Age-related changes in the mevalonate metabolism in vivo in chick kidneys

The mevalonate incorporation in vivo into total nonsaponifiable lipids by chick kidneys drastically increased after hatching, reaching similar levels to those previously observed in liver. Cholesterol was the major sterol formed from mevalonate from 11 days onward, while a fraction of polar nonsaponifiable lipid(s) was observed as the major compound(s) synthesized at 5-8 days. Relative percentages of squalene, squalene oxide(s) and lanosterol synthesized from mevalonate also increased between 11-18 days after hatching. Results in this paper demonstrate for the first time the accumulation of a fraction of nonsaponifiable lipid(s) identified as lanosterol derivatives and cholesterol precursors formed by kidneys from [5-14C]mevalonate in experiments carried out in vivo, as well as their evolution during postnatal period.     

白蚁抗病原微生物侵染机制研究进展

白蚁作为古老的社会性昆虫,在其生活史内易受到多种病原物的潜在威胁。本文主要根据白蚁的生理免疫及其社会性行为,如白蚁的细胞免疫和体液免疫以及相互清理等利他性行为,对其抵抗病原物侵染和扩散的个体生理免疫和群体行为防御策略进行了综述,以期为白蚁免疫策略的研究提供必要的信息。目前已从白蚁额腺和胸腺中分离到具抗菌活性的分泌物,且已明确白蚁能产生termicin、spinigerin等抗菌肽和转铁蛋白、溶菌酶等抗菌蛋白,抗菌肽的理化性质已得到初步研究。此外,白蚁的社会性行为( 如相互清理、交哺等) 能有效降低白蚁被病原物侵染的几率。运用RNA干扰等分子生物学方法证明利用病原微生物防治白蚁具有广阔的前景,但需要我们对白蚁抗菌肽的进化、各种抗菌肽和抗菌蛋白之间的相互作用以及各种生理免疫策略与群体内的社会性行为之间的联系进行更深入的研究。     

Variation in chromatin structure in two cell types from the same tissue: a short DNA repeat length in cerebral cortex neurons

We have used micrococcal nuclease as a probe of the repeating structure of chromatin in four nuclear populations from three tissues of the rabbit. Neuronal nuclei isolated from the cerebral cortex contain about 160 base pairs of DNA in the chromatin repeat unit, as compared with about 200 base pairs for nonastrocytic glial cell nuclei from the same tissue, neuronal nuclei from the cerebellum and liver nuclei. All four types of nuclei show the same features of nucleosomal organization as other eucaryotic nuclei so far studied: nucleosomes liberated by digestion with micrococcal nuclease give a “core particle” containing 140 base pairs as a metastable intermediate on further digestion and a series of single-strand DNA fragments which are mutiples of 10 bases after digestion with DNAase I. Nuclei from cerebral cortex neurons, which have a short repeat, are distinct from the others in being larger, in having a higher proportion of euchromatin (dispersed chromatin) as judged by microscopy and in being more active in RNA synthesis in vitro.     

Finding hidden females in a crowd: Mate recognition in fig wasps

Multi-species mating aggregations are crowded environments within which mate recognition must occur. Mating aggregations of fig wasps can consist of thousands of individuals of many species that attain sexual maturity simultaneously and mate in the same microenvironment, i.e, in syntopy, within the close confines of an enclosed globular inflorescence called a syconium – a system that has many signalling constraints such as darkness and crowding. All wasps develop within individual galled flowers. Since mating mostly occurs when females are still confined within their galls, male wasps have the additional burden of detecting conspecific females that are “hidden” behind barriers consisting of gall walls. In Ficus racemosa, we investigated signals used by pollinating fig wasp males to differentiate conspecific females from females of other syntopic fig wasp species. Male Ceratosolen fusciceps could detect conspecific females using cues from galls containing females, empty galls, as well as cues from gall volatiles and gall surface hydrocarbons.In many figs, syconia are pollinated by single foundress wasps, leading to high levels of wasp inbreeding due to sibmating. In F. racemosa, as most syconia contain many foundresses, we expected male pollinators to prefer non-sib females to female siblings to reduce inbreeding. We used galls containing females from non-natal figs as a proxy for non-sibs and those from natal figs as a proxy for sibling females. We found that males preferred galls of female pollinators from natal figs. However, males were undecided when given a choice between galls containing non-pollinator females from natal syconia and pollinator females from non-natal syconia, suggesting olfactory imprinting by the natal syconial environment.     

