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1.
Vet Microbiol ; 114(1-2): 123-33, 2006 Apr 16.
Artículo en Inglés | MEDLINE | ID: mdl-16386382

RESUMEN

The aim of this study was to determine the prevalence and the transmission routes of Arcobacter spp. in sows and their offspring on a breeding farm. Twelve Arcobacter-positive sows and their litters were studied for this purpose. Analysis of rectal samples showed a high prevalence of Arcobacter spp. among the sows (approximately 42% of the sows carried one or more Arcobacter species). Intermittent excretion of one particular species and shifts in excretion from one species to another were observed in individual animals over time. The detection of Arcobacter spp. in amniotic fluid of the sows and in rectal samples from newborn piglets (ranging from 38.5-83.3% per litter), as well as the high similarity between PFGE profiles of Arcobacter isolates from sows and their respective newborns indicated the existence of an intra-uterine transmission route for Arcobacter spp. Specific antibodies against Arcobacter spp. were detected in colostrum by Western blot. At 2 weeks of age, only a few piglets were positive for Arcobacter. The reappearance of Arcobacter in these piglets at Week 3 and the shift in the Arcobacter species detected (from a prominent presence of A. cryaerophilus at birth to the presence of A. skirrowii and A. butzleri at 3 weeks after birth) showed that a post-natal infection route from their mothers, newcomers or the environment to the piglets existed. Thus, in this manuscript the transmission of Arcobacter spp. (both vertical and horizontal) from carrying sows to their offspring is demonstrated.


Asunto(s)
Arcobacter , Transmisión de Enfermedad Infecciosa/veterinaria , Infecciones por Bacterias Gramnegativas/veterinaria , Transmisión Vertical de Enfermedad Infecciosa/veterinaria , Enfermedades de los Porcinos/microbiología , Enfermedades de los Porcinos/transmisión , Líquido Amniótico/microbiología , Animales , Anticuerpos Antibacterianos/análisis , Arcobacter/clasificación , Arcobacter/genética , Arcobacter/aislamiento & purificación , Western Blotting/veterinaria , Calostro/inmunología , Cartilla de ADN/química , ADN Bacteriano/química , Electroforesis en Gel de Campo Pulsado/métodos , Electroforesis en Gel de Campo Pulsado/veterinaria , Femenino , Infecciones por Bacterias Gramnegativas/epidemiología , Infecciones por Bacterias Gramnegativas/microbiología , Infecciones por Bacterias Gramnegativas/transmisión , Reacción en Cadena de la Polimerasa/métodos , Reacción en Cadena de la Polimerasa/normas , Reacción en Cadena de la Polimerasa/veterinaria , Recto/microbiología , Sensibilidad y Especificidad , Enfermedades de los Porcinos/epidemiología , Factores de Tiempo
2.
Biochim Biophys Acta ; 818(3): 343-51, 1985 Sep 10.
Artículo en Inglés | MEDLINE | ID: mdl-2412582

RESUMEN

A new method that involves the use of the cation exchange resin Dowex 50W-X4 to remove non-encapsulated drugs from liposome dispersions was investigated. Cytostatic drugs widely varying in their molecular structure can be removed from aqueous solutions by Dowex 50W-X4. The applicability of the resin to separate free from liposome-bound drugs was illustrated for a number of cytostatic drugs (cisplatin, doxorubicin, vincristine). The technique presented allows for a rapid, efficient and convenient procedure for the free drug removal from liposome dispersions without dilution of the liposomal preparation. Studies with liposome-encapsulated drugs will be facilitated by the use of this method, since it avoids many of the problems introduced by conventional methods as dialysis, gel filtration and centrifugation/washing. To elucidate the interaction mechanism of doxorubicin with Dowex 50W-X4, alternative adsorbents were studied for their doxorubicin binding properties. In the adsorption process of doxorubicin onto Dowex 50W-X4 both electrostatic (ion exchange) and hydrophobic effects play a role. The results indicate that hydrophobic contributions to the interaction are responsible for the high resistance offered by the binding forces against desorption of adsorbed doxorubicin. For other adsorbents the interactions are either mainly of an electrostatic or a hydrophobic nature.


Asunto(s)
Resinas de Intercambio Aniónico , Antineoplásicos/administración & dosificación , Resinas de Intercambio Iónico , Liposomas , Adsorción , Óxido de Aluminio , Cisplatino/administración & dosificación , Citarabina/administración & dosificación , Dextranos , Doxorrubicina/administración & dosificación , Metotrexato/administración & dosificación , Poliestirenos , Resinas Sintéticas , Dióxido de Silicio , Temperatura , Vincristina/administración & dosificación
3.
Biochim Biophys Acta ; 1419(2): 325-34, 1999 Jul 15.
Artículo en Inglés | MEDLINE | ID: mdl-10407083

