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Biomedical subjects

E Kremer

Publications and source records attributed to E Kremer.

At least 19 recordsLinked to original sources
PubMed2000

[1,3-Bis(diphenylphosphino)propane]trichlorooxorhenium(V).

Trichlorooxo[1,3-propanediylbis(diphenylphosphine)-P,P ']rhenium(V), [ReCl(3)O(C(27)H(26)P(2))], crystallizes with four formula units per unit cell. The crystal structure consists of neutral complexes of [ReOCl(3)(dppp)] [dppp is 1,3-bis(diphenylphosphino)propane] packed by H.pi-ring interactions. The Re atom is octahedrally coordinated to the oxo anion, three Cl atoms and two P atoms from the dppp ligand. The six-membered ring formed by the bidentate dppp ligand and the rhenium metal centre is in a chair conformation. The title compound is an intermediate in the synthesis of bis(dppp) complexes of rhenium.

Crystallography, X-Ray↗
PubMed2000

Sacrocaudal afferents induce rhythmic efferent bursting in isolated spinal cords of neonatal rats.

The ability of mammalian spinal cords to generate rhythmic motor behavior in nonlimb moving segments was examined in isolated spinal cords of neonatal rats. Stimulation of sacrocaudal afferents (SCA) induced alternating left-right bursts in lumbosacral efferents and in tail muscles. On each side of the tail, flexors, extensors, and abductors were coactive during each cycle of activity. This rhythm originated mainly in the sacrocaudal region because it persisted in sacrocaudal segments after surgical removal of the thoracolumbar cord. Sacrocaudal commissural pathways were sufficient to maintain the left-right alternation of lumbar efferent bursts, because their timing was unaltered after a complete thoracolumbar hemisection. The lumbar rhythm originated in part from sacrocaudal activity ascending in lateral and ventrolateral funiculi, because efferent bursts in rostral lumbar segments were nearly abolished on a particular side by lesions of the lateral quadrant of the cord at the L(4)-L(5) junction. Intracellular recordings from S(2)-S(3) motoneurons, obtained during the rhythm, revealed the presence of phasic oscillations of membrane potential superimposed on a tonic depolarization. Bursts of spikes occurred on the depolarizing phases of the oscillation. Between these bursts the membrane input conductance increased, and hyperpolarizing drive potentials were revealed. The inhibitory drive and the decreased input resistance coincided with contralateral efferent bursts, suggesting that crossed pathways controlled it. Our studies indicate that pattern generators are not restricted to limb-moving spinal segments and suggest that regional specializations of pattern-generating circuitry and their associated interneurons are responsible for the different motor patterns produced by the mammalian spinal cord.

Action Potentials↗
PubMed1999

Adenovirus-mediated gene transfer in dog prostate: a preclinical study of a relevant model system for gene therapy of human prostatic cancer.

This present study evaluates the potential of adenovirus-mediated gene transfer (AMGT) to the prostate of normal laboratory beagles. Many morphological and histological similarities can be noted between dog and human prostate. Moreover, dogs can spontaneously develop prostate cancer with a clinical and biological outcome identical to that in man. Firstly we showed the capacity of human adenovirus to infect canine prostatic cells in vitro. Secondly, we injected transrectally in the dogs' prostates 2x10(9) plaque forming units of a first generation recombinant adenovirus vector harboring the reporter gene beta-galactosidase (AdRSVbetagal). Seven days after the adenoviral delivery, we observed expression of the transgene in both prostates, and exclusively in epithelial cells. Despite a cellular and a humoral immune response, the infusion appeared safe, since the dogs had no fever and presented no urinary symptoms. This study constitutes the first evaluation of AMGT in dog prostate and provides a basis for gene therapy treatment of prostate carcinoma-bearing patients.

Journal Article↗
PubMed1998

GABA-receptor-independent dorsal root afferents depolarization in the neonatal rat spinal cord.

