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T G Cooper

Publications and source records attributed to T G Cooper.

At least 127 records · Page 7Linked to original sources
PubMed1995

Regulation of the transport of immunoglobulin G into the male rat reproductive tract.

The net movements of IgG in the rat testis and epididymis were studied using physiological techniques in conditions of anticipated subnormal organ concentrations of testosterone (T). The volume of extracellular fluid accessible to IgG per gram whole testis (Veq, microliter/g) did not change but that per gram interstitial tissue decreased in intact rats treated with T to suppress gonadotrophin secretion. Estimates of surface areas of the exchange vessels (S, cm2/g whole tissue) did not change in any of the organs studied. The speed at which equilibrium between tissue extracellular fluid and serum was reached (magnitude of K, min-1), lymph flow (QL, microliter/g/min) and estimates of microvascular permeability (P, nm/min) to IgG decreased in the testis of T-treated intact rats. In these animals, magnitude of K and P were higher than normal in the cauda. After castration, magnitude of K and P decreased to near zero in the caput and corpus regions. The changes in magnitude of K and P due to castration could partly be prevented by T supplementation in the caput, but not in the corpus, where Veq was supra-normal after T treatment. This difference in endothelial response between caput and corpus indicates differences in control of endothelium and epithelium along the length of the epididymal duct. In the testis of efferent duct-ligated (EDL) rats, P was lower than that in the intact rats, whereas in the corpus epididymidis of EDL rats, Veq and P were higher than normal. The supra-normal Veq values in the corpus epididymidis of EDL and castrated T-treated rats suggest that the epithelial barrier leaks in the corpus when luminal testosterone is low. It is concluded that during testosterone administration to normal rats, the testicular and epididymal epithelial barrier remains intact, that trans-endothelial transport of IgG and extracellular fluid volume decrease in the testicular interstitium and that in conditions of low serum T, the endothelium of microvessels in the caput and corpus epididymidis forms a barrier to IgG.

Animals↗
PubMed1995

Genetic evidence for Gln3p-independent, nitrogen catabolite repression-sensitive gene expression in Saccharomyces cerevisiae.

The expression of many nitrogen catabolic genes decreases to low levels when readily used nitrogen sources (e.g., asparagine and glutamine) are provided in the growth medium; this physiological response is termed nitrogen catabolite repression (NCR). Transcriptional activation of these genes is mediated by the cis-acting element UASNTR and the trans-acting factor Gln3p. A second protein encoded by URE2 possesses the genetic characteristics of a negative regulator of nitrogen catabolic gene expression. A third locus, DAL80, encodes a repressor that binds to sequences required for Gln3p-dependent transcription and may compete with Gln3p for binding to them. These observations are consistent with an NCR regulatory pathway with the structure environmental signal-->Ure2p-->(Gln3p/Dal80p)-->UASNTR operation-->NCR-sensitive gene expression. If NCR-sensitive gene expression occurs exclusively by this pathway, as has been thought to be the case, then the NCR sensitivity of a gene's expression should be abolished by a ure2 delta mutation. This expectation was not realized experimentally; the responses of highly NCR-sensitive genes to ure2 delta mutations varied widely. This suggested that NCR was not mediated exclusively through Ure2p and Gln3p. We tested this idea by assaying GAP1, CAN1, DAL5, PUT1, UGA1, and GLN1 expression in single, double, and triple mutants lacking Gln3p, Dal80p, and/or Ure2p. All of these genes were expressed in the triple mutant, and this expression was NCR sensitive for four of the six genes. These results indicate that the NCR regulatory network consists of multiple branches, with the Ure2p-Gln3p-UASNTR pathway representing only one of them.

Asparagine↗
PubMed1995

The Saccharomyces cerevisiae Leu3 protein activates expression of GDH1, a key gene in nitrogen assimilation.

