Search PubMed⌕ Search

Biomedical subjects

R Graf

Publications and source records attributed to R Graf.

At least 109 records · Page 6Linked to original sources

[Labrum lesions and residual dysplasia of the hip joint. Definition and prospectives].

Labral lesions are a sign of biomechanical decompensation of the hip joint and often represent the first clinical symptom of residual hip dysplasia (RHD) in the adult. Provocation tests (impingement, apprehension) are typical but not specific. Labral lesions and concomitant findings (intra- and extraosseous ganglia, stress bone marrow oedema) can be detected by magnetic resonance arthrography (MRA) with an accuracy of 91%. Primary therapeutic goal is the normalization of the underlying pathomorphology and instability by a redirectional acetabular osteotomy. There are several concepts concerning simultaneous arthrotomy at the time of osteotomy: no arthrotomy at all, selective arthrotomy, routine arthrotomy in every case. There are more clinical studies necessary before one of these concepts can be widely accepted and recommended. Based on preliminary results, palliative arthrotomy with partial labral resection but without corrective osteotomy in osteoarthritis secondary to residual hip dysplasia gives poor results; we therefore urgently dissuade from palliative labral surgery via arthrotomy. Whether labral surgery via arthroscopy might be a useful concept in symptomatic residual hip dysplasia, is still an open question. In this review article, the "state of the art" presented at the "Vienna Labral Symposium 1997" is reflected and summarized. At the end of this article, a "common statement" of the experts is published in English and German language.

Acetabulum↗

Signal peptide fragments of preprolactin and HIV-1 p-gp160 interact with calmodulin.

Secretory proteins and most membrane proteins are synthesized with a signal sequence that is usually cleaved from the nascent polypeptide during transport into the lumen of the endoplasmic reticulum. Using site-specific photo-crosslinking we have followed the fate of the signal sequence of preprolactin in a cell-free system. This signal sequence has an unusually long hydrophilic n-region containing several positively charged amino acid residues. We found that after cleavage by signal peptidase the signal sequence is in contact with lipids and subunits of the signal peptidase complex. The cleaved signal sequence is processed further and an N-terminal fragment is released into the cytosol. This signal peptide fragment was found to interact efficiently with calmodulin. Similar to preprolactin, the signal sequence of the HIV-1 envelope protein p-gp160 has the characteristic feature for calmodulin binding in its n-region. We found that a signal peptide fragment of p-gp160 was released into the cytosol and interacts with calmodulin. Our results suggest that signal peptide fragments of some cellular and viral proteins can interact with cytosolic target molecules. The functional consequences of such interactions remain to be established. However, our data suggest that signal sequences may be functionally more versatile than anticipated up to now.

Amino Acid Sequence↗

Nitric oxide generation from sodium nitroprusside and hydroxylamine in brain.

We aimed to demonstrate different modes of in situ nitric oxide (NO) generation from two NO donors in cerebral cortex of halothane anesthetized cats. NO donors were delivered by in vivo microdialysis into the cortical tissue where an NO electrode measured real-time changes in extracellular NO concentration. In vitro testing in Finger's solution revealed that sodium nitroprusside (SNP) liberates NO spontaneously in the presence of light, whereas hydroxylamine does not generate any NO under these conditions. Supplementation with 200 mM of the NO donors SNP and hydroxylamine evoked dose-related increases in NO concentration (52.0 +/- 6.7 nM, n = 5, and 14.0 +/- 3.8 nM, n = 5, respectively). The increase in NO concentration was significantly greater during SNP than during hydroxylamine delivery (p < 0.001). In contrast, CBF increased similarly under the influence of both NO donors. Electrocorticogram and cortical direct current potential remained unaffected. In conclusion, hydroxylamine is degraded into NO intracellularly and efficiently dilates cerebral blood vessels. SNP, in contrast, generates NO mainly in the extracellular space.

Animals↗

Molecular mechanism of membrane protein integration into the endoplasmic reticulum.

