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Biomedical subjects

C Schneider

Publications and source records attributed to C Schneider.

At least 109 records · Page 6Linked to original sources
PubMed2000

The growth suppressing gas1 product is a GPI-linked protein.

Growth arrest specific (gas) 1 gene product is expressed in non-transformed fibroblasts in response to stimuli driving cells into Go phase. Gas1 has been demonstrated to inhibit cell proliferation when over-expressed in proliferating fibroblasts. This activity depends on a function of the p53 protein independent of its transactivating ability. To better define the pathway leading from Gas1, which is located on the plasma membrane, to p53, we have undertaken a detailed characterization of its topology. We demonstrate that the protein undergoes cotranslational modifications in the endoplasmic reticulum, consisting of signal peptide cleavage, N-linked glycosylation and glycosyl-phosphatidylinositol anchor addition. Immunoelectron microscopy shows that, in its mature form, Gas1 is randomly distributed over the outer leaflet of the plasma membrane and that upon antibody-induced clustering it relocalizes to caveolae.

3T3 Cells↗
PubMed2000

Analysis of the domain requirement in Gas1 growth suppressing activity.

The product of the growth arrest specific gene, gas1, is a membrane-associated protein which activates a p53-dependent growth suppression signalling pathway. We have shown that Gas1 is linked to the plasma membrane through a glycosyl-phosphatidylinositol (GPI) anchor. Several GPI-anchored protein have been identified as part of receptor complexes either as co-receptors or as membrane bound ligands. In this report, we characterize the Gas1 domains required for its growth suppression function and demonstrate the dispensability of Gas1 GPI anchor.

3T3 Cells↗
PubMed2000

Large vasodilatations in skeletal muscle of resting conscious dogs and their contribution to blood pressure variability.

Large (up to +400 %) transient ( approximately 20 s) increases of blood flow were observed in the external iliac arteries of resting conscious dogs (n = 10) in the absence of major alerting or muscular activity. At the same time arterial pressure (AP) fell slightly while heart rate (HR) rose. The vasodilatations were resistant to atropine, ganglionic, beta-adrenergic and NO-synthase inhibition, but were suppressed by spinal or general anaesthesia. Vasodilatations of similar appearance were elicited by an alerting sound; these were abolished by atropine. The spontaneous vasodilatations occurred simultaneously and their magnitudes were well correlated between both legs, but were not correlated to the amount of concomitant activation of the surface electromyogram. The duration of this activation almost never outlasted 10 s. The reactive hyperaemia observed after a total occlusion of the artery even for 16 s was not large enough to explain the size of the spontaneous vasodilatations. Occlusion during peak flow of the vasodilatations did not affect the size of the reactive hyperaemia. Spectral analysis made separately for data segments with and without vasodilatation revealed that the vasodilatations substantially enhanced the variability of AP and HR at frequencies below approximately 0.1 Hz. In conclusion, large coordinated skeletal muscle vasodilatations were identified in resting conscious dogs, which are initiated neurally, but not by sympathetic-cholinergic or nitroxidergic fibres and which do not show any clear correlation to muscular contraction. The vasodilatations substantially affect the regulation of skeletal muscle blood flow and explain a significant portion of AP and HR variability.

Animals↗
PubMed2000

Cell-cycle regulation of the p53-inducible gene B99.

B99 is a p53-inducible gene whose accumulation upon p53 activation is restricted to late S/G2 cells. Here we have analyzed B99 regulation during the cell cycle in murine cells with or without functional p53. We report that B99 accumulates in late S/G2 phase, is phosphorylated in mitosis, and disappears in G1 phase, regardless of the status of p53. As a complement to this observation, we show that B99 is not induced by p53 in quiescent cells. Therefore, B99 expression is modulated both by cell-cycle regulatory mechanisms and by p53, and p53 can increase the cellular levels of B99 only during the window of the cell cycle when it is normally expressed. On the basis of these observations we rename B99 Gtse-1 (G-two- and S-phase-expressed).

3T3 Cells↗
PubMed2000

Cloning, chromosome mapping and functional characterization of a human homologue of murine gtse-1 (B99) gene.

