Testosterone and Androstenedione Blood Production Rates in Normal Women and Women with Idiopathic Hirsutism or Polycystic Ovaries

The average plasma testosterone concentration of women with either hirsutism or polycystic ovaries and hirsutism was higher (p < 0.01) than that of normal women although the ranges overlapped. Testosterone blood production rates averaged 830 +/- 120 SE and 1,180 +/- 310 SE mug per day in the two groups of hirsute women and 230 +/- 33 SE mug per day in normal women. The ranges did not overlap.The testosterone metabolic clearance rates of hirsute women (1,090 +/- 140 SE L per day) and of men (1,240 +/- 136 SE L per day) were significantly higher than those of normal women (590 +/- 44 SE L per day). These differences persisted when the metabolic clearance rates were corrected for surface area. We suggest that testosterone metabolic clearance rates vary directly with some function of testosterone production.The mean plasma androstenedione levels (2.8 +/- 0.35 SE and 2.8 +/- 0.30 SE mug per L) and production rates (6,060 +/- 450 SE and 7,360 +/- 345 SE mug per day) of the women with hirsutism or polycystic ovaries, respectively, were significantly higher than those of normal women (1.5 +/- 0.22 SE mug per L; 3,300 +/- 830 SE mug per day). The androstenedione metabolic clearance rates were the same in each group. Plasma androstenedione was the precursor of 49% of plasma testosterone in normal women and of 26% of plasma testosterone in hirsute women. Thus, 74% of the plasma testosterone in these subjects must have been either secreted or derived from a precursor that did not enter the plasma androstenedione pool.     

The metabolic clearance rate and origin of plasma dihydrotestosterone in man and its conversion to the 5α-androstanediols

Dihydrotestosterone metabolism was studied with a constant infusion technique in three men, three women, five hirsute women, and four estrogen-treated hirsute women. The mean dihydrotestosterone metabolic clearance rate was higher in men (336 liters/24 hr per m(2) [range, 239-448]) than in women (153 liters/24 hr per m(2) [range, 108-184]). The metabolic clearance rates in hirsute patients were intermediate between those men and women and were decreased by estrogen treatment. These observations demonstrate similarities in the metabolic rates of testosterone and dihydrotestosterone.The conversion of plasma testosterone and androstenedione to dihydrotestosterone was studied in men and hirsute women. Approximately 4 and 2% of plasma testosterone and androstenedione, respectively, were converted to plasma dihydrotestosterone in both groups. From these observations it was determined that a major fraction of plasma dihydrotestosterone was derived from these plasma precursors rather than from glandular secretion.Both 5alpha-androstan-3alpha,17beta-diol (3alpha-diol) and 5alpha-androstan-3beta,17beta-diol (3beta-diol) were identified in plasma during dihydrotestosterone and testosterone infusions. The conversion ratio of dihydrotestosterone to 3alpha-diol (C(BB) (DHT-3alpha)) was greater than the conversion ratio to the 3beta-isomer (C(BB) (DTH-3beta)) in all the patients studied. Both C(BB) (DHT-3alpha) and C(BB) (DHT-3beta) were higher in men (mean values of 0.151 [range, 0.110-0.222] and 0..031 [range, 0.022-0.042]) than in women (means of 0.044 [range, 0.037-0.048] and 0.012 [range 0.010-0.013]). A smaller fraction of testosterone was converted to 3alpha-diol and 3beta-diol.     

Androstenedione and Its Conversion to Plasma Testosterone in Congenital Adrenal Hyperplasia

The plasma concentration, production rate, and conversion ratio of androstenedione and testosterone were studied in seven children with congenital adrenal hyperplasia (CAH) of the 21-hydroxylase type. Plasma androstenedione and testosterone measured by double isotope derivative assay and estimated blood production rates were manyfold increased in the untreated state, markedly suppressed with glucocorticoid, and increased after the administration of ACTH.The metabolic clearance rate when corrected for body size and the conversion ratio of androstenedione to testosterone were similar to previously determined values in normal adults. Consideration of the androgen concentrations and conversion ratios indicates that in children with CAH, 76% of the plasma testosterone in prepubertal females and 36% in males are derived from peripheral conversion of blood androstenedione. The calculated amount of testosterone unaccounted for by peripheral conversion is similar to normal prepubertal values. This approach indicates that virilization in these children results from increased levels of testosterone but that the major source in CAH of this potent androgen is androstenedione secreted by the adrenal cortex.     