As proteins are integrated into the membrane of the endoplasmic reticulum, some hydrophilic polypeptide segments are transported through the translocation channel, others remain in the cytosol, and hydrophobic transmembrane sequences are released into the lipid phase. We have addressed the molecular mechanism by which these events occur. We demonstrate that both the lumenal and the cytosolic domains of a membrane protein are synthesized while the ribosome is membrane bound, so that even cytosolic domains come in contact with the translocation channel. We also find that, before translation of the protein is terminated, transmembrane sequences can laterally exit the translocation channel and enter the lipid environment. These results have significant implications for the folding and assembly of membrane proteins.

Animals↗

Direct and GTP-dependent interaction of ADP ribosylation factor 1 with coatomer subunit beta.

A site-directed photocrosslink approach was used to elucidate components that interact directly with ADP- ribosylation factor (ARF)-GTP during coat assembly. Two ARF mutants were generated that contain a photolabile amino acid at positions distant to each other within the ARF molecule. Here we show that one of the two positions specifically interacts with coatomer subunit beta both on Golgi membranes and in isolated coat protein complex type I (COPI)-coated vesicles. Thus, a direct and GTP-dependent interaction of coatomer via beta-coat protein complex (COP) with ARF is involved in the coating of COPI-coated vesicles. These data implicate a bivalent interaction of the complex with the donor membrane during vesicle formation.

ADP-Ribosylation Factors↗

Pancreatic stone protein (lithostathine), a physiologically relevant pancreatic calcium carbonate crystal inhibitor?

Apart from digestive enzymes, pancreatic juice contains several proteins that are not directly involved in digestion. One of these, lithostathine, has been reported to exhibit calcite crystal inhibitor activity in vitro. As pancreatic juice is supersaturated with respect to calcium carbonate, it was hypothesized that lithostathine stabilizes pancreatic juice. Lithostathine is cleaved by trace amounts of trypsin, resulting in a C-terminal polypeptide and an N-terminal undecapeptide, which has been identified as the active site of lithostathine regarding crystal inhibition. We produced rat lithostathine in a baculovirus expression system. In order to test its functional activity, the protein was purified using a nondenaturing multi-step procedure. In the low micromolar range, recombinant rat lithostathine in vitro exhibited calcite crystal inhibitor activity, confirming earlier reports. Limited tryptic proteolysis of recombinant lithostathine was performed, and the two cleavage products were separated; the C-terminal polypeptide was precipitated by centrifugation, and the N-terminal undecapeptide was purified by high performance liquid chromatography. Only the C-terminal peptide displayed measurable calcite crystal inhibitory activity. Furthermore, synthetic undecapeptides with identical sequence to the N-terminal undecapeptides of rat or human lithostathine were inactive. However, when tested in the same in vitro assays, other pancreatic or extra-pancreatic proteins show inhibitory activity in the same concentration range as lithostathine, and inorganic phosphate is active as well. Based on these findings it seems unlikely that lithostathine is a physiologically relevant calcite crystal inhibitor. The name "lithostathine" is therefore inappropriate, and the protein's key function remains to be elucidated.

Amino Acid Sequence↗

[A phase-I/II study on the local hyperthermia of cervical N2/N3 lymph node metastases].

BACKGROUND: Patients with advanced lymph node metastases from head and neck tumors at stage N2/N3 (i.e. UICC IV) present a difficult therapeutic problem. Despite combined radio-chemotherapy and hyperfractionated and/or accelerated fractionation regimens, local control of these tumors remains unsatisfactory. For this reason, the value of local radio wave/microwave hyperthermia was examined for this patient group in a phase I/II study.

Adult↗

[Hip ultrasonography. Basic principles and current aspects].

