Bibliothek

Notizen: Abstract: Amyloid β-peptide (Aβ) is the principal component of neuritic plaques in the brain in Alzheimer's disease (AD). Recent studies revealed that Aβ can be neurotoxic by a mechanism involving free radical production and loss of cellular ion homeostasis, thus implicating Aβ as a key factor in the pathogenesis of AD. However, other proteins are present in plaques in AD, including the protease thrombin and protease nexin-1 (PN1), a thrombin inhibitor. We therefore tested the hypothesis that thrombin and PN1 modify neuronal vulnerability to Aβ toxicity. In dissociated rat hippocampal cell cultures the toxicity of Aβ was significantly enhanced by coincubation with thrombin, whereas PN1 protected neurons against Aβ toxicity. Aβ induced an increase in levels of intracellular peroxides and calcium. Thrombin enhanced, and PN1 attenuated, the accumulation of peroxides and calcium induced by Aβ. Taken together, these data demonstrate that thrombin and PN1 have opposing effects on neuronal vulnerability to Aβ and suggest that thrombin and PN1 play roles in the pathogenesis of neuronal injury in AD.

Notizen: Abstract: Corticotropin-releasing factor (CRF) appears to integrate the endocrine, autonomic, immunologic, and behavioral responses of mammals to stress. To investigate further the role of CRF in the CNS, we have begun investigating the usefulness of “antisense knockdown” strategies directed against the CRF receptor using rat anterior pituitary gland primary cell cultures. The 15-mer antisense (5′ CTG-CGG-GCG-CCG-TCC 3′) and “scrambled” control (5′ CGT-CCG-CGC-GCT-GCG 3′) oligonucleotides were synthesized based on the rat CRF receptor sequence just downstream of the initiation codon. In each of four separate experiments, exposure to 10 µmol/L of antisense oligonucleotide for 40–67 h resulted in significant (17–36%) decreases in 125I-ovine CRF binding to pituitary cells as compared with either control (no oligonucleotide) or 10 µmol/L of “scrambled” oligonucleotide. Moreover, compared with scrambled oligonucleotide, exposure to 10 µmol/L of antisense oligonucleotide, which produced a 22% decrease in CRF receptor binding, also resulted in a significant attenuation of the adrenocorticotrophic hormone response following a 30-min challenge with 100 pmol/L of CRF. Thus, CRF receptor antisense oligonucleotides apparently reduce functional expression of CRF receptors. This technique may be useful in studying the kinetics of CRF receptor production and the physiological functions of CRF receptors within the CNS.

Notizen: Abstract: Evidence from in vitro studies suggests that excitotoxic neuronal degeneration can occur by either an acute or delayed mechanism. Studies of the acute mechanism in isolated chick embryo retina using histological methods indicate that this process is rapidly triggered by activation of glutamate receptors of either the N-methyl-d-aspartate (NMDA) or non-NMDA subtypes. The delayed mechanism, studied primarily in cortical and hippocampal cell cultures prepared from embryonic rodent brain, requires activation of NMDA receptors. In these cell culture systems, stimulation of non-NMDA receptors does not rapidly trigger delayed neuronal degeneration, or does so only indirectly, via activation of NMDA receptors secondary to glutamate release. To provide a more valid basis for comparison of these two mechanisms, we have modified the isolated chick embryo retina model to permit studies of delayed as well as acute excitotoxic neurodegeneration. Retinas maintained for 24 h exhibited no morphological or biochemical signs of damage. Retinal damage was assessed by measuring lactate dehydrogenase (LDH) present in the medium at various times after exposure to agonists and normalized to total LDH in each retina. Glutamate exposure (1 mM, 30 min) did not result in LDH release by the end of the exposure period, but LDH was released over the following 24 h. Briefer periods also led to substantial LDH release. Incubation in the presence of NMDA, or the non-NMDA agonists kainate (KA) or α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA), led rapidly to delayed LDH release. NMDA and AMPA were more potent than glutamate, but high concentrations of glutamate led to more LDH release than high concentrations of these agonists. KA was a powerful excitotoxin, providing more LDH release than glutamate, NMDA, or AMPA at every concentration tested. The delayed LDH release induced by glutamate involved activation of both NMDA and non-NMDA receptors, as a combination of receptor-selective antagonists was necessary to provide complete blockade. These results indicate that glutamate, NMDA, AMPA, and KA all cause delayed as well as acute excitotoxic damage in the retina. It is interesting that brief exposure to the non-NMDA receptor agonists, in relatively low concentrations, led to delayed LDH release. This is different than in other in vitro models of delayed excitotoxic neurodegeneration.

