Showing papers by "University of Düsseldorf published in 1989"

On the mechanism of NO release from sydnonimines.

Abstract: The vasodilator and antiaggregatory properties of sydnonimines like SIN-1 are thought to be due to their marked stimulatory action on soluble guanylate cyclase. Enzyme activation and consecutive cyclic GMP accumulation is mediated by the liberation of nitric oxide (NO) from the open-ring A forms of sydnonimines. The purpose of the present study was to investigate the mechanism of NO release from sydnonimines in direct comparison to their stimulatory effect at the target enzyme, soluble guanylate cyclase. All sydnonimines tested were found to spontaneously liberate NO, the rate of which closely correlated with the extent of enzyme activation. NO release occurred nonlinearly with time and became maximal at high sydnonimine concentration. The in vitro stability of the A forms neither correlated with the measured rate of NO release nor with enzyme activation, indicating that a direct stimulation of guanylate cyclase by the A forms is rather unlikely. Besides NO, all sydnonimines generated NO2- and NO3- at a nearly equimolar rate. The addition of cysteine induced a marked shift from NO3- to NO2- with a small reduction in NO release, which is paralleled by a weak rightward shift of the EC50 at the guanylate cyclase. All tested sydnonimines were found to consume molecular oxygen at rates that closely corresponded to the measured rates of NO formation. By a molar comparison, the amounts of consumed oxygen are clearly higher, as would be expected for the oxidative conversion of NO to NO2- and NO3-. Oxygen seems to be additionally involved in the induction of NO formation while being converted to superoxide (O2-). In accordance with an autocatalytic process, O2- further enhances sydnonimine decomposition, since in the presence of superoxide dismutase (SOD) the rate of SIN-1C and NO2-/NO3- formation from SIN-1A was reduced, whereas the rate of NO liberation seemingly increased. O2- has, however, no influence on the rate of hydrolysis of SIN-1 to SIN-1A. At the level of guanylate cyclase, the presence of SOD induced a leftward shift of the concentration-response curve to SIN-1, in agreement with an enhancement of efficacy of NO by blocking the NO-scavenging effect of O2-. An additional O2- generation markedly enhanced SIN-1A decomposition to NO2-/NO3- and reduced the apparent rate of NO formation. We conclude from our results that oxygen plays a key role in the decomposition of sydnonimines and thus in the formation of NO as their pharmacodynamically active principle. Oxygen attack most probably occurs by one-electron abstraction from the A form of the respective sydnonimine compound.(ABSTRACT TRUNCATED AT 400 WORDS)

Dehydrogenases for the synthesis of chiral compounds

Abstract: Redox reactions are important steps in the metabolism and energy conversion of living cells. Besides the enzymes necessary, a number of coenzymes are involved in such reactions: ferrodoxins, lipoic acid, NAD(H), NADP(H), flavins and cytochromes. The coenzymes differ in their redox potential, in the binding constants and the mode of regeneration [1]. NADH/NADPH are the most frequently encountered coenzymes. In general they dissociate easily and need a second reaction with another metabolite for regeneration. These properties are one of the means by which nature directs the flow of intermediates in response to biosynthetic needs. Because of the spectral properties of the coenzyme moiety, dehydrogenases have been extensively studied in the past [2] and have found widespread applications in clinical and food analysis [3].

The UG paradox in L2 acquisition

Abstract: There is a considerable amount of recent evidence that stable principles of Universal Grammar (UG) are available to adult second language (L2) learners in structuring their intuitions about the target language grammar. In contrast, however, there is also evidence from the acquisition of word order, agreement and negation in German that there are substantial differences between first language (L1) and L2 learners. In our view, these differences are due to UG principles guiding L1, but not L2 acquisition. We will show that alternative ways of accounting for the L1/L2 differences are not successful. Finally we will deal with the question of how our view can be reconciled with the idea that L2 learners can use UG principles to some extent in the evaluation of target sentences.

Antibodies against the human papillomavirus type 16 early proteins in human sera: correlation of anti-E7 reactivity with cervical cancer.

