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Biomedical subjects

C W Bardin

Publications and source records attributed to C W Bardin.

At least 19 recordsLinked to original sources
PubMed1992

A germ cell factor(s) modulates preproenkephalin gene expression in rat Sertoli cells.

Within the seminiferous tubule, both Sertoli and specific germ cells express opioid genes. Little is known about the paracrine regulation or role of opioid gene expression in the tubule. The present study shows that interactions among cells within the tubule may play a role in regulating preproenkephalin (PPenk) gene expression. Rat pachytene spermatocytes (PS) and round spermatids (RSd) were purified by centrifugal elutriation and established as primary cultures or co-cultured with Sertoli cells. The effects of germ cells or germ cell-conditioned media were studied to determine the expression of one of the opioid precursor genes in rat Sertoli cells, the PPenk gene. Following a 24 h co-culture with either PS or RSd, the expression of PPenk gene in Sertoli cells was increased 6.4- and 1.9-fold, respectively. Conditioned media obtained from either PS or RSd cultured for 20 h stimulated PPenk mRNA levels in Sertoli cells from as early as 2 h after exposure; maximum increases of 3.5- and 7.6-fold were observed at 12 h, respectively. The molecular weight of the germ cell factor(s) is greater than 30 kDa. 2 h after the addition of either PS- or RSd-conditioned media to Sertoli cells, small (2- to 2.6-fold, respectively) but significant (p less than 0.02) increases in extracellular cAMP levels were observed. Although both FSH and forskolin activated c-fos and PPenk gene expression in Sertoli cells, the germ cell factor(s) that stimulated PPenk mRNA levels did not affect the expression of this oncogene. These results indicate that germ cells interact with Sertoli cells, possibly by a protein(s) that acts as a short-loop paracrine factor, which regulates the expression of PPenk gene in Sertoli cells. These data suggest that stage-specific regulation of PPenk levels in Sertoli cells may occur in vivo.

Animals↗
PubMed1992

Binding of sex-hormone-binding globulin (SHBG) to testicular membranes and solubilized receptors.

UNLABELLED: Sex-hormone-binding globulin (SHBG) binds to a specific protein on the surface of prostate, epididymis, and a human breast cancer cell line (MCF-7), and is internalized by these cells. The present study demonstrated specific binding of SHBG to receptor on membranes prepared from rat testes. The binding was saturable, specific, and time and temperature dependent. Scatchard analysis of these binding studies suggested that SHBG binds to a single class of sites on testicular membranes with a Kd at 37 degrees C of 5 x 10(-8) M and a binding capacity of 30 +/- 0.6 pmol/mg protein. These binding characteristics are similar to the SHBG receptor on human prostate and MCF-7 cells. Solubilization of the receptor resulted in a 5-fold increase in its binding capacity (158 +/- 0.3 pmol/mg protein) and a 10-fold decrease in binding affinity (Kd at 37 degrees C = 6.5 x 10(-7) M). The apparent molecular weight of the testicular SHBG receptor, as estimated by gel filtration, was M(r) = 174,000. CONCLUSION: a specific binding site for SHBG was identified on testicular membranes. This binding site has been tentatively identified as a SHBG receptor based on its physical properties in testicular membrane preparations and following solubilization.

Animals↗
PubMed1992

Rat Sertoli cell clusterin, alpha 2-macroglobulin, and testins: biosynthesis and differential regulation by germ cells.