Splanchnic Extraction and Interconversion of Testosterone and Androstenedione in Man

A constant infusion of ³H-testosterone and ¹⁴C-androstenedione was administered to four human subjects, two males and two females, until the concentrations of radioactive testosterone and androstenedione in systemic plasma became constant. At that time the concentrations of radioactive testosterone and androstenedione in hepatic vein plasma were determined. Splanchnic extraction of testosterone and androstenedione and the contribution of the splanchnic system to the blood interconversion of testosterone and androstenedione were calculated.     

A comparative study of steroid concentrations in human adipose tissue and the peripheral circulation

Using radioimmunological methods, levels of cortisol, dehydroepiandrosterone, androstenedione, testosterone, oestrogens (oestradiol + oestrone), progesterone and 17 alpha-hydroxyprogesterone were determined in adipose tissue and peripheral blood obtained during surgical treatment of patients with non-endocrine diseases. The steroid content of human adipose tissue was observed to be extremely high relative to that in the general circulation, giving a tissue/serum ratio of 0.4 to 13.2. The concentration of steroids decreased in the following order: dehydroepiandrosterone greater than cortisol greater than androstenedione greater than progesterone greater than testosterone greater than 17 alpha-hydroxyprogesterone greater than oestradiol + oestrone. This sequence is different from that found in blood. When anthropometric variables were taken into consideration, the adipose tissue mass of severely obese subjects contained a steroid pool far greater than that in the total blood volume.     

A Study of the Endocrine Manifestations of Hepatic Cirrhosis

The clinical features and hormonal abnormalities were surveyedin 117 men with cirrhosis of the liver. Compared with healthymen of similar ages, the patients had significantly lower metabolicclearance rates, plasma production rates and total and freelevels of testosterone, reduced testosterone responses to humanchorionic gonadotrophin stimulation, higher oestradiol, luteinizinghormone and follicle stimulating hormone levels and higher bindingcapacities of sex steroid binding globulin. The peripheral conversionof testosterone to oestradiol was also found to be significantlyincreased. However, the metabolic clearance and plasma productionrates of oestradiol were not significantly different from thoseof healthy men. Patients who were severely ill with liver failureand one with haemochromatosis had low levels of luteinizinghormone and follicle stimulating hormone and sub-normal responsesto clomiphene and luteinizing hormone-releasing hormone. Higher plasma oestradiol levels were found in patients withgynaecomastia and spider naevi than in those without these signs.However, the clinical features of androgen deficiency—thatis, testicular atrophy, impotence and loss of secondary sexhair—were only poorly related to the low testosteronelevels, and production rates and longtitudinal studies indicatedthat the hormonal levels, endocrine features and severity ofthe liver disease could change independently. It is concluded that the clearance of oestradiol from plasmais not limited by liver disease in all patients, and that reduceddegradation of oestrogens is not the initial event in the sequenceleading to the hormonal abnormalities of cirrhosis. While gonadotrophindeficiency occurs with liver failure and in some patients withhaemo-chromatosis, the more usual findings are of elevated gonadotrophinlevels and a poor Leydig cell response to chorionic gonadotrophin.These suggest that the hypo-gonadism is primary in most patientswith cirrhosis. The causes of the high oestradiol levels werenot discovered. Increased peripheral conversion of precursorsto oestradiol or increased testicular secretion of oestradiolare possibilities. The high binding capacities of sex steroidbinding globulin were not significantly correlated with eitherthe low testosterone or high oestradiol level and the causeof this abnormality remains uncertain. The low metabolic clearancerates of testosterone appeared to result from the increasedplasma protein binding of testosterone. The discrepancies inthe expected relationships between the hormone and clinicalchanges suggest that factors other than those studied are alsoinvolved in the genesis of the endocrine features of hepaticcirrhosis.     

