Nyctohemeral rhythmicity of type II thyroxine 5′-deiodinase activity in the pineal gland but not in the Harderian gland of the Swiss mouse

Type II thyroxine 5-deiodinase (5-D) activity in both pineal and Harderian glands of the Swiss mouse was studied. Pineal 5-D activity exhibited a nyctohemeral profile with a maximal peak value at 05.00 h, which coincides with that for pineal melatonin production. However, no rhythm of 5-D activity in the Harderian gland could be found. In pineal gland, light at night inhibited the nocturnal increase in 5-D activity, while isoproterenol, a -adrenergic agonist, could not stimulate the enzyme. In the Harderian gland, neither darkness, nor light at night, or isoproterenol were capable of modifying basal values of 5-D activity.     

Parameters involved in the enhancement of monoclonal antibody targeting in vivo with recombinant interferon

Summary The effects of recombinant human leukocyte (clone A) interferon (rHu-IFN-A) were investigated on the expression of monoclonal antibody (MAb)-defined tumor antigens expressed on human mammary and colon carcinomas. The rHu-IFN-A treatment substantially increased the localization of radiolabeled MAb B6.2-F(ab)₂ to the transplantable Clouser human mammary carcinoma, as well as to the moderately differentiated human colon xenograft WiDr, when grown as s.c. tumors in athymic mice. In contrast, human tumor cell lines (i.e., LS174T, A375, etc.) that were unresponsive to the antigen-augmenting ability of rHu-IFN-A in vitro were also unresponsive in vivo, indicating a possible method of screening carcinoma cell populations for subsequent rHu-IFN-A adjuvant therapy prior to MAb administration. The method of delivery of rHu-IFN-A was also studied. The i.m. route resulted in a 3–4 h plasma half-life for rHu-IFN-A. The administration of rHu-IFN-A via an osmotic pump resulted in a stable circulating plasma titer of 400–800 antiviral units/ml for 7 days. Utilizing delivery of rHu-IFN-A by the constant infusion route, it was found that the increase in localization of ¹²⁵I-B6.2-F(ab)₂ was dependent on (1) the length of time of treatment and (2) the circulating plasma rHu-IFN-A levels. These results thus provide information useful for subsequent studies to determine the potential efficacy of adjuvant rHu-IFN-A treatment for MAb-targeted tumor diagnosis and treatment.     

Synthesis of N-linked pentasaccharides with isomeric glycosidic linkage

As part of a program to explore the structural requirement of N-glycans in the carbohydrate-mediated biological interactions, N-linked pentasaccharide core structure was stereochemically modified in terms of glycosidic linkage. Three isomers, -D-Man-(13)-[-D-Man-(16)]--D-Man-(14)--D-GlcNAc-(14)--D-GlcNAc-L-Asn, -D-Man-(13)-[-D-Man-(16)]--D-Man-(14)--D-GlcNAc-(14)--D-GlcNAc-L-Asn, and -D-Man-(13)-[-D-man-(16)]--D-Man-(14)--D-GlcNAc-(14)--D-GlcNAc-L-Asn, were synthesized. Synthesis of the pentasaccharide with natural linkage is also described.     

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.     

Comparative study on the chloroplast,mitochondrial and nuclear genome differentiation in two cultivated rice species,Oryza sativa and O. glaberrima,by RFLP analyses

