Electrical resistance and macromolecular permeability of brain endothelial monolayer cultures

Electrophysiological measurements were made on endothelial cells initially isolated as individual clones from bovine brain microvessels, and then grown as monolayers on a permeable support of glutaraldehyde-treated collagen gel. When transendothelial cell resistance (R) of the clones was measured, there was a range of values from a low of 157.4 +/- 4.5 omega.cm2 (n = 6) to a high of 783.2 +/- 7.0 omega.cm2 (n = 34). With the high-resistance cells, there was also a small potential difference of -0.46 +/- 0.03 mV luminal-side negative (n = 34). In comparison, endothelial cells from bovine aortas and rat epididymal fat pads cultured on the collagen gels had transendothelial R values of 13.5 +/- 0.2 (n = 62) and 0.45 +/- 0.03 (n = 10) omega.cm2, respectively. Exposure of the high-resistance brain endothelial cell monolayers to a Ca2+-free medium for 10 min decreased the R to 75% of the control values. Addition of Ca2+ back to the medium caused a return of the transendothelial R to control values within 1 h. Endothelial cells were also grown to confluency on microcarrier beads for permeability measurements to Evans blue dye-bovine serum albumin. Microcarriers with no cells (control) and microcarriers with bovine and epididymal endothelial cell monolayers showed no difference in the amount of adsorbed dye. Microcarriers with brain endothelial monolayers excluded up to 80% of the dye. This mammalian brain endothelial culture system will be a useful model for studies of the electrophysiological and permeability properties of the blood-brain barrier.     

Force recruitment during electrical nerve stimulation with multipolar intrafascicular electrodes

Experimentally, during electrical nerve stimulation, the influence is examined of two intrafascicular anodes on the force recruitment with one intrafascicular cathode. It is found that the anodes suppress recruitment and that this effect is more pronounced when the distance of the anodes to the cathode is decreased, or when the anodal currents are increased. The measured recruitment curve patterns can be qualitatively explained by a nerve stimulation model that calculates theoretical recruitment curves for intrafascicular multi-electrode configurations. Discrepancies between the experimental and the theoretical recruitment curves are seen, but these can be understood by taking into account a non-uniform fibre distribution.     

High survial rate of hamsters given intratracheal instillations of benzo[a]pyrene and ferric oxide and kept on a high {beta}-carotene diet

The study described in this paper was primarily conducted toidentify the cell types involved in the formation, progressionand regression of metaplastic changes in the respiratory tractepithelium of hamsters after intratracheal intubations withbenzo[a]pyrene Furthermore, the role of vitamin A and ß;-carotenein these processes was studied. In the course of the study aremarkable effect of dietary ß;-carotene on survivalof hamsters became a subject of investigation. Hamsters werefed diets with various levels of vitamin A or ß-caroteneand were treated intratracheally with a suspension of benzo[a]pyrenewith ferric oxide in saline. The tumour response of the respiratorytract was very low (2.8%) and hyper- and metaplasia of respiratoryepithelium were virtually absent. However, an interesting observationwas an exceptionally low mortality of only 2% after 69 weeksin the group of hamsters fed a high ß-carotene diet(1% w/w), whereas in the other groups mortality after 69 weeksamounted to 25%. Although the exact cause of death of most ofthe hamsters could not be established, a 40% reduction of lipidperoxidation in the livers was found in the high ß-carotenegroup. Moreover, In this group the degree and incidence of nephroslsand of focal mineralization of kidneys and heart were lowerthan in the other groups. These favourable effects of the highß-carotene diet may have contributed to the unusuallyhigh survival rate in hamsters fed this diet. Further studiesare planned to verify and study this observation.     

