rate of successful culture (94% vs 43%) and reached confluence in primary culture sooner (9.4 +/- 3 days vs 13.4 +/- 4.5 days). HSVEC were seeded at a density of 6 x 10(3) cells/cm2 on graft fragments coated with fibronectin (FN) or Transglutine (TGL), a biological glue. There was no HSVEC adhesion on Dacron or PTFE without protein pretreatment of the artificial surface. FN improved HSVEC adhesion but there was no cell growth. Adhesion, doubling time and cell density at confluence on PTFE pretreated with TGL were similar to those on conventional tissue culture polystyrene (TCP) pretreated with TGL or FN. HSVEC adhesion on Dacron pretreated with TGL was lower than on TCP pretreated with TGL; the doubling time was similar but the density at confluence was 40% lower. We conclude that pretreatment of vascular grafts with TGL, besides being an alternative to preclotting of the Dacron graft, allows adhesion and growth to confluence of HSVEC on these surfaces. 相似文献
8.
Gomes N Vassy J Lebos C Arbeille B Legrand C Fauvel-Lafeve F 《Clinical & experimental metastasis》2004,21(6):553-561
Tumour cell adhesion to vascular extracellular matrix (ECM), an important step of metastatic progression, is promoted by platelets.
The aim of our study was to investigate, in whole blood under venous and arterial shear conditions, the respective role of
tumour cell αvβ3 and platelet αIIbβ3integrins in MDA-MB-231 breast adenocarcinoma cell adhesion to human umbilical vein endothelial cell ECM. For that purpose,
blood containing MDA-MB-231 cells was incubated with non-peptide antagonists specific for platelet αIIbβ3 (lamifiban) or tumour cell αvβ3 (SB-273005). At 300 s−1, each antagonist used alone did not modify tumour cell adhesion, whereas, at 1500 s−1, tumour cell adhesion was decreased by 25% in presence of lamifiban indicating a role of platelet αIIbβ3 at higher shear rate. However, a combination of SB-273005 and lamifiban, or c7E3 Fab (a potent inhibitor of both αIIbβ3 and αvβ3) inhibited tumour cell adhesion by 40–45%, at either shear rate applied, indicating a cooperation between these two integrins
in MDA-MB-231 cell adhesion to ECM, as well as the participation of other adhesive receptors on tumour cells and/or platelets.
Thus, efficient anti-metastatic therapy should target multiple receptors on tumour cells and platelets.
This revised version was published online in July 2006 with corrections to the Cover Date. 相似文献
9.
《Biomaterials》2015
The behavior of cancer cells is strongly influenced by the properties of extracellular microenvironments, including topology, mechanics and composition. As topological and mechanical properties of the extracellular matrix are hard to access and control for in-depth studies of underlying mechanisms in vivo, defined biomimetic in vitro models are needed. Herein we show, how pore size and fibril diameter of collagen I networks distinctively regulate cancer cell morphology and invasion. Three-dimensional collagen I matrices with a tight control of pore size, fibril diameter and stiffness were reconstituted by adjustment of concentration and pH value during matrix reconstitution. At first, a detailed analysis of topology and mechanics of matrices using confocal laser scanning microscopy, image analysis tools and force spectroscopy indicate pore size and not fibril diameter as the major determinant of matrix elasticity. Secondly, by using two different breast cancer cell lines (MDA-MB-231 and MCF-7), we demonstrate collagen fibril diameter – and not pore size – to primarily regulate cell morphology, cluster formation and invasion. Invasiveness increased and clustering decreased with increasing fibril diameter for both, the highly invasive MDA-MB-231 cells with mesenchymal migratory phenotype and the MCF-7 cells with amoeboid migratory phenotype. As this behavior was independent of overall pore size, matrix elasticity is shown to be not the major determinant of the cell characteristics. Our work emphasizes the complex relationship between structural-mechanical properties of the extracellular matrix and invasive behavior of cancer cells. It suggests a correlation of migratory and invasive phenotype of cancer cells in dependence on topological and mechanical features of the length scale of single fibrils and not on coarse-grained network properties. 相似文献
10.
Flow and the associated shear stress have been shown to play an active role in the regulation of the structure and function
of endothelial cells (EC)in vitro. Although cultured EC subjected to flow exhibit an elongated morphology and a decreased cell growth rate rather like those
observedin vivo, there are differences in morphology and growth rate, as well as other characteristics, betweenin vitro andin vivo EC. This suggests that flow is only one of the many factors affecting EC differentiationin vivo. In this study, a co-culture model system was designed, which includes smooth muscle cells (SMC), a matrix of collagen type
I, and a confluent monolayer of EC, and this simplified model of the arterial wall was subjected to a steady, laminar shear
stress of 10 and 30 dyn/cm2. Under non-flow conditions, EC exhibited an elongated shape, but with a random orientation. In response to flow, there was
an alignment with the direction of flow. This alignment occurred more rapidly at 30 dyn/cm2 than at 10 dyn/cm2. The collagen matrix was found to be primordial in the maintenance of a quiescent endothelium, even in the absence of SMC
and flow, suggesting the importance of an organized extracellular matrix (ECM) in the differentiation of cellsin vivo. 相似文献
11.
