Biomedical subjects

J L Juengel

Publications and source records attributed to J L Juengel.

At least 19 recordsLinked to original sources

PubMed2003

Expression of the FcRn receptor (alpha and beta) gene homologues in the intestine of suckling brushtail possum (Trichosurus vulpecula) pouch young.

The neonatal IgG transporter FcRn consists of two chains, FcRn alpha and beta (also known as beta(2) microglobulin), and is involved in transferring IgG molecules across both mammary and intestinal epithelial cells. Developmental changes in FcRn IgG alpha and beta chain mRNA levels were investigated in the gut of brushtail possum (Trichosurus vulpecula) pouch young (PY) using Northern hybridisation. FcRn alpha transcripts were detected in the PY proximal intestine at all times examined, between days 1 and 195 of post-natal life, with increased levels detected from around day 110. The beta(2) microglobulin transcript levels in the PY proximal intestine were low to undetectable until day 110 of post-natal life and then increased dramatically after day 159. Both the FcRn alpha and beta gene transcripts were detected in a wide range of tissues in the adult possum (>365 days). Genomic sequences located 5' to the start of transcription of the FcRn alpha and beta(2) microglobulin genes were cloned and analysed for predicted cis-acting transcription control elements. Both the FcRn alpha and beta(2) microglobulin genomic sequences contained STAT5 binding motifs consistent with the transcription of both genes being modulated by prolactin. Using in situ hybridisation, the FcRn alpha and beta(2) microglobulin transcripts were localised to the epithelial cells of the PY intestine. However, no prolactin receptor transcripts were detected in the same epithelial cells suggesting that the observed changes in FcRn alpha and beta(2) microglobulin gene expression in the proximal intestine are not modulated directly by prolactin. The results are consistent with the hypothesis that changes in FcRn alpha and beta(2) microglobulin gene expression take place in the possum PY intestine to accommodate changes in maternal milk composition to meet the changing immunological demands of the PY.

Animals↗

PubMed2002

Bmp15 mutations and ovarian function.

BMP15, also known as growth and differentiation factor 9B (GDF9B), is a member of the transforming growth factor beta superfamily (TGFbeta) which in humans, rodents and sheep is expressed exclusively in the oocyte. BMP15 is closely related to GDF9, another oocyte-specific member of this superfamily which has been shown to be essential for early ovarian folliculogenesis. Inactivation of the BMP15 gene in mice has shown only minor effects on fertility. However, Inverdale and Hanna lines of sheep carry naturally occurring mutations in BMP15 which highlight differences in the action of this gene between mice and other mammals. Sheep which are heterozygous show an increase in ovulation rate whereas homozygotes are infertile. The granulosa cell receptor which mediates the BMP15 response has not yet been identified, but the discovery that a point mutation in the BMP1B receptor in Booroola sheep is responsible for increased ovulation rate highlights the importance of the TGFbeta signalling molecules in early folliculogenesis.

Animals↗

PubMed2001

Genetic mutations influencing ovulation rate in sheep.

Ovulation rate in mammals is determined by a complex exchange of endocrine signals between the pituitary gland and the ovary, and by paracrine signals within ovarian follicles between the oocyte and its adjacent somatic cells. One approach to identifying factors regulating ovulation rate is to find mutations that influence the target phenotype and, in this context, sheep are proving to be remarkable experimental models. Recently, in three sheep families, namely Inverdale, Hanna and Booroola, the inherited mutation was mapped to a specific region of the sheep X chromosome (Inverdale, Hanna) or sheep chromosome 6 (Booroola) and in each, a point mutation was identified in genes from the bone morphogenetic protein (BMP) relatives of the transforming growth factor beta superfamily or their receptors. In Inverdale (I) and Hanna (H) sheep, separate point mutations were identified in the BMP15 gene corresponding to sites in the mature peptide coding region of the BMP15 growth factor (also known as growth differentiation factor 9B; GDF9B). Expression of the BMP15 gene was located exclusively in oocytes from the primary stage of follicular growth. There is a complete block of normal follicular development in females carrying two copies of the Inverdale mutation (II), two copies of the Hanna mutation (HH), or one copy of each mutation (HI). Increased ovulation rates are found in females with only one copy of either mutation (I+ or H+). In Booroola sheep, a point mutation was identified in the highly conserved intracellular serine threonine kinase signalling domain of the BMP-1B receptor. Within the ovary, this gene is expressed in oocytes in primordial and pre-antral follicles and in granulosa cells from the primary stage of growth as well as in corpora lutea. The effect of the Booroola mutation is additive for ovulation rate: animals with one copy of the mutation have an ovulation rate of 3 or 4, whereas those with two copies have an ovulation rate of between 5 and 14. Physiological studies of the above mutations demonstrate that the oocyte plays an active role with respect to its adjacent somatic cells during follicular development and support the hypothesis that the oocyte has a significant influence on the number of follicles that proceed to ovulation.

