Estrogen-induced cytodifferentiation of the ovalbumin-secreting glands of the chick oviduct.

The histological, ultrastructural, and biochemical changes occurring during hormone-induced cytodifferentiation of the ovalbumin-secreting glands in the chick oviduct have been studied. Marked perivascular edema is an initial response of the immature oviduct stroma to diethylstilbestrol administration and is accompanied by an interstitial migration of mononuclear cells. Mitotic activity in the immature mucosal epithelium increases within 24 hr, and glands begin to develop on days 2-4 as budlike invaginations into the subepithelial stroma. An immediate intracellular effect of the hormone is aggregation of previously dispersed ribosomes. Ribosomal zones in the nucleolus gain prominence, and there is a progressive development of rough endoplasmic reticulum in the epithelial cells. Extensive profiles of endoplasmic reticulum are present in the gland cells by day 6. Fine apical progranules appear in the epithelial cells on day 2, and ovalbumin can be measured immunochemically by day 3 at about the same time that new species of nuclear RNA have been identified. Ovalbumin granules form within condensing vacuoles in the Golgi zone and begin to be released into the lumina of the gland acini at about day 6 of the treatment.

Characterization of the androgen receptor from a Syrian hamster ductus deferens tumor cell line (DDT1).

The hamster ductus deferens cloned tumor cell line (DDT1) contains a complex steroid receptor protein that binds 3H-labeled 5 alpha-dihydrotestosterone,. Diethylaminoethyl (DEAE) chromatography of cytosol from these cells yields two major receptor peaks of activity. Identification of this steroid binding protein as a cytoplasmic receptor was confirmed by salt dissociation on sucrose gradients, stability of the hormone-receptor complex at 0 degrees C, and the retention patterns on phosphocellulose and DEAE cellulose. Multiple forms of the receptor exist in a single homogeneous cell type. The data support the theory that steroid hormones bind to a cytoplasmic protein receptor composed of dissimilar subunits as the initial step in steroid hormone action.

Protein synthesis: differential stimulation of cell-specific proteins in epithelial cells of chick oviduct.

Immunofluorescent and radioautographic studies demonstrate that ovalbumin and avidin are cell-specific proteins synthesized by different epithelial cells in the chick oviduct mucosa. The mechanism of the selective induction of ovalbumin synthesis by estrogen and of avidin synthesis by progesterone may be through stimulation of specific target cells by these hormones.

Cell culture on artificial capillaries: an approach to tissue growth in vitro.

Artificial capillaries perfused with culture medium provide a matrix in which cells can attain tissue-like densities in vitro. Products secreted into the medium can be measured as indicators of cell function or may be recovered for other purposes without disturbing the culture.

Cortisol induction of growth hormone synthesis in a clonal line of rat pituitary tumor cells in culture.

Growth hormone synthesis increased markedly after addition of glucocorticoids at physiological concentrations to cultures of the GH(1) line of rat pituitary tumor cells. Stimulation of hormone protein synthesis by corticosteroids was selective, since (i) the rate of hormone synthesis increased while total protein synthesis decreased, and (ii) cortisol analogs, biologically inactive metabolites, and sex steroids did not induce growth hormone synthesis.

Metabolic clearance and production rates of human luteinizing hormone in pre- and postmenopausal women.

Metabolic clearance rates and production rates of human luteinizing hormone (HLH) were determined in pre- and postmenopausal women by the constant infusion technique. Highly purified HLH-(131)I was infused into the fasting subjects at a constant rate. Serial plasma samples were obtained and the radioactive hormone was precipitated by a double antibody technique. Plasma HLH-(131)I levels reached equilibrium by 4 hr after the infusion started. Metabolic clearance rates were: 24.4 +/- 1.8 (mean +/- SE) ml/min in five normal premenopausal women; 23.3 +/- 1.1 ml/min in five normal women taking norethinodrel and mestranol; and 25.6 +/- 4.1 ml/min in four postmenopausal women. Endogenous plasma HLH levels measured in the same subjects by radioimmunoassay immediately before infusion were 32.0 +/- 9.6 mU/ml in the normal women, 16.8 +/- 3.2 mU/ml in the women on oral contraceptives, and 99.2 +/- 23.2 mU/ml in the postmenopausal women. The corresponding HLH production rates were: 734 +/- 170 mU/min in the normal women: 387 +/- 86 mU/min in the women on norethinodrel and mestranol; and 2400 +/- 410 mU/min in the postmenopausal women. The metabolic clearance rate did not change after ovariectomy in one women, but the production rate rose from 583 to 1420 mU/min. Based on previously reported bioassay values for pituitary content and urinary excretion of HLH, the estimated turnover of HLH in the pituitary is about once per day and less than 5% of the total HLH produced appears in the urine in a biologically active form.