Murine Gtse-1 (G(2) and S phase expressed protein), previously named B99, is a wt-p53 inducible gene that encodes a microtubule-localized protein which is able to induce G(2)/M phase accumulation when ectopically expressed. Here we report the cloning and characterization of a new cDNA (GTSE-1) encoding a human homologue of the mouse Gtse-1 protein. Chromosome mapping of mouse and human genes assigned Gtse-1 to chromosome 15 and GTSE-1 to chromosome 22q13.2-q13.3 in a region with conserved synteny to that where Gtse-1 mapped. Analysis of the genomic structure revealed that GTSE-1 contains at least 11 exons and 10 introns, spanning approximately 33kb of genomic DNA. Similar to murine Gtse-1, the product of GTSE-1 localized to the microtubules, was able to delay G(2)/M progression when ectopically expressed and was cell cycle regulated. Taken together, these results indicate GTSE-1 as the human functional homologue of murine Gtse-1.

Amino Acid Sequence↗
PubMed2000

Synthesis and applications of stereospecifically (3)H-labeled arachidonic acids as mechanistic probes for lipoxygenase and cyclooxygenase catalysis.

Stereospecifically (3)H-labeled substrates are useful tools in studying the mechanism of hydrogen abstractions involved in the oxygenation of polyunsaturated fatty acids. Here, we describe modified methods for the synthesis of arachidonic acids labeled with a single chiral tritium on the methylene groups at carbons 10 or 13. The appropriate starting material is a ketooctadecanoic acid which is prepared from an unsaturated C18 fatty acid precursor or by total synthesis. The (3)H label is introduced by NaB(3)H(4) reduction and the resulting tritiated hydroxy fatty acid then is tosylated, separated into the enantiomers by chiral phase HPLC, and subsequently transformed into stearic acids. A variety of stereospecifically labeled unsaturated fatty acids are obtained using literature methods of microbial transformation with the fungus Saprolegnia parasitica. Two applications are described: (i) In incubations of [10S-(3)H]- and [10R-(3)H]arachidonic acids in human psoriatic scales we show that a 12R-lipoxygenase accounts not only for synthesis of the major product 12R-HETE, but it contributes also, through subsequent isomerization, to the minor amounts of 12S-HETE. (ii) The [10R-(3)H]- and [10S-(3)H]arachidonic acids were also used to demonstrate that prostaglandin ring formation by cyclooxygenases does not involve carbocation formation at C-10 of arachidonic acid as was hypothesized recently.

Animals↗
PubMed2000

Proximal myotonic myopathy: evidence for anticipation in families with linkage to chromosome 3q.

OBJECTIVE: To investigate anticipation in proximal myotonic myopathy (PROMM). BACKGROUND: PROMM is a recently described autosomal dominantly inherited disorder similar to but distinct from myotonic dystrophy (DM). DM belongs to the group of inherited disorders with anticipation caused by an unstable trinucleotide repeat expansion. In PROMM, no mutation has been identified, although PROMM has recently been mapped to a gene locus on chromosome 3q. METHODS: We investigated 10 German families with the PROMM phenotype and linkage to chromosome 3q. We based our analysis of anticipation on the age of disease onset. Anticipation was assumed if the offspring had first symptoms earlier in life than his or her affected parent. For statistical analysis Independence Estimating Equations (IEE) and a Monte-Carlo bootstrap were used. RESULTS: In 27 affected living parent-offspring pairs from these 10 families, the mean difference of disease onset was 18.8 years with either statistical analysis (p < 10-14 and p < 10-15). The mean disease onset interval in years was greater in father-offspring as compared to the mother-offspring pairs (p < 0.05; IEE). CONCLUSION: Our findings suggest the occurrence of anticipation in parent-offspring pairs from families with the PROMM phenotype and linkage to chromosome 3q. The different disease onset intervals in mother-offspring and father-offspring pairs could indicate a mild parent-of-origin effect. These observations are compatible with the suggestion that PROMM, like DM, may be a trinucleotide repeat associated disorder. In contrast to DM, anticipation in PROMM is milder, a congenital form does not seem to occur, and fertility does not appear to be affected.

Adolescent↗
PubMed2000

Experimental autoimmune myositis in the lewis rat: lack of spontaneous T-cell apoptosis and therapeutic response to glucocorticosteroid application.

Recently, it has been shown that inflammatory T-cells in human idiopathic myositis only very rarely undergo spontaneous apoptosis. The animal model of experimental autoimmune myositis (EAM) in the Lewis rat was chosen to investigate whether similar findings hold true in rat muscle and if glucocorticosteroids act by induction of T-cell apoptosis in inflammatory lesions. The rate of spontaneous T-cell apoptosis in rat EAM was low, even in muscle specimens with extensive inflammation. After intravenous glucocorticosteroid pulse therapy we found a dramatic increase in the rate of apoptotic T-cells in the inflamed muscles. Up to 50% of these apoptotic T-cells were CD8 positive apoptotic T-cells. T-cell apoptosis was significantly lower in similarly inflamed muscle specimens of the control group. We suggest that glucocorticosteroids induce apoptosis of endomysial T-cells in human idiopathic polymyositis. Glucocorticosteroid-induced apoptosis may be a candidate mechanism in the termination of inflammatory activity.

