Metal Cations Defibrillize the Amyloid Beta-Protein Fibrils

Amyloid beta-protein (A) is the major constituent of amyloid fibrils composing -amyloid plaques and cerebrovascular amyloid in Alzheimer's disease (AD). We studied the effect of metal cations on preformed fibrils of synthetic A by Thioflavin T (ThT) fluorescence spectroscopy and electronmicroscopy (EM) in negative staining. The amount of cross beta-pleated sheet structure of A 1–40 fibrils was found to decrease by metal cations in a concentration-dependent manner as measured by ThT fluorescence spectroscopy. The order of defibrillization of A 1–40 fibrils by metal cations was: Ca²⁺ and Zn²⁺ (IC₅₀ = 100 M) > Mg²⁺ (IC₅₀ = 300 M) > Al³⁺ (IC₅₀ =1.1 mM). EM analysis in negative staining showed that A 1–40 fibrils in the absence of cations were organized in a fine network with a little or no amorphous material. The addition of Ca²⁺, Mg²⁺, and Zn²⁺ to preformed A 1–40 fibrils defibrillized the fibrils or converted them into short rods or to amorphous material. Al³⁺ was less effective, and reduced the fibril network by about 80 % of that in the absence of any metal cation. Studies with A 1–42 showed that this peptide forms more dense network of fibrils as compared to A 1–40. Both ThT fluorescence spectroscopy and EM showed that similar to A 1–40, A 1–42 fibrils are also defibrillized in the presence of millimolar concentrations of Ca²⁺. These studies suggest that metal cations can defibrillize the fibrils of synthetic A.     

Über das Wachstum der Pankreaszellen von Säugetieren als Monolayer Cultures

Zusammenfassung Fetale menschliche Pankreaszellen bilden sowohl spontan auf zuerst gewachsenem Zellrasen (self-aggregation) als auch in Rollröhrchen (48 rph) (rotation-mediated-aggregation) histotypische Aggregate, die A- und B-Zellen enthalten. In der Nährlösung dieser 6–17 Tage lang gewachsenen Gewebestückchen konnte, Insulin (=IRI=immunoreactive-insulin) nachgewiesen werden.

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Monolayer cultures of pancreatic tissueII. Formation of histotypic aggregates from cell suspensions of fetal human pancreas

Summary Fetal Pancreas cells of man, whether in self-aggregation or rotation-mediated-aggregation, form spontaneously histotypical aggregates containing A- and B-cells. It is possible by means of an immunological method to determine insulin (=IRI=immuno-reactive-insulin) in the nutrient medium of these 6 to 17 days old tissue aggregates.

     

Isoform-Specific Membrane Translocation of Protein Kinase C After Ischemic Preconditioning

Mild cerebral anoxic/ischemic/stress insults promote tolerance and thereby protect the brain from subsequent lethal anoxic/ischemic insults. We examined whether specific activation of PKC , , , or isoforms is associated with ischemic preconditioning (IPC) in rat brain. IPC was produced by a 2-minute global cerebral ischemia. Membrane and cytosolic fractions of the hippocampi were immunoblotted using specific antibodies for PKC, , , and . PKC showed a significant translocation to the membrane fraction from 30 min to 4 h and PKC at 4 h following IPC. In contrast, the membrane/cytosol ratio of PKC showed a tendency to decrease at 30 min and 8 h, and the membrane/cytosol ratio of PKC was significantly decreased from 30 min to 24 h following IPC. These findings indicate PKC isoform-specific membrane translocations in the hippocampus after brief global brain ischemia and suggest that activation of PKC and PKC may be associated with IPC-induced tolerance in the rat hippocampus.     

