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Biomedical subjects

M C Romano

Publications and source records attributed to M C Romano.

At least 19 recordsLinked to original sources

Role of corticosterone on the development of passive electrical properties of cultured chick embryo neurons.

We worked with neuronal cells of chick embryo cultures. The cells were obtained from 7 days-old embryos and cultured for 12 days. The experimental group was submitted to a single dose of corticosterone during the first 48 hours of culture. We measured the membrane resting potential at different external potassium concentrations, as well as the passive electrical properties of the membrane, resistance, time constant and capacitance. We found that the control cells develop progressively their potassium permeability and the electrical properties of the membrane until reaching values in this parameters that were similar to those found in adult cells. Corticosterone application induced a significant enhancement in these parameters in 3 and 6 days-old cultures as compared with the control cultures. These results suggest that glucocorticoids accelerate the differentiation process of the neurons in culture when applied at very early stages of development. We discuss the possible mechanisms involved in this action.

Animals

An age-dependent thymic secretion modulates testicular function.

The acetone extract obtained from the thymuses of 14-day-old rats contains a factor that interacts with hCG in the adult testis cells and inhibits testosterone production. Experiments were designed to investigate the possible secretion of this factor. The media from the incubation of thymuses from 14-day-old rats were processed by molecular sieve chromatography and the fractions assayed using a bioassay with testicular cells in vitro. A fraction of approx. 30 kDa was found to inhibit the hCG-stimulated production of testosterone. In addition, the influence of age on the release of the active fraction was investigated. The inhibitory effect of this thymus product was greatest in the neonatal period (1-14 days) and declined thereafter towards the onset of puberty. The age-related decline of the inhibitory activity correlated with relative thymus weight and also with the amount of protein released to the incubation media. Thymic fraction activity is, however, present in the adult gland. These results suggest that the thymus secretes active agents that are able to modulate the response of testicular cells to hCG and that their release seems to be age-related.

Aging

Influence of the thymus on steroidogenesis by rat ovarian cells in vitro.

Thymic hormones and factors have been shown to modulate the function of other endocrine glands including the gonads. Absence of the thymus during development results in ovarian dysgenesis characterized by a decrease in the number of follicles and corpora lutea, bringing about severe changes in reproductive function. To examine whether thymic secretions might affect ovarian activity, whole dispersed ovarian cells obtained from immature rats pretreated with pregnant mare serum gonadotrophin were exposed to a thymus fraction of approximately 28 kDa and also to the media from incubated thymuses (TIM) and the conditioned media from cultured thymic reticuloepithelial cells (TCM). The thymic fraction caused a dose-dependent decrease in human chorionic gonadotrophin (hCG)-stimulated production of progesterone, oestradiol and testosterone, but had no effect on their synthesis in the absence of hCG. Similarly, hCG-induced production of these steroids was decreased by TIM and TCM. Progesterone secretion was the most markedly affected. These results suggest: (1) that the thymus contains a factor with a molecular weight of approximately 28 kDa which interacts with hCG in ovarian cells, (2) that the thymus can release active substances which modify steroid secretion by the ovary in vitro and (3) that the reticuloepithelial cells of the thymus are involved in the secretion of factors which modulate the stimulation by hCG of steroidogenesis in ovarian cells.

Animals

Prepubertal rat thymus secretes a factor that modulates gonadotropin secretion in cultured rat pituitary cells.

Thymus development and function are influenced by endocrine glands. On the other hand, the absence of thymus results in endocrine disturbances. The aim of the present paper is to investigate the direct influence of thymus products on the gonadotropin secretion of rat pituitary cells in culture. In this study, a chromatographic fraction obtained from the acetonic powder of thymus from 14-15 day-old rats (TF) increased the GnRH stimulated LH and FSH secretion from cultured pituitary cells, but had no effect on the basal secretion of these gonadotropins. The addition of incubation medium from thymus (TIM) to cultured pituitary cells further enhanced GnRH stimulated FSH and LH production, the effect was dose related. The GnRH potentiating effect of thymus incubation medium was also present in a chromatographic fraction of around 28 Kd obtained from this medium. These data suggest that the prepubertal thymus contains and secretes a factor that modulates pituitary cells secretion potentiating the Gn-RH effect.

Age Factors

Determination of microsomal lauric acid hydroxylase activity by HPLC with flow-through radiochemical quantitation.

An assay for the microsomal hydroxylation of lauric acid (LA), based on HPLC with flow-through radiochemical detection, has been developed. Conditions were optimized for resolution and quantitation of three microsomal metabolites of LA, one of which has not been reported previously as a metabolite of LA in mammalian microsomal incubations. These products, 12-(omega)-hydroxy-LA, 11-(omega-1)-hydroxy-LA, and a novel metabolite, 10-(omega-2)-hydroxy-LA, were isolated by HPLC and identified by gas chromatography/mass spectrometry. In the presence of NADPH, the formation of all three metabolites was linear with time and microsomal protein concentration. Hydrogen peroxide also supported the microsomal metabolism of LA, although the ratio of metabolites was substantially different than that produced by NADPH-supported microsomes. Several biochemical probes (metyrapone, alpha-naphthoflavone, 2-diethylaminoethyl-2,2-diphenylvalerate hydrochloride, and 10-undecynoic acid) were used to dissociate the three LA hydroxylase activities. These experiments suggest that the site-specific hydroxylation [omega-, (omega-1)-, (omega-2)-] of LA may be catalyzed by different isozymes of cytochrome P-450.

