Human rotavirus strains circulating in Venezuela after vaccine introduction: predominance of G2P[4] and reemergence of G1P[8].

BACKGROUND: Rotavirus (RV) is the most common cause of severe childhood diarrhea worldwide. Despite Venezuela was among the first developing countries to introduce RV vaccines into their national immunization schedules, RV is still contributing to the burden of diarrhea. Concerns exist about the selective pressure that RV vaccines could exert on the predominant types and/or emergence of new strains. RESULTS: To assess the impact of RV vaccines on the genotype distribution 1 year after the vaccination was implemented, a total of 912 fecal specimens, collected from children with acute gastroenteritis in Caracas from February 2007 to April 2008, were screened, of which 169 (18.5%) were confirmed to be RV positive by PAGE. Rotavirus-associated diarrhea occurred all year-round, although prevailed during the coolest and driest months among unvaccinated children under 24 months old. Of 165 RV strains genotyped for G (VP7) and P (VP4) by seminested multiplex RT-PCR, 77 (46.7%) were G2P[4] and 63 (38.2%) G1P[8]. G9P[8], G3P[8] and G2P[6] were found in a lower proportion (7.3%). Remarkable was also the detection of <5% of uncommon combinations (G8P[14], G8P[4], G1P[4] and G4P[4]) and 3.6% of mixed infections. A changing pattern of G/P-type distribution was observed during the season studied, with complete predominance of G2P[4] from February to June 2007 followed by its gradual decline and the reemergence of G1P[8], predominant since January 2008. Phylogenetic analysis of VP7 and VP4 genes revealed a high similarity among G2P[4] and global strains belonging to G2-II and P[4]-V lineages. The amino acid substitution 96D → N, related with reemergence of the G2 genotype elsewhere, was observed. The G1P[8] strains from Caracas were grouped into the lineages G1-I and P[8]-III, along with geographically remote G1P[8] rotaviruses, but they were rather distant from Rotarix® vaccine and pre-vaccine strains. Unique amino acid substitutions observed on neutralization domains of the VP7 sequence from Venezuelan post-vaccine G1P[8] could have conditioned their re-emergence and a more efficient dissemination into susceptible population. CONCLUSIONS: The results suggest that natural fluctuations of genotypes in combination with forces driving the genetic evolution could determine the spread of novel strains, whose long-term effect on the efficacy of available vaccines should be determined.

Tibolone induces serotonin, estrogen, and progesterone receptor expression but not contractile response to serotonin in the rat uterus.

Most studies on the effect of tibolone on the uterus have focused on the endometrium dismissing the importance of the myometrium. The aim of the present study was to investigate some estrogen-like actions of tibolone in the uterus assessed by: 1) the expression of estrogen, progesterone, and serotonin receptors, and 2) the myometrial contraction induced by serotonin. Estradiol (250 µg), progesterone (50 mg), or testosterone (25 mg) pellets were implanted to ovariectomized rats. Tibolone (0.5 mg/day) was orally administered. An implanted pellet containing vehicle or an equivalent volume of water p.o., were used as controls. Sixty days after beginning the treatments, rats were killed and uterus removed. One horn was processed to evaluate estrogen-alpha, progesterone A and B, and serotonin-2A receptors expression, and the other one was used for studying contraction to serotonin and 60 mM potassium solution. The present data showed that tibolone-induced expression of estrogen, progesterone, and serotonin receptors, but did not induce uterine contractile response to either serotonin or potassium solution. These findings suggest that, in the uterus, tibolone may exert molecular estrogenic actions such as the induction of receptor expression, but not a physiological response as the estrogen-dependent contraction to serotonin.

Socioeconomic impact on device-associated infections in limited-resource neonatal intensive care units: findings of the INICC.

