Labelling of liposomes with intercalating perylene fluorescent dyes

The high fluorescent potential and the exceptional photostability of lipophilic derivatives of perylene-3,4:9,10-bis(dicarboximides) are utilized for the fluorescence-labelling of liposomes. The preparation of the liposomes is affected by supersonic starting from a lipid mixture consisting of the matrix lipids soy lecithin, cholesterol, -tocopherol and the perylene dyes. From a multitude of perylene derivatives investigated only those are optimally incorporated inot the bilayer membrane of unilamellar liposomes which are substituted at both nitrogen atoms by one or two linear hydrocarbon groups. In order to attain an optimal fluorescent quantum yield, about 200 to 300 dye molecules can be incorporated per liposome. The liposomes thus obtained have a diameter of about 70 to 80 nm, are homogeneous and may be stored for more than seven months. Neither the fluorescent properties nor the stability of these liposomes are influenced by the additional incorporation of various ara C-derivatives and lipophilic anchor groups which subsequently enable the coupling of antibodies to the liposomes. As the water-insoluble perylene dyes are incorporated into the bilayer membrane, the aqueous inner volume of the liposomes remains available for a fruther utilization

Liposomes and Immunoliposomes as Carriers for Cytostatic Drugs, Magnetic Resonance Contrast Agents, and Fluorescent Chelates

Biological and medical applications of liposomes as carriers for cytostatic drugs, magnetic-resonance contrast agents, and fluorescent markers are presented. The cytostatic effects of liposomal preparations of lipophilic derivatives of cytosine arabinoside (ara-C), namely N4-oleyl-ara-C (NOAC) and N4-alkyl-ara-C are demonstrated in the mouse Ll210 tumor model. Liposomal drugs were more effective by factors of 2–10 as compared to ara-C administered in aqueous preparations. Synergistic effects of the combination of ara-C and mitoxantrone could be shown, again with significantly increased effects, when the drugs were administered in liposomes. A practical detergent-dialysis method for the preparation of large volumes of sterile liposomes for clinical use is presented. Clinical phase I/II studies of Iiposoma I NOAC and mitoxantrone are currently in progress. Methods of preparation of immunoliposomes, i.e. the coupling of antibodies or antibody subunits to liposome membranes, are discussed' and results of specific immunoliposome – cell binding are presented. Possibilities of increasing the blood circulation times of liposomes by incorporation of lipophilic derivatives of poly(ethylene glycol) into the liposome membranes are shown. The successful application of liposomes as carriers of paramagnetic metal complexes as contrast agents in magnetic-resonance imaging for liver and spleen is documented. Finally, the concept of liposomes as carriers for fluorescent Europium chelates as markers for time-resolved flow cytometric applications are presented

Pulmonary thrombembolism as cause of death on unenhanced postmortem 3T MRI

Objectives: To investigate unenhanced postmortem 3-T MR imaging (pmMRI) for the detection of pulmonary thrombembolism (PTE) as cause of death. Methods: In eight forensic cases dying from a possible cardiac cause but with homogeneous myocardium at cardiac pmMRI, additional T2w imaging of the pulmonary artery was performed before forensic autopsy. Imaging was carried out on a 3-T MR system in the axial and main pulmonary artery adapted oblique orientation in situ. In three cases axial T2w pmMRI of the lower legs was added. Validation of imaging findings was performed during forensic autopsy. Results: All eight cases showed homogeneous material of intermediate signal intensity within the main pulmonary artery and/or pulmonary artery branches. Autopsy confirmed the MR findings as pulmonary artery thrombembolism. At lower leg imaging unilateral dilated veins and subcutaneous oedema with or without homogeneous material of intermediate signal intensity within the popliteal vein were found. Conclusions: Unenhanced pmMRI demonstrates pulmonary thrombembolism in situ. PmMR may serve as an alternative to clinical autopsy, especially when consent cannot be obtained. Key Points: • Postmortem MRI (pmMRI) provides an alternative to clinical autopsy • Fatal pulmonary thrombembolism (PTE) can now be diagnosed using postmortem MRI (pmMRI). • Special attention has to be drawn to the differentiation of postmortem clot