RESUMEN

Rheumatoid arthritis (RA) is a prevalent and debilitating autoimmune disease that affects the joints. RA is characterized by an infiltration of the affected joint by blood-derived cells. In response to activation, these cells generate reactive oxygen species, resulting in an oxidative stress situation. One approach to counteract this oxidative stress situation is the use of antioxidants as therapeutic agents. The free radical scavenger enzyme superoxide dismutase (SOD) may be used as a therapeutic agent in rheumatoid arthritis, but its rapid elimination from the circulation is a major limitation. Targeted delivery of SOD may overcome this limitation. In this study, the utility of PEGylated liposomes (PEG-liposomes) for targeting SOD to arthritic sites was explored. The targeting of SOD to arthritic sites following intravenous administration of both PEG-liposomes and positively charged liposomes lacking PEG but containing stearylamine (SA-liposomes) in rats with adjuvant arthritis was studied. At 24 h post injection, the blood levels of long circulating liposomes with a mean size of 0.11 micrometer and 0.20 micrometer were 8- and 3-fold higher, respectively, as compared to the SA-liposomes. The majority of SOD administered in liposomal form remains within the liposomes when they circulate in the bloodstream. The highest target uptake was observed with PEG-liposomes with a mean size of 0.11 micrometer and the lowest uptake with the SA-liposomes. These results demonstrate that SOD can be targeted to inflamed sites most efficiently via small-sized PEG-liposomes. Small-sized PEG-coated liposomes are to be preferred if prolonged circulation and enhanced localization of SOD at arthritic sites are desired.


Asunto(s)
Artritis Experimental/tratamiento farmacológico , Superóxido Dismutasa/administración & dosificación , Animales , Artritis Experimental/diagnóstico por imagen , Artritis Experimental/etiología , Modelos Animales de Enfermedad , Portadores de Fármacos , Pie/diagnóstico por imagen , Corazón/diagnóstico por imagen , Radioisótopos de Indio , Inyecciones Intravenosas , Radioisótopos de Yodo , Liposomas/farmacocinética , Masculino , Mycobacterium , Tamaño de la Partícula , Ácido Pentético , Fosfatidiletanolaminas , Polietilenglicoles , Cintigrafía , Ratas , Ratas Wistar , Superóxido Dismutasa/farmacocinética , Factores de Tiempo , Distribución Tisular
4.
Biochim Biophys Acta ; 1419(2): 353-63, 1999 Jul 15.
Artículo en Inglés | MEDLINE | ID: mdl-10407086

RESUMEN

Previously, it was demonstrated that immunoliposomes, bearing anti-intercellular adhesion molecule-1 (ICAM-1) antibodies (mAb F10.2), can specifically bind to different cell types expressing ICAM-1. In this study, we have quantified the amount of immunoliposomes binding to IFN-gamma activated human bronchial epithelial cells (BEAS-2B) in vitro and studied the subsequent fate of cell-bound anti-ICAM-1 immunoliposomes. We demonstrate that binding of the immunoliposomes to the epithelial cells depends on the liposome concentration used. After binding to the cell surface, the anti-ICAM-1 immunoliposomes are rapidly internalised by the epithelial cells. Sixty percent of cell-bound immunoliposomes were internalised by the epithelial cells within 1 h of incubation at 37 degrees C. The results indicate that ICAM-1 targeted immunoliposomes may be used as carriers for the intracellular delivery of anti-inflammatory drugs to sites of inflammation characterised by an increased expression of ICAM-1.


Asunto(s)
Bronquios/inmunología , Molécula 1 de Adhesión Intercelular/inmunología , Liposomas/inmunología , Anticuerpos Monoclonales/inmunología , Adhesión Celular , Línea Celular , Portadores de Fármacos , Células Epiteliales/inmunología , Células Epiteliales/metabolismo , Fluoresceínas , Humanos , Inflamación/tratamiento farmacológico , Inflamación/inmunología , Molécula 1 de Adhesión Intercelular/metabolismo , Interferón gamma , Microscopía Confocal , Factores de Tiempo
5.
FEBS Lett ; 399(3): 232-6, 1996 Dec 16.
Artículo en Inglés | MEDLINE | ID: mdl-8985152

RESUMEN

A human anti-CD22 single chain (sc) Fv antibody fragment from a synthetic phage antibody display library was biosynthetically lipid-tagged by using Escherichia coli lipoprotein sequences. The purified anti-CD22 scFv lipoprotein was incorporated into liposomes by detergent dilution. Anti-CD22 immunoliposomes were shown to bind specifically in a dose- and time-dependent manner to CD22+ cell lines and CD22+ B-lymphocytes present in freshly isolated samples of blood mononuclear cells. The immunoliposomes were demonstrated to accumulate in intracellular compartments. Biosynthetically lipid-tagged human scFv antibody fragments isolated from phage display libraries may facilitate the construction of immunoliposomes with improved properties.


Asunto(s)
Bacteriófagos/genética , Moléculas de Adhesión Celular , Fragmentos de Inmunoglobulinas/genética , Lectinas , Lípidos/química , Liposomas , Antígenos CD/inmunología , Antígenos de Diferenciación de Linfocitos B/inmunología , Sitios de Unión , Vectores Genéticos , Humanos , Fragmentos de Inmunoglobulinas/química , Cinética , Lectina 2 Similar a Ig de Unión al Ácido Siálico
6.
FEBS Lett ; 357(2): 140-4, 1995 Jan 03.
Artículo en Inglés | MEDLINE | ID: mdl-7805880

RESUMEN

The anti-ICAM-1 monoclonal antibody F10.2 was conjugated to liposomes to target to cells expressing the cell adhesion molecule ICAM-1. We demonstrate that F10.2 immunoliposomes bind to human bronchial epithelial cells (BEAS-2B) and human umbilical vein endothelial cells (HUVEC) in a specific, dose- and time-dependent manner. It appears that the degree of ICAM-1 expression is the limiting factor in the degree of immunoliposome binding to the cells. These results are a first step in the strategy for specific drug delivery to target sites characterised by increased expression of adhesion molecules.