The Leu3 protein of Saccharomyces cerevisiae has been shown to be a transcriptional regulator of genes encoding enzymes of the branched-chain amino acid biosynthetic pathways. Leu3 binds to upstream activating sequences (UASLEU) found in the promoters of LEU1, LEU2, LEU4, ILV2, and ILV5. In vivo and in vitro studies have shown that activation by Leu3 requires the presence of alpha-isopropylmalate. In at least one case (LEU2), Leu3 actually represses basal-level transcription when alpha-isopropylmalate is absent. Following identification of a UASLEU-homologous sequence in the promoter of GDH1, the gene encoding NADP(+)-dependent glutamate dehydrogenase, we demonstrate that Leu3 specifically interacts with this UASLEU element. We then show that Leu3 is required for full activation of the GDH1 gene. First, the expression of a GDH1-lacZ fusion gene is three- to sixfold lower in a strain lacking the LEU3 gene than in an isogenic LEU3+ strain. Expression is restored to near-normal levels when the leu3 deletion cells are transformed with a LEU3-bearing plasmid. Second, a significant decrease in GDH1-lacZ expression is also seen when the UASLEU of the GDH1-lacZ construct is made nonfunctional by mutation. Third, the steady-state level of GDH1 mRNA decreases about threefold in leu3 null cells. The decrease in GDH1 expression in leu3 null cells is reflected in a diminished specific activity of NADP(+)-dependent glutamate dehydrogenase. We also demonstrate that the level of GDH1-lacZ expression correlates with the cells' ability to generate alpha-isopropylmalate and is lowest in cells unable to produce alpha-isopropylmalate. We conclude that GDH1, which plays an important role in the assimilation of ammonia in yeast cells, is, in part, activated by a Leu3-alpha-isopropylmalate complex. This conclusion suggests that Leu3 participates in transcriptional regulation beyond the branched-chain amino acid biosynthetic pathways.

Base Sequence↗
PubMed1995

The upstream region of the FOX3 gene encoding peroxisomal 3-oxoacyl-coenzyme A thiolase in Saccharomyces cerevisiae contains ABF1- and replication protein A-binding sites that participate in its regulation by glucose repression.

Expression of the FOX3 gene, which encodes yeast peroxisomal 3-oxoacyl-coenzyme A thiolase, can be induced by oleate and repressed by glucose. Previously, we have shown that induction was mediated by an oleate response element. Just upstream of this element a negatively acting control region that mediated glucose repression was found. In order to study this negative control region, we carried out DNA-binding assays and analyzed phenotypes of mutations in this region and in the trans-acting factor CAR80, which is identical to UME6. DNA-binding assays showed that two multifunctional yeast proteins, ABF1 and RP-A, interacted with the negative control element independently of the transcriptional activity of the FOX3 gene. ABF1 and RP-A, the latter being identical to BUF, were able to bind to DNA independently of one another but also simultaneously. The phenotypes of mutations in either DNA-binding sites of ABF1, RP-A, or both, which affected the DNA binding of these factors in vitro, indicated that these sites and the proteins that interact with them participate in glucose repression. The involvement of the RP-A site in glucose repression was further supported by our observation that the CAR80 gene product, which is required for repression mediated by the RP-A site, was essential for maintenance of glucose repression. In addition to the RP-A site in the FOX3 promoter, similar sequences were observed in other genes involved in peroxisomal function. RP-A proved to bind to all of these sequences, albeit with various affinities. From these results it is concluded that the ABF1 and RP-A sites are being required in concert to mediate glucose repression of the FOX3 gene. In addition, coordinated regulation of expression of genes involved in peroxisomal function in response to glucose is mediated by proteins associated with the RP-A site, probably RP-A and CAR80.

Acetyl-CoA C-Acyltransferase↗
PubMed1995

Effects of the male antifertility agent ornidazole on sperm function in vitro and in the female genital tract.

Infertility is induced in male rats by oral administration of 400 mg ornidazole kg-1 body mass day-1 within 10 days, without drastic effect on the motility of epididymal spermatozoa obtained after 15 days of treatment. Spermatozoa were recovered from the female tract 9 h after mating with males treated with ornidazole for 10 days to identify the cause of infertility. The number and motility of spermatozoa in the uterus indicated normal seminal deposition. Spermatozoa could pass the utero-tubal junction but total numbers of spermatozoa and their velocities in the oviductal isthmus were significantly lower than those recovered from females mated to fertile, control males fed vehicle alone. However, the number of spermatozoa recovered from the ampulla was not different from control and the number of spermatozoa that had penetrated the cumulus mass was lower and none of the eggs was fertilized by spermatozoa from rats treated with ornidazole. The ability of cauda epididymal spermatozoa from ornidazole-fed rats to penetrate viscous media in vitro was lower than that of the controls. When incubated in media containing different concentrations of substrates and ions, a reduction in movement of spermatozoa was observed when glucose was the only exogenous substrate or after preincubation in substrate-free medium. These results indicate that the antifertility action of orally administered ornidazole occurs via damage to spermatozoa in the upper regions of the female tract, possibly reflecting a failure in capacitation as a result of reduced energy production.

Animals↗
PubMed1994

DNA binding site specificity of the Neurospora global nitrogen regulatory protein NIT2: analysis with mutated binding sites.

