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1.
Kavanagh CA Gorelova TA Selezneva II Rochev YA Dawson KA Gallagher WM Gorelov AV Keenan AK 《Journal of biomedical materials research. Part A》2005,72(1):25-35
The aim of this study was to establish the capacity of thermoresponsive poly(N-isopropylacrylamide) copolymer films to deliver bioactive concentrations of vascular endothelial growth factor (VEGF165) to human aortic endothelial cells (HAEC) over an extended time period. Films were prepared using a 50:50 (w/w) mixture of non-crosslinkable and crosslinkable copolymers of the following monomer compositions (w/w): 85:15, N-isopropylacrylamide (NiPAAm):N-tert-butylacrylamide (NtBAAm); and 85:13:2 NiPAAm:NtBAAm:acrylamidobenzophenone (ABzPh, crosslinking agent), respectively. After crosslinking by UV irradiation, the ability of films to incorporate a fluorescently labeled carrier protein (FITC-labeled BSA, 1 mg loaded per film), at 4 degrees C, was first established. Incorporation into the matrix was confirmed by the observation that increasing film thickness from 5 to 10 microm increased release from collapsed films at 37 degrees C (1.76 +/- 0.15 and 10.98 +/- 3.38 microg/mL, respectively, at 24 h postloading) and that this difference was maintained at 5 days postloading (1.81 +/- 0.25 and 13.8 +/- 2.3 microg/mL, respectively). Incorporation was also confirmed by visualization using confocal microscopy. When 10-microm films were loaded with a BSA solution (1 mg/mL) containing VEGF165 (3 microg/mL), sustained release of VEGF165 was observed (10.75 +/- 3.11 ng at 24 h; a total of 31.32 +/- 8.50 ng over 7 days). Furthermore, eluted VEGF165 increased HAEC proliferation by 18.2% over control. The absence of cytotoxic species in medium released from the copolymer films was confirmed by the lack of effect of medium (incubated with copolymer films for 3 days) on HAEC viability. In conclusion this study has shown that NiPAAm:NtBAAm copolymers can be loaded with a therapeutic protein and can deliver bioactive concentrations to human vascular endothelial cells over an extended time period. 相似文献
2.
Wilson SJ Gorelov AV Rochev YA McGillicuddy F Dawson KA Gallagher WM Keenan AK 《Journal of biomedical materials research. Part A》2003,67(2):667-673
The aim of this study was to establish the capacity of thermoresponsive poly(N-isopropylacrylamide) copolymer films to deliver bioactive concentrations of an antimitotic agent to human vascular smooth muscle cells (HASMC) over an extended period of time. Copolymer films were prepared using a 50:50 (w/w) ratio of N-isopropylacrylamide (NiPAAm) monomer to the more hydrophobic N-tert-butylacrylamide (NtBAAm) and loaded with the antimitotic agent colchicine (0.1 micromol per film) at room temperature. Colchicine release from films was sustained over a 14-day period. At 24 h postloading, the concentration of colchicine in the medium overlying films was 2.12 +/- 0.16 microM; this fell to 0.20 +/- 0.01 microM at 7 days and decreased further to 0.12 +/- 0.01 microM after 14 days. Colchicine released from copolymer films inhibited proliferation when subsequently placed on HASMC: at 0.1 microM, released colchicine reduced proliferation to 18.5 +/- 0.8% of control cells (p < 0.001, n = 9). The antiproliferative effect of released colchicine was comparable to that of native colchicine, as observed in separate experiments. Furthermore, colchicine released from 50:50 polymer films inhibited the proliferation of cells grown in the same environment as the copolymer. Inhibition of cell proliferation was not due to the release of cytotoxic particles from the copolymer because medium incubated with copolymer for 5 days and then applied to HASMC did not alter cell viability. In conclusion, this study demonstrates that 50:50 NiPAAm:NtBAAm copolymers can deliver bioactive concentrations of the antimitotic agent colchicine to human vascular cells over an extended period of time. 相似文献
3.