Metabolic clearance rate and blood production rate of testosterone and dihydrotestosterone in normal subjects, during pregnancy, and in hyperthyroidism

The metabolic clearance rate (MCR) and blood production rate (BP) of testosterone (T) and dihydrotestosterone (DHT), the conversion of plasma testosterone to plasma dihydrotestosterone, and the renal clearance of androstenedione, testosterone, and dihydrotestosterone have been studied in man. In eight normal men, the MCR(T) (516+/-108 [SD] liters/m(2)/day) was significantly greater than the MCR(DHT) (391+/-71 [SD] liters/m(2)/day). In seven females, the MCR(T) (304+/-53 [SD] liters/m(2)/day) was also greater than the MCR(DHT) (209+/-45 [SD] liters/m(2)/day) and both values were less than their respective values in men (P < 0.001). In men the conversion of testosterone into dihydrotestosterone at 2.8+/-0.3% (SD) was greater than that found in females, 1.56+/-0.5% (SD) (P < 0.001). In five pregnant females the MCR(T) (192+/-36 [SD] liters/m(2)/day), the MCR(DHT) (89+/-30 [SD] liters/m(2)/day) and the conversion of testosterone into dihydrotestosterone (0.72+/-0.15%) (SD) were significantly less than the values found in nonpregnant women. In five females with hyperthyroidism, the MCR for testosterone and dihydrotestosterone were similar to those observed in pregnant females, but the conversion of testosterone into dihydrotestosterone (2.78+/-1.7%) (SD) was greater, and similar to that found in men. In men the production of dihydrotestosterone was 0.39+/-0.1 (SD) mg/day, 50% being derived from the transformation of plasma testosterone. In women the production of DHT was 0.05+/-0.028 (SD) mg/day, only 10% coming from testosterone. During pregnancy, the production of testosterone and dihydrotestosterone are similar to that in normal women. In three patients with testicular feminization syndrome (an adult with hyperthyroidism and two children) these two MCRs were greatly reduced compared to the normal females, but the conversion of testosterone into dihydrotestosterone was in the limits of normal male rangeIn the normal subjects the renal clearance of androstenedione was greater than that of testosterone and dihydrotestosterone. Less than 20% of the dihydrotestosterone and less than 10% of the androstenedione in the urine is derived from the plasma dihydrotestosterone and androstenedione.     

Effect of metabolic clearance rate and hepatic extraction of insulin on hepatic and peripheral contributions to hypoglycemia.

Effects of alterations in metabolic clearance rates, hepatic extraction, and plasma concentrations of insulin on hepatic and peripheral contribution to hypoglycemia and glucose counterregulation were studied in conscious dogs. Since insulin and sulfated insulin had markedly different metabolic clearance rates (34 +/- 1 vs. 16 +/- 1 ml/kg per min, respectively) and fractional hepatic extraction (42 +/- 1% vs. 15 +/- 2%, respectively), biologically equivalent amounts infused intraportally produced twofold higher hepatic vein and artery sulphated insulin concentrations and concentrations that were 30% higher in the portal vein. This significantly larger arterial/portal concentration ratio (0.67 vs. 0.45, respectively) permitted assessment of differential distribution of insulin on glucose turnover using [3-3H]glucose. Insulin and sulfated insulin (1 and 2 mU/kg per min) caused similar hypoglycemia. While insulin transiently suppressed glucose production and increased glucose disappearance, sulfated insulin had significantly greater effects on glucose disappearance and clearance, without suppression of glucose production. Despite similar hypoglycemia, sulfated insulin caused greater increment in glucagon. 3 mU/kg per min insulin caused more rapid and greater hypoglycemia, greater glucose clearance, and greater glucagon increments without suppression of glucose production, which indicates that with larger doses of insulin counterregulation can absolutely mask the suppressive effect of insulin. The effects of insulin and sulfated insulin were evaluated using euglycemic clamp to eliminate interference from stimulated counterregulation. Sequential infusion of 1 and 2 mU/kg per min of both insulins suppressed endogenous glucose production to 0 at 150 min, which indicates that the apparent lack of a hepatic effect of sulfated insulin during hypoglycemia was masked by greater counterregulation. This greater counterregulation may reflect greater peripheral glucose clearance, and prevented greater hypoglycemia than after the same insulin doses. The results indicate that the different rates of removal and the total metabolic clearance rate caused different concentrations and relative distribution between the portal and arterial blood compartments, leading to the significantly different contributions by the liver and peripheral tissues to the same hypoglycemia.     