Clusterin, alpha 2-macroglobulin and testins are three novel Sertoli cell proteins whose physiological functions may be related to cell-cell interactions in the seminiferous epithelium of the testis. We have demonstrated the biosynthesis of clusterin, alpha 2-macroglobulin, and testins by Sertoli cells in vitro using pulse-chase labeling analysis. For clusterin, two precursors with an apparent molecular weight (M(r)) of 72,000 (PH) and 66,000 (PL) were detected in the Sertoli cell cytosol in addition to the alpha (M(r) 43,000) and beta (M(r) 35,000) subunits of the mature protein. However, the precursors were not secreted into the medium since only the alpha and beta subunits of clusterin were detected. For alpha 2-macroglobulin and testins, no precursor molecules were detected either in the Sertoli cell cytosol or culture medium. The polarized secretory pattern of these proteins and their regulation by follicle stimulating hormone (FSH) and testosterone (T) were examined using a bicameral culture chamber that mimics the in vivo physiological conditions. Clusterin was secreted almost exclusively into the apical chamber of the bicameral culture unit with an apical:basal ratio of 30:1. In contrast, alpha 2-macroglobulin and testins had an apical:basal ratio of 1:1 and 1.5:1, respectively. Thus, the polarized secretory pattern for clusterin is different from alpha 2-macroglobulin and testins. It was noted that FSH and T, the known Sertoli cell regulators, did not affect the secretion of either clusterin or alpha 2-macroglobulin. Due to the morphological intimacy between Sertoli cells and germ cells in the adluminal compartment of the testis, the effects of germ cell-conditioned medium were investigated. Addition of germ cell-conditioned medium (1-30 micrograms protein) to the apical chamber of the bicameral culture unit caused a dose-dependent inhibition of clusterin and testins apical secretion and a slight but statistically significant stimulation of their basal secretion. In contrast, the secretion of alpha 2-macroglobulin by Sertoli cells was stimulated both apically and basally. These observations suggest that germ cell-conditioned medium contains a biological factor(s) that differentially regulates the bidirectional secretion of Sertoli cell proteins. These studies therefore reveal the complicated regulatory processes involved in cell-cell interactions in the seminiferous epithelium.

Animals↗
PubMed1992

Testins are localized to the junctional complexes of rat Sertoli and epididymal cells.

UNLABELLED: Testin I and Testin II were originally identified as Sertoli cell products with similar NH2-terminal amino acid sequences. Secretion of testins is stimulated by testosterone in Sertoli cell-enriched cultures. By contrast the secretion of testins from intact seminiferous tubules appears to be inversely related to germ cell number. In the present study testin antiserum that recognized both Testin I and Testin II ("testin") was used to localize these proteins in tissue secretions by immunofluorescence. Testin was localized at the base of the seminiferous epithelium at Sertoli-Sertoli junctions. Fluorescence also appeared to be located at the sites of interaction between spermatoids and Sertoli cells. A punctate pattern of fluorescence was also present in the cytoplasm of Leydig cells; without electron microscopic studies it was not possible to determine which structures the antibodies bound to in these cells. In the epididymis the reaction product was localized at the apices of the epithelial cells adjacent to the lumen at the sites of known junctional complexes. A variety of positive and negative controls indicated that staining was specific for testins. CONCLUSIONS: This is the first study to associate testins with junctional complexes. Relative to other junctional proteins, testins are unusual because of their small size and because they are secreted proteins.

Animals↗
PubMed1992

Effects of follicle-stimulating hormone on inhibin release by different testicular compartments in the adult ram.

The aim of this study was to determine the bidirectional release of immunoreactive inhibin-alpha (irINH-alpha) by different testicular compartments in the adult ram and to assess the effects of FSH on the distribution of inhibin in the testis. Immunoreactive INH-alpha was measured by RIA in fluid samples collected concurrently from the three testicular compartments--the seminiferous tubules, the interstitium, and the vascular system--through catheters inserted surgically into the rete testis, testicular lymphatic duct system, and spermatic veins, respectively. Overall, the concentration of irINH-alpha in rete testis fluid was 25 times the level in testicular lymph and over 500 times the concentration in peripheral blood. The pattern of irINH-alpha concentration in rete testis fluid was inversely related to that in testicular lymph, but i.v. administration of FSH had a decoupling effect on this relationship by depressing inhibin concentration in testicular lymph without affecting inhibin levels in rete testis fluid. Nevertheless, increased flow of testicular lymph more than compensated for the transient fall in irINH-alpha concentration so that, overall, the total output of inhibin via the testicular lymphatic duct system (and the vascular system) increased significantly. No persistent or significant changes were observed in the flow rate of rete testis fluid or concentration of irINH-alpha in the fluid after administration of FSH. The time frame for the response of the testis to FSH is indicative of the involvement of a mediator. Electrophoretic analysis of serially collected testicular lymph samples consistently revealed an FSH-induced release of a series of proteins in the M(r) range of 30,000-32,000.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Binding of an extracellular steroid-binding globulin to membranes and soluble receptors from human breast cancer cells (MCF-7 cells).

