Effect of Estrous Cycle Phases on Gene Expression of Bovine Oviduct Epithelial Cells

2021 ◽  
Author(s):  
Ricaurte Lopera Vasquez ◽  
Fabián Uribe-García ◽  
Iang Rondón-Barragán
Keyword(s):  
Gene Expression ◽  
Epithelial Cells ◽  
Estrous Cycle ◽  
Oviduct Epithelial Cells

scholarly journals 230TRANSCRIPTOMICS ANALYSIS OF CHANGES IN GENE EXPRESSION IN BOVINE OVIDUCT EPITHELIAL CELLS DURING THE ESTROUS CYCLE

2004 ◽  
Vol 16 (2) ◽  
pp. 236
Author(s):  
S. Bauersachs ◽  
S. Rehfeld ◽  
S. Koelle ◽  
S. Mallok ◽  
K. Prelle ◽  
...  
Keyword(s):  
Gene Expression ◽  
Epithelial Cells ◽  
Estrous Cycle ◽  
Cdna Array ◽  
Cdna Libraries ◽  
Cdna Clones ◽  
Ciliated Cells ◽  
Expression Levels ◽  
Functional Changes ◽  
Oviduct Epithelial Cells

The oviduct epithelium undergoes marked morphological and functional changes during the estrous cycle. It has been shown that a dramatic change in the frequencies of ciliated and non-ciliated cells occurs during the estrous cycle. At estrus the epithelium consists of secretory and ciliated cells and at diestrus mainly of ciliated cells. The oviduct provides the microenvironment for sperm capacitation, fertilization, and early cleavage-stage embryonic development. At the molecular level, only a few genes or proteins are known that change during the estrous cycle and which may be important for fertility, so as the bovine oviduct-specific glycoprotein, the major secretory protein in the oviduct. Therefore, we studied systematically the changes in gene expression in bovine ipsilateral oviduct epithelial cells at estrus and diestrus. To identify differentially expressed genes, a combination of subtracted cDNA libraries and cDNA array hybridization was used. Two subtracted libraries were produced to enrich cDNAs of upregulated genes at estrus and at diestrus. A total of 1536 cDNA clones of each library were analyzed with radioactively (33-P) labeled probes generated from the oviduct epithelial cells of six Simmental heifers, three of them slaughtered at Day 0 (estrus) and three at Day 12 after standing heat (diestrus). After normalization of the raw data and statistical analysis, all cDNAs showing significant differences in their expression levels at estrus compared to diestrus were sequenced. Sequencing revealed 84 different cDNAs;; 42 of them matched bovine genes or their human/mouse homologs with known functions, and 42 matched genes without a known function. Half of the genes (n=42) were expressed at a higher level at estrus;; for the other (n=42) expression levels were higher at diestrus. The regulated genes or their products represented a variety of functional classes, such as genes of the secretory pathway, genes involved in transcription regulation, cell-surface proteins, cell–cell interaction proteins, secreted proteins, members of signal transduction pathways, immune-related proteins, and some enzymes. The identification of genes differentially regulated in ipsilateral oviduct epithelial cells at estrus v. diestrus is the first step of a systematic analysis of differential gene expression during the estrous cycle. Further studies will follow, focusing on different compartments of the bovine oviduct and additional times of the estrous cycle.

scholarly journals Monitoring gene expression changes in bovine oviduct epithelial cells during the oestrous cycle

2004 ◽  
Vol 32 (2) ◽  
pp. 449-466 ◽  
Author(s):  
S Bauersachs ◽  
S Rehfeld ◽  
SE Ulbrich ◽  
S Mallok ◽  
K Prelle ◽  
...  
Keyword(s):  
Gene Expression ◽  
Epithelial Cells ◽  
Expression Profiles ◽  
Cdna Array ◽  
Oestrous Cycle ◽  
Cdna Clones ◽  
Rt Pcr ◽  
Expression Levels ◽  
Starting Point ◽  
Oviduct Epithelial Cells

The oviduct epithelium undergoes marked morphological and functional changes during the oestrous cycle. To study these changes at the level of the transcriptome we did a systematic gene expression analysis of bovine oviduct epithelial cells at oestrus and dioestrus using a combination of subtracted cDNA libraries and cDNA array hybridisation. A total of 3072 cDNA clones of two subtracted libraries were analysed by array hybridisation with cDNA probes derived from six cyclic heifers, three of them slaughtered at oestrus and three at dioestrus. Sequencing of cDNAs showing significant differences in their expression levels revealed 77 different cDNAs. Thirty-seven were expressed at a higher level at oestrus, for the other 40 genes expression levels were higher at dioestrus. The identified genes represented a variety of functional classes. During oestrus especially genes involved in the regulation of protein secretion and protein modification, and mRNAs of secreted proteins, were up-regulated, whereas during dioestrus particularly transcripts of genes involved in transcription regulation showed a slight up-regulation. The concentrations of seven selected transcripts were quantified by real-time RT-PCR to validate the cDNA array hybridisation data. For all seven transcripts, RT-PCR results were in excellent correlation (r>0.92) with the results obtained by array hybridisation. Our study is the first to analyse changes in gene expression profiles of bovine oviduct epithelial cells during different stages of the oestrous cycle, providing a starting point for the clarification of the key transcriptome changes in these cells.

scholarly journals 231 TEMPORAL AND SPATIAL GENE EXPRESSION ANALYSIS OF THE BOVINE OVIDUCT EPITHELIUM

