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J L Roberts

Publications and source records attributed to J L Roberts.

At least 181 records · Page 10Linked to original sources
PubMed1987

Dopamine regulates expression of the glandular-type kallikrein gene at the transcriptional level in the pituitary.

A glandular-like kallikrein enzyme, a member of a well-characterized family of specific arginyl endopeptidases that may be involved in prohormone processing, has previously been shown to be present in the anterior and neurointermediate lobes of the rat pituitary. We isolated glandular-like kallikrein cDNAs from cDNA libraries prepared from these two tissues. By nucleotide sequence, restriction endonuclease, solution hybridization/nuclease protection, and blot analyses, we showed that, of the 8-10 rat kallikrein-encoding genes, it is the true glandular kallikrein mRNA that is expressed in both pituitary lobes. RNA blot-hybridization analysis of anterior and neurointermediate lobe pituitary RNA revealed a kallikrein mRNA of approximately equal to 900 base pairs. As analyzed by blot-hybridization and solution hybridization/nuclease protection analyses, the true glandular kallikrein mRNA was present at low levels: approximately equal to 0.05% of total mRNA in both male and female neurointermediate lobes. Similar low levels of the glandular kallikrein mRNA were found in the male anterior lobe, whereas the levels were 10- to 15-fold higher in the female anterior lobe. In vivo administration of a dopamine agonist (bromocryptine) or antagonist (haloperidol) caused a decrease or increase, respectively, in the amount of true glandular kallikrein mRNA in the neurointermediate lobe of both sexes that closely paralleled changes in proopiomelanocortin mRNA levels. Bromocryptine decreased and haloperidol increased true glandular kallikrein mRNA levels in the female anterior lobe but had no effect in the male anterior lobe. Nuclear transcription run-on studies showed that the changes in mRNA were due, at least in part, to parallel effects of haloperidol on kallikrein gene transcription. Thus, these studies have demonstrated that the pituitary expresses the glandular-type member of the kallikrein gene family and that dopaminergic compounds elicit changes in kallikrein mRNA, at least in part, by modulating transcription. In the intermediate lobe, regulation of true glandular kallikrein gene expression is parallel to that of proopiomelanocortin gene expression, suggesting that the enzyme may play a physiological role in the production and/or secretion of the proopiomelanocortin peptides in this tissue.

Animals↗
PubMed1987

Complex transcriptional regulation by glucocorticoids and corticotropin-releasing hormone of proopiomelanocortin gene expression in rat pituitary cultures.

Proopiomelanocortin (POMC) peptide secretion from rat anterior pituitary corticotrophs and intermediate pituitary melanotrophs is stimulated by corticotropin-releasing hormone (CRH). CRH-stimulated secretion in the corticotrophs is inhibited by glucocorticoids in a complex fashion, involving both a fast, direct blockade of POMC secretion (minutes to hours) and a longer inhibitory action (hours to days) that decreases the amount of POMC peptide available for release. The current studies tested the ability of CRH to stimulate beta-endorphin (a peptide derived from POMC) secretion and POMC gene transcription in cultured anterior and neurointermediate lobe pituitary cells, and examined interactions between CRH and glucocorticoids in regulating POMC gene expression using an in vitro nuclear transcription run-on assay. In both tissues, CRH elicited a time-dependent stimulation of POMC gene transcription that was maximal at 60 min and remained elevated for at least 18 hr. Glucocorticoids rapidly inhibited POMC gene transcription fourfold in the anterior lobe with maximal effects within 20 min. Glucocorticoids also blocked CRH-stimulated POMC gene transcription in anterior pituitary cultures in a temporal manner paralleling their inhibitory effects on CRH-stimulated beta-endorphin secretion. In neurointermediate lobe cultures, the effects of glucocorticoids and CRH on POMC gene transcription were qualitatively similar to, but of lesser magnitude than those observed in the anterior lobe. These studies indicate that the regulation of POMC gene transcription by glucocorticoids and CRH is complex and that the two modulators do not function independently.