Studies of MCF-7 breast cancer cells demonstrated that sex hormone-binding globulin (SHBG) is internalized by receptor-mediated endocytosis. The present study demonstrated specific binding of SHBG to receptor on membranes isolated from MCF-7 cells. Scatchard analysis of these binding studies suggested that SHBG binds to a single class of sites on membranes. The analysis yielded a dissociation constant (Kd) at 37 C of 3 x 10(-8) M and a binding capacity of 48 +2- 0.12 pmol/mg protein. A procedure for solubilizing the SHBG receptor from MCF-7 membranes used buffers containing protease inhibitors, 10% glycerol, and 10 mM 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate. Solubilization of the receptor resulted in a 5-fold increase in its binding capacity (246 +/- 14 pmol/mg protein) and a 10-fold decrease in binding affinity (Kd at 37 C = 2 x 10(-7) M).

Binding, Competitive↗
PubMed1992

The biological activity of 7 alpha-methyl-19-nortestosterone is not amplified in male reproductive tract as is that of testosterone.

Based on the premise that testosterone, but not 7 alpha-methyl-androgens, is reduced at the 5 alpha-position in the prostate and seminal vesicles, the differential bioactivities of these androgens were investigated in castrated rats. The ability of 7 alpha-methyl-19-nortestosterone acetate (MENT) to increase the weights of ventral prostate and seminal vesicles of castrated rats was four times higher than that of testosterone, while its effect on the weights of bulbocavernosus plus levator ani muscles (muscle), was 10 times that of testosterone. MENT was also approximately 12 times more potent than testosterone in the suppression of serum gonadotropin levels. A dose of testosterone that maintains serum gonadotropin levels and muscle mass also maintains prostate and seminal vesicle weights in castrated rats. By contrast, a dose of MENT that maintains muscle and gonadotropins does not maintain prostate and seminal vesicles. The action of other 7 alpha-methylated androgens were similar to that of MENT. The importance of 5 alpha reductase in the differential action of testosterone and MENT on prostate was confirmed by using a 5 alpha-reductase inhibitor. The activity of testosterone was significantly suppressed in the ventral prostate and seminal vesicles but not on muscle by the 5 alpha-reductase inhibitor (N,N-diethyl-3-oxo-4-aza-5 alpha-androst-1-ene-17 beta-carboxamide). The enzyme inhibitor, however, had no influence on the activity of MENT on either tissue. In contrast, cyproterone acetate, an antiandrogen that competitively binds to the androgen receptors, inhibited the action of MENT and of testosterone on the prostate as well as on the muscle. In conclusion, these observations show that 7 alpha-methylated androgens can maintain muscle mass and normal gonadotropin levels in androgen deficient rats without hyperstimulating the prostate. These findings suggest that 7 alpha-methylated androgens may offer some health benefits to men who require androgen treatment.

3-Oxo-5-alpha-Steroid 4-Dehydrogenase↗
PubMed1992

Clusterin production in the obstructed rabbit kidney: correlations with loss of renal function.