Summary Restriction fragment length polymorphisms of chloroplast (ct), mitochondrial (mt) and nuclear DNA were investigated using eight cultivars of Oryza sativa and two cultivars of O. glaberrima. Relative variability in the nuclear and cytoplasmic genomes was estimated by a common measure, genetic distance. Based on the average genetic distances among ten cultivars for each genome, the evolutionary variabilities of the mitochondrial and nuclear genomes were found to be almost the same, whereas the variability of the chloroplast genome was less than half that of the other two genomes. Cluster analyses on ct and mt DNA variations revealed that chloroplast and mitochondrial genomes were conservative within a taxon and that their differentiations were well-paralleled with respect to each other. For nuclear DNA variation, an array of different degrees of differentiation was observed in O. sativa, in contrast with little variation in O. glaberrima. As a whole, differentiation between O. sativa and O. glaberrima was clearly observed in all three genomes. In O. sativa, no notable difference was found between the cultivars Japonica and Javanica, whereas a large differentiation was noticed between Japonica (including Javanica) and Indica. In all three genomes, the average genetic distances within Indica were much larger than those within Japonica (including Javanica), and almost similar between Japonica (including Javanica) and Indica. These facts indicate that differentiation in O. sativa was due mainly to Indica.     

Plasma α- and γ-tocopherol have different pattern during normal human pregnancy

Plasma - and -tocopherol were monitored in pregnant women throughout healthy gestational periods and after delivery and were compared with that of non pregnant women. The mean plasma -tocopherol and -tocopherol concentrations in non pregnant Saudi women (15.2 ± 1.3 and 1.8 ± 0.2 ol/l respectively) were found within normal range. The maternal plasma -tocopherol level steadily increased reaching maximum level (19.1 ± 1.6 mol/l) at late gestation and then gradually decreased after delivery. On the contrary, the optimum level of -tocopherol (2.1 ± 0.2 mol/l) was at mid gestation, followed by a progressive decrease until one month after delivery (1.5 ± 0.1 ol/l). This study shows that the maternal plasma - and -tocopherol have different profiles that may be attributed to their different responses to the changes in maternal lipids during pregnancy.     

Investigation by pyrolysis mass spectrometry,phage pattern and plasmid analysis of staphylococci that have reverted from ‘L’-phase to bacterial phase

No major differences have been found in series of Staphylococcus aureus strains which reverted from L-phase, either by pyrolysis mass spectrometry or by phage-typing or sensitivity testing. In L-phase they have been subcultured for a long time or transformed/reverted many times into/from L-phase. Plasmids were lost during transformations/reversions, but there was some difference between the tetracycline-connected plasmids on the one hand and the erythromycin-connected ones on the other.This investigation was supported by the Foundation for Fundamental Research on Matter (F.O.M.), subsidized by the Netherlands Organization for the Advancement of Pure Research (Z.W.O.).     

Effects of midazolam on the contraction and relaxation of segments of thoracic aorta stripped of endothelium and stimulated by adrenaline – experimental study in rabbits

To evaluate the effects of midazolam on the angiokinesis of segments of rabbits' thoracic aorta stripped of endothelium and stimulated by adrenaline.Two groups of aortic rings removed from albinic rabbits anesthetized with thiopental were used (Group I – 6 animals; Group II – 12 animals), stripped of endothelium, studied in an organ chamber, perfused by Krebs-Henseleit solution. The groups were stimulated by adrenaline, recording the maximum contraction and dT/dt at 12, 36, 60 and 120. When the plateau phase was reached, the vessel was washed with perfusion solution, recording relaxation at 2, 4 and 6. When the base values were reached, Group I underwent a new adrenergic stimulus; and Group II was stimulated with midazolam and then with adrenaline, and the same values were recorded. T test was applied as a statistical analysis when two variables were studied. When studying more than two variables the Anova test was used, supplemented by the Tuckey test.Group I did not show any significant difference between the two stimuli. Group II – the midazolam significantly reduced the maximum contraction induced by adrenaline (83.01 ± 4.11%) (p < 0.01). The dT/dt was reduced at 12 (57.06 ± 8.47%), and also at 36 (70.59 ± 5.26%). There was no significance at 60 and 120 (p < 0.01).The relaxation increased significantly at all measurements – at 2-adrenaline 39.31 ± 9.60%; adrenaline/midazolam: 44.06 ± 9.62% (p < 0.05). At 4-adrenaline: 53.08 ± 8.3%; adrenaline/midazolam: 61.68 ± 8.50% (p < 0.01). At 6-adrenaline: 76.26 ± 5.45%; adrenaline/midazolam: 84.20 ± 7.96% (p < 0.01).Midazolam significantly reduced the maximum contraction obtained by the adrenergic stimulus as well as the dT/dt in the initial phases of contraction. The relaxation speed also increased.     

