Alfred M. Bongiovanni

New solvent systems for the resolution of corticosteroids by paper chromatography.

Abstract A simplified procedure is described which permits employment of the Bush method for the separation of C-21 steroids on paper chromatograms at room temperature. Several new solvent systems have been devised, in which tert-butanol or a combination of tert-butanol and methanol constitutes the stationary phase. The new systems, used in combination with modified Bush systems, permit increased resolution of various steroid mixtures and increased structural characterization of unknown steroids, on the basis of their changing chromatographic behavior.

Ovarian tumors and cysts associated with sexual precocity. Report of 3 cases and review of the literature.

Summary Two cases of granulosa cell tumor and one of theca-lutein cyst of the ovary associated with sexual precocity are reported, which demonstrate the difficulty in distinguishing pseudoprecocity from true precocity in the female by the three criteria usually employed: the presence of a mass palpable bimanually, the excretion of large amounts of estrogens, and the absence of urinary gonadotropins when the precocity is produced by a granulosa cell tumor. One of the tumors completely lacked theca cells but contained abundant crystals usually thought to represent the source of estrogens. This finding casts doubt on the belief that the theca cells alone are responsible for estrogen production in ovarian tumors. A review of the literature reveals that granulosa cell tumors in children are less benign than is commonly believed. An analysis of the published cases of ovarian cysts associated with precocity suggests that not all are variants of true sexual precocity. The theca-lutein cyst reported is a possible example of an autonomous functional cyst, resembling five or six others described in the past.

INDUCED GENITAL ANOMALIES

The human female fetus has been reported to have varying degrees of virilization of the lower urogenital tract after maternal treatment with androgenic hormones with certain progestins, and rarely with estrogens. Paradoxically, three cases of feminization of male fetuses with varying degrees of hypospadias have been reported to occur after maternal treatment with large doses of either diethyl‐stilbestrol and progesterone, of progesterone alone, or of acetoxyprogesterone (Prodox). An analysis of the human reports and comparable experimental data suggest that the fetal effects of these agents are not analogous to their effects on the end organs in the postnatal animal. Rather, they appear to have some common dose‐dependent teratogenic action during critical periods of genital development in the two sexes. Genetic male infants born with a severe form of congenital adrenal hyperplasia due to a genetic deficiency in activity of 3β‐ hydroxysteroid dehydrogenase (3β‐enzyme) also have incomplete masculine development and hypospadias. Genetic females with this disease are born with a moderate degree of virilization, clitoral hypertrophy, and labial fusion. The 3β‐enzyme is essential in the early biosynthetic pathway of every biologically active steroid hormone.

Gonadotropin responses to luteinizing releasing factor in boys treated with cyclophosphamide for nephrotic syndrome

Cyclophosphamide therapy of the nephrotic syndrome has been associated with oligo- and azoospermia and with abnormalities of testicular histology in adults and pubertal boys. In 15 prepubertal boys, no abnormalities of basal serum levels of LH, FSH, or T were found when they were studied 8 months to 7 years after cyclophosphamide therapy. Five boys were pubertal during therapy were found to have elevated mean basal values of gonadotropins with normal testosterone levels and elevated LH responses to LRF; the FSH responses to LRF were elevated in four patients. One of four boys who were prepubertal during therapy but pubertal at the time of testing had an elevated basal LH and LH response to LRF. Three boys who were prepubertal at the times of therapy and testing had normal LH responses to LRF. The LRF test may provide a means of identifying the patient who has sustained testicular injury and who may require testicular biopsy.

Familial glucocorticoid insufficiency

A familial syndrome manifested by glucocorticoid insufficiency but with normal mineralocorticoid activity has been attributed to an inherited defect in the adrenocorticotrophic hormone-dependent system of the adrenal gland. 1 Five siblings with this syndrome have been studied. Two of these siblings had increased urinary excretion of 17-hydroxycorticosteroids in response to ACTH stimulation at an early age. Both children subsequently developed the clinical and biochemical changes of the syndrome. An infusion of ACTH to two children in this family caused an increase in urinary excretion of aldosterone. These findings suggest that the pathogenesis of the syndrome in this family is due to an inherited degenerative process and not to primary unresponsiveness to ACTH.

Determination, recovery, identification and renal clearance of conjugated adrenal corticoids in human peripheral blood.

Summary 1. An alternative method for the extraction and quantitation of free and conjugated corticosteroids in human plasma has been described. 2. With the use of the relatively pure monoglucuronide of tetrahy-dro-E, it has been possible to show that the recovery of conjugated corticosteroids is satisfactory within a broad range of concentrations. 3. Following the administration of large oral doses of free tetrahydro-E, very high levels of conjugated corticosteroids, in excess of 3 mg/100 ml of plasma, have resulted. The material has been definitively identified as tetrahydro-E. A second component, possibly the 20-reduced derivative has been detected. 4. The renal clearances of the free and conjugated steroids in human plasma have been determined.