Animals↗

PubMed2001

Highly prolific Booroola sheep have a mutation in the intracellular kinase domain of bone morphogenetic protein IB receptor (ALK-6) that is expressed in both oocytes and granulosa cells.

The Booroola fecundity gene (FecB) increases ovulation rate and litter size in sheep and is inherited as a single autosomal locus. The effect of FecB is additive for ovulation rate (increasing by about 1.6 corpora lutea per cycle for each copy) and has been mapped to sheep chromosome 6q23-31, which is syntenic to human chromosome 4q21-25. Bone morphogenetic protein IB (BMP-IB) receptor (also known as ALK-6), which binds members of the transforming growth factor-beta (TGF-beta) superfamily, is located in the region containing the FecB locus. Booroola sheep have a mutation (Q249R) in the highly conserved intracellular kinase signaling domain of the BMP-IB receptor. The mutation segregated with the FecB phenotype in the Booroola backcross and half-sib flocks of sheep with no recombinants. The mutation was not found in individuals from a number of sheep breeds not derived from the Booroola strain. BMPR-IB was expressed in the ovary and in situ hybridization revealed its specific location to the oocyte and the granulosa cell. Expression of mRNA encoding the BMP type II receptor was widespread throughout the ovary. The mutation in BMPR-IB found in Booroola sheep is the second reported defect in a gene from the TGF-beta pathway affecting fertility in sheep following the recent discovery of mutations in the growth factor, GDF9b/BMP15.

Activin Receptors↗

PubMed2001

Ontogeny of steroidogenesis in the fetal sheep gonad.

The aim of this study was to determine 1) the time of onset and cellular localization of gene expression for steroidogenic factor-1 (SF-1), steroidogenic acute regulatory protein, 3beta-hydroxysteroid dehydrogenase/Delta(5),Delta(4) isomerase (3beta-HSD), and the cytochrome P450 enzymes for cholesterol side-chain cleavage (P450(scc)), 17alpha-hydroxylase (P450(17alphaOH)), and aromatase (P450(arom)) during gonadal development; and 2) the amount of progesterone, androstenedione, testosterone, and 17beta-estradiol present in the fetal sheep gonad. Fetuses were collected on Days 24, 26, 28, 30, 32, 35, 40, 55, and 75 of gestation, and gene expression was determined by in situ hybridization. The steroid content of gonads collected on Days 30, 35, 55, and 75 of gestation was determined by RIA. Developing gonads collected from both male and female fetuses were steroidogenically active around the time of morphological sexual differentiation. In the female, the steroidogenic cells were initially located at the boundary of the cortex and medulla but become increasingly restricted to the mesonephric-derived cell streams. In the male, once tubules were identifiable, steroidogenesis was restricted to the interstitial regions. Interestingly, expression of both SF-1 and 3beta-HSD was observed prior to morphological sexual differentiation. In addition, expression of both of these genes was more widespread than the other genes in both males and females.

3-Hydroxysteroid Dehydrogenases↗

PubMed2001

Steroidogenic acute regulatory protein and peripheral-type benzodiazepine receptor associate at the mitochondrial membrane.