Translation of ovalbumin mRNA in Xenopus laevis oocytes. Characterization of the system and effects of estrogen on injected mRNA populations.

Ovalbumin messenger RNA (mRNAov) purified from hen oviduct was injected into Xenopus laevis oocytes. The oocytes were incubated in culture medium containing [3H]leucine. Analysis of the oocyte cytosol on Sephadex G-15O columns demonstrated a peak of radioactivity which cochromatographed with authentic ovalbumin. Radioactive protein contained in this peak was precipitated by ovalbumin antiserum, coelectrophoresed with ovalbumin on sodium dodecyl sulfate (SDS) and urea gels at pH 8.7, and eluted with the protein at the same pH (4.8) on CM-cellulose chromatography. Injection of increasing amounts of mRNAov was found to elicit a linear response in terms of ovalbumin synthesis. Moreover, there was linear incorporation of radioactivity into microinjected oocytes over a minimum period of 91 h. Less than 1 ng mRNAov was detected in this system. Ovalbumin mRNA activity was present in RNA preparations from chicks treated with estrogen but was undetectable in animals withdrawn from the hormone. This study constitutes an initial demonstration of a steroid hormone-induced alteration in mRNA population as assayed in intact viable heterologous cells.

Production rates and metabolic clearance rates of human follicle-stimulating hormone in premenopausal and postmenopausal women.

The production rates (PR) and the metabolic clearance rates (MCR) of human follicle-stimulating hormone (HFSH) were determined in six pre- and five postmenopausal women. Human FSH (PER-780) labeled with (131)I to specific activities of 50-150 muc/mug was used as a tracer. Both double antibody and trichloroacetic acid (TCA) precipitation techniques were used to determine HFSH-(131)I levels in infusate and plasma. In four of the subjects MCRs measured by both constant infusion and single injection techniques were the same. By constant infusion, plasma HFSH-(131)I levels reached equilibrium between 4-5 hr.MCRs in six premenopausal women were 14.2+/-1.1 (mean +/-SE) ml/min. MCRs in five postmenopausal women were 12.6 +/-1.1 ml/min. Simultaneous HFSH and human luteinizing hormone (HLH) MCRs were determined in a single patient using HFSH-(125)I and HLH-(131)I as tracers by both constant infusion and single injection methods. These studies showed that the MCR of HFSH was 10.8-11.1 ml/min, and the MCR of HLH was 18.5-19.4 ml/min. From these data and previous MCR and PR studies of HLH from this laboratory, it appears that the MCR of HFSH is about one-half that of HLH. Endogenous HFSH and HLH levels were measured by radioimmunoassay. The PRs of HFSH, calculated by the product of endogenous level and MCR, were 146 +/-27mU/min in the premenopausal women and 2141 +/-264 mM/min in the postmenopausal women. 24-hr PRs, based on these results, compared with reports of 24-hr urinary excretions of biologically active HFSH indicate that 3-5% of production is found in urine in biologically active form. After our single injections of HFSH-(131)I, 8-29% was recovered in urine over 24 hr.

Ectopic and eutopic secretion of chorionic gonadotropin and its sub-nits in vitro: comparison of clonal strains from carcinomas of lung and placenta.