Abstract: By Western blot technique, 519 samples of human sera were tested for the presence of antibodies to the human papillomavirus (HPV) type 16 proteins E4 and E7 that had been expressed in Escherichia coli as fusion proteins. Sera were obtained from patients attending the University hospitals for reasons unrelated to HPV infections (controls), from patients with HPV-associated lesions, as well as from patients suffering from cervical cancer. Within the control population, 18.1% of them had antibodies that reacted with the E4 protein, and 3.9% of them had antibodies that reacted with the E7 protein. No sex-specific difference in the antibody prevalence was observed. The highest proportion of anti-E4 antibody-positive individuals (40.7%) was observed in the age group between 11 and 20 years. The frequency of anti-E4-positive sera was threefold higher in patients with HPV-associated genital lesions than that in age-matched controls. Antibodies against the HPV16 E7 protein were found 14 times more frequently in patients with cervical cancer, compared with age- and sex-matched controls (P less than .00001). From these data, we concluded that anti-E4 antibodies may be correlated with virus replication and that anti-E7 antibodies may represent a marker for cervical cancer development.

Temperature-gradient gel electrophoresis of nucleic acids: analysis of conformational transitions, sequence variations, and protein-nucleic acid interactions.

Abstract: Temperature-gradient gel electrophoresis (TGGE) is applied to analyze conformational transitions and sequence variations of nucleic acids and protein-nucleic acid interactions. A linear and highly reproducible temperature-gradient is established perpendicular or parallel to the direction of the electrophoresis. The instrument consists of an electrically insulated metal plate, which is heated at one edge and cooled at the other edge by two thermostating baths and is used as an ancillary device for commercial horizontal gel electrophoresis instruments. Biopolymers are separated in TGGE according to size, shape and thermal stability of their conformational transitions. If the temperature-gradient is established perpendicular to the electrophoresis, monomolecular conformational transitions of nucleic acids show up as continuous transition curves; strand-separation leads to discontinuous transitions. In the studies on viroid RNA it was shown that natural circular viroid RNA undergoes one highly cooperative transition detected by TGGE as a drastic retardation in mobility. Oligomeric replication intermediates of viroids exhibit coexisting structures which could not be detected by any other technique. Double-stranded satellite RNA from cucumber mosaic virus is a mixture of sequence variants, all of which have the identical length of 335 nucleotides. In TGGE six different strains were resolved. Sequence variants of viroids were analyzed by hybridizing viroid RNA to (-)strand viroid RNA transcripts from viroid cDNA clones. Sequence variations lead to mismatches in the double strands and thereby to a shift of the transition curve to lower temperature. Mutations in plasmids, particularly in cloned inserts, were detected by mixing plasmids of two different clones, linearizing, denaturing, renaturing, and searching for shifts in the transition curves, which are generated by mismatch-formation during the renaturation of (+)- and (-)strands from different clones. Examples are given for different viroid clones and HIV-clones from one and the same patient. In another example, clones with point mutations from site-directed mutagenesis are analyzed and selected by TGGE. TGGE is also applied to study the effect of amino acid exchanges in the Tet repressor from E. coli on the thermal stability of the repressor and on the mode of binding of the repressor to the operator DNA. The results are discussed under the aspect that TGGE may be applied as routine analytical laboratory procedure.

Cerebral asymmetry: MR planimetry of the human planum temporale.

Abstract: A method is described for the imaging and planimetric measurement of the planum temporale (PT) using computer reformations from three-dimensional fast low angle shot magnetic resonance (MR) data. The procedure allows the determination of size and left-right asymmetry of this area on the posterior supratemporal surface, which has been assumed by previous investigators to be a structural correlate of speech lateralization. To establish a consistent method for future studies, special effort was invested in the elaboration of reliable imaging criteria for the borders of the PT. The authors report MR planimetric measurements of the PT in 10 human cadaver brains performed independently and in a "blinded" fashion by two observers. A high degree of interobserver agreement was found, demonstrating the reliability of the MR criteria for PT delineation. The planimetric data correlated with those from subsequent anatomic measurements following section of the same specimens. The left-right ratios of PT size were almost the same with both methods (mean 1.9). Magnetic resonance planimetry of the PT is a noninvasive, reliable, and quantitative procedure to determine gross cerebral asymmetry in the posterior speech area.

