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At least 1,693 records · Page 94Linked to original sources
PubMed1996

Characterization of a chicken luteinizing hormone receptor (cLH-R) complementary deoxyribonucleic acid, and expression of cLH-R messenger ribonucleic acid in the ovary.

Studies were conducted to characterize the chicken ovarian LH receptor (cLH-R) cDNA and to evaluate expression of cLH-R mRNA in follicles at different stages of development. A total of 1.89 kb of nucleic acid sequence corresponding to the cLH-R (1.79 kb of the predicted coding region) was isolated by a combination of reverse transcription-polymerase chain reaction and cDNA library screening techniques. Also of interest was the finding that two of three positive clones isolated from the hen ovarian cDNA library contained an 86-bp insert located in the extracellular domain within 69 bp of the putative transmembrane domain. This insert contains an inframe TGA stop codon, suggesting that an alternatively spliced transcript results in translation of a truncated protein corresponding to the extracellular domain of the cLH-R. Considering all protein domains thus far characterized, the deduced amino acid sequence of the cLH-R shares 73.2% and 74.2% identity with the rat and porcine LH-R sequences, respectively, with highest homology occurring within the seven transmembrane spanning regions (86-88% identity vs. mammalian sequences). Northern blot analysis determined that cLH-R mRNA levels in the theca layer tend to increase through follicle development to the second largest (F2) preovulatory follicle (p = 0.084), and to decrease in the largest preovulatory (F1) follicle (p < 0.02 vs. F2). By comparison, cLH-R mRNA levels are nondetectable (by Northern blot analysis) in granulosa cells from prehierarchal (3-8-mm diameter) follicles. Constitutive expression of cLH-R mRNA in granulosa cells is first detectable at the 9-12-mm diameter stage of follicle development, and levels are further increased in cells from large preovulatory (F1, F2, and F3) follicles (p < 0.01 vs. 9-12-mm stage). Collectively, these results are consistent with previous observations that granulosa cells from prehierarchal follicles fail to produce cAMP or steroids in response to short-term incubation with ovine LH, in vitro, and that granulosa cells acquire LH responsiveness only subsequent to follicle selection into the rapid growth phase.

Alternative Splicing↗
PubMed1996

Characterization of the chicken follicle-stimulating hormone receptor (cFSH-R) complementary deoxyribonucleic acid, and expression of cFSH-R messenger ribonucleic acid in the ovary.

Studies were conducted to characterize the chicken (c) FSH receptor (R) cDNA, and to evaluate expression of cFSH-R mRNA in the hen ovary at known stages during follicle development. A total of 2.5 kb of nucleic acid sequence including the complete cFSH-R coding region was isolated by a combination of the reverse-transcription polymerase chain reaction and 5'- and 3'-rapid amplification of cDNA ends techniques. Overall, the nucleic acid sequence homology of the cFSH-R cDNA coding region is 71.8% and 72.2% compared to the rat and bovine FSH-R, respectively, while the deduced amino acid sequence identity for the receptor protein (693 amino acids) is 71.9% and 72.4%, respectively. By comparison, the cFSH-R nucleic acid and amino acid sequences are 60.1% and 49.4% identical to the respective cLH-R sequences. Northern blot analysis detected a single 4.3-kb cFSH-R mRNA transcript, which was selectively expressed in ovarian (granulosa, theca, and stromal) tissues, but not the oviduct, adrenal, liver, muscle, or brain. As the follicle developed from the prehierarchical (6- to 8-mm diameter) to the largest preovulatory (F1 follicle) stage, cFSH-R mRNA levels progressively declined within both the granulosa and theca layers (p < 0.05). Moreover, cFSH-R mRNA levels were lower in whole atretic than in morphologically normal 3- to 5-mm follicles (p = 0.0015). The pattern of cFSH-R mRNA expression within the granulosa layer during follicle development was notably different from that of the recently reported cLH-R, in that cLH-R mRNA levels increase to become readily detectable coincident with dramatically increased steroidogenic capacity during the last few days before ovulation of the follicle. On the other hand, highest levels of cFSH-R mRNA in 6- to 8-mm (prehierarchical) follicles were consistent with a role for the cFSH-R in maintaining the viability of prehierarchical follicles and in initiating granulosa cell differentiation at the time when follicles are selected into the preovulatory hierarchy.

