RESUMO
A novel polyvinyl alcohol-amino multi-walled carbon nanotube (PVA-AMWCNT) nanocomposite microsphere was prepared successfully for the first time and used for endotoxin removal. The resulting AMWCNT modified PVA microsphere was characterized by SEM, Raman spectrum and fluorescence image, which indicated AMWCNT was dispersed into the macropores of PVA microsphere uniformly. The PVA-AMWCNT microspheres showed better adsorption capability and faster adsorption equilibrium for endotoxin in aqueous solution when compared to the PVA microsphere with polymyxin B (PMB) as ligand. More noteworthy, the PVA based microspheres had little nonspecific adsorption in simulated serum. Therefore, PVA-AMWCNT nanocomposite microsphere with an excellent haemocompatibility has a great potential application in clinical blood purification.
Assuntos
Endotoxinas/química , Endotoxinas/isolamento & purificação , Microesferas , Nanocompostos/química , Nanotubos de Carbono/química , Álcool de Polivinil/química , Adsorção , Hemoperfusão , Cinética , Membranas ArtificiaisRESUMO
Chitosan-carbon nanotube composite beads combines the advantages of chitosan in forming a stable biocompatible framework and carbon nanotube that provide nanometer effects (high strength and high specific surface area etc.). In this study, chitosan/amino multiwalled carbon nanotubes (CS/AMWCNT) composite beads was prepared by phase-inversion method, in which CS and AMWCNT was crosslinked by ethylene glycol diglycidyl ether (EGDE). The CS/AMWCNT nanocomposite beads produced has been characterized by BET, SEM, TGA, and Raman spectroscopy which exhibited enhanced thermal stability due to the incorporation of AMWCNT. Mechanical test results showed that mechanical strength of the CS/AMWCNT composite beads was significantly enhanced when comparing to unmodified chitosan beads, the breakage percentage decreased from 34.1% to 0.67%. The adsorption capacity for bilirubin was measured in PBS and BSA solutions, and the CS/AMWCNT composite beads with 5 wt% AMWCNT showed much higher adsorption capacity (12.7 mg/g in PBS and 7.6 mg/g in BSA) to bilirubin than chitosan beads (8.5 mg/g in PBS and 4.2 mg/g in BSA). Our nanocomposite beads with excellent hemocompatibility has a high potential application in blood purification as an efficient adsorbent for bilirubin. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 96-103, 2018.
Assuntos
Bilirrubina/química , Quitosana/química , Hemoperfusão , Teste de Materiais , Nanocompostos/química , Nanotubos de Carbono/química , Adsorção , Animais , CoelhosRESUMO
A novel nano-CaCO3/polystyrene nanocomposite adsorbent (NPS-8) was synthesized for efficient bilirubin removal from human plasma. A comparison with the polystyrene adsorbent (PS-8), which was without the incorporation of nano-CaCO3, revealed that NPS-8 had superior bilirubin adsorption capacity and mechanical strength. The resulting nano-CaCO3 reinforced PS-8 (NPS-8) was tested by transmission electron microscopy (TEM), scanning electron microscopy (SEM), mechanical strength test, and bilirubin adsorption assays. The adsorption results indicated that NPS-8 displayed better adsorption capacity for bilirubin (91%) than that of PS-8 (75.88%). The mechanical strength of NPS-8 was significantly greater than that of PS-8. In addition, both PS-8 and NPS-8 possessed good blood compatibility properties (a negligible hemolytic activity and platelet adhesion). Therefore, a conclusion could be drawn that NPS-8 has a high potential as an efficient bilirubin adsorbent for blood purification in clinical practice. At the same time, the success of organic-inorganic nanocomposite adsorbents might provide a new insight into the improvement of adsorbents in hemoperfusion.