Animals↗
PubMed1987

Primary adrenocortical carcinoma: sonographic evaluation with clinical and pathologic correlation in 26 patients.

The sonograms of 26 patients (19 adults and seven children) with pathologically proven diagnoses of primary adrenocortical carcinoma were evaluated. Clinical corroboration was obtained in all cases. The size of the lesions ranged from 3 to 22 cm. The five smaller lesions (3-6 cm) showed a homogeneous echo pattern, similar to renal cortical echogenicity. The 21 larger lesions varied in echo texture, having a heterogeneous appearance with focal or scattered echopenic or echogenic zones representing areas of tumor necrosis, hemorrhage, and/or, rarely (19%), calcification. Even the largest lesions were fairly well delineated, often with a lobulated border. Few (7/26 or 27%) showed a surrounding echogenic thin capsulelike rim. All five small lesions showed clinical evidence of endocrine activity. Larger lesions were hormonally active less often (9/21 or 43%). Twelve patients (46%) showed no sign of endocrine activity and presented with symptoms such as fever, weight loss, abdominal discomfort, abdominal mass, hematuria, and hypertension. In the pediatric and adolescent age group (0-16 years), all tumors were hormonally active, while only seven (37%) of tumors in the adult population (17-69 years) were hormonally active. Unfortunately no echo pattern was characteristic enough to allow differentiation of adrenal adenoma from carcinoma. Smaller lesions are more likely to be benign, and larger lesions with areas of necrosis, hemorrhage, and calcification are more likely to be malignant.

Adolescent↗
PubMed1986

Regulation of pro-opiomelanocortin gene transcription in individual cell nuclei.

A nonrepetitive complementary RNA probe specific for an intervening sequence of the rat pro-opiomelanocortin (POMC) gene primary transcript was used to analyze the hormonal regulation of POMC gene transcription in individual cell nuclei in the rat pituitary by in situ hybridization. This probe recognized only full-length POMC heterogeneous nuclear RNA, as verified by Northern blots of pituitary RNA. When pituitary sections were hybridized with this 3H-labeled POMC intron A probe, silver grains were predominantly localized over the nuclei of cells that expressed POMC in the anterior and intermediate lobes. Adrenalectomy increased both the average grain density over corticotroph nuclei and the number of cells in the anterior pituitary with significant numbers of silver grains over their nucleus. Dexamethasone administration to intact or adrenalectomized rats results in the rapid (within 30 minutes) disappearance of silver grains over the nuclei of corticotrophs in the anterior lobe, suggesting that POMC gene transcription had been inhibited. However, adrenalectomy or dexamethasone administration did not alter the silver grain density over nuclei of intermediate lobe melanotrophs. Thus, this in situ hybridization assay utilizing an intervening sequence-specific POMC probe can measure rapid physiological changes in POMC heterogeneous nuclear RNA in individual cell nuclei.

Animals↗
PubMed1986

Localization of proopiomelanocortin mRNA in functional subsets of neurons defined by their axonal projections.

In situ cDNA:mRNA hybridization is a technique that has been developed for the visualization of cDNA:mRNA hybrids in individual cells. To use this technique to answer questions of regulation in heterogeneous populations of cells in the brain, it must be combined with other procedures allowing for the identification of functional subgroups of neurons. We report here a procedure by which in situ cDNA:mRNA hybridization may be combined with retrograde axonal tracing using the fluorescent tracer fast blue. Using this technique, it now becomes possible to measure mRNA regulation in functional subsets of cells defined by their axonal projections.

Animals↗
PubMed1986

Brown adipose tissue triacylglycerol fatty acids of obese and lean mice: in situ and in transplants.