Clusterin, a protein associated with cell death, has been suggested as a marker of renal injury. Correlation of clusterin gene expression with changes in renal function and quantitative measurement of clusterin protein levels after ureteral obstruction have not been previously reported. With unilateral ureteral obstruction in rabbits as the experimental model, the time course of alterations in renal function, clusterin mRNA accumulation, and concentrations of clusterin protein in serum, urine, and renal tissue were investigated. RBF, GFR, and renal concentrating ability (percent sodium reabsorption and urine osmolarity) all decreased (P < 0.05) in the obstructed kidney from control values within 1 day of ureteral obstruction. Clusterin mRNA levels started to rise in the ipsilateral kidney within 12 h of ureteral obstruction and increased up to 10-fold above control levels after 3 days of obstruction. Hybridization histochemistry showed that clusterin mRNA was initially detectable in collecting ducts and distal tubules within 12 h of ureteral obstruction. After 7 days of obstruction, increased accumulation of clusterin mRNA was also detectable in proximal tubular epithelial cells. Clusterin gene expression remained elevated in collecting ducts after 60 days of obstruction. Clusterin expression in the contralateral kidney was increased twofold over control values after 12 h of obstruction. No increase in clusterin mRNA accumulation was detectable after 24 h in the contralateral kidney. Total clusterin protein in the obstructed kidney increased from 0.59 +/- 0.66 (mean +/- 1 SD) to 2.5 +/- 1.3 micrograms after 7 days of ureteral obstruction (P < 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Experience with 19-nortestosterone in the therapy of systemic lupus erythematosus: worsened disease after treatment with 19-nortestosterone in men and lack of improvement in women.

Three men and 4 women with systemic lupus erythematosus (SLE) received 100 mg of 19-nortestosterone decanoate in weekly intramuscular injections over a period of 3 to 24 months. During therapy in the men plasma luteotrophic hormone and free testosterone levels decreased while estrogen levels increased. Anti-DNA antibodies also increased for unknown reasons. Serological changes in two men coincided with an overall increase in the clinical activity of SLE and the appearance of new onset Raynaud's phenomenon and pleuropericardial disease. In contrast, women treated with 19-nortestosterone showed clinical stability despite the persistence of high titer antibodies to DNA, and were able to continue therapy for as long as 16 months in one case. These data indicate that men and women with SLE respond differently to synthetic androgen therapy. Additionally, dissociation of clinical signs and symptoms from serological variables seems possible.

Adult↗
PubMed1979

Ultrastructural changes in Leydig cells of streptozotocin-induced diabetic rats.

Hyperglycemia (experimental diabetes) was induced in adult male rats by destruction of the pancreatic beta cells with a single intravenous injection of streptozotocin (STZ). Testes from diabetic, from insulin-treated diabetic, and from sham-injected normal rats were fxed by vascular perfusion. The fine structure of Leydig cells was examined at two, three, and four weeks after the STZ injection in the untreated diabetic animals, and at four weeks in the controls and insulin-treated diabetic rats. A number of morphological changes was observed in Leydig cells of untreated diabetic animals. Most obvious of these was an accumulation of lipid droplets, not normally present in Leydig cells in adults of this species. Smooth endoplasmic reticulum (SER) was markedly reduced in Leydig cells of the hyperglycemic rats. Several types of intracellular bodies were seen exclusively in Leydig cells of the untreated diabetic animals. Many resembled secondary lysosomes or dense bodies, while others appeared to be autophagic vacuoles. In addition, a small, granule-containing lamellar structure was seen either within a typical dense body or free in the cytoplasm. Myelin-like structures were commonly observed within the cytoplasm of the Leydig cell or within mitochondria. The appearance of the mitochondria in diabetic rats was otherwise normal. The extracellular spaces surrounding Leydig cells from untreated hyperglycemic rats also contained large accumulations of myelin-like material. These structural changes appear to be direct consequences of the diabetic state of the animals, since the ultrastructure of insulin-treated diabetic rats did not differ from that of the controls. These findings may reflect an alteration or breakdown of Leydig cell components normally involved in the synthesis of androgen, and correlate with previous reports of lowered circulating levels of testosterone in diabetic rats.

Animals↗
PubMed1979

The use of medroxyprogesterone acetate to study progestin receptors in immature, pregnant, and adult rabbit uterus.