Laçin NT Utkan GG Kutsal T Pişkin E 《Journal of biomaterials science. Polymer edition》2012,23(5):577-592
Primary smooth muscle cells (SMC) isolated from the aorta of fetal calf were transfected with a green fluorescent protein (GFP)-encoding plasmid DNA, which was carried by a water-soluble and temperature-sensitive N-isopropylacrylamide-based (NIPAAm-based)-co-polymer, either poly(N-isopropylacrylamide-co-2-methacryloamidohistidine) (poly(NIPAAm-co-MAH)) or monosized PEGylated nanoparticle poly(styrene/poly(ethylene glycol) ethyl ether methacrylate/N-(3-(dimethylamino)propyl) methacrylamide) (poly(St/PEG-EEM/DMAPM)). Poly(NIPAAm-co-MAH) co-polymer was synthesized by solution polymerization of n-isopropylacrylamide (NIPAAm) and 2-methacrylamidohistidine (MAH). Monosized cationic nanoparticles were produced by emulsifier-free emulsion polymerization of styrene, PEG ethyl ether methacrylate and N-[3-(dimethyl-amino) propyl] methacrylamide, in the presence of a cationic initiator, 2,2-azobis (2-methylpropionamidine) dihydrochloride. The structure of poly(St/PEG-EEM/DMAPM) and poly(NIPAAm-co-MAH) was confirmed by(1) H-NMR and FT-IR spectroscopy. Particle size/size distribution and surface charges of both carriers were measured by Zeta Sizer. The LCST behavior of poly(NIPAAm-co-MAH) co-polymer was followed spectrophotometrically. Poly(St/PEG-EEM/DMAPM) nanoparticles, with an average size of 78 nm and zeta potential of 54.4 mV, and an average size of 200 nm with a zeta potential of 54.2 mV, and poly(NIPAAm-co-MAH) were used in the transfection studies. The cytotoxicity of the vectors was tested using the MTT method. According to conditions for the transfection study (polymer/cell ratio and polymer-cell incubation period), cell loss was only 4 and 15% with poly(St/PEG-EEM/DMAPM) sized 78 and 200 nm, respectively. Poly(NIPAAm-co-MAH) cytotoxicity was insignificant. Poly(NIPAAm-co-MAH) uptake efficiency in SMCs was around 85%, but gene expression efficiency were low compared to poly(St/PEG-EEM/DMAPM)/pEGFP-N2 conjugates because of the low zeta potential of the co-polymer. Polymer uptake efficiencies of the nanoparticles were 90-95%. GFP expression efficiency was 68 and 64% after transfection with pEGFP-N2 conjugate with 78 and 200 nm sized poly(St/PEG-EEM/DMAPM) nanoparticles. 相似文献
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5.
Poly(gamma-glutamic acid) nanoparticles as an efficient antigen delivery and adjuvant system: potential for an AIDS vaccine 总被引:9,自引:0,他引:9
Antigen delivery systems using polymeric nanoparticles are of special interest as stable protein-based antigen carriers. In the present study, novel biodegradable poly(gamma-glutamic acid) (gamma-PGA) nanoparticles were examined for their antigen delivery and immunostimulatory activities in vitro and in vivo. The uptake of ovalbumin by dendritic cells was markedly enhanced by gamma-PGA nanoparticles, and the ovalbumin was gradually released from gamma-PGA nanoparticles into the cells. In addition, gamma-PGA nanoparticles appeared to have great potential as an adjuvant, because they could induce the maturation of dendritic cells. Although not only ovalbumin-encapsulating nanoparticles (OVA-NPs) but also a simple mixture of ovalbumin and nanoparticles induced dendritic cell maturation, the only dendritic cells exposed to OVA-NPs could strongly activate antigen-specific interferon (IFN)-gamma-producing T cells. Subcutaneous immunization of mice with human immunodeficiency virus type 1 (HIV-1) p24-encapsulating nanoparticles activated antigen-specific IFN-gamma-producing T cells in spleen cells and induced p24-specific serum antibodies, as compared to immunization with p24 alone. Like ovalbumin, a mixture of p24 and nanoparticles also induced antigen-specific serum antibodies but did not activate IFN-gamma-producing T cells in spleen cells, suggesting that nanoparticles play a critical role in inducing cellular immune responses. Furthermore, gamma-PGA nanoparticles had a capacity comparable to that of the complete Freund's adjuvant (CFA) in inducing p24-specific serum antibody. However, unlike CFA, they predominantly activated p24-specific IFN-gamma-producing T cells. Thus, gamma-PGA nanoparticles encapsulating various antigens may have great potential as novel and efficient protein-based vaccines against infectious diseases, including HIV-1 infection. 相似文献
6.