The triacylglycerols of white adipose tissue (WAT) from animals with high rates of lipogenesis, such as obese hyperglycemic mice or hypothalamically lesioned rats, contain high proportions of palmitoleic acid (16:1) and low proportions of linoleic acid (18:2). These differences appear to result from dilution of dietary 18:2 by synthesized fatty acids, particularly 16:1. To test this we have investigated the triacylglycerol fatty acid composition of brown and white adipose tissue of lean and obese mice, as brown adipose tissue (BAT) has a higher lipogenic rate than WAT and lipogenesis is faster in obese than in lean mice. Between three and eight weeks of age the proportions of fatty acids in the tissues changed, with a marked fall in milk-derived lauric and myristic acids. From 8 to 16 weeks they were more stable and the proportions of 16:1 and 18:2 in the different tissues were as expected, with the highest and lowest proportions, respectively, in BAT from obese mice. When BAT from obese mice was transplanted under the kidney capsule of lean mice, or vice versa, for one month, the fatty acid composition of the grafts changed toward that of the host BAT. The proportions of 18:2 and, to a lesser extent, 16:1 were slightly higher in the grafts than in the hosts but since this also occurred in lean-to-lean and obese-to-obese grafts it was probably a transplantation artifact. Overall, the results confirm that the physiological environment, rather than the source of the adipose tissue, is the major determinant of its fatty acid composition.

Adipose Tissue↗
PubMed1986

In situ cDNA:mRNA hybridization: development of a technique to measure mRNA levels in individual cells.

In this article we have described our protocol for in situ cDNA:mRNA hybridization and a variety of methodological considerations we have entertained to optimize the procedure. It should be stressed that each different system will have its own special characteristics and require optimization to obtain maximal and reproducible signals. We believe that by following a methodological logic as outlined here, researchers should be able to establish the in situ cDNA:mRNA hybridization protocol with their probes and tissue systems.

Animals↗
PubMed1986

Lipomatous tumors of the liver: evaluation with CT and US.

Seven cases of lipomatous masses within the liver parenchyma were demonstrated with computed tomography (CT). Five of these cases were obtained from a retrospective review of 50 cases of renal angiomyolipoma in which the liver was adequately demonstrated. The other two cases were from the files of the Armed Forces Institute of Pathology and had no associated renal lesions. Three of the five cases were associated with tuberous sclerosis. In all seven cases, the fatty tumors appeared on CT scans as a well-defined, 0.8-13-cm mass, with attenuation coefficients of less than -30 HU. On ultrasound studies, the lesions were well circumscribed, highly echogenic, and similar to hemangiomas. While distinctly rare lesions, these lipomatous masses are not as unusual as the literature would indicate. One may anticipate such masses in patients with renal angiomyolipomas and in a relatively high percentage of those with tuberous sclerosis.

Adult↗
PubMed1986

Control of respiratory activity of the genioglossus muscle in micrognathic infants.

The genioglossus (GG) muscle activity of four infants with micrognathia and obstructive sleep apnea was recorded to assess the role of this tongue muscle in upper airway maintenance. Respiratory air flow, esophageal pressure, and intramuscular GG electromyograms (EMG) were recorded during wakefulness and sleep. Both tonic and phasic inspiratory GG-EMG activity was recorded in each of the infants. On occasion, no phasic GG activity could be recorded; these silent periods were unassociated with respiratory embarrassment. GG activity increased during sigh breaths. GG activity also increased when the infants spontaneously changed from oral to nasal breathing and, in two infants, with neck flexion associated with complete upper airway obstruction, suggesting that GG-EMG activity is influenced by sudden changes in upper airway resistance. During sleep, the GG-EMG activity significantly increased with 5% CO2 breathing (P less than or equal to 0.001). With nasal airway occlusion during sleep, the GG-EMG activity increased with the first occluded breath and progressively increased during the subsequent occluded breaths, indicating mechanoreceptor and suggesting chemoreceptor modulation. During nasal occlusion trials, there was a progressive increase in phasic inspiratory activity of the GG-EMG that was greater than that of the diaphragm activity (as reflected by esophageal pressure excursions). When pharyngeal airway closure occurred during a nasal occlusion trial, the negative pressure at which the pharyngeal airway closed (upper airway closing pressure) correlated with the GG-EMG activity at the time of closure, suggesting that the GG muscle contributes to maintaining pharyngeal airway patency in the micrognathic infant.