Notizen: Objective To evaluate the effect of 75 mg oestradiol implants on the bone mass of older postmenopausal women with osteoporosis.Design One year prospective-controlled study.Subjects Thirty-two postmenopausal women over 60 years of age and more than five years postmenopausal with low bone density were recruited. Thirty women completed treatment for one year with 75 mg oestradiol implants. The changes in bone density were compared with a control group of 14 women. The side effects with treatment were documented.Main outcome measures Dual energy X-ray absorptiometry of the lumbar spine and proximal femur using Hologic 1000 QDR before treatment and at one year. Plasma oestradiol assays were performed before and after one year.Results The median percentage changes in the treated group after one year was 12.6% at the lumbar spine and 5.22 % at the total hip. The increase in vertebral bone density was greatest in women with low initial bone density (r = 0.35, P 〈 0.05) and those with highest treatment plasma oestradiol levels (r = 0.47, 〉P 〈 0.01). Side effects were common but most were transient and mild in nature.Conclusion 75 mg oestradiol implants significantly increase the bone mineral density at the spine and hip of older postmenopausal women with established osteoporosis.

Notizen: Abstract: The invertebrate phototransduction system is a valuable model of the ubiquitous inositol lipid signalling system. Taking advantage of the ability to obtain relatively large amounts of retinal material from the cephalopod eye, partial protein sequence data were obtained for a 92-kDa component isolated from a detergent-insensitive cytoskeletal fraction of a squid retinal microvillar membrane preparation. Degenerate oligonucleotides, designed on the basis of these sequence data, were used to isolate a full-length cDNA, encoding the 92-kDa component, using both cDNA library screening and 5′-rapid amplification of cDNA ends (5′-RACE) techniques. Comparison of the amino acid sequence encoded by this cDNA with entries in the OWL composite protein sequence database reveals greatest sequence similarity with the products of the Drosophila trp and trpl genes. Greatest variation from the Drosophila Trp protein is seen in the carboxyl-terminal region, which is considerably truncated in the squid protein and which accounts for most of the substantial difference in molecular weight seen between these proteins. This variation may be significant as the carboxyl-terminal domain has been shown to be in the regulation of several ligand-gated channels. The carboxyl-terminal domain has been expressed and shown to interact with calmodulin in a calcium-dependent fashion, thereby supporting this hypothesis. The likely occurrence of other homologues in a variety of systems suggests that this is a novel and important family of regulated ion channels involved in calcium signalling.

Notizen: Objective To determine the efficacy of a 100 μg twice weekly dose of Estraderm TTS compared with a 200 μg dose in the treatment of severe PMS, and to determine the overall acceptability of the treatment. To determine the serum oestradiol levels produced by the two doses of Estraderm and to discover whether the lower dose suppresses ovulation.Design Main: randomised, prospective, comparative study. Subsidiary: cross-sectional and prospective.Setting Premenstrual syndrome clinic in teaching hospital.Subjects Women with severe PMS confirmed by prospective daily symptom recording.Interventions Estraderm TTS® at a dose of either 100 or 200 μg twice weekly continuously with either dydrogesterone 10 mg or medroxyprogesterone acetate 5 mg, from day 17 to day 26 of each cycle.Main outcome measures Main: change in total, exponentially smoothed, average maximum score (total-ESAmax) of 10 common premenstrual syndrome symptoms derived from Trigg's trend analysis and patient satisfaction. Subsidiary: plasma oestradiol and day 21 progesterone levels.Results Main: no difference in change in total-ESAmax between Estraderm 100 μg and 200 μg groups. Greater drop-out rate and greater incidence of side effects attributed to oestrogen in higher dosage group. Satisfaction rate of 45% to 57% at eight months. Subsidiary: 1. Mean (95% CI) oestradiol level of 300 pmol/l (255 to 345) with Estraderm 100 Fg and 573 (494 to 693) with Estraderm 200 pg; 2. Estraderm 100 μg suppresses mid-luteal progesterone from a mean (95% CI) of 353 (28.4 to 427) to 3.4 (2.4 to 4.5).Conclusions Estraderm TTS 100 μg twice weekly is as effective as 200 μg twice weekly in reducing symptom levels in severe premenstrual syndrome but is better tolerated. Estraderm 100 μg suppresses ovulation and results in a mean plasma oestradiol level similar to that observed in a spontaneous cycle.