3α-Androstanediol Kinetics in Man

Kinetics of 5alpha-androstane-3alpha, 17beta-diol (3alpha-diol) were studied in man. Clearance rates were determined by both the constant infusion and single injection techniques. Production rates were calculated as the product of clearance rate data and plasma values in the a.m. obtained by a radioimmunoassay specific for 3alpha-diol. Mean metabolic clearance rates were 1,776+/-492 (SD) liters/day in males and 1,297+/-219 (SD) liters/day in females. Metabolic clearance rates by single injection were similar. Calculated production rates are 208+/-26 (SD) mug/day in males and 35+/-11 mug/day in females, which are significantly different. Hepatic extraction of 3alpha-diol determined by hepatic vein catheterization during constant infusion was 76% which was greater than expected from information on in vitro binding in plasma. The kinetic data is of interest since 3alpha-diol has a calculated inner pool (V(1)) volume of 12-14 liters, similar to 17beta-hydroxyandrost-4-en-3-one (testosterone) and 5alpha-androstan-17beta-ol-3-one (dihydrotestosterone), but the calculated outer pool (V(2)) of 33.5 liters is very large as are the metabolic rate and transfer constant. In contrast to testosterone and dihydrotestosterone, 3alpha-diol, although bound to sex hormone binding globulin, has a high metabolic clearance of which a large fraction represents extrahepatic (splanchnic) metabolism. A production rate of 3alpha-diol similar to dihydrotestosterone together with rather unique kinetic characteristics encourages further investigation of the biological role of this potent androgen.     

Metabolism of testosterone and androstenedione in normal and ovariectomized women

Metabolic clearance rates of testosterone (MCR^T) and androstenedione (MCR^A) were determined twice during the same cycle in six normal women, using a constant infusion of testosterone-³H and androstenedione-¹⁴C. Nonlabeled steroids served as internal standards. Plasma concentrations of testosterone (i^T) and androstenedione (i^A) were measured, and the blood production of testosterone (P_B^T) and of androstenedione (P_B^A) were calculated. The interconversions of these two steroids were also estimated. Six ovariectomized women were studied in the same manner. For testosterone, the mean i^T in the normal women was not significantly different from that in the ovariectomized subjects, whereas the MCR^T and P_B^T were significantly lower in the ovariectomized subjects. For androstenedione, the mean MCR^A values of the two groups of subjects were not different, whereas the i^A and P_B^A in the normal women were about double those in the ovariectomized subjects. In comparing the follicular and luteal phases of the menstrual cycle in four of six subjects there was no difference in i^T, MCR^T, or P_B^T, whereas i^A, MCR^A, and P_B^A were increased in the luteal phase. In one ovariectomized woman infused with testosterone and androstenedione at physiologic levels, MCR^T doubled but MCR^A remained the same. After six wk on estrogen, the same subject did not show any change in MCR^T after infusion of testosterone. It is suggested that MCR^T depends on P_B^T and on plasma binding of testosterone which is partly estrogen dependent.     

