Expression of the ovine stem cell factor gene during folliculogenesis in late fetal and adult ovaries

1997 ◽  
Vol 18 (2) ◽  
pp. 127-135 ◽  
Author(s):  
D J Tisdall ◽  
L D Quirke ◽  
P Smith ◽  
K P McNatty
Keyword(s):  
Gene Expression ◽  
Stem Cell ◽  
Granulosa Cells ◽  
Stem Cell Factor ◽  
Ovarian Follicle ◽  
Northern Analysis ◽  
Nucleotide Sequencing ◽  
Sequence Difference ◽  
Follicle Growth ◽  
Fetal Sheep

ABSTRACT Two ovine stem cell factor (oSCF) cDNAs (822 bp and 738 bp) were generated from ovarian follicle mRNA by RT-PCR. Nucleotide sequencing revealed that the oSCF 822 bp cDNA encodes a precursor protein of 274 amino acids. An amino acid change 109E to 109Q was the only sequence difference from that previously described for this species. The smaller (738 bp) oSCF cDNA was shown by nucleotide sequencing to be an mRNA splice variant, equivalent to that found in other mammals, in which an exon (84 bp) encoding a potential proteolytic cleavage site is removed. Northern analysis revealed a single transcript of approximately 6·5 kb in follicles, corpora lutea and stroma of mid-luteal sheep ovaries. In situ hybridization was used to detect oSCF mRNA within ovaries of fetal sheep on days 90, 100, 120 and 135 of gestation (term=147) and of adult sheep within the breeding season. In fetal and adult ovaries, oSCF mRNA was detected in the granulosa cells of follicles at all stages of follicle growth (primordial through to antral). The SCF gene was also expressed in granulosa cells of atretic follicles but appeared to be down-regulated in the cumulus cells surrounding the oocyte at more advanced stages of atresia. In fetal ovaries at day 90 of gestation (90DG), oSCF was expressed in the subepithelial mesenchymal cells of the ovarian cortex. By 100DG the gene expression in the subepithelial cells became restricted to a narrow region below the epithelium, and areas of expression were observed in groups of cells around isolated oocytes, primordial and primary follicles. oSCF gene expression also occurred in the surface epithelial cells of 90DG ovaries, the expression was absent from these cells by 135DG and in adult ovaries. Localization of oSCF mRNA was observed in the ovarian rete and endothelial cells of blood vessels of fetal ovaries. These results suggest that oSCF may have an important and continuous role in the development and/or maintenance of germ cells during follicle growth and atresia in sheep.

Cloning, sequencing and expression of stem cell factor (c-kit ligand) cDNA of brushtail possum (Trichosurus vulpecula)

1996 ◽  
Vol 8 (4) ◽  
pp. 789 ◽  
Author(s):  
PJ Greenwood ◽  
C Seamer ◽  
DJ Tisdall
Keyword(s):  
Stem Cell ◽  
Amino Acid ◽  
Stem Cell Factor ◽  
Biological Activities ◽  
Amino Acid Level ◽  
Amino Acid Sequences ◽  
Northern Analysis ◽  
Nucleotide Sequencing ◽  
Brushtail Possum ◽  
Factor C

By means of reverse transcription polymerase chain reaction (RT-PCR), three stem cell factor (SCF) cDNAs (822-738 bp in size) were amplified from brushtail possum ovarian poly (A)+ RNA. The largest and smallest of these cDNAs were cloned and sequenced. Characterization of these cDNAs has revealed that possum SCF has approximately 75% and 66% homology to SCF of eutherian mammals at the nucleotide level and the predicted amino acid level respectively. Nucleotide sequencing shows that the 738-bp cDNA represents an mRNA splice variant, equivalent to that found in eutherian mammals, in which an exon (84 bp) encoding a potential proteolytic cleavage site is removed. Comparison of the predicted possum SCF amino acid sequence with the predicted SCF amino acid sequences from eutherian mammals reveals conservation of all cysteine residues and 3 of 4 potential N-linked glycosylation sites. In addition, the hydropathicity profile of the possum SCF protein is similar to that of eutherian SCF suggesting that protein conformation is conserved. Northern analysis was used to characterize possum SCF gene expression in adult ovary and testis. A major transcript of 9 kb was observed in both ovarian and testicular tissue. The conservation of the SCF gene and its predicted protein, suggests that SCF in the possum has similar biological activities to SCF in eutherian mammals.

