["Follow up" strategy in the management of hospital stay]. / Estrategia de Follow up en la gestión de la estancia hospitalaria.

Background: In countries with emerging economies, the adequate and efficient management of resources is a priority, through strategies to reduce prolonged stay, increase the availability of beds, maximize profitability and reduce iatrogenic complications. Objective: The purpose of the study was to evaluate the effect of the "Follow up" strategy (FU) on the main indicators of the hospitalization process. Material and methods: A cross-sectional, comparative study was developed to evaluate the impact of the FU strategy on the indicators: hospital admissions and discharges, average days of hospital stay (DEH), percentage of hospital occupancy (OH), bed substitution interval (ISC), bed turnover rate (CRI) and prolonged hospital stay (EHP). Results: The FU was associated with a reduction in DEH [5.7 (5.5-6.1) vs. 6.5 days (6.1-6.9), p = 0.01]; ISC [0.6 (0.4-0.8) vs. 1.2 (0.8-1.3), p = 0.01] and EHP [23.6 (21.6-24.7) vs. 26.3% (24.4-28.7), p = 0.02] compared to the control group, with an increase in existence [1436 (1381-1472) vs. 1347 patient days (1280-1402), p = 0.02], respectively. There was no significant difference in the number of admissions, discharges or in the IRC. Conclusions: The FU reduces the average number of days of hospital stay, the rate of bed substitution and prolonged stay.

Introducción: en los países con economías emergentes es prioritaria la gestión adecuada y eficiente de los recursos hospitalarios. Las estrategias de gestión pueden reducir la estancia prolongada, aumentar la disponibilidad de camas, maximizar la rentabilidad y reducir las complicaciones iatrogénicas. Objetivo: el propósito del estudio fue evaluar el efecto de la estrategia de Follow up (FU) en los principales indicadores del proceso de hospitalización. Material y métodos: se desarrolló un estudio transversal, comparativo, para evaluar el impacto de la estrategia de FU en los indicadores: ingresos y egresos hospitalarios, promedio de días de estancia hospitalaria (DEH), porcentaje de ocupación hospitalaria (OH), intervalo de sustitución de camas (ISC), índice de rotación de camas (IRC) y estancia hospitalaria prolongada (EHP). Resultados: la estrategia de FU se asoció con una reducción de los DEH [5.7 (5.5-6.1) frente a 6.5 días (6.1-6.9), p = 0.01]; ISC [0.6 (0.4-0.8) frente a 1.2 (0.8-1.3), p = 0.01] y EHP [23.6 (21.6-24.7) frente a 26.3% (24.4-28.7), p = 0.02] respecto al grupo control, con incremento de la existencia [1436 (1381-1472) frente a 1347 días paciente (1280-1402), p = 0.02], respectivamente. No hubo diferencia significativa en el número de ingresos, egresos ni en el IRC. Conclusiones: la estrategia de FU disminuyó el promedio de días de estancia hospitalaria, el índice de sustitución de camas y la estancia prolongada.

50 Aniversario de la Escuela de Biología

Editorial. 50 Aniversario de la Escuela de Biología La historia de la Escuela de Biología de la Universidad de San Carlos de Guatemala inicia con la inspiración de un hombre visionario, quien desde el año 1968, siendo director del departamento de Biología de la Facultad de Ciencias Químicas y Farmacia, elaboró una propuesta para su creación. En su momento, esta no fue aprobada, por razones de presupuesto, administrativas y de otra índole, pero este visionario no dio marcha atrás en su lucha por sentar las bases para una formación sólida en el campo de la Biología, tan necesaria en nuestro país.