Animals

Biochemical characterization of hepatic microsomal leukotriene B4 hydroxylases.

omega-Hydroxylation of leukotriene B4 (LTB4) has been reported in human and rodent polymorphonuclear leukocytes; preliminary information indicates that this metabolism is cytochrome P-450 dependent. Therefore, these studies were initiated to characterize the cytochrome P-450-dependent metabolism of LTB4 in other tissues. LTB4 was metabolized by rat hepatic microsomes to two products, 20-hydroxy(omega)-LTB4 and 19-hydroxy(omega-1)-LTB4. The formation of these metabolites was both oxygen and NADPH dependent indicating that a monooxygenase(s) was responsible for these reactions. The apparent Km and Vmax for LTB4 omega-hydroxylase were 40.28 microM and 1202 pmol/min/mg of protein, respectively. In contrast, the apparent Km and Vmax for LTB4 (omega-1)-hydroxylase were 61.52 microM and 73.50 pmol/min/mg of protein, respectively. Both LTB4 omega- and (omega-1)-hydroxylases were inhibited by metyrapone in a concentration-dependent fashion. However, SK&F 525A inhibited LTB4 (omega-1)- but not omega-hydroxylase. In contrast, alpha-naphthoflavone decreased LTB4 omega- but not (omega-1)-hydroxylase activities. The differences in the Km apparent for substrate as well as the differential inhibition by inhibitors of cytochrome P-450 suggest that the omega- and (omega-1)-hydroxylations of LTB4 in hepatic microsomes are mediated by different isozymes of P-450. Furthermore, several additional characteristics of LTB4 hydroxylases indicate that these isozymes of P-450 may be different from those which catalyze similar reactions on medium-chain fatty acids, such as laurate and prostaglandins.

Animals

Corticosterone therapy during gestation: effects on the development of rat cerebellum.

Corticosterone (1 microgram/g) was administered to the pregnant rat mother at 17, 18 and 19 days of gestation. The pups were killed at birth or at 6 or 12 days of age and the morphological and biochemical development of the brain, with special emphasis in the cerebellum was studied. The brain and cerebellar weight was slightly diminished in the corticosterone treated animals. Corticosterone produces changes in the pattern of development of the cerebellar layers, causing an accelerated decrement of the external granular layer at 12 days of life. The total protein content of the cerebellum was increased in the hormone treated pups at birth, 6 and 12 postnatal days. Newborn corticosterone treated animals showed a decreased DNA content, but this phenomenon was completely reversed at 12 days. It was concluded that corticosterone given to the pregnant mother influences the time pattern of the development of the brain and the cerebellum.

Animals

Human chorionic gonadotropin binding to rat testis receptors is inhibited by a thymus factor.

An interrelationship between immune and reproductive systems has been postulated, and involves, among others, bidirectional effects between gonads and thymus. To this effect a rat thymus fraction of about 28000 mol wt has been reported to inhibit the effect of hCG on in vitro suspension of Leydig cells. We have investigated the antigonadotropin activity of thymus extracts on rat testis receptors. Acetonic powder obtained from thymus of 14 day-old rats was separated by molecular sieve chromatography. The effect of the collected fractions on the 125I-hCG binding to receptor sites in rat testes was evaluated. A fraction corresponding to 27000-28000 mol wt named thymus factor (TF), was found to inhibit the binding activity of 125I-hCG to its testicular receptor. The inhibitory effect of TF on hCG binding is dose related. By Scatchard analysis a competitive interaction at the receptor level between TF and hCG was demonstrated. The Ka values of hCG binding were diminished in the presence of TF while no significative changes were detected in the number of receptor sites. Present results strongly suggest a modulation function of TF at the testis receptor level.

Animals

Prednisone effects on postnatal brain development of rats following maternal therapy.

16-beta-Metilprednisone was administered in the last week of pregnancy in order to study its effects on postnatal brain development of the litters. Two groups of pregnant inbred rats were used for the experiments. Group I received 1.25 mg/kg/day 16-beta-metilprednisone. Group II served as untreated controls. The litters were killed either within 24 hr after birth, at 17 days, or at 45 days of age. Treated litters (animals whose mothers received the steroid) had a decreased body weight at 17 days of age. Wet brain weight was decreased in the same group. At 17 and 45 days of life treated rats showed a dry brain weight lower than that of controls. Newborn and 17-day-old rats whose mothers received the steroid showed a decreased content of brain proteins. Total lipids, cholesterol, and phospholipids were reduced in the brain of treated rats at 45 days of life. Brain phospholipids were also decreased in 17-day-old rats. These results show that 16-beta-metilprednisone administration to pregnant rats produces long lasting changes in brain chemical composition.

Animals