Medizin - Open Access LMU - Teil 07/22

An abnormal ratio between Na+ and K+ conductances seems to be the cause for the depolarization and paralysis of skeletal muscle in primary hypokalemic periodic paralysis. Recently we have shown that the k+ channel opener cromakalim hyperpolarizes mammalian skeletal muscle fibers. Now we have studied the effects of this drug on the twitch force of muscle biopsies from normal and diseased human skeletal muscle. Cromakalim had little effect on the twitch force of normal muscle whereas it strongly improved the contraction force of fibers from patients suffering from hypokalemic periodic paralysis. Recordings of intracellular K+ and Cl- activities in human muscle and isolated rat soleus muscle support the view that cromakalim enhances the membrane K+ conductance (gK+). These data indicate that K+ channel openers may have a beneficial effect in primary hypokalemic periodic paralysis.

The reasons for the resistance to ischaemia of peripheral nerves in diabetics are not well understood. We have now explored whether axonal depolarization underlies this phenomenon, as has previously been proposed. Resistance to ischaemia was determined by the new method of “threshold tracking”. This method revealed an increase in excitability of the peroneal nerve at the popliteal fossa during ischaemia, and a decrease in excitability in the post-ischaemic period. The extent of these alterations in 28 type 1 diabetics without peripheral neuropathy showed a strong correlation with the mean blood glucose concentrations during the last 24 h before examination. To test whether the ischaemic resistance was related to membrane potential, we also measured axonal superexcitability in 11 selected diabetics, since it has been shown that post-spike changes in excitability depend on membrane potential. Changes in excitability of the peroneal nerve were measured in the period between 10 and 30 msec following a conditioning supramaximal compound action potential. Under resting conditions, no differences in the post-spike superexcitability were found between controls and diabetics, despite striking differences in their responses to a 10-min pressure cuff. These observations indicate that membrane depolarization is not involved in the resistance to ischaemia of motor axons in diabetic subjects.

N-terminal (atrial natriuretic factor (ANF) 1-98) and C-terminal (ANF 99-126) fragments of proatrial natriuretic factor (NTA and CTA, respectively) were determined in plasma of healthy subjects adopting different postures and in patients with cirrhosis. Seven healthy subjects were investigated while seated and 30 min after assuming a horizontal position. NTA plasma concentrations increased in subjects in the horizontal position (from 734±250 (SE) fmol/ml to 9021227 fmol/ml; p

1. The vascular actions of atrial natriuretic factor (ANF) have been assessed with other vasoactive agents on the hepatic arterial and portal vascular beds of the anaesthetized dog. 2. Intra-arterial bolus injections of ANF (0.1-50 nmol) caused graded increases in hepatic arterial blood flow representing a vasodilatation of relatively short duration. Vasoconstriction was never observed. 3. The maximum increase in hepatic arterial blood was the same for ANF and isoprenaline (Iso) i.e. approximately 60-70% increase over control flow. 4. On a molar basis, ANF was less potent than Iso although over the higher dose range (10(-9)-10(-7) mol) its vasodilator activity exceeded that of the endogenous vasodilator adrenaline. 5. Intraportal bolus injections (1.0-50 nmol) of ANF did not alter portal inflow resistance since no changes in portal inflow pressure occurred when the portal circuit was perfused at constant inflow volume. 6. This differential action of ANF on the hepatic arterial and portal vascular beds may provide a change in total liver blood flow in favour of the arterial component. 7. ANF, by altering hepatic haemodynamics to favour formation of trans-sinusoidal fluid exchange, may provide a temporary expansion of the extravascular fluid reservoir to buffer any increased venous pressure. However, chronically elevated plasma levels of ANF would encourage the formation of ascitic fluid.

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/5904/1/5904.pdf Gerbes, Alexander L.; McEnroe, G. A.; Vollmar, Angelika M.; Kollenda, Margit C.