NIT2, a positive-acting regulatory protein in Neurospora crassa, activates the expression of a series of unlinked structural genes that encode nitrogen catabolic enzymes. NIT2 binding sites in the promoter regions of nit3, alc and lao have at least two GATA sequence elements. We have examined the binding affinity of the NIT2 protein for the yeast DAL5 wild-type upstream activation sequence UASNTR, which contains two GATA elements, and for a series of mutated binding sites, each differing from the wild-type site by a single base. Substitution for individual nucleotides within 5' or 3' sequences that flank the GATA elements had only modest effects upon NIT2 binding. In contrast, nearly all substitutions within the GATA elements almost completely eliminated NIT2 binding, demonstrating the importance of the GATA sequence for NIT2 binding. Four high-affinity binding sites for the NIT2 protein were found within a central region of the nit-2 gene itself.

Base Sequence↗
PubMed1994

Maturation of hamster epididymal sperm motility and influence of the thiol status of hamster and rat spermatozoa on their motility patterns.

A method for objective quantification of hamster sperm movement parameters as an indicator of maturation along the epididymis was established using a computerised system. Analysis of spermatozoa released into medium from five epididymal regions showed that the most drastic increases in percentage motility and curvilinear velocity (VCL) occurred from the distal corpus to the beginning of the proximal cauda and in straight-line velocity (VSL) from the beginning to a more distal site within the proximal cauda region. Both high osmolarity (400 mOsm/kg) and the thiol-oxidising agent diamide (10 microM) increased flagellar straightness of distal corpus spermatozoa, but VSL was increased only with the latter. The thiol-reducing agent dithiothreitol (DTT, 1mM) stimulated and maintained percentage motility and velocities of spermatozoa from the caput, stimulated only percentage motility of distal corpus sperm, but decreased velocities of those from the proximal cauda in prolonged incubation. In rats, diamide increased path straightness but not velocities of caput spermatozoa and yet caused immotility within 15 min, whereas DTT prolonged the maintenance of in vitro motility. The slight increases in kinematic parameters in the presence of DTT were enhanced by a 2-min preincubation with diamide. The finding that the effects of DTT and diamide were not compensatory suggests that the influence of the SH/S-S status on sperm movement is multifaceted, with decreasing sensitivity to stimulation upon sperm maturation.

Animals↗
PubMed1994

Immunology of the testicular excurrent ducts.

The sperm autoantigen concentration in the epididymis equals or exceeds that in the testis. This makes the epididymis a probable site of initiation of an antisperm autoimmune response. The mechanisms regulating antisperm antibody formation in the testicular excurrent ducts and some related aspects with clinical interest are reviewed.

Animals↗
PubMed1994

Magnetic resonance signals in healing menisci: an experimental study in dogs.

This study examined the magnetic resonance imaging (MRI) signal from healing menisci in adult dogs. Complete, full-thickness radial tears were created in the medial menisci of adult dogs and allowed to heal spontaneously. Menisci were harvested and examined at 8, 12, and 26 weeks after injury using a spin-echo imaging sequence. After imaging, the menisci were processed for routine histologic examination. The results of the study demonstrated that the repair tissue consistently yielded an increased MRI signal as compared with the normal meniscal tissue. This increased signal persisted at 26 weeks even though the repair tissue had modulated from fibrovascular scar tissue into fibrocartilage. The results of this study confirm the clinical impression that normal fibrovascular repair tissue in the healing meniscus can emit persistently increased signals on MRI examination. However, the long-term history of these signals remains unknown.

Animals↗
PubMed1994

Basal cells of the human epididymis--antigenic and ultrastructural similarities to tissue-fixed macrophages.

Very little is known about the basal cells in the epididymal epithelium. Their function is unclear, although they are present in all mammalian epididymides studied. The corpus epididymides from five patients undergoing castration because of prostatic carcinoma were fixed and processed for electron microscopy. Basal cells were characterized by a slightly heterochromatic nucleus with prominent nucleolus, pale round mitochondria, dispersed endoplasmic reticulum, and sparse Golgi apparatus; they were often rich in lipofuscin inclusions, possibly originating from principal cells. Some peritubular macrophages in close proximity to the epithelium were structurally similar to basal cells. Immunohistochemical staining revealed in the epididymides of another ten patients that the basal cells were recognized by the monoclonal antibody (mAb) 25F9 against mature, tissue-fixed macrophages but not by mAbs 27E10 or RM3/1, which were against activated macrophages usually found in acute or late inflammation, respectively. On the basis of the present findings, as a working hypothesis a scavenging role of the basal cells in a local immune defense mechanism is proposed, in which antigenic products (possibly of sperm degradation), taken up by the principal cells, would be phagocytosed by the basal cells. It could be inferred that when the basal cells are overloaded, they would leave the epithelium to be replenished by tissue-fixed macrophages.