Steroidogenic acute regulatory protein (StAR) and peripheral-type benzodiazepine receptor (PBR) have both been implicated in the transport of cholesterol across mitochondrial membranes in steroidogenic cells. Therefore, we hypothesized that StAR and PBR were associated in this process. To test this hypothesis, we measured fluorescence energy transfer (FET) between these proteins by fusing enhanced green fluorescent protein (GFP, donor fluorophore) and yellow fluorescent protein (YFP, acceptor fluorophore) to the C-terminus of ovine StAR (37GFP) and ovine PBR (PBRYFP), respectively. These intrinsically fluorescent proteins were stably transfected into Cos-7 cells and determined to be biologically active. For FET to occur the appropriate fluorescent molecules need to be <100 A from each other. We observed 22.0 +/- 0.9% energy transfer efficiency for 37GFP and PBRYFP, a 4.9 fold increase above non-specific energy transfer between free GFP and PBRYFP (p <.0001). Thus, it appears that StAR and PBR are closely associated in mitochondrial membranes and that these molecules may interact in the transportation of cholesterol.

Animals↗

PubMed2000

Growth and paracrine factors regulating follicular formation and cellular function.

The purpose of this paper is to review, using fetal sheep as the animal model, aspects of ovarian development related to follicular formation and to report on the identity of growth and paracrine factors which might be involved in this process. Before follicular formation there is a massive and sustained colonisation of the fetal ovary by mesonephric cells, which become a precursor source of follicular cells. From within the ovarian medulla, somatic 'cell-streams' branch into the cortex around nests of oogonia and oocytes. These 'cell-streams', which contain elongated cells with either flattened or cuboidal shaped nuclei, express steroidogenic factor-1 (SF-1), steroid acute regulatory protein (StAR), 3beta-hydroxysteroid dehydrogenase (3beta-HSD), cytochrome P450(scc), and P450(aromatase) mRNA and/or protein. Follicles form from the association of an oocyte with the 'cell-stream' with either a single layer of flattened cells (i.e. type 1 follicle) or with a mixture of flattened and cuboidal cells (i.e. type 1a follicle). These newly-formed follicles have between 3 and 57 somatic cells (i.e. granulosa cells) and contain oocytes which vary in diameter between 23 and 52 microm. Newly formed and early growing follicles have been identified with growth factors or growth factor receptors in either the oocytes or granulosa cells. Many of the growth factors are from the TGFbeta superfamily and are expressed in a cell- and stage-specific manner.

Animals↗

PubMed2000

Mutations in an oocyte-derived growth factor gene (BMP15) cause increased ovulation rate and infertility in a dosage-sensitive manner.

Multiple ovulations are uncommon in humans, cattle and many breeds of sheep. Pituitary gonadotrophins and as yet unidentified ovarian factors precisely regulate follicular development so that, normally, only one follicle is selected to ovulate. The Inverdale (FecXI) sheep, however, carries a naturally occurring X-linked mutation that causes increased ovulation rate and twin and triplet births in heterozygotes (FecXI/FecX+; ref. 1), but primary ovarian failure in homozygotes (FecXI/FecXI; ref. 2). Germ-cell development, formation of the follicle and the earliest stages of follicular growth are normal in FecXI/FecXI sheep, but follicular development beyond the primary stage is impaired. A second family unrelated to the Inverdale sheep also has the same X-linked phenotype (Hanna, FecXH). Crossing FecXI with FecXH animals produces FecXI/FecXH infertile females phenotypically indistinguishable from FecXI/FecXI females. We report here that the FecXI locus maps to an orthologous chromosomal region syntenic to human Xp11.2-11.4, which contains BMP15, encoding bone morphogenetic protein 15 (also known as growth differentiation factor 9B (GDF9B)). Whereas BMP15 is a member of the transforming growth factor beta (TGFbeta) superfamily and is specifically expressed in oocytes, its function is unknown. We show that independent germline point mutations exist in FecXI and FecXH carriers. These findings establish that BMP15 is essential for female fertility and that natural mutations in an ovary-derived factor can cause both increased ovulation rate and infertility phenotypes in a dosage-sensitive manner.

Amino Acid Sequence↗

PubMed2000

Effect of dose of prostaglandin F(2alpha) on steroidogenic components and oligonucleosomes in ovine luteal tissue.