Effect of ethanol administration on metabolism of lipids in heart and aorta in isoproterenol induced myocardial infarction in rats

Effect of ethanol administration on the severity of myocardial infarction induced by isoproterenol in rats was studied. Even though serum CPK and GOT levels as well as the extent of myocardial damage as revealed by histopathological studies, were similar, the survival rate was higher in rats administered ethanol. Concentration of cholesterol and triglycerides in the serum and heart in rats given ethanol and isoproterenol seems to be the additive effect caused individually by ethanol and isoproterenol. Myocardial alcohol dehydrogenase and aldehyde dehydrogenase both showed increased activity in rats treated with ethanol. The rate of recovery from myocardial infarction however, was slower in rats treated with ethanol as judged from the serum CPK value.     

In vitro systems in pneumocystis research

Most groups involved in Pneumocystis research need large quantities of well preserved, viable Pneumocystis organisms free of host cell contamination. Biological, biochemical, immunological, genetic or other studies on Pneumocystis usually involve the separation of Pneumocystis from lung tissue as well as elimination of host cell debris from parasite extracts. In other investigations, such as transmission, infectivity, life cycle, biochemical, in vitro culture or drug-screening studies, viable and infectious Pneumocystis organisms are urgently required. However, there is no generally accepted methodology for obtaining Pneumocystis from experimental hosts or from human clinical samples; methods are still far from reaching standardization, as discussed here by the members of the European Concerted Action (ECA) on Pneumocystis carinii, which is co-ordinated by Eduardo Dei-Cas and Jean-Charles Cailliez.     

Gene flow in pollen in commercial almond orchards

 Honeybee-assisted gene flow by pollen is of enormous benefit to world food production. Molecular genetic markers were used to detect the pollen-contributing parent in embryos of seeds after fruit set and, subsequently, to deduce the direction and distance of gene flow by pollen as assisted by bees in an almond orchard. It was shown that gene flow by pollen resulting in nut set takes place most strongly between neighbouring sides of adjacent cross-compatible pairs of trees. Examination of pellets extracted from pollen traps fitted to hives for the isozyme markers showed that most of the pellets were composed of pollen from only one cultivar or another, in accord with the hypothesis that the honeybee predominantly visits only one cultivar, either single trees or along rows of one cultivar, and that cross-pollination results from accidental or rare visits involving two cultivars. The results suggest that orchards should be designed to bring cross-compatible pairs of cultivars as close together as possible. Received: 26 April 1996 / Revision accepted: 16 August 1996     

The role of lipocortin I in macrophage-mediated immunosuppression in tumor-bearing mice

The soluble mediators and/or mechanisms involved in immunosuppression in tumor-bearing hosts are not well characterized, although macrophages have long been recognized as major participants. We have investigated the role of lipocortin I, a phospholipid-binding protein, in macrophage-mediated immunosuppression in tumor-bearing mice. Proliferation of splenic lymphocytes in response to the mitogens (PHA, Con A, LPS, and PWM) was severely suppressed in tumor (Sqc-NH-1 carcinoma)-bearing mice. This immunosuppression was associated with a decrease in T and B lymphocytes and an increase in macrophages in these spleens. Mac-2+ macrophages were found only in spleens from tumor-bearing mice. Splenic macrophages from tumor-bearing, but not normal, mice were responsible for this immunosuppression, as revealed by negative and positive selection experiments. The levels of lipocortin I mRNA expression were markedly increased in peripheral blood cells from tumor-bearing mice as compared with those from normal mice. Lipocortin I mRNA was strongly induced in splenic mononuclear cells from tumor-bearing mice. Furthermore, these cells displayed increased expression of lipocortin I protein, as judged by Western blot analysis with polyclonal anti-lipocortin I serum. Some nonimmune organs such as the heart, submaxillary gland, muscle, and bladder also displayed increased levels of lipocortin I mRNA expression in tumor-bearing mice. Mac-2+ macrophages among the splenic mononuclear cells in tumor-bearing mice expressed lipocortin I mRNA, as judged by negative and positive selection experiments. Most of these Mac-2+ macrophages also had Mac-1 and Mac-3 Ag. Lipocortin I protein was increased in the serum of tumor-bearing mice as compared with normal mice. The culture supernatants of splenic cells from tumor-bearing mice suppressed the mitogenic responses of splenic cells from normal mice, and addition of anti-lipocortin I antiserum inhibited this suppression. Furthermore, recombinant mouse lipocortin I suppressed mitogenic responses of splenic cells from normal mice. In summary, Mac-2+ macrophage-derived lipocortin I was largely involved in immunosuppression in tumor-bearing mice.     