Assuntos
Bilirrubina/isolamento & purificação , Carbonato de Cálcio/química , Nanocompostos/química , Poliestirenos/química , Adsorção , Bilirrubina/sangue , Bilirrubina/química , Remoção de Componentes Sanguíneos/métodos , Humanos , Microscopia Eletrônica de Varredura , Microscopia Eletrônica de Transmissão , Nanocompostos/ultraestruturaRESUMO
PS-DVB/nano-CaCO3, a novel abundant mesoporous structured polymer nano-composite, was successfully synthesized via suspension polymerization. Characterization of this type of bead nano-composite demonstrated that it exhibits significantly enhanced TNF-α adsorption from blood plasma and possesses good mechanical strength.
Assuntos
Carbonato de Cálcio/química , Nanocompostos/química , Poliestirenos/química , Fator de Necrose Tumoral alfa/química , Fator de Necrose Tumoral alfa/isolamento & purificação , Compostos de Vinila/química , Humanos , Tamanho da Partícula , Polimerização , Porosidade , Propriedades de SuperfícieRESUMO
A multi-functional polystyrene based adsorbent (NKU-9) with a unique mesoporous and a high surface area was prepared by suspension polymerization for removal of therapeutic toxins in blood purification. The adsorbent produced had an almost equal amount of mesopore distribution in the range from 2 to 50 nm. The adsorption of serum toxins with different molecular weights were examined by in vitro adsorption assays and compared with some clinical currently used adsorbents such as HA-330, Cytosorb and BL-300 which are produced by China, America and Japan, respectively. Test results indicated that the adsorption rate for pentobarbital by NKU-9 was 81.24% which is nearly as high as HA-330 (81.44%). The latter adsorbent is currently used for acute detoxification treatment in China. To reach adsorption equilibrium, NKU-9 was faster than HA-330, which implies short treatment time. For the removal of middle molecular toxins such as ß2-microglobulin (98.88%), NKU-9 performed better adsorptive selectivity than Cytosorb (92.80%). In addition, NKU-9 showed high performance for the removal of albumin-bound toxins (e.g., bilirubin), and its adsorption rate for total bilirubin (80.79%) in plasma was 8.4% higher than that of anion exchange resin BL-300 which is currently used to eliminate bilirubin in clinic. Therefore, our results indicate that the newly developed adsorbent with a wide distribution and almost equal amount of mesopores is a multifunctional adsorbent for high efficient removal of serum toxins with different molecular weights which might be an excellent blood purification adsorbent especially to treat diseases that conventional medical methods are low or not efficient.
RESUMO
Strong base anion exchange resin (e.g., BL-300) is currently used to eliminate bilirubin from blood in clinical hemoperfusion. However, there is a potential weakness of the resin to remove heparin and activation of coagulation. To overcome this weakness, we prepared a novel non-ionic macroporous polystyrene adsorbent (SZ-9) with rich mesopores and high surface area by suspension polymerization. Test results indicated that SZ-9 performed better adsorption capacity for bilirubin than BL-300. In addition, better blood compatibility was observed during whole blood hemoperfusion. Therefore, adsorbent SZ-9 might be an efficient alternative to anion exchange resin for the removal of bilirubin in hemoperfusion.
Assuntos
Hemoperfusão/métodos , Falência Hepática/sangue , Falência Hepática/terapia , Poliestirenos/química , Animais , Bilirrubina/sangue , Bilirrubina/isolamento & purificação , Teste de Materiais , Porosidade , CoelhosRESUMO
To enhance vascularization of hydrophobic implants in vivo, a VEGF-Fc fusion protein consisting of vascular endothelial growth factor (VEGF) fused to the immunoglobulin G Fc domain was prepared as an artificial extracellular matrix (ECM). VEGF-Fc was stably immobilized on a polystyrene plate due to the hydrophobicity of the Fc domain, and significantly enhanced the adhesion of human umbilical vein endothelial cells (HUVECs). Additionally, the use of VEGF-Fc as an ECM markedly promoted the proliferation of HUVECs longer than 72 h and induced the reorganization of actin filaments into larger stress fibers within these cells. The VEGF-Fc fusion protein may be a promising artificial ECM for enhancing endothelial cell growth.