Defective remethylation of homocysteine is related to decreased synthesis of coenzymes B2 in thyroidectomized rats

Summary. We investigated the influence of hypothyroidism on homocysteine metabolism in rats, focusing on a hypothetical deficient synthesis of FAD by riboflavin kinases. Animals were allocated in control group (n=7), thyroidectomized rats (n=6), rats with diet deficient in vitamin B2, B9, B12, choline and methionine (n=7), thyroidectomized rats with deficient diet (n=9). Homocysteine was decreased in operated rats (2.6±1.01 vs. 4.05±1.0mol/L, P=0.02) and increased in deficient diet rats (29.56±4.52 vs. 4.05±1.0mol/L, P=0.001), when compared to control group. Erythrocyte-Glutathione-Reductase-Activation-Coefficient (index of FAD deficiency) was increased in thyroidectomized or deficient diet rats (P=0.004 for both). Methylenetetrahydrofolate-reductase and methionine-synthase activities were decreased in thyroidectomized rats but not in those subjected to deficient diet. Cystathionine--synthase was increased only in operated rats. Taken together, these results showed a defective re-methylation in surgical hypothyroidism, which was due in part to a defective synthesis of vitamin B2 coenzymes. This defective pathway was overcompensated by the increased Cystathionine--synthase activity.     

The taxonomic significance of the trunk limbs of the chydoridae (Cladocera)

Summary The differences in the structure of the trunk limbs allow to outline three sections of Chydoridae (see table I and fig. 1), coinciding with the sections distinguished according to the structure of the head pores.

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Chydoridae (Cladocera)

Chydoridae (. ), , .

     

Degradation of 5-pregnene-3β, 20β-diol by a Nocardia sp.

Summary With growing cells of a Nocardia sp., isolated from soil, the degradation of 5-pregnene-3, 20-diol into 3-[5-oxo-7a-methyl-1 (1-hydroxo)-ethyl-3a-perhydroindane-4]-propionic acid was investigated. The results show that iron is essential for production of the perhydroindanpropionic acid, that this production is greatly enhanced by the presence of calcium and that it is maximal in the pH range 7.0–7.5.Abbreviations used in the text PD 5-pregnene-3, 20-diol (pregnendiol) - PDSA 3-[5-oxo-7a-methyl-1(1-hydroxo)-ethyl-3a-perhydroindane-4]-propionic acid (pregnendiol-secoacid) - PSA 3-[5-oxo-7a-methyl-1-acetyl-3a-perhydroindane-4]-propionic acid (progesterone-secoacid) - EDTA Ethylendiamintetracetic acid - DMSO Dimethylsulfoxide     

Stereospecificity and electrogenicity of amino acid transport in Riccia fluitans

In the aquatic liverwort Riccia fluitans, membrane depolarization (_m), change in membrane conductance (g_m), and current-voltage (I-V) characteristics in the presence of different amino acids as well as the uptake of ¹⁴C-labeled amino acids were measured. L-isomers of the tested amino acids generate larger electrical effects (_m, g_m) than D-isomers, and the I-V characteristics show that the positive electrical inward-current of 20 mA m^-2 generated by 0.5 mM D-serine is only about 50% of the current generated by adding 0.5 mM L-serine. Whereas - and -amino acids rapidly depolarize the membrane to the same extend, with -aminobutyric acid (-AB) and dipeptides no significant electrical effects have been measured. The uptake kinetics of ¹⁴C-labeled amino acids display three components: (I) A saturable high-affinity component with K_s-values of 48 M D-alanine, 12 M -aminoisobutyric acid (AIB), 9 M L-alanine, 8 M L-proline, and 6 M L-serine, respectively; (2) an apparently linear low-affinity component, and (3) an also linear but unspecific component at concentrations >20 times the given K_s-value. Uptake of ¹⁴C-labeled AIB can be inhibited competitively by all tested neutral amino acids, the L-isomers being more effective than the D-isomers, as well as by ammonium or methylamine. Vice versa, AIB competitively inhibits uptake of L-serine and L-alanine. It is concluded that an uncharged stereospecific carrier for the investigated amino acids exists in the plasmalemma of Riccia fluitans. Accumulation ratios of about 50 suggest secondary active transport driven by a transmembrane electro-chemical gradient (mainly _m) which is generated by the electrogenic proton pump. It is suggested that this carrier binds to the amino group forming either a charged binary complex with positively charged amines (Felle 1980), or an uncharged complex with -AB or dipeptides, whereas electrogenic transport of - and -amino acids is mediated by a ternary carrier complex, probably charged by a proton.Symbols and Abbreviations _m membrane potential (mV) - E_co equilibrium potential (mV) of the transport system - g_m membrane (slope) conductance (Sm^-2) - g_m change in g_m - I-V curve current-voltage curve - AIB -aminoisobutytric acid - -AB -aminobutyric acid     