Covalent modification of epithelial fatty acid-binding protein by 4-hydroxynonenal in vitro and in vivo. Evidence for a role in antioxidant biology

4-Hydroxynonenal (4-HNE) is a cytotoxic alpha,beta-unsaturated acyl aldehyde that is naturally produced from lipid peroxidation and cleavage in response to oxidative stress and aging. Such reactive lipids covalently modify cellular target proteins, thereby affecting biological structure and function. Herein we report the identification of the epithelial fatty acid-binding protein (E-FABP) as a molecular target for 4-HNE modification both in vitro and in vivo. 4-HNE covalently modified (t(12) < 60 s) E-FABP in vitro, as revealed by a combination of matrix-assisted laser desorption ionization-time of flight mass spectrometry and immunochemical reactivity using antibodies directed to 4-HNE-protein conjugates. Identification of Cys-120 as the major site of modification was determined through tandem mass spectral sequencing of tryptic peptides, as well as analysis of E-FABP mutants C120A, C127A, and C120A/C127A. The in vitro modification of Cys-120 by 4-HNE was relatively insensitive to pH (6.4-8.4), and temperature (4-37 degrees C) but was markedly potentiated by noncovalently bound fatty acids. 4-HNE-modified E-FABP was more stable than unmodified E-FABP to chemical denaturation by guanidine hydrochloride, as assessed by changes in intrinsic tryptophan fluorescence. Analysis of soluble protein extracts from rat retina with antibodies directed to 4-HNE-protein conjugates revealed immunoreactivity with a 15-kDa protein that was identified by electrospray ionization and matrix-assisted laser desorption ionization-time of flight mass spectrometry as E-FABP. Evaluation of retinal pigment epithelial cell extracts derived from E-FABP null mice by two-dimensional gel electrophoresis using anti-4-HNE antibodies revealed increased modification in the null cells relative to those from wild type cells. These results indicate that E-FABP is a molecular target for 4-HNE modification and the hypothesis that E-FABP functions as an antioxidant protein by scavenging reactive lipids through covalent modification of Cys-120.     

Lipidg Peroxidation in Liver and Ehrlich Ascites Cell Mitochondria

Ehrlich ascites cell mitochondria are highly resistant to lipid peroxidation as compared to liver mitochondria from host animals. Succinate protects mitochondria from peroxidative damage, proteins from crosslinks, enzymes from inactivation of the enzymes and membranes from permeability changes. The sensitivity of Ehrlich ascites cell mitochondrial membranes to lipid peroxidation is significantly increased in sub-mitochondrial particles. Lipid peroxidation in tumour mitochondrial membranes can not be diminished by succinate as effectively as in liver mitochondria. Ascites cell mitochondria seems to be protected very efficiently from peroxidative damage by a glutathione-dependent mechanism.     

Glyceraldehyde metabolism in human erythrocytes in comparison with that of glucose and dihydroxyacetone

Metabolism of D-glyceraldehyde in human erythrocytes in comparison with that of glucose and dihydroxyacetone was studied. Both trioses were metabolized to produce L-lactate at rates comparable to that of L-lactate formation from glucose. Almost complete inactivation of glyceraldehyde-3-phosphate dehydrogenase by treatment of cells with iodoacetate resulted in a 95% decrease in L-lactate formation from the ketotriose as well as from glucose, whereas L-lactate formation from the aldotriose was only partially reduced (60%). D-Lactate was produced faster from either the aldotriose or the ketotriose than from glucose, but the ability of the two trioses to produce D-lactate was far lower than that to produce L-lactate. Almost complete inhibition of aldehyde dehydrogenase by disulfiram and of both aldose reductase and aldehyde reductase II by sorbinil, had no effect on L-lactate formation from D-glyceraldehyde. The present study suggests that D-glyceraldehyde is metabolized via two or more pathways including the glycolytic pathway after its phosphorylation by triokinase, and that neither oxidation to D-glyceric acid nor reduction to glycerol is a prerequisite for D-glyceraldehyde metabolism.     

Role of the mukB gene in chromosome and plasmid partition in Escherichia coli

The intracellular locations of oriC and oriR1, the replication origins of the chromosome and plasmid R1, respectively, were visualized by fluorescence in situ hybridization (FISH) in exponentially growing populations of Escherichia coli. The locations of oriC and oriR1 (from a Par+ R1 plasmid) were unique and different in the wild-type host. In a mukB mutant, the positions were perturbed for both origins. The position of oriR1 from a plasmid with active partition (Par+) in the mukB host was as randomized as that of oriR1 from the Par- plasmid in a wild-type host. However, this mukB-induced randomization did not result in unstable inheritance of the Par+ plasmid, as measured by the conventional segregation assay. This might result from the preferential association of the Par+ plasmid with the bigger, decondensed nucleoid-containing daughters during cell division of MukB- cells, whereas the Par- plasmids were distributed at random and were lost by frequently ending up in anucleate cells.     

