Biomedical subjects

R R Tremblay

Publications and source records attributed to R R Tremblay.

At least 91 records · Page 5Linked to original sources

PubMed1985

Major contribution of epididymis to alpha-glucosidase content of ram seminal plasma.

Acid alpha-glucosidase and L-carnitine (a well-known epididymal marker) were measured in rete testis and epididymal fluids of adult male rams. These fluids were collected by selective catheterization or by a micropuncture technique, respectively. Both parameters remained at a low and constant level in rete testis and all along caput and corpus epididymidis. Then they increased at equivalent rates in cauda epididymidis to much higher levels than those in seminal plasma (5 mU/mg protein and 10 mM, respectively). An optimum pH study of alpha-glucosidase activity in these fluids showed two well-separated peaks in rete testis and caput epididymal fluids around pH 4 and 7, respectively, but only a single peak at pH 4 in cauda epididymidis that was comparable to the one in seminal plasma. Sucrose density gradient fractions analyzed for their enzyme content in the absence or presence of sodium dodecyl sulfate (1% w/v), a selective inhibitor of acid alpha-glucosidase activity, allowed the demonstration of two enzyme forms at pH 6.8 in rete testis fluid sedimenting in the 7S and 4S regions of the gradient, while a unique 4S form was encountered in cauda epididymidis and in seminal plasma. Although the fate of the minor 7S component of the rete testis fluid in its epididymal transit is presently unknown, similarities between the enzyme in cauda epididymidis and seminal plasma are strong enough to support the hypothesis that epididymis contributes primarily to the acid alpha-glucosidase content of ram seminal plasma.

Animals↗

PubMed1985

In vitro translation of mRNA for arginine esterase, the major secretory protein of dog prostate, and in vitro processing of the translation product.

Poly(A)+ rich RNA was isolated from prostate of adult dogs and translated in the rabbit reticulocyte lysate cell-free protein-synthesizing system. Two-dimensional gel electrophoresis of the translation products showed that a protein with a molecular weight of 31 000 was predominantly synthesized. This protein was immunoprecipitated with antibodies directed against purified arginine esterase from dog seminal plasma. mRNA isolated from the prostate of animals castrated for 1 or 2 weeks was unable to direct the synthesis of arginine esterase. However, the synthesis of the enzyme could be stimulated by androgens in castrated animals, presumably by increasing prostatic concentrations of arginine esterase mRNA. The single chain translation product could be further processed in vitro by the addition of dog pancreas microsomes and purified arginine esterase. This procedure yielded split chains of arginine esterase which had identical electrophoretic mobilities as seminal plasma enzyme by two-dimensional gel electrophoresis. When prostatic tissue slices were incubated with tunicamycin, the unglycosylated arginine esterase obtained had a lower molecular weight than the in vitro translation product, suggesting that a signal peptide had been removed in the living cells. These results indicate that arginine esterase processing may include the following steps: removal of a signal peptide, glycosylation, and splitting of the polypeptide chain by active arginine esterase in the secretory granules or outside the cell.

Animals↗

PubMed1985

Arginine esterase from isolated dog prostate secretory granules is fully active enzymatically.

We have isolated secretory granules from dog prostate homogenates and have determined whether a major portion of arginine esterase was localized in this fraction and if it was enzymatically active. Secretory granules were purified by density gradient centrifugation on sucrose, metrizamide, or Percoll. A major proportion of whole prostate homogenate arginine esterase was found in the granule fractions. Furthermore, the specific enzymatic activity in the granules was similar to the one observed in seminal plasma. No evidence could be found for the existence of significant amount of a zymogen inactive form of arginine esterase. These results suggest that arginine esterase could be active within the secretory granules in vivo and that it could hydrolyze protein substrates contained in this organelle.

Acid Phosphatase↗

PubMed1985

Nature of the residual alpha-1,4-glucosidase activity in the seminal plasma of vasectomized men.