PURPOSE: To evaluate the impact of country socioeconomic status and hospital type on device-associated healthcare-associated infections (DA-HAIs) in neonatal intensive care units (NICUs). METHODS: Data were collected on DA-HAIs from September 2003 to February 2010 on 13,251 patients in 30 NICUs in 15 countries. DA-HAIs were defined using criteria formulated by the Centers for Disease Control and Prevention. Country socioeconomic status was defined using World Bank criteria. RESULTS: Central-line-associated bloodstream infection (CLA-BSI) rates in NICU patients were significantly lower in private than academic hospitals (10.8 vs. 14.3 CLA-BSI per 1,000 catheter-days; p < 0.03), but not different in public and academic hospitals (14.6 vs. 14.3 CLA-BSI per 1,000 catheter-days; p = 0.86). NICU patient CLA-BSI rates were significantly higher in low-income countries than in lower-middle-income countries or upper-middle-income countries [37.0 vs. 11.9 (p < 0.02) vs. 17.6 (p < 0.05) CLA-BSIs per 1,000 catheter-days, respectively]. Ventilator-associated-pneumonia (VAP) rates in NICU patients were significantly higher in academic hospitals than in private or public hospitals [13.2 vs. 2.4 (p < 0.001) vs. 4.9 (p < 0.001) VAPs per 1,000 ventilator days, respectively]. Lower-middle-income countries had significantly higher VAP rates than low-income countries (11.8 vs. 3.8 per 1,000 ventilator-days; p < 0.001), but VAP rates were not different in low-income countries and upper-middle-income countries (3.8 vs. 6.7 per 1,000 ventilator-days; p = 0.57). When examined by hospital type, overall crude mortality for NICU patients without DA-HAIs was significantly higher in academic and public hospitals than in private hospitals (5.8 vs. 12.5%; p < 0.001). In contrast, NICU patient mortality among those with DA-HAIs was not different regardless of hospital type or country socioeconomic level. CONCLUSIONS: Hospital type and country socioeconomic level influence DA-HAI rates and overall mortality in developing countries.

Effects of single sessions of low-intensity continuous and moderate-intensity intermittent exercise on blood lipids in the same endurance runners.

The factors responsible for the acute effects of exercise on blood lipids are not well known, and there have been few studies comparing different kinds of exercise in the same population. The concentration of blood lipids was evaluated in this study at the end and at post-24h of two 14km/90min single exercise sessions: continuous exercise (CE) at 44.5+/-5.6% VO(2max) and intermittent exercise (IE) at 39-72% VO(2max), in subjects with high levels of aerobic training. Fourteen male athletes (endurance runners) took part in this study and each completed a 24h dietary record. The O(2) uptake and CO(2) production were recorded, and blood lactate and blood lipids were measured. The results showed that triacylglycerols were not modified by any kind of exercise. Total cholesterol was increased at the end of both exercises: 7.04% for CE (p<0.001) and 4.23% for IE (p=0.001). High-density lipoprotein cholesterol was increased at the end of IE: 11.38% (p=0.03) and low-density lipoprotein cholesterol was increased only at the end of CE: 7.45% (p=0.006). The increase of lipids for CE was negatively correlated with aerobic fitness indicators (heart rate and %HRmax at lactate threshold), and was positively associated with energy expenditure. For IE, %HRmax and lactate were negatively correlated, and the respiratory exchange ratio was positively correlated, with the lipid increase. We conclude that in trained male athletes, a 14km run in 90min induced different changes of lipid profile if the exercise was done continuously or intermittently, and that in CE the extent of these increases was influenced by aerobic fitness.

Stability of plasma for add-on PT and APTT tests.

We conducted studies to determine at what time point an add-on prothrombin time (PT) or activated partial thromboplastin time (APTT) test can be honored on specimens that have been received in the laboratory hours earlier without yielding results with clinically significant differences from those if the test had been performed on the original unstored plasma. PT and APTT tests were performed on blood samples from 20 healthy subjects, 30 patients receiving warfarin, and 30 patients receiving heparin anticoagulation therapy. The tests were performed on plasma prepared initially after the samples were obtained. The same tests were assayed on plasma that had been left on spun-down blood cells at room temperature for 2, 4, and 8 hours. We found that the PT of the majority of plasma samples from healthy subjects and from patients receiving oral anticoagulant therapy tended to become shorter on storage. However, the difference in PT values was small and had no clinical significance. In most cases, the APTT values for the stored plasma from healthy subjects tended to increase with time. Except in one specimen in which the 8-hour add-on APTT was 1.2 seconds longer than the APTT result for the original sample, all others had APTT results less than 1.2 seconds longer than the original values. In patients receiving heparin, the differences in APTT values between the initial and add-on tests were larger than those observed for healthy subjects. However, those differences are not beyond what we would accept for duplicate checks for heparinized samples with high APTT values. Unlike samples from healthy subjects, there was no obvious trend of time-related prolongation of the APTT in heparinized plasma. These results led us to believe that within an 8-hour period and with plasma on spun-down cells at room temperature, add-on tests for PT and APTT could be performed with results similar to what would be obtained from testing unstored samples.