Williams C Xie AW Emani S Yamato M Okano T Emani SM Wong JY 《Tissue engineering. Part A》2012,18(9-10):986-998
In pediatric patients requiring vascular reconstruction, the development of a cell-based tissue-engineered vascular patch (TEVP) has great potential to overcome current issues with nonliving graft materials. Determining the optimal cell source is especially critical to TEVP success. In this study, we compared the ability of human aortic smooth muscle cells (HuAoSMCs) and human mesenchymal stem cells (hMSCs) to form cell sheets on thermoresponsive poly(N-isopropylacrylamide) (PIPAAm) substrates. hMSCs treated with transforming growth factor beta 1 (TGFβ1) and ascorbic acid (AA) had higher expression of SMC-specific proteins compared to HuAoSMCs. hMSCs also had larger cell area and grew to confluence more quickly on PIPAAm than did HuAoSMCs. hMSCs typically formed cell sheets in 2-3 weeks and had greater wet tissue weight and collagen content compared with HuAoSMC sheets, which generally required growth for up to 5 weeks. Assays for calcification and alkaline phosphatase activity revealed that the osteogenic potential of TGFβ1+AA-treated hMSCs was low; however, Alcian Blue staining suggested high chondrogenic behavior of TGFβ1+AA-treated hMSCs. Although hMSCs are promising for cell-based TEVPs in their ability to form robust tissue with significant extracellular matrix content, improved control over hMSC behavior will be required for long-term TEVP success. 相似文献
7.
p75(NTR) mediates neurotrophin-induced apoptosis of vascular smooth muscle cells 总被引:4,自引:0,他引:4
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Wang S Bray P McCaffrey T March K Hempstead BL Kraemer R 《The American journal of pathology》2000,157(4):1247-1258
The development of atherosclerotic lesions results from aberrant cell migration, proliferation, and extracellular matrix production. In advanced lesions, however, cellular apoptosis, leading to lesion remodeling, predominates. During lesion formation, the neurotrophins and the neurotrophin receptor tyrosine kinases, trks B and C, are induced and mediate smooth muscle cell migration. Here we demonstrate that a second neurotrophin receptor, p75(NTR), is expressed by established human atherosclerotic lesions and late lesions that develop after balloon injury of the rat thoracic aorta. The p75(NTR), a member of the tumor necrosis factor/FAS receptor family, can modulate trk receptor function as well as initiate cell death when expressed in cells of the nervous system that lack kinase-active trk receptors. p75(NTR) expression colocalizes to neointimal cells, which express smooth muscle cell alpha-actin and are expressed by cultured human endarterectomy-derived cells (HEDC). Areas of the plaque expressing p75(NTR) demonstrate increased TUNEL positivity, and HEDC undergo apoptosis in response to the neurotrophins. Finally, neurotrophins also induced apoptosis of a smooth muscle cell line genetically manipulated to express p75(NTR), but lacking trk receptor expression. These studies identify the regulated expression of neurotrophins and p75(NTR) as an inducer of smooth muscle cell apoptosis in atherosclerotic lesions. 相似文献
8.
Marrow-derived cells as vehicles for delivery of gene therapy to pulmonary epithelium 总被引:18,自引:0,他引:18
Grove JE Lutzko C Priller J Henegariu O Theise ND Kohn DB Krause DS 《American journal of respiratory cell and molecular biology》2002,27(6):645-651
Gene therapy application to pulmonary airways and alveolar spaces holds tremendous promise for the treatment of lung diseases. However, safe and effective long-term gene expression using viral and nonviral vectors has not yet been achieved. Adenoviral vectors, with a natural affinity for airway epithelia, have been partially effective, but are inflammatory and induce only transient gene expression. We investigate the novel approach of using retrovirally transduced multipotent bone marrow-derived stem cells (BMSC) to deliver gene therapy to lung epithelium. We have shown previously that up to 20% of lung epithelial cells can be derived from marrow following BMSC transplantation. Here, irradiated female mice were transplanted with male marrow that had been transduced with retrovirus encoding eGFP. Transgene expressing lung epithelial cells were present in all recipients analyzed at 2, 5, or 11 mo after transplant (n = 10), demonstrating that highly plastic BMSC can be stably transduced in vitro and retain their ability to differentiate into lung epithelium while maintaining long-term transgene expression. 相似文献
9.
