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Biomedical subjects

N A Musto

Publications and source records attributed to N A Musto.

At least 55 records · Page 3Linked to original sources
PubMed1983

Spermatogenesis in vitro: completion of meiosis and early spermiogenesis.

In vitro formation of haploid spermatids has not been convincingly demonstrated in mammals. To investigate this problem we selected defined segments of rat seminiferous tubules containing late pachytene and diakinetic primary spermatocytes (Stages XII and XIII of the cycle) for culture in a chemically defined medium. After 2 days, most spermatocytes completed both meiotic divisions, and by 6 days the tubular epithelium developed morphologic characteristics of Stage V in which the newly formed spermatids had acrosomic systems characteristic of step 5 spermiogenesis. The seminiferous tubules also differentiated biochemically as evidenced by increased production of proteins characteristically secreted by Stage V. Since this in vitro differentiation of the germinal epithelium occurred in the absence of testosterone and FSH, we conclude that late pachytene spermatocytes and their associated Sertoli cells have all the information required for both meiotic divisions and early spermiogenesis.

Animals↗
PubMed1983

Radioimmunoassay of testosterone-estradiol-binding globulin in humans: a reassessment of normal values.

Antiserum against human testosterone-estradiol-binding globulin (hTeBG) was prepared in rabbits, and its specificity was demonstrated by crossed immunoelectrophoresis. A RIA for the measurement of hTeBG in serum was developed. With this assay, hTeBG was readily measured in 1-2 microliters serum. Interassay coefficients of variation for pools of serum from men, women, and women in late pregnancy were 7.7%, 5.5%, and 6.1%, respectively. Interassay coefficients of variation for the same pools were 10%, 12%, and 12%, respectively. The TeBG levels in a number of nonselected subjected determined by the present method show good correlations with those obtained by steady state polyacrylamide gel electrophoresis and dextran-coated charcoal assay. The concentrations of TeBG determined by the RIA in sera from men, women, and women in late pregnancy were 18 +/- 9 (n = 12), 54 +/- 13 (n = 8), and 374 +/- 55 (n = 6) pmol/ml (mean +/- SD), respectively.

Amniotic Fluid↗
PubMed1983

The hormonal and cellular control of Sertoli cell secretion.

Sertoli cells synthesize and secrete a number of cell-specific products including androgen binding protein (ABP), as well as "serum proteins" such as transferrin. The secretion of these proteins is regulated by extra-testicular hormones such as FSH and insulin; Leydig cell-produced steroids and proopiomelano-cortin-derived peptides; and the presence of peritubular myoid cells and/or factors secreted by these cells. Many of the Sertoli cell proteins are secreted in a cyclic fashion during the different stages of the spermatogenic cycle suggesting communication between Sertoli cells and developing germ cells. The availability of quantitative measurements for Sertoli cell-specific proteins such as ABP make it feasible to follow Sertoli cell function in vivo by measuring these products in serum. A bidirectional secretion of proteins by Sertoli cells is proposed to explain the presence of specific peptides in the male reproductive tract and blood.

Androgen-Binding Protein↗
PubMed1982

Purification and characterization of androgen binding protein from rabbit epididymis.

A method for the purification of androgen binding protein (ABP) from the rabbit epididymis is presented. Epididymal extracts were submitted to sequential ammonium sulfate precipitation, androgen affinity chromatography, concanavalin A (Con A) affinity chromatography, and preparative polyacrylamide gel electrophoresis. Since the blood protein testosterone-estradiol binding globulin (TeBG) was a possible component of the epididymal extract, ABP was differentiated and separated from TeBG by affinity chromatography on Con A-Sepharose since the latter protein was shown to be completely absorbed by the lectin while the former was not. The final product was shown to be pure by polyacrylamide gel electrophoresis. Electrophoresis in the presence of sodium dodecyl sulfate revealed that ABP is comprised of subunits.

Androgen-Binding Protein↗
PubMed1982

Subunit structure of rabbit testosterone estradiol-binding globulin.

Testosterone estradiol-binding globulin (rbTeBG) of rabbit plasma was purified to homogeneity by the sequential use of ammonium sulfate precipitation, affinity chromatography on 5 alpha-dihydrotestosterone-Sepharose, reactive blue-Sepharose, and finally preparative polyacrylamide gel electrophoresis. An overall purification of greater than 3000-fold was achieved with a recovery of 32%. This purified material was shown to possess the following physical characteristics: an equilibrium dissociation constant at 4 degrees C for dihydrotestosterone estimated to be 0.59 X 10(-9) M and a half-time of dissociation of rbTeBG-dihydrotestosterone complex of approximately 9 min at 4 degrees C. The purified protein shows considerable microheterogeneity with regard to size on sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis and to charge on isoelectric focusing gels. Analytical gel electrophoresis in the presence of SDS revealed two molecular weight components of 41,000 and 45,000. Size analysis on native gels and cross-linking studies yields a molecular weight of 101,600 and 83,000, respectively. These size components were shown to be very closely related with regard to primary structure. rbTeBG was completely absorbed by concanavalin A Sepharose resin, demonstrating the glycoprotein nature of rbTeBG. Highly purified rbTeBG and unpurified rbTeBG from rabbit plasma could be photolabeled with [3H]delta 6-testosterone and when analyzed on SDS-polyacrylamide gel electrophoresis (PAGE), two major bands of radioactivity with molecular weights of 41,000 and 45,000 appeared in both samples.

Amino Acids↗
PubMed1981

Origin of the heavy and light protomers of androgen-binding protein from the rat testis.