Dorsal root afferent depolarization and antidromic firing were studied in isolated spinal cords of neonatal rats. Spontaneous firing accompanied by occasional bursts could be recorded from most dorsal roots in the majority of the cords. The afferent bursts were enhanced after elevation of the extracellular potassium concentration ([K+]e) by 1-2 mM. More substantial afferent bursts were produced when the cords were isolated with intact brain stems. Rhythmic afferent bursts could be recorded from dorsal roots in some of the cords during motor rhythm induced by bath-applied serotonin and N-methyl--aspartate (NMDA). Bilaterally synchronous afferent bursts were produced in pairs of dorsal roots after replacing the NaCl in the perfusate with sodium-2-hydroxyethansulfonate or after application of the gamma-aminobutyric acid-A (GABAA) receptor antagonist bicuculline with or without serotonin (5-HT) and NMDA. Antidromic afferent bursts also could be elicited under these conditions by stimulation of adjacent dorsal roots, ventrolateral funiculus axons, or ventral white commissural (VWC) fibers. The antidromic bursts were superimposed on prolonged dorsal root potentials (DRPs) and accompanied by a prolonged increase in intraspinal afferent excitability. Surgical manipulations of the cord revealed that afferent firing in the presence of bicuculline persisted in the hemicords after hemisection and still was observed after removal of their ventral horns. Cutting the VWC throughout its length did not perturb the bilateral synchronicity of the discharge. These findings suggest that the activity of dorsal horn neurons is sufficient to produce the discharge and that the bilateral synchronicity can be maintained by cross connectivity that is relayed from side to side dorsal to the VWC. Antagonists of GABAB, 5-HT2/5-HT1C, or glutamate metabotropic group II and III receptors could not abolish afferent depolarization in the presence of bicuculline. Depolarization comparable in amplitude to DRPs, could be produced in tetrodotoxin-treated cords by elevation of [K+]e to the levels reported to develop in the neonatal rat spinal cord in response to dorsal root stimulation. A mechanism involving potassium transients produced by neuronal activity therefore is suggested to be the major cause of the GABA-independent afferent depolarization reported in our study. Possible implications of potassium transients in the developing and the adult mammalian spinal cord are discussed.

Action Potentials↗
PubMed1997

Prevention of bleomycin-induced pulmonary fibrosis after adenovirus-mediated transfer of the bacterial bleomycin resistance gene.

A serious limitation in the use of the DNA-cleaving, antitumoral-antibiotic, bleomycin during chemotherapy is pulmonary toxicity. Lung injury induced by bleomycin is characterized by an increased deposition of interstitial extracellular matrix proteins in the alveolar wall that compromises respiratory function. Several drugs have been tested in animal models to prevent the pulmonary toxicity of bleomycin, but have not led to a useful clinical treatment because of their adverse effects on other tissues. We have shown that transgenic mice expressing Streptoalloteichus hindustanus (Sh) ble bleomycin resistance protein in pulmonary epithelial cells in the lungs are protected against bleomycin-induced toxicity in lungs. In the present study, we used intranasal administration by adenovirus-mediated gene transfer of the bleomycin resistance Sh ble gene to mouse lung for prevention of bleomycin-induced pulmonary fibrosis. We constructed recombinant adenoviruses Ad.CMVble and Ad.RSVble harboring the bleomycin resistance Sh ble gene under the control of the cytomegalovirus early promoter and the Rous sarcoma virus early promoter, respectively. Transgene expression was detected in epithelia of conducting airways and alveolar septa by immunostaining with a rabbit polyclonal antibody directed against the bleomycin resistance protein and persisted for the duration of drug treatment; i.e., up to 17 d. No toxic effect was seen in adenovirus-treated mice. Pretreatment of mice with Ad.CMVble or Ad.RSVble completely prevented collagen deposition 42-133 d after bleomycin treatment, as measured by lung OH-proline content. Histologic studies indicated that there was little or no lung injury in the adenovirus/bleomycin-treated mice compared with the bleomycin-treated mice. These observations may lead to new approaches for the prevention of bleomycin-induced pulmonary fibrosis.

Acetyltransferases↗
PubMed1997

Localization of the spinal network associated with generation of hindlimb locomotion in the neonatal rat and organization of its transverse coupling system.