Animals↗
PubMed2000

Shared and unique roles of CAP23 and GAP43 in actin regulation, neurite outgrowth, and anatomical plasticity.

CAP23 is a major cortical cytoskeleton-associated and calmodulin binding protein that is widely and abundantly expressed during development, maintained in selected brain structures in the adult, and reinduced during nerve regeneration. Overexpression of CAP23 in adult neurons of transgenic mice promotes nerve sprouting, but the role of this protein in process outgrowth was not clear. Here, we show that CAP23 is functionally related to GAP43, and plays a critical role to regulate nerve sprouting and the actin cytoskeleton. Knockout mice lacking CAP23 exhibited a pronounced and complex phenotype, including a defect to produce stimulus-induced nerve sprouting at the adult neuromuscular junction. This sprouting deficit was rescued by transgenic overexpression of either CAP23 or GAP43 in adult motoneurons. Knockin mice expressing GAP43 instead of CAP23 were essentially normal, indicating that, although these proteins do not share homologous sequences, GAP43 can functionally substitute for CAP23 in vivo. Cultured sensory neurons lacking CAP23 exhibited striking alterations in neurite outgrowth that were phenocopied by low doses of cytochalasin D. A detailed analysis of such cultures revealed common and unique functions of CAP23 and GAP43 on the actin cytoskeleton and neurite outgrowth. The results provide compelling experimental evidence for the notion that CAP23 and GAP43 are functionally related intrinsic determinants of anatomical plasticity, and suggest that these proteins function by locally promoting subplasmalemmal actin cytoskeleton accumulation.

Actins↗
PubMed2000

Early and selective loss of neuromuscular synapse subtypes with low sprouting competence in motoneuron diseases.

The addition or loss of synapses in response to changes in activity, disease, or aging is a major aspect of nervous system plasticity in the adult. The mechanisms that affect the turnover and maintenance of synapses in the adult are poorly understood and are difficult to investigate in the brain. Here, we exploited a unique anatomical arrangement in the neuromuscular system to determine whether subtypes of synapses can differ in anatomical plasticity and vulnerability. In three genetic mouse models of motoneuron disease of diverse origin and severity, we observed a gradual and selective loss of synaptic connections that begun long before the onset of clinical deficits and correlated with the timing of disease progression. A subgroup of fast-type (fast-fatiguable) neuromuscular synapses was highly vulnerable and was lost very early on. In contrast, slow-type synapses resisted up to the terminal phase of the disease. Muscle-specific differences were also evident. Similar selective losses were detected in aged mice. These selective vulnerability properties of synapses coincided with hitherto unrecognized major differences in stimulus-induced anatomical plasticity that could also be revealed in healthy mice. Using paralysis and/or growth-associated protein 43 overexpression to induce synaptic sprouting, we found that slow-type, disease-resistant synapses were particularly plastic. In contrast, fast-type synapses with the highest vulnerability failed to exhibit any stimulus-induced change. The results reveal pronounced subtype specificity in the anatomical plasticity and susceptibility to loss of neuromuscular synapses and suggest that degenerative motoneuron diseases involve a common early pathway of selective and progressive synaptic weakening also associated with aging.

Aging↗
PubMed2000

Spatial requirements for 15-(R)-hydroxy-5Z,8Z,11Z, 13E-eicosatetraenoic acid synthesis within the cyclooxygenase active site of murine COX-2. Why acetylated COX-1 does not synthesize 15-(R)-hete.