Asynchrony between pronuclear development and protein synthetic changes in zygotes of the mouse derived from oocytes aged postovulation in vivo and fertilized in vivo

The pattern of proteins synthesized by one-cell embryos derived from unaged oocytes and oocytes aged postovulation in vivo was analyzed by means of 35S-methionine labeling and gel electrophoresis. The oocytes were obtained after ovulation induction by an injection of luteinizing hormone-releasing hormone (LHRH) at proestrus or after a superovulation procedure. The analysis was performed in unfertilized aged and unaged secondary oocytes and in zygotes derived from them. The patterns of proteins synthesized by secondary oocytes from all experimental groups were very similar: The oocytes showed a predominant synthesis of 35 kDa proteins. Zygotes from aged as well as unaged LHRH-induced oocytes also showed a predominant synthesis of one group of polypeptides with a relative molecular weight of about 35 kDa. The proteins of the 35 kDa protein complex migrated in an upper (u), middle (m), or lower (l) band in 10% polyacrylamide sodium dodecyl sulfate (SDS) gels. The u- and m-band 35 kDa proteins were shown to be synthesized by secondary oocytes. Early pronuclear zygotes from unaged LHRH-induced oocytes synthesized u- and m- but no l-band 35 kDa proteins. In contrast, part (38%, n = 47) of the early pronuclear zygotes from aged LHRH-induced oocytes did synthesize the l-band 35 kDa proteins. Late pronuclear zygotes (LPZ) from aged as well as unaged oocytes synthesized predominantly the l-band 35 kDa proteins. However, although only 5.8% (n = 51) of LPZ from unaged oocytes synthesize m- and l-band 35 kDa proteins, these bands of proteins are present in 25% (n = 24) of the LPZ from aged oocytes.(ABSTRACT TRUNCATED AT 250 WORDS)     

Role of sestrin2 in peroxide signaling in macrophages

Reactive oxygen species not only serve as signaling molecules, they also contribute to oxidative stress and cell damage. The thioredoxin and glutaredoxin systems form along with peroxiredoxins a precisely regulated defense system to maintain the cellular redox homeostasis. There is evidence that nitric oxide (NO) protects cells from oxidative stress by preventing inactivation of peroxiredoxins by sulfinylation. Here we demonstrate that NO and hypoxia upregulate Sestrin2 by HIF-1-dependent and additional mechanisms and that Sestrin2 contributes to preventing peroxiredoxins from sulfinylation. We conclude that Sestrin2 plays a role in peroxide defense as a reductase for peroxiredoxins.     

Evaluation of the bioavailability of radionuclides from soil in cattle by an in vitro method

Factors have been examined which influence the radionuclides sorption from soil particles by fluid imitating rumen liquid of the cattle. It is noted that the extent of extractability of 137Cs from soil is influenced mainly by presence of potassium ions in modelling liquid. Major factor influencing the release of 90Sr from soil is pH value of the medium. The presence of heavy metals salts affected the release of radionuclides from soil particles. The data observed make it possible to use the modelling solutions to predict bioavailability of radionuclides from soil as well as to predict possible contamination of animal products (milk and meat).     

Assessment of poliovirus eradication in Japan: genomic analysis of polioviruses isolated from river water and sewage in toyama prefecture

Seventy-eight poliovirus strains isolated from river water and sewage in Toyama Prefecture, Japan, during 1993 to 1995 were characterized by the PCR-restriction fragment length polymorphism (RFLP) method and by partially sequencing the VP3 and VP1 regions of the viral genome. Of these isolates, 36 were identified as Sabin vaccine strains, and 42 were identified as vaccine variant strains that had less than 1.4% nucleotide divergence from the Sabin strains, including 7 isolates with patterns different from those of Sabin strains as determined by PCR-RFLP analysis. These findings suggest that wild-type poliovirus was not circulating in Toyama Prefecture.     