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/5907/1/5907.pdf Jüngst, Dieter; Mezger, J.; Xie, Yining; Gerbes, Alexander L. ddc:610, Medizin

In order to assess the effect of food ingestion on splanchnic disposal of human alpha-atrial natriuretic peptide (ANF), hepatic-intestinal removal of ANF was determined before and after a test meal. Hepatic venous and arterial plasma samples were obtained from six subjects, most of whom had only disorders of minor degree. Hepatic blood flow (HBF) increased significantly after meal ingestion (1.10 ± 0.17 [SEM] to 1.51 ± 0.26 L/min, P < .01). Baseline arterial ANF (10.9 ± 3.1 pmol/L) did not change significantly. In contrast, hepatic venous ANF increased after meal intake (5.7 ± 2.0 to 8.4 ± 1.9 pmol/L, P < .05), and accordingly the splanchnic fractional extraction decreased (0.53 ± 0.09 to 0.35 ± 0.08), although this was not statistically significant. Splanchnic clearance of ANF increased from 347 ± 90 mL/min to a maximal value of 615 ± 158 mL/min (P < .05). Splanchnic removal of ANF was 3.0 ± 0.5 pmol/min before and increased to a maximum value (7.1 ± 2.2 pmol/min, P < .05) 35 minutes after ingestion of the meal. Our results showed enhanced splanchnic removal of ANF after food intake. This is due to increased hepatic-intestinal clearance of the peptide consequent on increased splanchnic blood flow, rather than altered fractional extraction of ANF.

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/5993/1/5993.pdf Sauerbruch, Tilman; Kratzer, M.; Gerbes, Alexander L.; Scheurlen, C.; Fischer, G. ddc

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/6052/1/6052.pdf Scherbaum, Werner A.; Heinze, Eberhard; Hauner, Hans; Bornstein, Stefan; Fetzer, A.; Yassin, Nuha; Speck, U.; Glück, Michael; Seißler, Jochen

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/6063/1/Gerbes_Alexander_6063.pdf Vollmar, Angelika M.; Gerbes, Alexander L. ddc:610, Medizin

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/6100/1/6100.pdf Paumgartner, Gustav; Gerbes, Alexander L. ddc:610, Medizin

The effects of the cholinoceptor agonist, carbachol (CCh), were examined in the rat hippocampal slice preparation. Intracellular recordings from CA1 pyramidal neurones revealed that CCh (1–3 μM) inhibited excitatory postsynaptic responses evoked by stimulation of the Schaffer collateral/commissural pathway while, at the same time, direct excitability was enhanced. Extracellularly, CCh produced a concentration-dependent reduction of the amplitude of the field excitatory postsynaptic potential (field EPSP) recorded in the CA1 apical dendritic region. The muscarinic receptor antagonist, pirenzepine, competitively antagonized the effects of CCh on the field EPSP with a pA2 of 7.4. These results confirm earlier reports of a presynaptic inhibitory action of CCh in the hippocampal CA1 region and provide strong evidence that this effect is mediated by muscarinic receptors of the M1 subtype.

1. Intracellular microelectrodes were used to obtain recordings from neurons in layer II/III of rat frontal cortex. A bipolar electrode positioned in layer IV of the neocortex was used to evoke postsynaptic potentials. Graded series of stimulation were employed to selectively activate different classes of postsynaptic responses. The sensitivity of postsynaptic potentials and iontophoretically applied neurotransmitters to the non-N-methyl-D-asparate (NMDA) antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) was examined. 2. As reported previously, low-intensity electrical stimulation of cortical layer IV evoked short-latency early excitatory postsynaptic potentials (eEPSPs) in layer II/III neurons. CNQX reversibly antagonized eEPSPs in a dose-dependent manner. Stimulation at intensities just subthreshold for activation of inhibitory postsynaptic potentials (IPSPs) produced long-latency (10 to 40-ms) EPSPs (late EPSPs or 1EPSPs). CNQX was effective in blocking 1EPSPs. 3. With the use of stimulus intensities at or just below threshold for evoking an action potential, complex synaptic potentials consisting of EPSP-IPSP sequences were observed. Both early, Cl(-)-dependent and late, K(+)-dependent IPSPs were reduced by CNQX. This effect was reversible on washing. This disinhibition could lead to enhanced excitability in the presence of CNQX. 4. Iontophoretic application of quisqualate produced a membrane depolarization with superimposed action potentials, whereas NMDA depolarized the membrane potential and evoked bursts of action potentials. At concentrations up to 5 microM, CNQX selectively antagonized quisqualate responses. NMDA responses were reduced by 10 microM CNQX. D-Serine (0.5-2 mM), an agonist at the glycine regulatory site on the NMDA receptor, reversed the CNQX depression of NMDA responses.