The two isoforms of cyclooxygenase, COX-1 and COX-2, are acetylated by aspirin at Ser-530 and Ser-516, respectively, in the cyclooxygenase active site. Acetylated COX-2 is essentially a lipoxygenase, making 15-(R)-hydroxyeicosatetraenoic acid (15-HETE) and 11-(R)-hydroxyeicosatetraenoic acid (11-HETE), whereas acetylated COX-1 is unable to oxidize arachidonic acid to any products. Because the COX isoforms are structurally similar and share approximately 60% amino acid identity, we postulated that differences within the cyclooxygenase active sites must account for the inability of acetylated COX-1 to make 11- and 15-HETE. Residues Val-434, Arg-513, and Val-523 were predicted by comparison of the COX-1 and -2 crystal structures to account for spatial and flexibility differences observed between the COX isoforms. Site-directed mutagenesis of Val-434, Arg-513, and Val-523 in mouse COX-2 to their COX-1 equivalents resulted in abrogation of 11- and 15-HETE production after aspirin treatment, confirming the hypothesis that these residues are the major isoform selectivity determinants regulating HETE production. The ability of aspirin-treated R513H mCOX-2 to make 15-HETE, although in reduced amounts, indicates that this residue is not an alternate binding site for the carboxylate of arachidonate and that it is not the only specificity determinant regulating HETE production. Further experiments were undertaken to ascertain whether the steric bulk imparted by the acetyl moiety on Ser-530 prevented the omega-end of arachidonic acid from binding within the top channel cavity in mCOX-2. Site-directed mutagenesis was performed to change Val-228, which resides at the junction of the main cyclooxygenase channel and the top channel, and Gly-533, which is in the top channel. Both V228F and G533A produced wild type-like product profiles, but, upon acetylation, neither was able to make HETE products. This suggests that arachidonic acid orientates in a L-shaped binding configuration in the production of both prostaglandin and HETE products.

Acetylation↗
PubMed2000

Stereospecificity of hydrogen abstraction in the conversion of arachidonic acid to 15R-HETE by aspirin-treated cyclooxygenase-2. Implications for the alignment of substrate in the active site.

The initial and rate-limiting step in prostaglandin biosynthesis is stereoselective removal of the pro-S hydrogen from the 13-carbon of arachidonic acid. This is followed by oxygenation at C-11, formation of the five-membered ring, and a second oxygenation at C-15 to yield the endoperoxide product, prostaglandin G(2). Aspirin treatment of cyclooxygenase-2 is known to acetylate an active site serine, block prostaglandin biosynthesis, and give 15R-hydroxyeicosatetraenoic acid (15R-HETE) as the only product. 15R-HETE and prostaglandins have opposite stereoconfigurations of the 15-hydroxyl. To understand the changes that lead to 15R-HETE synthesis in aspirin-treated COX-2, we employed pro-R- and pro-S-labeled [13-(3)H]arachidonic acids to investigate the selectivity of the initial hydrogen abstraction. Remarkably, aspirin-treated COX-2 formed 15R-HETE with removal of the pro-S hydrogen at C-13 (3-9% retention of pro-S tritium label), the same stereoselectivity as in the formation of prostaglandins by native cyclooxygenase. To account for this result and the change in oxygenase specificity, we suggest that the bulky serine acetyl group forces a realignment of the omega end of the arachidonic acid carbon chain. This can rationalize abstraction of the C-13 pro-S hydrogen, the blocking of prostaglandin synthesis, and the formation of 15R-HETE as the sole enzymatic product.

Acetylation↗
PubMed2000

Lack of association between serotonin transporter gene promoter variants and autistic disorder in two ethnically distinct samples.

Family-based studies performed to date provide conflicting evidence of linkage/association between autistic disorder and either the "short" [Cook et al., 1997: Mol Psychiatry 2:247-250] or the "long" [Klauck et al., 1997: Hum Mol Genet 6:2233-2238] allele of a polymorphic repeat located in the serotonin transporter (5-HTT) gene promoter region, affecting 5-HTT gene expression [Lesch et al., 1996: Science 274:1527-1531]. The present study was designed to assess linkage and linkage disequilibrium in two new ethnically distinct samples of families with primary autistic probands. The 5-HTT promoter repeat was genotyped in 54 singleton families collected in Italy and in 32 singleton and 5 multiplex families collected in the U.S.A., yielding a total sample of 98 trios. Linkage/association between 5-HTT gene promoter alleles and autistic disorder was assessed using the transmission/disequilibrium test (TDT) and the haplotype-based haplotype relative risk (HHRR). Both the Italian and the American samples, either singly or combined, displayed no evidence of linkage/association between 5-HTT gene promoter alleles and autistic disorder. Our findings do not support prominent contributions of 5-HTT gene variants to the pathogenesis of idiopathic infantile autism. Heterogeneity in pathogenetic mechanisms underlying the disease may require that linkage/association studies be targeted toward patient subgroups isolated on the basis of specific biochemical markers, such as serotonin (5-HT) blood levels. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:123-127, 2000.

Adolescent↗
PubMed2000

p53 is involved in the p120E4F-mediated growth arrest.