Asunto(s)
Anticuerpos Monoclonales/inmunología , Sistemas de Liberación de Medicamentos , Molécula 1 de Adhesión Intercelular/inmunología , Bronquios/citología , Bronquios/inmunología , Línea Celular , Endotelio Vascular/citología , Endotelio Vascular/inmunología , Humanos , Liposomas , Venas Umbilicales
7.
J Nucl Med ; 36(9): 1639-44, 1995 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-7658225

RESUMEN

UNLABELLED: To determine the potential of sterically stabilized liposomes to image infectious and inflammatory foci, the in vivo behavior of 111In-labeled PEGylated (coated with polyethylene glycol) liposomes was studied in a rat model. METHODS: Indium-111-PEGylated lipsomes were administered intravenously to rats infected with S. aureus in the left calf muscle. The distribution of the radiolabel was studied by gamma counting of dissected tissues and gamma camera imaging for 48 hr. As a reference agent, the preparation of 111In-IgG was included in these studies. RESULTS: Clearance of the PEGylated liposomes from the blood compartment was similar to the clearance of 111In-IgG in this model (t1/2 approximately 20 hr). Uptake of the radiolabel in the abscess with the 111In-liposomes was twice as high as the uptake following injection of 111In-IgG (2.7%ID/g versus 1.1%ID/g at 48 hr postinjection). Tissue counting revealed that abscess-to-muscle ratios reached values up to 20 and 34 (24 and 48 p.i., respectively). As early as 1 hr postinjection, the abscess could be visualized scintigraphically. CONCLUSION: The in vivo characteristics of this liposomal formulation in this rat model indicate that sterically stabilized liposomes labeled with gamma emitters might be a valuable addition to the arsenal of radiopharmaceuticals currently used for infection imaging.


Asunto(s)
Infección Focal/diagnóstico por imagen , Radioisótopos de Indio , Absceso/diagnóstico por imagen , Animales , Miembro Posterior , Liposomas , Masculino , Cintigrafía , Ratas , Ratas Wistar , Infecciones Estafilocócicas/diagnóstico por imagen
8.
J Nucl Med ; 38(3): 489-93, 1997 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-9074545

RESUMEN

UNLABELLED: In previous studies we have shown that liposomes sterically stabilized with polyethylene glycol (PEG), preferentially localize in infectious and inflammatory foci. In this study, we further optimized the formulation of PEG liposomes for infection imaging in a rat model. METHODS: The biodistribution and imaging characteristics of different liposomal formulations labeled with 99mTc were determined in rats with S. aureus infection of the left calf muscle. The influence of liposomal size (mean diameter varying from 90 nm to 220 nm) as well as circulation time (modulated by inclusion of 0-10 mole% phosphatidylserine) were studied. RESULTS: The smallest liposomes displayed improved characteristics for infection imaging: 90-nm liposomes revealed the highest abscess uptake (1.6% +/- 0.4% ID/g, 24 hr postinjection) in combination with the lowest splenic accumulation (6.9% +/- 0.7% ID/g, 24 hr postinjection) as compared to the larger sized preparations. Enhanced abscess-to-blood ratios (4.0 versus 1.3 at 24 hr postinjection) were obtained by including 1.0 mole% phosphatidylserine in the lipid bilayer of the PEG liposomes. However, enhanced blood clearance of these liposomes reduced their absolute abscess uptake. CONCLUSION: These results indicate that the in vivo behavior of PEG liposomes can be modulated to optimize their characteristics for infection imaging.


Asunto(s)
Absceso/diagnóstico por imagen , Infección Focal/diagnóstico por imagen , Músculo Esquelético/diagnóstico por imagen , Polietilenglicoles/administración & dosificación , Tecnecio , Animales , Portadores de Fármacos , Liposomas , Masculino , Tasa de Depuración Metabólica , Músculo Esquelético/metabolismo , Tamaño de la Partícula , Polietilenglicoles/farmacocinética , Cintigrafía , Ratas , Ratas Wistar , Tecnecio/farmacocinética , Distribución Tisular
9.
J Nucl Med ; 37(8): 1392-7, 1996 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-8708782

