Adult Male Serum Research Articles

The effect of acute physical and mental stress on soluble cellular adhesion molecule concentration

AimsThis study investigated the impact of acute physical and mental stress on serum concentrations of vascular cell adhesion molecule (VCAM)-1 and CX3CL1/fractalkine. Materials and methodsMale volunteers (n=20; 21.3±0.55years of age) completed a graded treadmill test to exhaustion and a 20-minute mental stress task (Stroop Color-Word Test, mental arithmetic) on separate, non-consecutive days. Heart rate (HR) was measured at baseline and throughout exercise and mental stress. Blood was collected at baseline (PRE), immediately following (POST) and 30min after (POST30) exercise and mental stress. Soluble VCAM-1 and fractalkine were quantified in participant serum via enzyme-linked immunosorbent assays. Key findingsBoth treadmill exercise and the mental stress task significantly increased participant HR; although, exercise resulted in a substantially greater increase in participant HR compared to mental stress (197.82±11.99 vs. 38.67±3.10% [p<0.001]). VCAM-1 (815.74±139.55 vs. 738.67±131.59ng/mL [p=0.002]) and fractalkine (1.032±0.33 vs. 0.59±0.20ng/mL [p<0.001]) were significantly elevated in participant serum POST maximal exercise before returning to values similar to baseline at POST30. The acute mental stress task did not significantly alter serum VCAM-1 or fractalkine at any time point. SignificanceIn conclusion, maximal aerobic exercise results in a significant elevation of the soluble adhesion molecules VCAM-1 and fractalkine in the serum of adult males that does not occur following laboratory-induced mental stress. The findings of the current investigation may suggest a novel protective role for acute aerobic exercise in vascular health via exercise-induced CAM proteolysis.

Xenoandrogenic Activity in Serum Differs across European and Inuit Populations

BackgroundAnimal and in vitro studies have indicated that human male reproductive disorders can arise as a result of disrupted androgen receptor (AR) signalling by persistent organic pollutants (POPs). Our aim in the present study was to compare serum xenoandrogenic activity between study groups with different POP exposures and to evaluate correlations to the POP proxy markers 2,2′,4,4′,5,5′-hexachlorobiphenyl (CB-153) and 1,1-dichloro-2,2-bis(p-chlorophenyl)-ethylene (p,p′-DDE).MethodsWe determined xenoandrogenic activity in the serum fraction containing the lipophilic POPs but free of endogenous hormones. Adult male serum (n = 261) from Greenland, Sweden, Warsaw (Poland), and Kharkiv (Ukraine) was analyzed. Xenoandrogenic activity was determined as the effect of serum extract alone (XAR) and in the presence of the synthetic AR agonist R1881 (XARcomp) on AR transactivated luciferase activity.ResultsThe study groups differed significantly with respect to XARcomp activity, which was increased in the Inuits and decreased in the European study groups; we observed no difference for XAR activity. We found the highest level of the AR antagonist p,p′-DDE in Kharkiv, and accordingly, this study group showed the highest percent of serum samples with decreased XARcomp activities. Furthermore, the percentage of serum samples with decreased XARcomp activities followed the p,p′-DDE serum level for the European study groups. No correlations between serum XAR or XARcomp activities and the two POP markers were revealed.ConclusionsThe differences in XARcomp serum activity between the study groups suggest differences in chemical exposure profiles, genetics, and/or lifestyle factors.

AN ANALOGUE-BASED FREE TESTOSTERONE RADIOIMMUNOASSAY DETECTS AND TRACKS TOTAL TESTOSTERONE CONCENTRATIONS.