Infección por virus de Epstein-Barr y hepatitis aguda colestásica / Infection with Epstein-Barr virus and acute cholestatic hepatitis

El Virus de Epstein-Barr (VEB) es un herpes virus cuyo medio de transmisión es a través de secreciones de una persona portadora del virus, siendo el hombre el único huésped. La primo infección por lo general es asintomática o puede manifestarse como mononucleosis infecciosa con la triada clásica de fiebre, faringitis y adenopatías. Esta cursa con elevación leve y autolimitada de transaminasas, por lo que solo un 5% de los casos se ha asociado con hepatitis aguda colestásica. Presentamos a un paciente con una infección por virus de Epstein-Barr y hepatitis aguda colestásica con historia de aparición de una masa cervical lateral derecha. Al examen físico evidencia ictericia a nivel de escleras, mucosas y ambos miembros superiores. Niveles de bilirrubina en sangre elevados. Paciente con ultrasonido hepático y vías biliares normal, colangiopancreatografía retrograda endoscópica normal por lo que se procede a realizar pruebas serológicas para VEB siendo esta positiva. Se da tratamiento con ganciclovir, mejorando pruebas de función hepática y disminuyendo ictericia, teniendo así una evolución favorable del paciente...(AU)

Epstein-Barr Virus (EBV) is a herpes virus, whose means of transmission is through secretions of a person carrying the virus, the man being the only host. The cousin infection is usually asymptomatic or may manifest as infectious mononucleosis with the classical triad of fever, pharyngitis and lymphadenopathy. This is a mild and self-limiting elevation of transaminases, which means that only 5% of the cases have been associated with acute cholestasis hepatitis. We present a patient with an Epstein-Barr virus infection and acute cholestasis hepatitis with a history of the appearance of a right lateral cervical mass. Physical examination shows jaundice at the level of sclera, mucosa and both upper limbs. Elevated blood bilirubin levels. Patient with hepatic ultrasound and normal bile ducts, normal endoscopic retrograde cholangiopancreatography, so serological tests for EBV are performed and this is positive. Ganciclovir is given, improving liver function tests and decreasing jaundice, thus having a favorable evolution of the patient...(AU)

Porcine embryo production following in vitro fertilization and intracytoplasmic sperm injection from vitrified immature oocytes matured with a granulosa cell co-culture system.

This study was designed to evaluate the capacity of vitrified-warmed porcine immature oocytes to mature and to be fertilized using in vitro fertilization or intracytoplasmic sperm injection, and to determine the subsequent embryo development. Immature oocytes were vitrified using ethylene glycol and dimethylsulphoxide as cryoprotectants and the Cryolock method. After warming oocytes were cultured 44 h for maturation. Oocytes were randomly distributed in three treatment groups and subjected to in vitro fertilization (Experiment 1) or intracytoplasmic sperm injection (Experiment 2) procedures. The results indicate that the embryo development was higher in denuded oocytes co-cultured with granulosa cells (NkO-CC group) fertilized by in vitro fertilization or intracytoplasmic sperm injection compared to cumulus-cell oocyte complexes (COCs group), showing no significant differences with control. Vitrified denuded oocytes matured with a co-culture system NkO-CC group, displayed higher cleavage rate and blastocyst production than vitrified COCs group. Blastocysts were successfully obtained after IVF and ICSI procedures; however, the development to the blastocyst stage was better after IVF. These results show that the vitrification-warming media, the employment of a granulosa cell co-culture system and the Cryolock method during vitrification, increased the nuclear and cytoplasmic maturation of vitrified porcine immature oocytes. Further experiments are required to enhance porcine embryo production after vitrification.

In vivo and in vitro estrogenic profile of 17ß-amino-1,3,5(10)estratrien-3-ol.