To determine whether prostaglandin (PG) F(2alpha) had a dose-dependent effect upon secretion of progesterone, oligonucleosome formation, or loss of luteal weight, ewes on Day 9 or 10 of the estrous cycle were administered 0, 3, 10, or 30 mg PGF(2alpha) per 60 kg BW (i.v.), and luteal tissue was collected 9 and 24 h after injection. All doses of PGF(2alpha) decreased (P < 0. 05) concentrations of progesterone in sera by 9 h; however, in ewes treated with 3 mg PGF(2alpha), concentrations of progesterone were similar to control values at 24 h and higher (P < 0.05) than those in the 10- or 30-mg groups. Concentrations of progesterone in sera over all dose levels were highly correlated to luteal concentrations of mRNA encoding steroidogenic acute regulatory protein (P < 0.001), cytochrome P450 side-chain cleavage (P < 0.02), and 3beta-hydroxysteroid dehydrogenase (P < 0.01). Corpora lutea collected at 24 h from ewes treated with the 10- and 30-mg doses of PGF(2alpha) weighed less (P < 0.05) than those from controls. Oligonucleosomes were not present in luteal tissues from control ewes. Surprisingly, all doses of PGF(2alpha)-induced oligonucleosomes in a majority of animals at 9 h and in a majority of ewes treated with 10 and 30 mg of PGF(2alpha) at 24 h. In conclusion, 3 mg of PGF(2alpha) per 60 kg BW transiently decreased serum concentrations of progesterone and induced oligonucleosome formation, but did not result in reduced luteal weight. The 10- and 30-mg doses of PGF(2alpha) decreased secretion of progesterone and induced oligonucleosome formation and luteolysis.

3-Hydroxysteroid Dehydrogenases↗

PubMed2000

Gene expression in abnormal ovarian structures of ewes homozygous for the inverdale prolificacy gene.

Animals heterozygous (I+) for the Inverdale prolificacy gene (FecX(I)) have an increased ovulation rate whereas those homozygous (II) for FecX(I) are infertile with "streak" ovaries and follicular development arrested at the primary (type 2 follicle) stage. The streak ovaries also contain small oocyte-free nodules with granulosa-like cells and often tumor-like structures. It has been hypothesized that these abnormal structures are of granulosa cell origin, and the aim of this study was to determine whether genes normally expressed in granulosa cells are also expressed in the nodules and tumor-like structures. The mRNAs encoding c-kit and its ligand stem cell factor (SCF), FSH receptor (FSH-R), follistatin, alpha-inhibin subunit, and the beta(A)- and beta(B)-activin/inhibin subunits were localized in ovaries of ewes with 0 (++), 1 (I+), or 2 (II) copies of the FecX(I) gene (n = 4-9 animals per genotype per gene) using in situ hybridization. Ontogeny of expression of all mRNAs examined was similar between ++ and I+ ewes. Expression of c-kit mRNA was observed in the oocyte of all follicular types present in ++, I+, and II ewes. Moreover, granulosa cells of type 2 (II) and type 2 and larger follicles (++, I+) expressed SCF mRNA. The mRNAs encoding FSH-R, follistatin, alpha-inhibin subunit, and beta(B)-activin/inhibin subunit were identified in type 3 and larger follicles of ++ and I+ ewes but not in follicles of II ewes that were only at the type 1, 1a, or 2 stages of development. However, the cells within the oocyte-free nodules of II ewes expressed all of these genes. The mRNAs encoding c-kit and beta(A)-activin/inhibin subunit were not observed in granulosa cells until antrum formation (type 5 follicles) or in the nodules of II ewes. Tumors from 4 ewes were obtained and classified as cystic, semisolid, or solid structures containing granulosa-like cells or as solid structures containing predominately fibroblast- and luteal-like cells. Often, two tumors were present on the same ovary. Tumors containing granulosa-like cells (n = 3-4 per gene) expressed the mRNAs encoding alpha-inhibin subunit, beta(A)-, and beta(B)-activin/inhibin subunits, follistatin, and the FSH-R but did not contain detectable amounts of mRNA for c-kit or SCF. Tumors composed predominately of fibroblast- and luteal-like cells expressed very low levels of SCF mRNA; of the other mRNAs examined, none were detected. Also, none of the genes examined were found to be expressed by the surface epithelium, theca externa, fibroblast, or vascular cells within the ovary of animals of any genotype. These findings are consistent with the hypothesis that the somatic cells in oocyte-free nodules and tumor-like tissue in II ewes originate from the granulosa cells of the small follicles.