FSH receptor gene expression during ovarian follicle development in sheep

1995 ◽  
Vol 15 (3) ◽  
pp. 273-281 ◽  
Author(s):  
D J Tisdall ◽  
K Watanabe ◽  
N L Hudson ◽  
P Smith ◽  
K P McNatty
Keyword(s):  
Gene Expression ◽  
Granulosa Cells ◽  
Ovarian Follicle ◽  
Receptor Gene ◽  
Fsh Receptor ◽  
Follicular Growth ◽  
Fetal Sheep ◽  
Adult Sheep ◽  
Receptor Gene Expression ◽  
Fsh Receptor Gene

ABSTRACT A key question in elucidating the role of FSH in ovarian function is to determine when during follicular growth the FSH receptor first appears. The aim of this study was to examine the site and time of FSH receptor gene expression during early follicular growth. This study was carried out on ovaries of adult sheep during the luteal and prostaglandin-induced follicular phase of the oestrous cycle and also on ovaries of fetal sheep at 90, 100, 120 and 135 days of gestation (term=day 147). Using reverse transcription-PCR and a set of PCR primers spanning exons 8/9/10, two partial FSH receptor cDNAs (500 and 310 bp) were isolated from adult sheep ovary. It was shown by sequencing that exon 8 was deleted in the 310 bp cDNA, implying that this was part of an alternatively spliced FSH receptor transcript. Using RNA in situ hybridisation on ovaries of adult sheep, FSH receptor mRNA was observed in granulosa cells of early preantral follicles with one to two cell layers and it was seen that gene expression continued throughout folliculogenesis into advanced stages of atresia. Moreover, in the fetus, FSH receptor gene expression was detected in follicles with two or more layers of granulosa cells in ovaries taken at 100, 120 and 135 days of gestation. These results suggest that the FSH receptor gene is expressed after the granulosa cells of a folllicle have begun to divide but not during the earliest stages of follicle growth, namely the transformation of a primordial follicle to a primary follicle.

Anti-Müllerian Hormone Regulates Stem Cell Factor via cAMP/PKA Signaling Pathway in Human Granulosa Cells by Inhibiting the Phosphorylation of CREB

2020 ◽  
Vol 27 (1) ◽  
pp. 325-333
Author(s):  
Yun-Xing Fu ◽  
Fei-Miao Wang ◽  
Xiao-E Ou-yang ◽  
Hui-Min Yang ◽  
Ting Hu ◽  
...  
Keyword(s):  
Stem Cell ◽  
Signaling Pathway ◽  
Granulosa Cells ◽  
Stem Cell Factor ◽  
Human Granulosa Cells

KIT Exon 8 Mutations Associated with Core Binding Factor (CBF) - Acute Myeloid Leukemia (AML) Cause Hypersensitivity of KIT to Its Natural Ligand Stem Cell Factor.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 92-92
Author(s):  
Tobias M. Kohl ◽  
Susanne Schnittger ◽  
Wolfgang Hiddemann ◽  
Karsten Spiekermann
Keyword(s):  
Acute Myeloid Leukemia ◽  
Stem Cell ◽  
Stem Cell Factor ◽  
Myeloid Leukemia ◽  
Biological Effects ◽  
Nucleotide Sequencing ◽  
Core Binding Factor ◽  
Binding Factor ◽  
Significant Difference ◽  
Acute Myeloid