Effect of glycerol on gluconeogenesis in isolated rabbit kidney cortex tubules

In renal tubules isolated from fed rabbits glycerol is not utilized as a glucose precursor, probably due to the rate-limiting transfer of reducing equivalents from cytosol to mitochondria. Pyruvate and glutamate stimulated an incorporation of [14C]glycerol to glucose by 50- and 10-fold, respectively, indicating that glycerol is utilized as a gluconeogenic substrate under these conditions. Glycerol at concentration of 1.5 mM resulted in an acceleration of both glucose formation and incorporation of [14C]pyruvate and [14C]glutamate into glucose by 2- and 9-fold, respectively, while it decreased the rates of these processes from lactate as a substrate. In the presence of fructose, glycerol decreased the ATP level, limiting the rate of fructose phosphorylation and glucose synthesis. As concluded from the 'cross-over' plots, the ratios of both 3-hydroxybutyrate/acetoacetate and glycerol 3-phosphate/dihydroxyacetone phosphate, as well as from experiments performed with methylene blue and acetoacetate, the stimulatory effect of glycerol on glucose formation from pyruvate and glutamate may result from an acceleration of fluxes through the first steps of gluconeogenesis as well as glyceraldehyde-3-phosphate dehydrogenase. As inhibition by glycerol of gluconeogenesis from lactate is probably due to a marked elevation of the cytosolic NADH/NAD+ ratio resulting in a decline of flux through lactate dehydrogenase.     

Cerebroside elicitors found in diverse phytopathogens activate defense responses in rice plants

Cerebrosides A and C, compounds categorized as glycosphingolipids, were isolated in our previous study from the rice blast fungus (Magnaporthe grisea) as novel elicitors which induce the synthesis of rice phytoalexins. In this paper, these cerebroside elicitors showed phytoalexin-inducing activity when applied to plants by spray treatment and also induced the expression of pathogenesis-related (PR) proteins in rice leaves. This elicitor activity of the cerebrosides showed the structural specificity as that for the induction of phytoalexins. Ceramides prepared from the cerebrosides by removal of glucose also showed the elicitor activity even in lower level compared to the cerebrosides. In field experiments, the cerebroside elicitors effectively protected rice plants against the rice blast fungus, an economically devastating agent of disease of rice in Japan. The cerebrosides elicitors protected rice plants from other disease as well and were found to occur in a wide range of different phytopathogens, indicating that cerebrosides function as general elicitors in a wide variety of rice-pathogen interactions.     

Palmitate oxidation by in vivo and in vitro grown Mycobacterium lepraemurium.

Oxidation of palmitate by Mycobacterium lepraemurium isolated from C3H mice lepromata (in vivo) and also grown on Ogawa egg-yolk medium (in vitro) was investigated. Palmitate was found to be oxidized, after a lag period of about 8 h, by both the in vivo and in vitro grown bacilli. Cell-free extracts prepared from in vivo and in vitro grown cells catalysed an active oxidation of palmitate after a lag period of 3-4 h. The amount of ATP increased, with the increase in time during oxidation of palmitate by the cell-free extracts. The generation of ATP was strongly inhibited by the inhibitors rotenone, antimycin A and cyanide as well as by the uncouplers 2,4-dinitrophenol and 2,6-dibromophenol. These results indicated that oxidation of palmitate by the in vivo and in vitro grown M. lepraemurium is mediated through the respiratory chain using oxygen as the terminal electron acceptor.     

Cyclical mechanical stretch-induced apoptosis in myocytes from young rats but necrosis in myocytes from old rats

Mechanical load as stimulus for apoptosis and necrosis could be responsible for the loss of cardiomyocytes. Ventricular myocytes from young (3 mo) and old (14-24 mo) rats underwent cyclical mechanical stretch (CMS; 5% elongation, 1 Hz) for 24 h. Spontaneous apoptosis was in myocytes from young rats 0.33 +/- 0.12% and from old rats 1.05 +/- 0.35% [Tdt-mediated dUTP nick-end labeling (TUNEL) assay]; associated with a decrease of Bcl-2. CMS increased the apoptosis to 0.58 +/- 0.18% in myocytes from young rats. Western blot analysis showed that CMS reduced Bcl-2 and increased p53 (young rats). Bax was not changed by CMS. These were confirmed by cytochrome c release (31 +/- 13%) and by the enrichment of cytosolic nucleosomes (11 +/- 8%). CMS did not influence the apoptosis in myocytes from old rats (TUNEL assay, Bcl-2, Bax, or p53). CMS did not cause necrosis in myocytes from young rats. CMS increased the number of necrotic cells by showing the cell membrane rupture in myocytes from old rats (50 +/- 13% 5-hexadecanoylaminofluorescein-positive and 38 +/- 6% propidium iodide-positive cells) as well as by measuring the lactate dehydrogenase release. The results suggest that CMS-induced apoptosis in myocytes of young rats but necrosis in myocytes from old rats, which could be attributed to more stress sensitivity of cells from old rats.