Use of commercially available cell culture inserts for primary culture and electrophysiologic studies of guinea pig gastric mucous epithelial cells

Summary The adherence, growth, and electrophysiologic properties of guinea pig gastric mucous epithelial cells were investigated using porous membrane filters. We also tested three commercially available Ussing-type chambers that were designed to be used with the various porous membrane supports. Overall, the 0.45-µm Falcon-Cyclopore porous membrane was found to be very favorable for the consistent attachment and growth of our cells. This same filter also gave good results in the detection of periodic acid Schiff-positive mucous glycoprotein and Nile red neutral lipid fluorescence in the gastric mucous cells. Our cells grew poorly on collagen-coated Costar-Snapwells and Millipore Millicell-CM porous filters. For measurement of transepithelial potential difference resistance, and short-circuit current, the Costar-Snapwell with the Costar-Snapwell Diffusion-chamber system was superior in design and operation when compared to the Costar Transwell-COL, Falcon-Cyclopore, or Anotec-Anocell porous inserts used with conventional Ussing-chambers. The gastric mucous cells grew best on ICN-Cellagen membranes, but these filters routinely detached from their plastic holder and therefore could not be used for Ussing-chamber studies. The large 24.5-mm, 0.40-µm pore size Costar-Transwell-COL and the 24.1-mm, 0.45-µm Falcon-Cyclopore membranes gave good results when used in a modified horizontal-chamber for microelectrode analysis of membrane potentials and resistances of the gastric mucous cell monolayers.     

Lymphocyte Populations and Cellular Immune Reactions in Juvenile Rheumatoid Arthritis

Ten patients with juvenile rheumatoid arthritis and age- and sex-matched healthy controls were investigated in pairs. The patients were found to have both normal proportions and normal absolute numbers of T lymphocytes, B lymphocytes, and the Fc-receptor-bearing lymphoid cells in peripheral blood. No abnormality of mitogen-induced lymphocyte transformation was observed. Lymphocyte-mediated cytotoxicity induced by phytohemagglutinin (PHA) or anti-target cell antibodies was also found to be normal. As in an earlier study, impaired delayed hyper-sensitivity by skin testing was observed in the patient group, thus indicating a dissociation between in vivo and in vitro parameters of lympboid cell function.     

Treatment of motoneuron degeneration by intracerebroventricular delivery of VEGF in a rat model of ALS

Neurotrophin treatment has so far failed to prolong the survival of individuals affected with amyotrophic lateral sclerosis (ALS), an incurable motoneuron degenerative disorder. Here we show that intracerebroventricular (i.c.v.) delivery of recombinant vascular endothelial growth factor (Vegf) in a SOD1(G93A) rat model of ALS delays onset of paralysis by 17 d, improves motor performance and prolongs survival by 22 d, representing the largest effects in animal models of ALS achieved by protein delivery. By protecting cervical motoneurons, i.c.v. delivery of Vegf is particularly effective in rats with the most severe form of ALS with forelimb onset. Vegf has direct neuroprotective effects on motoneurons in vivo, because neuronal expression of a transgene expressing the Vegf receptor prolongs the survival of SOD1(G93A) mice. On i.c.v. delivery, Vegf is anterogradely transported and preserves neuromuscular junctions in SOD1(G93A) rats. Our findings in preclinical rodent models of ALS may have implications for treatment of neurodegenerative disease in general.     

Chest radiography in general practice: indications, diagnostic yield and consequences for patient management

BACKGROUND: Chest radiography (CXR) is frequently performed in Western societies. There is insufficient knowledge of its diagnostic value in terms of changes in patient management decisions in primary care. AIM: To assess the influence of CXR on patient management in general practice. DESIGN OF STUDY: Prospective cohort study. SETTING: Seventy-eight GPs and three general hospitals in the Netherlands. METHOD: Patients (n = 792) aged > or =18 years referred by their GPs for CXR were included. The main outcome was change in patient management assessed by means of questionnaires filled in by GPs before and after CXR. RESULTS: Mean age of the patients was 57.3+/-16.2 years and 53% were male. Clinically relevant abnormalities were found in 24% of the CXRs. Patient management changed in 60% of the patients following CXR. Main changes included: fewer referrals to a medical specialist (from 26 to 12%); reduction in initiation or change in therapy (from 24 to 15%); and more frequent reassurance (from 25 to 46%). However, this reassurance was not perceived as such in a quarter of these patients. A change in patient management occurred significantly more frequently in patients with complaints of cough (67%), those who exhibited abnormalities during physical examination (69%), or those with a suspected diagnosis of pneumonia (68%). CONCLUSION: Patient management by the GP changed in 60% of patients following CXR. CXR substantially reduced the number of referrals and initiation or change in therapy, and more patients were reassured by their GP. Thus, CXR is an important diagnostic tool for GPs and seems a cost-effective diagnostic test.     