17ß-amino-1,3,5(10)estratrien-3-ol (17ßAE2), is the 17ß-aminoestrogens prototype possessing anticoagulant activity, contrasting with the procoagulant effects of 17ß-estradiol (17ßE2). Its estrogenicity profile has not been reported, and it was evaluated by uterotrophic assay, estrogen receptor binding affinity and its ability to induce gene transcription of the human estrogen receptor (hER)α mediated in a Saccharomyces cerevisiae yeast expression system. Additionally, 17ßAE2 and 17αAE2 were compared with 17ßE2 in HeLa cells co-transfected with expression vectors for hERα or hERß subtypes and for an estrogen-responsive reporter gene. Immature female CD1 mice and Wistar rats (21 days old) were treated for three days with 17ßAE2 (10-5000 µg/kg), 17ßE2 (0.001-1000 µg/kg) or vehicle (propylenglycol 10 ml/kg) and uterine weights were estimated. 17ßAE2 increased uterine weight in a dose-dependent manner. The effective dose (ED)50 uterine weight values: 17ßAE2=552 and 764 µg/kg (17ßE2=4.8 and 16 µg/kg) and their relative uterotrophic potency were 0.86 and 2.1 (17ßE2=100) in mice and rats, respectively. 17ßAE2 competed with [(3)H]E2 for the estrogen receptor. The 17ßAE2 relative binding affinities (RBAs) were: 0.074; Ki=2.2×10(-6)M (17ßE2=100; Ki=1.6×10(-9)M); 0.029 and Ki=3.8×10(-6)M (17ßE2=100; Ki=1.1×10(-9)M) for mice and rats uteri respectively. 17ßAE2 activated hERα-mediated ß-galactosidase transcription activity in the yeast system co-transfected with hERα gene. 17ßAE2 effective concentration (EC)50=1.82 µM (17ßE2=2.14 nM) with a relative potency of 0.12 (17ßE2=100). These transactivation effects were abolished by the antagonist fulvestrant (ICI 182,780), similarly to 17ßE2. 17ßAE2 and 17αAE2 bind with low relative affinity to hERα and hERß. Both induced hER-mediated reporter gene transactivation in a dose-response manner. The overall results provide evidence that 17ßAE2 has a weak agonist estrogenic action greatly mediated through the hERß and to a lesser extent the hERα at genomic level.

Co-culture with granulosa cells improve the in vitro maturation ability of porcine immature oocytes vitrified with cryolock.

This study was designed to evaluate the efficiency of two oocyte vitrification-warming procedures using two different devices: Superfine Open Pulled Straws (SOPS) and Cryolock, as well as the effect of the co-culture of vitrified immature oocytes with fresh granulosa cells to improve in vitro maturation (IVM). Immature oocytes were vitrified with two procedures: A) Oocytes were exposed to an increasing concentration of ethylene glycol (EG) from 4% to 35% with 0.5 M trehalose. They then, were loaded in SOPS or Cryolock. For warming, oocytes were exposed to decreasing concentrations of trehalose 0.3, 0.2 and 0.1 M for IVM. B) Oocytes were exposed to two mixtures of EG and dimethylsulfoxide (Me2SO), at 7.5% and 16%, both with 0.4 M of sucrose and then loaded in SOPS or Cryolock and stored in liquid nitrogen. For warming, oocytes were exposed to a single concentration of sucrose 0.5M. After warming, viability was determined; and after 44 h of IVM both viability and meiotic stages were evaluated. The results indicate no significant differences between procedures A and B with SOPS in all maturation stages, reaching a maximum maturation rate of 21%. As to Cryolock, significant differences were observed between both procedures, being procedure B, more efficient with a yield of 38% in MII stage and increased to 49% due to the co-culture with fresh granulosa cells. In conclusion, viability and maturation rates were improved with Cryolock and procedure B with the co-culture system in vitrified immature oocytes.

Litter size determines circadian periodicity of nursing in rabbits.

Nursing in rabbits occurs inside the nest with circadian periodicity. To determine the contribution of suckling stimulation in regulating such periodicity, we varied the size of the litters provided (1, 2, 4, or 6-8 pups). Nursing does, kept under a 14:10 (L:D) photoperiod, were continuously videotaped from parturition into lactation day 15. Although parturitions occurred throughout the day, a significant negative linear correlation (p < 0.0001; r = -0.68) was evident between time of delivery and time of nursing on lactation day 1, regardless of newborn number: longer intervals between these two events were seen in does delivering in the early morning than in those that gave birth late in the day. In rabbits suckling 6-8 pups, a Rayleigh analysis revealed that the population vector best describing their nursing pattern (across lactation days 1-15) had a phase angle = 58° (corresponding to solar time 0352 h and rho = 0.78; p < 0.001). In contrast, the nursing pattern of does nursing litters smaller than 6 pups did not show circadian periodicity; rather, mothers showed multiple entrances into the nest box throughout the day. Cluster analysis revealed that the main equilibrium point of intervals between suckling bouts shifted from 24 h (6-8 pups) to 6 h (4 and 2 pups) and to as low as 4 h with 1 pup. In the groups nursing 2, 4, or 6-8 pups, most nursing episodes were followed by food and water intake. Those mothers also showed self-grooming of the ventrum and nipples after nursing. The incidence of these behaviors was lower in does nursing 1 pup. In conclusion, nursing in rabbits spontaneously occurs with circadian periodicity, but it is largely modulated by a threshold of suckling stimulation.

