The Absence of ER-β Results in Altered Gene Expression in Ovarian Granulosa Cells Isolated From In Vivo Preovulatory Follicles

Abstract Determining the spatial and temporal expression of genes involved in the ovulatory pathway is critical for the understanding of the role of each estrogen receptor in the modulation of folliculogenesis and ovulation. Estrogen receptor (ER)-β is highly expressed in ovarian granulosa cells, and mice lacking ER-β are subfertile due to inefficient ovulation. Previous work has focused on isolated granulosa cells or cultured follicles and, although informative, provides confounding results due to the heterogeneous cell types present including granulosa and theca cells and oocytes and exposure to in vitro conditions. Herein we isolated preovulatory granulosa cells from wild-type (WT) and ERβ-null mice using laser capture microdissection to examine the genomic transcriptional response downstream of pregnant mare serum gonadotropin (mimicking FSH) and pregnant mare serum gonadotropin/human chorionic gonadotropin (mimicking LH) stimulation. This allows for a direct comparison of in vivo granulosa cells at the same stage of development from both WT and ERβ-null ovaries. ERβ-null granulosa cells showed altered expression of genes known to be regulated by FSH (Akap12 and Runx2) as well as not previously reported (Arnt2 and Pou5f1) in WT granulosa cells. Our analysis also identified 304 genes not previously associated with ERβ in granulosa cells. LH-responsive genes including Abcb1b and Fam110c show reduced expression in ERβ-null granulosa cells; however, novel genes including Rassf2 and Megf10 were also identified as being downstream of LH signaling in granulosa cells. Collectively, our data suggest that granulosa cells from ERβ-null ovaries may not be appropriately differentiated and are unable to respond properly to gonadotropin stimulation.

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Transcriptomic analysis of expression of genes regulating cell cycle progression in porcine ovarian granulosa cells during short-term in vitro primary culture

Histochemistry and Cell Biology ◽

10.1007/s00418-020-01860-2 ◽

2020 ◽

Vol 153 (6) ◽

pp. 397-412 ◽

Cited By ~ 2

Author(s):

Magdalena Kulus ◽

Wiesława Kranc ◽

Patrycja Sujka-Kordowska ◽

Piotr Celichowski ◽

Aneta Konwerska ◽

...

Keyword(s):

Cell Cycle ◽

Primary Culture ◽

Granulosa Cells ◽

Cell Cycle Progression ◽

Transcriptomic Analysis ◽

Short Term ◽

Cycle Progression ◽

Ovarian Granulosa Cells ◽

Expression Of Genes

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Correlations between Low Doses of Zearalenone, Its Carryover Factor and Estrogen Receptor Expression in Different Segments of the Intestines in Pre-Pubertal Gilts—A Study Protocol

Toxins ◽

10.3390/toxins13060379 ◽

2021 ◽

Vol 13 (6) ◽

pp. 379

Author(s):

Magdalena Gajęcka ◽

Magdalena Mróz ◽

Paweł Brzuzan ◽

Ewa Onyszek ◽

Łukasz Zielonka ◽

...

Keyword(s):

Estrogen Receptor ◽

Mrna Expression ◽

Estrogen Receptor Beta ◽

Receptor Expression ◽

Control Group ◽

Plant Materials ◽

Expression Of Genes ◽

Horizontal Part

Plant materials can be contaminated with Fusarium mycotoxins and their derivatives, whose toxic effects on humans and animals may remain subclinical. Zearalenone (ZEN), a low-molecular-weight compound, is produced by molds in crop plants as a secondary metabolite. The objective of this study will be to analyze the in vivo correlations between very low monotonic doses of ZEN (5, 10, and 15 μg ZEN/kg body weight—BW for 42 days) and the carryover of this mycotoxin and its selected metabolites from the intestinal contents to the intestinal walls, the mRNA expression of estrogen receptor alfa (ERα) and estrogen receptor beta (ERβ) genes, and the mRNA expression of genes modulating selected colon enzymes (CYP1A1 and GSTP1) in the intestinal mucosa of pre-pubertal gilts. An in vivo experiment will be performed on 60 clinically healthy animals with initial BW of 14.5 ± 2 kg. The gilts will be randomly divided into a control group (group C, n = 15) and three experimental groups (group ZEN5, group ZEN10, and group ZEN15; n = 15). Group ZEN5 will be administered per os 5 μg ZEN/kg BW (MABEL), group ZEN10—10 μg ZEN/kg BW (NOAEL), and group ZEN15—15 µg ZEN/kg BW (low LOAEL). In each group, five animals will be euthanized on analytical dates 1 (exposure day 7), 2 (exposure day 21), and 3 (exposure day 42). Samples for in vitro analyses will be collected from an intestinal segment resected from the following regions: the third (horizontal) part of the duodenum, jejunum, ileum, cecum, ascending colon, transverse colon, and descending colon. The experimental material will be collected under special conditions, and it will be transported to specialist laboratories where samples will be obtained for further analyses.