Hip sonography is the subject of controversial discussion in several countries. This is because of the different demands made on the diagnosis. In principle the techniques described by Harke, Terjesen and Suzuki are satisfied by a differentiation of luxation from non-luxated joints. An evaluation of the hyalin cartilaginous preformed acetabular roof does not exist. Physiological variations dependent on age cannot be differentiated from real dysplasias by the techniques mentioned above. The sonographic examination technique in German-speaking countries evaluates the maturation of the joint and quantifies the cartilage and the bony acetabular roof. Additionally, this coverage is seen in relation to the age, and the deformation of the acetabulum in luxated joints is analysed. Physiological elastic deflection of the hyalin cartilaginous part of the acetabulum can be separated from real instability signs. Oblique positions of the probe during examination of the baby can lead to diagnostic errors, so special examination techniques have to be used. 3D-sonograms may improve the diagnostic possibilities. A general screening of newborns in use in Austria since 1992 has demonstrated that the rate of treatment can be decreased by about 50% and sonography, used correctly, can avoid overtreatment.

Acetabulum↗

[Ultrasonography-guided therapy].

Therapy for hip dysplasia and DDH by means of sonography. Numerous methods are available for treating "congenital" dysplasia of the hip (CDH). The important point seems to be the differentiation between dysplastic hip joints with and without displaced femoral heads. A historical review of the diagnosis of so-called "congenital dysplasia" and dislocation of the hip and different indications for therapeutic maneuvers is given. Generally, a three-step plan of treatment is recommended: dislocated heads must be reduced, after which retention is needed until the hip joint is stable. This is followed by maturation period, until the nonossified hyalin cartilagenous performed acetabular roof is ossified. All hip joints, regardless of age, can be assigned to one of these phases. In all cases, measures are taken to prevent ischemic necrosis of the femoral head. To prevent the residual dysplasia of the acetabulum roof and femoral head necrosis, the reduction-retention position should be more than 90 degrees flexion in the hip joint, but not more than 45-50 degrees abduction. In each therapeutic phase there is a different biomechanical situation in the hip joint. The appropriate orthosis for treatment is chosen according to the particular biomechanical situation. Each sonographic type can be set up according to the our classification in the three-phase treatment system. In this way the adequate orthosis for treatment in each treatment-phase can be chosen.

Braces↗

Molecular anatomy of the perivascular sheath in human placental stem villi: the contractile apparatus and its association to the extracellular matrix.

In previous studies, we have shown that smooth muscle cells and myofibroblast subpopulations of the perivascular stem villous sheath of the human placenta contain focal adhesion plaques and talin immunoreactivity. The close association of these cells to elastic and collagen fibres have led to the assumption of a functional myofibroelastic unit within the perivascular stem villous sheath. Interactions between the extracellular matrix and smooth muscle cells depend on a variety of structural protein assemblies. In the present study, we examined, by immunocytochemistry, whether the molecular assembly of extracellular matrix proteins and molecules of focal adhesions, known to be essential for signal transduction in smooth muscle cells, are also found in smooth muscle cells of the perivascular stem villous sheath of the human placenta. Vascular and extravascular smooth muscle cells were immunoreactive for alpha-actinin, vinculin, paxillin and tensin, the integrin chains alpha1 and beta1, and the basement membrane components laminin and heparan/-chondroitin sulfate proteoglycan perlecan. pp125(FAK) did not react. In the extracellular matrix of blood vessel walls and the perivascular stem villous sheath, we found immunoreactivity of fibronectin and collagen types I, VI and undulin (collagen type XIV). From our data we conclude that within the perivascular stem villous sheath, there exists a system of signal transduction molecules, indicating a cross talk between the smooth muscle cells of this sheath and their surrounding extracellular matrix.

Actins↗

Identification of three allatostatins and their cDNA from the mosquito Aedes aegypti.