Vasectomy leads to a drastic decrease of neutral alpha-1,4-glucosidase from human seminal plasma. The nature of this residual enzyme activity has been ascertained according to optimum pH and sucrose density gradient analysis with or without inhibitors of neutral (maltotriose) or acid (sodium dodecyl sulfate) alpha-1,4-glucosidase. Data from the present study provide strong evidence that the enzyme content of the seminal plasma is of mixed nature after exclusion of the epididymis.

Adult↗

PubMed1985

Hormonal steroids indirectly influence insulin binding in rat testis.

Pituitary LH is a key factor in controlling Leydig cell activity. In order to verify how steroids produced by the male gonads could influence testicular insulin binding, different steroids at varying doses were administered. Testosterone propionate (Tp) as well as estradiol-17 beta (E2-17 beta) and 5 alpha-dihydrotestosterone (DHT) led to a 50% decrease of insulin binding in testis while no effect was noted when HCG was administered concomitantly with DHT. LH receptors were parallely measured: DHT and E2-17 beta significantly depressed testis LH binding. It is concluded that steroids play a role in the regulation of testis membrane insulin receptor via their feedback mechanism on pituitary LH.

Animals↗

PubMed1985

Biochemical characteristics of the proteins secreted by dog prostate, a review.

Proteins secreted by dog prostate differ from those of other species and in particular from those of man, both in terms of quantity and quality. These differences can be observed in the electrophoretic pattern of proteins in polyacrylamide gels and in enzymes such as phosphatases, glycohydrolases and proteases. For instance, canine acid phosphatase, although quite similar biochemically to the human enzyme, is about one hundred fold less concentrated in both prostatic tissue and seminal plasma than in the human. By contrast, arginine esterase, which is virtually absent in human prostatic secretion, comprises more than 90% of total proteins secreted by dog prostate. The most recent data on each of these enzymes, enzyme classes and proteins will be reviewed. Despite recent advancement of knowledge on the biochemistry of these constituents, their biological function as well as their contribution to the pathogenesis of prostatic diseases still remain a matter of speculation. For both of these aspects, dog appears to be a particularly interesting model.

Acid Phosphatase↗

PubMed1984

The major androgen-dependent protease in dog prostate belongs to the kallikrein family: confirmation by partial amino acid sequencing.

Canine prostate fluids and seminal plasma contain a major androgen-dependent protein which was identified as a proteolytic enzyme exhibiting an Arg-esterase activity. This protease, as characterized, is shown to be present as a two-chain structure held together by at least one disulfide bridge and composed of approximately 220 amino acids. Amino acid sequence determination of both chains has revealed a clear homology to other known amino acid sequences of serine proteases. Furthermore, the comparison of the presented 58 amino acids of the Arg-esterase with the other sequences revealed a very strong homology (larger than 50%) to members of the kallikrein family. The two chain structure could thus result from autolysis of a single chain enzyme in the 'kallikrein autolysis loop'. Amino acid composition of the canine prostatic enzyme suggests that it is related, but not identical, to pancreatic canine kallikrein.

Amino Acid Sequence↗

PubMed1984

Involution of spontaneous benign prostatic hyperplasia in the dog under the influence of chronic treatment with a LHRH agonist.

Eight dogs with spontaneous benign prostatic hyperplasia were treated with daily subcutaneous injections of 25 micrograms of Buserelin, (D-Ser(TBU)6,des-Gly-NH2(10))ethylamide during 3 months. Prostate weight before treatment was estimated by tridimensional measurements during open surgery. After 3 months of treatment, prostate weight was decreased by 50-60% In two castrated control animals, prostate weight decreased about 70% after 3 months. Histological pattern in prostates from castrated or Buserelin-treated animals was similar and indicated considerable atrophy of glandular epithelium and predominance of stroma. At the same time, testes weight decreased with complete disappearance of spermatogenic activity. Acid phosphatase and arginine esterase activity levels, two markers of androgenic action in the prostate, were similar after Buserelin or castration. Steroid receptors for androgens, estrogens, and progesterone also behaved similarly under the influence of Buserelin and castration.