We compared rates of secretion in vitro of human chorionic gonadotropin (HCG) and its subunits alpha and beta by established clonal cell lines of a bronchogenic carcinoma (ChaGo) and a choriocarcinoma (JEG). Clones showing the highest secretion rates of either HCG or its subunits were studied: ChaGo-K1, a new clonal strain, and ChaGo-C5 and JEG-3, two previously reported clonal lines. Cells were grown under identical conditions in the same laboratory. Hormone and subunit concentrations were measured by radioimmunoassays. ChaGo-K1 and ChaGo-C5 secreted only alpha-subunit whereas JEG-3 secreted only HCG. Average peak secretion rates in picomoles/day/mg protein were: for ChaGo-K1, HCG less than 0.3, alpha=290, and beta less than 0.5; for ChaGo-C5, HCG less than 0.3, alpha=21, and beta less than 0.5; and for JEG-3, HCG=18, alpha less than 0.7, and beta less than 0.5. The ChaGo-K1 secretion rate of alpha was greater than that of any of out previously reported ChaGo clones. Significant quantities of estradiol and progesterone accumulated in the media of all three cell lines; however, only JEG-3 secreted detectable quantities of placental lactogen. Thus under identical culture conditions, a bronchogenic carcinoma clonal line secreted only alpha-subunit, whereas a choriocarcinoma line secreted only HCG; these findings implied major differences in cellular control mechanisms. Moreover, the ectopic secretions of alpha exceeded the eutopic trophoblastic secretion of HCG, which suggested that in certain cases ectopic protein production may be even more efficient than nonectopic production.

A human trophoblastic isozyme (lactate dehydrogenase-Z) associated with choriocarcinoma.

A unique electrophoretic form of lactate dehydrogenase (LDH-Z), formerly observed in a choriocarcinoma cell line (JEG-3) and first-trimester placenta, has been shown to be the same as that produced in hydatidiform mole and term placenta. We have also observed LDH-Z in second-trimester placenta, choriocarcinoma metastasized to the liver, and five of five additional independently derived (from different patients) choriocarcinoma cell lines. The only exception to the production of LDH-Z in a choriocarcinoma was in the cell line BEWO, which was established from the same tumor as JEG-3. Since BEWO has been in culture for over 200 passages more than any of the independently derived lines, its lack of LDH-Z is viewed as being consistent with the loss of certain expressional characteristics upon such long-term culture. Analysis of JEG-3 subclones revealed the expression of LDH-Z to be independent of the products of the LDH-A genetic locus. Present data do not allow us to determine whether LDH-Z is the product of a newly discovered LDH locus or is a modified form of the product of the LDH-B locus. LDH-Z has not been observed in other human tissues, nor have we observed it in homogenates prepared from over 60 cell lines established from a wide variety of human neoplasms. We therefore conclude that LDH-Z is an isozyme associated with human choriocarcinoma and is indicative of the trophoblastic origin of the cells.

Effects of guanadrel on patients with thyrotoxicosis.

Eleven patients with Graves' disease were treated with guanadrel sulfate and observed for changes in neuromuscular and cardiovascular manifestations. No notable changes in pulse rate or muscle strength were detected in either these patients during a three-day pretreatment period or in five control patients with Graves' disease receiving placebo for six days. Thyroid hormone levels were not altered by seven days of guanadrel sulfate therapy (5 to 20 mg orally every six hours), and no adverse side effects were encountered. Mean supine resting pulse fell from 102 +/- 6 (mean +/- SEM) to 90 +/- 3 beats per minute (P less than .02). The patients' proximal and distal muscle strengths were initially decreased, when compared with healthy subjects, and improved substantially with guanadrel therapy. We conclude that guanadrel sulfate may be useful in the symptomatic management of patients with thyrotoxicosis.

Pituitary apoplexy following chlorpromazine stimulation.

Chlorpromazine is frequently administered to patients with hyperprolactinemia to stimulate an increase in the serum levels of prolactin. A patient with a prolactin secreting adenoma is described in whom pituitary apoplexy developed in association with a hypotensive episode following the administration of 25 mg of chlorpromazine. Prolactin levels fell from more than 2,000 ng/ml to 340 ng/ml following infarction of the pituitary tumor. Pituitary apoplexy should be considered as a rare complication of chlorpromazine stimulation in a patient with a pituitary tumor.

The coexistence of androgen and glucocorticoid receptors in the DDT1 cloned cell line.

