Identification of the peptide bond cleaved during activation of human Clr

CNBr cleavage of unreduced proenzyme Clr yielded fragment CP2b, isolated by gel filtration and highpressure gel permeation chromatography. This fragment (˜ M_τ 55000) comprised at least 4 disulphidelinked peptides, which were separated by gel filtration after reduction and alkylation. Peptide CP2bRA4, overlapping the A- and B-chain regions in proenzyme Clr was digested by V8 staphylococcal protease, and the digest separated by reversed-phase HPLC. N-terminal sequence analysis of peptide CP2bRA4SP9 established that Clr activation involves the cleavage of a single Arg-Ile bond, located in the sequence: Gln-Arg-Gln-Arg-Ile-Ile-Gly-Gly     

Embryonic development times of entomostracan zooplankton from Lake le Roux (Orange River,South Africa), and their possible relationships to seasonal succession

Embryonic development times of the principal entomostracan zooplankton of Lake le Roux, Orange River, South Africa, are described. Durations decreased monotonically with temperature over the thermal tolerance range and are described using Blehrádek's equation. The present data are compared with other observations on rate-temperature responses of warm-water and temperate zooplankton. Small, but ecologically significant, differences are discerned. The possible significance of species-specific rate-temperature responses to the phenomenon of seasonal succession is explored.     

Partial Purification and Properties of Human Brain Aldehyde Dehydrogenases

Acetaldehyde and biogenic aldehydes were used as substrates to investigate the subcellular distribution of aldehyde dehydrogenase activity in autopsied human brain. With 10 microM acetaldehyde as substrate, over 50% of the total activity was found in the mitochondrial fraction and 38% was associated with the cytosol. However, with 4 microM 3,4-dihydroxyphenylacetaldehyde and 10 microM indoleacetaldehyde as substrates, 40-50% of the total activity was found in the soluble fraction, the mitochondrial fraction accounting for only 15-30% of the total activity. These data suggested the presence of distinct aldehyde dehydrogenase isozymes in the different compartments. The mitochondrial and cytosolic fractions were, therefore, subjected to salt fractionation and ion-exchange chromatography to purify further the isozymes present in both fractions. The kinetic data on the partially purified isozymes revealed the presence of a low Km isozyme in both the mitochondria and the cytosol, with Km values for acetaldehyde of 1.7 microM and 10.2 microM, respectively. However, the cytosolic isozyme exhibited lower Km values for the biogenic aldehydes. Both isozymes were activated by Mg2+ and Ca2+ in phosphate buffers (pH 7.4). Also, high Km isozymes were found in the mitochondria and in the microsomes.     

Immunoassay of the Neuronal and Neuroendocrine Marker PGP 9.5 in Human Tissues

An inhibitory, coated-well immunoassay for the neurone-specific protein PGP 9.5 has been devised and used to measure the concentrations of the protein in human tissues. Concentrations of PGP 9.5 between 40 ng/ml and 10 micrograms/ml could be measured using this assay. In brain PGP 9.5 was present at 100.58 +/- 16.18 micrograms/mg protein. Of the other organs examined only kidney and testis showed significant concentrations of PGP 9.5 (3.97 +/- 0.87 microgram/mg protein and 3.25 +/- 0.36 microgram/mg protein, respectively). All other organs contained less than 2% of the brain level. The tissue levels determined by coated-well immunoassay confirmed the tissue specificity of PGP 9.5 originally determined by high-resolution two-dimensional gel electrophoresis.     

Mobility of the conceptus and uterine contractions in the mare

Location of the embryonic vesicle within the uterus of mares was recorded every. five minutes for two consecutive hours (25 location determinations per trial) in three experiments. In Experiment 1 (n=7), the number of location changes among nine uterine segments (three body segments and three segments for each horn) was greater (P<0.05) on Day 13 than on Day 10. The vesicle was located in the body more frequently (P<0.05) and tended (P<0.1) to move to a more caudal position more frequently on Day 10 than on Day 13. Fixation occurred on Day 15 in four of seven mares and on Day 16 in the remaining three mares. The number of location changes was not significantly different between two days prior to fixation and one day prior to fixation. In Experiment 2, the effect of clenbuterol, a B₂ sympathomimetic blocker of uterine contractions, was studied on Days 12 or 13 of pregnancy. Location changes occurred less frequently (P<0.05) in treated mares (n=9) than in controls (n=10), indicating involvement of uterine contractions in the mobility of the embryonic vesicle. In Experiment 3, when the initial direction of location changes was caudal within a horn and cranial within the uterine body, the vesicle was more likely (P<0.05) to continue moving in the same direction than in the opposite direction. However, when the direction within a horn was cranial, the next location change was as likely to be in the opposite direction as in the same direction (not significantly different from equality). When the direction within the uterine body was caudal, the next location change was more likely (P<0.05) to be in the opposite direction.     