Aged↗
PubMed1994

Improvement in quality of cryopreserved human spermatozoa by swim-up before freezing.

Selecting a population of spermatozoa by the swim-up technique yields, after freezing and thawing, a population of cells that contains proportionally more spermatozoa which are morphologically normal, fewer spermatozoa with damaged tail membranes, and a greater percentage of progressively motile spermatozoa with greater velocities and amplitudes of head displacement than those obtained after freezing and thawing the same semen samples in the normal way. This pattern was found for the semen from 10 patients and five volunteers. However, the cells selected by swim-up were as susceptible to the stresses caused by freezing and thawing as unselected spermatozoa in the original semen sample, and the improvement came from the better quality of the initial sample.

Adult↗
PubMed1994

Development of sperm motility patterns in the murine epididymis.

The maturation of sperm motility in the epididymis of the mouse was assessed using a computer-assisted sperm analysis system. Spermatozoa were immotile in the most proximal regions of the epididymis but developed motility rapidly in the proximal caput epididymis; the percentage motility remained high thereafter. In the caput, flagellar beat was erratic with little progression, but in the proximal corpus region circular movement patterns were reflected in reduced linearity (LIN) and straightness (STR) of the sperm tracks, although velocities were little changed and wobble (WOB) increased. In the mid-corpus region, however, all velocities, LIN, STR and WOB, increased markedly. In more distal regions there was little change in these parameters. Distribution curves of the kinematic parameters of spermatozoa obtained from each region indicated that the most heterogeneous population was that from the mid-corpus epididymis; the most homogeneous was that from the mid-cauda region. Individual sperm tracks revealed slowly progressing spermatozoa in the distal caput, transforming to motion in small circles, interrupted by more linear progression. More distally, linear progression was interrupted by looping movements and a generally progressive path was observed thereafter, with less deviation from the average path as the spermatozoa matured. Spermatozoa displaying motion compatible with passing the uterotubal junction were first found in the proximal corpus epididymis, in agreement with earlier in-vivo fertilization studies on where fertilizing capacity is achieved with epididymal spermatozoa.

Animals↗
PubMed1994

The UGA4 UASNTR site required for GLN3-dependent transcriptional activation also mediates DAL80-responsive regulation and DAL80 protein binding in Saccharomyces cerevisiae.

Expression of the nitrogen catabolic genes in Saccharomyces cerevisiae, including those of the gamma-aminobutyric acid (UGA) and allantoin (DAL) pathways, is regulated positively by the GLN3 protein and negatively by the DAL80 protein. The deduced sequences of the DAL80 and GLN3 proteins contain a zinc finger motif homologous to those shown to bind GATA sequences. In addition, DAL80 protein has been directly shown to bind to a pair of GATA-containing sequences (URSGATA) in vitro, and a pair of GATA-containing sequences (UASNTR) is required for GLN3-dependent transcriptional activation in a heterologous expression vector. We demonstrate here that the GATA-containing sites upstream of UGA4 required for optimal GLN3-dependent transcriptional activation also mediate DAL80 protein binding in vitro and DAL80-responsive regulation in vivo.

4-Aminobutyrate Transaminase↗
PubMed1994

The URE2 protein regulates nitrogen catabolic gene expression through the GATAA-containing UASNTR element in Saccharomyces cerevisiae.

Many of the gene products that participate in nitrogen metabolism are sensitive to nitrogen catabolite repression (NCR), i.e., their expression is decreased to low levels when readily used nitrogen sources such as asparagine are provided. Previous work has shown this NCR sensitivity requires the cis-acting UASNTR element and trans-acting GLN3. Here, we extend the analysis to include the response of their expression to deletion of the URE2 locus. The expression of these nitrogen catabolic genes becomes, to various degrees, NCR insensitive in the ure2 deletion. This response is shown to be mediated through the GATAA-containing UASNTR element and supports the current idea that the NCR regulatory circuit involves the following steps: environmental signal-->URE2-->GLN3-->UASNTR operation-->NCR-sensitive gene expression. The various responses of the nitrogen catabolic genes' expression to deletion of the URE2 locus also indicate that not all NCR is mediated through URE2.

