Biomedical subjects
G Butler
Publications and source records attributed to G Butler.
The calcium binding proteins parvalbumin and calbindin-D 28K form complementary patterns in the cat superior colliculus.
Parvalbumin (PV) and calbindin-D 28K (CaBP) are calcium binding proteins involved in calcium regulation in the brain. In some regions they coexist in the same neuron, while in other regions they are found in different cell types. We have studied the distribution and morphology of PV labeled neurons in the cat superior colliculus (SC) with antibody immunocytochemistry and compared this labeling to that of CaBP. PV neurons were concentrated in a dense tier within the deep superficial gray and upper optic layers. Scattered PV neurons also were found within the deep layers of SC. By contrast, CaBP neurons were concentrated in three tiers: one within the zonal and upper superficial gray layers, a second within the deep optic and upper intermediate gray layers (IGL), and a third within the deep gray layer. The distribution of PV neurons is thus complementary to that of CaBP neurons, with the CaBP cell tiers bordering the dense tier of PV neurons. PV neurons varied in size and morphology. The average diameter of labeled cells was 20 microns, almost twice the size of CaBP neurons. The cells were predominantly round, vertical fusiform, or stellate, and included the very large neurons found scattered in the IGL. Horseradish peroxidase injections into the lateral geniculate nucleus, the lateral posterior nucleus, the opposite superior colliculus, the dorsal lateral pontine gray nucleus, and two descending pathways--the crossed predorsal bundle and the tecto-ponto-bulbar tracts--each labeled SC neurons that were also labeled by PV. A large percentage (84%) of projection neurons contained PV. This result also differs from CaBP neurons in SC, most of which are interneurons. Two antigen double-label experiments did not produce any cells that contained both PV and CaBP. The two calcium binding proteins thus reveal a unique sublaminar organization in SC that consists of alternating small cell interneuron groups and large cell projection neuron groups.
Parallel pathways of gene regulation: homologous regulators SWI5 and ACE2 differentially control transcription of HO and chitinase.
Two independent pathways of transcriptional regulation that show functional homology have been identified in yeast. It has been demonstrated previously that SWI5 encodes a zinc finger DNA-binding protein whose transcription and cellular localization both are cell cycle regulated. We show that ACE2, whose zinc finger region is nearly identical to that of SWI5, shows patterns of cell cycle-regulated transcription and nuclear localization similar to those seen previously for SWI5. Despite their similarities, SWI5 and ACE2 function in separate pathways of transcriptional regulation. SWI5 is a transcriptional activator of the HO endonuclease gene, whereas ACE2 is not. In contrast, ACE2 is a transcriptional activator of the CTS1 gene (which encodes chitinase), whereas SWI5 is not. An additional parallel between the SWI5/HO pathway and the ACE2/CTS1 pathway is that HO and CTS1 both are cell cycle regulated in the same way, and HO and CTS1 both require the SWI4 and SWI6 transcriptional activators. Overproduction of either SWI5 or ACE2 permits transcriptional activation of the target gene from the other pathway, suggesting that the DNA-binding proteins are capable of binding in vivo to promoters that they do not usually activate. Chimeric SWI5/ACE2 protein fusion experiments suggest that promoter specificity resides in a domain distinct from the zinc finger domain.
Feed efficiency, carcass characteristics, and sensory quality of lambs, with or without prolific ancestry, fed diets with different protein supplements.