Mode of uptake of sterol by Arthrobacter simplex

The mechanism of uptake of water-insoluble -sitosterol by a newly isolated strain of Arthrobacter simplex SS-7 was studied. The production of an extracellular sterol-pseudosolubilizing protein during growth of A. simplex on -sitosterol was demonstrated by isolating the factor from the cell-free supernatant and its subsequent purification by Sephadex G-150 column chromatography. The M _r of the purified sterol-pseudosolubilizing protein determined by SDS–PAGE was 19kDa. The rate of sterol pseudosolubilization (5.2×10^–3g l^–1h^–1) could not adequately account for the rate of sterol uptake (72×10^–3g l^–1h^–1) and the specific growth rate (56×10^–3 h^–1). However in the unfavourable growth condition, when the cells were treated with sodium azide at the level of 30–60% of MIC, the sterol pseudosolubilization accounted for nearly 74% of the total growth containing 96% free cells. Cellular adherence to substrate particles was found to play an active role in the normal growth of the strain on -sitosterol. Unlike sodium acetate-grown cells, whose surface activity was negligible (60mNm^–1), the sterol-grown cells had strong surface activity (40mNm^–1). The high lipid content and long chain fatty acids in the cell-wall of -sitosterol-grown cells probably contribute to the high sterol adherence activity of the cells.     

The cytology of phytophthora infestans

Summary Phytophthora infestans has three kinds of somatic nuclei: an oval shaped nucleus (approx. 3.1×2.7 ) which stains diffusely except for a crescent shaped Feulgen positive cap which stains intensely; a granular nucleus whose contents are organized into a fairly constant number of stained bodies, and, a deeply staining condensed nucleus. The capped nucleus is thought to be metabolic or resting and the granular nucleus is thought to be dividing as it is most commonly found in hyphal tips. Attenuated forms of all three kinds of nuclei are found.Nuclear division is mitotic and intranuclear. Eight—ten chromosomes are seen at metaphase.Sporangia have a mean of 6.3 nuclei which is constant for age and strain of culture. Sporangia become multinucleate as a result of nuclear migration and not by division in the developing sporangium. Zoospores are usually uninucleate.The nuclear cap is persistent throughout nuclear division when it also divides. It is associated with flagella production and nuclear migration and has some of the properties of a blepharoplast.     

Existence and uniqueness of a sharp travelling wave in degenerate non-linear diffusion Fisher-KPP equations

In this paper we use a dynamical systems approach to prove the existence of a unique critical value c ^* of the speed c for which the degenerate density-dependent diffusion equation u _ct = [D(u)u _x ] _x + g(u) has: 1. no travelling wave solutions for 0 < c < c ^*, 2. a travelling wave solution u(x, t) = (x - c ^* t) of sharp type satisfying (– ) = 1, () = 0 ^*; '(^*–) = – c ^*/D'(0), '(^*+) = 0 and 3. a continuum of travelling wave solutions of monotone decreasing front type for each c > c ^*. These fronts satisfy the boundary conditions (– ) = 1, '(– ) = (+ ) = '(+ ) = 0. We illustrate our analytical results with some numerical solutions.     