Allantoin↗
PubMed1994

Changes in magnetic resonance images of muscle depend on exercise intensity and duration, not work.

Echo-planar magnetic resonance imaging was used to study the effect of exercise rate and duration on magnetic resonance imaging signal intensity (SI) of anterior tibialis muscle in normal human subjects (mean age 35 yr, n = 6). Axial midcalf echo-planar images (repetition time/echo time = 6,000/45, acquisition time = 80 ms) were acquired every 6 s for 1 min before and during 15 min of dynamic ankle dorsiflexion exercise (peak force 36% of 1 repetition maximum) at 10, 20, and 30 contractions/min. At each rate, muscle SI rose along an approximately exponential time course (mean time constant 1.8 min) toward a plateau that was linearly dependent on force times contraction rate (r = 0.64, P < 0.01) but varied significantly among subjects. The results confirm previous reports that changes in muscle SI correlate with exercise intensity, but not with total work performed, over a submaximal range of exercise intensities.

Adult↗
PubMed1994

Induction of reversible infertility in male rats by oral ornidazole and its effects on sperm motility and epididymal secretions.

Ornidazole (400 mg kg-1 day-1) given by oral gavage rendered male rats infertile by 6.6 +/- 0.7 days (mean +/- SEM, n = 9, range 3-10) after beginning the treatment and fertility returned within 5-10 days after treatment with ornidazole for 6-7 days. At 200 mg ornidazole kg-1 day-1, fertility was reduced but total infertility was not achieved. No differences were found in the percentage motility of spermatozoa recovered from any region of the epididymides of ornidazole-treated rats compared with controls. However, computer aided sperm analysis revealed significantly lower straight-line and average path velocities in ornidazole-treated animals (400 mg kg-1 day-1) for spermatozoa from the distal regions of the tract than for controls. Curvilinear velocity was significantly lower than that of controls in the distal corpus and cauda regions. The motility characteristics of spermatozoa from animals receiving 200 mg ornidazole kg-1 day-1 were lower than, but not significantly different from, motility in controls. There were no differences between the total protein, L-carnitine, glycerophosphocholine or total alpha-glucosidase content in epididymal homogenates from fertile control and infertile ornidazole-treated animals. Spermatozoa released from the cauda epididymidis of untreated rats into ornidazole solutions displayed no changes in the percentage motility up to 20 mmol l-1 and were only depressed at 50 mmol l-1. All velocities revealed a biphasic response with an initial increase in motility and then inhibition at higher concentrations, but a significant difference from velocities in the absence of orindazole was evident only for straight line velocity (VSL) at 50 mmol l-1.(ABSTRACT TRUNCATED AT 250 WORDS)

Administration, Oral↗
PubMed1994

Study of the role of epididymal alpha-glucosidase in the fertility of male rats by the administration of the enzyme inhibitor castanospermine.

The activity of epididymal alpha-glucosidase in adult rats was rapidly suppressed to histochemically undetectable levels within 2 days by the continuous release of the enzyme inhibitor castanospermine via a peritoneal osmotic pump at a rate of 100-200 nmol h-1. It was established that mating activities overnight depleted 72% of the spermatozoa in the distal cauda, which was replenished in 2 days, and that fertility began to decline 3 weeks after efferent duct ligation. Male rats of proven mating proficiency and fertility were treated with castanospermine, or buffered saline as control, for up to 30 days and enzyme inhibition was confirmed at the end of treatment by histochemistry. Fertility was normal at the first mating test on day 7, significantly decreased at the second mating on day 9, but recovered in a stepwise manner at subsequent matings on days 12 and 14. Delaying the third mating until day 25 did not sustain the transient subfertility. However, prolonging sperm storage in the distal cauda epididymides and preventing replenishment with freshly matured spermatozoa, by efferent duct ligation for 14 days performed on day 15 during castanospermine administration, caused a decrease in fertility and a change in the kinematics of epididymal spermatozoa of the castanospermine-treated group. In control rats, binding of epididymal spermatozoa to Vicia faba, a lectin specific for glucose and glucosamine, and mannose and mannosamine residues, decreased from the proximal caput to the distal corpus coincident with the increase in alpha-glucosidase activity on the epithelial brush border. Lectin binding then increased in the cauda where enzyme activity was absent. However, castanospermine treatment did not significantly alter this binding profile. The findings suggest that epididymal alpha-glucosidase does not play a crucial role in the development of sperm fertilizing capacity, but may be involved in the preparation of spermatozoa for storage.

Animals↗