Activins↗

PubMed2000

Prostaglandin metabolism in the ovine corpus luteum: catabolism of prostaglandin F(2alpha) (PGF(2alpha)) coincides with resistance of the corpus luteum to PGF(2alpha).

To examine possible mechanisms involved in resistance of the ovine corpus luteum to the luteolytic activity of prostaglandin (PG)F(2alpha), the enzymatic activity of 15-hydroxyprostaglandin dehydrogenase (PGDH) and the quantity of mRNA encoding PGDH and cyclooxygenase (COX-2) were determined in ovine corpora lutea on Days 4 and 13 of the estrous cycle and Day 13 of pregnancy. The corpus luteum is resistant to the action of PGF(2alpha) on Days 4 of the estrous cycle and 13 of pregnancy while on Day 13 of the estrous cycle the corpus luteum is sensitive to the actions PGF(2alpha). Enzymatic activity of PGDH, measured by rate of conversion of PGF(2alpha) to PGFM, was greater in corpora lutea on Day 4 of the estrous cycle (P < 0.05) and Day 13 of pregnancy (P < 0.05) than on Day 13 of the estrous cycle. Levels of mRNA encoding PGDH were also greater in corpora lutea on Day 4 of the estrous cycle (P < 0. 01) and Day 13 of pregnancy (P < 0.01) than on Day 13 of the estrous cycle. Thus, during the early estrous cycle and early pregnancy, the corpus luteum has a greater capacity to catabolize PGF, which may play a role in the resistance of the corpus luteum to the actions of this hormone. Levels of mRNA encoding COX-2 were undetectable in corpora lutea collected on Day 13 of the estrous cycle but were 11 +/- 4 and 44 +/- 28 amol/microgram poly(A)(+) RNA in corpora lutea collected on Day 4 of the estrous cycle and Day 13 of pregnancy, respectively. These data suggest that there is a greater capacity to synthesize PGF(2alpha), early in the estrous cycle and early in pregnancy than on Day 13 of the estrous cycle. In conclusion, enzymatic activity of PGDH may play an important role in the mechanism involved in luteal resistance to the luteolytic effects of PGF(2alpha).

Animals↗

PubMed2000

Mechanisms controlling the function and life span of the corpus luteum.

The primary function of the corpus luteum is secretion of the hormone progesterone, which is required for maintenance of normal pregnancy in mammals. The corpus luteum develops from residual follicular granulosal and thecal cells after ovulation. Luteinizing hormone (LH) from the anterior pituitary is important for normal development and function of the corpus luteum in most mammals, although growth hormone, prolactin, and estradiol also play a role in several species. The mature corpus luteum is composed of at least two steroidogenic cell types based on morphological and biochemical criteria and on the follicular source of origin. Small luteal cells appear to be of thecal cell origin and respond to LH with increased secretion of progesterone. LH directly stimulates the secretion of progesterone from small luteal cells via activation of the protein kinase A second messenger pathway. Large luteal cells are of granulosal cell origin and contain receptors for PGF(2alpha) and appear to mediate the luteolytic actions of this hormone. If pregnancy does not occur, the corpus luteum must regress to allow follicular growth and ovulation and the reproductive cycle begins again. Luteal regression is initiated by PGF(2alpha) of uterine origin in most subprimate species. The role played by PGF(2alpha) in primates remains controversial. In primates, if PGF(2alpha) plays a role in luteolysis, it appears to be of ovarian origin. The antisteroidogenic effects of PGF(2alpha) appear to be mediated by the protein kinase C second messenger pathway, whereas loss of luteal cells appears to follow an influx of calcium, activation of endonucleases, and an apoptotic form of cell death. If the female becomes pregnant, continued secretion of progesterone from the corpus luteum is required to provide an appropriate uterine environment for maintenance of pregnancy. The mechanisms whereby the pregnant uterus signals the corpus luteum that a conceptus is present varies from secretion of a chorionic gonadotropin (primates and equids), to secretion of an antiluteolytic factor (domestic ruminants), and to a neuroendocrine reflex arc that modifies the secretory patterns of hormones from the anterior pituitary (most rodents).

Animals↗

PubMed1999

Molecular regulation of luteal progesterone synthesis in domestic ruminants.