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Functional Implication of Netrin Expression in Malignant Melanoma

Analytical Cellular Pathology ◽

10.1155/2009/954172 ◽

2009 ◽

Vol 31 (6) ◽

pp. 415-422

Author(s):

Simone Kaufmann ◽

Silke Kuphal ◽

Thomas Schubert ◽

Anja K. Bosserhoff

Keyword(s):

Malignant Melanoma ◽

Melanoma Cells ◽

Transcriptional Level ◽

Altered Expression ◽

Invasion And Migration ◽

Expression Of Genes ◽

Functional Relevance ◽

And Migration

Background: Malignant melanoma cells are known to have altered expression of genes supporting proliferation and invasion, however, the expression of molecules of the Netrin family of repellent factors has not been analyzed in melanomas until now.Results: Here, we show that Netrin-1 expression is strongly induced in melanoma cells compared to melanocytes in vivo and in vitro controlled at the transcriptional level via ETS-1. In addition, the expression of the netrin receptor UNC5B was induced and that of UNC5C was reduced in the tumor cells. In order to determine the functional relevance of Netrin-1 expression in malignant melanoma, Netrin expression in melanoma cells was reduced by siRNA attempts and primary human melanocytes were treated with recombinant Netrin-1. The cells showed no changes in proliferation or apoptosis, however, a strong reduction of migratory properties was observed in the melanoma cells after reduction of Netrin expression whereas melanocyte migration was strongly induced by treatment with Netrin.Conclusions: Our study suggests that Netrin-1 promotes melanoma cell invasion and migration and therefore has an important role in the progression of malignant melanoma.

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The effect of pregnant mare serum gonadotropin on mouse embryos fertilized in vivo or in vitro

Journal of In Vitro Fertilization and Embryo Transfer ◽

10.1007/bf01133247 ◽

1986 ◽

Vol 3 (6) ◽

pp. 353-357 ◽

Cited By ~ 24

Author(s):

Fumihiko Sato ◽

Richard P. Marrs

Keyword(s):

Mouse Embryos ◽

Pregnant Mare Serum Gonadotropin ◽

Serum Gonadotropin

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Meat and Livestock Association Plenary Lecture 2005. Oocyte signalling molecules and their effects on reproduction in ruminants

Reproduction Fertility and Development ◽

10.1071/rd05104 ◽

2006 ◽

Vol 18 (4) ◽

pp. 403 ◽

Cited By ~ 34

Author(s):

Kenneth P. McNatty ◽

Stephen Lawrence ◽

Nigel P. Groome ◽

Mohammed F. Meerasahib ◽

Norma L. Hudson ◽

...

Keyword(s):