RESUMEN

UNLABELLED: The performance of 99mTc Stealth liposomes was investigated in various rat models. METHODS: Preformed polyethyleneglycol-containing liposomes with encapsulated reduced glutathione, were radiolabeled using the lipophilic 99mTc-HMPAO. The labeled liposomes were intravenously administered to rats with focal S. aureus or E. coli infection, or turpentine-induced inflammation. For comparison, Tc-99m-nanocolloid- and 99mTc-labeled nonspecific IgG were tested. In rats with Pneumocystis carinii pneumonia (PCP), Tc-99m-liposomes were directly compared to In-111 labeled nonspecific IgG. RESULTS: Technetium-99m-liposomes accumulated in the infectious and inflammatory muscle foci over 24 hr (0.59% injected dose per gram tissue (%ID/g) for S. aureus; 1.18 %ID/g for turpentine). Abscess-to-muscle ratios increased to values as high as 24.0, 41.7 and 44.5 for the respective models at 24 hr postinjection. Technetium-99m-liposomes visualized the foci as early as 1 hr postinjection. Technetium-99m-IgG visualized S. aureus infection, but abscess-to-muscle ratios and abscess uptake at the later time points were significantly lower. Technetium-99m-nanocolloid failed to visualize any of the muscle foci. In PCP however, 99mTc-liposomes did not show preferential localization in the infection. The control agent 111In-IgG showed a significant, two-fold increase in lung uptake. CONCLUSION: Technetium-99m-Stealth liposomes preferentially accumulated in abscesses, leading to very high target-to-nontarget ratios. This property appears to be related to a process based on uptake of long-circulating particles. In a specific type of infection, i.c. PCP, 99mTc-liposomes did not accumulate in diseased lung tissue, thus mimicking the in vivo behavior of labeled leukocytes.


Asunto(s)
Absceso/diagnóstico por imagen , Infecciones por Escherichia coli/diagnóstico por imagen , Inflamación/diagnóstico por imagen , Liposomas , Compuestos de Organotecnecio , Oximas , Neumonía por Pneumocystis/diagnóstico por imagen , Radioinmunodetección/métodos , Infecciones Estafilocócicas/diagnóstico por imagen , Animales , Femenino , Inmunoglobulina G , Radioisótopos de Indio , Masculino , Ratas , Ratas Sprague-Dawley , Ratas Wistar , Agregado de Albúmina Marcado con Tecnecio Tc 99m , Exametazima de Tecnecio Tc 99m , Distribución Tisular
10.
J Nucl Med ; 38(10): 1596-600, 1997 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-9379199

RESUMEN

UNLABELLED: Scintigraphic techniques are frequently used for evaluation of inflammatory bowel disease. The radiopharmaceutical of choice is labeled leukocytes. In this study, two new agents, 111In-labeled polyethylene glycol-coated liposomes and 111In-labeled human nonspecific gamma globulin (immunoglobulin G; IgG), were compared with 111In-leukocytes in a rabbit model of colitis. METHODS: In rabbits, acute colitis was induced by colonic instillation of trinitrobenzene sulfonic acid at 25 cm from the anal sphincter. After 24 hr, 15 MBq of the radiopharmaceuticals was injected intravenously in groups of four rabbits. Twenty-four hours after injection, the animals were killed and macroscopic abnormalities were scored in seven consecutive affected colonic segments of 5 cm each (0 = normal, 1 = inflammation, 2 = ulcers). The ex vivo uptake was measured in the normal ascending colon and the affected colonic segments. The colitis index (CI, affected-to-normal colon-uptake ratio) was calculated. RESULTS: Histologically, an acute, patchy, transmural colitis was observed at the site of instillation and the distal colon. The CI of all agents in colitis lesions correlated with the severity of the abnormalities. With increasing severity, the CI for liposomes was 1.86 +/- 0.24, 4.88 +/- 0.42 and 7.42 +/- 0.54 (r2 = 0.68, p < 0.001); for leukocytes 1.77 +/- 0.32, 3.10 +/- 0.58 and 5.54 +/- 0.83 (r2 = 0.31, p < 0.01); for IgG 1.60 +/- 0.29, 2.81 +/- 0.21 and 2.65 +/- 0.21 (r2 = 0.29, p < 0.02). CONCLUSION: Indium-111-labeled-leukocytes, -IgG and -liposomes all show increased uptake in inflamed colonic tissue. Indium-111-liposomes showed the highest CI, which correlates best with the morphological abnormalities. Indium-111-leukocytes and 111In-liposomes are superior to 111In-IgG for this indication.


Asunto(s)
Colitis/diagnóstico por imagen , Radioisótopos de Indio , Radiofármacos , Animales , Colitis/inducido químicamente , Colon/diagnóstico por imagen , Femenino , Humanos , Inmunoglobulina G , Leucocitos , Liposomas/farmacocinética , Polietilenglicoles , Conejos , Cintigrafía , Radiofármacos/farmacocinética , Distribución Tisular , Ácido Trinitrobencenosulfónico
11.
Nucl Med Commun ; 17(9): 742-8, 1996 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-8895901

RESUMEN

Indium-111 (111In) and technetium-99m (99Tcm) Stealth liposomes were compared with 111In- and 99Tcm-labelled white blood cells (WBC) in experimental infection in a rabbit model. Preformed polyethylene glycol-coated liposomes and separated WBC were radiolabelled with either 111In-oxine or 99Tcm-hexamethylpropyleneamine oxime (99TcM-HMPAO). After the intravenous administration of one of the four radiopharmaceuticals to rabbits with focal Staphylococcus aureus infection, scintigraphic images were recorded at various time points post-injection and the biodistribution of the radiopharmaceuticals was determined. At 4 h post-injection, uptake of 111In-WBC in the abscess was significantly higher than that of the three other products. AT later time points, 111In-WBC, 111In-liposome and 99Tcm-liposome uptake in the abscess were similar. In contrast, a 20 h post-injection, uptake of 99Tcm-WBC was significantly lower. The abscess-to-background ratios showed a similar pattern to the absolute abscess uptake: initial high values for 111In-WBC, a more gradual increase over time of the liposome preparations to the level of 111In-WBC and persistently low values for 99Tcm-WBC. Clearance from the blood of both labelled WBC preparations was significantly faster and splenic uptake significantly higher compared with those of the labelled liposomes. In conclusion, given the similar in vivo characteristics of labelled liposomes and labelled WBC, labelled liposomes may be an attractive replacement for labelled WBC, providing a continuously available, high-quality, 99Tcm-labelled radiopharmaceutical that can be prepared easily without any need to handle blood.