T Isomura A M Dvorak R I Garcia H F Dvorak S J Galli 《Laboratory investigation; a journal of technical methods and pathology》1986,55(6):703-716
Aortic endothelial cell clones were established from adult inbred strain 2 guinea pigs by using a simple method requiring neither feeder layers nor growth factors other than isologous serum. One clone has been maintained in continuous culture for 26 passages over a 9-month period. Cloned endothelial cells grew in a cobblestone pattern at confluence, formed intercellular junctions, and expressed factor-VIII-related antigen by immunofluorescence microscopy. The cloned cells also contained high levels of angiotensin-converting enzyme activity, although the level of enzyme activity expressed by different cloned cell lines varied. Cloned endothelial cells contained numerous Weibel-Palade bodies which appeared larger and less electron-dense than those of strain 2 guinea pig aortic endothelial cells in vivo. These clones may serve as an in vitro model to investigate the formation and function of this organelle. Cloned populations of strain 2 guinea pig vascular endothelial cells maintained in medium supplemented only with strain 2 guinea pig serum also provide an immunologically isologous system ideal for investigating interactions between the vascular endothelium and immunocompetent leukocytes in vitro. 相似文献
12.
Sho E Komatsu M Sho M Nanjo H Singh TM Xu C Masuda H Zarins CK 《Experimental and molecular pathology》2003,75(1):1-11
Endothelial cell activation and proliferation are the essential steps in flow-induced arterial remodeling. We investigated endothelial cell turnover in the early stages of high-flow in the rabbit common carotid arteries using an arteriovenous fistula (AVF) model by kinetic investigation of cell proliferation and cell molecular analysis. BrdU was administrated to label endothelial cells (ECs) in DNA synthetic phase (S-phase) of the cell mitotic cycle. Pulse labeling revealed that ECs entered S-phase at 1.5 days of AVF (0.93 +/- 0.19%). Endothelial cell labeling index (EC-LI) peaked at 2 days of AVF (8.90 +/- 0.87%) with a high index of endothelial cell mitosis (EC-MI, 1.67 +/- 0.47%). Endothelial cell density increased remarkably at 3 days of AVF with a significant decrease in EC-LI (54%) and EC-MI (60%). Study of kinetics of EC proliferation revealed that endothelial cells took 16-24 h to finish one cycle of cell mitosis. Tracking investigation of pulse BrdU-labeled endothelial cells at 1.5 days showed that more than 66% of endothelial cells were BrdU-labeled 1.5 days after labeling. VEGF, integrin alphanubeta3, PECAM-1, and VE-cadherin were upregulated significantly preceding endothelial cell proliferation and kept at high levels during endothelial cell proliferation. These data suggest that endothelial cell proliferation is the initial step in flow-induced arterial remodeling. Hemodynamic forces may drive endothelial cell downstream migration. Expression of VEGF and cell junction molecules contribute to flow-induced arterial remodeling. 相似文献
13.
Alberto Giudiceandrea MD Henryk J. Salacinski PhD Nigel R. M. Tai FRCS Geoffrey Punshon BSc George Hamilton MD Alexander M. Seifalian PhD 《Journal of artificial organs》2000,3(1):16-24
The objective of this study was to develop and validate a flow system capable of simulating in vitro the pulsatile-flow waveform,
pressures, and degree of oxygenation of physiological arterial circulation in vivo. We used this model to accurately determine
endothelial cell adhesiveness on seeded vascular grafts exposed to physiological shear stress. The system consisted of a pulsatile-flow
phantom capable of reproducing physiological arterial flow-including a reverse-flow component-which was filled with a nutritive,
oxygenated solution that was of the same viscosity as whole blood. Real-time monitoring of all flow variables allowed continual
adjustment of parameters and maintenance of physiological shear stress. To assess cell retention, human umbilical vein endothelial
cells were radiolabeled with111indium, seeded onto compliant polyurethane vascular graft segments at a density of 1.8×106 cells/cm2, mounted in series with the flow circuit, and exposed to arterial shear stress for an 8-h period (n=6). Dynamic scintigraphy images were acquired in real time using a nuclear medicine gamma camera system during the 8-h perfusion
period. The perfusion medium was tested for cytotoxicity, and the viability of seeded cells was assessed by Alamar Blue assay
and scanning electron microscopy. Using the model, the shear stress and flow waveforms of human femoral arterial circulation
were successfully simulated. The perfusate (viscosity 0.035 poise) was determined to be noncytotoxic. Postperfusion examination
by scanning electron microscopy for known morphological indicators and Alamar Blue colorimetric assay for metabolic activity
demonstrated normal cell appearance and activity. This system allows complete simulation of the hemodynamic variables encountered
at the luminal surface of the femoral artery, while providing a nontoxic supportive milieu. Investigation of the behavior
of seeded endothelial cells can be reliably and accurately undertaken with this system.