Animals↗

PubMed1984

Single case report of prostate adenocarcinoma in a dog castrated three months previously. Morphological, biochemical, and endocrine determinations.

During the course of another investigation, three dogs had been castrated 3 months previously. Upon completion of the experiment, it was discovered that one dog presented a spontaneous prostatic adenocarcinoma of intraalveolar proliferative type at histology. Prostate weight of this dog before castration was estimated to be 22 g by tridimensional measurement at laparotomy and remained relatively constant (19 g) 3 months after castration. These results indicate that if regression had occurred in some cell populations (androgen-dependent) it was only partial and masked by growth of androgen-independent cells. Analysis of 12 individual steroids in peripheral blood and in prostatic tissue attested of a normal adrenal secretory activity. A series of 15 hydrolytic enzymes along with receptors for androgen, estrogen, and progesterone, were determined in prostatic tissue obtained at sacrifice. Enzymatic activities were those of typical epithelial cells, and most of them remained relatively high despite low levels of circulating testosterone. However, two markers of androgen action in dog prostate, acid phosphatase and arginine esterase, were significantly reduced. Receptor levels were similar to those of castrated animals. Thus, cancer cells had probably retained some androgen sensitivity.

Adenocarcinoma↗

PubMed1984

Characteristics of the inhibitory effect of chronic treatment with an LHRH agonist on testicular steroidogenesis in the dog.

Daily subcutaneous administration for 3 months of the potent LHRH agonist (D-Ser(TBU)6, des-Gly-NH2(10] LHRH ethylamide (25 micrograms) to adult dogs having spontaneous benign prostate hyperplasia (BPH) causes a marked inhibition of testicular androstenedione and testosterone secretion. This inhibition of delta 4-androgen secretion is accompanied by a decrease of testicular progestin precursors and 5 alpha-androgen metabolites, thus suggesting that, in dog, the loss of testicular steroidogenic activity, induced by the administration of an LHRH agonist, is due to a total inhibition of testicular steroidogenesis. In plasma, the concentration of both testosterone and dihydrotestosterone is also markedly depressed while androstane-3 alpha, 17 beta-diol levels remain unchanged. Measurement of prostatic steroid content has shown that administration of the LHRH agonist as well as castration is associated with a marked decrease in androstenedione, testosterone, and dihydrotestosterone levels in prostate while there is a small inhibition of androst-5-ene-3 beta, 17 beta-diol, androstane-3 beta, 17 beta-diol, dehydroepiandrosterone, and estrone concentrations in this tissue. The present data show that treatment with an LHRH agonist in the dog causes a marked inhibition of testicular steroid secretion similar to the one observed in adult men, and suggest that steroids from adrenal origin may also be involved in prostatic function.

17-alpha-Hydroxyprogesterone↗

PubMed1984

Synthesis of acid phosphatase and other proteins by human prostatic tissue in vitro.

We have studied the synthesis of proteins by normal and hyperplastic human prostatic tissue incubated in vitro in the presence of [35S]methionine. The overall pattern of newly synthesized proteins was similar in individuals with an age ranging from 15 to 68 years. The pattern of labeled proteins was quite different from that of total proteins stained with Coomassie blue in two-dimensional polyacrylamide gels, since the major stained proteins was not labeled. Among the most heavily labeled proteins (about a dozen) were several spots representing charge isoforms with molecular weights ranging from 46 000 to 51 000, and these were the only proteins immunoprecipitated by a polyclonal antibody developed against purified acid phosphatase. The other heavily labeled proteins had molecular weights ranging from 26 000 to 72 000. These results show that tissue slices can be used to study the synthesis and processing of acid phosphatase, the major secretory product of the prostate, and of other unidentified proteins.