The ovarian and cervical regions of the rat uterus display a different contractile response to serotonin and prostaglandin F2alpha. I. The estrous cycle.

In pharmacological studies using isolated tissues, the sensitivity to different agonists may vary depending on the anatomical region. The aim of the present study was to evaluate the in vitro contractile response to serotonin, prostaglandin F2alpha, and oxytocin of the ovarian and the cervical uterine segments isolated from rats in the four different stages of the rat estrous cycle. Non-cumulative curves were recorded for both, the ovarian and the cervical uterine segments. The cervical portion displayed a higher contractile response to serotonin and a lower response to PGF2alpha than the ovarian portion. Oxytocin induced similar responses in both uterine segments. The uterine ovarian segment displayed a similar sensitivity to serotonin in all the estrous cycle stages, whereas in the cervical segment, influenced by estrogens in diestrus and proestrus, an increase in contractility was observed. According to these findings, serotonin might participate in the spermatozoa transport toward the oviduct. The higher response of the ovarian portion to prostaglandin F2alpha is in line with its role during labor and delivery.

Xanthorrhizol induces endothelium-independent relaxation of rat thoracic aorta.

Xanthorrhizol, a bisabolene isolated from the medicinal plant Iostephane heterophylla, was assayed on rat thoracic aorta rings to elucidate its effect and likely mechanism of action, by measuring changes of isometric tension. Xanthorrhizol (1, 3, 10, 30 and 100 microg/mL) significantly inhibited precontractions induced by KCI-; (60mM), noradrenaline (10(-6) M) or CaCl2 (1.0 mM). Increasing concentrations of external calcium antagonized the inhibitory effect on KCl-induced contractions. The vasorelaxing effect of xanthorrhizol was not affected by indomethacin (10 microM) or L-NAME (100 microM) in intact rat thoracic aorta rings precontracted by noradrenaline, which suggested that the effect was not mediated through either endothelium-derived prostacyclin (PGI2) or nitric oxide release from endothelial cells. Endothelium removal did not affect the relaxation induced by xanthorrhizol on rat thoracic aorta rings, discarding the participation of any substance released by the endothelium. Xanthorrhizol inhibitory effect was greater on KCI- and CaCl2-induced contractions than on those induced by noradrenaline. Xanthorrhizol inhibitory effect in rat thoracic aorta is likely explained for interference with calcium availability by inhibiting calcium influx through both voltage- and receptor-operated channels.

The androgenic effect of norethisterone and 5alpha-norethisterone on the contractile response of the rat vas deferens to methoxamine and serotonin.

Norethisterone (NET) and its metabolite 5alpha-norethisterone (5alpha-NET) are competitors for the androgen receptor. The sensitivity of the rat vas deferens to the contractile action of methoxamine and serotonin is regulated by hormonal and anatomical factors. The aim of this study was to evaluate the ability of NET and 5alpha-NET to induce the androgen-regulated contractile response to methoxamine and serotonin in the epididymal and prostatic portions of rat vas deferens. Adult male rats either intact, castrated or steroid-treated castrated were used. The contractility was recorded isometrically, and non-cumulative concentration-response curves to either methoxamine or serotonin were obtained. NET and 5alpha-NET partially restored the sensitivity to methoxamine and serotonin in the epididymal portion of castrated rats. The maximal responses to both agonists were significantly higher than those observed in castrated rats, and significantly lower than the responses observed in either intact or androgen-treated castrated rats. The prostatic portion was less responsive to both agonists than the epididymal portion, in all groups but castrated rats, as castration induced sensitivity to both agonists. NET and 5alpha-NET displayed a partial though similar androgenic activity in the rat vas deferens. These results contrast with previous reports where a decrease of androgenic effect due to the 5alpha-reduction of NET has been found.