Asunto(s)
Radioisótopos de Indio , Liposomas , Infecciones Estafilocócicas/diagnóstico por imagen , Tecnecio , Absceso/diagnóstico por imagen , Animales , Modelos Animales de Enfermedad , Estudios de Evaluación como Asunto , Femenino , Radioisótopos de Indio/farmacocinética , Leucocitos , Enfermedades Musculares/diagnóstico por imagen , Conejos , Cintigrafía , Tecnecio/farmacocinética , Factores de Tiempo
12.
Lab Anim ; 26(4): 269-80, 1992 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-1447905

RESUMEN

Recent reports demonstrate a hypocholesterolaemic effect of daily subcutaneous injections of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors in different rat models of hyperlipidaemia. However, this effect is not seen after oral administration of HMG-CoA reductase inhibitors in rats. We found that oral administration of the HMG-CoA reductase inhibitor Simvastatin also had no effect on plasma cholesterol in severely hyperlipidaemic Nagase analbuminaemic rats (NAR). Simvastatin (an apolar compound dissolved in propylene glycol) was infused continuously for 28 days into the subcutis of control Sprague-Dawley rats (SDR) and NAR using an implanted osmotic pump. All doses which were effective in reducing cholesterol in the NAR (reductions up to approximately 60%), reduced apolipoprotein AI but not apolipoprotein B and caused a severe inflammatory reaction in the dermis. Similar toxicity was observed in the SDR. Subcutaneous administration of the vehicle (propylene glycol) did not cause this reaction and did not affect plasma lipids. Administration of Lovastatin in osmotic pumps resulted in a similar inflammatory reaction. Incorporation of Simvastatin into liposomes did not diminish the toxic effect. On the other hand, infusion of Pravastatin (a polar HMG-CoA reductase inhibitor dissolved in isotonic saline) caused no changes in the dermis and had no effect on plasma lipids in NAR or SDR. Liver microsomes prepared from the Pravastatin-treated rats demonstrated a 3- to 4-fold increase in HMG-CoA reductase activity as compared to untreated rats, confirming uptake of the drug. We conclude that continuous subcutaneous administration of the HMG-CoA reductase inhibitors Simvastatin, Lovastatin and Pravastatin for 28 days may not reduce plasma cholesterol in rats by a mechanism which is related to inhibition of HMG-CoA reductase activity in the liver. The decrease of plasma cholesterol effected by subcutaneous infusion of Simvastatin or Lovastatin in NAR coincides with, and may be related to inflammatory changes caused by administering these compounds into the dermis.


Asunto(s)
Anticolesterolemiantes/administración & dosificación , Inhibidores de Hidroximetilglutaril-CoA Reductasas , Lovastatina/análogos & derivados , Lovastatina/administración & dosificación , Pravastatina/administración & dosificación , Administración Oral , Animales , Anticolesterolemiantes/toxicidad , Femenino , Hipercolesterolemia/sangre , Hipercolesterolemia/tratamiento farmacológico , Hipercolesterolemia/patología , Bombas de Infusión Implantables , Inyecciones Subcutáneas , Lovastatina/toxicidad , Pravastatina/toxicidad , Ratas , Ratas Mutantes , Ratas Sprague-Dawley , Simvastatina
13.
Equine Vet J ; 44(2): 196-202, 2012 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-21696436

RESUMEN

REASONS FOR PERFORMING STUDY: Liposomes are phospholipid nanoparticles that extravasate at sites of increased vascular permeability. They have potential in equine medicine for targeted drug delivery and diagnostic imaging of infectious, inflammatory and neoplastic lesions. OBJECTIVES: This study evaluates the safety and biodistribution of i.v. polyethyleneglycol(PEG) liposomes in normal horses. METHODS: PEG-liposomes were prepared by the film hydration method and labelled using (99m) Tc-hexamethyl-propylene-amine-oxime. A single dose of 0.24 µmol/kg bwt (99m) Tc-PEG-liposomes and 2.4 µmol/kg bwt unlabelled PEG-liposomes was administered to 10 conscious horses via i.v. infusion at a rate of 6 µmol/min for the first 15 min and 60 µmol/min thereafter. Clinical parameters, haematology, plasma biochemistry and serum complement activity were monitored serially. Scintigraphic imaging was performed at 1, 12 and 21 h post infusion (PI). Six horses were subjected to euthanasia at 24 h PI. The percentage injected dose per kilogram of tissue was calculated for multiple organs. Results were analysed using repeated measures ANOVA. RESULTS: Horses did not demonstrate adverse reactions during or after liposome infusion. There was a significant elevation in heart rate and respiratory rate at 20 and 25 min PI. No significant complement consumption was detected, although a trend for decreased total haemolytic complement values at 20 min PI was present. Scintigraphic studies revealed a prolonged vascular phase that lasted to 21 h PI, with a reproducible pattern of organ distribution. Biodistribution studies revealed the highest concentrations of radiopharmaceutical within the lung, kidney, liver and spleen. CONCLUSIONS: Intravenous liposome administration appears to be safe in horses. When administered in combination with PEG-liposomes, (99m) Tc-PEG-liposomes have long circulating characteristics and a reproducible pattern of organ distribution in horses. POTENTIAL RELEVANCE: Radiolabelled liposomes may be useful for detecting infection, inflammation and neoplasia in the horse. Liposomes have significant potential for targeted drug delivery in the horse. This study establishes the scintigraphic findings and tissue distribution of 99mTc-PEG-liposomes after i.v. administration in healthy horses.