Three allatostatins have been isolated from the mosquito Aedes aegypti. These peptides have the following structures: Ser-Pro-Lys-Tyr-Asn-Phc-Gly-Leu-amide, Leu-Pro-His-Tyr-Asn-Phe-Gly-Leu-amide, and Arg-Val-Tyr-Asp-Phe-Gly-Leu-amide. A cDNA encoding these peptides was isolated from an abdominal ganglia cDNA library and sequenced. It was found to encode two additional allatostatins: Ala-Ser-Ala-Tyr-Arg-Tyr-His-Phe-Gly-Leu-amide and Leu-Pro-Asn-Arg-Tyr-Asn-Phe-Gly-Leu-amide. Northern analysis of whole mosquito mRNA revealed a single prepro-allatostatin message of around 3,000 bases. Identification of a partial prepro-allatostatin cDNA from a midgut cDNA library shows that the same gene is also expressed in the mosquito midgut.

Aedes↗

Binding of antibodies against high and low molecular weight cytokeratin proteins in the human placenta with special reference to infarcts, proliferation and differentiation processes.

Recent immunocytochemical studies have shown that placental villous trophoblasts contain the high molecular weight cytokeratin (CK) proteins 5/6 and 17. In the case of CK 17, trophoblastic immunostaining was positive in villi covered by fibrinoid. CKs 5/6 and 17 are expressed by hyperproliferative cells. The aim of this investigation was to examine the location of these CKs in placental infarcts, known to be demarcated by fibrinoid and hyperproliferative trophoblasts. The results were compared with those obtained by immunostaining against Ki-67, tenascin and alpha 1-, alpha 6- and beta 1-integrins, which are involved in cell proliferation, differentiation and regenerative processes. Furthermore, the expression of the single CKs 7, 8, 10, 13, 14, 18 and 19 was investigated by immunocytochemistry and immunoblotting. While low and high molecular weight CKs were present in villous and extravillous trophoblasts, only low molecular weight CKs were detected in vascular and extravascular placental smooth muscle cells. Placental infarcts revealed different immunoreactivities in the infarct margin and centre: high molecular CKs, tenascin, Ki-67 and oncofoetal fibronectin predominated in the infarct margin, low molecular CKs, fibrin and integrins in the centre. The expression of tenascin and a defined change in the expression of CK 17 indicates villous repair and hyperproliferative mechanisms in placental infarcts.

Antibody Formation↗

Repeat positron emission tomographic studies in transient middle cerebral artery occlusion in cats: residual perfusion and efficacy of postischemic reperfusion.

The wider clinical acceptance of thrombolytic therapy for ischemic stroke has focused more attention on experimental models of reversible focal ischemia. Such models enable the study of the effect of ischemia of various durations and of reperfusion on the development of infarctions. We used high-resolution positron emission tomography (PET) to assess cerebral blood flow (CBF), cerebral metabolic rate of oxygen (CMRO2), oxygen extraction fraction (OEF), and cerebral metabolic rate of glucose (CMRglc) before, during, and up to 24 h after middle cerebral artery occlusion (MCAO) in cats. After determination of resting values, the MCA was occluded by a transorbital device. The MCA was reopened after 30 min in five, after 60 min in 11, and after 120 min in two cats. Whereas all cats survived 30-min MCAO, six died after 60-min and one after 120-min MCAO during 6-20 h of reperfusion. In those cats surviving the first day, infarct size was determined on serial histologic sections. The arterial occlusion immediately reduced CBF in the MCA territory to < 40% of control, while CMRO2 was less affected, causing an increase in OEF. Whereas in the cats surviving 24 h of reperfusion after 60- and 120-min MCAO, OEF remained elevated throughout the ischemic episode, the initial OEF increase had already disappeared during the later period of ischemia in those cats that died during the reperfusion period. After 30-min MCAO, the reperfusion period was characterized by a transient reactive hyperemia and fast normalization of CBF, CMRO2, and CMRglc, and no or only small infarcts in the deep nuclei were found in histology. After 60- and 120-min MCAO, the extent of hyperperfusion was related to the severity of ischemia, decreased CMRO2 and CMRglc persisted, and cortical/subcortical infarcts of varying sizes developed. A clear difference was found in the flow/metabolic pattern between surviving and dying cats: In cats dying during the observation period, extended postischemic hyperperfusion accompanied large defects in CMRO2 and CMRglc, large infarcts developed, and intracranial pressure increased fatally. In those surviving the day after MCAO, increased OEF persisted over the ischemic episode, postischemic hyperperfusion was less severe and shorter, and the perfusional and metabolic defects as well as the final infarcts were smaller. These results stress the importance of the severity of ischemia for the further course after reperfusion and help to explain the diverging outcome after thrombolysis, where a relation between the residual flow and the effectiveness of reperfusion was also observed.