Acid Phosphatase↗

PubMed1984

Biochemical properties and kinetic parameters of monoamine oxydase in human seminal plasma.

The present study demonstrates that monoamine oxydase (MAO) is present in the human seminal plasma and that its activity was higher in infertile than in fertile men. The Km and Vmax of the enzyme were, respectively, 3.0 X 10(-3) M and 625 eta mol of deaminated 5-HT mg of prot-1 min-1. The enzyme was activated by pyridoxal-5' phosphate but inhibited by usual MAO inhibitors such as pargyline-HCl, iproniazid-PO4 and clorgyline. These findings suggest a relationship between the synthesis and degradation of biogenic amines and testicular function.

Enzyme Activation↗

PubMed1983

Effect of castration and steroid treatments on the activity of some hydrolytic enzymes in dog prostate.

Adult mongrel dogs were castrated and treated by intramuscular injections of 5 alpha-androstane-3 alpha,17 beta-diol (androstanediol) alone or in combination with estradiol in order to find convenient enzymatic markers of hormone action in prostate. The activities of 15 hydrolytic enzymes were determined. Arginine esterase, acid sulfatase, and acid phosphatase were found to be the most sensitive markers of testicular hormones since they were decreased 18-, 5- and 5-fold respectively after 1 month of castration. The enzyme activities returned to precastration levels after 2 weeks of injection of androstanediol to castrated animals. The effect of androstanediol on the majority of the remaining enzymes was small. In general, the activities obtained after androstanediol treatment in combination with estradiol were similar to those obtained with androstanediol alone. Finally, beta-glucuronidase and neutral sulfatase were increased after castration, a finding that suggests that these enzymes are constituents of stromal cells. These studies will provide a basis for future studies of hormone action in the dog prostate.

Acid Phosphatase↗

PubMed1983

Determination of occupied cytoplasmic glucocorticoid receptor sites by an exchange assay in rat muscles.

We have established an assay to determine total cytoplasmic glucocorticoid receptor sites in heart, vastus and levator-ani-bulbocavernosus muscles from rats. We have tested the exchange of endogenously bound hormone with labeled triamcinolone acetonide by performing long incubation at 0-4 degrees C. This procedure appeared to produce an adequate exchange of exogenously added unlabeled corticosterone in the range of 16 nM (normal unstressed concentration of corticosterone in plasma) with vastus and levator ani-bulbocavernosus muscles. Beyond that level, the displacement of unlabeled corticosterone by labeled triamcinolone acetonide was less efficient. By contrast, in heart and especially in liver cytosols, the exchange was adequate even in the presence of high concentrations of corticosterone (64 and 460 nM respectively). Using the exchange assay conditions (20 h of incubation at 0-4 degrees C with 16 nM of labeled triamcinolone acetonide), we found no significant difference between glucocorticoid receptor levels in intact male and female rats. In both sexes, these glucocorticoid receptor values were not affected by gonadectomy, but increased more than 2-fold after adrenalectomy. No seasonal variations were recorded in intact or adrenalectomized rats.

Adrenalectomy↗

PubMed1983

Comparative study of alpha-glucosidase activities in blood and seminal plasma of rams.

1. This study was undertaken to characterize the alpha-glucosidase present in blood and seminal plasma of two strains of pure-bred rams which are known as seasonal breeders. 2. pH profiles and activity levels were investigated in blood and seminal plasma using a sensitive spectrophotometric assay with para-nitrophenyl-alpha-D-glucoside as substrate. 3. According to their pH optimum, blood plasma and seminal plasma alpha-glucosidases were typically neutral and acid enzymes and significant differences were recorded in their physico-chemical properties, establishing the tissue specificity of the enzyme. 4. Notwithstanding the tissue under study, the nature of the alpha-glucosidase activity was similar in both strains of pure-bred rams.