The actions of the phosphodiesterase inhibitor denbufylline on the excitability of hippocampal neurons were investigated by means of extracellular and intracellular recordings. Denbufylline, which has been shown to selectively inhibit a low KM, Ca2+/calmodulin-independent phosphodiesterase isozyme, concentration-dependently increased the amplitude of the extracellularly recorded CAI population spike evoked by electrical stimulation of the Schaffer collateral/commissural pathway. Concentration-response-curves yielded an EC50 for denbufylline of 0.76 M. In comparison, the nonselective phosphodiesterase inhibitor 3-isobutyl-lmethylxanthine (IBMX) also produced an increase in the amplitude of the population spike. From the concentration-response-curve, which was steeper than that of denbufylline, an EC50 for IBMX of 1.04 M was obtained. However, despite their similar EC50 values, denbufylline was found to be significantly more potent at lower concentrations (

The actions of the reductant ascorbic acid on rat neocortical neurons in vitro was investigated by means of intracellular recordings. At a concentration (500 μM), which reduced the magnitude of dopamine degradation in oxygen-saturated saline solutions by about 50%, ascorbic acid reversibly depressed synaptic potentials and enhanced direct excitability of cortical neurons. The latter effect was not reversible within the observation period. Ascorbic acid did not alter membrane potential and input resistance of the neurons. On the basis of our results we conclude that ascorbic acid is not a useful reductant to avoid oxidation of catecholamines in oxygen-saturated solutions used in electrophysiological experiments in vitro.

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/6124/1/Kellerer_Albrecht_6124.pdf Kellerer, Albrecht M.; Ponsel, G.; Wachsmann, M.; Smida, J.; Trutschler, K.; Hieber

Die neue Bewertung der Krebsmortalität der Überlebenden von Hiroshima und Nagasaki führt zu einer erhöhten Risikoschätzung für Strahlen-Kanzerogenese.

The current definitions of microdosimetric and dosimetric quantities use the notion of 'ionizing radiation'. However, this notion is not rigorously defined, and its definition would require the somewhat arbitrary choice of specified energy cut-off values for different types of particles. Instead of choosing fixed cut-off values one can extend the system of definitions by admitting the free selection of a category of types and energies of particles that are taken to be part of the field. In this way one extends the system of dosimetric quantities. Kerma and absorbed dose appear then as special cases of a more general dosimetric quantity, and an analogue to kerma can be obtained for charged particle fields; it is termed cema. A modification that is suitable for electron fields is termed reduced cema.

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/6127/1/6127.pdf Kellerer, Albrecht M. ddc:610, Medizin

The microdosimetric quantities energy imparted, lineal energy, and specific energy are defined with reference to certain volumes but are quantified in terms of frequency distributions of possible values without regard to spatial interrelations. Computer simulations of the patterns of energy deposits seem, therefore, only loosely related to the microdosimetric distributions. In a more general formulation one treats the specific energy and the related microdosimetric quantities as point functions; one deals then with the spatial distribution of their random values and not merely with the frequency of different values. A further extension of the formalism admits reference regions of vanishing size; the inchoate distribution of energy deposits is then the limit case of specific energy. The definitions are related to Matheron's concept of the regularisation of a spatial variable; this is a convolution process that permits a flexible mathematical treatment. One resulting possibility is the definition of specific energy with reference not to the conventional geometry of a sphere or a cylinder but to a disperse region of support. This extension provides distributions of specific energy that are relevant to diffusion or transport processes and it can help to free biophysical models from a one-sided fixation on the concept of geometric targets. The formalism is applied also to the definition of the proximity functions and the related spatial autocorrelation functions.

Applications of the variance-covariance technique are presented that illustrate the potential of the method. The dose mean lineal energy, yD, can be determined in time-varying radiation fields where the fluctuations of the dose rate are substantially in excess of the stochastic fluctuations of the energy imparted. An added advantage is, that yD is little influenced by noise that affects both detectors simultaneously. The variance-covariance method is thus stable with respect to dose rate fluctuations and other temporal variations of a radiation field, and to various influences of noise and electronic artefacts. The dose mean lineal energy obtained at different simulated diameters agrees well with experimental data by other authors and can be determined for small simulated diameters (below 100 nm).