Asunto(s)
Caballos/metabolismo , Liposomas/farmacocinética , Polietilenglicoles/farmacocinética , Radiofármacos/farmacocinética , Exametazima de Tecnecio Tc 99m/farmacocinética , Animales , Femenino , Inyecciones Intravenosas , Liposomas/administración & dosificación , Liposomas/efectos adversos , Liposomas/química , Masculino , Polietilenglicoles/administración & dosificación , Polietilenglicoles/efectos adversos , Polietilenglicoles/química , Radiofármacos/administración & dosificación , Radiofármacos/efectos adversos , Radiofármacos/química , Exametazima de Tecnecio Tc 99m/administración & dosificación , Exametazima de Tecnecio Tc 99m/efectos adversos , Exametazima de Tecnecio Tc 99m/química , Distribución Tisular
14.
Vaccine ; 13(1): 45-53, 1995 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-7762277

RESUMEN

The objective of this study was the comparison of the quantity and quality of the humoral immune response against inactivated whole human immunodeficiency virus type 2. Twenty-four different adjuvants were tested. The toxicity of these preparations was determined. Due to the large number of experiments, the animal model used was the NMRI mouse. Sera were assayed for the presence of antibodies by ELISA and Western blot. The toxicity of the adjuvants was determined by observing lethal side-effects occurring within two days after injection of the immunogenic preparations. The results show that polymethylmethacrylate nanoprticles were the best overall adjuvant, inducing the highest titres of antibodies compared with other adjuvants as well as producing a significantly better immune response against a number of individual HIV-2 antigens without any observable toxic side-effects. However, this adjuvant did not induce antibodies against the outer envelope protein gp120, whereas such antibodies were induced by aluminium compounds, Freund's complete and incomplete adjuvants, and fumed silica (Aerosil). This result suggests the necessity of using vaccines with two or more different adjuvants in order to induce the required immune response against physically different antigens.


Asunto(s)
Vacunas contra el SIDA/inmunología , Adyuvantes Inmunológicos/farmacología , Reacciones Antígeno-Anticuerpo , Antígenos Virales/sangre , VIH-2 , Adyuvantes Inmunológicos/efectos adversos , Animales , Formación de Anticuerpos , Western Blotting , Femenino , Ratones , Ratones Endogámicos , Volumetría , Vacunas de Productos Inactivados/inmunología
15.
Ann Rheum Dis ; 56(6): 369-73, 1997 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-9227166

RESUMEN

OBJECTIVES: Liposomes sterically stabilised with polyethylene glycol (PEG) labelled with technetium-99m were tested for their ability to image adjuvant arthritis in a rat model. METHODS: Adjuvant arthritis was induced in the ankle joint of the left hind foot by injection of Mycobacterium butyricum in Freund's incomplete adjuvant in the foot pad. Seven days later animals received the following radiopharmaceuticals labelled with 99mTc (a) non-PEG-liposomes, (b) PEG-liposomes or (c) non-specific human polyclonal IgG. For each of the radiopharmaceuticals the in vivo distribution of the radiolabel was monitored both scintigraphically as well as by counting the dissected tissues at two, eight, and 24 hours after injection. RESULTS: The pharmacokinetics of the radiopharmaceuticals differed considerably (halflife in the blood: PEG-liposomes (18 hours) > 99mTc-IgG (3 hours) > non-PEG liposomes (1 hour)). The inflamed focus was visualised with each of the agents. The uptake of each of the radiopharmaceuticals in the inflamed ankle region correlated with their residence time in the blood (inflamed joint uptake: PEG liposomes (1.15% injected dose (ID)/ g) > 99mTc-IgG (0.35% ID/g) > non-PEG-liposomes (0.05% ID/g)). Quantitative analysis of the images showed that the inflamed ankle to background ratio was highest with the PEG-liposomes (7.5 at 24 hours after injection), while with the other two agents this ratio did not exceed 4. CONCLUSION: This study shows that 99mTc-labelled PEG-liposomes may be an excellent agent to visualise arthritis. Increased label uptake in the inflamed joint and increased target to background ratios can be obtained with PEG-liposomes because of their long circulating properties. In addition to their use as vehicles for scintigraphic imaging of arthritis PEG-liposomes might also be used for the site specific delivery of antirheumatic drugs.