The hamster ductus deferens cloned tumor cell line (DDT1) has been shown to contain both androgen and glucocorticoid binding activity. The androgen receptor binding site concentration is 1.07 x 10(-13) mol of testosterone/mg protein, and testosterone (T) binds with a Kd of 4.3 x 10(-10) M. Dihydrotestosterone (DHT) is also bound to the receptor with a Kd of 2.99 x 10(-10) M and the binding site concentration is 1.33 x 10(-13) mol/mg protein. The order of steroid binding affinity is DHT greater than T greater than Estradiol greater than Progesterone. Cortisol, dexamethasone, and triamcinolone acetonide do not inhibit the androgen binding in vivo or in vitro. In a cell free system antiandrogens inhibit the binding of DHT. The DDT1 cells have a separate receptor for cortisol which binds at saturation 3.44 x 10(-13) mol cortisol/mg protein and has a Kd of 4.54 x 10(-9) M. These studies provide evidence that these endocrine target cells contain specific high affinity receptors for more than one type of steroid. The glucocorticoid receptor may be important for maintaining essential undifferentiated functions while the DHT receptor gives the specific characteristics of sex hormone responsive tissues.

Investigation of choriocarcinoma clonal cell lines in vitro and choriocarcinoma transplants in the hamster for the secretion of a thyroid-stimulating factor.

Six choriocarcinoma cell lines that secrete chorionic gonadotropin (hCG) in tissue culture were investigated for the production of thyroid stimulators. In the mouse thyrotropin bioassay, no thyroid-stimulating activity was found in cell culture media even after 30-fold concentration by the method of Bates. Further, concentrates of culture media did not react in a porcine thyrotropin radioimmunoassay in which hCT cross-reacted completely. Thyroid function was also assessed in hamsters bearing hCG-secreting human choriocarinoma transplants. Even though the thyroid-stimulating factor present in commercial urinary hCG preparations caused an increase in PB131I in the hamster, the presence of the choriocarcinoma had no effect on PB131I. From these results, it appears that human choriocarcinoma cell lines maintained in tissue culture and human choriocarcinoma serially transplanted in the hamster are not suitable models for the study of the secretion of a thyroid-stimulating factor by trophoblastic tissues.

The potential for an androgen male contraceptive.

PIP: 20 healthy men, 23-34 years of age, were selected to study the potential for using an exogenous androgen as a reversible male contraceptive. The experimental design consisted of a 10-week control period, an initial 12-week period with weekly 200 mg injections of testosterone enanthate (TE), a 36-week period with 200 mg injections of TE every 3 weeks, and a 28-week recovery period. Serum concentrations of testosterone were increased about 50% during the initial treatment period, and gonadotropins became very low or undetectable and essentially unresponsive to LRH. Sperm density fell, and 11 of the 19 men evaluated at the end of the weekly injection period had less that 1 million sperm/ml semen. Serum follicle stimulating hormone (FSH) and luteinizing hormone (LH) and sperm density rose when TE was given every 3 weeks, and on this regimen only 1 man consistently had suppressed gonadotropins and sperm counts of less than 1 million/ml. Each man's sperm count during the recovery period exceeded 25 million/ml and 15 of the 17 men had average values which were similiar to or exceeded their average control values. There were no serious side effects during the study period. The mean concentration of hemoglobin increased from 15.1 + or - 0.2 g/dl to about 16.0 g/dl and the serum glutamic oxalopyruvic transaminase increased from 44 + or - 4.5 to more than 60 centaunits during treatment. Serum concentration of estradiol increased from 34.1 + or - 2.6 to 59.4 + or - 6.2 pg/ml when testosterone levels were increased. There were no significant changes in carbohydrate tolerance, cholesterol, triglyceride, or clotting studies. The regimen failed to uniformly achieve azoospermia or ligospermia of sufficient degree to assure its effectiveness as a contraceptive agent.^ieng

Somatomedin C in treated acromegaly: poor correlation with growth hormone and clinical response.