Assuntos
Proliferação de Células , Células Endoteliais/fisiologia , Técnicas de Cultura de Células/métodos , Matriz Extracelular/genética , Matriz Extracelular/metabolismo , Humanos , Interações Hidrofóbicas e Hidrofílicas , Fragmentos Fc das Imunoglobulinas/genética , Fragmentos Fc das Imunoglobulinas/metabolismo , Poliestirenos/química , Ligação Proteica , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Fatores de Tempo , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
Various types of porous resin adsorbents based on polystyrene, agarose, and cellulose as matrixes coupling with DNA, amino acids and other biological active molecules as ligands were extensively studied in China. Molecular recognition between the ligand and pathogenic molecule was investigated. Several commercialized products are now widely used in hospitals all over China. Whole blood hemoperfusion is used to treat patients suffering from autoimmune diseases, uremia acute intoxication, and hyperbilirubinemia. Clinical performances of hundreds and thousands of patients treated by whole blood sorption therapy show that the therapy is safe, efficient, and cost-effective.
Assuntos
Hemoperfusão/métodos , Imunoadsorventes , Troca Plasmática/métodos , Resinas Sintéticas/química , Resinas Sintéticas/síntese química , Desintoxicação por Sorção/métodos , Adsorção , Sítios de Ligação de Anticorpos/imunologia , Celulose/síntese química , Celulose/química , China , Análise Custo-Benefício , DNA/imunologia , Humanos , Polimetil Metacrilato/síntese química , Polimetil Metacrilato/química , Poliestirenos/síntese química , Poliestirenos/química , Álcool de Polivinil/síntese química , Álcool de Polivinil/química , Sefarose/síntese química , Sefarose/químicaRESUMO
The whole blood immuno-adsorption (WBIA) system, using an adsorbent to remove pathogenic antibodies of Myasthenia Gravis (MG), was studied. Cellulose-tryptophan adsorbent was synthesized and purified in our lab. Experimental autoimmune myasthenia gravis (EAMG) rabbits were passively transferred with immunoglobulin from patients with myasthenia gravis. The rabbits underwent extracorporeal whole blood adsorption for 2 hours. Results showed no significant damage to blood cells and no changes in the concentrations of electrolytes. Total protein decreased by 12.6% (P<0.05) and globulin protein decreased 21.9% (P<0.05). The overall removal of antibodies against nicotinic acetylcholine receptor (nAChR) was 49.85%. The percentage of decrement of compound muscle action potential in 3, 5, 10 Hz of EAMG rabbits all dropped down after the treatment. The quantity of neuromuscular junctions per unit area (25 mm(2)) increased significantly after treatment (P<0.05). In conclusion, the adsorbent was biocompatible, safe for whole blood immuno-adsorption. Whole blood immuno-adsorption improved clinical manifestation and neuromuscular function of the passively transferred EAMG rabbits.
Assuntos
Celulose/química , Circulação Extracorpórea , Imunoglobulinas/imunologia , Imunoglobulinas/isolamento & purificação , Miastenia Gravis/imunologia , Miastenia Gravis/terapia , Triptofano/química , Adolescente , Adsorção , Adulto , Animais , Materiais Biocompatíveis/química , Criança , Feminino , Humanos , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Imunoglobulina G/isolamento & purificação , Imunoglobulinas/sangue , Masculino , Pessoa de Meia-Idade , Miastenia Gravis/fisiopatologia , Junção Neuromuscular/fisiopatologia , Coelhos , Receptores Nicotínicos/imunologia , Adulto JovemRESUMO
The aim of the present study was to investigated the construction of polycaprolactone-lecithin (PCL-L) electrospun fibers as a novel scaffold material for a tissue-engineered ureter. The effect of bone marrow mesenchymal stem cells (BM-MSCs) on the neovascularization of the scaffolds and the viability of planted urothelial cells (UCs) on PCL-L were also studied. UCs were obtained from New Zealand rabbit bladders, cultured and then seeded onto the lumen of the tubular scaffolds before being subcutaneously transplanted into the space of nude mice. The cultured UCs showed vacuolar degeneration after 7 days of transplantation and they gradually degraded thereafter. To facilitate the regeneration of the tissue-engineered ureter and the survival of UCs in the implant, MSCs were seeded into the tubular grafts by rolling up the nanofibrous membrane, followed by the seeding of UCs. This facilitated the survival of the UCs, which formed several cellular layers after 30 days. The mean microvessel density was significantly increased in tissues seeded with MSCs. Cell-tracking experiments revealed that the transplanted MSCs did not integrate directly into capillaries for angiogenesis. Our results demonstrated that the PCL-L electrospun fibrous scaffold has a high potential for a tissue-engineered ureter especially when seeded with BM-MSCs, which enhanced angiogenesis.