Asunto(s)
Artritis Experimental/diagnóstico por imagen , Liposomas , Compuestos de Organotecnecio , Oximas , Animales , Articulación del Tobillo/diagnóstico por imagen , Articulación del Tobillo/metabolismo , Artritis Experimental/metabolismo , Inmunoglobulinas/metabolismo , Liposomas/química , Liposomas/farmacocinética , Masculino , Compuestos de Organotecnecio/farmacocinética , Oximas/farmacocinética , Polietilenglicoles , Cintigrafía , Ratas , Ratas Wistar , Tecnecio/metabolismo , Tecnecio/farmacocinética , Exametazima de Tecnecio Tc 99m , Distribución Tisular
16.
Biochemistry ; 32(17): 4641-9, 1993 May 04.
Artículo en Inglés | MEDLINE | ID: mdl-8485142

RESUMEN

The role of electrostatics in the adsorption process of proteins to preformed negatively-charged (phosphatidylcholine/phosphatidylglycerol) and neutral (phosphatidylcholine) liposomes was studied. The interaction was monitored at low ionic strength for a set of model proteins as a function of pH. The adsorption behavior of trypsin inhibitor (pI = 4.6), myoglobin (pI = 7.4), ribonuclease (pI = 9.6), and lysozyme (pI = 10.7) with preformed liposomes was investigated, along with changes in the electrophoretic mobility of liposomes through the adsorption of charged proteins. Mean protein charge was determined by acid/base titration. Significant adsorption of the proteins to negatively-charged liposomes was only found at pH values where the number of positive charge moieties exceeds the number of negative charge moieties on the protein by at least three charge units. Negligible adsorption to liposomes composed of zwitterionic lipids was observed in the pH range tested (4-9). The absolute value of the electrophoretic mobilities of negatively-charged, empty liposomes decreased after adsorption of positively-charged proteins. With increasing protein to phospholipid ratio, the drop in the electrophoretic mobility leveled off and reached a plateau; protein adsorption profiles showed a similar shape. Analysis of the data demonstrated that neutralization of the liposome charge due to the adsorption of the positively-charged proteins is the controlling factor in their adsorption. The plateau level reached depended on the type of protein and the pH of the incubation medium. This pH dependency could be ascribed to the mean positive charge of the protein.(ABSTRACT TRUNCATED AT 250 WORDS)


Asunto(s)
Electroforesis , Globulinas/química , Liposomas/química , Adsorción , Electroquímica , Concentración de Iones de Hidrógeno , Punto Isoeléctrico , Muramidasa/química , Mioglobina/química , Concentración Osmolar , Ribonucleasas/química , Solubilidad , Inhibidores de Tripsina/química , Agua
17.
Biochemistry ; 36(41): 12583-91, 1997 Oct 14.
Artículo en Inglés | MEDLINE | ID: mdl-9376364

RESUMEN

An antibody-peptide model system was used to study the binding characteristics between a bactericidal antibody (MN12H2) and the P1. 16 epitope of class 1 outer membrane protein PorA of Neisseria meningitidis by means of a thermodynamic approach. A series of four linear peptides and three "head-to-tail" cyclic peptides (with ring sizes of 9, 15 and 17 amino acids) were synthesized and evaluated as ligands. The peptides contain a fluorescein label and the core determinant amino acid sequence TKDTNNN (residues 180-186) of the PorA P1.16 epitope of meningococcal strain H44/76. Thermodynamic data of the binding of the peptide homologs of the epitope by MN12H2 were assessed by measuring affinity constants (Ka) over a temperature range of 4-55 degrees C, using fluorescence spectroscopy. Curvilinear plots of ln Ka versus T (K) revealed strong temperature dependencies of enthalpy (DeltaH) and entropy (DeltaS). The Gibbs free energy change (DeltaG) was only weakly temperature dependent. The large negative enthalpy value indicated the importance of polar interactions in the binding of both linear and cyclic peptides by MN12H2. Sturtevant's analysis of the thermodynamic parameters showed large unfavorable vibrational contributions to the binding for all linear peptides [Sturtevant, J. M. (1977) Proc. Natl. Acad. Sci.U.S.A. 74, 2236-2240]. The large hydrophobic contribution compensating these vibrational modes was partially attributed to aspecific interaction of the fluorescein label with the antibody. Binding of MN12H2 to conformationally restricted epitope sequences was characterized by a dramatic reduction in the size of unfavorable vibrational components of the thermodynamic parameters. Substitution of individual charged amino acids of the P1.16 epitope sequence revealed that aspartate-182 was essential for the binding. The pH profile observed for the MN12H2-peptide complexes with a midpoint pH of approximately 8.5 suggests a positively charged histidine from the antibody binding site to be involved in a charge interaction with Asp-182. These findings are consistent with the results from the crystal structure of the Fab fragment of MN12H2 in complex with a linear fluorescein-conjugated peptide homolog of the P1.16 epitope [van den Elsen et al. (1997) Proteins (in press)], thereby identifying the basis of an increased incidence of endemic disease in England and Wales since 1981 caused by a mutant meningococcal strain.