Medroxyprogesterone acetate (MPA)3, a potent synthetic progestin, was employed as the 3H-ligand to investigate the status of the rabbit uterine progestin receptor under various hormonal conditions. MPA dissociated slowly from the cytosol and nuclear progestin receptor obtained from ovariectomized, estrogen-primed rabbits. This slow rate of dissociation permitted a partial characterization of the receptors in pregnant and immature, unprimed rabbit uteri. High affinity progestin binding to 6--7 S and 4 S macromolecular species was detected in cytosol and nuclear extracts, respectively, from these animals. In contrast to the 3 peaks of cytosol receptor activity from estrogen-primed animals which were eluted stepwise from DEAE-cellulose columns, predominantly 2 peaks were obtained with cytosols from pregnant and immature, unprimed uteri. Further studies are required to determine to what extent progestin action is modified through interaction of these binding components with themselves and other cellular proteins.

Aging↗
PubMed1979

In vivo metabolism and binding of 6 alpha-methylprogesterone; a progestin with anti-androgenic and synandrogenic activities.

The synthetic progestin 6 alpha-methylprogesterone (6MP) was shown to have androgenic, synandrogenic and anti-androgenic actions on mouse kidney. In vitro studies indicated that 6MP competes with testosterone for binding sites on the androgen receptor in kidney cytosol and nuclei. The in vivo distribution of 6MP and testosterone differed following subcutaneous administration: Testosterone levels were 10 and 100-fold higher than those of 6MP in plasma and nuclei from prostate-seminal vesicle. In kidney nuclei, the concentrations of these two steroids were similar. In addition, kidney nuclei bound a hydroxylated metabolite of 6MP as well as the parent compound, whereas only 6MP was found in nuclei from prostate-seminal vesicle. In vivo competition studies indicated that nonradioactive 6MP and testosterone decreased the uptake of 3H-testosterone by nuclei of kidney and prostate-seminal vesicle. By contrast, 3H-6MP uptake in kidney nuclei was potentiated by the prior administration of 6MP or testosterone. These results suggest that factors in addition to the androgen receptor may be involved in the binding and mechanism of action of 6MP on mouse kidney.

Animals↗
PubMed1979

Metabolic mimicry of Bartter's syndrome by covert vomiting: utility of urinary chloride determinations.

Bartter's syndrome characteristically exhibits the constellation of hypokalemic alkalosis with moderate kaliuresis, normotensive hyperreninemia, hyperaldosteronism, urinary hyperexcretion of prostaglandin E (PGE) and vascular hyporesponsivity to pressor agents. We describe precise biochemical mimicry of these metabolic abnormalities in a patient with surreptitious repetitive vomiting, in whom simple urinary chloride excretion data subsequently excluded the diagnosis of Bartter's syndrome.

Adult↗
PubMed1979

The neuroendocrinology of male reproduction.

Androgen biosynthesis occurs in Leydig cells and spermatogenesis in the seminiferous tubules, but both are under control of the anterior pituitary through its secretion of gonadotropins. The hypothalamus is primarly involved in feedback control of gonadotropin secretion in response to varying blood levels of testicular hormones. Another pituitary steroid, prolactin, also is involved in male behavior.

Androgens↗
PubMed1979

Protein carboxyl-methylation in rat testes: a study of inherited and X-ray-induced seminiferous tubule failure.

Protein carboxyl-methylase (PCM), the enzyme that transfers methyl groups from S-adenosyl-methionine to free carboxyl groups on proteins, is highly localized in testes. The cellular distribution of PCM and its substrates, the methyl acceptor proteins, was investigated. Separation of testicular cells on an albumin gravity gradient revealed the preferential localization of both enzyme and substrates in spermatids. In young rats, PCM activity increases with age coincidently with germ cell maturation. Rats which are heterozygous for the Hre gene (Hre/+) are infertile as a result of germ cell depletion. In these animals, testicular PCM specific activity and total activity were, respectively, 4--6 and 40--50 times lower than in normal testes. Enzyme activity in testes from animals with x-ray-induced germ cell depletion was also very low. These observations suggest that PCM is located in germ cells.

Animals↗