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R Graf

Publications and source records attributed to R Graf.

At least 91 records · Page 5Linked to original sources
PubMed1999

The anchoring zone in the human placental amnion: bunches of oxytalan and collagen connect mesoderm and epithelium.

This study deals with the examination of the elastic fibre system as well as collagen fibrils and collagen type IV in the amnion of the human chorionic plate of uncomplicated pregnancies at term. In organs other than placenta, the elastic fibre system comprises elastic fibres, elaunin and oxytalan microfibrils. The investigation was performed by light and electron microscopy and immunocytochemistry. Abundant oxytalan fibres were present in all amnionic layers, while no elastic fibres were found. Oxytalan microfibrils formed a broad sub-epithelial layer and were intermingled with collagen fibrils in the subjacent compact layer and in the amnionic mesoderm. Light microscopically, bunches containing orcein-stained oxytalan and collagen-type-IV-immuno-stained microfibrils were seen rising from the amnionic mesoderm perpendicularly towards the epithelial layer, where they obviously inserted. It can be assumed that the subepithelial microfibrillar layer and the following compact layer form an anchoring zone between the amnionic mesoderm and the epithelium that may contribute to the maintenance of strength. The ultrastructure of the bunches clearly showed collagen fibrils mixed with oxytalan microfibrils. No collagen type I-immunostaining was found in the bunches. After pretreatment of cryostat sections with elastase, oxytalan-orcein-staining was absent, but collagen type IV-immunoreactivity was not altered. Furthermore, after oxytalan-orcein-staining resp. anti-collagen type IV incubation, all positive fibres revealed an identical morphological pattern. We propose that oxytalan and collagen type IV may represent further members of the microfibril complex.

Adult↗
PubMed1999

Increase of segments of elastic-type blood vessel walls in fetal placental stem villi during pre-eclampsia at term.

In recent studies we described the presence of elastic-type blood vessels within trunci and rami chorii of human placental stem villi. For systemic and pulmonary hypertension it is known that elastic fibres are enhanced in arteries. The aim of our study was, therefore, to examine whether pre-eclampsia may lead to an increase of elastic tissue fibres in blood vessel walls of placental stem villi and whether there are differences in the thickness of blood vessel walls within these villi when compared to normotensive pregnant women. Twenty-six women with uncomplicated pregnancies and 25 patients with pre-eclampsia were investigated. Unfixed cryostat serial sections were processed for conventional orcein staining and for the demonstration of alpha-actin-immunoreactivity. The intensity of orcein staining of stem villus blood vessel walls was evaluated by a semiquantitative score method. Significant higher intensities of orcein staining (P<0.00001) were calculated for blood vessel walls of placentae with pre-eclampsia. The amount of thick stem villus vessels (>41 microm) increased during pre-eclampsia from 39 gestational weeks onwards. Our study demonstrates that segments of thick blood vessel walls and elastic-type vessel walls are increased in placental stem villi of patients with pre-eclampsia. This reaction may protect the fetal placental vessels and avert an increase of the fetal hypertension.

Chorionic Villi↗
PubMed1999

Cyclin A-dependent kinase activity affects chromatin binding of ORC, Cdc6, and MCM in egg extracts of Xenopus laevis.

The initiation of DNA replication in eukaryotes requires the loading of the origin recognition complex (ORC), Cdc6, and minichromosome maintenance (MCM) proteins onto chromatin to form the preinitiation complex. In Xenopus egg extract, the proteins Orc1, Orc2, Cdc6, and Mcm4 are underphosphorylated in interphase and hyperphosphorylated in metaphase extract. We find that chromatin binding of ORC, Cdc6, and MCM proteins does not require cyclin-dependent kinase activities. High cyclin A-dependent kinase activity inhibits the binding and promotes the release of Xenopus ORC, Cdc6, and MCM from sperm chromatin, but has no effect on chromatin binding of control proteins. Cyclin A together with ORC, Cdc6 and MCM proteins is bound to sperm chromatin in DNA replicating pseudonuclei. In contrast, high cyclin E/cdk2 was not detected on chromatin, but was found soluble in the nucleoplasm. High cyclin E kinase activity allows the binding of Xenopus ORC and Cdc6, but not MCM, to sperm chromatin, even though the kinase does not phosphorylate MCM directly. We conclude that chromatin-bound cyclin A kinase controls DNA replication by protein phosphorylation and chromatin release of Cdc6 and MCM, whereas soluble cyclin E kinase prevents rereplication during the cell cycle by the inhibition of premature MCM chromatin association.