Granulosa Cells ◽

Point Mutations ◽

Follicular Development ◽

Ovis Aries ◽

Ovulation Rate ◽

Active Immunisation ◽

Signalling Molecules ◽

Stage Of Development

Sheep (Ovis aries) are a highly diverse species, with more than 900 different breeds that vary significantly in their physiological characteristics, including ovulation rate and fecundity. From examination of inherited patterns of ovulation rate, several breeds have been identified with point mutations in two growth factor genes that are expressed in oocytes. Currently, five different point mutations have been identified in the BMP15 (GDF9b) gene and one in GDF9. Animals heterozygous for the GDF9 and/or the BMP15 mutations have higher ovulation rates than their wild-type counterparts. In contrast, those homozygous for any of the aforementioned BMP15 or GDF9 mutations are sterile owing to arrested follicular development. In bovine and ovine ovaries, GDF9 was expressed exclusively in oocytes throughout follicular growth from the primordial stage of development, whereas in sheep BMP15 was expressed exclusively in oocytes from the primary stage: no data for the ontogeny of BMP15 expression are currently available for cattle. In vitro, ovine growth differentiation factor 9 (oGDF9) has no effect on 3H-thymidine incorporation by either bovine or ovine granulosa cells, whereas ovine bone morphogenetic protein 15 (oBMP15) has modest (1.2- to 1.6-fold; P < 0.05) stimulatory effects. Ovine GDF9 or oBMP15 alone inhibited progesterone production by bovine granulosa cells, whereas in ovine cells only oGDF9 was inhibitory. The effects of oGDF9 and oBMP15 together were often cooperative and not always the same as those observed for each factor alone. Active immunisation of ewes with BMP15 and/or GDF9 peptides affected ovarian follicular development and ovulation rate. Depending on the GDF9 and/or BMP15 vaccine formulation, ovulation rate was either increased or suppressed. A primary and single booster immunisation of ewes with a BMP15 peptide in a water-based adjuvant has led to 19–40% increases in lambs born per ewe lambing. Collectively, the evidence suggests that oocyte signalling molecules have profound effects on reproduction in mammals, including rodents, humans and ruminants. Moreover, in vivo manipulation of these oocyte signalling molecules provides new opportunities for the management of the fertility of ruminants.

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KDM5 inhibition offers a novel therapeutic strategy for the treatment of KMT2D mutant lymphomas

Blood ◽

10.1182/blood.2020008743 ◽

2021 ◽

Author(s):

James Andrew Heward ◽

Lola Koniali ◽

Annalisa D'Avola ◽

Karina Close ◽

Alison Yeomans ◽

...

Keyword(s):

Therapeutic Strategy ◽

Cell Signalling ◽

Loss Of Function ◽

H3k4 Methylation ◽

Altered Expression ◽

Bcl2 Family ◽

Expression Of Genes ◽

Altered Gene

Loss-of-function mutations in KMT2D are a striking feature of the germinal centre (GC) lymphomas, resulting in decreased H3K4-methylation and altered gene expression. We hypothesised that inhibition of the KDM5 family, which demethylates H3K4me3/me2, would re-establish H3K4-methylation and restore the expression of genes repressed upon loss of KMT2D. KDM5-inhibition increased H3K4me3 levels and caused an anti-proliferative response in vitro, which was markedly greater in both endogenous and CRISPR-edited KMT2D mutant DLBCL cell lines, while tumour growth was inhibited in KMT2D mutant xenografts in vivo. KDM5-inhibition reactivated both KMT2D-dependent and -independent genes, resulting in diminished B-cell signalling and altered expression of BCL2 family members, including BCL2 itself. KDM5-inhibition may offer an effective therapeutic strategy for ameliorating KMT2D loss-of-function mutations in GC-lymphomas.

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KDM5 inhibition offers a novel therapeutic strategy for the treatment of KMT2D mutant lymphomas

10.1101/2020.06.30.177477 ◽

2020 ◽

Author(s):

James A Heward ◽

Lola Konali ◽

Annalisa D’Avola ◽

Karina Close ◽

Alison Yeomans ◽

...

Keyword(s):

Therapeutic Strategy ◽

Cell Receptor ◽

Loss Of Function ◽

H3k4 Methylation ◽

Altered Expression ◽

Bcl2 Family ◽

Expression Of Genes ◽

Altered Gene

AbstractLoss-of-function mutations in KMT2D are a striking feature of the germinal centre (GC) lymphomas, resulting in decreased H3K4 methylation and altered gene expression. We hypothesised that inhibition of the KDM5 family, which demethylates H3K4me3/me2, would re-establish H3K4 methylation and restore the expression of genes repressed upon loss of KMT2D. KDM5-inhibition increased H3K4me3 levels and caused an anti-proliferative response in vitro, which was markedly greater in both endogenous and CRISPR-edited KMT2D mutant DLBCL cell lines, whilst tumour growth was inhibited in KMT2D mutant xenografts in vivo. KDM5-inhibition reactivated both KMT2D-dependent and -independent genes, resulting in diminished B-cell receptor signalling and altered expression of BCL2 family members, including BCL2 itself, allowing it to synergise with agents targeting these pathways. KDM5-inhibition may offer an effective therapeutic strategy for ameliorating KMT2D loss-of-function mutations in GC-lymphomas.Statement of significanceWe detail a novel way of reverting the effects of loss-of-function mutations in the histone methyltransferase KMT2D by inhibiting the KDM5 demethylase family, increasing levels of H3K4me3 and restoring expression of KMT2D regulated genes.

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