In vivo profile of the anticoagulant effect of 17ß-amino-1,3,5(10)estratrien-3-ol.

The anticoagulant activity of 17ß-amino-1,3,5(10)estratrien-3-ol (AE(2)) was established for the first time. Experiment 1: mice groups were treated with a single subcutaneous (s.c.) AE(2) injection (0.5, 1, 2, 4, and 8 mg/100 g BW) or vehicle (propylenglycol; 0.5 ml/100 g). After 24 h, AE(2) produced dose-dependent blood clotting time increases related to control, Emax=+121% (P<0.01) finishing the sixth day. Experiment 2: four groups received a single s.c. administration of AE(2) (4 or 8 mg/100g BW) or 17ß-estradiol (E(2); 3mg/100g BW) or vehicle. After 24 and 48 h post-administration, the times of blood clotting, prothrombin, thrombin, and activated partial thromboplastin and fibrinogen concentrations were assessed. Both AE(2) doses increased blood clotting and fibrinogen similarly, blood clotting time: 64, 94%; fibrinogen: 71, 107% (P<0.01). Prothrombin, activated partial thromboplastin and thrombin times, increased 13-15%, 27-55%, and 15-29%, respectively (P<0.01). Meanwhile E(2) decreased blood clotting 20% (P<0.01) and thrombin 23% (P<0.01) after 48 h. Experiment 3: for five consecutive days, mice received AE(2) or E(2) (0.1, 1, 10, 100, and 1000 µg/kg/day), or vehicle. Blood clotting time was assessed at 1, 2, 3, 4, 5, 8, and 11 days after treatment. AE(2) at all doses were anticoagulant for 2-3 days after administration whereas E(2) was procoagulant for 8-11 days. These opposite effects were: AE(2) Emax=+29%; E(2) Emax=-30%; (P<0.01). AE(2) is the parent compound of the 17ß-aminoestrogens, with the largest and longest anticoagulant effect until now reported.

Neonatal rat osteoblasts bioconvert testosterone to non-phenolic metabolites with estrogen-like effects on bone cell proliferation and differentiation.

Testosterone (T) restores bone mass loss in postmenopausal women and osteoporotic men mainly through its bioconversion to estradiol (E2). In target tissues, T is also biotransformed to the A-ring-reduced metabolites 3α,5α-androstanediol (3α,5α-diol) and 3ß,5α-androstanediol (3ß,5α-diol), which are potent estrogen receptor (ER) agonists; however, their biological role in bone has not been completely elucidated. To assess if osteoblasts bioconvert T to 3α,5α-diol and to 3ß,5α-diol, we studied in cultured neonatal rat osteoblasts the metabolism of [14C]-labeled T. In addition, the intrinsic estrogenic potency of diols on cell proliferation and differentiation in neonatal calvarial rat osteoblasts was also investigated. Osteoblast function was assessed by determining cell DNA, cell-associated osteocalcin, and calcium content, as well as alkaline phosphatase activity and Alp1 gene expression. The results demonstrated that diols were the major bioconversion products of T, with dihydrotestosterone being an obligatory intermediary, thus demonstrating in the rat osteoblasts the activities of 5α-steroid reductase and 3α- and 3ß-hydroxysteroid dehydrogenases. The most important finding was that 3ß,5α- and 3α,5α-diols induced osteoblast proliferation and differentiation, mimicking the effect of E2. The observation that osteoblast differentiation induced by diols was abolished by the presence of the antiestrogen ICI 182,780, but not by the antiandrogen 2-hydroxyflutamide, suggests that diols effects are mediated through an ER mechanism. The osteoblast capability to bioconvert T into diols with intrinsic estrogen-like potency offers new insights to understand the mechanism of action of T on bone cells and provides new avenues for hormone replacement therapy to maintain bone mass density.