Cleft Palate↗
PubMed1986

A diurnal rhythm in proopiomelanocortin messenger ribonucleic acid that varies concomitantly with the content and secretion of beta-endorphin in the intermediate lobe of the rat pituitary.

The present study investigated whether diurnal variations in the secretion of alpha MSH and beta-endorphin from the intermediate lobe (IL) of the pituitary are associated with parallel changes in the synthesis of mRNA specifically encoding proopiomelanocortin (POMC). The results demonstrate that concomitant diurnal variations occur in both plasma and IL concentrations of immunoreactive beta-endorphin and alpha MSH. Plasma and IL peptide levels were relatively constant during daylight hours (0600-1800 h), but increased after the onset of darkness and reached maximal concentrations at 0200 h. To examine the possibility that this diurnal rhythm in the content and secretion of POMC-derived peptides resulted from diurnal changes in the biosynthesis of POMC, the concentration and rate of synthesis of POMC mRNA were examined. POMC mRNA levels were elevated during the dark period, reaching a maximum level at 0200 h that was 2-fold higher than that occurring during the light period. POMC mRNA synthesis, determined by measuring the number of RNA polymerase II complexes transcribing the POMC gene in isolated cell nuclei, also varied diurnally, with maximum transcription rates occurring at 1800 h, thus preceding maximal increases in POMC mRNA content and POMC peptide secretion by 8 h. Together, these data indicate that diurnal variations in the content and secretion of POMC-derived peptides are associated with parallel changes in POMC mRNA concentrations and are preceded by similar changes in POMC gene transcription.

Animals↗
PubMed1986

Coordinate regulation of peptide acetyltransferase activity and proopiomelanocortin gene expression in the intermediate lobe of the rat pituitary.

Proopiomelanocortin (POMC) is posttranslationally processed in the intermediate lobe of the pituitary to both N-terminally acetylated and nonacetylated forms of alpha MSH and beta-endorphin (beta END). N-Acetylation substantially modifies the physiological responses produced by both peptides, suggesting that the activity of the peptide acetyltransferase, which posttranslationally acetylates beta END and des-acetyl-alpha MSH, may play an important role in defining the biological activity of the secretory products of the intermediate pituitary lobe. The present results demonstrate that peptide acetyltransferase activity is induced by treating rats chronically with the dopamine receptor antagonist haloperidol. Haloperidol administration produced parallel and essentially equivalent increases in acetyltransferase activity, POMC mRNA levels, and the content and secretion of POMC-derived peptides in the neurointermediate pituitary. Time-course and dose-response studies further demonstrated that acetyltransferase activity covaried with POMC mRNA and peptide levels. Chronic treatment with the dopamine receptor agonist bromocriptine had the opposite effects; it lowered acetyltransferase activity, POMC mRNA levels, and alpha MSH and beta END immunoreactivities. Subcellular fractionation showed that acetyltransferase activity was localized in three subcellular compartments corresponding in density to secretory vesicles, rough endoplasmic reticulum and Golgi apparatus, and cytosol. Haloperidol treatment significantly increased the specific activity of the secretory vesicle-associated acetyltransferase without affecting the specific activity of the enzymes present in the endoplasmic reticulum or cytosol. Together, these data indicate that peptide acetyltransferase activity and POMC biosynthesis are coregulated.

Acetylation↗
PubMed1986

The regulation of granulosa cell proopiomelanocortin messenger ribonucleic acid by androgens and gonadotropins.