Data were collected on 130 intact male lambs fed diets based on roughages supplemented with fish meal, soybean meal, or corn gluten-blood meal and slaughtered at 43 kg live weight. A nonsupplemented group served as a control. The lambs represented Romanov (R), Finnsheep (F), a new breed developed in Canada (DLS), Coopworth (C), and Suffolk (S), three first crosses of DLS with R, F, and Booroola Merino (B), and their backcrosses to DLS. Supplemented lambs consumed 16 to 22% less (P less than .05) silage than control lambs. Average daily gains of lambs fed fish meal (226 g) and corn gluten-blood meal (217 g) were higher and feed conversion ratio (4.99 and 5.11) lower than that of lambs fed soybean meal (189 g and 5.48) or control (186 g and 5.76) diets (P less than .05). The cost of feed per kilogram of gain or per kilogram of lean produced was similar in the four treatments but was between C $.23 and .65 cheaper in the protein-supplemented groups when the number of days to reach slaughter weight was considered. The effect of diet on carcass traits and meat quality were minimal. Meat of lambs fed the three protein supplements was less juicy than that of control lambs. With a few exceptions, most of the significant differences among genetic groups in growth, carcass, and sensory traits were mainly between prolific (R and F) and meat-type breeds (C, S, and DLS). Gain in weight was highest in S lambs (199 g/d), but F and R first crosses were the youngest at slaughter (196 and 198 d). The F lambs had higher dressing and kidney fat percentages than meat-type breeds. The DLS lambs had the largest longissimus muscle area (14.0 cm2), whereas C had the smallest (10.7 cm2). The B crosses had larger longissimus muscle area than R and F crosses. The R lambs had more lean and less fat in the 12th rib, whereas C lambs had the lowest lean and a high bone percentage. The toughest and the most tender roasts were those of R and B crossbred lambs, respectively. Roasts from F lambs had the most intense lamb flavor.
Elimination of mycoplasmas from cell cultures by a novel soft agar technique.
Mycoplasmal infection of cell cultures remains a significant threat to diagnostic and research procedures. In certain defined situations, curing of mycoplasmal infected cultures is a reasonable exercise. Four methods of curing were compared: treatment with BM-cycline, 5 bromouracil, use of specific antisera and treatment of infected cells suspended in soft agar with antibiotics. Antisera treatments were of low efficiency of curing: 50%. None of nine infected cell lines treated with 5-bromouracil were consistently cured of mycoplasmas. The use of BM-cycline was effective for some, but not all lines and required long periods of treatment, 12-21 days. 35 naturally or deliberately infected cultures were treated in soft agar a total of 119 times. This procedure which consisted of suspending infected cultures in soft agar containing appropriate antibiotics resulted in successful mycoplasmal elimination 118/119 times. This soft agar technique took 1-3 days. In separate studies, it was shown that certain Mycoplasma fermentans strains were resisted to this and other curing methods. This may be due to their intracellular location. Such strains may be more amenable to antibiotics that penetrate mammalian cells. It is concluded that the soft agar technique is a rapid, efficient and reliable method to eliminate cell culture mycoplasmas.
Comparison of behavior therapy and cognitive behavior therapy in the treatment of generalized anxiety disorder.
In a controlled clinical trial, 57 Ss meeting DSM-III-R criteria for generalized anxiety disorder, and fulfilling an additional severity criterion, were randomly allocated to cognitive behavior therapy (CBT), behavior therapy (BT), or a waiting-list control group. Individual treatment lasted 4-12 sessions; independent assessments were made before treatment, after treatment, and 6 months later, and additional follow-up data were collected after an interval of approximately 18 months. Results show a clear advantage for CBT over BT. A consistent pattern of change favoring CBT was evident in measures of anxiety, depression, and cognition. Ss were lost from the BT group, but there was no attrition from the CBT group. Treatment integrity was double-checked in England and in Holland, and special efforts were made to reduce error variance. Possible explanations for the superiority of CBT are discussed.
ACE2, an activator of yeast metallothionein expression which is homologous to SWI5.
Transcription of the Saccharomyces cerevisiae metallothionein gene CUP1 is induced in response to high environmental levels of copper. Induction requires the ACE1 gene product, which binds to specific sites in the promoter region of the CUP1 gene. In this study, we found that deleting the entire coding sequence of the ACE1 gene resulted in a decrease in basal-level transcription of CUP1 to low but detectable levels and conferred a copper-sensitive phenotype to the cells. We have isolated a gene, designated ACE2, which when present on a high-copy-number plasmid suppresses the copper-sensitive phenotype of an ace1-deletion strain. The presence of multiple copies of the ACE2 gene enhanced expression of an unlinked CUP1-lacZ fusion integrated in the yeast genome and resulted in an increase in the steady-state levels of CUP1 mRNA in an ace1-deletion background. A large deletion of the coding region of the genomic copy of ACE2 resulted in a decrease in steady-state levels of CUP1 mRNA, indicating that ACE2 plays a role in regulating basal-level expression of CUP1. The ACE2 open reading frame encodes a polypeptide of 770 amino acids, with putative zinc finger structures near the carboxyl terminus. This protein is 37% identical to the SWI5 gene product, an activator of HO gene transcription in S. cerevisiae, suggesting that ACE2 and SWI5 may have functional similarities.