Introduction This free testosterone (FTe) assay is routinely applied to whole serum. In whole male serum 1.5 to 2.0% of total testosterone (TTe) is FTe. (The remainder is bound to Te binding serum proteins.) The FTe assay is based on a radiolabeled Te analogue (a Te conjugate) that competes with unlabeled Te for binding to a Te-specific antibody. Two questions were asked about the assay. First, how does the assay respond when only protein-bound Te and TTe are diluted while FTe is constant? Second, how does the assay respond when FTe replaces protein-bound Te while TTe remains constant? Analogue-based FTe estimates and TTe determinations were compared. Materials and Methods Equilibrium dialysis (Nichols Institute Diagnostics) was applied to normal adult male serum. The free Te assay was applied to dialysate. A total Te assay was applied to retentate. Te was added to an aliquot of dialysate until total Te matched retentate total Te. One aliquot of serum retentate was progressively diluted with serum dialysate (no Te added). Thus, protein-bound and total Te were decreased while free Te remained constant. Another aliquot of serum retentate was progressively diluted with the dialysate containing added Te. Thus, protein-bound Te decreased and FTe increased while TTe remained constant. The FTe RIA and the TTe RIA (both from Diagnostic Products Company, Los Angeles, CA) were applied to all Te solutions. Results Free Te was not detected in serum dialysate. When TTe and protein-bound Te were progressively reduced while free Te was constant, TTe determinations progressively declined 95%, from 581 to 29 ng/dL. The median was 167 ng/dL. The analogue-based FTe determinations also progressively declined ≈95% from 1.28 to r 2 .98). When protein-bound Te was progressively replaced with FTe while total Te was constant (648 ng/dL ± 14), free Te determinations were similarly constant (1.48 ng/dL ± 0.02). Conclusions These analogue-based free Te estimates were not traceable to serum free Te concentrations. Free Te determinations were traceable to total Te concentrations and total Te concentrations only. The forms of Te tracked and the concentrations of Te detected by this assay provide direct evidence of protein-bound Te cross-reactivity. No evidence was found to support the concept that this free Te RIA tracks or detects serum free Te.

Paracrine and endocrine roles of insulin-like factor 3

Insulin-like factor 3 (INSL3) is expressed in Leydig cells of the testis and theca cells of the ovary. This peptide affects testicular descent by acting on gubernaculum via its specific receptor leucine-rich repeat-containing G protein-coupled receptor 8 (LGR8). From initial animal data showing the cryptorchid phenotype of Insl3/Lgr8 mutants, an extensive search for mutations in INSL3 and LGR8 genes was undertaken in human patients with cryptorchidism, and a frequency of mutation of 4-5% has been detected. However, definitive proofs of a causative role for some of these mutations are still lacking. More recent data suggest additional paracrine (in the testis and ovary) and endocrine actions of INSL3 in adults. INSL3 circulates at high concentrations in serum of adult males and its production is dependent on the differentiation effect of LH. Therefore, INSL3 is increasingly used as a specific marker of Leydig cell differentiation and function.

Investigation, Treatment, and Monitoring of Late‐Onset Hypogonadism in Males: ISA, ISSAM, and EAU Recommendations

Demographic data clearly demonstrate that the percentage of the population in the older age group is increasing. Androgen deficiency in the aging male has become a topic of increasing interest and debate throughout the world. Cross-sectional and longitudinal data indicate that the testosterone falls progressively with age and that a significant percentage of men over the age of 60 years have serum testosterone levels that are below the lower limits of young adult (age 20–30 years) men (1–4). The principal questions raised by these observations are whether older hypogonadal men will benefit from testosterone treatment and what will be the risks associated with such intervention. The past decade has brought evidence of benefit of androgen treatment of hypogonadal men on multiple target organs and the recent studies show short-term beneficial effects of testosterone in older men that are similar to those in younger men. This has been comprehensively reviewed and summarized by the Institute of Medicine in ‘Testosterone and Aging: Clinical Research Directions’ (5). Long-term data on the effects of testosterone treatment in the older population are limited mainly to effects on body composition and bone mass (6–11). Key questions of the effects of testosterone on patient reported outcomes and functional benefits that may retard physical or mental frailty of the elderly or improve the quality of life are not yet available. Specific risk data on the prostate and cardiovascular systems are needed.