Regulation of progesterone secretion from the corpus luteum during the oestrous cycle requires the integration of multiple signals to achieve the appropriate amount of progesterone to maximize reproductive efficiency. Development of a mature corpus luteum capable of secreting sufficient amounts of progesterone is dependent upon the pituitary hormones LH and growth hormone (GH). Continued secretion of progesterone from the mature corpus luteum is also dependent upon pituitary hormones. If pregnancy does not occur, prostaglandin F2 alpha (PGF2 alpha) of uterine origin causes a precipitous decrease in progesterone secretion and demise of the corpus luteum. A major point of regulation of progesterone secretion by both luteotrophic and luteolytic hormones appears to be regulation of transport of cholesterol through the mitochondrial membranes to cytochrome P450scc. It is likely that both luteotrophic and luteolytic hormones regulate steroidogenic acute regulatory protein (StAR), which facilitates transport. Regulation may be occurring through increases or decreases in gene transcription, translation efficiency or post-translational modifications such as phosphorylation. Thus, although synthesis of progesterone is a complex process, both positive and negative regulation of the process appears to occur primarily at a single step (transport of cholesterol to the inner mitochondrial membrane) in the pathway.

Animals↗

PubMed1998

Concentration of mRNA encoding 3 beta-hydroxysteroid dehydrogenase/delta 5,delta 4 isomerase (3 beta-HSD) and 3 beta-HSD enzyme activity following treatment of ewes with prostaglandin F2 alpha.

The objectives of these experiments were (1) to determine if prostaglandin F2 alpha (PGF2 alpha) decreased mRNA encoding 3 beta-hydroxysteroid dehydrogenase/d5,delta 4 isomerase (3 beta-HSD) specifically in large steroidogenic luteal cells, which contain the high affinity receptors for PGF2 alpha; and (2) to determine if the decreased concentration of mRNA encoding 3 beta-HSD following administration of PGF2 alpha was associated with a decrease in 3 beta-HSD enzyme activity. Ewes on days 11 or 12 of the estrous cycle were administered PGF2 alpha (25 mg i.v. followed by 10 mg i.m. 2 h later) and corpora lutea collected 4, 12, 24, or 48 h later (n = 4-5/time). Corpora lutea were also collected from non-injected (n = 4) or saline-injected (n = 4) control ewes. Administration of PGF2 decreased (P < 0.05) steady-state concentrations of mRNA encoding 3 beta-HSD to 35, 15, 9, and 5 percent of the concentrations in the control group at 4, 12, 24, and 48 h, respectively. Concentrations of mRNA encoding 3 beta-HSD in large luteal cells were decreased to 43% of controls 4 h following injection, which was similar to the decrease seen in steady-state concentrations of this mRNA in total luteal mRNA (35%). However, 3 beta-HSD enzyme activity was not significantly decreased by 48 h after PGF2 alpha injection. Thus, the dramatic decreased in mRNA encoding 3 beta-HSD was not associated with an immediate decrease in 3 beta-HSD enzyme activity and, therefore, does not appear to be responsible for the acute decrease in secretion of progesterone from ovine luteal tissue during PGF2 alpha-induced luteolysis.

Animals↗

PubMed1998

Luteal expression of steroidogenic factor-1 mRNA during the estrous cycle and in response to luteotropic and luteolytic stimuli in ewes.