The segmental organization of the hindlimb locomotor pattern generators and the coordination of rhythmic motor activity were studied in isolated spinal cords of the neonatal rat. All lumbar segments and many thoracic and sacral segments of the cord exhibited an alternating left-right rhythm in the presence of serotonin (5-HT) and N-methyl-D-aspartate (NDMA). Other thoracic segments exhibited a synchronized left-right rhythm or an irregular bursting activity. Transection of the cord at the thoracolumbar or lumbosacral junction abolished the rhythmicity of nonlumbar segments and had no affect on the rhythmicity of lumbar segments. A fast alternating rhythm persisted in rostral lumbar segments after transection of the cord at mid-L3. A much slower alternating rhythm was found in the detached caudal lumbar segments after elevation of the NMDA concentration. These findings suggest that neurogenesis of hindlimb locomotion is not restricted to L1/L2, and that the lumbar pattern generators exhibited rostrocaudal specialization. An alternating left-right rhythm persisted in lumbar cords of midsagittally split preparations that were kept with either L1, L2, L3, or L4 as the only bilaterally intact segment. An alternating rhythm persisted also in preparations that were midsagittally split up to T13-T12, or down to L4. Extension of these lesions led to a bilaterally synchronous rhythm or to left-right independent rhythms in the lumbar cord. These results indicated that the transverse coupling system in the caudal-thoracic and lumbar segments in specialized and that left-right alternation in the lumbar cord can be carried out by the cross connectivity, which is relayed at least through the T12-L4 segments. Bath application of the glycine receptor antagonist strychnine, or the gamma-aminobutyric acid-A (GABAA) receptor blocker bicuculline, induced in the presence of NMDA and 5-HT a bilaterally synchronous rhythm in any intact or detached segment of the cord and in midsagittally split preparations with few bilaterally intact upper thoracic or lower sacral segments. A strychnine-resistant left-right alternating rhythm was found in the presence of 5-HT and NMDA in preparations that were treated with the non-NMDA receptor blocker 6-cyano-7-nitroquinoxaline (CNQX) before and during the application of strychnine. Subsequent washout of CNQX immediately induced a bilateral synchronous rhythm. These results suggest that the phase relation between the hemicords during the rhythm is determined by a dynamic interplay between the excitatory and inhibitory cross connectivity, and that this interplay can be modulated experimentally. Local application of strychnine to L2 kept bilaterally intact in midsagittally split preparations perturbed but did not completely block the alternating pattern of the rhythm induced by 5-HT and NMDA. Local application of bicuculline under the same conditions prolonged the cycle time and had no effect on left-right alternation. These results, together with those described above, suggest that left-right alternation is mediated mainly by strychnine-sensitive glycine receptors with possible contribution of strychnine-resistant glycine receptors and/or GABAA receptors.

6-Cyano-7-nitroquinoxaline-2,3-dione↗
PubMed1996

Strategies for cancer gene therapy using adenoviral vectors.

Modification of tumor cells using gene transfer either to enhance host immunity or to act directly on tumor cells is being intensively studied in animal models. Remarkable results have yielded to approved clinical protocols in the treatment of cancer patients using this approach. Several methods of gene delivery have been developed. This article is particularly devoted to the interest of the use of adenoviral vectors in the different strategies of cancer gene therapy.

Adenoviridae↗
PubMed1995

Conformational analyses on soluble and surface bound osteopontin.

Immunohistology of calvarial sections revealed that staining with monoclonal anti-osteopontin antibodies (clone MPIIIB10) is minimal unless sections are first treated with EDTA. In contrast, following treatment of sections with EDTA, strong staining of mineralizing osteoid areas and osteoblast-like cells was noted (Fig. 1B). Immunostaining for osteopontin appeared to be specific in that controls which substituted rabbit IgG or normal mouse ascites fluid for monoclonal antibody, or which omitted monoclonal antibody uniformly gave background results (Fig. 1C). In an effort to circumvent problems of antibody accessibility we examined the immunoreactivity of OP when adsorbed to plastic and hydroxyapatite surfaces. Although OP bound to plastic surfaces is reactive with MPIIIB10 antibodies, OP adsorbed to hydroxyapatite crystal surfaces is not recognized by these antibodies as assessed by two detection methods. These results demonstrate that most or all of OP bound to hydroxyapatite exhibits a different conformation than when bound to plastic surfaces. On the basis of immunohistologic results with calvarial sections, we suggest that the conformation of native OP in bone and of isolated OP adsorbed to hydroxyapatite may be similar. Finally, solution circular dichroism and Fourier-transformed infrared spectroscopic studies indicate that the conformation of bone OP is dependent upon its concentration, and, secondarily to the presence or absence of calcium ion. With both spectroscopic methods, addition of calcium appeared to increase the extent of disordered structure. We suggest that these findings support our hypothesis that bone matrix proteins exhibit a different conformation when adsorbed on hydroxyapatite crystal surfaces. Assumption of a more organized secondary structure in concentrated OP solutions (i.e., 15 mg/ml) is consistent with these results in that local concentrations of OP within a semisolid matrix may approach or exceed levels used here.

Animals↗
PubMed1995

Inhibitory effects of pentoxifylline on LPS-induced leukocyte adhesion and macromolecular extravasation in the microcirculation.