Animals↗
PubMed1999

Breakdown of calcium homeostasis in relation to tissue depolarization: comparison between gray and white matter ischemia.

In vitro studies suggest that ischemic injury of cerebral white matter is mediated by nonsynaptic cellular mechanisms, such as Ca2+ entry into axons through reversal of the Na+ -Ca2+ exchanger. The authors investigated extracellular Ca2+ concentration in relation to tissue depolarization (direct current potential) in vivo using ion-selective electrodes in cortical gray and subcortical white matter of alpha-chloralose-anesthetized cats during 120 minutes of global cerebral ischemia. On induction of ischemia, regional CBF, as measured by hydrogen clearance, ceased. The direct current potential decreased rapidly within minutes in gray matter and with little time delay in white matter. Extracellular Ca2+ concentration decreased just as quickly in gray matter. In white matter, in contrast, extracellular Ca2+ increased in the first 20 to 30 minutes, and a delayed and much slower decline, compared with gray matter, was observed thereafter, reaching a minimal level only about 60 minutes after occlusion. Our results suggest that smaller and delayed transmembrane shifts of Ca2+ are correlates of delayed ischemic membrane dysfunction in central white matter tracts, which may be explained by a lack of synaptic mechanisms.

Action Potentials↗
PubMed1999

Regulation of PSP/reg in rat pancreas: immediate and steady-state adaptation to different diets.

Pancreatic stone protein/reg protein (PSP/reg) is a secretory pancreatic protein of hitherto unknown function. It is precursor to a spontaneously precipitating peptide called pancreatic thread protein, which is found in protein plugs within the pancreatic ductal system. Increasing PSP/reg concentrations in pancreatic juice might augment the risk of intraductal plug formation and therefore be a condition predisposing to chronic pancreatitis. Malnutrition is associated with a high incidence of chronic pancreatitis in tropical countries. In a diet study with rats, we tested the hypothesis that protein malnutrition leads to increased PSP/reg concentrations in pancreatic juice. A highly sensitive and reliable enzyme-linked immunosorbent assay (ELISA) for rat PSP/reg was newly established. Male Sprague-Dawley rats were allocated to three nearly isocaloric experimental diets, which contained 0, 45, or 82% casein, respectively, or to a control diet (22% casein). We evaluated PSP/reg expression under these four dietary conditions on the RNA and on the protein level, performing a time-course study over a period of 28 days. Our results demonstrate that PSP/reg expression is not increased because of a protein-deficient diet if investigated under steady-state conditions. After a temporary increase in PSP/reg levels due to a carbohydrate-deficient high-protein diet, we could not find signs of a diet-dependent regulation of this protein. The regulation of PSP/reg thus differs from that of most other pancreatic secretory proteins. Our findings contradict earlier reports that had drawn conclusions based solely on messenger RNA levels.

Adaptation, Physiological↗
PubMed1999

Which targets are relevant for therapy of acute ischemic stroke?

BACKGROUND: The efficiency of various strategies of neuroprotection is well documented in animal experiments but is thus far disappointing in ischemic stroke, for which only early reperfusion induced by thrombolysis has improved clinical outcome. This discrepancy between expectation from experimental research and clinical reality may be related to differences in the pathogenetic factors contributing to infarction. SUMMARY OF COMMENT: Positron emission tomography cerebral blood flow studies within 3 hours of onset were used to identify the various compartments of the infarct outlined on MRI 2 to 3 weeks after a hemispheric stroke in 10 patients. Critical hypoperfusion below the viability threshold accounted for the largest proportion (mean, 70%) of the final infarct, whereas penumbral tissue (18%) and initially sufficiently perfused tissue (12%) were responsible for considerably smaller portions of the final infarct. CONCLUSIONS: These results indicate that early critical flow disturbance leading to rapid cell damage is the predominant cause of infarction, while secondary and delayed pathobiochemical processes in borderline or initially sufficiently perfused regions contribute only little to the final infarct. Therefore, emerging therapeutic strategies should be targeted to the initially critically perfused tissue subcompartments. Clinical drug trials might benefit from stratification of patients for target tissue compartments applying functional imaging.