Comparative studies of the preparation of immunoliposomes with the use of two bifunctional coupling agents and investigation of in vitro immunoliposome-target cell binding by cytofluorometry and electron microscopy

The two coupling agents SPDP (N-succinimidyl-3-(2-pyridyldithio)propionate) and SATA (N-succinimidyl-S-acetylthioacetate) were compared in their efficiency and feasibility to couple monoclonal antibodies (Abs) via thioether linkage to liposomes functionalized by various lipophilic maleimide compounds like Full-size image methyl ester (MP-PL), N-(3-maleimidopropionyl)phosphatidylethanolamide (MP-PE), Full-size image methyl ester (EMC-PL), and N-(6-maleimidocaproyl)phosphatidylethanolamine (EMC-PE). The composition of the liposomes was soy phosphatidylcholine (SPC), cholesterol, maleimide compounds and -tocopherol (1:0.2:0.02:0.01, mol parts), plus N4-oleylcytosine arabinoside (NOAC) as cytostatic prodrug (0.2 mol parts) and a new, lipophilic and highly fluorescent dye N,N′-bis(1-hexylheptyl)-3,4:9,10-perylenebis(dicarboximide) (BHPD, 0.006 mol parts). From the maleimide derivatives MP-PL was the most effective in terms of preservation of the coupling activity in dependence of liposome storage. The coupling of the monoclonal A B8-24.3 (mouse IgG2b, MHC class I, anti H-2kb) and IB16-6 (rat IgG2a, anti B16 mouse melanoma) to the drug carrying liposomes was more effective and easier to accomplish with SATA as compared to SPDP. Coupling rates of 60–65% were obtained with SATA at molar ratios of 12 SATA:1 Ab:40 maleimide spacer groups on the surface of one liposome. The highest coupling rates with SPDP were obtained at the ratio of 24 SPDP:1 Ab:40 liposomal maleimide groups, with an Ab binding efficiency of only 20–25%. The optimal in vitro binding conditions to specific target cells (EL4 for B8-24.3-liposomes and B16-F10 for IB16-6-liposomes) were determined by cytofluorometric measurement of the liposomal BHPD fluorescence with SATA linked Abs. Optimal immunoliposome binding to specific epitopes on the target cells was achieved with 1–2 Ab molecules coupled to one liposome, with immunoliposome concentrations of 20–130 nM and with a small incubation volume of 0.3–0.4 ml. The specificity of the binding of B8-24.3-liposomes to EL4 target cells was visualized by scanning electron microscopy. Antibody mediated endocytic uptake of immunoliposomes could be demonstrated by transmission electron microscopy

Multidimensional Participation in Hybrid Wireless Communities

Wireless communities have been long considered an interesting approach to provide mobile Internet, but the key issue is whether they are able to attract and retain a critical mass of active members. It is therefore crucial to understand what motivates and dissuades people from joining and participating in them, especially with the development of mainstream 3G technologies, in order to evaluate their potential development. This paper analyzes motivations and barriers influencing participation in a large wireless community – Fon – based on a survey of 268 members. Two distinct forms of participation driven by different motivations emerge: a ‘participation by sharing’ driven by idealistic motivation and a ‘social participation’ driven by social motives and technical interest. Utilitarian motivations do not play a major role for active participation despite being crucial in attracting members to the community. Accordingly, the way hybrid wireless communities are currently designed (hardly offering occasions for a social usage experience, experimentation and with decreasing utilitarian benefits due the development of 3G technologies) is casting serious doubts about a possible potential development above the status of a niche complement to the dominant cellular technologies

Prenatal and postnatal maternal contributions in the infection model of schizophrenia