Spirulina maxima prevents fatty liver formation in CD-1 male and female mice with experimental diabetes.

The dietary administration of 5% Spirulina maxima (SM) during four weeks to diabetic mice, starting one week after a single dose of alloxan, 250 mg/Kg body weight, prevented fatty liver production in male and female animals. The main action of SM was on triacylglycerol levels in serum and liver. There was also a moderate hypoglycemia in male mice. The thiobarbituric acid reactive substances also decreased in serum and liver after SM administration. There was also a decrease in the percentage of HDL in diabetic mice that was reverted by the SM administration. The sum of LDL + VLDL percentages was also partially normalized in diabetic animals by the SM administration. An additional observation was the lower incidence of adherences between the liver and the intestine loops in the diabetic mice treated with SM compared with diabetic mice without SM. Male and female mice showed differences to diabetes susceptibility and response to SM, the female being more resistant to diabetes induction by alloxan and more responsive to the beneficial effects of SM. It is worth future work of SM on humans looking for better quality of life and longer survival of diabetic patients.

Effects of acute and chronic estrogenic treatment on vasomotor responses of aortic rings from ovariectomized rats.

The effects of either chronic or acute estrogenic treatment on the "in vitro" vasomotor responses to phenylephrine (10(-9)-10(-5) M) and to carbachol (10(-9)-10(-5) M) of aortic rings excised from ovariectomized rats were analyzed. Chronic estrogenic treatment consisted in a single subcutaneous dose of 1 mumol estradiol 17-stearate. Effects of acute estrogenic treatment were evaluated by recording the responses of aortic rings excised from untreated ovariectomized rats both before and after the addition of 17 beta-estradiol to the superfusing solutions. In order to identify the endothelium-dependent responses each experiment was performed simultaneously on pairs of rings from the same aorta, one with and the other without functional endothelium. The contractile responses to phenylephrine of endothelium-intact vessels were attenuated by chronic estrogenic treatment; this attenuation was further increased by preincubation of the vessels with indomethacin and was reverted by N omega-nitro-L-arginine methyl ester. Either chronic or acute estrogenic treatment enhanced the carbachol-induced endothelium dependent relaxation of phenylephrine-precontracted rings. The results may be explained by assuming that estrogens increase the basal release of both nitric oxide and a cyclooxygenase-dependent vasoconstricting prostanoid as well as the receptor-mediated release of nitric oxide from the endothelium of the rat aorta.

Preventive effect of Spirulina maxima on the fatty liver induced by a fructose-rich diet in the rat, a preliminary report.

Cyanobacteria Spirulina maxima from Texcoco Lake in Mexico was administered as a 5% component of a purified diet, to Wistar rats together with a high percentage of fructose (60%) and its effect on several lipid fractions of plasma and liver was studied and compared to those of rats fed purified diets containing 60% of glucose or 60% of fructose. A preventive effect of Spirulina maxima on the fructose-induced increase of the liver triglycerides level was observed together with an elevation of the phospholipid concentration in this tissue. On the other hand Spirulina maxima produced a plasma cholesterol level even lower than that observed in the control group.

Effects of dietary Spirulina maxima on endothelium dependent vasomotor responses of rat aortic rings.