Assuntos
Lecitinas , Transplante de Células-Tronco Mesenquimais , Poliésteres , Alicerces Teciduais/química , Transplantes , Ureter/cirurgia , Bexiga Urinária/transplante , Animais , Transplante de Medula Óssea , Sobrevivência de Enxerto , Nanofibras/química , Coelhos , Ratos , Ratos Nus , Bexiga Urinária/citologia , Urotélio/citologia , Urotélio/transplanteRESUMO
A new low-density lipoprotein (LDL) adsorbent with phosphate groups as the ligand was prepared in this study. Macroporous poly(vinyl acetate-co-triallyl isocyanurate) microspheres were prepared using a free-radical suspension polymerization method. A hydrolysis reaction in sodium hydroxide/methanol changed the materials into poly(vinyl alcohol) (PVA) microspheres. Further reaction with phosphorus oxychloride in anhydrous DMF led to the LDL adsorbent PVA-phosphate microspheres. The preparation conditions such as reaction time, temperature and the amount of phosphorus oxychloride were optimized. The adsorption of plasma lipoproteins was examined by in vitro adsorption assays. The influence of adsorption time, plasma volume and ionic strength on the adsorption capacity was investigated. The circulation adsorption showed that the pathogenic lipoproteins in the plasma such as total cholesterol (TC), LDL and triglyceride (TG) could be removed markedly, in which the removal percentages were 42.9%, 45.0% and 44.74%, respectively. However, the reduction of high-density lipoprotein (HDL) and other normal plasma components was very slight. For in vivo experiment, rabbits were fed with high-cholesterol food to develop a hyperlipidemia model and treated by extracorporeal blood perfusion using the PVA-phosphate columns. Eight hyperlipidemia rabbits were treated with the PVA-phosphate adsorbent, and the removal of TC, LDL and TG was 45.03 +/- 6.64%, 48.97 +/- 9.92% and 35.42 +/- 14.17%, respectively. The sterilization and storage tests showed that the adsorbent was chemically and functionally stable. It could be easily sterilized by a common method and stored for months without loss of adsorption capacity. Therefore, this new PVA-phosphate-based LDL adsorbent may have potential for application in LDL apheresis.
Assuntos
Remoção de Componentes Sanguíneos/instrumentação , Microesferas , Álcool de Polivinil/química , Adsorção , Animais , Radicais Livres , Hiperlipidemias/metabolismo , Ligantes , Lipoproteínas LDL/metabolismo , Modelos Químicos , Perfusão , Fosfatos/química , Compostos de Fósforo/química , Porosidade , CoelhosRESUMO
Kidney failure is a common disease with high frequency. Food-grade recombinant bacteria that can effectively remove urea has great potential for treatment of renal failure. A nonpathogenic strain, L. lactis MG1363, was transformed with plasmid pMG36eure, which carries urease gene. The expression of transgene urease in genetically modified L. lactis MG1363 and the urease activity in removal of urea were investigated. It was found that the removal of urea by recombinant L. lactis MG1363 was pH- and nickel-dependent. At pH 6.5 and in the presence of 250 microM of NiSO4, 50 approximately 60% of urea could be removed in 24 hours. The urea removal activity was also evaluated in imitative gastroenteric environment. After being exposed to acidic solution (pH2.5-4.0) for 2 hours, the cells were then grown in a medium containing 0.1 cfu/ml bile acid salt, 30 mg/dl urea, and 250 microM NiSO4 at pH 6.8. The concentration of urea decreased over time, and the removal was about 30% at 10 hours and 65% at 24 hours, respectively. The safety tests were performed by feeding normal rats with either L. lactis MG1363 or recombinant L. lactis MG1363. The two materials did not cause any changes in blood cells and blood biochemical indexes. There were no differences in terms of body weight and water/food consumption between the two materials. These results indicate the safety, feasibility, and capacity of urease gene modified Lactococcus Lactis in removal of urea under the gastroenteric circumstances. Further investigation may generate a food-grade strain for treatment of chronic renal failure.