Pentoxifylline (PTX) has been shown to combat effectively endotoxin induced symptoms of shock or inflammation by reducing both leukocyte activation and endogenous cytokine formation. With regard to blood perfusion, inflammation is defined as a local reaction to injury of the living microvasculature and its content. Leukocyte margination, rolling, adhesion, and emigration is mediated by adhesion molecules along the endothelium of postcapillary venules and is considered to be an important step in the inflammatory response. Changes in the vascular integrity can be estimated in terms of increased extravasation of macromolecules. Using intravital microscopy with the help of an analogous video image processing system we measured the effect of PTX on lipopolysaccharide (LPS, 15 mg/kg i.v.) induced leukocyte adhesion and extravasation of FITC-rat serum albumin (FITC-RSA) in rat mesenteric venules. The changes in vascular permeability correlates significantly (r = 0.75) with a locally increased number of adherent leukocytes. PTX significantly inhibits both leukocyte adhesion and extravasation of FITC-RSA dose dependently. Our results indicate that PTX effectively preserves vascular integrity in the microcirculation by acting primarily on LPS-induced leukocyte adhesion.

Animals↗
PubMed1994

Implications of FRA16A structure for the mechanism of chromosomal fragile site genesis.

Fragile sites are chemically induced nonstaining gaps in chromosomes. Different fragile sites vary in frequency in the population and in the chemistry of their induction. DNA sequences encompassing and including the rare, autosomal, folate-sensitive fragile site, FRA16A, were isolated by positional cloning. The molecular basis of FRA16A was found to be expansion of a normally polymorphic p(CCG)n repeat. This repeat was adjacent to a CpG island that was methylated in fragile site-expressing individuals. The FRA16A locus in individuals who do not express the fragile site is not a site of DNA methylation (imprinting), which suggests that the methylation associated with fragile sites may be a consequence and not a cause of their genesis.

Alleles↗
PubMed1994

99mTc-ADP: a potential agent for in vivo tumor detection.

The possibility of using 99mTc-labelled nucleotides as tumour seeking agents has been proposed by different research groups. We have recently reported the preparation of a 99mTc-ADP complex with a high radiochemical purity (> 95%), good in vitro and in vivo stability and promising biodistribution results when injected in mice bearing spontaneous mammary adenocarcinomas. Here we report the results of further investigations in animals with spontaneous neoplastic processes, including whole-body autoradiography in mice (20 minutes and 60 minutes post injection) and gamma-camera imaging studies in Wistar rats. Dynamic studies (up to 45 minutes) and static images (up to 18 hours) were acquired to determine the pharmacokinetics of 99mTc-ADP and the tumour/muscle and tumour/blood ratios. Blood-pool studies were also performed as a control. Tumours were visualized by autoradiography as was to be expected from the biodistribution studies. Dynamic studies showed a rapid blood clearance and a behaviour that fitted to a tricompartimental model. Radioactivity was rapidly taken up by the kidneys and excreted in the urine. No evidence of in vivo instability of the complex was observed. Tumour uptake reached the maximum values after 20 minutes post-injection. Tumour/blood and tumour/muscle ratios improved over time, enhancing tumour visualization. The best images were obtained after 3 hours post injection. In summary, our studies suggest that 99mTc-ADP is a promising radiopharmaceutical for tumour diagnosis.

Adenosine Diphosphate↗
PubMed1993

A cytokine receptor gene cluster in the X-Y pseudoautosomal region?

The receptors for interleukin-3 (IL-3), IL-5, and granulocyte-macrophage colony-stimulating factor (GM-CSF) are heterodimers comprised of ligand specific alpha chains and a common beta chain. The genes encoding the IL-5 receptor alpha chain and the common beta chain reside on chromosome 3 and 22 respectively, while the GM-CSF receptor alpha chain gene (CSF2RA) has been mapped to the pseudoautosomal region (PAR) of the sex chromosomes, which is a 2.6-Mb stretch of homologous sequence at the tips of the short arms within which a single obligatory recombination occurs during male meiosis. We have mapped the gene encoding the IL-3 receptor alpha chain (IL3RA) to the sex chromosomes by polymerase chain reaction (PCR) analysis of human-mouse or human-chinese hamster cell hybrids, and to Yp13.3 and Xp22.3 using fluorescence in situ hybridization. To explore the possibility that IL3RA is located within the pseudoautosomal region we screened the Centre d'Etude du Polymorphisme Humain (CEPH) pedigrees for an informative-restriction fragment-length polymorphism (RFLP) that showed male meiotic recombination. Two informative CEPH pedigrees were identified that displayed this phenomenon, confirming the psuedoautosomal location of IL3RA. Using long-range restriction mapping we have found that IL3RA maps to the same 190-kb restriction fragment as CSF2RA, suggesting that a cytokine receptor gene cluster may reside in the PAR.