Migration of human vascular endothelial and smooth muscle cells. 总被引:7,自引:0,他引:7
G Thorgeirsson A L Robertson D H Cowan 《Laboratory investigation; a journal of technical methods and pathology》1979,41(1):51-62
Migration of endothelial and smooth muscle cells was studied in vitro by measuring the increase in surface area at specific time intervals of confluent cell colonies advancing under agarose gels that contained both Morgan's medium 199 and variuos types of sera. First passage cultures of endothelial cells or 3 to 6 passage smooth muscle cells were plated into wells punched in agarose gels, at a seeding density of 50,000 cells per well. At zero time the size of the cell colonies was 35.4 sq. mm. +/- standard error 0.1. Irradiation (1500 rads) did not affect the expansion of the cell colonies although 3H-thymidine uptake was inhibited. Endothelial cells migrated under the agarose gels concentrically as contiguous sheets. When exposed to either 20 per cent platelet-poor plasma serum, platelet-rich plasma serum, or whole blood serum, the average increase in surface area was approximately 9 sq. mm. per day. In contrast, arterial smooth muscle cell colonies expanded with an increment of approximately 9 sq. mm. per day when exposed to 10 per cent platelet-poor plasma serum but 12 sq. mm. per day when exposed to 10 per cent platelet-rich plasma serum (p less than 0.001). Platelet factors also had stimulatory effects on the migration of venous smooth muscle cells. Cytochalasin B, dibutyryl cyclic AMP, and theophylline inhibited the migration of both endothelial and smooth muscle cells, but the latter responded more to the inhibitory effects of all three agents. It is concluded that in contrast to vascular smooth muscle, endothelial cells do not require platelet factors for migration and are less responsive to specific inhibitors affecting cell movement. 相似文献
10.
Three-dimensional microchannels in biodegradable polymeric films for control orientation and phenotype of vascular smooth muscle cells 总被引:1,自引:0,他引:1
Shen JY Chan-Park MB He B Zhu AP Zhu X Beuerman RW Yang EB Chen W Chan V 《Tissue engineering》2006,12(8):2229-2240
The poor mechanical strength and vasoactivity of current small-diameter tissue engineered blood vessels (TEBVs) remain unsolved problems. Given the plasticity of smooth muscle cells (SMCs), 1 of the main limitations of current scaffolding techniques is the difficulty in controlling SMC phenotype shifts in vitro. A synthetic phenotype allows the cells to rapidly proliferate and produce extracellular matrix (ECM), whereas a shift to contractile phenotype with organized ECM ultimately provides a functional blood vessel. In this study, 3D deep (65 microm) and wide microchannels separated by high-aspect ratio (8) microwalls were successfully ultraviolet (UV) microembossed using a liquid UV polymerizable biodegradable macromer (poly(epsilon-caprolactone-r-L-lactide-r-glycolide) diacrylate) and the in vitro guidance effects of varying channel width (40-160 microm) on SMCs were verified. The results show that SMCs cultured in the wider microchannels (80-160 microm wide) switch from fibroblast morphology and random orientation to spindle-shaped morphology, and align along the direction of the microchannel nearing confluence achieved with similar cell density to unpatterned film. Further, an enhanced expression of smooth muscle alpha-actin of SMCs grown on micropatterns was found nearing confluence, which demonstrates a phenotype shift to a more contractile phenotype. These films are flexible and can be folded into tubular and lamellar structures for tissue engineering of small-diameter TEBVs as well as other organs such as esophagus or intestine. These results suggest that these micropatterned synthetic biodegradable scaffolds may be useful for guiding SMCs to grow into functional, small-diameter vascular grafts. 相似文献
11.
The mixture of proteins secreted by neonatal rat aorta smooth muscle cells cultured in the presence of beta-aminopropionitrile was readily oxidized and polymerized upon incubation with purified or crude preparations of lysyl oxidase. Western blot analysis indicated that these substrates included 30-60kDa protein bands reactive with anti-elastin, presumed to be fragments derived from tropoelastin. Thus, truncated, elastin-like as well as other proteins accumulate in the media of these cultures which, in toto, can serve as a conveniently prepared, highly efficient substrate for the routine assay of lysyl oxidase activity. 相似文献
12.