Epidemiological studies have indicated that the risk of schizophrenia is enhanced by prenatal maternal infection with viral or bacterial pathogens. Recent experimentation in rodents has yielded additional support for a causal relationship between prenatal immune challenge and the emergence of psychosis-related abnormalities in brain and behaviour in later life. However, little is known about the putative roles of maternal postnatal factors in triggering and modulating the emergence of psychopathology following prenatal immunological stimulation. Here, we aimed to dissect the relative contributions of prenatal inflammatory events and postnatal maternal factors in precipitating juvenile and adult psychopathology in the resulting offspring with a cross-fostering design. Pregnant mice were exposed to the viral mimic, polyriboinosinic-polyribocytidilic acid (PolyI:C; at 5mg/kg, intravenously), or vehicle treatment on gestation day 9, and offspring born to PolyI:C- and vehicle-treated dams were then simultaneously cross-fostered to surrogate rearing mothers, which had either experienced inflammatory or vehicle treatment during pregnancy. Prenatal PolyI:C administration did not affect the expression of latent inhibition (LI) at a juvenile stage of development, but led to the post-pubertal emergence of LI disruption in both aversive classical and instrumental conditioning regardless of the postnatal rearing condition. In addition, deficits in conditioning as such led to a pre- and post-pubertal loss of LI in prenatal control animals that were adopted by PolyI:C-treated surrogate mothers. Our findings thus indicate that the adoption of prenatally immune-challenged neonates by control surrogate mothers does not possess any protective effects against the subsequent emergence of psychopathology in adulthood. At the same time, however, the present study highlights for the first time that the adoption of prenatal control animals by immune-challenged rearing mothers is sufficient to precipitate learning disabilities in the juvenile and adult offsprin

Neonatological and pulmonological management of bilateral pulmonary sequestration in a neonate

Background: Bronchopulmonary sequestration is a lung malformation characterized by nonfunctioning lung tissue without primary communication with the tracheobronchial tree. Intrauterine complications such as mediastinal shift, pleural effusion or fetal hydrothorax can be present. We present the case of a newborn with bilateral intralobar pulmonary sequestration. Methods: Prenatal ultrasonography in a primigravida at 20 weeks of gestation revealed echogenic masses in the right fetal hemithorax with mediastinal shift towards the left side. Serial ultrasound confirmed persistence of the lesion with otherwise appropriate fetal development. Delivery was uneventful and physical examination revealed an isolated intermittent tachypnea. Chest CT scan and CT angiography showed a bilateral intrathoracic lesion with arterial supply from the aorta. Baby lung function testing suggested possible multiple functional compartments. Results: Right and left thoracotomy was performed at the age of 7 months. A bilateral intralobar sequestration with vascularisation from the aorta was resected. Pathological and histological examination of the resected tissue confirmed the surgical diagnosis. At the age of 24 months, the child was doing well without pulmonary complications. Conclusions: Bilateral pulmonary sequestration requires intensive prenatal and postnatal surveillance. Though given the fact of a bilateral pulmonary sequestration, postnatal outcome showed similar favourable characteristics to an unilateral presentation. Baby lung function testing could provide additional information for optimal postnatal management and timing of surgical interventio

Incidental occult gunshot wound detected by postmortem computed tomography

The body of a 59year old woman underwent postmortem computed tomography (PMCT) examination prior to forensic autopsy, using a 256 slice multidetector row computed tomography scanner. A large left tension pneumothorax detected on the PMCT was considered to be a likely cause of death and this was confirmed at autopsy. In addition there was an unsuspected PMCT finding of a probable gunshot injury traversing the right orbit, facial bones and frontal sinus. The autopsy technique was adjusted accordingly and PMCT findings confirmed. PMCT in this case was not only diagnostic of cause of death, but also revealed retained projectile fragments of an old gunshot wound to the face. Without prior imaging such findings would have been undetected at autopsy. This case further underscores the contribution of routine PMCT examination to forensic autopsy practic