Acute Disease↗
PubMed1999

Placental glucose transporter expression is regulated by glucocorticoids.

Although glucocorticoids play important roles in development and fetal programming, they are widely used for treatment of a variety of diseases during pregnancy. In various tissues, glucocorticoids down-regulate glucose transport systems; however, their effects on glucose transporters in the placenta are unknown. In the present study, the glucose carrier proteins GLUT1 and GLUT3 were localized in the trophoblast and endothelium of the human, rat, and mouse placenta. Subsequently, it was investigated whether glucocorticoids affect messenger ribonucleic acid and protein expression of these molecules by Northern and Western blotting using 1) human term placental trophoblast cells cultured in the presence or absence of 0.5, 5, and 50 micromol/L triamcinolone; 2) placentas of rats that received a single i.p. dose of 0.38 mg/kg triamcinolone; and 3) placentas of transgenic mice bearing an antisense glucocorticoid receptor gene construct. In all of these systems, both glucose transporters were significantly down-regulated (P < 0.05), with the exception of increased GLUT3 messenger ribonucleic acid and protein levels in transgenic mice. The results demonstrate that triamcinolone is a potent regulator of placental GLUT1 and GLUT3 expression involving the glucocorticoid receptor. We speculate that impaired expression of placental glucose transporters after glucocorticoid administration might contribute to the adverse side-effects, the foremost of which is a growth-retarded fetus, of this treatment during pregnancy.

Animals↗
PubMed1998

Nucleotide sequence and deduced amino acid sequence of a putative asparagine synthetase in the mosquito Aedes aegypti (L.).

A cDNA was cloned from a Aedes aegypti head cDNA library, containing the complete coding sequence for an asparagine synthetase homolog. The predicted polypeptide sequence exhibits high homology with different proteins of the 'purF' glutamine amidotransferase enzyme family. The aminoterminal region, containing Cys-1 which is crucial to perform the glutaminase reaction, was highly conserved among the asparagine synthetase family. Subsequent expression of the cDNA yielded a 54,000 Da protein corresponding to the molecular weight of other asparagine synthetases.

Aedes↗
PubMed1998

Identification of a steroidogenic neurohormone in female mosquitoes.

In the female mosquito, Aedes aegypti, neurohormones are released from the brain in response to a blood meal and stimulate the ovaries to secrete ecdysteroid hormones, which modulate yolk protein synthesis in the fat body. Neuropeptides with this bioactivity were isolated from head extracts, and partial sequences from these peptides when aligned gave a 31-residue sequence at the amino terminus. Oligonucleotide primers for this sequence were used to amplify with the polymerase chain reaction a genomic DNA product that hybridized to a clone from a head cDNA library. The cDNA encodes a 149-residue preprohormone that is processed into an 86-residue peptide, as indicated by the mass value obtained from the native peptide, with the expected amino-terminal sequence. After modification, the cDNA for the putative neurohormone was expressed in a bacterial system, and the purified peptide had high specific activity in bioassays, thus confirming that it is a steroidogenic gonadotropin, the first to be identified for invertebrates.

Aedes↗
PubMed1998

Localization of the mosquito insulin receptor homolog (MIR) in reproducing yellow fever mosquitoes (Aedes aegypti).

The female mosquito takes a blood meal to produce a batch of eggs. Initiation of egg maturation and growth of oocytes is governed by several endocrine factors. Peptide factors from the brain are involved in this process and some are also responsible for the induction of ecdysone secretion. The latter appears to be required to maintain a high rate of vitellogenin synthesis. By analogy with the known functions of insulin-like molecules (e.g. bombyxins) which in insects activate the secretion of ecdysteroids, we have postulated that there is an insulin receptor homolog responsible for activation of endysone secretion in the ovary. We have recently cloned the mosquito homolog (MIR) and are now investigating its spatial and temporal distribution. Here, we have localized the insulin receptor (MIR) both at the mRNA and protein level using in situ-hybridization and immunocytochemistry. The receptor is expressed before a blood meal mainly in the nurse cells of ovaries. After a meal, follicle and nurse cells contain mRNA coding for the receptor. The intensity of expression rises in the follicle cells until they degenerate during choriogenesis. Immunocytochemical localization confirms the in situ data: the protein is present before and after a meal. Both methods confirm our previous findings by Northern blot analysis, in which the ovary was found to be the main source of the receptor mRNA. The dynamics of receptor mRNA are related to the dynamics of ecdysone secretion and its action on physiological processes.