Linear energy transfer, L, is currently used as reference parameter for the quality factor in radiation protection, and this practice is likely to be continued in an impending revision of the quality factor. But the numerical convention is not, at present, actually applied to photon or electron fields; their quality factor is, instead, summarily equated to unity. The current trend of tightened dose limits in radiation protection may create the need for precise computations of quality factors even for photon and electron radiations. Such computations can, however, not be performed in terms of unrestricted linear energy transfer. The reasons for this difficulty are explained, and the formulae for the quality factor as an integral over electron fluence and restricted linear energy transfer are given. A correct energy balance in this and in a variety of linked dosimetric relations requires a modification of the definition of restricted linear energy transfer. For large energy cut-off values the correction is of minor importance, but for small cut-off values - such as those invoked in biophysical considerations - the modification is essential.

A generalised formulation of microdosimetry clarifies the linkages between the spatial distribution of energy deposits, their proximity function, and the specific energy. The role of the proximity function suggest that it may replace, for various purposes, the inchoate distribution. The Fourier transform of the proximity function is the product of the Fourier transform of the inchoate distribution with its conjugate. This operation causes the loss of phase information, and the reconstruction problem - the reconstruction of the inchoate distribution from its proximity function - can, therefore, not be resolved by a mere deconvolution. For any finite point pattern one can, however, show that its proximity function permits, in principle, the reconstruction. Numerical examples with 2-dimensional patterns of up to 30 points have consistently led to unique solutions, apart from reflections. While there is a finite algorithm, it is readily seen that the number of steps becomes excessive when the number of points in the pattern increases. The reconstruction problem can, thus, be solved in principle but not necessarily in practice. A more general approach must thus be based on numerical optimisation. The algorithm starts with an assumed initial point pattern and utilises a suitable measure for the difference between its proximity function and that of the original pattern. Minimising this difference can lead to the original or to a similar pattern. With simple algorithms one obtains convergence only for patterns of few points; but improved optimisation methods are likely to provide more general solutions.

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/6143/1/6143.pdf Paumgartner, Gustav; Gerbes, Alexander L. ddc:610, Medizin

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/6183/1/6183.pdf Müller-Lissner, S.; Riepl, Rudolf L.

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/6196/1/6196.pdf Chmelevsky, D.; Spiess, H.; Kellerer, Albrecht M. ddc:610, Mediz

More than 50 bone sarcomas have occurred among a collective of about 800 patients who had been injected in Germany after World War II with large activities of radium-224 for the intended treatment of bone tuberculosis and ankylosing spondylitis.^In an earlier analysis it was concluded that, at equal mean absorbed doses in the skeleton, patients with longer exposure time had a higher incidence of bone sarcomas.^The previous analysis was based on approximations; in particular, it did not account for the varying times at risk of the individual patients.^In view of the implications of a reverse protraction factor for basic considerations in radiation protection, the need was therefore felt to reevaluate the data from the continued follow-up by more rigorous statistical methods.^A first step of the analysis demonstrates the existence of the reverse dose-rate effect in terms of a suitably constructed rank-order test.^In a second step of the analysis it is concluded that the data are consistent with a linear no-threshold dose dependence under the condition of constant exposure time, while there is a steeper than linear dependence on dose when the exposure times increase proportionally to dose.^A maximum likelihood fit of the data is then performed in terms of a proportional hazards model that includes the individual parameters, dose, treatment duration, and age at treatment.^The fit indicates proportionality of the tumor rates to mean skeletal dose with an added factor (1 + 0.18.tau), where tau is the treatment time in months.^This indicates that a protraction of the injections over 15 months instead of 5 months doubles the risk of bone sarcoma.

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/6225/1/6225.pdf Stellmach, R.; Berghaus, Alexander

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/6244/1/6244.pdf Berghaus, Alexander ddc:610, Medizin 0

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/6261/1/6261.pdf Berghaus, Alexander; Gundlach, P. ddc:610, Medizin

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/6262/1/6262.pdf Berghaus, Alexander; Tausch-Treml, R. ddc:610, Medizin

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/6446/1/6446.pdf Eisenmenger, Wolfgang ddc:610, Medizin

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/6543/1/6543.pdf Jüngst, Dieter

Although the relative contribution of different immune effector functions to clearing tissues of cytomegalovirus is controversial, the contribution of CD8+ T lymphocytes has generally been accepted as essential. In this report, we show that under certain conditions the CD8+ T-lymphocyte subset can be dispensable for clearance of cytomegalovirus. Mice depleted of the CD8+ T-lymphocyte subset eliminated infectious virus with a clearance kinetics similar to that of normal mice. Adoptive transfer studies revealed that the limitation of virus spread required the cooperation between the CD4+ subset and other cells. Comparison between protective functions generated in fully immunocompetent and in CD8- mice demonstrated that elimination of the CD8+ subset before infection altered the quality of the antiviral immune response. The compensatory protective activity gained by CD4+ cells in CD8- mice was absent in normal mice recovering from virus infection.