Presented at the Sixth International Society for Applied Cardiovascular Biology, Munich, Germany, 5–7 March, 1998 相似文献
14.
15.
16.
The usefulness of nanoscale topography in improving vascular response in vitro was established previously on hydrothermally modified titanium surfaces. To propose this strategy of surface modification for translation onto clinically used metallic stents, it is imperative that the surface should be also hemocompatible: an essential attribute for any blood-contacting device. The present in vitro study focuses on a detailed hemocompatibility evaluation of titania nanostructures created through an alkaline hydrothermal route on metallic Ti stent prototypes. Direct interactions of TiO2 nanocues of various morphologies with whole blood were studied under static conditions as well as using an in vitro circulation model mimicking arterial flow, with respect to a polished Ti control. Nanomodified stent surfaces upon contact with human blood showed negligible hemolysis under constant shear and static conditions. Coagulation profile testing indicated that surface roughness of nanomodified stents induced no alterations in the normal clotting times, with insignificant thrombus formation and minimal inflammatory reaction. Endothelialized nanomodified Ti surfaces were found to inhibit both activation as well as aggregation of platelets compared with the control surface, with the endothelium formed on the nanosurfaces having an increased expression of anti-thrombogenic genes. Such a nanotextured Ti surface, which is anti-thrombogenic and promotes endothelialization, would be a cost-effective alternative to drug-eluting stents or polymer-coated stents for overcoming in-stent restenosis. 相似文献
17.
This study examined the effects of 4 mg nicotine and placebo upon problem solving performance in word and number tasks, and subsequent recall and recognition of the answers to these problems. The results demonstrated that the drug had no effect upon the subject's ability to generate the correct answers to the problems, but that immediate and delayed recall and recognition were significantly impaired. These data clearly do not support the view that nicotine, without exception, enhances information processing, and it was suggested that the effects of nicotine upon information retrieval may be specific to tasks which assess episodic memory in the absence of retrieval cues or a problem solving context. 相似文献
18.
Angiogenesis occurs through a cascade of events controlled by complex multiple signals that are orchestrated according to specific spatial patterns and temporal sequences. Vascularization is a central issue in most tissue engineering applications. However, only a better insight into spatio-temporal signal presentation can help in controlling and guiding angiogenesis in vivo. To this end, versatile and accessible material platforms are required in order to study angiogenic events in a systematic way. In this work we report a three-dimensional porous polyethylene glycol (PEG) diacrylate hydrogel bioactivated with heparin that is able to deliver vascular endothelial growth factor (VEGF) in a sustained and controlled manner. The efficiency of the material has been tested both in vitro and in vivo. In particular, the VEGF released from the hydrogel induces cell proliferation when tested on HUVECs, retains its bioactivity up to 21days, as demonstrated by Matrigel assay, and, when implanted on a chorion allantoic membrane, the hydrogel shows superior angiogenic potential in stimulating new vessel formation compared with unfunctionalized hydrogels. Moreover, in the light of potential tissue regeneration studies, the proposed hydrogel has been modified with adhesion peptides (RGD) to enable cell colonization. The porous hydrogel reported here can be used as a valid tool to characterize angiogenesis, and, possibly, other biological processes, in different experimental set-ups. 相似文献
19.
Coculture of endothelial and smooth muscle cells on a collagen membrane in the development of a small-diameter vascular graft 总被引:5,自引:0,他引:5
In this study, we have evaluated the feasibility of developing a biodegradable collagenous small diameter vascular graft of 2mm diameter and 1cm length. In brief, bi-layer type I collagen membrane was fabricated under vacuum suction and lyophilization methods. The smooth muscle cells were inoculated into the lower side of the porous membrane, while endothelial cells were seeded onto upper smooth side of the membrane. After cultured for 7 days, the vascular substitute was either harvested for in vitro examination or in vivo implanted in the subcutaneous layer for biocompatibility test. The tubular vascular prosthesis was then used as a temporary absorbable guide that served as an in vivo vascular graft to promote the complete regeneration of rat inferior vena cava. After implantation for 12 weeks, a thin continuous layer of endothelial cells and smooth muscle cells were lined with the vascular lumen and tunic media, respectively. Histology results showed that there were no signs of significant thrombogeneity and intima hyperplasia. This tissue engineered vascular substitute not only had enough tensile strength and good biocompatibility, but also advanced vascular regeneration. In the future, we suggest that this biodegradable vascular substitute will provide with the possibility in application on small diameter prosthetic grafts in artificial blood vessels. 相似文献