The INSL3-LGR8 Hormonal System in Humans: Testicular Descent, Cryptorchidism and Testicular Functions

INSL3 is a member of the relaxin-insulin family, and it is expressed in pre- and postnatal Leydig cells of the testis in a variety of mammalian species, including humans. This peptide affects testicular descent during embryonic development by acting on gubernaculum via its specific receptor LGR8. From initial animal data showing the cryptorchid phenotype of Insl3/Lgr8 mutants, an extensive search for mutations in the INSL3 and LGR8 genes was undertaken in human patients with cryptorchidism. Six mutations in INSL3 and one in LGR8 genes have been detected exclusively in men with undescended testes, with an overall frequency of mutation in cryptorchid or ex-cryptorchid men of 4-5%. Definitive proofs of a causative role for many of these mutations are still lacking. However, the specific association with cryptorchidism actually suggests that they might be responsible for the phenotype. Apart from the role in testicular descent and cryptorchidism, more recent data suggest additional yet unidentified endocrine and paracrine actions in adults. INSL3 is produced constitutively, but in a differentiation-dependent manner by the adult Leydig cells, and its production and secretion is dependent on LH. INSL3 circulates at high concentrations in serum of adult males and it is increasingly used as a specific marker of Leydig cell differentiation and function. Research is needed to clarify the possible paracrine role of the INSL3/LGR8 system in the testis and ovary, and endocrine effects in many tissues where LGR8 expression has been identified.

The effective free fraction of estradiol and xenoestrogens in human serum measured by whole cell uptake assays: physiology of delivery modifies estrogenic activity.

The biological activity of natural estrogens is influenced by the degree to which they bind to serum proteins. To determine directly how serum affected the uptake of estradiol, we compared the whole cell uptake of [3H]estradiol in intact MCF-7 human breast cancer cells from serum-free medium with the uptake from 100% serum from adult men. In estrogen receptor saturation assays, 28.9 times more estradiol was required in serum to occupy the same number of estrogen receptors as was required in serum-free medium (SFM), suggesting that the effective free fraction of estradiol in adult male serum was 3.46% (1/28.9). Since most xenoestrogens are not available in tritium-labeled form, the cell uptake of unlabeled xenoestrogens could not be measured directly with saturation analysis. Therefore, we developed the relative binding affinity-serum modified access (RBA-SMA) assay to determine the effect of serum on the access of nonradioactive xenoestrogens to estrogen receptors within intact MCF-7 cells. Serum modified access (SMA) was calculated by dividing the relative binding affinity (RBA, relative to estradiol) measured in 100% serum, by the RBA measured in serum-free medium. An SMA > 1 indicated that the xenoestrogen had greater access to estrogen receptors than estradiol from serum. In contrast, an SMA < 1 indicated that the xenoestrogen had less access to estrogen receptors from serum than did estradiol. The synthetic estrogen diethylstilbestrol (DES) binds poorly to sex hormone binding globulin (SHBG), and DES showed enhanced access in serum, SMA = 6.2. Additional calculations through the Ki (inhibition constant) indicated that this corresponded to an effective free fraction of 26.9% for DES in serum. The phytoestrogens, coumestrol, genistein, and equol, showed substantial enhanced access in serum, over 10-fold relative to estradiol (SMA = 12.1, 10.3, and 11.3, respectively), and effective free fractions in serum of 47.8, 45.8, and 49.7%, respectively. Since most in vitro assays of xenoestrogens do not address how serum influences their bioactivity, the estrogenic activity of these phytoestrogens would be underestimated. Conversely, biochanin A showed decreased access from serum (SMA = 0.44) and had an effective free fraction of 2.4%; its estrogenic activity would be overestimated in serum-free assays.