Sensorimotor disturbances in patients with lesions of the parietal cortex

Abstract: Somatosensory and motor disturbances of hand function were examined in 9 patients with parietal lobe lesions. A quantitative score was used for the elaboration of sensorimotor profiles displaying the relative degree of functional impairment. In patients with anterior parietal lobe lesions somaesthesis was clearly more disturbed than motor function. Simple aspects of somaesthesis (surface sensibility, two-point discrimination, position sense) were disturbed to about the same degree as complex somatosensory (tactile recognition) tasks. On the other hand, patients with lesions of the posterior parietal lobe showed preferential impairment of complex somatosensory and motor functions (exploratory and manipulative finger movements). In 4 patients, analysis of motor behaviour by means of an optoelectronic system showed that reaching, formation of hand aperture and target acquisition were less disturbed than manipulative behaviour. Finger movement trajectories during dynamic digital palpation of objects were grossly deranged in the patients with posterior parietal damage. The temporal characteristics of the finger movements during active touch were completely destroyed. This leads to a breakdown of the finely tuned digital scanning process required for the sequential sampling of mechanoreceptive information. Remarkably, these patients could produce the exploratory finger movements imitatively. The motor disability of the parietal hand does not lie in the loss of the kinetic memory to perform these movements, but in the loss of their evocation by appropriate sensory stimuli. This deficit is not due to a lack of somatosensory information because that may be relatively well preserved. It is concluded that the motor disturbance in posterior parietal lobe disease lies essentially in the conception and execution of the spatiotemporal movement patterns necessary to bring those receptors into action which would normally provide the information about tactile objects. This illustrates the intricate mutual dependence of the spatiotemporal organization of receptor activation by movement and of the formation of movement trajectories on the basis of adequate sensory processing.

Macrophage infiltration precedes and is a prerequisite for lymphocytic insulitis in pancreatic islets of pre-diabetic BB rats

Abstract: We have analysed whether infiltration of macrophages and lymphocyte subtypes into pancreatic islets of diabetes prone BB rats occurs at random or whether insulitis requires a specific sequence of events. Serial sections from more than 700 islets of diabetes prone BB rats (70–150 days of age) were analysed for infiltrating immunocytes and expression of major histocompatibility complex antigens by 4–11 different monoclonal antibodies. In parallel, electron microscopy was performed in a fraction of islets. Part of the animals had been treated with macrophage toxic silica particles. A specific non-random sequence of events was identified and 4 stages of islet inflammation were recognised. Stages 1 a and 1 b are defined by macrophage (ED1+, W3/25+. Ox3/6/17+, ED2−) infiltration and concomitant enhanced major histocompatibility complex class I antigen expression initially at one pole or at the periphery of islets. T-, NK- and B-lymphocytes are absent (<1 cell per mean islet section). In stage 2, more macrophages are infiltrating and concomitantly Ox19+-T-lymphocytes and Ox8+-granular (NK-) lymphocytes are observed. In stage 3, additional massive infiltration of Ox12+-B-lymphocytes is noted. Silica treatment of BB rats largely prevented macrophage infiltration. Concomitantly islets were free of lymphocytes. Thus, macrophage infiltration clearly precedes T- and NK-lymphocyte and later B-lymphocyte infiltration. Lymphocytes do not infiltrate islets in the absence of prior macrophage invasion.

Photoinhibition at chilling temperature : Fluorescence characteristics of unhardened and cold-acclimated spinach leaves.