Animals↗

Reduction of infarct volume by halothane: effect on cerebral blood flow or perifocal spreading depression-like depolarizations.

Halothane is a strong inhibitor of potassium evoked spreading depression (SD) in cats. In the current study, we investigate halothane effects on induction of perifocal SD-like depolarizations, CBF, and infarct evolution in focal ischemia. Calomel and platinum electrodes measured cortical direct current potential and CBF in ectosylvian, suprasylvian, and marginal gyri. Left middle cerebral artery occlusion (MCAO) induced permanent focal ischemia for 16 hours in artificially ventilated cats (30% oxygen, 70% nitrous oxide) under halothane (0.75%, n = 8) or alpha-chloralose anesthesia (60 mg/kg intravenously, n = 7). Under alpha-chloralose, MCAO induced severe ischemia in ectosylvian and suprasylvian gyri(mean CBF < 10 mL/100 g/min), and direct current potentials turned immediately into terminal depolarization. In marginal gyri, CBF reduction was mild (more than 20 mL/100 g/min), and in six of seven animals, frequent SD-like depolarizations turned into terminal depolarization at a later stage of the experiments. Under halothane, MCAO induced severe ischemia (less than 10 mL/100 g/min) and immediate terminal depolarization only in ectosylvian gyrus. In suprasylvian gyrus, residual CBF remained significantly higher (more than 10 mL/100 g/min) than under alpha-chloralose, whereas in marginal gyri, CBF did not differ between groups. Compared with chloralose, the number of transient depolarizations was significantly reduced in marginal gyrus, and in suprasylvian gyrus transient but significantly longer depolarizations than in marginal gyrus were recorded. Except for one animal, transient depolarizations did not turn into terminal depolarization under halothane, and infarct volume reduction was particularly seen in suprasylvian gyrus. We conclude that halothane, the most commonly used anesthetic in studies of experimental brain ischemia, has protective properties, which may depend on both cerebrovascular and electrophysiologic influences.

Anesthetics, Inhalation↗

Profiles of cortical tissue depolarization in cat focal cerebral ischemia in relation to calcium ion homeostasis and nitric oxide production.