Animals↗

PubMed1983

Effect of testicular hormones on synthesis of soluble proteins by dog prostate slices.

We have submitted adult mongrel dogs to various endocrine manipulations. Prostate slices from these animals were then incubated in vitro in the presence of [3H]leucine or [35S]methionine. We have analyzed the cytosolic proteins by polyacrylamide gel electrophoresis. In intact adult uncastrated dogs, the radioactive amino acids were incorporated into three major bands having respective molecular weights (MW) of 32 000, 16 000, and 15 000 in one-dimensional gels in presence of sodium dodecyl sulfate and mercaptoethanol. Two-dimensional electrophoresis revealed heterogeneity of each of these bands, both in isoelectric focussing (IEF) or nonequilibrium pH gel electrophoresis (NEpHGE) conditions. The 32 000 MW proteins showed five to six major radioactive spots and the 15 000 - 16 000 MW proteins showed six to seven spots by IEF. However, the highest incorporation of radioactivity occurred in a 16 000 MW protein seen only in NEpHGE. The lower MW proteins corresponded to some of the major proteins of dog seminal plasma as observed by immunoprecipitation of prostate proteins with antibodies against whole seminal plasma. By contrast, the 32 000 MW proteins were minor proteins of prostate cytosol and seminal plasma by Coomassie blue staining. Castration for 2 weeks completely abolished the synthesis of all these proteins. When castrated animals were treated with 5 alpha-androstane-3 alpha-17 beta-diol (10 mg/day for 2 weeks), the pattern of protein synthesis returned to the one observed in intact uncastrated animals. These observations show that testicular androgens control the synthesis of dog prostate major secretory proteins.

Androstane-3,17-diol↗

PubMed1983

Presence of alpha-glucosidases in the male reproductive system of the rat and hormonal influences.

The use of a synthetic substrate (p-nitrophenyl-alpha-D-glucoside) to measure alpha-glucosidase activity has allowed us to demonstrate the presence of acid and neutral alpha-glucosidases in the reproductive organs of the male rat. Both enzymes increased in the epididymis, particularly in the caput segment, along with initiation of spermatogenesis at puberty; it then started decreasing after 12 weeks of life. Similar variations were not recorded in testis, prostate, and seminal vesicles. Castration led to a significant decrease of acid and neutral alpha-glucosidases in all accessory reproductive organs, but administration of testosterone proprionate (50 micrograms/day for 10 days) restored the enzyme activity to its original level. When estradiol-17 beta (5 mg) was administered simultaneously with testosterone (500 micrograms), the antagonistic effect of estradiol on testosterone was particularly evident on the levels of neutral alpha-glucosidases which reached the castration range, while the acid alpha-glucosidase remained unchanged in epididymis, prostate, and seminal vesicles. These results show that both acid and neutral alpha-glucosidases may be influenced by gonadal hormones in the male rat.

Animals↗

PubMed1983

Effect of glucocorticoid administration in vivo on insulin binding in rat testis.

We have studied the effects of adrenalectomy and glucocorticoid injections on insulin binding in membranes from rat testis and liver. Glucocorticoids were administered for 7 days to adrenalectomized rats at daily doses of 30 or 300 micrograms for dexamethasone and 100 or 1000 micrograms for prednisolone. Glucocorticoids, at the selected doses, were associated with 5- to 10-fold increases in basal insulin levels with no significant changes in glucose concentrations. As previously shown by other studies, down-regulation of insulin receptors was observed in liver membrane particularly at the higher dose of steroids. Such an effect was not found in the testis. By contrast, the number of high-affinity sites in the testis was slightly increased with the higher doses of dexamethasone and prednisolone. However, percent 125I-labelled insulin binding was not significantly changed after corticotherapy. These results are in accord with our previous studies and suggest that the testicular receptor for insulin is not affected by mild to moderate changes of insulin concentrations, but that it can be modulated by glucocorticoids through other mechanisms.

Animals↗