The regulation of murine cytomegalovirus early (E) gene expression was studied in the cell line B25, which is stably transfected with the immediate-early ie1/ie3 gene complex. Infection of B25 cells in the presence of the protein synthesis inhibitor cycloheximide resulted in the expression of some E genes, whereas for the expression of other E genes prior protein synthesis was still mandatory, thus showing differences in the expression requirements of individual E genes. Transcription unit e1, a member of the E genes induced by immediate-early products of the ie1/ie3 gene complex, was characterized. It is located between map units 0.709 and 0.721 of the genome of murine cytomegalovirus strain Smith. A 2.6-kilobase RNA specified in this region is spliced from three exons of 912, 177, and 1,007 or 1,020 nucleotides, which are separated by introns of 93 and 326 nucleotides. The second AUG located in the first exon 119 nucleotides downstream of the 5' cap site is followed by an open reading frame of 990 nucleotides. The predicted polypeptide of 330 amino acids has a calculated molecular mass of 36.4 kilodaltons. Transfection with e1 revealed three antigenically related proteins of 36, 37, and 38 kilodaltons; these proteins probably represent differently modified forms of the predicted protein. These three proteins are phosphorylated and are associated with intranuclear inclusion bodies. A 33-kilodalton protein also derived from e1 was identified as a product of nonspliced transcripts. Comparison of amino acid sequences revealed homology between the murine cytomegalovirus transcription unit e1 and a human cytomegalovirus E transcription unit.

The properties of transmembrane and soluble transplantation antigens were compared with respect to the induction of tolerance and the selection of the T-cell repertoire. For this purpose, transgenic (H-2b x H-2d)F1 mice were constructed that carry integrated copies of a modified H-2Kk gene resulting in the secretion from various cell types including thymocytes of soluble H-2Kk molecules. Despite the presence of H-2Kk antigen, these mice were still able to generate an H-2Kk-specific T-cell response. This response was comparable to that produced by normal littermates when stimulated with cells expressing membrane H-2Kk in a mixed lymphocyte reaction. In contrast, only transgenic mice failed to generate a cytolytic T-cell response to soluble H-2Kk antigen expressed by recombinant vaccinia virus and presented by the H-2Db molecule. These data imply the presence of two populations of alloreactive cytolytic T cells. A small fraction of T cells recognizes alloantigen as antigenic peptide(s) presented by other major histocompatibility complex class I molecules and tolerance can be induced in this population by soluble alloantigen. The majority of T cells, however, require the whole cell membrane-expressed class I molecule for recognition. This population is not affected by tolerance induction to the soluble major histocompatibility complex class I molecule.

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/6779/1/6779.pdf Jüngst, Dieter; Pauletzki, J.

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/6877/1/6877.pdf Remien, J.; Liebl, B.; Hauck, R.; Emslander, H.-P.; Haen, E.; Langenmayer, Irmgard

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/6999/1/6999.pdf Kaufmann, W.; Winkelmann, W.; Cohen, S.; Hackenberg, E.; Allolio, B.; Reincke, Martin ddc:610, Medizin

We describe a series of 18 patients with an intrasellar mass incidentally discovered by computed tomography or magnetic resonance imaging. The average size of the mass was 13 mm, with a range from 5 to 25 mm. Initial ophthalmologic examination revealed bitemporal hemianopia in 2 patients. Results of routine endocrine testing showed partial hypopituitarism in 5 patients and growth hormone hypersecretion without signs and symptoms of acromegaly in 1 patient. Four patients underwent neurosurgery. Histologically, one chondroid chordoma and three pituitary adenomas were found. In the remaining 14 patients treated conservatively, repeated computed tomography and magnetic resonance imaging revealed no significant change in tumor size at the time of follow-up (median, 22 months). Our results suggest that the "incidentaloma" of the pituitary gland is a benign condition that does not necessarily require neurosurgical intervention.