Receptor-independent low density lipoprotein transport in the rat in vivo. Quantitation, characterization, and metabolic consequences.

Receptor-independent low density lipoprotein (LDL) transport plays a critical role in the regulation of plasma cholesterol levels; hence, these studies were done to characterize this process in the tissues of the rat. High rates of receptor-independent clearance were found in the spleen, but other organs, like liver, gastrointestinal tract, and endocrine glands manifested lower clearance rates that varied from 3 to 9 microliter/h per g, while the rates in nervous tissue, muscle, and adipose tissue were less than 1 microliter/h per g. Receptor-dependent uptake was much higher in liver (85 microliter/h per g) and adrenal gland (219 microliter/h per g), but was also low in most other tissues. At normal plasma LDL concentrations, 67% of the receptor-dependent transport in the whole animal was accounted for by LDL uptake in the liver. In contrast, the receptor-independent uptake found in the whole animal took place in many organs, including skeletal muscle (20%), liver (16%), small bowel (15%), skin (10%), and spleen (7%). Furthermore, in liver, the rate of cholesterol synthesis could be varied 11-fold, yet the rate of receptor-independent LDL clearance remained constant at approximately 8 microliter/h per g. When the circulating levels of LDL were systematically increased, receptor-independent LDL clearance also remained constant, so that hepatic LDL-cholesterol uptake by this mechanism increased linearly, from 1 to 20 micrograms/h per g, as the plasma LDL-cholesterol level was increased from 10 to 250 mg/dl. Finally, when equal amounts of LDL-cholesterol were delivered into the liver by either the receptor-dependent or receptor-independent mechanism, there was significant suppression of cholesterol synthesis and an increase in cholesteryl esters. Thus, in any situation in which receptor-dependent LDL degradation is lost, cholesterol balance in the whole animal and across individual organs is maintained by receptor-independent mechanisms, although when the new steady state is achieved, circulating levels of LDL must necessarily be very much increased.     

Maximal activity of phosphate-dependent glutaminase and glutamine metabolism in septic rats

The activity of phosphate-dependent glutaminase and glutamine metabolism by tissues known markedly to utilize or synthesize glutamine (or both) were studied in rats made septic by cecal ligation and puncture technique and compared with the same measures in rats that underwent sham operation (laparotomy). Blood glucose level was not markedly different in septic rats, but lactate, pyruvate, alanine, and glutamine levels were markedly increased. Conversely, blood ketone body concentrations were significantly decreased in septic rats. Both plasma insulin and glucagon levels were markedly elevated in response to sepsis. The maximal activity of phosphate-dependent glutaminase was decreased in the small intestine, increased in the kidney and mesenteric lymph nodes, and unchanged in the liver of septic rats. Arteriovenous concentration difference measurements across the gut showed a decrease in the net glutamine removed from the circulation in septic rats. Arteriovenous concentration difference measurements for glutamine showed that both renal uptake and skeletal muscle release of the amino acid were increased in response to sepsis, whereas measurements across the hepatic bed showed a net uptake of glutamine in septic rats. Enterocytes isolated from septic rats exhibited a decreased rate of utilization of glutamine and production of glutamate, alanine, and ammonia, whereas lymphocytes isolated from septic rats showed an enhanced rate of utilization of glutamine and production of glutamate, aspartate, and ammonia. It is concluded that, during sepsis, glutamine uptake and metabolism are enhanced in renal and lymphoid tissue but decreased in that of the small intestine, with increased rates of release by skeletal muscle; however, the liver appears to utilize glutamine in septic rats.     