The aim of this study was to evaluate the effects of Spirulina maxima on vasomotor responses of aorta rings from male Wistar rats fed on a purified diet. For this purpose, the animals (weighing 200-240 g) were allocated randomly in two groups. One receiving purified control diet (A) and the other receiving purified diet containing 5% Spirulina (B). Purified diets were according to American Institute of Nutrition guidelines and adjusted to Spirulina protein content. All animals were fed (20 g/day/rat) during two weeks, receiving water ad libitum and 12 h. light-dark cycles. Spirulina maxima effects were evaluated by concentration-response (CR) curves of aorta rings with or without endothelium to phenylephrine (PE), both in presence and absence of indomethacin (Indom) or indomethacin plus L-NAME (Indom. + L-NAME), and to carbachol (CCh). Aorta rings with endothelium from group B showed, relative to corresponding rings from group A: 1) a significant decrease in the maximal tension developed in response to PE. 2) this decrease was reverted by Indom. 3) Indom. + L-NAME induced an additional increase in the contractile responses to PE. 4) a significant shift to the left of the CR curve to CCh. No significant differences were observed in the tension developed in response to PE in rings without endothelium from either group. These results suggest that Spirulina maxima may decrease vascular tone by increasing the synthesis and release of both a vasodilating cyclooxygenase-dependent product of arachidonic acid and nitric oxide, as well as by decreasing the synthesis and release of a vasoconstricting eicosanoid from the endothelial cells.

Arthrospira maxima prevents the acute fatty liver induced by the administration of simvastatin, ethanol and a hypercholesterolemic diet to mice.

An evident fatty liver, corroborated morphologically and chemically, was produced in CD-1 mice after five daily doses of simvastatin 75 mg/Kg body weight, a hypercholesterolemic diet and 20 percent ethanol in the drinking water. After treating the animals, they presented serum triacylglycerols levels five times higher than the control mice, total lipids, cholesterol and triacylglycerols in the liver were 2, 2 and 1.5 times higher, respectively, than in control animals. When Arthrospira maxima was given with diet two weeks prior the onset of fatty liver induction, there was a decrement of liver total lipids (40%), liver triacylglycerols (50%) and serum triacylglycerols (50%) compared to the animals with the same treatment but without Arthrospira maxima. In addition to the mentioned protective effect, the administration of this algae, produced a significant increase (45%) in serum high density lipoproteins. The mechanism for this protective effect was not established in these experiments.

High lovastatin doses combined with hypercholesterolemic diet induce hepatic damage and are lethal to the CD-1 mouse.

The addition of 1% lovastatin (LVT) to hypercholesterolemic diets [1% cholesterol or 1% cholesterol plus 0.1% sodium deoxycholate (HD)] induced hepatic damage and was lethal to CD-1 mice in the first days of treatment; the females were more resistant than males. LVT or HD administered alone was harmless to male or female mice. After a 3-day treatment all groups that received LVT (1%, 0.1% or 0.05%) plus HD showed a higher percentage of liver weight, with respect to whole body weight. Cholesterol serum levels increased in males with HD, but remained low in female mice. In the liver, total lipids and cholesterol levels increased in male mice with HD, but cholesterol remained unchanged in females. The addition of LVT to HD prevented the increase of serum and liver cholesterol levels in male mice. These results allow us to propose the CD-1 male-mouse as a model to evaluate the toxicity of LVT or other vastatins.

Comparative toxicity of high doses of vastatins currently used by clinicians, in CD-1 male mice fed with a hypercholesterolemic diet.

The CD-1 male-mouse model was employed to evaluate comparatively the toxicity of four vastatins (VTS) currently used in clinical medicine: lovastatin (LVT), simvastatin (SVT), pravastatin (PVT) and fluvastatin (FVT). Each vastatin was used orally in doses of 500 mg/Kg body weight/day, in animals with a hypercholesterolemic diet (HD) 5 days, or with a control diet (CD) 30 days. The association of high doses of VTS + HD produced a significant increase in liver weight and liver weight to body weight ratio in animals with SVT and FVT. Cholesterol (Chol) and triacylglycerols (TAG) in the liver increased significantly with FVT but not with the other VTS; Chol increased and TAG decreased in serum very significantly with FVT and SVT. The serum aminotransferases increased quite significantly with FVT but not with other VTS. In the experiment with high doses of VTS + CD, the animals receiving SVT or FVT showed a moderate loss of body weight. Liver weight and liver weight to body weight ratios were similar among all groups. Liver Chol showed a significant decrease with all VTS. Serum Chol decreased moderately with LVT and FVT. TAG in serum and liver showed a moderate decrease with all VTS. The serum aminotransferases were not modified by any vastatin. Our results indicate that high doses of VTS in male mice with a hypercholesterolemic diet result in a decreasing toxicity as follows: FVT>SVT>LVT>PVT.