Abstract: The effects of moderate light at chilling temperature on the photosynthesis of unhardened (acclimated to +18° C) and hardened (cold-acclimated) spinach (Spinacea oleracea L.) leaves were studied by means of fluorescence-induction measurements at 20° C and 77K and by determination of quantum yield of O2 evolution. Exposure to 550 μmol photons·m(-2)·s(-1) at +4° C induced a strong photoinhibition in the unhardened leaves within a few hours. Photoinhibition manifested by a decline in quantum yield was characterized by an increase in initial fluorescence (F o) and a decrease in variable fluorescence (F v) and in the ratio of variable to maximum fluorescence (F V/F M), both at 77K and 20° C. The decline in quantum yield was more closely related to the decrease in the F V/F M ratio measured at 20° C, as compared with F V/F M at 77K. Quenching of the variable fluorescence of photosystem II was accompanied by a decline in photosystem-I fluorescence at 77K, indicating increased thermal de-excitation of pigments as the main consequence of the light treatment. All these changes detected in fluorescence parameters as well as in the quantum yield of O2 evolution were fully reversible within 1-3 h at a higher temperature in low light. The fast recovery led us to the view that this photoinhibition represents a regulatory mechanism protecting the photosynthetic apparatus from the adverse effects of excess light by increasing thermal energy dissipation. Long-term cold acclimation probably enforces other protective mechanisms, as the hardened leaves were insensitive to the same light treatment that induced strong inhibition of photosynthesis in unhardened leaves.

Effects of dietary long-chain polyunsaturated fatty acids on the essential fatty acid status of premature infants

Abstract: The effect of different diets on the percentage content of long-chain polyunsaturated fatty acids (LCP; metabolites of linoleic and alpha-linolenic acids) in plasma lipids was studied in 29 premature infants on days 4 and 21 of life. Eleven infants were fed human milk which supplies LCP (1.7% of the fatty acids), 10 a commercially available milk formula without LCP, and 8 a new formula enriched with LCP of the omega-6 and the omega-3 series (0.5% LCP). LCP values in plasma lipids remained stable during the observation period in infants fed human milk. In contrast, LCP decreased markedly in plasma lipids of infants fed the conventional formula. Since the precursor fatty acids linoleic and alpha-linolenic acids were high in their diet and plasma, this finding indicates that premature infants have a limited capacity for LCP biosynthesis and may require their dietary supplementation. Infants fed the LCP enriched formula had significantly higher LCP proportions in plasma lipids than infants given the conventional formula, but less than infants fed human milk. Our results demonstrate that small concentrations of dietary LCP have marked effects on plasma lipid composition, particularly on phospholipids, suggesting that dietary LCP are preferentially channelled into structural lipids. We conclude that the essential fatty acid status of formula-fed premature infants can be improved by a supplementation of omega-6- and omega-3-LCP.

Cerebral cortical localization: application and validation of the proportional grid system in MR imaging.

Abstract: The Talairach stereotactic proportional grid method for telencephalic localization was applied to magnetic resonance imaging of 30 brain hemispheres of healthy adult volunteers. Using sagittal images, outlines of the following characteristic principal sulci were identified on the brain surface to validate the method: callosal sulcus, parietooccipital sulcus, marginal sulcus, superomedial portion of central and postcentral sulcus, and posterior sylvian fissure. The maximal variation zones of sulcus location obtained by superim-position of the individual grid data on a standard proportional grid were similar to those originally reported by Talairach from pneumoencephalographic studies. However, the central sulcus, which was only studied postmortem by Talairach, was located 0.5–1 cm more posteriorly in the present study. Magnetic resonance also provided more detailed data on the considerable variation of the terminal parts of the sylvian fissure and on the well-known left-right asymmetries. The left sylvian fissure extended farther posteriorly in 11 of 15 brains. With the exception of this posterior perisylvian region, the proportional grid proved to be valid for indirect cerebral cortical localization in the regions studied.

Spontaneous cytotoxicity of macrophages against pancreatic islet cells.

Abstract: Activated peritoneal macrophages were found to lyse syngeneic [3H]leucine-labeled pancreatic islet cells or rat insulinoma cells after 15 h of coculture at 37 degrees C Lysis was verified by electron microscopic analysis Islet cell lysis was dependent on the T:E ratio and was comparable with P815 and L929 tumor cells used as targets The cytotoxic activity was localized in the adherent fraction of Corynebacterium parvum activated peritoneal cells and was destroyed by incubation of cells with macrophage-toxic silica particles Syngeneic thyrocytes and hepatocytes were found to be resistant to the cytolytic action of activated macrophages It has been shown previously that macrophages contribute to pancreatic islet inflammation The present in vitro analysis demonstrates that macrophages can function as effector cells in islet destruction

Comparison of the renal effects of angiotensin converting enzyme inhibitor and calcium antagonist in hypertensive type 2 (non-insulin-dependent) diabetic patients with microalbuminuria: a randomised controlled trial.