Assuntos
Terapia Biológica/métodos , Lactococcus lactis/genética , Ureia/metabolismo , Urease/metabolismo , Animais , Engenharia Genética , Concentração de Íons de Hidrogênio , Nefropatias/terapia , Níquel/farmacologia , Ratos , Proteínas Recombinantes , Transformação Bacteriana , Transgenes , Resultado do Tratamento , Urease/administração & dosagem , Urease/genética , Urease/uso terapêuticoRESUMO
Polyamidoamine (PAMAM) dendrimers represent one of the most efficient polymeric gene carriers. This study describes a new family of PAMAM dendrimers that can be synthesized using a Pentaerythritol derivative (PD) as a core that possesses 12 branches. This new approach in the synthesis of divergent dendrimers provided a rapid increase in the number of branches, which made it easier to obtain dendrimers with high generation and large enough molecular size. The PD dendrimers of generations 3-5 synthesized in this study could efficiently condense DNA into nanoscale complexes with slightly positive charges. Their transfection efficiency was evaluated in different cell lines. These PD dendrimers were found to show higher transfection efficiency, but much lower cytotoxicity, than the commercial nonviral gene carriers polyethyleneimine (PEI), polylysine (PLL), and PAMAM dendrimers with an ethylenediamine core (generations 5 and 7). The results indicate that, with high transfection efficiency and low cytotoxicity, the PD dendrimers hold promise as novel nonviral gene carriers.
Assuntos
DNA/genética , Dendrímeros/química , Vetores Genéticos/química , Substâncias Macromoleculares/química , Plasmídeos/genética , Propilenoglicóis/química , Animais , Células COS , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Chlorocebus aethiops , DNA/administração & dosagem , Dendrímeros/síntese química , Dendrímeros/farmacologia , Sistemas de Liberação de Medicamentos , Avaliação Pré-Clínica de Medicamentos , Técnicas de Transferência de Genes , Humanos , Substâncias Macromoleculares/síntese química , Substâncias Macromoleculares/farmacologia , Camundongos , Células NIH 3T3 , Tamanho da Partícula , Plasmídeos/administração & dosagem , Propriedades de Superfície , TransfecçãoRESUMO
Despite their increasing uses for cardiovascular and cerebrovascular tissue engineering, synthetic polymeric conduits still have their limitations in clinical applications, particularly in small vessels, mainly due to the thrombus formation. Seeding the synthetic scaffolds with endothelial cells (ECs) will potentially solve this problem, but this endothelialization approach demands synthetic materials with better hemacompatibility and cell affinity. To improve the currently used materials and screen for better surface properties, we synthesized copolymer of poly(lactide-co-beta-malic acid) (PLMA), and its derivatives with pendant hydroxyl arms (PLMAHE) or extended carboxyl arms (PLMA-ECA). We analyzed their physical and chemical properties, their hydrophilicity, and their degradation in physiological conditions. More importantly, their blood compatibility was investigated by the measurements of prothrombin time, activated partial thromboplastin time, and interaction with platelets; their cell affinity and cell growth potentials were observed using the human umbilical vein EC cultures. Results from these experiments showed that the copolymer with the carboxyl arms attracted little platelets, and exhibited better cell affinity and supported the cell proliferation, thus demonstrating the potential usefulness of PLMA-ECA for tissue engineering. We speculate that this novel material will offer new opportunities for the design of better vascular-engineered scaffolds owing to its improved biological and chemical properties.