In vivo and in vitro evaluation of the estrogenic properties of the 17ß-(butylamino)-1,3,5(10)-estratrien-3-ol (buame) related to 17ß-estradiol.

BACKGROUND: Buame [17ß-(butylamino)-1,3,5(10)-estratrien-3-ol] possesses anticoagulant and antiplatelet activities that are potentially antithrombotic. Since its estrogenicity is unknown, it was evaluated by established methods. METHODS: Buame (10, 100, 500, and 1,000 µg/kg), 17ß-estradiol (E(2)) (100 µg/kg), or propylene glycol (10 ml/kg) were subcutaneously (sc) administered for three days to immature Wistar female rats (21 days old). The relative uterotrophic effect to E(2) was 78 (E(2) = 100) with a relative uterotrophic potency of 1.48 (E(2) = 100). Adult ovariectomized Wistar rats received an sc injection at 8:00 h (reversed cycle) of: 7.5 µg of E(2) (≈ 30 µg/kg), buame (≈ 750, 1,500, 3,000 µg/kg), or corn oil (≈ 1.2 ml/kg). After 24 h, progesterone (4-5 mg/kg) was administered. Sexual receptivity was assessed 5 to 7 h later, and the lordosis quotient (LQ; number lordosis/number mounts x 100) was evaluated. RESULTS: Buame induced lordosis (LQmax 85 ± 9; ED50 952 ± 19 µg/kg) and E(2) LQmax 56 ± 8; ED50 10 ± 2 µg/kg; the relative LQ-potency was 0.51 (E(2) = 100). Buame competed with [(3)H]E(2) for the estrogen receptor (Buame RBA= 0.15 and Ki = 5.9 x 10(-7) M; E(2) RBA= 100;Ki = 6.6 x 10(-9) M). Buame increased MCF-7 cells proliferation, from 10(-11) to 10(-)9 M, its proliferative effect was 1.73-1.79 (E(2) = 3.0-3.9); its relative proliferative effect to E(2) was 33-40% (E(2) = 100%) and relative potency 10.4-10.7 (E(2) = 100). Tamoxifen and fulvestrant (ICI 182,780) inhibited buame's proliferation indicating mediation through estrogen receptors in this response. CONCLUSION: Buame is therefore an estrogen partial agonist with a weak estrogenic activity.

Bioconversion of norethisterone, a progesterone receptor agonist into estrogen receptor agonists in osteoblastic cells.

A number of clinical studies have demonstrated that norethisterone (NET), a potent synthetic progestin, restores postmenopausal bone loss, although its mode of action on bone cells is not fully understood, while the effect of naturally occurring progesterone in bone has remained controversial. A recent report claims that the potent effects of NET on osteoblastic cell proliferation and differentiation, mimicking the action of estrogens, are mediated by non-phenolic NET derivatives. To determine whether osteoblasts possess the enzymes required to bioconvert a progesterone receptor (PR) agonist into A-ring reduced metabolites with affinity to bind estrogen receptor (ER), we studied the in vitro metabolism of [(3)H]-labeled NET in cultured neonatal rat osteoblasts and the interaction of its metabolic conversion products with cytosolic -osteoblast ER, employing a competition analysis. Results indicated that NET was extensively bioconverted (36.4%) to 5 alpha-reduced metabolites, including 5 alpha-dihydro NET, 3 alpha,5 alpha-tetrahydro NET (3 alpha,5 alpha-NET) and 3beta,5 alpha-tetrahydro NET (3beta,5 alpha-NET), demonstrating the activities of 5 alpha-steroid reductase and two enzymes of the aldo-keto reductases family. Expression of Srd5a1 in neonatal osteoblast was well demonstrated, whereas Srd5a2 expression was not detected. The most striking finding was that 3beta,5 alpha-NET and 3 alpha,5 alpha-NET were efficient competitors of [(3)H]-estradiol for osteoblast ER binding sites, exhibiting affinities similar to that of estradiol. The results support the concept that the interplay of 5 alpha-steroid reductase and aldo-keto reductases in osteoblastic cells, acting as an intracrine modulator system is capable to bioconvert a PR agonist into ER agonists, offering an explanation of the molecular mechanisms NET uses to enhance osteoblastic cell activities.