Amino Acid Sequence↗
PubMed1975

Ribonucleic acid destruction and synthesis during intraperiplasmic growth of Bdellovibrio bacteriovorus.

During growth of Bdellovibrio bacteriovorus on (2-14C)uracil-labeled Escherichia coli approximately 50% of the radioactivity is incorporated by the bdellovibrio and most of the remainder is released as free nucleic acid bases. Kinetic studies showed that 50 and 30S ribosomal particles and 23 and 16S ribosomal ribonucleic acid (RNA) of E. coli are almost completely degraded by the first 90 min in a 210- to 240-min bdellovibrio developmental cycle. Synthesis of bdellovibrio ribosomal RNA was first detected after 90 min. The specific activity and the ratio of radioactivity in the bases of the synthesized bdellovibrio RNA was essentially the same as those of the substrate E. coli. The total radioactivity of the bdellovibrio deoxyribonucleic acid (DNA) exceeded that in the DNA of the substrate E. coli cell, and the ratio of radioactivity of cytosine to thymine residues differed. Intraperiplasmic growth of B. bacteriovorus in the presence of added nucleoside monophosphates (singly or in combination) significantly decreased the uptake of radioactivity from (2-14C)uracil-labeled E. coli; nucleosides or nucleic acid bases did not. It is concluded that the RNA of the substrate cell, in the form of nucleoside monophosphates, is the major or exclusive precursor of the bdellovirbrio RNA and also serves as a precursor for some of the bdellovibrio DNA.

Bacteria↗
PubMed1972

Covalent linkage between ribonucleic Acid primer and deoxyribonucleic Acid product of the avian myeloblastosis virus deoxyribonucleic Acid polymerase.

Initiation of deoxyribonucleic acid (DNA) synthesis by the avian myeloblastosis virus DNA polymerase was previously suggested to involve a ribonucleic acid (RNA) primer, the initial product being a DNA molecule joined by a phosphodiester bond to the RNA primer. The existence and nature of such an RNA-DNA joint was investigated by assaying for transfer of a (32)P atom from an alpha-(32)P-deoxyribonucleotide to a 2'(3')-ribonucleotide after alkaline hydrolysis of the polymerase product. Such a transfer was observed, but only from alpha-(32)P-deoxyadenosine triphosphate and only to 2'(3')-adenosine monophosphate. This same transfer was observed in both the endogenous DNA polymerase reaction of purified virions and the reconstructed reaction of purified DNA polymerase plus purified 60 to 70S viral RNA. These results indicate a high level of specificity for the initiation process and support the idea of a low-molecular-weight initiator RNA as part of the 60 to 70S RNA complex.

Journal Article↗
PubMed1985

Modulation of procollagen gene expression by retinoids. Inhibition of collagen production by retinoic acid accompanied by reduced type I procollagen messenger ribonucleic acid levels in human skin fibroblast cultures.