Steroidogenic factor-1 (SF-1) is a transcription factor involved in regulating basal and/or cAMP-induced increases in expression of several components of the steroidogenic pathway, including cytochrome P450 side-chain cleavage (P450scc), steroidogenic acute regulatory protein (StAR), and 3beta-hydroxysteroid dehydrogenase/delta5, delta4 isomerase (3beta-HSD). In experiment 1, on days 3, 6, 9, 12, and 15 of the estrous cycle, steady-state concentrations (fmol/microg poly A+ RNA) of SF-1 mRNA in luteal tissue were 0.09 +/- 0.01, 0.17 +/- 0.01, 0.24 +/- 0.03, 0.30 +/- 0.09, and 0.20 +/- 0.05, respectively (estrus = day 0; n = 4/d). Concentrations of SF-1 mRNA increased (p < 0.05) between days 3 and 12, but were not different among the other days of the estrous cycle. Luteal concentrations of SF-1 mRNA and concentrations of progesterone in sera were highly correlated (p < 0.01; r = 0.72). In experiment 2, ewes on days 11 or 12 of the estrous cycle were injected with 25 mg prostaglandin F2alpha (PGF2alpha) into the jugular vein followed by an injection of 10 mg PGF2alpha i.m. 2 h later. Corpora lutea were collected 4, 12, and 24 h after the first injection of PGF2alpha (n = 4-5 ewes/time). Control luteal tissue was collected from ewes on days 11-13 of the estrous cycle, which had not been injected (n = 4) or had been injected with saline 24 h previously (n = 4). Steady-state concentrations of SF-1 mRNA had decreased (p < 0.05) to 48% of control values by 4 h after injection, and remained low at 12 and 24 h. In experiment 3, ewes on days 9-12 of the estrous cycle were administered PGF2alpha (1 micromol), phorbol 12-myristate 13-acetate (PMA; 2 micromol), luteinizing hormone (LH; 20 microg), forskolin (50 microg), or vehicle (1 mL saline) directly into the ovarian artery. Corpora lutea were collected 0 (noninfused) 4, 12, or 24 h later (n = 3-4 animals/treatment/time) for quantification of SF-1 mRNA. Steady-state concentrations of mRNA encoding SF-1 were not affected by infusion of PGF2alpha or PMA, although concentrations of mRNA encoding StAR and 3beta-HSD were decreased (p < 0.05) by these treatments. Concentrations of mRNA encoding SF-1 were increased (p < 0.05) to 157 and 149% of control values by LH and forskolin, respectively, 12 h following infusion and returned to control values by 24 h following either treatment. In contrast, infusion of LH or forskolin did not change concentrations of mRNA encoding StAR, P450scc, or 3beta-HSD. In summary, during the estrous cycle, the pattern of expression of SF-1 mRNA was similar to the pattern of concentrations of progesterone in serum and expression of mRNA encoding P450scc, but differed from that previously shown for 3beta-HSD and StAR mRNA. The effects of administration of PGF2alpha on concentrations of SF-1 mRNA appeared to be dose-dependent. However, acute effects of PGF2alpha on mRNA encoding 3beta-HSD and StAR were observed when concentrations of mRNA encoding SF-1 were not influenced. In addition, although LH or forskolin increased luteal SF-1 mRNA 12 h following infusion, no increases in mRNA encoding StAR, P450scc, or 3beta-HSD were observed. Thus, during the midluteal phase of the estrous cycle, neither luteotropic nor luteolytic hormones appear to coordinately regulate mRNA encoding SF-1 and mRNA encoding StAR, P450scc, or 3beta-HSD.

3-Hydroxysteroid Dehydrogenases↗

PubMed1998

Steady-state concentrations of mRNA encoding two inhibitors of protein kinase C in ovine luteal tissue.

Prostaglandin F2 alpha (PGF2 alpha) decreases secretion of progesterone from the corpus luteum in domestic ruminants. However, it is less effective during the early part of the oestrous cycle (Louis et al., 1973) and at the time of maternal recognition of pregnancy (Silvia and Niswender, 1984; Lacroix and Kann, 1986). Decreased luteal responsiveness may be due to failure of PGF2 alpha to activate fully its normal second messenger system, protein kinase C (PKC). Alternatively, increased resistance of the corpus luteum to PGF2 alpha might be attributable to greater concentrations of recently identified biological inhibitors of PKC. These possibilities were addressed by measuring steady-state concentrations of mRNA encoding PGF2 alpha receptor and two inhibitors of PKC, protein kinase C inhibitor-1 (PKCI-1) and kinase C inhibitor protein-1 (KCIP-1, brain 14-3-3 protein), in corpora lutea collected from ewes on days 4, 10 and 15 of the oestrous cycle (n = 5 per day) and day 15 of pregnancy (n = 7). There were no differences in mean concentrations of mRNA encoding PGF2 alpha receptor among the groups. However, concentrations of mRNA encoding both inhibitors of PKC were higher (P < 0.01) on day 4 of the oestrous cycle compared with the other groups. Treatment of ewes with a luteolytic dose of PGF2 alpha, which activates PKC, did not change concentrations of mRNA encoding either PKCI-1 or KCIP-I up to 24 h later. Luteal expression of mRNA encoding the PKC inhibitors and PGF2 alpha receptor was also examined in ewes treated with oestradiol in vivo for 16 h in the midluteal phase. High concentrations of oestradiol in serum (20 and 70 pg ml-1) did not influence quantities of any of the mRNAs examined. Therefore, an increase in PKC inhibitors may be involved in resistance of the corpus luteum to PGF2 alpha during the early part of the oestrous cycle but does not appear to mediate the increased resistance of the corpus luteum to PGF2 alpha during maternal recognition of pregnancy. Neither PGF2 alpha nor oestradiol affected steady-state concentrations of mRNAs encoding PKCI-1 or KCIP-I.