Journal Article↗
PubMed1998

A temporal profile of the endocrine control of trypsin synthesis in the yellow fever mosquito, Aedes aegypti.

Trypsin synthesis and secretion is induced after the female mosquito takes a blood meal. Its peak activity has been shown to be proportional to the amount and quality of food uptake. Further regulatory elements, hormones from the brain and the ovary, increase the synthethic rate of trypsin in the midgut by a factor of two. We investigated the temporal effect of removing the humoral factors by decapitation and ovariectomy. Trypsin synthesis was reduced to less than half its normal output when the operations were performed prior, or immediately after the blood meal. Postponing decapitation resulted in an increased activity. However, the dependence on hormones extended up to 14-16hrs after a meal, when maximal synthethic rates are assumed. Similarly, ovariectomy had a prolonged effect on trypsin synthesis. Finally, the lack of hormones reduced the synthetic capacity of the midgut even when small blood meals were given. We conclude that for continued efficient trypsin synthesis, humoral stimulation is necessary but is not part of the feedback mechanism that links the presence of food with the amount of trypsin secreted.

Journal Article↗
PubMed1998

Selective residual dipolar couplings in cross-linked elastomers by 1H double-quantum NMR spectroscopy.

1H double-quantum (DQ) solid-state NMR spectroscopy under fast magic-angle spinning (MAS) is introduced as a new spectroscopic tool for the investigation of the structure and local chain dynamics of elastomers. Dipolar connectivities between the protons of the various functional groups can be directly established from the highly resolved DQ solid-state NMR spectra as is shown for a series of cross-linked poly(styrene-co-butadiene). More quantitatively, residual dipolar couplings within and between the functional groups are evaluated selectively from the build-up curves of the double-quantum signals in the limit of the spin-pair approximation. In particular, the CH-CH and the CH2-CH couplings of butadiene, which both act predominantly along the chain-segment direction, have been measured relative to the CH2 coupling. The total build-up intensity is correlated with the cross-link density.

Cross-Linking Reagents↗
PubMed1998

Quantitative neuroimaging for the evaluation of the effect of stroke treatment.

A prerequisite for the successful treatment of acute ischaemic stroke is the existence of viable tissue that is morphologically intact but functionally impaired due to a flow decrease below a certain threshold. At this stage, tissue at risk of infarction can be identified only by functional imaging. This penumbral tissue can be classified as having a critical flow decrease with preservation of oxygen consumption and therefore increased oxygen extraction. Such 'misery-perfused' tissue has been observed consistently in the first few hours following ischaemic stroke but usually develops into necrosed tissue at follow-up observations. Several studies have indicated that penumbral tissue can be identified up to 17 h or even 48 h after stroke in the border zones of ischaemic tissue and that this condition is occasionally reversible without resulting in permanent infarction. Positron emission tomography studies of cerebral blood flow (CBF), cerebral metabolic rate of oxygen (CMRO2) and cerebral metabolic rate of glucose (CMRglc) can be used to demonstrate the effect of treatment on functional variables within tissue showing perfusional disturbances. Such studies have shown the value of these markers of ischaemia, which also correlate with clinical efficacy. However, when therapeutic strategies, such as thrombolysis, do not permit arterial blood sampling, quantitative determinations of CBF and CMRO2 are not feasible. In such cases relative indices, such as those for CBF, must be applied. Such qualitative assessments of perfusion, which were calibrated in an independent cohort of patients with acute stroke, were used to demonstrate the effect of early systemic treatment of acute ischaemia with recombinant tissue plasminogen activator. By applying operationally defined thresholds for tissue viability and the penumbra, and co-registering these tissue compartments to infarcted and non-infarcted tissue on late magnetic resonance imaging, the proportions of at-risk tissue salvaged from infarction could be revealed in individual patients. In the future, functional imaging modalities that could eventually include tracers for neuronal integrity could be used to select patients for thrombolytic therapy. In some instances such techniques may permit the extension of the critical time period for inclusion of patients to aggressive stroke management strategies.