Triggering and predisposing factors in the “Red” decline syndrome of Norway spruce (Picea abies)

Summary Analysis of 62 mature Norway spruce (Picea abies provenance Viborg) trees growing in a Danish plantation was undertaken along with analysis of their nutrient contents (N, P, K, Ca, Mg, Fe, Mn, Cu, Zn, B and Na), in each of the three youngest needle age classes, from branches of four exposure directions near the tree top. The aim was to investigate if one among the studied possible predisposing factors was also a triggering factor in the 1989 outbreak of the Red Norway spruce decline in Denmark. Neither nutrient imbalance or deficiency, nor excessive N-deposition or salt-stress were indicated as triggering factors in 1989. The Red syndrome, noticeable for the bright red colour of the current-year needles, was found to be an extension of the European type Novel Decline. Red syndrome is similar to previously reported phenomena of top-dying and sub top-dying, in that it had fewer needle age classes and significantly higher contents of mobile cations (and Ca) in the younger needle classes. Tree ring analysis suggested that the Red syndrome was initiated in the early 1980s, when the trees experienced adverse climatic conditions. Because of this long-term development of the Red Norway spruce decline syndrome, it is concluded that a triggering factor is of minor importance relative to the multitude of predisposing factors.     

An elongated segment of DNA observed between two human α globin genes

Summary Detailed restriction enzyme analysis of the DNA from a Chinese female showed that one of her chromosomes had a >17.5 kb deletion of DNA, including the , 2, and 1 globin genes, which is present in many Southeast Asians with an -thalassemia-1 chromosome. Her normal chromosome had the expected cluster of -like globin genes (5----2-1-3), but the segment of DNA between the two globin genes was elongated by some 0.5–0.7 kb. Analyses of various restriction sites suggested that this normal variant of the human globin gene complex is due to a crossover between a normal chromosome with () and a chromosome with an -thalassemia-2 (–^3.7) and an -21-hybrid gene.     

Psychological changes associated with self-regulatory treatments of irritable bowel syndrome

The psychological side effects of self-regulatory treatment (a combination of relaxation, thermal biofeedback, and cognitive therapy) for irritable bowel syndrome (IBS) were compared among 20 successfully treated patients, 12 unsuccessfully treated patients, and 9 patients who merely monitored symptoms for 12 weeks. Pretreatment and posttreatment scores on the Beck Depression Inventory, State-Trait Anxiety Inventory, and Psychosomatic Symptom Checklist were examined. Successfully treated patients had significant (p<.01) reductions on all measures and significantly greater reductions on depression and state anxiety than the symptom monitoring group. Interestingly, the failures also showed a significant (p=.027) reduction in trait anxiety and no significant increases on other measures.     

The filtration apparatus of Cladocera: Filter mesh-sizes and their implications on food selectivity

Summary The filtering apparatus of eleven Cladoceran species was studied. The distances between the setulae, which act as filters, were measured. Among adult individuals, they vary from 0.2 m in Diaphanosoma brachyurum to 4.7 m in Sida crystallina. Species can be grouped according to the mesh-sizes, as fine mesh filter-feeders: Diaphanosoma brachyurum, Ceriodaphnia quadrangula, Chydorus sphaericus, Daphnia cucullata and Daphnia magna; medium mesh filter-feeders: Daphnia galeata, D. hyalina. D. pulicaria, Bosmina coregoni, and coarse mesh filter-feeders: Holopedium gibberum and Sida crystallina. In Daphnia hyalina, the distances between setulae increase from 0.3–0.4 m in small juveniles, to 0.8–2.0 m in adults. In Daphnia magna, the mesh-size of the filter does not increase significantly with growth. There is good evidence that the relative abundance of the filter-feeding types varies with the trophic state of the lake. In oligotrophic lakes the coarse mesh filter-feeders usually dominate throughout the year. The seasonal succession of zooplankton species in eutrophic lakes can be interpreted as a succession of feeding types; during winter coarse mesh filter-feeders dominate, while fine mesh filter-feeders are most abundant during summer phytoplankton blooms. Our results support the hypothesis that the species composition of filter-feeding zooplankton is strongly influenced by the amount of suspended bacteria which are available as food only for filter-feeding species with fine meshes.     