Production of Congenital Adrenal Cortical Hyperplasia, Hypospadias, And Clitoral Hypertrophy (Adrenogenital Syndrome) in Rats by Inactivation of 3β-Hydroxysteroid Dehydrogenase.∗

Summary Varying daily doses of a steroidal inhibitor of 3β-hydroxysteroid dehydrogenase (2α-cyano-4,4,17α-trimethylandrost-5-en-17β-ol-3-one), were administered to pregnant rats on the 15 th to 20th days of gestation to determine whether this inhibitor could produce the biological manifestations of the human disease associated with deficient activity of this enzyme. A single dose of the inhibitor was also administered to pregnant rats on various days to determine the critical periods for production of these manifestations. Infants with this disorder have hyperplastic adrenal cortices deficient in activity of the 3β-enzyme, incomplete masculine development in males resulting in hypospadias with shortening of the anogenital distance, and clitoral hypertrophy without urethral orificial displacement in females. The experimental fetuses had adrenal cortical hyperplasia (average of 5.9 ± 1.32 mg against 2.2 ± 0.26 mg controls), deficient histochemical activity of the 3β-enzyme in the adrenals and testes, increased activity of glucose-6-phosphate dehydrogenase in adrenals, testes, and liver and no change in histochemical activities of 3α- or 17β-enzymes in these tissues. The anogenital distance in gonadal males was reduced from the normal of 2.88 ± .12 mm to 1.26 ± .08 mm in proportion to dose. The gonadal females had clitoral hypertrophy but no change in anogenital distance from the normal of 1 mm. The optimal period for production of hypospadias by the inhibitor corresponds to the early appearance of strong activity of the 3β-enzyme in the fetal testis. The optimal day for the production of adrenal hyperplasia and of clitoral hypertrophy corresponds to the time of maximal adrenal activity of the enzyme. This study thus demonstrates an experimental model for production of the congenital manifestations of this inborn error of metabolism.

Urinary vanilmandelic acid (VMA) excretion in children: use of a simple semiquantitative test.

A simplified colorimetric method is described for use in detecting excessive excretion of urinary VMA in children with suspected neurogenic tumors. Elevated levels are demonstrated in patients with neuroblastomas (13), ganglioneuroblastomas (3), ganglioneuromas (7), a pheochromocytoma (1), and a chemodectoma (1). Other patients studied included those with chronic diarrhea (15), hypertension (7), various tumors (10), cystic fibrosis of the pancreas (12), hypoxia (10), familial dysautonomia (2), and glycogen storage disease (1). In this miscellaneous group values were usually within the normal range with the exception of a few cases which are discussed in more detail.

Treatment of juvenile thyrotoxicosis

Individuality of treatment and a certain latitude in choice of therapy in the management of juvenile thyrotoxicosis are advocated. Eighteen such patients, illustrative of this approach, are reviewed. Remission occurred in seven during medical therapy (thiouracil derivatives). Eight underwent subtotal thyroidectomy; 2 because of relapse after adequate periods of treatment, 3 because of toxic reactions to the drugs, with 2 there was inability to adhere rigidly to the treatment regimen, and in 1 instance because idiopathic neutropenia precluded trial of antithyroid medication. Inflexibility in the management of juvenile thyrotoxicosis is countered by compromise proposals which include initial use of antithyroid drugs and the later choice of definitive therapy based upon individual circumstances.

The measurement of plasma Δ4-androstenedione by competitive protein binding

Abstract A sensitive competitive protein binding (CPB) method capable of measuring Δ 4 -androstenedione (Δ 4 ) in 3 ml of plasma is presented. This method utilizes third trimester pregnancy plasma as a source of testosterone binding globulin and the conversion of Δ 4 to testosterone (T) by potassium borohydride. Dextran coated charcoal is used to separate the free steroid from the bound. Analysis of twenty standard curves in the range of 0–1 ng T provided coefficients of variation from 5.9% at 0.2 ng T to 7.9% at 1.0 ng T. The mean recovery of tritiated Δ 4 added to plasma as a tracer was 31.6%. Normal values for adult males and females as determined by this CPB method fall within the range of published values as determined by double isotope derivative assays (DIA) and gas liquid chromatography (GLC). Compared to the DIA and GLC methods, the blank of this method is higher and the measurement of replicate samples is slightly less precise.