We have previously demonstrated the presence of proopiomelanocortin (POMC) messenger RNA (mRNA) in rat granulosa cells. This study examines the unique, tissue-specific regulation of granulosa cell POMC mRNA levels by hormones with established regulatory effects on these cells. Northern blot analysis of immature rat ovarian RNA revealed the presence of a single mRNA of approximately 900 base pairs in length. The levels of ovarian POMC mRNA increased approximately 6-fold 48 h after priming with PMSG when expressed per microgram of total RNA. Granulosa cells were isolated from immature PMSG-primed rats and cultured under serum-free conditions in the presence and absence of various hormones to determine their effect on POMC mRNA levels. Treatment of the cells for 48 h with LH (100 ng/ml), androstenedione (10(-7) M), or LH plus androstenedione elevated POMC mRNA levels. LH alone elicited a 5-fold increase in POMC mRNA, and androstenedione elicited a 10-fold increase; the combination treatment led to a 47-fold increase. The effects of the nonaromatizable androgen dihydrotestosterone (DHT) were also evaluated. Treatment for 48 h with DHT (10(-7) M) elicited a 40-fold increase in POMC mRNA levels. In vivo experiments measuring ovarian POMC mRNA from immature female rats corroborated the in vitro results. Animals injected sc with DHT (500 micrograms) demonstrated 5-fold increases in ovarian POMC mRNA when expressed per microgram of total RNA. These studies provide both in vitro and in vivo evidence that granulosa cell POMC mRNA is under unique hormonal regulation by both androgens and gonadotropins.

Androgens↗
PubMed1986

Prolactin messenger ribonucleic acid levels in the normal and hypogonadal mouse pituitary gland.

The effects of estrogen on the synthesis of pituitary PRL mRNA and PRL and the release of PRL into plasma were investigated in male and female normal and hypogonadal (hpg) mice. Because the hpg mouse is totally deficient in hypothalamic GnRH, it provides an excellent model for investigating the effects of estrogen and GnRH on PRL synthesis and release in the presence of a totally inactive gonadotropin-gonadal system. Estrogen stimulated the synthesis and release of pituitary PRL in both sexes of normal and hpg mice, and this marked increase in PRL synthesis correlated with an equally dramatic increase in pituitary PRL mRNA levels. Administration of GnRH alone produced a slight but significant increase in pituitary PRL content, which is consistent with an action of GnRH on prolactotropes, either directly or by way of a paracrine action involving gonadotropes.

Animals↗
PubMed1986

Characterization of proopiomelanocortin mRNA detected by in situ hybridization.

In situ hybridization is a method for detecting specific nucleotide sequences by using a labeled complementary nucleic acid probe. We have characterized in situ hybridization in rat pituitary using a cDNA probe directed against a portion of proopiomelanocortin (POMC) mRNA to show its specificity in tissue sections. This characterization includes the size of the target mRNA from tissue processed for in situ hybridization, the dissociation pattern of the cDNA:mRNA hybrid, and quantitation of changes in POMC mRNA levels in the rat pituitary intermediate lobe detected both by in situ hybridization and "dot blot" techniques.

Animals↗
PubMed1985

Reactivity and activation of dioxygen-derived species in aprotic media (a model matrix for biomembranes).