EFFECTS ON PUBERTAL GROWTH AND REPRODUCTION IN RATS EXPOSED TO LEAD PERINATALLY OR CONTINUOUSLY THROUGHOUT DEVELOPMENT

The reproductive, endocrine, and growth effects of developmental lead exposure were assessed using a rat model in which 0.6% lead acetate (w/v) was administered in the drinking water ad libitum during different developmental periods to determine if lead actions were a result of direct effects of continuous exposure to the metal ion or secondary to disrupted neonatal "endocrine imprinting." Sprague Dawley rats were exposed to lead: (1) from gestational d 5 through birth; (2) during pregnancy and lactation; (3) during lactation only; (4) from birth through adulthood; or (5) from gestational d 5 through adulthood. Lead effects were measured on the development of aspects of the reproductive system, adult sex steroid levels, and growth rates in both male and female animals. The relative weights of male secondary sex organs in adult offspring were not significantly affected in any of the lead-treated groups. In contrast, female pups exposed to lead from birth through adulthood or from gestational day 5 through adulthood were observed to have significantly delayed vaginal opening and disrupted estrus cycling. These effects on female reproductive physiology were not observed in animals where lead exposure was confined only to pregnancy or lactation. Significant suppression of adult mean serum testosterone levels was only observed in male pups exposed to lead continuously from gestational age 5 d throughout life. Lead decreased birth weight in all animals exposed in utero and mean body weights were significantly decreased in all lead-treated groups up to weaning. Analysis of growth curves revealed that all lead-treated groups had significantly reduced growth rates during lactation. However, in addition, in male pups exposed to lead during pregnancy and lactation, from birth or from gestational age 5 d, growth rates were also significantly reduced during puberty. Postpubertal growth rates were unaffected in any lead-treated group. Thus, delayed female reproductive development and suppression of adult male serum testosterone concentration required continuous exposure to the heavy metal. Little evidence was observed for an alteration of "endocrine imprinting" by lead on either reproductive or growth parameters. Exposure during early development (pregnancy and lactation) resulted in no permanent effects in this model other than small (10%) decreases in the body weight of pups postpuberty.

Development, validation and application of a two-site enzyme-linked immunosorbent assay for activin-AB.

Monoclonal antibodies, specific for the beta A and beta B subunits of activin, were used to develop a new two-site ELISA for activin-AB. The assay had a detection limit of 0.19 ng/ml. High concentrations of activin-AB were found in bovine, ovine and porcine follicular fluids (FF), with less in human FF (1310, 1730, 688 and 7 ng/ml respectively). Recovery of spiked activin-AB standard from human, bovine and ovine FFs and from homogenized human placental extracts averaged 91%, 115%, 115% and 94% respectively. Within-plate coefficients of variation for different concentration of activin-AB were between 1.3% and 2.67%. The between-plate coefficient of variation was 5.5%. Cross-reactivity experiments showed the high specificity of the assay for activin-AB, with inhibin-A, inhibin-B, follistatin, activin-A and activin-B all cross-reacting < 0.2%. Incubation with high concentrations of follistatin (500 ng/ml) prior to assay did not affect the recovery of activin-AB. Samples of bovine, porcine, ovine and human FF gave dose responses parallel to that of the standard, as did bovine granulosa cell-conditioned media. In human and porcine FF, levels of activin-A and activin-AB were similar whereas, in bovine and ovine FF, activin-A levels were approximately threefold higher than activin-A, nearly all of the endogenous activin-AB in bovine FF was detected in the eluate from gel permeation chromatography with an M(r) of > 700000 indicating its association with higher molecular weight binding protein(s). By contrast, after denaturation, immunoreactive activin-AB was detected with an M(r) of approximately 25000 consistent with the complete dissociation from binding proteins. Activin-A was detected in relatively high concentrations in human FF (approximately 5 ng/ml), homogenized placental extracts (4.35-95.5 ng/g), sera from pregnant women (> 4 ng/ml) and amniotic fluid (3-13 ng/ml), and in much lower concentrations in postmenopausal serum (500 pg/ ml), normal cycle serum (100-200 pg/ml), serum from gonadotrophin-treated women (200 pg/ml), and normal adult male serum (225 pg/ml). Activin-A was also found in the culture media from explants of human amnion, chorion, maternal decidua and placenta. In marked contrast, activin-AB was undetectable (< 0.19 ng/ml) in all of these samples with the exception of human FF (approximately 7 ng/ml). In conclusion, we have developed a sensitive and specific ELISA to measure total (bound+free) activin-AB. Preliminary results show a more restricted distribution of this isoform compared with activin-A. The presence of high levels of both activin-A and activin-AB in FF suggests a function for both isoforms in the developing ovarian follicle.