Heat shock transcription factor activates transcription of the yeast metallothionein gene.
In the yeast Saccharomyces cerevisiae, transcription of the metallothionein gene CUP1 is induced by copper and silver. Strains with a complete deletion of the ACE1 gene, the copper-dependent activator of CUP1 transcription, are hypersensitive to copper. These strains have a low but significant basal level of CUP1 transcription. To identify genes which mediate basal transcription of CUP1 or which activate CUP1 in response to other stimuli, we isolated an extragenic suppressor of an ace1 deletion. We demonstrate that a single amino acid substitution in the heat shock transcription factor (HSF) DNA-binding domain dramatically enhances CUP1 transcription while reducing transcription of the SSA3 gene, a member of the yeast hsp70 gene family. These results indicate that yeast metallothionein transcription is under HSF control and that metallothionein biosynthesis is important in response to heat shock stress. Furthermore, our results suggest that HSF may modulate the magnitude of individual heat shock gene transcription by subtle differences in its interaction with heat shock elements and that a single-amino-acid change can dramatically alter the activity of the factor for different target genes.
The cytochrome b of the sea urchin Paracentrotus lividus is naturally resistant to myxothiazol and mucidin.
The ubiquinol:cytochrome c reductase activity of Paracentrotus lividus mitochondria is relatively insensitive to the specific inhibitors myxothiazol and mucidin. The I50 of myxothiazol and mucidin are three and two orders of magnitude higher, respectively, in P. lividus than in bovine heart mitochondria. The natural resistance of the P. lividus reductase to these inhibitors can be correlated with a single amino replacement, an alanine for a glycine at position 143, in the sequence of cytochrome b. This position is located in a conserved region of the molecule, believed to be important in the oxidation of ubiquinol by the reductase.
TUF factor binds to the upstream region of the pyruvate decarboxylase structural gene (PDC1) of Saccharomyces cerevisiae.
The upstream activation site of the pyruvate decarboxylase gene, PDC1, of Saccharomyces cerevisiae contains an RPG box, and mediates the increase in expression of a PDC1-lacZ fusion gene during growth on glucose. Oligonucleotide replacement experiments indicate that the RPG box functions as an absolute activator of expression, but other elements (possibly CTTCC repeats) are required for carbon source regulation, and maximal expression. Gel retardation and oligonucleotide competition experiments suggest that the DNA binding factor TUF interacts with the RPG box in the upstream region of PDC1. Binding of TUF factor is not carbon source dependent in in vitro experiments, and is probably not responsible for glucose induction of PDC1 expression.
Recovery of an epitope recognized by a novel monoclonal antibody from airway lavage during experimental induction of chronic bronchitis.
Prolonged exposure of dogs to high concentrations of SO2 gas results in a syndrome with many of the characteristics of human chronic bronchitis, including cough and chronic mucous hypersecretion as well as airway obstruction. We developed and used a novel monoclonal antibody, GB-4B, raised against epithelial glycoprotein isolated from human hypersecretory mucus to probe airway lavage samples from dogs before and during prolonged exposure to SO2 gas. There were relatively low mean titers of the epitope recognized by GB-4B in airway lavage fluid as evidenced by enzyme-linked immunosorbent assay before exposure to SO2 gas. After 25 to 50 wk of SO2 exposure, the dogs showed a significant increase in pulmonary resistance and there was a significant increase in the titer of the epitope in the airway lavage fluid. Using the same antibody immunohistochemical analysis of airway tissues from SO2-exposed dogs revealed patchy staining of the mucous glands and airway secretory cells and dense staining along the airway surface; airway tissue from control dogs and one SO2-exposed dog whose lavage fluid did not contain the epitope showed little or no staining. These data demonstrate that similar mucin epitopes appear in airway lavage fluid under hypersecretory conditions in both animals and humans. The epitope may have utility as a marker of chronic mucous hypersecretion.
Plasma and tissue vitamin E concentrations in sheep after administration of a single intraperitoneal dose of dl-alpha-tocopherol.