Abstract: Seven of eight hypertensive Type 2 (non-insulin-dependent) diabetic patients with microalbuminuria completed a randomised crossover trial to compare the renal effects of angiotensin converting enzyme inhibitor (enalapril) and calcium antagonist (nicardipine). Four-week fixed oral maintenance dosages of enalapril (10–20 mg/day) and nicardipine (60–120 mg/day) significantly (p<0.05) lowered the systolic and diastolic blood pressures without altering renal blood flow, glomerular filtration rate and filtration fraction. Both drugs significantly reduced (p<0.05) urinary albumin excretion rate and fractional clearance of albumin to similar extents. Total renal vascular resistance decreased significantly by nicardipine (p<0.05) and non-significantly by enalapril. Plasma osmotic pressure, plasma aldosterone concentration, total serum protein concentration, serum electrolytes and HbA1c remained unchanged by these drugs, whereas plasma renin activity was significantly higher (p<0.05) in the enalapril than in the control and nicardipine phases. These results suggest that both drugs have similar renal function preserving effects with a concomitant hypotensive action in hypertensive Type 2 diabetic patients with microalbuminuria, and that the angiotensin converting enzyme inhibitor may not have advantageous renal effects when compared to the calcium antagonist and vice versa. Both drugs might be useful for treatment of high blood pressure in hypertensive diabetic patients, if long-term studies of these drugs can be shown to benefit the patients over other conventional antihypertensive therapies.

Imaging of viroids in nuclei from tomato leaf tissue by in situ hybridization and confocal laser scanning microscopy.

Abstract: The intracellular localization of viroids has been investigated by viroid-specific in situ hybridization and analysis by digital microscopy of the distribution of the fluorescent hybridization signals. Isolated nuclei from green leaf tissue of tomato plants infected with potato spindle tuber viroid (PSTVd) were bound to microscope slides, fixed with formaldehyde and hybridized with biotinylated transcripts of cloned PSTVd cDNA. The bound probe was detected with lissamine--rhodamine conjugated streptavidin. Nucleoli were identified by immunofluorescence using the monoclonal antibody Bv96 and a secondary FITC-conjugated antibody. In plants infected with either a lethal or an intermediate PSTVd strain, the highest intensity of fluorescence that arose from hybridization with the probe specific for the viroid (+)strand was found in the nucleoli, confirming results of previous fractionation studies. A similar distribution was found for (-)strand replication intermediates of PSTVd using specific (+)strand transcripts as hybridization probes. In order to determine if viroids are located at the surface or in the interior of the nucleoli, the distribution of the fluorescence hybridization signals was studied with a confocal laser scanning microscope (CLSM). It was shown by three-dimensional reconstruction that viroids are neither restricted to the surface of the nucleoli nor to a peripheral zone, but are instead homogeneously distributed throughout the nucleolus. The functional implications of the intranucleolar location of viroids and their replication intermediates are discussed with respect to proposed mechanisms of viroid replication and pathogenesis.

Induction of glia-derived nexin after lesion of a peripheral nerve.

Abstract: GLIA-derived nexin1 (GDN), also known as protease nexin I2,3, is a serine protease inhibitor of deduced relative molecular mass 41,700, identified in conditioned media of glioma cells by its neurite-promoting activity4. GDN can promote neurite outgrowth in vitro from neuroblastoma cells5,6, sympathetic neurons7 and hippocampal neurons (L. Farmer et al., manuscript in preparation). In vivo, GDN is constitutively expressed in all parts of the olfactory system8, where axonal regeneration and neurogenesis occur continuously throughout life. This observation indicates that GDN could be important for axonal regeneration in vivo. To investigate this possibility, we have taken advantage of the fact that damage to nerves in the peripheral nervous system leads to their regeneration, whereas in the central nervous system no such regeneration can occur. Here we report that after lesion of the rat sciatic nerve there is a large transient increase in the amount of GDN messenger RNA and of released GDN. The cells showing GDN immunoreactivity are mainly localized distal to the lesion site. These results further support the suggestion that GDN is important for axonal regeneration in vivo, and indicate that protease inhibitors could have a role in Wallerian degeneration and peripheral nerve regeneration.