Recent clinical observations have suggested that retinoids, which are in frequent use in dermatology, can affect the connective tissue metabolism in skin and other tissues. In this study, we examined the effects of several retinoids on the metabolism of collagen by human skin fibroblasts in culture. Incubation of cultured fibroblasts with all-trans-retinoic acid or 13-cis-retinoic acid, in 10(-5) M or higher concentrations, markedly reduced the procollagen production, as measured by synthesis of radioactive hydroxyproline. The effect was selective in that little, if any, inhibition was noted in the incorporation of [3H]leucine into the noncollagenous proteins, when the cells were incubated with the retinoids in 10(-5) M concentration. Similar reduction in procollagen production was noted with retinol and retinal, whereas an aromatic analogue of retinoic acid ethyl ester (RO-10-9359) resulted in a slight increase in procollagen production in these cultures. The reduction in procollagen production by all-trans-retinoic acid was accompanied by a similar reduction in pro alpha 2(I) of type I procollagen specific messenger RNA (mRNA), as detected by dot blot and Northern blot hybridizations. Hybridizations with human fibronectin and beta-actin specific DNA probes indicated that the levels of the corresponding mRNAs were not affected by the retinoids, further suggesting selectivity in the inhibition of procollagen gene expression. Further control experiments indicated that all-trans-retinoic acid, under the culture conditions employed, did not affect the posttranslational hydroxylation of prolyl residues, the mannosylation of newly synthesized procollagen, the specific radioactivity of the intracellular prolyltransfer RNA pool, or DNA replication. All-trans-retinoic acid also elicited a reduction in trypsin-activatable collagenase, but not in the activity of prolyl hydroxylase or an elastaselike neutral protease in the fibroblast cultures. Incubation of three fibroblast lines established from human keloids with all-trans-retinoic acid or 13-cis-retinoic acid also resulted in a marked reduction in procollagen production. The results, therefore, suggest that further development of retinoids might provide a novel means of modulating collagen gene expression in patients with various diseases affecting the connective tissues.

Cells, Cultured↗
PubMed2004

Valproic acid inhibits leptin secretion and reduces leptin messenger ribonucleic acid levels in adipocytes.

Treatment of epilepsy or bipolar disorder with valproic acid (VPA) induces weight gain and increased serum levels for the satiety hormone, leptin, through an unidentified mechanism. In this study we tested the effects of VPA, a short-chain branched fatty acid (C8:0), on leptin biology and fatty acid metabolism in 3T3-L1 adipocytes. VPA significantly reduced leptin secretion in a dose-dependent manner. Because fatty acid accumulation has been hypothesized to block leptin secretion, we tested the effect of VPA on fatty acid metabolism. Using 14C-radiolabeled VPA, we found that the 14C was mainly incorporated into triacylglycerol. VPA did not alter lipogenesis from acetate, nor did it change the amount of intracellular free fatty acids available for triacylglycerol synthesis. Decreased leptin secretion was accompanied by a reduction in leptin mRNA, even though VPA treatment did not alter the protein levels for known transcription factors affecting leptin transcription including: CCAAT/enhancer binding protein-alpha, peroxisome proliferator-activated receptor-gamma, or steroid regulatory element binding protein 1a. VPA altered levels of leptin mRNA independent of de novo protein synthesis without affecting leptin mRNA degradation. This report demonstrates that VPA decreases leptin secretion and mRNA levels in adipocytes in vitro, suggesting that VPA therapy may be associated with altered leptin homeostasis contributing to weight gain in vivo.

3T3-L1 Cells↗
PubMed1989

Retinoic acid selectively stimulates growth hormone secretion and messenger ribonucleic acid levels in rat pituitary cells.

Hydrocortisone (HCT) is known to stimulate rat GH gene expression. As the retinoic acid receptor cDNA bears a 15% homology to the hydrocortisone receptor cDNA, the effects of retinoic acid were tested in GH3 rat pituitary cells. Retinoic acid selectively stimulated GH secretion after 24 h. Maximal induction of basal GH secretion (220% of controls) was achieved with 1 microM retinoic acid after 72 h. HCT (50 nM) alone stimulated GH secretion 3.5-fold during 72-h incubation, and retinoic acid (1 microM) with HCT caused a further 7-fold increase in GH secretion. The stimulating effects of retinoic acid on GH were observed when cells were incubated in defined serum-free medium, 4% fetal calf serum, or Dulbecco's Modified Eagle's Medium containing HCT. Northern gel hybridization showed that steady state levels of GH mRNA transcripts in these cells were also selectively induced by retinoic acid. PRL secretion and mRNA levels, however, were not appreciably altered by retinoic acid. The results show that retinoic acid selectively stimulates basal and HCT-induced GH secretion and mRNA levels in these cells in a dose- and time-dependent manner. This selective action of retinoic acid appears to occur independently of the presence of serum in the culture medium.