Uptake of Zn++ and aflatoxin from perlite and liquid culture by Zea mays seedlings

The present paper reports our attempts to determine whether the inclusion of 0.0014 mM Zn⁺⁺ within a hydroponic culture medium affects the ability of 12-day-old Zea mays, cv. SS-522 to take-up [³H]-aflatoxin B₁. Data from the corollary experiment, i.e., whether inclusion of aflatoxin affects the ability of Zea mays, cvs. Truckers White, X-Sweet and Merit to take-up ⁶⁵ZnCl₂ are presented also. This report is a preliminary to one regarding an in-progress analysis of whether pollutant levels of Zn⁺⁺ affect aflatoxin uptake and distribution. In the absence of irrigating seedlings, which were grown in Perlite containing ⁶⁵ZnCl₂, with a solution containing mixed aflatoxins, the stem contained the greatest amount of label with root plus seed the next highest and the leaf the least for each of the cvs. In contrast, when the seedlings were irrigated with a solution containing mixed aflatoxins, the root plus seed contained either an amount nearly identical to (cv. Truckers White) or in excess of that within the stem (cvs. X-Sweet and Merit). Calculation of the percentages of aflatoxin-induced diminutions in leaf, stem and root label suggested that the aflatoxins interfered with the translocation of ⁶⁵ZnCl₂ from the root to the stem and leaf, at least for cvs X-Sweet and Merit. When 0.0014mM Zn⁺⁺ as ZnSO₄ was added to an incubation medium in which 12-day-old seedlings were suspended and plant growth assessed over 72 hours, a 15% increase (significant at 0.05 level) in seedling height over that of Zn⁺⁺-deficient plants was observed. No differences in [³H]-aflatoxin B₁ uptake were noted between those seedlings which were grown in either Zn⁺⁺-containing or lacking media. Less than one % of the[³H]-aflatoxin B₁ which was taken-up was recovered within chloroform extracts of the seedlings. The distributions of radioactivity from [³H]-aflatoxin B₁ for leaf, stem, seed and root were 0, 57, 26 and 19% and 0, 26, 58 and 18% for Zn⁺⁺-containing and -lacking media, respectively.     

Localization of the gene for human heart fatty acid binding protein to chromosome 1p32-1p33

Among 639 spontaneous abortions between the 8th and 14th week of gestation 342 (53.5%) revealed an abnormal karyotype. While the rate of trisomies distinctly increased with advancing maternal age, a decrease in the rate of 45,X conceptuses and polyploidies was observed among abortions from older women. The overall relation of XXXXXXYY among the tetraploidies was 1411 and that of XXXXXYXYY among the triploidies was 26 361. However, when the latter was related to maternal age, a reversal of the XXXXXY ratio of 12 in the younger to 21 in the older age groups became evident. Furthermore a decrease in the rate of paternally derived partial hydatidiform moles was found among the triploid abortion specimens from older women. From these observations we conclude that digyny plays a major role in the origin of triploidy in the increased maternal age groups, while diandry related to immaturity of oocytes and impairment of oocyte cortical function is more frequent in triploid abortions from younger women.     

Genetics of resistance to Puccinia graminis tritici in ‘Chris’ and ‘W3746’ wheats

Summary Chris wheat possessed genes Sr5, Sr7a, Sr8a, Sr9g and Sr12. W3746, derived from the cross Chris/Baart, possessed Sr7a and Sr12. The response conferred by Sr7a was influenced by the genetic background. Although Sr7a or Sr12 alone conferred no observable resistance upon adult plants, the adult resistances of Chris and W3746 to predominant pathotypes appeared to be associated with the interaction of Sr7a and Sr12, or genes at closely linked loci.     