Effects of intrathecal administration of adrenergic agonists on the frequency of copulatory pelvic thrusting of the male rat.

The effects of the intrathecal perispinal administration of adrenergic agonists on the characteristics of frequency, duration, and vigor of pelvic thrusting displayed by male rats during copulation was assessed by an accelerometric technique. A different dose of one drug (noradrenaline, clonidine or isoproterenol) and saline as control was administered at the lumbosacral level of the spinal cord to sexually active male rats in tests of sexual behavior performed at weekly intervals. The intrathecal administration of noradrenaline (alpha-adrenoceptor agonist) increased the frequency of pelvic thrusting in mount and intromission responses, whereas both the alpha 2-adrenoceptor agonist clonidine (25 micrograms) and the beta-adrenoceptor agonist isoproterenol (40 micrograms) reduced the frequency of pelvic thrusting in these responses as compared to values obtained under the intrathecal administration of saline. On the other hand, the duration of the thrusting trains and the potency or vigor of pelvic thrusting in mounts and intromissions did not differ from values obtained under saline treatment. These findings indicate a possible participation of noradrenaline in the modulation of the spinal mechanisms involved in the generation of rhythmic pelvic thrusting performed by the male rat during copulation.

Estrogenic actions of norethisterone and its A-ring reduced metabolites. Induction of in vitro uterine sensitivity to serotonin.

The estrogen-like effects of norethisterone (NET) seem to be mediated by the interaction of 3 beta 5 alpha- and 3 alpha 5 alpha-tetrahydronorethisterone (3 beta 5 alpha- and 3 alpha 5 alpha-NET, respectively) with the estrogen receptor. Considering that the in vitro uterine contractile response to serotonin (5-HT) is specifically dependent on estrogen, the aim of the present study was to investigate whether NET and its A-ring reduced metabolites administered in vivo to ovariectomized rats induce uterine sensitivity to 5-HT in vitro. The administration of 3 beta NET in vivo, which is the NET metabolite with the highest affinity for the estrogen receptor, induced a maximal contractile response to serotonin similar to that of 17 beta-estradiol treatment. The other metabolites induced less uterine activity. According to the effective dose 50, the order of estrogenic potency was 17 beta-estradiol > 3 beta 5 alpha NET > 3 alpha 5 alpha NET > NET > 5 alpha NET. The estrogenic effect of 3 beta 5 alpha- and 3 alpha 5 alpha NET may be exerted through their interaction with the estrogen receptor, whereas NET and 5 alpha NET, which do not bind to the estrogen receptor and display a minor estrogenic activity, require prior bioconversion to 3 beta 5 alpha NET and perhaps to 3 alpha 5 alpha NET. The A-ring reduced metabolites of NET, mainly the 3 beta 5 alpha NET, may be exerting estrogenic responses and modulating uterine activity when administered in vivo.

Comparative analysis of erythrocyte fatty acid composition in a sample of Mexico City children and young adults on a free diet.

In the present study the influence of age on red blood cell fatty acid (RBCFA) composition was analyzed in a sample of Mexico City children and young people on a free diet, as there is scarce information about RBCFA composition in the Mexican population. Erythrocyte lipids were extracted with isopropyl alcohol and fatty acid methyl esters were prepared to be analyzed by gas liquid chromatography. The 1- to 2-year-old group showed a higher percent level of C18:0 (34.75 +/- 2.5 vs. 29.67 +/- 1.3, p < 0.002) and lower of C16:1 (0.58 +/- 0.2 vs. 1.09 +/- 0.2, p < 0.05), C20:4 (14.08 +/- 4.1 vs, 18.20 +/- 1.2, p < 0.05) and C22:5 (2.79 +/- 1.7 vs. 7.68 +/- 0.8, p < 0.001) than the 20- to 25-year-old group. Both groups showed a very low linoleic acid proportion, children 0.48% and young adults 0.54%. The unsaturated/saturated fatty acid ratio was found to be 0.55 +/- 0.2 in children and 0.91 +/- 0.1 in adults (p < 0.001). These findings indicate the presence of factors related to age that affect the fatty acid composition in the erythrocyte membrane different from diet habits in the sample analyzed. Results are compared with reports in the literature.