The presence of müllerian inhibiting substance in human seminal plasma.

Müllerian inhibiting substance (MIS), produced by testicular Sertoli cells, is present in adult male serum. The first aim of this study was to determine if MIS is present in seminal plasma. Using an enzyme-linked immunosorbent assay (ELISA), we measured MIS concentrations in seminal plasma from 23 donors exhibiting normal (WHO criteria) sperm qualities, and 169 patients with subnormal sperm parameters. The second aim of this study was to examine a potential relationship between MIS and sperm motility. MIS concentrations in seminal plasma ranged from 0.5 to 3.6 ng/ml in donors and from 0.5 to 17.8 ng/ml in patients. Motility index (MI, mean +/- SEM) for all patient samples was lower compared with donors (113.3 +/- 3.2 and 198.3 +/- 13.5, P < 0.00001), while mean MIS concentration +/- SEM) was higher (4.2 +/- 0.3 and 1.4 +/- 0.2, P < 0.0003). When the patients were stratified into Groups I (motility < 50%, n = 42) and II (motility > 50%, n = 127), the MI (mean +/- SEM) values were 62.3 +/- 3.8 and 130.2 +/- 2.7 respectively (P < 0.0001 for both compared with donors) and mean MIS concentrations (+/- SEM) were 5.4 +/- 0.6 and 3.9 +/- 0.3, respectively (P < 0.0001 and P < 0.001 compared with donors). The inverse relationship between MIS concentration in seminal plasma and motility index suggests that MIS may have a function in modulating motility.

Retardation of fetal brain cell growth during maternal starvation: circulating factors versus altered cellular response.

Maternal starvation inhibits fetal brain development during late gestation in the rat. To determine whether intrinsic or extrinsic factors might be the principal contributor to altered growth, brain cells from 20 day fetuses were cultured in a 96 well plate with MEM and 10% adult rat serum. Tissue growth was monitored by spectrophotometric measurement of the mitochondrial reduction of a chromagen 3-(4,5 dimethylthiazol-2-yl)-2,5 diphenyl tetrazolium bromide (MTT). After 1, 4 or 6 days incubation, MTT activity in non confluent cultures was shown to be directly related to tissue mass. When fetal brain cell cultures were incubated with 1% and 10% concentrations of adult rat serum, an 11-fold increase in MTT activity paralleled a 15-fold increase in tritiated thymidine incorporation. The impact of maternal starvation on fetal brain cell growth was examined by measuring MTT activity in fetal brain cells from fed and starved mothers. When cultures were incubated for 6 days with graded concentrations of fed adult serum (1.25-10%), the MTT response was slightly but consistently lower in cells from starved when compared with cells from fed mothers. By contrast, a marked difference in MTT activity which was paralleled by a lower DNA content became apparent when fetal rat brain cells were incubated with starved adult serum. Fetal serum and adult male serum were found to support growth equally well, while incubation of fetal brain cells with maternal sera resulted in lower MTT values than with the corresponding fetal sera. When cells were incubated with fetal sera pooled from starved mothers, MTT activity was decreased by 42 to 45%.(ABSTRACT TRUNCATED AT 250 WORDS)

Chromatofocusing profile of purified human α-fetoprotein and albumin differs from those of crude samples: Effect of protein concentration on the elution of the sample