In aprotic media the electrochemical reduction of dioxygen yields superoxide ion (O2-), which is an effective Brønsted base, nucleophile, one-electron reductant, and one-electron oxidant of reduced transition metal ions. With electrophilic substrates (organic halides and carbonyl carbons) O2- displaces a leaving group to form a peroxy radical (ROO.) in the primary process. Superoxide ion oxidizes the activated hydrogen atoms of ascorbic acid, catechols, hydrophenazines and hydroflavins. Combination of O2- with 1,2-diphenylhydrazine yields the anion radical of azobenzene, which reacts with O2 to give azobenzene and O2- (an example of O2--induced autoxidation). With phenylhydrazine, O2- produces phenyl radicals. The in situ formation of HO2. (O2- plus a proton source) results in H-atom abstraction from allylic and other groups with weak heteroatom--H bonds (binding energy (b.e.) less than 335 kJ). This is a competitive process with the facile second-order disproportionation of HO2. to H2O2 and O2 (kbi approximately equal to 10(4) mol-1 s-1 in Me2SO). Addition of [FeII(MeCN)4] (ClO4)2 to solutions of hydrogen peroxide in dry acetonitrile catalyses a rapid disproportionation of H2O2 via the initial formation of an adduct [FeII(H2O2)2+----Fe(O)(H2O)2+], which oxidizes a second H2O2 to oxygen. In the presence of organic substrates such as 1,4-cyclohexadiene, 1,2-diphenylhydrazine, catechols and thiols the FeII-H2O2/MeCN system yields dehydrogenated products; with alcohols, aldehydes, methylstyrene, thioethers, sulphoxides, and phosphines, the FeII(H2O2)2+ adduct promotes their monoxygenation. The product from the FeO2+-H2O2 reaction, [FeII(H2O2)22+], exhibits chemistry that is closely similar to that for singlet oxygen (1O2), which has been confirmed by the stoichiometric dioxygenation of diphenylisobenzofuran, 9,10-diphenylanthracene, rubrene and electron-rich unsaturated carbon-carbon bonds (Ph2C = CPh2, PhC = CPh and cis-PhCH = CHPh). In dry ligand-free acetonitrile (MeCN), anhydrous ferric chloride (FeIIICl3) activates hydrogen peroxide for the efficient epoxidation of alkenes. The FeIIICl3 further catalyses the dimerization of the resulting epoxides to dioxanes. These observations indicate that strong Lewis acids that are coordinatively unsaturated, [FeII(MeCN)4]2+ and [FeIIICl3], activate H2O2 to form an effective oxygenation and dehydrogenation agent.(ABSTRACT TRUNCATED AT 400 WORDS)

Cell Membrane↗
PubMed1985

mRNA sequence predictions from homologous protein sequences.

A model has been developed that permits the prediction of mRNA nucleic acid sequence from the sequences of the translated proteins. The model relies on the information obtained from the comparison of protein sequences in related species to reduce the number of possible codons for those amino acids where mutations are observed. The predictions so obtained have been tested by applying the model to proteins whose mRNA sequences are known. The model's predictions have been found to be 100% accurate if three or more different amino acids are known at a given position and if the protein sequences are restricted to relatively closely related species (within the same class). The use of this model may permit a reduction of the mRNA sequence degeneracy and therefore be helpful in the synthesis of cDNA probes or for the prediction of restriction endonuclease sites. Computer programs have been developed to ease the use of the model.

Amino Acid Sequence↗
PubMed1985

Ocular immune reactants in patients dying with systemic lupus erythematosus.

Immunopathologic studies were performed on ocular tissue obtained at autopsy in five patients with systemic lupus erythematosus (SLE). The immunopathologic findings were correlated with histopathologic and clinical features of SLE. Immune reactants were demonstrated by direct immunofluorescence in all patients in a granular pattern suggesting immune complex aggregates; control ocular tissues from four patients without SLE were negative. Ultrastructural studies demonstrated corresponding electron-dense aggregates in a majority of samples. Histologic and gross anatomic evidence of inflammation were generally more focal and less frequent than the distribution of immune reactants. Most of the immune deposits were localized in the walls of blood vessels of the conjunctiva, ciliary body, retina, choroid, and sclera. Diffuse deposits were also found in association with basement membranes in ciliary body (two of five patients) and along the epithelial basement membrane in the cornea (two of five). Immune deposits in peripheral nerves of the ciliary body and conjunctiva in one patient with visual symptoms contained immunoglobulins A and E. The most intense and widespread ocular immune deposits were observed in patients with persistently increased serologic and clinical activity of their systemic disease. These results suggest a role for immune complex localization in the pathogenesis of the ocular lesions of SLE.

Adult↗