Induction of male sexual behavior by norethisterone: role of its A-ring reduced metabolites.

The estrogenic and androgenic potencies of norethisterone (NET), a synthetic nonaromatizable progestin, and three of its reduced metabolites (5 alpha-NET; 3 alpha, 5 alpha-NET; 3 beta, 5 alpha-NET) were assessed by their ability to restore male sexual behavior in castrated male rats following their chronic administration in combination with either 5 alpha-dihydrotestosterone (DHT) or estradiol (E2), or when given alone. Full restoration of mating was achieved when 3 beta, 5 alpha-NET was administered with DHT, indicating an estrogenic effect of this compound. Lower estrogenic effects were noticed with 3 alpha, 5 alpha-NET and 5 alpha-NET, while NET had very little estrogenic potency. The only effective compound to restore ejaculation, when administered with E2, was NET, indicating its androgen-like intrinsic potency. When administered alone, NET exerted the most potent effect on male behavior, followed by 5 alpha-NET, while the tetrahydro derivatives were ineffective. The observation that NET alone restored male sexual activity at a level identical to that induced by testosterone demonstrated an androgenic-estrogenic activity of this progestin exerted through its intrinsic androgenic effect, and the estrogenic effect of its tetrahydro derivatives. Overall results indicated that the metabolism of NET modulates its mode of action at the brain, and support the concept that both estrogenic and androgenic effects are required for mating activation.

Steroid metabolism in granulosa and theca interna cells from preovulatory follicles of domestic hen (Gallus domesticus).

The capability of granulosa and theca interna cells, from preovulatory follicles of the domestic hen, to metabolize steroid precursors was evaluated. Granulosa and theca interna cells were isolated from ovarian preovulatory follicles at three different developmental stages: F1, F3 and F5. Tritiated pregnenolone (P5), progesterone (P4), dehydroepiandrosterone (DHEA), androstenedione (A4) and testosterone (T) were employed as precursors and their metabolic products were evaluated. The major metabolite of P5 by granulosa cells was P4, but we also observed low amounts of 5beta-pregnandione. DHEA metabolism by granulosa cells yielded mainly A4, and minute quantities of 5beta-androstan-3,17-dione (5beta-dione) were detected. The only significant metabolite obtained in granulosa cells from A4 was 5beta-dione, whereas T was only transformed into A4. On the other hand, P5 metabolism by theca interna cells yielded A4 as the main product, also P4, 17alpha-OHP4, 17alpha-OHP5, 5beta-pregnandione, and DHEA, were found. When DHEA was the precursor A4 was produced in higher amounts than 5beta-dione. A4 was mainly transformed into 5beta-dione. In similar conditions, T was transformed into A4. These results show that granulosa cells have enzymatic activities of 3beta-hydroxysteroid dehydrogenase/5-4 isomerase (3beta-HSD from P5 and DHEA), 17beta-hydroxysteroid dehydrogenase (17beta-HSD from T) and 5beta-reductase (from P5, DHEA and A4). Whereas theca interna cells have enzymatic activities of cytochrome P450c17 (from P5 and P4), 3beta-HSD (from P5 and DHEA), 17beta-HSD (from T) and 5beta-reductase (from P4, DHEA and A4). These data support the concept that theca interna cells have the ability to synthesize androgens from progestins produced in granulosa cells. In addition, since theca interna cells did not show the capacity to aromatize androgens suggests that interaction between theca interna and theca externa cells occurs in vivo, thus confirming the three cell model for estrogen production. Furthermore, the fact that other metabolites were produced both in granulosa and theca interna cells, but in a different extent, suggests that complex mechanisms are participating in the regulation of steroid synthesis in avian ovary follicles.