Androgen-binding protein (ABP) isolated from rat epididymides by androgen affinity chromatography is a dimer with a native molecular weight of 85,000. When fractionated on sodium dodecyl sulfate gels two components were identified which were present in a 3:1 ratio; these were designated heavy (H) and light (L) (Mr of 45,000 and 41,000), respectively. The fact that H and L both have steroid binding sites and that they can be cross-linked suggests that both are protomers of native ABP. NH2-terminal analysis, amino acid analysis, and peptide maps suggest an extensive homology between H and L. In addition, peptide maps of H and L photolabeled with delta 6-[3H]testosterone suggest that the binding sites on these protomers are identical. ABP was also purified 33,800-fold from testes. The H and L protomers from this preparation were identical with those from the epididymides as judged by peptide maps. In addition, [35S]methionine was incorporated into the H and L synthesized by Sertoli cells in the same 3:1 ratio as in highly purified ABP from testis and epididymis. These observations suggest that L is not derived from H following secretion. Selective incorporation of [3H] fucose into the H protomer suggests that differences in carbohydrate composition account for at least part of the difference between H and L. These results support the conclusion that native ABP is composed of two kinds of protomers which exist in a ratio of 3:1. These protomers are similar polypeptides that differ partly in carbohydrate composition.

Androgen-Binding Protein↗
PubMed1981

Sertoli cells secrete both testis-specific and serum proteins.

The secretions of the Sertoli cell were examined with two polyvalent antisera--one prepared against proteins in rat serum and the other against testis-specific proteins in rete testis fluid. These antisera detected 12 serum and 9 testis-specific proteins in rete testis fluid. To determine the origin of these proteins, primary cultures enriched in Sertoli cells were incubated with [35S]methionine, and the radiolabeled proteins in the medium were immunoprecipitated. Gel electrophoresis of the two immunoprecipitates resolved eight serum and nine testis-specific proteins. These two sets of proteins were specifically bound to their respective antiserum and were immunologically distinct. Medium from Sertoli cell cultures contained 10 times more of the testis-specific proteins than did cultures enriched for testicular myoid or interstitial cells. The concentration of the serum proteins in Sertoli cell medium was 5 and 10 times greater, respectively, than in myoid or interstitial cell preparations. The proteins from Sertoli cells were next characterized on two-dimensional gels. Seven of the proteins recognized by antiserum against serum proteins had identical molecular weights and isoelectric points as serum proteins. Three of these proteins were ceruloplasmin, transferrin, and glycoprotein 2. In addition to the proteins immunoprecipitated by the two antisera, more than 60 other proteins were detected on two-dimensional gels of the total secretory proteins. We conclude that the Sertoli cell secretes many proteins, some of which are specific to the testis and others of which are similar to serum proteins.

Animals↗
PubMed1981

Immunocytochemical localization of androgen-binding protein in the male rat reproductive tract.

The localization of androgen-binding protein (ABP) in the reproductive tract of young adult male rats was studied with the peroxidase-antiperoxidase technique using frozen sections and light microscopy. Within the seminiferous tubules, a positive reaction was noted in the apical portion of the epithelium, apparently in spermatids and/or Sertoli cells. ABP was localized in granules in the apical cytoplasm of the principal epithelial cells of the proximal part of the caput epididymis and in the epithelial cells of the ductuli efferentes. The cells in the distal part of the caput as well as the corpus and cauda of the epididymis did not contain ABP. Numerous coated vesicles and multivesicular bodies were present in the supranuclear cytoplasm of the epididymal epithelium where ABP was taken up. The results indicate that ABP is taken up from the lumen by epithelial cells of the ductuli efferentes and proximal part of the caput epididymis.

Androgen-Binding Protein↗
PubMed1977

In vitro metabolism of progestins. III. The metabolic clearance rate of medroxyprogesterone acetate in monkeys.

3H-medroxyprogesterone acetate (MPA) was synthesized by selective catalytic tritiation of the delta1-olefinic bond of 6alpha-methyl-17alpha-hydroxy-pregna-1,4-diene-3,20-dione acetate. The metabolic clearance rate of MPA (MCRMPA) was determined in 9 female Rhesus monkeys by the single injection technique. The blood MCRMPA was 201 +/- 19 L/day (42.2 +/- 4.0 L/day/kg) which was approximately the same as that reported for progesterone clearance in the monkey. We conclude that a greater biological activity of MPA compared to progesterone cannot be related to its prolonged retention in the blood. Although both MPA and amino-glutethimide alter the rate of steroid metabolism in some species, neither of these agents influences the metabolic clearance rate of 3H-MPA in the monkey.

Aminoglutethimide↗
PubMed1977

Characteristics of semen collected from the cauda epididymis of conscious rams.

A technique which permits collection of the total output of spermatozoa and fluid from the cauda epididymidis of conscious rams is described. The volume of epididymal semen collected varied in an approximately inverse proportions with the frequency of collection, but there was little variation in the concentration of spermatozoa. Chemical analyses showed that cell-free cauda epididymidis fluid contained lactic acid at various concentrations, only traces of glucose but relatively high amounts of phospholipid, pregenenolone, androgen-binding protein and 5alpha-dihydrotestosterone (5alpha-DHT). Testicular spermatozoa maintained their metabolic activity and cellular integrity when stored in cauda epididymidis fluid in vitro at low temperature. After exposure to cauda epididymidis fluid for 14 days, testicular spermatozoa were predominantly glycolytic in their mode of glucose dissimilation and in this respect they resembled ejaculated spermatozoa.

Animals↗