Base Sequence↗
PubMed1992

Evidence of founder chromosomes in fragile X syndrome.

The mutation responsible for fragile X syndrome and myotonic dystrophy involves the amplification of a simple trinucleotide repeat sequence, which increases in successive generations of affected pedigrees accounting for increasing penetrance of both disorders. This common molecular basis suggests that the two diseases may share other genetic features, but whereas myotonic dystrophy exhibits a significant founder chromosome effect, fragile X syndrome apparently has a very high mutation frequency. By haplotype analysis of microsatellite markers which flank the fragile X unstable element, we have uncovered evidence of founder chromosomes of the fragile X 'mutation'. Disorders caused by heritable unstable elements may therefore exhibit common genetic properties including anticipation and founder chromosomes.

Base Sequence↗
PubMed1992

Fragile-X syndrome: unique genetics of the heritable unstable element.

The fragile site at Xq27.3 is an unstable microsatellite repeat, p(CCG)n. In fragile-X syndrome pedigrees, this sequence exhibits variable amplification, the length of which correlates with fragile-site expression. There is a direct relationship between increased p(CCG)n copy number and propensity for instability: individuals having large amplifications exhibit somatic variation due to increased instability. The instability of the p(CCG)n repeat, when transmitted through affected pedigrees, explains the unusual segregation patterns of fragile-X phenotype, referred to as the Sherman paradox. All individuals of fragile-X genotype were found (where testing was possible) to have a parent with amplified p(CCG)n repeat, indicating that few, if any, cases of fragile-X syndrome are not familial.

Blotting, Southern↗
PubMed1991

Hereditary unstable DNA: a new explanation for some old genetic questions?

Fragile X syndrome, associated with the fragile X chromosome, is the most common cause of familial mental retardation. The condition is characterised by a heritable DNA sequence that consists of an abnormal number of CCG repeats, and which is unstable in both mitosis and meiosis. We suggest that such heritable unstable DNA sequences could be present in other parts of the genome and that these might explain a number of genetic events that are not well understood in terms of classic genetic mechanisms. Such poorly explained observations include anticipation, incomplete penetrance, variable expression, and possibly imprinting, variegation, and multifactorial inheritance.

Chromosome Fragility↗
PubMed1991

Fragile X genotype characterized by an unstable region of DNA.

DNA sequences have been located at the fragile X site by in situ hybridization and by the mapping of breakpoints in two somatic cell hybrids that were constructed to break at the fragile site. These hybrids were found to have breakpoints in a common 5-kilobase Eco RI restriction fragment. When this fragment was used as a probe on the chromosomal DNA of normal and fragile X genotype individuals, alterations in the mobility of the sequences detected by the probe were found only in fragile X genotype DNA. These sequences were of an increased size in all fragile X individuals and varied within families, indicating that the region was unstable. This probe provides a means with which to analyze fragile X pedigrees and is a diagnostic reagent for the fragile X genotype.

Chromosome Mapping↗
PubMed1991

Fragile X syndrome: genetic localisation by linkage mapping of two microsatellite repeats FRAXAC1 and FRAXAC2 which immediately flank the fragile site.

We report the genetic localisation of the fragile site at Xq27.3 associated with fragile X syndrome. The position of the fragile site within the multipoint linkage map was determined using two polymorphic microsatellite AC repeat markers FRAXAC1 and FRAXAC2. These markers were physically located within 10 kilobases and on either side of the p(CCG)n repeat responsible for the fragile site. FRAXAC1 has five alleles with heterozygosity of 44% and is in strong linkage disequilibrium with FRAXAC2 which has eight alleles and a heterozygosity of 71%. No recombination was observed either between these markers in 40 normal CEPH pedigrees or with the fragile X in affected pedigrees. These markers provide the means for accurate diagnosis of the fragile X genotype in families by rapid polymerase chain reaction analysis and were used to position the fragile X within the multipoint map of the X chromosome to a position 3.7 cM distal to DXS297 and 1.2 cM proximal to DXS296.

Base Sequence↗