《Journal of biomaterials science. Polymer edition》2013,24(7):809-827
A series of poly(N-isopropylacrylamide)-grafted gelatins (PNIPAM gelatins) of three different graft densities (approx. 11, 22 and 34 graft chains per gelatin molecule) and three different molecular weights of their graft chains (molecular weight approximately 1.2 × 104, 5.0 × 104 and 1.3 × 105 g/mol) were prepared by multiple derivatization of dithiocarbamyl (DC) group in a gelatin molecule and subsequent iniferter (acts as an initiator, transfer-agent and terminator)-based photopolymerization of NIPAM. The weight ratio of PNIPAM graft chains to gelatin (P/G) varied from 1.4 to 49. Aqueous solutions of PNIPAM-gelatins showed thermo-responsiveness, depended on the graft density and the molecular weight of PNIPAM graft chain or P/G. Aqueous solutions (10 or 20%, w/v) of PNIPAM-gelatins with P/G of more than 5.8 were converted to gels at 37°C. Focal plane images of PNIPAM-gelatin gels by confocal laser scanning microscopy revealed that the size of hydrophobically clustered aggregates increased with P/G, whereas the space of microvoids decreased with concentration. Compressive strain–stress measurements revealed that compressive strength of PNIPAM-gelatin increased with P/G. Bovine smooth muscle cells (SMCs)-entrapped gels were produced from PNIPAM-gelatin-containing cell-suspended medium solutions at 37°C. The entrapped cells proliferated in the gel with P/G of more than 12. A higher cell proliferativity was obtained at low concentration (5%, w/v) and higher P/G (> 18). Tissue formation composed of proliferative SMCs and cell-secreted extracellular matrices (collagen) was obtained at 14 days incubation. The inter-relationship between the molecular parameters of PNIPAM-gelatin, internal structural features and cell proliferation potential was discussed. 相似文献
13.
A novel transgenic marker for migrating limb muscle precursors and for vascular smooth muscle cells. 总被引:4,自引:0,他引:4
A Tidhar M Reichenstein D Cohen A Faerman N G Copeland D J Gilbert N A Jenkins M Shani 《Developmental dynamics》2001,220(1):60-73
A unique pattern of LacZ expression was found in a transgenic mouse line, likely due to regulatory elements at the site of integration. Two new genes flanking the transgene were identified. At early stages of development, the transgene is transiently expressed in ventro-lateral demomyotomal cells migrating from the somites into the limb buds. At late developmental stages and in the adult, lacZ staining marks vascular smooth muscle cells throughout the vascular bed, with the exception of the major elastic arteries, and in pericytes. No expression was detected in skeletal and smooth muscles. Different patterns of expression in vascular smooth muscles was observed at distinct levels of the vascular tree, in arteries as well as in veins. Vessel injury, resulting in stimulation of smooth muscle cells proliferation and migration, is associated with transgene down-regulation. After the formation of neointima thickening, it is reactivated. This transgenic insertion may therefore be used as a useful marker to identify novel physiological cues or genetic elements involved in the regulation of the vascular smooth muscle phenotype(s). It may also provide an experimental tool for studying vasculature and the involvement of pericytes in regulating microvascular homeostasis. 相似文献
14.
K Norioka M Hara M Harigai A Kitani T Hirose W Hirose K Suzuki M Kawakami M Kawagoe H Nakamura 《Autoimmunity》1990,7(1):41-50
Systemic vasculitis is an inflammatory disorder of blood vessels characterized by a perivascular mononuclear cell infiltration around the vessel and fibrinoid necrosis within vessel walls. Interleukin-1 (IL-1) is a multipotent inflammatory mediator and affects several properties of vascular cells. To determine whether IL-1 could contribute to the pathogenesis of vascular diseases, we examined the effect of IL-1 on B cell stimulatory factor-2/interleukin-6 (IL-6) production by cultured human vascular smooth muscle cells (SMC) and the proliferation of these cells. Supernatants of SMC stimulated IgM synthesis of human B cell line. SKW6-CL4 cells. This activity was increased (1.7 to 2.6-fold) when SMC were pretreated with IL-1 or calcium ionophore A23187 for 48 h, and was completely blocked by rabbit anti-human IL-6 antibodies. These IL-6 activities of the SMC supernatants were also assessed by using an IL-6 dependent murine hybridoma cell line. MH-60. BSF-2. In addition, we observed that pretreatment of SMC with IL-1 for 48 h stimulated growth of SMC during the 96 h incubations, as assessed by cell number (p less than 0.05). These results suggest that IL-1 may contribute to the pathogenesis of inflammatory and immunological vasculitis by the augmentation of IL-6 release and growth of SMC. 相似文献
15.