The effects of synthetic 19-norprogestins on osteoblastic cell function are mediated by their non-phenolic reduced metabolites.

The key role of estrogens on osteoblastic cell function is well documented; however, the role of progesterone (P) and synthetic progestins remains controversial. While several reports indicate that P has no significant effects on bone cells, a number of clinical studies have shown that 19-norprogestins restore postmenopausal bone loss. The mechanisms by which 19-norprogestins induce estrogen-like effects on bone cells are not fully understood. To assess whether the actions of 19-norprogestins on osteoblasts are mediated by their non-phenolic metabolites, we studied the effects of norethisterone (NET), levonorgestrel (LNG), and two of their A-ring reduced derivatives upon cell proliferation and differentiation in neonatal rat osteoblasts. Osteoblast function was assessed by determining cell DNA, cell-associated osteocalcin and calcium content, alkaline phosphatase activity, and mineral deposition. P failed to induce changes on osteoblasts, while NET and LNG exerted a number of actions. The most striking finding was that the 3beta,5alpha- and 3alpha,5alpha-tetrahydro derivatives of NET and LNG induced osteoblast proliferation and differentiation with higher potency than those exerted by their parent compounds, mimicking the effects of estradiol. Interestingly, osteoblast differentiation and mineral deposition induced by NET and LNG were abolished by finasteride, a 5alpha-reductases inhibitor, while the potent effect on osteoblast proliferation induced by progestin derivatives was abolished by a steroidal antiestrogen. Results demonstrate that A-ring reduced derivatives of NET and LNG exhibit intrinsic estrogen-like potency on rat osteoblasts, offering a plausible explanation for the mechanism of action of 19-norprogestins in bone restoration in postmenopausal women and providing new insights for hormone replacement therapy research.

Enhanced formation of non-phenolic androgen metabolites with intrinsic oestrogen-like gene transactivation potency in human breast cancer cells: a distinctive metabolic pattern.

Breast cancer is a sex steroid hormone-dependent malignant neoplasia. The role of oestradiol in this malignancy has been well documented; however, the involvement of androgens has remained controversial. To determine the role of non-phenolic androgen metabolites in human breast cancer, we studied the metabolism of [(14)C] testosterone and [(14)C] androstenedione in oestrogen-dependent MCF-7 cells and non-oestrogen-dependent MDA-MB 231 cells, at different substrate concentrations (1-10 muM) and time periods (30 min-48 h). Cultured non-oestrogen-dependent HeLa and yeast cells served as controls. Metabolites were identified and quantified by reverse isotope dilution. A distinctive pattern of androgen metabolism was identified in MCF-7 cells, being the 5alpha-androstane-3alpha,17beta-diol (3alpha,5alpha-diol) and its 3beta epimer (3beta,5alpha-diol), the major conversion products of testosterone (48.3%), with 5alpha-dihydrotestosterone as intermediary. The formation of 3alpha,5alpha-diol and 3beta,5alpha-diol (diols) was substrate concentration- and time-dependent, and abolished by finasteride. In contrast, very little of any diol formation was observed in MDA-MB 231, HeLa and yeast cell incubations. Additional enzyme gene expression studies revealed an overexpression of 5alpha-steroid reductase type-1 in MCF-7 cells, as compared with MDA-MB 231 cells. The oestrogen-like activities of diols were assessed in HeLa cells co-transfected with expression vectors for alpha or beta subtypes of the human oestrogen receptor (hER) genes and for an oestrogen-responsive reporter gene. The results show that 3beta, 5alpha-diol and to a lesser extent 3alpha,5alpha-diol bind with high relative affinity to hERalpha and hERbeta. Both diols induced hER-mediated reporter gene transactivation in a dose-response manner, similar to that induced by oestradiol, though with lower potency, an effect that was abolished by ICI-182 780. Furthermore, 3beta,5alpha-diol and to lesser extent 3alpha,5alpha-diol induced MCF-7 cell proliferation. The overall results demonstrated that MCF-7 cells exhibit enhanced expression and activity of androgen-metabolising enzymes, leading to rapid and large diol formation, and provide evidence that these androgen metabolites exert a potent oestrogen-agonistic effect, at genomic level, in oestrogen-dependent breast cancer cells. The data suggest that diols may act as in situ intracrine factors in breast cancer and that its formation can be pharmacologically inhibited.