Assuntos
Materiais Biocompatíveis , Células Endoteliais/metabolismo , Poliésteres/química , Polímeros/química , Engenharia Tecidual/métodos , Plaquetas/efeitos dos fármacos , Plaquetas/metabolismo , Plaquetas/ultraestrutura , Adesão Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Células Endoteliais/efeitos dos fármacos , Endotélio Vascular/citologia , Humanos , Ácido Láctico/química , Malatos/síntese química , Malatos/química , Poliésteres/síntese química , Poliésteres/farmacologia , Polímeros/síntese química , Tempo de Protrombina , Propriedades de Superfície , Veias Umbilicais/citologia , Água/químicaRESUMO
AIMS: Controversy exists regarding the effects of granulocyte colony-stimulating factor (G-CSF) on post-infarction remodelling, which is regulated by matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs). The aim of this study was to investigate the impact of G-CSF administration on cardiac MMP/TIMP ratios and long-term remodelling in a rat model of acute myocardial infarction (MI). METHODS AND RESULTS: Sprague-Dawley rats underwent coronary ligation to produce MI. Rats surviving the MI for 3 h were randomized to receive G-CSF (50 microg/kg/day for 5 consecutive days, n = 16) or saline (n = 10). Sham-operated animals received no treatment (n = 10). G-CSF injection significantly increased circulating white blood cells, neutrophils, and monocytes. Western blotting revealed that the ratios of MMP-2/TIMP-1 and MMP-9/TIMP-1 were significantly decreased in the infarcted myocardium. At 3 months, echocardiographic and haemodynamic examinations showed that the G-CSF treatment induced left ventricular (LV) enlargement and dysfunction. Histological analysis revealed that the extent of myocardial fibrosis and infarct size were larger in the G-CSF group than in the Saline group. Furthermore, G-CSF treated animals showed a significantly lower post-MI survival during the study period. CONCLUSION: Decrease of cardiac MMP/TIMP ratios by G-CSF after infarction may be important as a mechanism in promotion of myocardial fibrosis, which further facilitates infarct expansion and LV dysfunction.
Assuntos
Oclusão Coronária/complicações , Fator Estimulador de Colônias de Granulócitos/efeitos adversos , Fator Estimulador de Colônias de Granulócitos/farmacologia , Infarto do Miocárdio/etiologia , Infarto do Miocárdio/patologia , Miocárdio/patologia , Animais , Pressão Sanguínea/efeitos dos fármacos , Modelos Animais de Doenças , Feminino , Fibrose , Ventrículos do Coração/diagnóstico por imagem , Ventrículos do Coração/metabolismo , Ventrículos do Coração/patologia , Leucócitos/patologia , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Monócitos/patologia , Infarto do Miocárdio/metabolismo , Miocárdio/metabolismo , Neutrófilos/patologia , Ratos , Ratos Sprague-Dawley , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Ultrassonografia , Remodelação Ventricular/efeitos dos fármacosRESUMO
G250 is a tumor associated antigen that is found on > 90% of renal cell carcinoma (RCC). In order to develop a highly targeting gene vector for RCC gene therapy, G250 monoclonal antibody was prepared, purified and characterized. The antibody was chemically bound to Polyethylenimine (PEI) to form the IgG-PEI conjugate. The conjugate is capable of forming DNA complexes in the size of nano meters and with a narrow size distribution. The targeting effect and transfection efficiency were tested on five cell lines, ketr 3, Hela, ACHN, HepG2, and smooth muscle cells. The transfection was quantitatively determined by fluorescence activated cell sorting (FACS) and luciferase assay. The FACS results show that for G250 positive cells ketr 3 and Hela, the transfection efficiency of IgG-PEI are 2-fold higher than that of PEI. But for G250 negative cells, antibody modification has no effect on transfection. The expression of luciferase in ketr 3 cells which is expressed as enzyme activity is 15-fold and 61-fold higher than that in ACHN and SMC, respectively. In the presence of free antibody, the targeting effect of IgG-PEI is impaired and the transfection efficiency is normalized. It indicates that G250 antibody is an ideal targeting ligand for delivery of genes into RCC. Application of this IgG-PEI conjugate in RCC gene therapy will be of great interest.