Abstract Mutations in the extracellular portion of the KIT receptor tyrosine kinase (exon 8 mutations) are strongly associated with core binding factor (CBF) - acute myeloid leukemia (AML), but the functional role of these mutations has not been elucidated. In 93% of cases, codon Asp419 is deleted and exon 8 mutations were reported to confer an impaired prognosis to patients with CBF-AML. In this study, we are the first to report pro-proliferative and antiapoptotic potential of representative KIT exon 8 mutations in a cell culture model and to show a significant difference to KIT wildtype (KIT-WT). Three representative exon 8 mutants including a single deletion of codon 419 were created by in vitro site-directed mutagenesis. The integrity of all constructs was assessed by complete nucleotide sequencing. After stable expression in IL-3 dependent Ba/F3 cells (confirmed by FACS analysis and immunoblotting), exon 8 KIT mutants were characterized by a hypersensitivity to stem cell factor (SCF) stimulation in terms of proliferation and resistance to apoptotic cell death. The differences to KIT-WT occurred in the physiological range of SCF from 1 to 10ng/ml. The proliferative response caused by stimulation with SCF was reversed in KIT-WT and exon 8 mutants in the presence of Imatinib® (Novartis) in contrast to the activation loop mutant D816V which could not be inhibited. These biological effects were confirmed by demonstrating increased phosphorylation of the KIT downstream targets mitogen-activated protein kinase (MAPK) and AKT after SCF stimulation compared to the KIT-WT receptor. Furthermore, the MEK inhibitor PD98059 and the PI3 kinase inhibitor LY294002 resulted in a dose dependent inhibition of SCF induced proliferation in exon 8 mutants. Our data show that KIT exon 8 mutations represent gain-of-function mutations by inducing receptor hypersensitivity to its ligand SCF by activation of MAPK and PI3K and might represent a new molecular target for treatment of CBF leukemias.

Localization of ovine follistatin and α and βA inhibin mRNA in the sheep ovary during the oestrous cycle

1994 ◽  
Vol 12 (2) ◽  
pp. 181-193 ◽  
Author(s):  
D J Tisdall ◽  
N Hudson ◽  
P Smith ◽  
K P McNatty
Keyword(s):  
Gene Expression ◽  
Granulosa Cells ◽  
Ovarian Follicle ◽  
Indirect Evidence ◽  
Oestrous Cycle ◽  
Corpora Lutea ◽  
Α Subunit ◽  
Ovarian Follicle Development ◽  
Antral Follicles

ABSTRACT The sites of follistatin and α and βA inhibin gene expression were examined by in situ hybridization in sheep ovaries during the early and mid-luteal phases (days 3 and 10) of the oestrous cycle and a prostaglandin F2α (PGF2α)-induced follicular phase. Follistatin mRNA was detected in the granulosa cells of preantral, antral and early atretic follicles at all stages of the oestrous cycle, and in the corpora lutea at the early and mid-luteal stages of the cycle. However, only low levels of expression of follistatin were observed in the presumptive preovulatory follicle at 56 h after treatment with PGF2α. Both α and βA inhibin were shown to be expressed in ovaries at all stages of the oestrous cycle. In situ hybridization localized α subunit mRNA to the granulosa cells of most, but not all, healthy antral follicles, and to no other ovarian cell type. In contrast, expression of the βA subunit was confined to a few medium-to-large healthy antral follicles. In antral follicles expressing βA inhibin, mRNAs for α inhibin and follistatin were always detected, but the converse was not true. Unlike follistatin, no α and βA inhibin expression was seen in preantral follicles, developing corpora lutea, or follicles undergoing atresia. These results show that, in the adult sheep ovary, follistatin gene expression is a constitutive event in all growing follicles from the early preantral stage, and also provide indirect evidence of the involvement of follistatin, but not inhibin or activin, in the early stages of ovarian follicle development in sheep.

scholarly journals Association of stem cell factor gene expression with severity and atopic state in patients with bronchial asthma

2017 ◽  
Vol 18 (1) ◽  
Author(s):  
Safaa I. Tayel ◽  
Sally M. El-Hefnway ◽  
Eman M. Abd El Gayed ◽  
Gehan A. Abdelaal
Keyword(s):  
Gene Expression ◽  
Stem Cell ◽  
Bronchial Asthma ◽  
Stem Cell Factor ◽  
Factor Gene ◽  
Atopic State

scholarly journals Inhibition of the stem cell factor 248 isoform attenuates the development of pulmonary remodeling disease