Synthetic 19-nortestosterone derivatives as estrogen receptor alpha subtype-selective ligands induce similar receptor conformational changes and steroid receptor coactivator recruitment than natural estrogens.

The binding of estradiol (E(2)) to estrogen receptors (ER) is followed by conformational changes resulting in coactivator or corepressor recruitment that influences gene transcription. A series of synthetic A-ring reduced 19-nortestosterone-derived progestins has the capacity to selectively bind and activate transcription through the ERalpha. Herein, the molecular mechanisms involved in ER subtype-selective interactions of these compounds as assessed by their effects upon both ERalpha and ERbeta structural conformation and their ability to induce recruitment of steroid receptor coactivator-1 (SRC-1) to ERalpha were investigated. The results demonstrated that all synthetic A-ring 3beta,5alpha-tetrahydro-reduced derivatives of 19-nortestosterone induced an ERalpha trypsin digestion pattern similar to that seen with E(2), without effects upon ERbeta. In addition, these compounds had the ability to recruit SRC-1 to the ligand-binding domain of ERalpha similar to E(2). Our data indicate that A-ring 3beta,5alpha-tetrahydro-reduced 19-nortestosterone-derived progestins behave as selective ERalpha agonists with ligand-receptor structural and functional responses similar to those induced with natural E(2).

Comparative evaluation of androgen and progesterone receptor transcription selectivity indices of 19-nortestosterone-derived progestins.

Synthetic 19-nortestosterone-derived progestins show affinity for the androgen receptor (AR) and retain varying degrees of androgenic activity. In this study, AR- and progesterone receptor (PR)-dependent transcriptional activation induced by norethisterone (NET), levonorgestrel (LNG) and gestodene (GSD), and their 5alpha-reduced derivatives, including limited trypsin digestion of AR in the presence of natural and synthetic progestins were investigated. The results confirmed the progestogenic activity of the three 19-nortestosterone derivatives, which decreases after reduction of the 4-ene-double bound. These compounds were able to activate AR-dependent reporter gene expression, LNG and GSD being the stronger activators. 5alpha-Reduction of LNG and GSD did not change their androgenic transcriptional activity; however, the activation of AR by 5alpha-NET was four-fold higher than NET. The highest selectivity transcriptional index, as a measure of progestogenicity versus androgenicity, was obtained for NET. The 5alpha-reduced derivatives had values significantly lower than those of their parent compounds. Non-reduced and 5alpha-reduced 19-nortestosterone progestins induced virtually identical proteolysis fragmentation patterns of the AR to those observed with DHT.

Vasodilating effect of norethisterone and its 5 alpha metabolites: a novel nongenomic action.

Estrogens are generally administered in hormone replacement therapy in combination with synthetic progestins. Studies of cardiovascular risk factors in postmenopausal women have shown a variety of responses according to the molecular structure of the progestin used in hormone replacement therapy schemes. The present study sets out to determine the vasoactive effects of norethisterone and its 5alpha-dihydro (5alpha-norethisterone) and -tetrahydro (3alpha,5alpha-norethisterone and 3beta,5alpha-norethisterone) metabolites in isolated precontracted rat thoracic aorta. The addition of norethisterone and 3alpha,5alpha-norethisterone in rat aorta exhibited a potent, concentration-response inhibition of noradrenaline-induced contraction, while 5alpha- and 3beta,5alpha-norethisterone had very little, if any, vasorelaxing effect. Relaxation to norethisterone and 3alpha,5alpha-norethisterone had very rapid time-courses and it was neither affected by the absence of endothelium nor by the inhibitor of nitric oxide synthase, Nomega-nitro-L-arginine methyl ester (L-NAME). The addition of specific anti-androgen, anti-progestin and anti-estrogen compounds and protein synthesis inhibitors did not preclude the vasorelaxing effect of norethisterone and its 3alpha,5alpha-reduced metabolite. The results strongly suggest that these effects are not mediated by nuclear sex steroid hormone receptors. The overall data document a novel nongenomic endothelium-independent vasorelaxing action of a 19-nor synthetic progestin and one of its A-ring-reduced derivatives.