Animals↗
PubMed1992

Cloning of an ovine 11 beta-hydroxysteroid dehydrogenase complementary deoxyribonucleic acid: tissue and temporal distribution of its messenger ribonucleic acid during fetal and neonatal development.

Glucocorticoids promote the development of many organ systems vital for extrauterine survival, and fetal cortisol provides the trigger for birth in sheep. The activity of glucocorticoids may be influenced at a cellular level by 11 beta-hydroxysteroid dehydrogenase (11 beta-HSD), which is responsible for the interconversion of cortisol and cortisone. To examine 11 beta-HSD gene expression during fetal development, two overlapping clones which yield a 1.4 kilobase (kb) complementary DNA encoding sheep 11 beta-HSD from a liver library were isolated by using a rat 11 beta-HSD cDNA as the probe. This cDNA contains a 879 base pair open reading frame for a protein of 292 amino acids that has more than 70% sequence identity to rat and human 11 beta-HSDs. To define the tissue distribution of 11 beta-HSD messenger RNA in sheep, selected tissues were collected from one fetus at day 130 and term (approximately 145 days), and from a nonpregnant ewe. Cellular RNA was extracted and subjected to Northern blot analysis, and a single 1.8 kb transcript was detected in the fetal and adult liver, lung, hypothalamus, anterior pituitary, and placenta. This was undetectable in adrenals and kidneys, but a smaller (1.5 kb) transcript was present in fetal and adult kidney RNA. The relative abundance of 11 beta-HSD mRNA was greatest in fetal and adult livers, and it was much higher in adult liver, lung, and kidney than in the corresponding fetal tissues. To examine whether 11 beta-HSD gene expression is developmentally regulated in the fetal sheep, liver, lung, and kidney tissues were taken from fetuses at day 60-70, day 100-110, day 125-130, at term, and from newborn lambs (24-48 h old). In the lung and kidney, the relative abundance of 11 beta-HSD mRNA did not change from day 60 to term but increased in the lungs of newborn lambs. In contrast, 11 beta-HSD mRNA levels in the liver increased between day 125 and term and rose further in the newborn. Collectively, these results demonstrate that 11 beta-HSD gene expression in sheep is regulated in a tissue-specific and developmentally programmed manner.

11-beta-Hydroxysteroid Dehydrogenases↗
PubMed2006

Androgen receptor immunostaining and androgen receptor messenger ribonucleic acid expression are increased in cremaster muscles associated with undescended testis.

OBJECTIVES: To determine and compare androgen receptor (AR) immunostaining and AR messenger ribonucleic acid (mRNA) expression in cremaster muscles associated with descended or undescended testis. METHODS: Eight boys with descended testis but with inguinal hernia and 8 boys with undescended testis were evaluated. Serum follicle-stimulating hormone (FSH), luteinizing hormone (LH), testosterone, and free testosterone levels were determined, and samples of cremaster muscles were immunostained for AR. Groups were compared by unpaired t tests and Fisher's exact tests; P values of <0.05 were considered significant. Samples of cremaster muscles were obtained from another 5 boys with descended testis but with inguinal hernia and 5 boys with undescended testis. The expression of AR mRNA in those samples was determined by semiquantitative reverse transcriptase polymerase chain reaction. RESULTS: Serum FSH, LH, testosterone, and free testosterone levels were similar among groups. None of the samples from boys with descended testis showed positive staining, but 4 of 8 samples from boys with undescended testis stained positive for AR. Androgen receptor mRNA transcript levels were approximately 10 times lower in cremaster muscles of boys with descended testis compared with those in boys with undescended testis. CONCLUSIONS: Despite similar serum hormone levels, more AR expression in cremaster muscles associated with undescended testis might represent evidence of being subjected to a lesser degree of androgenic effects.

Child, Preschool↗