Blood plasma and tissue Vitamin E concentrations were determined in 20 sheep following a single i.p. dose of 5 g of dl-alpha-tocopherol. In addition, five sheep were used as controls (no treatment and killed at d 0). From the 20 vitamin E-dosed sheep, 4 were slaughtered on d 3, 6, 10, 15 and 28 after dosing. There was a significant time effect in alpha-tocopherol concentrations in all tissues. In most tissues, the peak alpha-tocopherol concentration was at 3 d postdosing. Uptake varied among the different tissues examined. Three days postdosing, a large uptake of vitamin E by the liver was observed; this supports the concept that hepatic tissues are a target organ for vitamin E action. Also at 3 d uptake was pronounced in spleen and lung. Vitamin E concentrations in the other body tissues at d 3 postdosing increased considerably, but to a lesser degree than those in liver, spleen and lung. Vitamin E concentration in all tissues declined 3 d after i.p. dosing.
Plasma vitamin E response in sheep dosed intraruminally or intraduodenally with various alpha-tocopherol compounds.
A total of 24 sheep were used to study two routes of vitamin E administration: intraruminal (IR) and intraduodenal (ID). For each route of administration, 12 sheep were divided into 3 groups of 4, and each group received one of the following vitamin E treatments (mg/kg body weight): treatment 1, D-alpha-tocopherol (60 mg); treatment 2, D-alpha-tocopherol acetate (70 mg); treatment 3, DL-alpha-tocopherol (90 mg). The results showed that two major indices of bioavailability, i.e. concentration maximum and the area under the plasma concentration time-curve, were higher following IR administration than ID administration for all three vitamin E compounds used in this experiment. In sheep dosed intraruminally no difference was observed between the three vitamin E preparations in their plasma vitamin E response. For the ID administration the peak of plasma vitamin E concentration was lower after D-alpha-tocopherol acetate dosing than with the other two preparations. The results indicated a beneficial effect of rumen environment on the availability of vitamin E supplements.
Identification of an upstream activation site in the pyruvate decarboxylase structural gene (PDC1) of Saccharomyces cerevisiae.
The upstream region of the Saccharomyces cerevisiae pyruvate decarboxylase structural gene, PDC1, has been isolated and fused to the indicator gene Escherichia coli lacZ. 1.2 kb of the upstream region has been sequenced. The PDC1-lacZ fusion has been integrated at the ura3-52 locus in the yeast genome, and has a basal level of expression on ethanol. On glucose media this level is increased 30-50 fold. An upstream activation site, UASpdc, between 793 and 535 bp upstream from the ATG of PDC1, which mediates the response to glucose has been identified by deletion analysis. The UASpdc contains a consensus RPG box, originally identified in ribosomal protein genes (Leer et al. 1985). The function of UASpdc is independent of distance from the ATG. There is also an upstream repressing sequence located between 535 and 385 bp upstream from the translational start of PDC1.
Predicting response to anxiety management in patients with generalised anxiety disorders.
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Klinefelter's syndrome: an analysis of the origin of the additional sex chromosome using molecular probes.
The results of our study of the origin of the additional X chromosome in 39 males with a 47,XXY chromosome constitution are reported. We used a total of 20 X-linked RFLPs and successfully determined the origin of all 32 patients in whom DNA from both parents was available, and a further 3 in whom DNA was available from the patient and mother only. Males whose additional X chromosome was maternal in origin were further investigated using an X-linked centromere specific probe to determine the cell division at which the error occurred. Our results showed 53% of the non-disjunction to be attributable to pat mei I errors, 34% to mat mei I errors, 9% to mat mei II errors and 3% to a post-zygotic mitotic error. In the great majority of patients resulting from an error of maternal meiosis there was clear evidence of recombination involving the non-disjoined chromosomes, suggesting that absence of recombination is not an important aetiological factor in non-disjunction of the X chromosome in female meiosis. There was no alteration of parental age associated with the paternally derived 47,XXY males but a marked increase in maternal age among the maternally derived 47,XXY males, the increase being associated with mat mei I but not mat mei II errors. The proportion of paternally and maternally derived cases was similar among different ascertainment classes, suggesting that there is no dramatic effect of parental origin of the additional X chromosome on the phenotype of 47,XXY males.
Proper drug testing procedures require lab analysis accuracy.
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Anxiety management: developing effective strategies.
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