Comparison of translocation of different types of microorganisms from the intestinal tract of burned mice

The aim of this study was to compare the ability of various microorganisms to translocate from the intestine to the mesenteric lymph nodes (MLNs), liver, and spleen in a burned mouse model. Balb/c mice were gavaged with 1 x 10(9) or 1 x 10(10) of one of 11 different microorganisms. All animals were then given a 20% burn. Survival after 10 days showed no significant difference between any of the groups at the 10(10) dose. At the 10(9) dose, significantly higher survival rates were found in three of the 11 strains. Microbial translocation (gavage of 10(10) 111In-labeled organisms) and host's ability to kill translocated bacteria (viable bacteria in tissues) were measured followed by burn injury and sacrifice four hours later. Translocation and killing of Staphylococcus epidermidis and Escherichia coli was high in the MLNs compared with all other groups but translocation was lower to the liver. Klebsiella, Pseudomonas, and Serratia translocated more evenly to all the tissues. However, these groups showed very high clearance of bacteria in the liver and spleen except for Klebsiella and one strain of Pseudomonas in the spleen. Candida showed poor translocation to all of the tissues and high clearance. It is concluded that various strains of bacteria translocate from the intestine to a similar degree after injury, but the tissues to which they translocate and the rate at which they are killed are somewhat strain dependent.     

Source of raised serum estrogens in male rats with portal bypass.

We sought to establish the mechanism for the raised serum estrogen levels that occur in male rats with portal hypertension and resultant portal bypass. Using the portal vein ligated (PVL) rat model, we evaluated plasma steroid hormone concentrations, metabolic clearance rate (MCR) of estradiol, and hepatic metabolism of androstenedione to estrogens and other products. In contrast to serum testosterone levels that were reduced, serum androstenedione levels were normal in the PVL rat. Estradiol MCR was measured by a constant intravenous infusion technique and was found to be similar in PVL and control animals. Androstenedione MCR was determined during constant intravenous infusion of [3H]androstenedione, and the resultant radiolabeled steroids present in plasma were separated by thin layer chromatography. The MCR of androstenedione was not diminished in PVL rats compared with controls. However, there was a sevenfold increase in the plasma estradiol derived from [3H]androstenedione in rats with portal bypass. Examination of radiolabel excreted in bile during infusion of [3H]androstenedione showed that 25-46% of this steroid was converted to estradiol in PVL rats compared with less than 3% in control male rats (P less than 0.001). Moreover, there was a selective reduction in the excretion of 16 alpha-hydroxyandrostenedione, a finding which suggested that the metabolism of androstenedione via this pathway was decreased. Androstenedione 16 alpha-hydroxylation is known to be catalyzed by a male-specific cytochrome P-450 isoform, P-450UT-A. We conclude that raised plasma estradiol levels after portal bypass in male rats are due to increased production rates, resulting in turn from enhanced aromatization of androstenedione to estradiol. On the basis of the observed specific changes in androstenedione hydroxylation pathways, it is proposed that alterations in levels of sex-specific forms of cytochrome P-450 occur in male rats with portal bypass and could account for the enhanced formation of estradiol.     

Estimation of urinary unconjugated androstenedione, dehydroepiandrosterone, testosterone, cortisol, aldosterone and 18-hydroxycorticosterone as a potential tool for assessing adrenal status

A method is described for the simultaneous assay of non-conjugated androstenedione, dehydroepiandrosterone, testosterone, cortisol, aldosterone and 18-hydroxycorticosterone in urine. The method involves solid-phase extraction, automatic high performance liquid chromatography and subsequent radioimmunological quantitation of the individual steroids. Excretion rates of these urinary free steroids were determined in normal males and females. There were no significant sex differences in excretion rates, although both urinary free testosterone and dehydroepiandrosterone were distinctly lower in females than in males. Representative measurements of the excretion rates of patients with Cushing's disease, Addison's disease, ectopic corticotropin syndrome and hirsutism were made. The present method has been shown to be well suited for routine purposes. Its final diagnostic significance for monitoring alterations in glucocorticoid, mineralocorticoid and androgenic activity of the adrenal cortex has yet to be explored.     

Infusion of tumor necrosis factor/cachectin promotes muscle catabolism in the rat. A synergistic effect with interleukin 1.