In vivo and in vitro estrogen bioactivities of alkyl parabens.

The alkyl esters of p-hydroxybenzoic acid known as parabens (Pbens) are used as preservatives in food, pharmaceutical and cosmetic formulations. They have been reported as estrogenic. Here, we present evidence for the in vivo and in vitro bioactivities and receptor binding affinities of methylparaben (MePben), ethylparaben (EtPben), propylparaben (PrPben), and butylparaben (BuPben) compared with those of estradiol (E2). Estrogenicity was studied using the uterotrophic assay in immature (Im) and adult ovariectomized (Ovx) CD1 mice, and in immature female Wistar rats (IW). Animals were subcutaneously (sc) treated for three consecutive days with different molar equivalent doses ranging from 3.62 to 1086 micromol/kg body weight of Pbens, E2 (0.036 micromol/kg), or vehicle. Pbens increased uterine weight in Im and Ovx animals and their relative uterotrophic effect to E2 (100) (RUEE2) were from 34 to 91. The relative uterotrophic potencies related to E2 (100) (RUPE2) of these compounds were from 0.003 to 0.007. The E2 ED50 for CD1 animals able to increase the uterine weight was 7 microg/kg (0.9-55 confidence limits); and that of Pbens ranged from 18 to 74 mg/kg. In IW rats, the ED50 were from 33 to 338 mg/kg. All Pbens, except MePb, competed with [3H]E2 for the estrogen receptor binding sites. The uterotrophic effects of Pbens in Im mice have a positive correlation with the side-chain length of the ester group of these compounds. The E2 and Pbens relative binding affinities (RBA) and Ki values correlated to their estrogenic activity. The NOELs values for Pbens uterotrophic activity in Im were from 0.6 to 6.5 mg/kg per day; and Ovx from 6 to 55 mg/kg. The NOELs IW ranged from 16.5 to 70 mg/kg indicating that Im were more susceptible than Ovx and IW to these effects. The data shown here confirm the estrogenicity of Pbens.

The intrinsic transcriptional estrogenic activity of a non-phenolic derivative of levonorgestrel is mediated via the estrogen receptor-alpha.

Levonorgestrel (LNG), a 19-nor-testosterone derivative, is widely used in contraceptive formulations. This compound does not bind to the estrogen receptor (ER), but it shows estrogen-like effects under in vivo and in vitro conditions. The estrogenicity of LNG may be attributed to its bio-transformation into non-phenolic metabolites. In this study, the ability of A-ring reduced LNG metabolites to activate transcription via an estrogenic mechanism of action, including differences between ER alpha and ER beta subtypes, were investigated. Transactivation assays were performed in HeLa cells transfected with expression vectors for ER alpha and ER beta and an estrogen-responsive reporter gene. Cells were also transfected with expression vectors for both progesterone receptor (PR) isoforms (A or B). As expected, the tetrahydro derivatives of LNG (3 alpha,5 alpha- and 3 beta,5 alpha-LNG) showed significantly lower PR-mediated transcriptional activities through both isoforms when compared with progesterone (P(4)) and LNG. In contrast, the 3 beta,5 alpha-tetrahydro derivative resulted in a significant activation of estrogen-dependent gene transcription. This effect was selectively confined to the ER alpha, since little if any activity could be observed with the ER beta and no antagonistic activities were demonstrated. This study provides structural and molecular clues for the well documented in vitro and in vivo intrinsic estrogenicity of 19-nor-testosterone-derived progestins and ligand requirements for ER alpha recognition.