A series of poly(N-isopropylacrylamide)-grafted gelatins (PNIPAM gelatins) of three different graft densities (approx. 11, 22 and 34 graft chains per gelatin molecule) and three different molecular weights of their graft chains (molecular weight approximately 1.2 x 10(4), 5.0 x 10(4) and 1.3 x 10(5) g/mol) were prepared by multiple derivatization of dithiocarbamyl (DC) group in a gelatin molecule and subsequent iniferter (acts as an initiator, transfer-agent and terminator)-based photopolymerization of NIPAM. The weight ratio of PNIPAM graft chains to gelatin (P/G) varied from 1.4 to 49. Aqueous solutions of PNIPAM-gelatins showed thermo-responsiveness, depended on the graft density and the molecular weight of PNIPAM graft chain or P/G. Aqueous solutions (10 or 20%, w/v) of PNIPAM-gelatins with P/G of more than 5.8 were converted to gels at 37 degrees C. Focal plane images of PNIPAM-gelatin gels by confocal laser scanning microscopy revealed that the size of hydrophobically clustered aggregates increased with P/G, whereas the space of microvoids decreased with concentration. Compressive strain-stress measurements revealed that compressive strength of PNIPAM-gelatin increased with P/G. Bovine smooth muscle cells (SMCs)-entrapped gels were produced from PNIPAM-gelatin-containing cell-suspended medium solutions at 37 degrees C. The entrapped cells proliferated in the gel with P/G of more than 12. A higher cell proliferativity was obtained at low concentration (5%, w/v) and higher P/G (>18). Tissue formation composed of proliferative SMCs and cell-secreted extracellular matrices (collagen) was obtained at 14 days incubation. The inter-relationship between the molecular parameters of PNIPAM-gelatin, internal structural features and cell proliferation potential was discussed. 相似文献
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Localized drug delivery from drug-eluting stents has been accepted as one of the most promising treatment methods for preventing restenosis after stenting. However, thrombosis, inflammation, and restenosis are still major problems for the utility of cardiovascular prostheses such as vascular grafts and stents. Epigallocatechin-3-O-gallate (EGCG), a major polyphenolic constituent of green tea, has been shown to have anti-thrombotic, anti-inflammatory and anti-proliferative activities. It was hypothesized that controlled release of EGCG from biodegradable poly(lactide-co-epsilon-caprolactone, PLCL) stent coatings would suppress migration and invasion of vascular smooth muscle cells (VSMCs) as well as platelet-mediated thrombosis. EGCG-releasing PLCL (E-PLCL) was prepared by blending PLCL with 5% EGCG. The surface morphology, roughness and melting temperature of PLCL were not changed despite EGCG addition. EGCG did, however, EGCG appreciably increase the hydrophilicity of PLCL. EGCG was found to be uniformly dispersed throughout E-PLCL without direct chemical interactions with PLCL. E-PLCL displayed diffusion controlled release of EGCG release for periods up to 34 days. E-PLCL significantly suppressed the migration and invasion of VSMCs as well as the adhesion and activation of platelets. E-PLCL coatings were able to smooth the surface of bare stents with neither cracks nor webbings after balloon expansion. The structural integrity of coatings was sufficient to resist delamination or destruction during 90% dilatation. These results suggest that EGCG-releasing polymers can be effectively applied for fabricating an EGCG-eluting vascular stent to prevent in-stent restenosis and thrombosis. 相似文献
18.