The structure of the incompatibility factors of Schizophyllum commune: Constitution of the three classes of B factors

Summary The B factors of Schizophyllum commune are of 3 classes: The high recombining class I has 7 alleles and 7 alleles; the low recombining classes are class II with 7 allels and probably 2 alleles and class III with probably 2 (or also 2) alleles and 7 allels. A fourth hypothetical class (-) was not found and either does not exist or is indistinguishable from class III by the tests employed. The and alleles differ from and by either (a) mutations affecting both mating specificity and recombination frequency, or (b) deletions involving most of the B region.The research was supported by a grant from the Atomic Energy Commission of the U.S. No. (30-1)-3875 and was performed at the Biological Laboratories, Harvard University, Cambridge, Mass., U.S.A.     

Photoreversion und Photoprotektion bei UV-induzierten Mutationen des nichtphotoreaktivierbaren Bacteriums E.coli phr?

Summary Stationary phase cells of strain phr^–/MC 2 ofE. coli are not photoreactivable but the frequency of UV-induced mutations to low Streptomycine-resistance (S 3, 3/ml) is decreased strongly by illumination with light of fluorescence tubes (310 to 500 nm) after UV-irradiation. Also dark-reversion (DRM) of these mutations due to keeping UV-irradiated cells in saline is observed. Illumination before UV-irradiation decreases the frequency of the mutations (photoprotection against mutation=PPM) to the same extent as the combined action of photoreversion (PRM) and DRM. The lag-phase of cell division is prolonged strongly by illumination from 80 min without light to 150 min by the light-dose of highest activity. The additional lag is nearly the same if the illumination is done before, after or without UV-irradiation; this lag is about additive to the small lag caused by UV. Pre-illumination of the stationary-phase cells does not cause photoprotection against killing (PP), it even decreases the survival after high UV-doses. The observations support the hypothesis that PRM in this strain may be indirect, i.e. caused by the light-induced additional division lag which enhances the dark repair of UV-premutations. Also spontaneous premutations which are apparently present in the stationary-phase cells seem to be influenced by the light in this way.     

Ultrastructure and behaviour of the spindle pole body of the aphid-pathogenic fungusErynia neoaphidis

Summary A detailed account of the ultrastructure and behaviour of the spindle pole body (SPB) of the entomophthoraceous fungusErynia neoaphidis is presented for the first time.The SPB consists of extranuclear (ENC) and intranuclear (INC) components. The ENC is a saucepan-shaped structure which lies in a pocket of the nuclear envelope. It is composed of a forked, fibrillar handle and a shallow, cylindrical pan. The pan has a wall of two layers, both of which are thickened with a regular periodicity so that they appear to be beaded. It is postulated that the pan is formed from rough endoplasmic reticulum and that it synthesizes the amorphous, electron-dense material coating the ENC.The INC is a saucer-shaped, electron-dense plaque in which the ends of the spindle microtubules terminate. During metaphase, a clear zone separates the INC from the nuclear envelope and persists until telophase. The roles of the amorphous, electron-dense material and the clear zone as well as the method of SPB replication are discussed.     