Multiple shoot regeneration from root organ cultures of Populus alba x P. grandidentata

Excised roots of various ages from Crandon and Hansen clones of Populus alba x P. grandidentata were tested for their regeneration capacity. Sixty-day-old excised roots that contained root tips were found to be most suitable. The highest number of shoots (an average of 111 shoots/root segment with Crandon and 98 with Hansen) was obtained by adding 22M and 14M zeatin to the medium, respectively. The two clones of hybrid poplar responded similarly to growth regulator treatments; however, the number of shoots produced was greater from the root organs derived from Crandon clones. Regenerated shoots were rooted in basal Woody Plant Medium without any growth regulators. Successful transplantation into soil and growth was achieved with all plants.     

MHC class II sequences of an HLA-DR2 narcoleptic

Narcolepsy has a 98% association with the DR2-Dw2/DQw1 haplotype. To establish if a disease-specific allele is present in narcolepsy, a cDNA library was made from a B-cell line from a DR2,4/DQw1,3 narcoleptic. Clones encoding the two expressed DR2 chains, along with DQw1 and chains, were isolated and completely sequenced. The coding regions of these four genes were similar to published nucleotide and protein sequences from corresponding healthy controls, with some minor exceptions. The 3 untranslated region of one of the DR2 genes in the narcoleptic was extended by 42 bp. Complete sequences were not available for DQw1.2 or from healthy individuals, but first domain nucleotide sequences showed only a single nonproductive difference in DQ. Partial protein sequences of both DQ and from published data were identical. Although the effects of minor differences cannot be ruled out completely, it is concluded that there are probably no narcolepsy-specific DR or DQ / sequences, and that the alleles found in narcolepsy are representative of those found in the healthy population.     

Rapid plant regeneration of pea using thidiazuron

A simple and rapid pea regeneration procedure was developed. An average of up to 20 shoots formed from hypocotyl explants of cvs. Sugar Ann and Patriot cultured on Murashige and Skoog basal medium supplemented with 0.5 or 1.0 M thidiazuron (N-phenyl-N-1,2,3-thiadiazol-5-ylurea). Hypocotyls of Puget and Sugar Daddy did not respond. Regenerated shoots rooted rapidly when cultured on Murashige and Skoog basal medium containing either 2.0 M -naphthaleneacetic acid or 1.0–2.0 M indole-3-butyric acid. Seeds were harvested from regenerated plants after only 9–11 weeks.Abbreviations BAP 6-benzyladenine - 2,4-d 2,4-dichlorophenoxy acetic acid - GA₃ gibberellic acid - IBA indole-3-butyric acid - MS Murashige and Skoog (1962) medium - NAA -naphthaleneacetic acid - TDZ thidiazuron (N-phenyl-N-1,2,3-thiadiazol-5-ylurea)     

Action of nereistoxin on recombinant neuronal nicotinic acetylcholine receptors expressed in<Emphasis Type="Italic"> Xenopus laevis</Emphasis> oocytes

Nereistoxin (NTX), a natural neurotoxin from the salivary glands of the marine annelid worm Lumbriconereis heteropoda, is highly toxic to insects. Its synthetic analogue, Cartap, was the first commercial insecticide based on a natural product. We have used voltage-clamp electrophysiology to compare the actions of NTX on recombinant nicotinic acetylcholine receptors (nicotinic AChRs) expressed in Xenopus laevis oocytes following nuclear injection of cDNAs. The recombinant nicotinic AChRs investigated were chicken 7, chicken 42 and the Drosophila melanogaster/chicken hybrid receptors SAD/2 and ALS/2. No agonist action of NTX (0.1–100 M) was observed on chicken 7, chicken 42 and the Drosophila/chicken hybrid nicotinic AChRs. Currents elicited by ACh were reduced in amplitude by NTX in a dose-dependent manner. The toxin was slightly more potent on recombinant Drosophila/vertebrate hybrid receptors than on vertebrate homomeric (7) or heteromeric (42) nicotinic AChRs. Block by NTX of the chicken 7, chicken 42 and the SAD/2 and ALS/2 Drosophila/chicken hybrid receptors is in all cases non-competitive. Thus, the site of action on nicotinic AChRs of NTX, to which the insecticide Cartap is metabolised in insects, differs from that of the major nicotinic AChR-active insecticide, imidacloprid.     