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/7028/1/7028.pdf Winkelmann, Werner; Allolio, Bruno; Reincke, Martin ddc:610, Medizin

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/7030/1/7030.pdf Allolio, B.; Winkelmann, W.; Reincke, Martin; Arlt, W. ddc:610, Medizin

The pregnancy-specific glycoprotein (PSG) genes encode a group of proteins which are found in large amounts in placenta and maternal serum. In situ hybridization analyses of metaphase chromosomes reveal that all the human pregnancy-specific glycoprotein (PSG) genes are located on the long arm of chromosome 19 (19q13.2–13.3), overlapping the region containing the closely-related carcinoembryonic antigen (CEA) gene subgroup. Higher resolution analyses indicate that the PSG genes are closely linked within an 800kb SacII restriction endonuclease fragment. This has been confirmed through restriction endonuclease mapping and DNA sequence analyses of isolated genomic clones, which show that at least some of these genes are located in very close proximity. Further, these studies have helped to identify a new member of the PSG gene sub-family (PSG7). DNA/RNA hybridization analyses, using gene-specific oligonucleotide probes based on published sequences, showed that five from six PSG genes tested are coordinately transcribed in the placenta. Due to the close proximity of these genes and their coordinated expression pattern, common transcriptional regulatory elements may exist.

The gene encoding the human tumor marker carcinoembryonic antigen (CEA) belongs to a gene family which can be subdivided into the CEA and the pregnancy-specific glycoprotein subgroups. The corresponding proteins are members of the immunoglobulin superfamily, characterized through the presence of one IgV-like domain and a varying number of IgC-like domains. Since the function of the CEA family is not well understood, we decided to establish an animal model in the rat to study its tissue- specific and developmental stage-dependent expression. To this end, we have screened an 18-day rat placenta cDNA library with a recently isolated fragment of a rat CEA-related gene. Two overlapping clones containing the complete coding region for a putative 709 amino acid protein (rnCGM1; Mr = 78,310) have been characterized. In contrast to all members of the human CEA family, this rat CEA-related protein consists of five IgV-like domains and only one IgC-like domain. This novel structure, which has been confirmed at the genomic level might have important functional implications. Due to the rapid evolutionary divergence of the rat and human CEA gene families it is not possible to assign rnCGM1 to its human counterpart. However, the predominant expression of the rnCGM1 gene in the placenta suggests that it could be analogous to one of the human pregnancy-specific glycoprotein genes.

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/7082/1/the_time_course_of_retrograde_transsynaptic_transport_7082.pdf Büttner-Ennever, J. A.; Horn, Anja K. E.

Carcinoembryonic antigen (CEA) is a widely used tumor marker, especially in the surveillance of colonic cancer patients. Although CEA is also present in some normal tissues, it is apparently expressed at higher levels in tumorous tissues than in corresponding normal tissues. As a first step toward analyzing the regulation of expression of CEA at the transcriptional level, we have isolated and characterized a cosmid clone (cosCEA1), which contains the entire coding region of the CEA gene. A close correlation exists between the exon and deduced immunoglobulin-like domain borders. We have determined a cluster of transcriptional starts for CEA and the closely related nonspecific cross-reacting antigen (NCA) gene and have sequenced their putative promoters. Regions of sequence homology are found as far as approximately 500 nucleotides upstream from the translational starts of these genes, but farther upstream they diverge completely. In both cases we were unable to find classic TATA or CAAT boxes at their expected positions. To characterize the CEA and NCA promoters, we carried out transient transfection assays with promoter-indicator gene constructs in the CEA-producing adenocarcinoma cell line SW403, as well as in nonproducing HeLa cells. A CEA gene promoter construct, containing approximately 400 nucleotides upstream from the translational start, showed nine times higher activity in the SW403 than in the HeLa cell line. This indicates that cis-acting sequences which convey cell type-specific expression of the CEA gene are contained within this region.

Mon, 1 Jan 1990 12:00:00 +0100 https://epub.ub.uni-muenchen.de/7127/1/7127.pdf Zimmermann, Wolfgang; Barnett, T.