Chromatofocusing was utilized to characterize charge microheterogeneity of purified human α-fetoprotein (AFP) and human serum albumin (HSA). Crude cord blood samples yielded three isoforms: AFP-IA, IB, and II, with p Is 4,57 (52%), 4,27 (43%), and <4,00 (5%), respectively. In contrast, 10 μg of purified AFP or 250,000 cpm of 125I-AFP eluted entirely as isoform AFP-II. 125I-AFP focused in the presence of crude cord blood, amniotic fluid, adult male serum, or 25 mg purified HSA resulted in elution profiles similar to those of crude cord blood. Pure AFP focused along with 0.1, 1.0, 5.0, or 10 mg HSA showed a gradual shift from AFP-II to AFP-I. With ≥5 mg HSA, isoform I was further resolve into AFP-IA and IB. Similarly, 250,000 cpm of 125I-HSA, which also eluted entirely as isoform II, showed a gradual shift to isoform I when increasing concentrations of unlabeled HSA were added. The resolution of isoform HSA-I in HSA-IA, IB, and IC was again improved with ≥5 mg unlabeled HSA. When carrier proteins of varying p I values were chromatofocused along with purified AFP, it was observed that only those proteins withp Is in the range of AFP caused significant alteration in the relative distribution of AFP. We conclude that sample protein concentration and composition must be carefully considered when chromatofocusing is being used for purified samples and when the elution profiles of samples from different origins and varying protein concentrations are being compared.

The impact of maternal serum on development of enolase activity in fetal rat brain cell culture

The effect of gestational age on serum-mediated changes in enolase activity was tested in a fetal rat brain cell culture. After 6 days exposure to graded concentrations (10 and 20%) of nonpregnant female rat sera, enolase activity in brain cell cultures increased from 2.83 ± 0.03 to 3.74 ± 0.19 μmol/min/mg protein, P < 0.01. By contrast, similar concentrations of 20-day maternal serum progressively decreased enzyme activity from 1.52 ± 0.14 to 1.19 ± 0.08 μmol/min/mg protein. The inhibitory effect was apparent at 14 days gestation and became progressively greater during late gestation to reach a maximum at 20 days. Combining equal concentrations of 20-day pregnant with either nonpregnant or adult male serum neutralized the inhibitory activity. When serum from 20-day pregnant rats was partitioned by a dialysis membrane with a 50,000 MW pore size, inhibitory activity could be similarly neutralized by male or nonpregnant female serum. When 20-day maternal serum was passed successively through filters with a greater than 300,000, 100,000, and 50,000 MW exclusion, the inhibitory activity was apparent in all fractions excluded by a molecular weight of 50,000. No inhibition was apparent in fractions that were not excluded by 50,000 MW pore size. Inhibition of enolase activity was greatest in the fraction with MW greater than 300,000. Binding of IGF II could also be demonstrated in this fraction. Binding of IGF II was evident in the fraction greater than 100,000 MW but could not be demonstrated in fractions with a lower molecular weight. The presence of mRNA for IGF II in 20-day fetal rat brain cell cultures was evident when total cellular RNA was analyzed by an RNAase protection assay. It is proposed that a high-molecular-weight component of maternal serum in late gestation can bind endogenously generated IGF II. Such binding, by depleting the necessary growth factors, could inhibit in vitro growth and development of enolase activity.

Long-Term Administration of Gonadotropin-Releasing Hormone in Men with Idiopathic Hypogonadotropic Hypogonadism

The effect of long-term administration of gonadotropin-releasing hormone (GnRH) for induction and maintenance of sexual maturation was characterized in 23 men with idiopathic hypogonadotropic hypogonadism. Twenty-two men achieved normal adult male serum testosterone concentrations (575 +/- 33 ng/dL; p less than 0.0001 compared with the baseline mean of 61 +/- 6 ng/dL) that were sustained in 21 men for up to 36 months with bolus doses of GnRH varying from 25 to 300 ng/kg body weight administered every 2 hours. Pulsatile luteinizing hormone (LH) secretion occurred in all 23 men, with mean levels of LH (14.7 +/- 1.3 mlU/mL) and follicle-stimulating hormone (11.3 +/- 1.3 mlU/mL) within or above the normal range for adult men. Mature sperm were observed in the ejaculates of 20 men, with counts ranging from less than 1 X 10(6) to 96 X 10(6)/mL. Increasing responsiveness of the pituitary-gonadal axis to GnRH was shown in 6 men. Men with idiopathic hypogonadotropic hypogonadism present a useful model to study the onset and maintenance of reproductive function in men.