Asunto(s)
Anticuerpos Antibacterianos/química , Antígenos Bacterianos/química , Neisseria meningitidis/inmunología , Porinas/química , Animales , Anticuerpos Antibacterianos/inmunología , Anticuerpos Antibacterianos/metabolismo , Anticuerpos Monoclonales/química , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales/metabolismo , Reacciones Antígeno-Anticuerpo , Antígenos Bacterianos/inmunología , Antígenos Bacterianos/metabolismo , Ratones , Porinas/inmunología , Porinas/metabolismo , Termodinámica
18.
J Immunol ; 160(9): 4449-56, 1998 May 01.
Artículo en Inglés | MEDLINE | ID: mdl-9574550

RESUMEN

Synthetic peptide-based vaccines have been shown to induce potent protective and therapeutic T cell-mediated immunity in preclinical animal models and are now being evaluated in clinical phase I/II studies for their efficacy against tumors or infectious diseases. However, such vaccines might also specifically tolerize T cells causing enhanced tumor outgrowth, as shown by vaccination with two CTL epitopes derived from the adenovirus type 5 early region 1 (Ad5E1) oncogenes. We now report that modification of the Ad5E1 peptide vaccine either through incorporation of the peptides into liposomes or by ligation of the peptides to lipid tails, another vaccine formulation being tested in the clinic, fails to convert immunosuppression into effective antitumor vaccination. Inclusion of a helper T cell epitope into the vaccine likewise induces enhanced tumor outgrowth and thus does not diminish the capacity of the peptides to tolerize Ad5E1-specific CTL. In contrast, the Ad5E1-derived peptides evoke a strong tumor-protective CTL response when presented on dendritic cells (DC), indicating that the in vivo CTL-tolerizing potential of these peptides is converted to specific immunostimulation when presented on DC. These findings have important implications for the development of peptide-based immune intervention strategies and emphasize the superior nature of Ag-pulsed DC over other peptide-based vaccination protocols as well as the crucial importance of the mode of peptide-Ag delivery in setting the balance between T cell stimulation and tolerization.


Asunto(s)
Presentación de Antígeno , Citotoxicidad Inmunológica , Células Dendríticas/inmunología , Neoplasias Experimentales/inmunología , Péptidos/inmunología , Receptores de Antígenos de Linfocitos T/inmunología , Linfocitos T Citotóxicos/inmunología , Vacunación , Secuencia de Aminoácidos , Animales , Portadores de Fármacos , Tolerancia Inmunológica , Liposomas , Ratones , Ratones Endogámicos C57BL , Datos de Secuencia Molecular , Neoplasias Experimentales/patología , Péptidos/administración & dosificación , Receptores de Antígenos de Linfocitos T/administración & dosificación , Células Tumorales Cultivadas , Escape del Tumor/inmunología , Vacunación/efectos adversos
19.
Arthritis Rheum ; 43(9): 1951-9, 2000 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-11014344

RESUMEN

OBJECTIVE: To assess whether intraarticular (IA) administration of clodronate liposomes results in local macrophage depletion in patients with rheumatoid arthritis (RA). Primary goals were to address both the immunohistologic and potential toxic effects of this approach. Moreover, the correlation between immunohistologic findings and clinical assessments of disease activity and cartilage damage were assessed. METHODS: An open study was conducted in consecutive RA patients who were scheduled for knee joint replacement in our department. Synovial biopsy tissue was obtained from the knee joint at 2 weeks before and at the time of surgery. This protocol was controlled for safety and immunohistologic concordance in 6 patients. One week before surgery, 10 patients received a single IA dose of clodronate liposomes. Staining of synovial tissue for cell markers (CD68, CD14, CD3, CD38) and adhesion molecules (vascular cell adhesion molecule 1 [VCAM-1], intercellular adhesion molecule 1 [ICAM-1]) was assessed by 2 blinded observers. Local and systemic parameters of disease activity were measured before each intervention. Cartilage damage was scored using standard radiologic techniques at baseline and during surgery. RESULTS: A single IA dose of clodronate liposomes significantly reduced the number of CD68-positive cells (P = 0.005) and the expression of ICAM-1 and VCAM-1 in the synovial lining (P = 0.013 and P = 0.039, respectively). The intervention did not affect fibroblast-like synoviocytes, T cells, or plasma cells. No immunohistologic changes were observed in the control group. The procedure was well tolerated. The levels of ICAM-1 and VCAM-1 in the sublining layers correlated with the extent of macroscopic synovitis (P < 0.0005 and P < 0.005, respectively). The expression of ICAM-1 and CD14 in the sublining correlated with the levels of C-reactive protein (P < 0.0005 and P < 0.01, respectively). Cartilage destruction was correlated only with the expression of CD68 in the sublining (P = 0.02). CONCLUSION: A single IA administration of clodronate liposomes leads to macrophage depletion and decreased expression of adhesion molecules in the synovial lining in patients with longstanding RA. The procedure is well tolerated, and its therapeutic potential is currently under investigation. The expression of adhesion molecules in the sublining layers reflects ongoing inflammation.


Asunto(s)
Artritis Reumatoide/patología , Ácido Clodrónico/farmacología , Macrófagos/efectos de los fármacos , Macrófagos/fisiología , Adulto , Anciano , Biomarcadores/análisis , Biopsia , Ácido Clodrónico/administración & dosificación , Femenino , Humanos , Inmunohistoquímica , Liposomas , Masculino , Persona de Mediana Edad , Membrana Sinovial/patología , Factores de Tiempo
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