Aldimine to ketoamine isomerization (Amadori rearrangement) potential at the individual nonenzymic glycation sites of hemoglobin a: Preferential inhibition of glycation by nucleophiles at sites of low isomerization potential

The relative roles of the two structural aspects of nonenzymic glycation sites of hemoglobin A, namely the ease with which the amino groups could form the aldimine adducts and the propensity of the microenvironments of the respective aldimines to facilitate the Amadori rearrangement, in dictating the site selectivity of nonenzymic glycation with aldotriose has been investigated. The chemical reactivity of the amino groups of hemoglobin A forin vitro reductive glycation with aldotriose is distinct from that in the nonreductive mode. The reactivity of amino groups of hemoglobin A toward reductive glycation (i.e., propensity for aldimine formation) decreases in the order Val-1(), Val-1(), Lys-66(), Lys-61(), and Lys-16(). The overall reactivity of hemoglobin A toward nonreductive glycation decreased in the order Lys-16(), Val-1(), Lys-66(), Lys-82(), Lys-61(), and Val-1(). Since the aldimine is the common intermediate for both the reductive and nonreductive modification, the differential selectivity of protein for the two modes of glycation is clearly a reflection of the propensity of the microenvironments of nonenzymic glycation sites to facilitate the isomerization reaction (i.e., Amadori rearrangement). A semiquantitative estimate of this propensity of the microenvironment of the nonenzymic glycation sites has been obtained by comparing the nonreductive (nonenzymic) and reductive modification at individual glycation sites. The microenvironment of Lys-16() is very efficient in facilitating the rearrangement and the relative efficiency decreases in the order Lys-16(), Lys-82(), Lys-66(), Lys-61(), Val-1(), and Val-1(). The propensity of the microenvironment of Lys-16() to facilitate the Amadori rearrangement of the aldimine is about three orders of magnitude higher than that of Val-1() and is about 50 times higher than that of Val-1(). The extent of nonenzymic glycation at the individual sites is modulated by various factors, such as thepH, concentration of aldotriose, and the concentration of the protein. The nucleophiles—such as tris, glycine ethyl ester, and amino guanidine—inhibit the glycation by trapping the aldotriose. The nonenzymic glycation inhibitory power of nucleophile is directly related to its propensity to form aldimine. Thus, the extent of inhibition of nonenzymic glycation at a given site by a nucleophile directly reflects the relative role ofpK _a of the site in dictating the glycation at that site. The nonenzymic glycation of an amino group of a protein is an additive/synergestic consequence of the propensity of the site to form aldimine adducts on one hand, and the propensity of its microenvironment to facilitate the isomerization of the aldimines to ketoamines on the other. The isomerization potential of microenvironment plays the dominant role in dictating the site specificity of the nonenzymic glycation of proteins.     

Microsatellite markers in the hexaploid Prunus domestica species and parentage lineage of three European plum cultivars using nuclear and chloroplast simple-sequence repeats