SDS Induced Structural Changes in α-Crystallin and It’s Effect on Refolding

-Crystallin, a major eye lens protein and a key member of the small heat shock protein family, acts like a chaperone by preventing aggregation of substrate proteins. One of the hallmarks of most small heat shock proteins is their existence as a large oligomer, the role of which in its function is not understood at present. We have studied the role of the oligomer in the stability of its structure against SDS induced destabilization by CD measurements. -Crystallin from bovine source as well as recombinant preparation was used for this purpose. As SDS concentration was gradually increased, the -sheet structure was diminished followed by concomitant increase in the -helical structure. The quaternary structural changes in presence of SDS were also monitored by light scattering, polarization and anisotropy measurements. It was found that the breakdown of the oligomeric structure was nearly complete above 1 mM SDS concentration. The results were compared with that of a monomeric -crystallin, which is also a major -sheet protein like -crystallin. When -crystallin was first converted into monomeric random coil structure in presence of 6 M urea and allowed to refold in SDS solution, amount of -helix was more than that incubated directly in the same concentration of SDS. The results show that -crystallin attains extra structural stability against external stress due to its oligomeric structure. The implication for the extra stability is discussed in reference to its function as molecular chaperone.     

Freeze-fracture studies on the cockroach hemocyte membrane complex: Symmetric and asymmetric membrane particle distributions

Summary Freeze-fracture studies were conducted on the membranes of normal cockroach hemocytes. The plasmalemma is asymmetric with the A fracture face containing 80–100 Å membrane intercalated particles at a concentration of 2500/². The B fracture face contains 120–150 Å particles with a relatively low density (800/²). The nuclear envelope displays an asymmetry with the A fracture face containing 1500 particles/² and the B face containing 300/ ². No significant particle size differences were observed in nuclear envelope fracture faces. Two types of symmetric membranes were also found in these cells. Both A and B fracture faces of the membrane surrounding the numerous cytoplasmic inclusion bodies contain particle sizes and concentrations similar to the B face of the plasmalemma. A second type of symmetry was observed in cells apparently engaged in exocytosis. Vesicles (0.1 D) from this process were completely particle free on both fracture faces. Such particle free vesicles could be found in the cytoplasm, attached to the plasmalemma, or completely separated from the cell.Supported by a Pharmaceutical Manufacturers Association Foundation Fellowship.The author wishes to thank Ms. Annalena K. Charla for assistance in plate preparation, Dr. Julius Schultz and the Papanicolaou Cancer Research Institute for use of the freeze-etch device, and Dr. David Smith for the electron microscope facilities.     

Cytokine production in whole blood cell cultures of patients undergoing therapy with biological response modifiers or 5-fluorouracil

We have measured the levels of interferon (IFN), tumor necrosis factor (TNF), interleukin-1 (IL-1), IL-1, and IL-2 in the whole blood cell culture supernatants of 43 tumor patients undergoing a treatment with biological response modifiers or a conventional therapy with 5-fluorouracil and leucovorin. In the blood cell cultures of the 16 patients who received 5-fluorouracil and leucovorin IFN levels decreased (P0.01) and TNF levels rose (P0.05) during each therapy cycle. However, in the blood samples a declining number of total leukocytes and lymphocytes was measured (P0.05). Progressive disease could be correlated to a tendency towards lower IFN levels in the pretherapeutic cultures of these patients. The second group analyzed consisted of 8 patients receiving a low-dose IL-1 therapy. In this group we found either an unchanged or an augmented IFN production of the blood cells during treatment. In the group of 13 patients receiving low-dose recombinant IL-2 (4.5×10⁶IU m^–2 day^–1) significantly increasing IFN levels were seen in the blood cell cultures during the therapy (P0.05), although total leukocyte counts decreased. In this group, 4 had stable disease for at least 2 months and 9 patients had tumor progression under therapy. In the cultures of the latter a tendency towards lower IFN values was found. Finally, the cytokine production in the blood cell cultures of 6 patients receiving a combination therapy of IFN and high-dose IL-2 was studied. During this therapy a dramatically reduced production not only of IFN but also of all other measured cytokines was found. In this group all patients had tumor progression under therapy. It is concluded that the measurements of cytokine production in a reproducible whole blood culture system may be useful for monitoring immunological therapies and may help us to find out which doses of biological response modifiers have enhancing or suppressive effects on the functions of the immune cells.