Cortical depolarization was investigated in a topographic gradient of ischemic density after 1-hour transient middle cerebral artery occlusion in halothane-anesthetized cats. A laser Doppler flow probe, an ion-selective microelectrode, and a nitric oxide (NO) electrode measured regional CBF (rCBF), direct current (DC) potential, extracellular Ca2+ concentration ([Ca2+]o), and NO concentration in ectosylvian and suprasylvian gyri of nine animals. Recordings revealed 12 of 18 sites with persistent negative shifts of the DC potential, severe rCBF reduction, and a drop of [Ca2+]o characteristic for core regions of focal ischemia. Among these sites, two types were distinguished by further analysis. In Type 1 (n = 5), rapid, negative DC shifts resembled anoxic depolarization as described for complete global ischemia. In this type, ischemia was most severe (8.9 +/- 2.5% of control rCBF), [Ca2+]o dropped fast and deepest (0.48 +/- 0.20 mmol/L), and NO concentration increased transiently (36.1 +/- 24.0 nmol/L at 2.5 minutes), and decreased thereafter. In Type 2 (n = 7), the DC potential fell gradually over the first half of the ischemic episode, rCBF and [Ca2+]o reductions were smaller than in Type 1 (16.2 +/- 8.2%; 0.77 +/- 0.41 mmol/L), and NO increased continuously during ischemia (53.1 +/- 60.4 nmol/L at 60 minutes) suggesting that in this type NO most likely exerts its diverse actions on ischemia-threatened tissue. In the remaining six recording sites, a third type (Type 3) attributable to the ischemic periphery was characterized by minimal DC shifts, mild ischemia (37.2 +/- 13.3%), nonsignificant alterations of [Ca2+]o, but decreased NO concentrations during middle cerebral artery occlusion. Reperfusion returned the various parameters to baseline levels within 1 hour, the recovery of [Ca2+]o and NO concentration being delayed in Type 1. An NO synthase inhibitor (N(G)-nitro-L-arginine, 50 mg/kg intravenously; four animals) abolished NO elevation during ischemia. In conclusion, even in the core of focal cerebral ischemia and reperfusion, different ischemic densities produce different types of cortical tissue manifesting distinctive chronological profiles of depolarization, Ca2+ influx, and NO synthesis.

Animals↗

Advantages and disadvantages of various access routes in sonographic diagnosis of dysplasia and luxation in the infant hip.

The purpose of the study is to clarify the access route that is most suitable in sonography of the infant hip for diagnosis of luxation and dysplasia. The process of luxation gives rise to deformation of the hyaline cartilaginous preformed part as well as the osseous acetabulum. These deformations are located in the superoposterior part of the acetabulum. With a posterior exposure only the luxated head of the femur can be imaged sonographically, but the dysplastic acetabulum cannot be diagnosed. In anterior exposures the pathological changes in the superior part of the acetabulum are detected poorly or not at all. The lateral exposure can image all deformations of the acetabulum. The strictly standardized lateral sonographic exposure is hence recommended using a standardized plane of measurement.

Acetabulum↗

Insulin-mediated secretion of ecdysteroids from mosquito ovaries and molecular cloning of the insulin receptor homologue from ovaries of bloodfed Aedes aegypti.

The reproductive cycle of female mosquitoes is activated by ingestion of blood from vertebrate hosts. Shortly after feeding, neurohormones are released from the brain neurosecretory system and stimulate the ovaries to secrete ecdysteroids, which are necessary for vitellogenesis by the fat body. Because bombyxins, which are insulin-like peptides, stimulate ecdysteriodgenesis in silkworm larvae, we tested porcine insulin and found that it activates ecdysteroidogenesis and protein synthesis in ovaries isolated from unfed mosquitoes. To further characterize the regulation of ecdysteroidogenesis in female mosquitoes, we cloned the mosquito insulin receptor (MIR) homologue from ovarian mRNA. The sequence of the extracellular domain shows moderate homologies with vertebrate and Drosophila insulin receptor homologues, as well as with the insulin receptor-related receptor, the latter being an "orphan" receptor with an unknown function. In the intracellular domain, high homologies are observed, particularly in those subdomains that are responsible for ATP binding and kinase activity. Northern blot analysis of MIR demonstrated a highly specific expression in ovaries, and cloning experiments indicated its presence in the brain. Recombinant MIR extracts from a baculovirus expression system contained high constitutive kinase activity in the presence of manganese or magnesium. Activation was independent of a ligand. SDS-gel analysis suggested that the recombinant receptor was not post-translationally processed into an alpha- and beta-subunit as was expected from a putative cleavage signal. Enzymatic properties of the proreceptor are presented: the Km for ATP was between 15 and 50 microM in the presence of a synthetic substrate: maximal kinase activity to 100-fold over basic activity was reached in the presence of 1 mM manganese. Stimulation of key oogenic processes by porcine insulin and identification of a MIR indicate that insulin-like neurohormones may have an important regulatory role in mosquito oogenesis.

Aedes↗