Assuntos
Anticorpos Monoclonais/farmacologia , Carcinoma de Células Renais/terapia , DNA/farmacologia , Terapia Genética , Imunoglobulina G/farmacologia , Neoplasias Renais/terapia , Polietilenoimina/farmacologia , Animais , Anticorpos Monoclonais/química , Antígenos de Neoplasias/metabolismo , Anidrase Carbônica IX , Anidrases Carbônicas/metabolismo , Carcinoma de Células Renais/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , DNA/química , Feminino , Técnicas de Transferência de Genes , Células HeLa , Humanos , Imunoglobulina G/química , Neoplasias Renais/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Miócitos de Músculo Liso/metabolismo , Células NIH 3T3 , Plasmídeos/genética , Polietilenoimina/química , CoelhosRESUMO
Agar-gel resin with tryptophan as ligand was synthesized as an adsorbent for the removal of Immunoglobulin E (IgE) in bronchial asthma. Factors affecting the adsorption properties of the adsorbent, such as the different kind of amino acid linked to the carrier, NaOH concentration and amount of epichlorohydrin used in activation, were studied in detail. The effect of spacer on the adsorption capacity was also investigated. Results showed that the agar-tryptophan adsorbent with a spacer could enhance the adsorption capacity to 70.73% (1309.403IU/g agar-gel), which was higher than that without spacers (53%). The selectivity and hemocompatibility of the adsorbent were also examined, which showed good selectivity for IgE and satisfactory blood compatibility.
Assuntos
Asma/imunologia , Imunoglobulina E/química , Imunoglobulina E/isolamento & purificação , Adsorção , Ágar/química , Aminoácidos/química , Soluções Tampão , Custos e Análise de Custo , Géis/química , Histocompatibilidade , Concentração de Íons de Hidrogênio , Imunoglobulina E/imunologia , Ligantes , Anidridos Ftálicos/química , Sensibilidade e Especificidade , Hidróxido de Sódio/química , Anidridos Succínicos/química , Triptofano/químicaRESUMO
Acquired or congenital abnormalities may lead to urethral damage or loss, often requiring surgical reconstruction. Urethrocutaneous fistula and strictures are common complications, due to inadequate blood supply. Thus, adequate blood supply is a key factor for successful urethral tissue reconstruction. In this study, urethral grafts were prepared by seeding rabbit bladder urothelial cells (UCs) modified with human vascular endothelial growth factor (VEGF(165)) gene in the decellularized artery matrix. A retroviral pMSCV-VEGF(165)-GFP vector was cloned by insertion of VEGF open reading frame into the vector pMSCV-GFP (murine stem cell virus [MSCV]; green fluorescent protein [GFP]). Retrovirus was generated using package cell line 293T. Rabbit UCs were expanded ex vivo and modified with either MSCV-VEGF(165)-GFP or control MSCV-GFP retrovirus. Transduction efficiency was analyzed by fluorescence-activated cell sorting. The expression of VEGF(165) was examined by immunofluorescence, reverse transcript-polymerase chain reaction, Western blot, and enzyme-linked immunosorbent assay (ELISA). Decellularized rabbit artery matrix was seeded with genetically modified UCs and was subsequently cultured for 1 week prior to subcutaneous implantation into nude mice. Four weeks after implantation, the implants were harvested and analyzed by fluorescence microscopy, and by histologic and immunohistochemical staining. Ex vivo transduction efficiency of UCs was greater than 50% when concentrated retrovirus was used. The modified cells expressed both VEGF and GFP protein. Furthermore, the VEGF-modified UCs secreted VEGF in a time-dependent manner. Scanning electron microscopy and histochemical analysis of cross sections of the cultured urethral grafts showed that the seeded cells were attached and proliferated on the luminal surface of the decellularized artery matrix. In the subcutaneously implanted vessels, VEGF-modified cells significantly enhanced neovascularization and the formation of a urethral layer compared to GFP-modified cells. These results indicate that VEGF gene therapy may be a suitable approach to increase the blood supply in tissue engineering for treatment of urethral damage or loss.