In a previous study, cDNA microsatellite markers were described in apricot (Prunus armeniaca L.). Specific PCR primers were designed to amplify the microsatellite-containing regions from genomic DNA in different Prunus species. In the present work, cDNA microsatellite markers were developed in the hexaploid Prunus domestica L. species and polymorphism was ascertained in a segregating plum population. Co-dominant mendelian segregation of alleles was demonstrated and microsatellite polymorphism displayed up to 6 alleles per SSR locus per individual. Parentage lineage of three full-sib European plum cultivars (cv. Cacanska najbolja, Cacanska rana and Cacanska lepotica) was reconstructed by the analysis of the above nuclear SSR markers, completed by four chloroplastic microsatellite loci. The six most informative nuclear loci enabled discrimination between the three Cacak cultivars and unrelated individuals as well as the previously proposed parents, Wangenheim and Pozegaca. Data obtained support previous evidence that these cultivars originated from the Stanley cultivar. However, SSR analysis finally excluded Wangenheim as the other possible parent. Based on the results obtained with nuclear and chloroplast SSR loci, we propose the origin of those three Cacak cultivars in a cross between Stanley as the mother plant and Ruth gerstetter as the pollinator. Furthermore, we demonstrate the utility of these apricot SSR markers for genotype fingerprinting of the hexaploid plum cultivars.     

Selection of microorganisms which produce raw-starch degrading enzymes

Summary Microorganisms which produce strong raw-starch degrading enzymes were isolated from soil using a medium containing a unique carbon source, -amylase resistant starch (-RS), which is insoluble in water and hardly digested with Bacillus amyloliquefaciens -amylase. Among the isolates, three strains showing high activities were characterized. Two of them, K-27 (fungus) and K-28 (yeast), produced -amylase and glucoamylase, and the final product from starch was only glucose. The third strain, K-2, was a bacterium and produced -amylase, which produced glucose and malto-oligosaccharides from starch. The enzyme preparation of these strains degraded raw corn starch rapidly.     

Enhanced power of QTL detection for Fusarium head blight resistance in wheat by means of codominant scoring of hemizygous molecular markers

Based on different marker information content mapping of QTLs for Fusarium head blight resistance in wheat was compared with regard to number and consistency of detected QTLs as well as QTL positions and effects. Therefore, two linkage maps, obtained by dominant and codominant genotyping of hemizygous markers, were constructed with 211 AFLPs, 37 SSRs and the barley RGA marker XaACT/CAA. The codominant marker set comprised 59% codominant markers, whereas the dominant map consisted of only 13%. A segregating wheat population of 94 F₄-RILs was used for QTL analysis. Fusarium head blight resistance was estimated in field trials in six environments. Conventional dominant marker scoring found seven QTLs. The phenotypic variations explained by QTLs detected in single environment analyses ranged from 11.1 to 44.6%. QTL analysis performed with the codominant marker set confirmed not only all QTL positions as revealed by dominant QTL analysis', but also 12 additional QTLs were found. QTLs in single environments explained 36.3 up to 55.7% of the phenotypic variation. In the QTL analysis across all environments, none of the QTLs could be confirmed using dominant marker scoring. However, by codominant QTL analysis' environment-specific QTLs were retrieved. STS marker XaACT/CAA was found to be significantly associated with FHB resistance only by codominant scoring. Support intervals of QTLs commonly found in both marker sets averaged to 10.3 cM in the dominant QTL analysis', whereas the length was shortened to 8.9 cM by codominant genotyping. The advantages of extracting codominant information from dominant markers are discussed.     

Viability and activity in readily culturable bacteria: a review and discussion of the practical issues

In microbiology the terms viability and culturability are often equated. However, in recent years the apparently self-contradictory expression viable-but-nonculturable (VBNC) has been applied to cells with various and often poorly defined physiological attributes but which, nonetheless, could not be cultured by methods normally appropriate to the organism concerned. These attributes include apparent cell integrity, the possession of some form of measurable cellular activity and the apparent capacity to regain culturability. We review the evidence relating to putative VBNC cells and stress our view that most of the reports claiming a return to culturability have failed to exclude the regrowth of a limited number of cells which had never lost culturability. We argue that failure to differentiate clearly between use of the terms viability and culturability in an operational versus a conceptual sense is fuelling the current debate, and conclude with a number of proposals that are designed to help clarify the major issues involved. In particular, we suggest an alternative operational terminology that replaces VBNC with expressions that are internally consistent.