Human seminal alpha-inhibins: detection in human pituitary, hypothalamus, and serum by immunoreactivity.

An antiserum generated in rabbits against human seminal alpha-inhibin-52 has been used to develop a sensitive radioimmunoassay for the detection of alpha-inhibins. The alpha-inhibin-52 antiserum reacts with alpha-inhibin-92 and alpha-inhibin-31 with equal avidity. These peptides were found to be present in human pituitary, hypothalamus, and serum. In exclusion chromatography on Sephadex G-100, the immunoreactive material eluted in a large molecular size region. Immunoblot analysis of column-derived fractions of these extracts revealed the presence of alpha-inhibin-92. The mean concentrations of immunoreactive alpha-inhibin were found to be 7.2 ng/ml in normal adult male serum, 70.3 ng/g (wet weight) of pituitary, and 12.9 ng/g (wet weight) of hypothalamus. This communication reports on the evidence for the existence of gonadal peptides in the brain.

Measurement of testosterone binding sites.

ABSTRACT A Sephadex assay technique, in use in our laboratory for the study of steroid-protein interactions, allows expression of the molar concentration of a testosterone-binding component of human serum of high binding affinity in terms of the molar concentration of testosterone-binding sites: examples with model calculations are provided. The presence in the assay system of competing protein of low binding affinity (i. e. albumin) and of competing steroid (i. e. endogenous, radioinert testosterone) is taken into account when estimating the testosterone-binding component. With the aid of some approximations, the distribution of bound testosterone between the testosterone-binding component and albumin is readily calculated. From this estimate and the total testosterone concentration, the serum concentration of unbound testosterone (Su) is calculated. The concentration of the testosterone-binding component is thus estimated to be of the order of 8 × 10−7M and 1 × 10−7M in late pregnancy serum and adult male serum, respectively, assuming that there is only one binding site for testosterone in the protein molecule. The testosteronebinding component appears to play a role at least equal to that of albumin in binding endogenous testosterone in adult male serum; the testosterone-binding component is clearly dominant in this role in late pregnancy serum. The values for Su at 25° are calculated to be of the order of 3 × 10−10M and 1 × 10−11M in adult male serum and late pregnancy serum, respectively, corresponding to 1.7% and 0.27% of the total testosterone concentration, respectively. It is suggested that the physiologically effective level of circulating androgen may correlate better with the serum concentration of unbound testosterone (Su) than with that of total testosterone.

The growth of mouse prostate during culture in vitro in chemically defined and natural media, and after transplantation in vivo: The effects of insulin and normal human serum

1. 1. Organ cultures of mouse ventral prostate in Trowell's chemically defined medium T8, which contains insulin, showed irregular epithelial hyperplasia after 5 to 7 days. 2. 2. Cultures in serum-containing media or in T8 without insulin retained their normal structure but a similar hyperplasia was produced in cultures in serum-containing media by adding insulin to the medium. 3. 3. The hyperplasia regressed if the cultures were transferred to serumcontaining media without insulin, or transplanted isologously to normal young adult mice. 4. 4. The hyperplastic cells persisted if transferred to an insulin-free chemically defined medium, perhaps because of residual hormone in the cells. 5. 5. Insulin therefore under the conditions of these experiments has a growth-stimulating effect on the epithelium of the mouse ventral prostate. 6. 6. There is a factor present in normal human adult male serum and in normal young adult male mice which destroys the hyperplastic cells and restores the prostatic glands to their normal structure.