Assuntos
Terapia Genética/métodos , Engenharia Tecidual/métodos , Uretra/irrigação sanguínea , Uretra/crescimento & desenvolvimento , Urotélio/citologia , Urotélio/transplante , Fator A de Crescimento do Endotélio Vascular/genética , Animais , Artérias/anatomia & histologia , Artérias/fisiologia , Técnicas de Transferência de Genes , Humanos , Camundongos , Coelhos , Alicerces Teciduais , Uretra/anormalidades , Uretra/citologia , Urotélio/irrigação sanguínea , Urotélio/crescimento & desenvolvimentoRESUMO
The current work investigated whether 1,25-dihydroxyvitamin D(3)(1,25-(OH)(2)D(3)) can promote the neovascularization of tissue-engineered bone. Human osteoblast-like cells (HOB) and endothelial cells (EC) were isolated and cultured. HOB and EC were inoculated at the ratio of 2:1 onto the coral-derived hydroxyapatite (CHA) scaffolds coated with and without 1,25-(OH)(2)D(3). Tissue-engineered bones were cultured for 3 days before implantation into the backs of nude mice. Four and 8 weeks after the operation, the retrieved scaffolds and cells were examined histologically and by scanning electron microscope, and the vascular area was measured. The immature bone grew into the pores of CHA scaffolds in both groups. At each time interval, there was a conspicuous neovascularization in the 1,25-(OH)(2)D(3) treatment group, with a larger amount of new capillaries accompanying immature bone. In the 1,25-(OH)(2)D(3) group, scanning electron microscopy revealed luminal sprouting from the larger vessels. Maturation of the new bone was paralleled by the occurrence of the new capillaries. The vascular areas were 28.74% +/- 7.81% and 19.52% +/- 4.57% at 4-week intervals (p < 0.05) and 24.66% +/- 7.38% and 17.84% +/- 5.22% at 8-week intervals (p < 0.05) in test and control groups, respectively. These results imply that 1,25-dihydroxyvitamin D(3) may be useful as a cytokine for tissue engineering bone for neovascularization.
Assuntos
Osso e Ossos/irrigação sanguínea , Osso e Ossos/efeitos dos fármacos , Comunicação Celular/efeitos dos fármacos , Células Endoteliais/efeitos dos fármacos , Neovascularização Fisiológica/efeitos dos fármacos , Osteoblastos/efeitos dos fármacos , Vitamina D/análogos & derivados , Animais , Osso e Ossos/citologia , Osso e Ossos/ultraestrutura , Células Cultivadas , Células Endoteliais/citologia , Humanos , Camundongos , Camundongos Nus , Osteoblastos/citologia , Vitamina D/farmacologiaRESUMO
Using chitosan as raw materials, a suitable size (300-500 mum) of porous microcarrier was fabricated by suspension crosslinking and lyophilizing method, which made the carrier has an average pore size of 50 microm and 86% porosity. The microcarrier was modified with lactose and maltose respectively. Various factors that influenced the preparation of microcarrier were studied and the reaction conditions were optimized. Rat hepatocytes cultured on modified microcarrier retained a spherical shape which is similar to those in vivo and formed aggregates. The metabolic activities of cells on lactose-modified were higher than those on maltose-modified microcarrier. The highest albumin secretion reached 54.8 microg/10(6 )cells/d, and the highest urea synthesis reached 4.65 micromol/10(6)cells/d, which may be promoted by the formation of cellular aggregates. In conclusion, lactose-modified porous microcarrier is promising scaffold for hepatocytes culture.