A transitory but significant stimulation of mitochondrial activity, increase of reactive oxygen species (ROS) and oxidative stress were previously observed in L929 fibroblasts cultured on poly(epsilon-caprolactone) (PCL) films. ROS, mainly formed in mitochondria, play a physiological role but an excessive production can promote endothelial dysfunction, cause oxidative injury to vascular cells, oxidize lipoproteins and accelerate atherothrombogenesis. On the other hand, mitochondria have a crucial position in programmed cell death control and are responsible for ATP synthesis through the coupling of oxidative phosphorylation to respiration. This coupling requires the existence of a mitochondrial membrane potential (Deltapsi(m)). The aim of the present study was to evaluate by flow cytometry the ROS content and Deltapsi(m) of both endothelial (EC) and smooth muscle cells (SMC) cultured on PCL films as a potential substrate for vascular graft development. Cell size, internal complexity and cell cycle were also analyzed to detect the possible appearance of the subG(1) cell fraction, characteristic of apoptotic cells. The effect of treating PCL films with NaOH before culture was also studied. PCL decreases the ROS content of EC during the culture but produces an increase of these levels in SMC after 7 days. PCL also induces variations of Deltapsi(m) which show a significant parallelism with the changes observed in ROS levels proving the importance and sensitivity of these measurements as indicators of the mitochondrial function. The treatment of PCL with NaOH decreases these effects demonstrating the benefits of increasing the surface hydrophilicity before cell culture which improves cell adhesion and proliferation and reduces oxidative stress. Since no important changes have been detected in subG(1) fraction of EC and SMC cultured on either PCL or PCL-NaOH, the changes of Deltapsi(m) observed in the present study cannot be related to apoptosis. These results confirm the potential utility of PCL as a suitable scaffold in Vascular Tissue Engineering. 相似文献
19.
K P Dingemans C A Wagenvoort 《Laboratory investigation; a journal of technical methods and pathology》1976,35(3):205-212
The pulmonary vessels of rats treated with fulvine were studied electron microscopically for morphologic signs of contraction of smooth muscle cells. Except for a number of indirect indications of vasoconstriction such as medial smooth muscle cell hypertrophy and excessive crenation of elastic laminae, conspicuous smooth muscle cell excrescences were observed and were interpreted to be a direct result of contraction. The close relationship between contraction and smooth muscle cell excrescences was confirmed by their simultaneous occurrence within 1 minute after administration of histamine to isolated perfused guinea pig lungs. The images of pulmonary vessels of rats with prolonged survival times after fulvine administration suggested a gradual increase in the size of the excrescences with a simultaneous degeneration of their cytoplasmic content, and in some cases their eventual detachment from the main cell body. The latter changes were possibly associated with the widespread vasculitis that often occurred at longer intervals after fulvine application. The smooth muscle cell excrescences in pulmonary veins were generally much more prominent than those in pulmonary arteries. This difference was probably caused by the more rigid structure of the arterial wall whcih prevented the formation of large excrescences. The mechanism of the formation of smooth muscle cell excrescences, their possible general validity as markers of vascular smooth muscle cell contraction, and the implications for the mechanism of action of fulvine are briefly discussed. 相似文献
20.
H Palmer P Libby 《Laboratory investigation; a journal of technical methods and pathology》1992,66(6):715-721
Positive and negative signals regulate the proliferation in vitro of vascular smooth muscle cells (SMC), a principle cell type in the blood vessel wall. Immune interferon (IFN-gamma, a type II IFN) retards the growth of human SMC, but the effect of type I IFN (IFN-alpha or beta) is unknown. Furthermore, the capacity of SMC to produce IFN is uncharacterized. If type I IFN alters SMC growth and is produced by this cell type, an autocrine inhibitory loop could operate in vascular growth control. To test this possibility, we compared the effects of IFN-alpha, beta, and gamma on the growth of SMC stimulated by platelet-derived growth factor, interleukin-1 or tumor necrosis factor alpha. IFN-beta and IFN-gamma, but not IFN-alpha, consistently retarded growth of SMC cultures (measured by net DNA accumulation and cell number). We investigated whether SMC could produce IFN-beta, a mediator characteristically produced by fibroblasts. Vascular SMC treated with poly(I):poly(C) or tumor necrosis factor-alpha expressed IFN-beta mRNA. SMC treated with poly(I):poly(C) or Newcastle Disease virus elaborated biologically active IFN-beta as well. Our results establish that IFN-beta inhibits human vascular SMC growth and that these cells can express the IFN-beta gene. These findings show that human vascular SMC have the capacity of producing a potential autocrine growth regulator. 相似文献