Control of cell growth and division by the p53 tumor suppressor protein requires its abilities to transactivate and repress specific target genes and to associate in complex with other proteins. Here we demonstrate that p53 binds to the E1A-regulated transcription factor p120E4F, a transcriptional repressor of the adenovirus E4 promoter. The interaction involves carboxy-terminal half of p120E4F and sequences located at the end of the sequence-specific DNA-binding domain of p53. Ectopic expression of p120E4F leads to a block of cell proliferation in several human and murine cell lines and this effect requires the association with wild-type (wt) p53. Although p120E4F can also bind to mutant p53, the growth suppression induced by overexpression of the protein is severely reduced in a cell line that contains mutant p53. These data suggest that p120E4F may represent an important element within the complex network of p53 checkpoint functions.

3T3 Cells↗
PubMed2000

Laparoscopic abdominoperineal resection: early postoperative results of a prospective study involving 116 patients. The Laparoscopic Colorectal Surgery Study Group.

PURPOSE: Although laparoscopic colorectal surgery is attracting ever more attention, its use for curative treatment of colorectal carcinoma in particular continues to be controversial. The present study was an attempt to analyze the results of the perioperative course, oncologic quality, and preliminary long-term results. METHOD: The data considered here were collected within the framework of a prospective, observational study initiated on August 1, 1995, and involving a total of 18 institutions in Germany and Austria. At the end of three years, the results are now being presented selectively, i.e., focusing only on abdominoperineal resection. RESULTS: A total of 116 patients underwent laparoscopic abdominoperineal resections, 98 (84.5 percent) of which were performed with curative intent. The mean operating time was 226 (confidence interval, 140-365) minutes. Seven patients (6 percent) experienced an intraoperative complication, which in more than one-half of the cases was a vascular injury involving the presacral venous plexus; the conversion rate was 3.4 percent. Postoperatively, 40 patients developed 97 complications--including those of a very minor nature--giving an overall morbidity rate of 34.4 percent. Reoperation in six patients (5.2 percent) had to be performed for an afterbleed in one-half of the cases and ileus in the other one-half. Postoperative mortality was a low 1.7 percent. In most of the curative resections, an oncologically radical operation with high transection of the inferior mesenteric artery and a complete dissection of the pelvis down to the floor was performed. The median number of lymph nodes investigated was 11.5, and there was wide fluctuation in the numbers among the individual institutions. Tumor cell dissemination occurred intraoperatively in five patients. In the meantime, 79 patients (81 percent) underwent at least one follow-up examination, the mean follow-up period being 491 days. Seven patients developed a local recurrence, and a further six patients developed distant metastases. For recurrence-free survival rate, the Kaplan-Meier estimation calculated a probability of 71 percent. CONCLUSION: Not all of the reservations about laparoscopic abdominoperineal resection, in particular with regard to resection with curative intent, have yet been eliminated. The present study does, however, show that a laparoscopic approach can in principle meet oncologic requirements of radicality and, with regard to the postoperative course, is associated with considerable benefits to the patient.

Abdomen↗
PubMed2000

Magnetic resonance pharmacological stress for detecting coronary disease. Comparison with echocardiography.

Stress testing is the cornerstone in the diagnosis of patients with suspected coronary artery disease (CAD). Although exercise ECG remains the primary approach for the detection of ischemia in patients with chest pain syndromes, its sensitivity and specificity is limited and exercise ECG does not provide detailed information about the localisation and extent of CAD. Stress echocardiography has been used for the detection of ischemia for more than a decade and has become an increasingly popular noninvasive method for the detection of CAD. In experienced hands wall motion analysis based on stress echocardiography has proved to be as sensitive and specific for the detection of myocardial ischemia as scintigraphic techniques. Recent technical improvements, namely the availability of ultrafast imaging sequences with a significant reduction of imaging time have initiated several studies which examined the combination of pharmacological stress and magnetic resonance imaging (MRI) for the detection of suspected CAD. The most well developed stress-MRI technique is wall motion imaging during dobutamine stress. This technique is analogous to stress echocardiography, but MRI has the inherent advantages of better resolution, higher reproducibility and true long and short axis imaging with contiguous parallel slices. However, the clinical impact of MRI for the diagnosis of CAD is still low. Further technical developments including real time imaging and a reliable automated quantitative analysis of left ventricular function are required before stress-MRI becomes a serious challenge to stress-echocardiography in the clinical arena. Currently, only a few MRI facilities and physicians are dedicated to pharmacological stress testing with MRI and the future clinical impact of this promising technique will depend on its potential to provide information beyond myocardial function including perfusion, metabolism and coronary anatomy in form of a "one-stop"-shop for the cardiac patient.

Cardiotonic Agents↗