2020 ◽  
Vol 318 (1) ◽  
pp. L200-L211
Author(s):  
Andrew Rasky ◽  
David M. Habiel ◽  
Susan Morris ◽  
Matthew Schaller ◽  
Bethany B. Moore ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cell ◽  
Stem Cell Factor ◽  
Mesenchymal Cell ◽  
Lung Fibroblasts ◽  
Normal Lung ◽  
Expression Array ◽  
Lung Remodeling ◽  
Lung Biopsies

Stem cell factor (SCF) and its receptor c-kit have been implicated in inflammation, tissue remodeling, and fibrosis. Ingenuity Integrated Pathway Analysis of gene expression array data sets showed an upregulation of SCF transcripts in idiopathic pulmonary fibrosis (IPF) lung biopsies compared with tissue from nonfibrotic lungs that are further increased in rapid progressive disease. SCF248, a cleavable isoform of SCF, was abundantly and preferentially expressed in human lung fibroblasts and fibrotic mouse lungs relative to the SCF220 isoform. In fibroblast-mast cell coculture studies, blockade of SCF248 using a novel isoform-specific anti-SCF248 monoclonal antibody (anti-SCF248), attenuated the expression of COL1A1, COL3A1, and FN1 transcripts in cocultured IPF but not normal lung fibroblasts. Administration of anti-SCF248 on days 8 and 12 after bleomycin instillation in mice significantly reduced fibrotic lung remodeling and col1al, fn1, acta2, tgfb, and ccl2 transcript expression. In addition, bleomycin increased numbers of c-kit+ mast cells, eosinophils, and ILC2 in lungs of mice, whereas they were not significantly increased in anti-SCF248-treated animals. Finally, mesenchymal cell-specific deletion of SCF significantly attenuated bleomycin-mediated lung fibrosis and associated fibrotic gene expression. Collectively, these data demonstrate that SCF is upregulated in diseased IPF lungs and blocking SCF248 isoform significantly ameliorates fibrotic lung remodeling in vivo suggesting that it may be a therapeutic target for fibrotic lung diseases.

268. Suppressors of cytokine signalling (SOCS): roles in ovarian follicle activation

2008 ◽  
Vol 20 (9) ◽  
pp. 68
Author(s):  
R. Keightley ◽  
E. McLaughlin ◽  
S. D. Roman ◽  
R. L. Robker ◽  
D. L. Russell
Keyword(s):  
Granulosa Cells ◽  
Ovarian Follicle ◽  
Expression Patterns ◽  
Neonatal Mouse ◽  
Mrna Levels ◽  
Follicle Growth ◽  
Oocyte Growth ◽  
Primordial Follicles ◽  
Gene And Protein Expression ◽  
Follicle Activation

Oocytes are sequestered in primordial follicles before birth and remain quiescent in the ovary for decades, until recruited into the growing pool throughout the reproductive years. Therefore activation of follicle growth is a major biological checkpoint that controls female reproductive potential. However we are only just beginning to elucidate the cellular mechanisms required, for either maintenance of the quiescent primordial pool, or initiation of follicle growth. Analysis of microarray data derived from neonatal mouse ovaries indicated that members of the Suppressors of Cytokine Signalling SOCS family of proteins may play pivotal roles in folliculogenesis. We undertook a detailed analysis of gene and protein expression patterns of the eight members of the SOCS family, namely CIS and SOCS1–7, within adult and neonatal mouse ovaries. Quantitative real time PCR and immunohistochemistry was performed to determine mRNA levels and cellular localisation in the ovaries of cycling and new born animals. SOCS proteins were expressed largely within the oocytes of developing follicles and in the granulosa cells of the larger preovulatory follicles. Expression of SOCS4 in the granulosa cells and SOCS5 within the oocyte was coincident with the activation of oocyte growth and the differentiation of squamous pregranulosa to cuboidal granulosa cells. Our investigation has identified a role for the SOCS family proteins within the ovary and SOCS4 and SOCS5 as major regulators of cytokine signalling pathways in follicle activation and development.

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