Immunocytochemical and immunochemical techniques were used to study the expression of the neural cell adhesion molecule (NCAM) by human lung cancer cell lines. Intense surface staining for NCAM was found at light and electron microscopic levels on small cell lung cancer cells. The NCAM polypeptide of Mr 140000 (NCAM 140) was detected by immunoblotting in all of 7 small cell lung cancer cell lines examined and in one out of two of the closely related large cell cancer cell lines: it was not detected in cell lines obtained from one patient with a mesothelioma, in two cases of adenocarcinoma, nor in two cases of squamous cell cancer. In contrast, neuron-specific enolase was found by immunoblotting in all the lung cancer cell lines tested and synaptophysin in all but the adenocarcinoma cell lines. These antigens were localized intracellularly. The specific expression of NCAM 140 by human small and large cell lung carcinomas suggests its potential as a diagnostic marker.

We have analyzed the expression of the intracellular marker protein neuron specific enolase (NSE), synaptophysin (SYN) and of the cell surface marker NCAM (neural cell adhesion molecule) in both normal human hypophysis and in pituitary adenomas in order to explore their potential use as diagnostic tools. All adenomas (4 prolactinomas, 3 growth hormone (GH) producing adenomas and 4 inactive adenomas) showed SYN and NSE immunoreactivity on tissue sections and this was confirmed by immunoblots. NCAM 140 (an isoform of NCAM with molecular mass 140 kDa) was detected by immunoblotting in normal human adenohypophysis, in all GH adenomas, and in three out of four inactive adenomas, but not in prolactinomas. Using highly sensitive techniques, NCAM immunoreactivity was observed by electron microscopy in all adenomas. These data indicate that NCAM 140 is a constituent of the cell surface of endocrine cells in both normal human adenohypophysis and its tumors. Since prolactinomas express very low levels of NCAM 140 compared to other hypophyseal tumors its virtual absence could be used for differential diagnosis. A combined analysis of NCAM, SYN and NSE could be useful to characterize inactive adenomas which are not immunoreactive for pituitary hormones and which may contain no or only low levels of the alpha chain of the glycoprotein hormones.

The intracellular action on exocytosis of botulinim A toxin and constituent chains was studied using permeabilized isolated nerve endings from the rat neural lobe. The release of the neuropeptide vasopressin was measured by radioimmunoassay. In the presence of the reducing agent dithiothreitol, the two-chain form of botulinum A toxin inhibited vasopressin release induced by 10 μM free calcium. Half maximal inhibition was obtained with 15 nM botulinum A toxin. In the absence of the heavy chain the light chain of the toxin strongly inhibited exocytosis with a half maximal effect of 2.5 nM. The inhibitory effects on secretion could be prevented by incubating the light chain with an immune serum against botulinum A toxin. The heavy chain of botulinum A toxin did not affect vasopressin release. However, it prevented the inhibitory effects of the light chain on stimulated exocytosis. It is concluded that botulinum A toxin inhibits the calcium-dependent step leading to exocytosis by interfering with a target present in the isolated and permeabilized nerve terminals. The functional domain of this neurotoxin, which is responsible for the inhibition of vasopressin release, is present in its light chain.

The olfactory bulb of the rat contains chromogranin A at a similar level as the adrenal gland or the hypophysis as revealed by immunoblots. Olfactory chromogranin A also displays the same size as chromogranin A of endocrine cells. In the hippocampus and other brain regions, we could not detect chromogranin A by immunoblotting. In contrast, chromogranin A messenger ribonucleic acid (using S1 nuclease protection assays) was observed in all brain regions examined, including the olfactory bulb. By in situ hybridization histochemistry with a complementary ribonucleic acid probe (280 nucleotides), and by immunocytochemistry, chromogranin A synthesis could be localized to cell bodies of the mitral cell layer, of the external plexiform layer and of the periglomerular region of the olfactory bulb. Immunocytochemically, chromogranin A was also detected in the central projection areas of mitral and tufted cells in the primary olfactory cortex and the anterior amygdaloid area but not in the olfactory glomeruli, where the incoming olfactory nerve fibers of the primary olfactory neurons establish synaptic contacts. Taken together the data show that chromogranin A, following biosynthesis in the perikarya of the mitral and tufted cells, is specifically transported into their axonal terminals but not into their primary dendrites. We propose that the rat olfactory system could serve as a model for the study of chromogranin A regulation and function in neurons.