Inhibition of Antibody-Dependent Human Lymphocyte-Mediated Cytotoxicity by Immunoglobulin Classes, IgG Subclasses, and IgG Fragments

The cytotoxicity of human peripheral blood lymphocytes against chicken erythrocytes sensitized by rabbit antibodies was inhibited by human immunoglobulin and immunoglobulin fragments. Myeloma proteins isolated in dimeric state or aggregated by heat treatment inhibited better than the corresponding monomeric proteins. Strong inhibition was observed with IgG1 and IgG3, and with IgG₂ after aggregation, while IgG₄ inhibited very little. No inhibition was found with IgM, IgA. IgD and IgE. The F(ab')₂. and Fab fragments of IgG inhibited poorly or not at all. While- considerable inhibition was observed with the Fc fragment, the pFc' fragment, which roughly corresponds to the C-terminal half of the Fc portion, showed little inhibitory capacity. A fragment isolated from IgG3, containing an extension of the N-terminal part of Fc (the Fch fragment), was an even better inhibitor than tin Fc fragment. The inhibitory capacity of the Fch and Fc fragments was greatly diminished following partial reduction and alkylation On the basis of the inhibitory pattern of IgG fragments, it is suggested that the region on the immunoglobulin molecule involved in binding to the Fc receptor of the effector lymphocytic cell may be located within the CH2 domain.     

Age-related loss of synaptophysin immunoreactive presynaptic boutons within the hippocampus of APP751SL, PS1M146L, and APP751SL/PS1M146L transgenic mice

Neuron and synapse loss are important features of the neuropathology of Alzheimer's disease (AD). Recently, we observed substantial age-related hippocampal neuron loss in APP751SL/PS1M146L transgenic mice but not in PS1M146L mice. Here, we investigated APP751SL mice, PS1M146L mice, and APP751SL/PS1M146L mice for age-related alterations in synaptic integrity within hippocampal stratum moleculare of the dentate gyrus (SM), stratum lucidum of area CA3 (SL), and stratum radiatum of area CA1-2 (SR) by analyzing densities and numbers of synaptophysin-immunoreactive presynaptic boutons (SIPBs). Wild-type mice, APP751SL mice and PS1M146L mice showed similar amounts of age-related SIPB loss within SM, and no SIPB loss within SL. Both APP751SL mice and PS1M146L mice showed age-related SIPB loss within SR. Importantly, APP751SL/PS1M146L) mice displayed the severest age-related SIPB loss within SM, SL, and SR, even in regions free of extracellular Abeta deposits. Together, these mouse models offer a unique framework to study the impact of several molecular and cellular events caused by mutant APP and/or mutant PS1 on age-related alterations in synaptic integrity. The observation of age-related SIPB loss within SR of PS1M146L mice supports a role of mutant PS1 in neurodegeneration apart from its contribution to alterations in Abeta generation.     

Long-term adhesion and survival of dissociated cortical neurons on miniaturised chemical patterns

The influence of neuron-adhesive pattern geometry on long-term adhesion, survival and pattern compliance of cortical neuronal tissue was studied over a period of 15 days. The results are relevant for a successful, long-term integration of neuronal cells with electrodes from micro-electronic devices. Microwells (depth 0.5μm), with diameters of 25, 50, 100 and 150 μm and spacing distances of 15, 30, 60 and 90 μm, were etched in a neuron-repellent fluorocarbon (FC) layer and coated with neuron-adhesive polyethylenimine (PEI). Results showed that adhesion, survival and compliance to the underlying patterns were geometry-and time-dependent. After 1 day, adhesion was inversely proportional to the diameter of the microwells, thus favouring the 25 μm microwells. However, adhesion was best on 50 μm microwells after 15 days. Survival of neurons was limited on 25 μm microwells (viability function V(D, T) was 0.08), as opposed to the better survival on 150 μm microwells (V(D, T) was 0.25) after 15 days. In summary, the study shows that the chemical patterns with microwells of 150 μm diameter (90 μm spacing gap) are most suitable for application on neuro-electronic devices owing to the better long-term survival and high pattern compliance of the neuronal cells.