To determine the usefulness of commercially available somatomedin C levels in the evaluation of the treatment of acromegaly, 15 patients were tested at 0.25-15.4 yr after onset of therapy. Clinical response, as determined by a numerical scoring system, was compared with RIA of GH and somatomedin C. Symptomatic response was poorly correlated with somatomedin C (r = 0.033) as well as with GH (r = 0.24). The correlation of GH and somatomedin C was also poor (r = 0.46, P greater than 0.05). Eighty-three percent of patients with clinical improvement had GH less than or equal to 10 ng/ml, 50% had GH less than or equal to 5 ng/ml, while 42% had somatomedin C less than or equal to 3.0 U/ml. All patients who were evaluated at 1 yr or less after therapy had elevated somatomedin C levels with normal or near normal GH values. In contrast only 2 of 11 patients evaluated at more than 1 yr after therapy had a mild persistence of somatomedin C elevation with normal GH levels. Determination of somatomedin c costs more than GH determinations and appears to offer no apparent advantage over GH in following patients treated for acromegaly.

Giant invasive prolactinomas.

Two of the largest prolactinomas ever documented that have been followed for nine and 10 years, respectively, demonstrate how aggressive prolactinomas may become and how difficult invasive prolactinomas are to treat. One of these prolactinomas invaded both internal auditory canals and simultaneously grew inferiorly, reducing the bony support of the skull and necessitating the patient to utilize both hands to hold his head up. The second patient's prolactinoma invaded the sphenoidal, ethmoidal, and cavernous sinuses. Both of these patients had neurosurgical debulking of their tumors followed by radiation therapy. Neither patient's prolactin levels decreased significantly during their first five years post-surgically, at which time bromocriptine was added. Since then, there has been a gradual lowering of serum prolactin levels and a decrease in the size of these tumors. These cases demonstrate that prolonged treatment and very large doses of bromocriptine may be necessary for tumor reduction in patients with invasive prolactinomas.

Hyperalimentation as cause of markedly worsened metabolic control in a patient with autoantibodies to the insulin receptor.

Patients with type B insulin resistance and acanthosis nigricans have autoantibodies to their insulin receptors and usually have signs and symptoms of other autoimmune diseases. The first case demonstrating that hyperalimentation markedly disturbs blood glucose control in type B insulin-resistant patients is described. Neither prednisone, insulin (up to 240 units per hour), nor tolbutamide appeared to help this patient's metabolic control. After the addition of cyclophosphamide for one week, the anti-insulin receptor autoantibody titer dropped from greater than 1:1,000 to 1:1. Six months later, the patient had a complete remission, which is rare, with only three other reported remissions in these patients with type B insulin resistance.

Recent advances in the control and function of the anterior pituitary.

Major advances in our understanding of the synthesis and release of anterior pituitary hormones have been made over the past several years. Neurons of the hypothalamus have been found to serve as "neuroendocrine transducers" in that they have both electrical and secretory functions. Peptidergic neurons respond to appropriate stimuli with a release of hypothalamic factors into the hypophyseal-portal system. These factors or hormones ultimately control the endocrine function of anterior pituitary cells. Three hormones, Thyrotropin Releasing Hormone (TRH), Gonadotropin Releasing Hormone (GnRH or LHRH) and somatostatin have been identified, synthesized and tested for clinical applications. The clinical assessment of pituitary function has been greatly improved by new and improved radioimmunoassays. One of the recent clinical advances in the area of pituitary disease has been the determination of the relatively high frequency of prolactinomas. Prolactin secreting microadenomas are an important and treatable cause of amenorrhea and infertility in young women. In addition, many pituitary tumors previously believed to be non-functional or "chromophobe adenomas" appear to be prolactinomas. Many new diagnostic and therapeutic techniques are continuing to be developed to improve our management of patients with hypothalamic-pituitary disease.

Changing structure to improve function: one academic health center's experience.

Academic health centers (AHCs) have been under siege for the past few years, with decreased federal and state funding for educational and research programs and increasing competition in the health care marketplace. In addition, many AHCs are burdened with the bureaucratic red tape of large educational institutions, which makes agility in responding to a demanding health care market difficult. The authors describe the response to these threats by Oregon Health Sciences University (OHSU), an approach that has been different from those of most similar institutions. OHSU chose to change its structure from being part of the state system of higher education to being an independent public corporation. The authors outline the political process of building widespread support for the legislation passed in 1995, the key features of the restructuring, the challenges faced before and after the transition to a public corporation, and lessons learned in this metamorphosis to a new form.