To improve our understanding of the metabolic role of cytokines in protein wasting, we estimated the rates of protein synthesis and degradation in muscle and liver tissues in intact rats treated with several doses of recombinant IL 1 and/or tumor necrosis factor (TNF)/cachectin. Protein breakdown in muscle and liver were derived in vivo from the relationship between [14C]leucine distribution and tissue dilution in reference to circulating leucine. Synthesis was derived from the relationship between [14C]leucine appearance in the protein-bound and free-tissue leucine pools. To specifically relate changes in leucine tracer metabolism to protein dynamics, we separately measured the effect of these cytokines on blood flow to different tissues. The increase in dilution of the tissue-free [14C]leucine by TNF and TNF/IL 1 mixture, but not by IL 1 alone, could not be explained by a hemodynamic effect of these cytokines. Rather, this finding indicated that muscle proteolysis is enhanced by TNF and synergistically augmented by the addition of IL 1. Compatible with these data was the finding that more prolonged infusions of recombinant TNF/cachectin and the combination with IL 1 increased urinary nitrogen excretion. Changes in [14C]leucine dilution in the liver were less pronounced than those in skeletal muscle and consistent with net anabolic effect of TNF on liver protein. We conclude that rats exposed systemically to sublethal doses of TNF respond with increasing muscle and decreasing liver proteolysis, similar to that observed in inflammation and in cancer.     

Metabolism of prostaglandins A1 and E1 in man.

To investigate the in vivo whole blood metabolic clearance rates and sites of metabolism of prostaglandins A1 and E1 in man, constant infusions of the tritiated compounds were administered to normal subjects and to patients undergoing cardiac catheterization. The whole blood metabolic clearance rate of [3H]prostaglandin A1 in eight men was 5,003 +/- 864 liters/day (SD) or 2,546 +/- 513 liters/day per m2 (SD). Nonradioactive prostaglandin A1 was similarly infused in two subjects, and the metabolic clearance rates were determined, utilizing a specific radioimmunoassay. The clearance rates with this method correlated closely with those determined by the isotope infusions. Extraction studies of prostaglandin A1 showed that pulmonary, splanchnic, renal, and extremity perfusions resulted in 8.1 +/- 4.1, 56.1 +/- 10.1, 50.3 +/- 3.4, and 34.4 +/- 5.9% (SEM) removal, respectively. With [3H]=prostaglandin E1, the whole blood metabolic clearance rate was determined from the pulmonary artery concentration in three patients and averaged 4,832 +/- 1,518 liters/day (SD) or 2,686 +/- 654 liters/day per m2 (SD). Pulmonary extraction was 67.8 +/- 6.8% (SEM) and extremity removal averaged 6.6 +/- 4.9% (SEM). These results indicate that A prostaglandins are metabolized by several organs, such as the liver and kidney, and possibly by intravascular pathways as well. In man, the E prostaglandins are primarily metabolized by the lung, but extraction is not complete and approximately one-third may escape lung metabolism. Thus, these findings suggest that both E and A prostaglandins in the venous circulation may reach the systemic circulation in man.     