Long-term safety, efficacy and side-effects of continuous subcutaneous insulin infusion treatment for type 1 (insulin-dependent) diabetes mellitus: a one centre experience.

Abstract: A follow-up study of 116 Type 1 (insulin-dependent) diabetic patients on long-term continuous subcutaneous insulin infusion was conducted after 4.5 +/- 0.2 years. The average HbA1c-value of these patients decreased by 1% to 6.7 +/- 0.1% during this observation period. Typical side effects of continuous subcutaneous insulin infusion such as skin inflammation at the catheter insertion site occurred with similar frequency as has been reported previously by other authors. Diabetic ketoacidosis (0.14 per patient year) and disabling hypoglycaemia (0.1 per patient year, including 0.05 hypoglycaemic coma per patient-year) occurred at substantially lower rates than in other comparable studies with Type 1 diabetic patients at a similar degree of metabolic control. Subgroup evaluation suggested that a normal (less than 5.6%) HbA1c-value at follow-up was associated with increased incidence of disabling hypoglycaemia, whereas poor metabolic control (HbA1c greater than 7.5%) was associated with increased rates of skin complications and hospital treatment for ketoacidosis. Thus, under the policies of this diabetes centre, continuous subcutaneous insulin infusion has proved to be beneficial to a large proportion of experienced adult Type 1 diabetic patients, who voluntarily had opted for, and continued with, this particular mode of insulin treatment.

Immunotoxicology: suppressive and stimulatory effects of drugs and environmental chemicals on the immune system - A discussion

Abstract: The fundamental characteristic of the adaptive immune system which has evolved in the vertebrates is the ability to recognise, and subsequently destroy, "foreign", and potentially harmful, antigens. The selective advantage which the immune system confers is the capacity to resist infectious, and possibly malignant, disease. It has been apparent for many years that individuals in whom immune function is impaired, due either to a congenital defect or to other factors such as treatment with certain immunosuppressive drugs, exhibit an increased susceptibility to infection and, in some cases, an elevated risk of developing at least some forms of malignancy. There is an increasing awareness from rodent studies that a variety of drugs and environmental chemicals have the potential to unintentionally impair components of the immune system. Risk assessment, based upon data from chemically induced changes in one or more parameters of immune function, is, however, dependent upon a knowledge of the functional reserve of the immune system. One of the objectives of the meeting from which this report derives was to examine what sources of information are available, and what experimental protocols can be employed, to permit accurate evaluation of immunological reserve. Although, under normal circumstances, the immune system selectively and specifically recognises foreign antigen, it is clear that the potential to recognise "self" is present and that in certain circumstances this potential is realised. Antibodies directed against normal tissue antigens have been shown to be associated with, and in some instances the presumptive cause of, "autoimmune" disease. There is a growing list of drugs and chemicals which are capable of eliciting autoantibodies and pathological autoimmune reactions. A second purpose of this meeting and of this report was to review the current state of knowledge regarding drug- and chemical-induced autoimmunity.