丁酸对体外培养的人鼻咽癌上皮细胞的影响

无论是自发的、病毒引起的或致癌物诱发的恶性转化的哺乳类细胞的体外培养,其形态多发生改变,总是变得近似圆形,边缘突起短而少,细胞致密和折光性强,同时失去生长接触抑制,降低细胞与细胞之间和细胞与生长底物之间的粘着性等特性。近年报道了关于短链脂肪酸如丁酸(或丁酸钠)对细胞能产生明显的影响,能抑制培养细胞的分裂,可诱发一些上皮性细胞产生形态的改变,可使转化的细胞     

Ecdysteroid levels during adult reproductive and embryonic developmental stages of Sarcophaga bullata (Sarcophagidae: Diptera)

Ecdysteroid levels were determined during the period of the adult reproductive cycle in the ovovivparous fleshfly Sarcophaga bullata. Low levels were found in males during the 10 days following eclosion while the entire adult female showed a significant peak of ecdysteroid activity at 190 h post eclosion (i.e. during embryogenesis). When the female reproductive tract was analyzed it was observed that the ovaries became vitellogenic a short time after a protein meal was offered at 96 h post eclosion: at 140 h, ecdysteroid activity was recorded in the developing oöcytes. The major peak of ecdysteroids during the reproductive cycle was found in developing embryos at 190 h. The significance of these releases of ecdysteroids is discussed in relation to major embryonic developmental events.     

Amino Acids in Plasma and CSF and Monoamine Metabolites in CSF: Interrelationship in Healthy Subjects

Plasma and cerebrospinal fluid (CSF) concentrations of amino acids were measured in 65 healthy volunteers (50 men and 15 women). The CSF levels of the monoamine metabolites homovanillic acid (HVA), 3-methoxy-4-hydroxyphenylethylene glycol (MOPEG), and 5-hydroxyindoleacetic acid (5-HIAA) were also determined. Sex differences were observed in both plasma and CSF amino acid levels as well as in the relationship between these concentrations. No significant correlations were observed between the CSF levels of HVA and 5-HIAA, and the concentrations of their precursor amino acids in either plasma or CSF. The MOPEG level in CSF correlated positively with the plasma concentrations of several amino acids.     

Human Brain Phenol Sulfotransferase: Biochemical Properties and Regional Localization

Phenol sulfotransferase (PST) catalyzes the sulfate conjugation of catecholamines and phenol and catechol drugs. The human blood platelet contains a thermolabile (TL) form of PST that catalyzes the sulfate conjugation of dopamine and other monoamines and a thermostable (TS) form that catalyzes the sulfate conjugation of micromolar concentrations of phenol and p-nitrophenol. Experiments were performed to determine whether the brain contains forms of PST analogous to the TL and TS forms found in the human platelet, and to determine whether there are regional variations in human brain PST activity. We found that the human brain contains at least two forms of PST, forms that are similar to the platelet TS and TL forms of the enzyme with respect to substrate specificity, apparent Km constants, thermal stability, and sensitivity to inhibitors. Optimal conditions were determined for the measurement of these two activities in brain homogenates. The stability of PST activities in the human brain after death was determined in five samples of cerebral cortex that were obtained during clinically indicated neurosurgical procedures. An average of 76 +/- 8% and 80 +/- 9% (mean +/- SEM) of the basal TL and TS PST activities, respectively, remained in these five samples of cerebral cortex after 8 h of storage under simulated post-mortem conditions. Six human brains were then obtained less that 8 h after death from patients who had no neurological disease prior to death. The mean activities of the TL and TS forms of PST were measured in 17 different regions of the six brains. If the pituitary was excluded from consideration, TL and TS PST activities both varied approximately fivefold among these regions, and both activities were highest in cerebral cortex. However, the average TS activity in the anterior pituitary, a tissue of non-neural origin embryologically, was 6.5-fold greater than the highest average TS PST activity found in cerebral cortex.     

Evidence for Feedback Regulation of Glutamate Decarboxylase by γ-Aminobutyric Acid

Abstract: The concentration of γ-aminobutyric acid (GABA) in the human ovary and the capacity of a membrane preparation from the same organ to bind [³H]GABA specifically were examined. The GABA concentration in the ovary was found to be 214 ± 66 nmol/g frozen tissue (mean ± SEM of six independent determinations). Moreover, a single population of high-affinity GABA binding sites has been identified in the ovarian membranes. The apparent dissociation constant ( K _d) and maximum binding capacity ( B _max) were 38.3 n M and 676 fmol/mg protein, respectively. The specific binding of [³H]GABA was displaced by muscimol, unlabelled GABA, or (+)bicuculline, but was unaffected by (±)baclofen and picrotoxin. The present results show that GABA and an extremely high density of GABA_A receptor binding sites are present in the human ovary, indicating a physiological significance of this amino acid in the female reproductive system.