The Source of Plasma Dihydrotestosterone in Man

The source of plasma dihydrotestosterone (DHT) (17beta-hydroxy-5alpha-androstan-3-one) in humans has been investigated by infusing two potential peripheral precursors, testosterone (T) and androstenedione (A). Metabolic clearance rates (MCR), conversion ratios (CR), transfer constants (rho), and blood production rates (P(B)) were calculated. Plasma testosterone and dihydrotestosterone were measured by competitive binding techniques. The MCR(DHT) was 652 +/-35 (SD) liters/day in five males and 314 +/-63 (SD) liters/day in four adult females. In each individual, the MCR(DHT) was significantly lower than MCR(T) as predicted by testosterone-binding protein affinity studies. The P(B) (DHT) was 302 +/-65 (SD) mug/day in males and 56 +/-26 mug/day in females. Testosterone and androstenedione are precursors (prehormones) for plasma dihydrotestosterone. The conversion ratio CR(BB) (T-DHT), calculated as the ratio of counts per minute per liter of plasma of product to precursor after infusion of labeled precursor, was 5.6 +/-0.6 (SD)% (six subjects) in the male and 3.5 +/-0.4 (SD)% (four subjects) in the female. CR(BB) (A-DHT) after androstenedione infusion to three female subjects averaged 9.2%. No dihydrotestosterone back conversion was detected (< 0.2%). The transfer constants were [rho]BB(T-DHT), 3.9 +/-1.0% (male) and 1.7 +/-0.6% (female), and [rho]BB(A-DHT) average was 13.3% in three female subjects. Using either plasma testosterone and dihydrotestosterone values from our subjects and mean androstenedione values as reported in the literature, approximate contributions can be calculated. Testosterone conversion accounts for at least 70% of plasma DHT in the male, but less than 20% in the normal female. Androstenedione appears to be a major prehormone of plasma dihydrotestosterone accounting for at least two-thirds plasma dihydrotestosterone by peripheral conversion in adult females. In three normal women undergoing tubal ligation, there was an unimpressive gradient between ovarian vein and peripheral plasma dihydrotestosterone. It is suggested that dihydrotestosterone in the blood does not arise from direct secretion but may reflect events occurring in peripheral androgen target tissues.     

The effect of storage and temperature on the analysis of steroids in plasma and blood

In the analysis of steroid hormones careful attention is usually paid to blood collection and plasma storage. However, the appropriate care of samples cannot always be assured in routine work with steroids. Therefore, the stability of cortisol, aldosterone, 17-hydroxyprogesterone, testosterone, androstenedione, dehydroepiandrosteronesulphate, oestrone, oestradiol, sex hormone binding globulin (SHBG) and, the binding of testosterone and cortisol to plasma proteins in blood and plasma were studied before and after various handling procedures. Ten cycles of alternate freezing and thawing of plasma did not significantly affect the levels of the steroids or their plasma binding. The greatest differences, compared with controls, were seen for aldosterone (-6.2%) and oestradiol (-5.3%). Plasma storage at -28 degrees C was hardly superior to a 4 days storage at 4 degrees C (refrigerator) or 22 degrees C (room temperature). Although androstenedione (-10.9%), oestrone (-10.2%) and oestradiol (-12.2%) levels decreased by more than 10%, the means of all analyses were still in the 2 SD range. Even SHBG and the steroid binding were only slightly affected by temperature. When whole blood was stored at 4 degrees C or 22 degrees C, the resulting values differed from those obtained with plasma, but the differences were usually less than 10%. Although the levels were within the 2 SD range, whole blood showed a decrease of 12.3% for aldosterone and 14.5% for androstenedione. In contrast, plasma binding of testosterone (25.9%) and cortisol (15.1%) were substantially affected by storage at 22 degrees C in whole blood. It is concluded that repeated freezing and thawing of plasma, or storage at various temperatures have only a small effect on the measured levels of steroids and their plasma binding. Although it is not advisable, even whole blood may be used for the analysis of steroid concentrations.     

A method for the assay of testosterone and androstenedione in human female plasma by competitive protein binding

A competitive protein-binding method for the simultaneous determination of testosterone and androstenedione in human female plasma is described. After extraction from plasma, test osterone and androstenedione are separated by paper chromatography. Androstenedione is reduced with sodium borohydride and the testosterone generated is separated from other reduction products by paper chromatography. The binding property of the testosterone-binding globulin, which has a high binding affinity for steroids with an unhindered 17β-hydroxyl group, is used as an assay end-point. The reliability criteria of the method in terms of precision, accuracy, sensitivity and specificity have been evaluated. The mean level of testosterone and androstenedione in random plasma samples from normal females in the 15–50 years age group is 33.5 ± 9.4 (SD) ng/100 ml and 105 ±35.6 ng/100 ml respectively.