Fat content and fatty acid composition of infant formulas

Abstract: Fat content and fatty acid composition of 25 commercial infant formulas sold in the Federal Republic of Germany and of 3 home-made milk formulas were analysed, using gravimetry of extracted lipids and high-resolution capillary gas-chromatography. Results were compared with the composition of human milk. Fat contents of all commercial formulas were similar to human milk values and met current recommendations, but 2 home-made preparations were at the upper and lower limits of the recommended range. Milk formulas tended to contain higher percentages of saturated and lower ones of cis-monounsaturated and trans-isomeric fatty acids than human milk. Linoleic acid (C18:2n-6) content was similar to human milk in most products but deviated clearly from recommended values in 2 home-made mixtures. Alpha-linolenic acid (C18:3n-3) values were often low in formulas, resulting a high n-6/n-3-ratios. In contrast to human milk, all formulas contained only minor amounts of the physiologically important long-chain polyunsaturated fatty acids with 20 and 22 carbon atoms. Some seasonal variation in the content of palmitic (C16:0), oleic (C18:1n-9), linoleic and trans-fatty acids was found when five arbitrarily selected adapted formulas were analysed repeatedly over one year. The composition of a home-made formula made from fresh cow's milk was markedly different in winter and in summer, when percentages of saturated and trans-fatty acids were higher and of linoleic acid were lower. We conclude that the composition of most commercial formulas is better suited to meet the lipid requirements of young infants than the home-made preparations investigated.(ABSTRACT TRUNCATED AT 250 WORDS)

The genetics of Crohn disease: Complex segregation analysis of a family study with 265 patients with Crohn disease and 5,387 relatives

Abstract: We have analysed the pedigrees of 265 probands with Crohn disease, collected from a specialty clinic at the University of Dusseldorf. Complex segregation analysis suggests the presence of a recessive gene with incomplete penetrance for susceptibility to the disease with no residual causes of family resemblance. However, a proportion of the isolated cases are probably due to phenocopies, this proportion being greatest among cases with an advanced age of onset.

Astroglia-released neurite growth-inducing activity for embryonic hippocampal neurons is associated with laminin bound in a sulfated complex and free fibronectin

Abstract: Neurons from embryonic (E18) rat hippocampus were chosen to identify and characterize neurite growth-stimulating proteins accumulating in serum-free conditioned media (CM) obtained from primary or secondary cultures of cerebral astrocytes (less than 5% nonglial cells) using a quantitative cell culture bioassay. CM were fractionated by FPLC on an anion exchange column (Mono Q) and by gel filtration (Superose 6). Column fractions were analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis, immunoblotting and enzyme-linked immunosorbent assay (ELISA) using antibodies to laminin (LN) and fibronectin (FN). The neurite-promoting activity (NPA) was tested by incubating aliquots of the eluted fractions with poly-L-lysine precoated glass coverslips prior to addition of neurons suspended in chemically defined medium. We provide evidence that the NPA in astroglial CM could be assigned mainly to a negatively charged, highly sulfated LN complex consisting predominantly of the B-chains of LN and presumably a sulfated proteoglycan that was sensitive for chondroitinase and to a lower degree to heparinase degradation. In addition, a smaller proportion of the NPA was associated with uncomplexed LN and free FN. FN reached approximately 10 times the concentration of LN in astroglial CM. As revealed by immunofluorescence microscopy, both LN and FN are simultaneously expressed by cultured astrocytes; however, only the production of FN, measured by ELISA, increased during the time astrocytes were in culture, whereas the release of LN remained unchanged. We conclude that, besides the most active LN complex, FN bound to a polycationic matrix is able to induce neurite growth in hippocampal neurons in vitro.

Immunological alterations inducible by mercury compounds. III. H-2A acts as an immune response and H-2E as an immune "suppression" locus for HgCl2-induced antinucleolar autoantibodies.

Abstract: In responder mouse strains repeated injections of subtoxic doses of HgCl2 induce formation of antinuclear autoantibodies (ANA) and antinucleolar autoantibodies (ANolA). Others have shown that responsiveness to HgCl2-induced formation of ANA and ANolA is linked to H-2. Here, we extend these studies to a variety of mouse strains not tested previously. After confirming that strain B10.S (H-2s) is a high responder we have shown that strains B10.D2 (H-2d) and B10.BR (H-2k) are nonresponders. By comparing a panel of strains carrying appropriate intra-H-2 recombinant haplotypes derived from d, k and s, we were able to map responsiveness to As. Interestingly, among four strains all of which were As, and thus responsive, only the two H-2E- ones, B10.S and B10.RSD2, were high responders whereas the two H-2E+ ones, B10.HTT and B10.S(9R), were significantly less responsive. Thus, the genetics of HgCl2-induced autoantibody formation follow the rules established for immune responses to a variety of different antigens in that expression of H-2E “suppressed” the response.