14-3-3 Proteins↗

PubMed1997

Effects of ovariectomy and hypothalamic-pituitary disconnection on amounts of steroidogenic factor-1 mRNA in the ovine anterior pituitary gland.

Steroidogenic factor-1 (SF-1) is a transcription factor involved in regulation of steroidogenic enzymes. Recent evidence indicates that SF-1 is also important in the anterior pituitary gland, where it may influence gene expression in gonadotropes. We isolated a cDNA encoding ovine SF-1 and demonstrated that the SF-1 gene is expressed in the anterior pituitary gland of sheep. SF-1 transcripts and luteinizing hormone (LH) were colocalized in gonadotropes by in situ hybridization and immunohistochemistry, respectively. To test the hypothesis that GnRH stimulates pituitary expression of ovine SF-1 mRNA, ewes were ovariectomized to increase endogenous secretion of GnRH. Compared to ovary-intact ewes, ovariectomy resulted in three- and fourfold increases in steady-state amounts of mRNA encoding SF-1 and LH beta subunit, respectively. In ovariectomized ewes in which delivery of GnRH to the anterior pituitary gland was prevented by hypothalamic-pituitary disconnection (HPD), steady-state amounts of mRNA encoding SF-1 and LH beta-subunit were decreased. These results provide evidence that pituitary SF-1 gene expression in sheep is regulated by GnRH. Coordinate regulation of mRNAs encoding SF-1 and LH beta-subunit raises the possibility that SF-1 may be an important transcriptional regulator of LH beta-subunit gene expression in ovine gonadotropes.

Animals↗

PubMed1997

Hormonal regulation of monocyte chemoattractant protein-1 messenger ribonucleic acid expression in corpora lutea.

Monocyte chemoattractant protein-1 (MCP-1) is a potent chemokine that attracts monocytes and macrophages. It is known that macrophages accumulate in the corpus luteum (CL) during luteal regression in many species. In this study, we investigated the regulation of MCP-1 mRNA in ovine and bovine CL during prostaglandin (PG) F2alpha-induced luteolysis, after LH treatment, or after pharmacologic activation of the protein kinase (PK) A or PKC intracellular effector systems. In experiment 1, ewes on day 11 or 12 of the estrous cycle were infused with saline or PGF2alpha. PGF2alpha increased MCP-1 mRNA at 1 and 4 h after treatment. MCP-1 mRNA returned to basal level at 12 h and increased again at 24 h post treatment. In experiment 2, ewes received saline, PGF2alpha, phorbol 12-myristate 13-acetate (PMA), luteinizing hormone (LH), or forskolin infusion and CL were collected at 0 (untreated), 4, 12, or 24 h after infusion. Similar to experiment 1, PGF2alpha induced MCP-1 mRNA at 4 and 24 h post treatment. PMA increased mRNA for MCP-1 at 4, 12, and 24 h. Treatment with LH or forskolin transiently decreased MCP-1 mRNA expression. In experiment 3, cows were treated with a luteolytic dose (25 mg) of PGF2alpha on day 4 or day 11 of estrous cycle and expression of MCP-1 mRNA was quantified. Steady-state concentrations of mRNA for MCP-1 were induced by PGF2alpha treatment only in mid-cycle CL but not in early CL. In summary, administration of PGF2alpha or activation of PKC induced MCP-1 mRNA expression. Expression of MCP-1 may be important for stimulating immune processes during luteal regression.

Animals↗