Animals↗
PubMed1998

Are there differences in the effects of long-acting calcium antagonists on ambulatory blood pressure? Extended-release nisoldipine versus amlodipine as a model.

BACKGROUND: Twenty-four-hour ambulatory blood pressure monitoring (ABPM), which provides important information regarding mean 24 h efficacy, variability of effect during sleeping-awake cycles, and effects on the early morning surge in blood pressure, is a sensitive method for evaluating efficacy of antihypertensive agents. Extended-release nisoldipine and amlodipine are long-acting dihydropyridine calcium antagonists used for the treatment of hypertension. Because these agents have different pharmacokinetic profiles, 24 h ABPM could provide clues regarding their different effects on blood pressure. OBJECTIVE: To assess the effects of extended-release nisoldipine and amlodipine on 24 h ambulatory blood pressure control and heart rate. METHODS: After completion of a 3-4 week placebo run-in period, 100 patients were randomly allocated to double-blind treatment with 10-40 mg extended-release nisoldipine or 2.5-10 mg amlodipine for 8 weeks, starting at the lowest dose. Medications were titrated at 2-week intervals on the basis of office blood pressures in seated patients. Twenty-four-hour ABPM was performed at placebo baseline and at the end of double-blind therapy. RESULTS: Extended-release nisoldipine and amlodipine provided equivalent mean 24 h changes in blood pressure [systolic blood pressure (SBP)/diastolic blood pressure decreases by 9.8/7.1 and 8.0/6.0 mmHg, respectively] and heart rate. These two treatments also provided similar changes in blood pressure at trough (22-24 h after dosing; decreases by 10.4/7.2 and 10.1/7.3 mmHg, respectively). The antihypertensive effects of amlodipine during the awake and sleeping intervals were similar (decreases by 9.6/5.9 and 9.9/5.8 mmHg, respectively, NS); whereas the effect of nisoldipine during the awake interval was significantly greater than its effect during the sleeping interval (decreases by 12.4/8.0 and 8.9/4.3 mmHg, respectively, P = 0.08/0.01). Furthermore, extended-release nisoldipine, but not amlodipine, blunted the rate of rise in early morning SBP. CONCLUSIONS: Extended-release nisoldipine and amlodipine have similar effects on mean 24 h and trough blood pressures. However, different effects during the sleeping and awake intervals and on the rate of rise in early morning SBP were observed with nisoldipine.

Journal Article↗
PubMed1998

[The acetabular labrum in infants].

The knowledge of embryology and early development of the hip joint (and especially the "Anlage" of the acetabular labrum) is necessary to correctly understand further growth disturbances and developmental dysplasias and dislocations of the hip joint. "Teratologic" luxations--based on damages of the fetal "Anlage"--should be distinguished from "developing deformations" of originally normal-shaped hip joints. By using modern imaging techniques, especially sonography and MRI, the morphologic relationships of the acetabular labrum in centered and decentered hip joints could be clarified. The tip and the basis of the labrum and their relations with the hyalin-preformed growth zone cartilage of the acetabular roof are changing during the process of decentering and can be clearly visualized by sonographic means. To understand the morphologic changes in decentered hip joints, a clear and consistent terminology should be used: The term "limbus" is misleading and should be avoided; one should use the terms "acetabular labrum" and "hyalin-preformed cartilaginous acetabular roof" for the two histologic subdivisions of the acetabular roof cartilage. We do not know much about size and shape, about nutrition and vascularity of the acetabular labrum, and only few facts about the junction zone of the labral attachment to the hyalin cartilage acetabular roof, and so on. We also do not have any clear evidence about possible micro-damages of the labral-capsular-complex during successful closed reduction and their sequelae in childhood and adolescence, possibly being one cause of labral lesions in adults. Further basic research in this field seems useful and necessary.

Acetabulum↗