2005 ◽  
Vol 17 (2) ◽  
pp. 266
Author(s):  
S. Rehfeld ◽  
S. Bauersachs ◽  
H. Blum ◽  
S. Mallok ◽  
H. Wenigerkind ◽  
...  
Keyword(s):  
Gene Expression ◽  
Epithelial Cells ◽  
Estrous Cycle ◽  
Secretory Pathway ◽  
Separate Analysis ◽  
Physiological Changes ◽  
Immune Functions ◽  
Tissue Samples ◽  
Oviduct Epithelium ◽  
Maternal Communication

The fallopian tube plays a central role in reproduction, providing the appropriate environmental conditions for oocyte maturation as well as for sperm capacitation. Furthermore, fertilization and the first cleavage stages of embryonic development take place in the oviduct. At the molecular level, only fragmentary data are available regarding the physiological changes in the oviduct epithelium during the estrous cycle. Therefore, we performed a systematic study of gene expression changes in bovine ipsilateral oviduct epithelial cells derived from either the ampulla or the isthmus part of the oviduct at four different time points of the estrous cycle. A cDNA array consisting of approximately 400 candidate genes, primarily identified in different studies in the context of gene expression regulation in the oviduct, was designed and hybridized with 33P-labeled cDNA probes prepared from 28 different tissue samples. These tissue samples were collected from cyclic Simmental heifers at Day 0 (n = 3), Day 3.5 (n = 3), Day 12 (n = 4) and Day 18 (n = 4) of the estrous cycle. Ipsilateral epithelial cells were separately collected from ampulla and isthmus. After array evaluation (AIDA Image Analyzer, version 3.41, Raytest, Straubenhardt, Germany), the raw data were normalized to internal reference cDNAs on the arrays. Statistical analysis was done using ANOVA and the Tukey post-hoc test (GeneSpring® version 6.1, Silicon Genetics, Redwood City, CA, USA). For selected genes, differential expression was verified by real-time RT-PCR. A simplified Gene Ontology was built for the genes present on the array and a pathway analysis was performed to elucidate gene networks involved in the regulation of oviduct epithelial cell function. The expression patterns of two functional groups of genes are presented here: genes that are related to immune functions and genes of the secretory pathway or encoding secreted proteins. Messenger-RNA levels of immune-related genes were higher in epithelial cells of the ampulla compared to the isthmus part of the oviduct. This implies that certain immune functions may be differentially regulated in ampulla and isthmus. Furthermore, mRNAs of genes of the secretory pathway showed highest levels mainly in the ampulla around time of estrus, which may be explained with the increase of the secretory activity in the epithelium of the ampulla beginning at pre-estrus. In general, this study shows the importance of a separate analysis of the oviduct compartments and the influence of the estrous cycle on the expression level of a variety of genes. In the context of fertilization and early embryo-maternal communication, these results could provide an insight into the physiological changes during the estrous cycle, which are the bases for these processes. This work was supported by the Deutsche Forschungsgemeinschaft (Research Unit “Mechanisms of Embryo-Maternal Communication”; FOR 478/1).

scholarly journals 329 A NOVEL SUSPENSION CULTURE SYSTEM FOR BOVINE OVIDUCT EPITHELIAL CELLS

2005 ◽  
Vol 17 (2) ◽  
pp. 315
Author(s):  
R. Rottmayer ◽  
S.E. Ulbrich ◽  
S. Koelle ◽  
K. Prelle ◽  
H.H.D. Meyer ◽  
...  
Keyword(s):  
Gene Expression ◽  
Epithelial Cells ◽  
Suspension Culture ◽  
Culture System ◽  
Cell Counting ◽  
Blue Staining ◽  
Significant Difference ◽  
Oviduct Epithelial Cells ◽  
Maternal Communication

Bovine oviduct epithelial cells (BOEC) are difficult to culture without dedifferentiation. Frequently observed changes in morphology during culture suggest that gene expression patterns are affected as well. We explored a novel short-term culture system for BOEC – suitable for co-culture experiments with embryos – and evaluated the cells with respect to morphological criteria, candidate gene expression, and hormone responsiveness. Simmental heifers were slaughtered on Day 3.5 after standing heat and BOEC were obtained by squeezing along the ampulla with forceps. The cell sheets were separated mechanically by repeated passages through syringes and pipetting, and recovered by sedimentation. Cells from the ipsi- and contralateral oviduct were cultured separately at a density of 106 cells per well in 24-well plates with 800 μL TCM-199 supplemented with 2% OCS (estrous cow serum, as used in embryo culture) or CS 3.5 (cow serum, Day 3.5 after standing heat, adequate to the cycle stage in which cells were obtained) and 0.25 mg/mL gentamicin. For cell counting, an aliquot was further disaggregated by passing 15 times through a 30-gauge needle to achieve a single-cell suspension. Culture took place at 38°C in a humidified atmosphere of 5% CO2 in air. Cells were examined by light microscopy at seeding and after 6, 12, 24, and 48 h. Cell aggregates showed a worm-like structure, displaying numerous vigorously beating cilia on their surfaces throughout the culture period. Trypan blue staining indicated that cells contained in aggregates were viable while single cells stained predominantly positive (non-viable). The purity of the epithelial cell culture was >95%, as determined by immunohistochemistry using antibodies against vimentin and cytokeratin. For electron microscopic investigations, BOEC were sampled at seeding and after 24 h in culture. Cultured BOEC showed a morphology highly similar to that of BOEC in vivo. Both secretory cells with numerous secretory granules and mitochondria, and ciliated cells with long, well developed and actively moving kinocilia were visible. RT-PCR data for candidate genes (ERα, ERβ, HMGCR, PHGPx, PR) obtained from BOEC samples at seeding and after 6, 12, 24, and 48 h in culture showed that gene expression was stable for the majority of transcripts after 6 h in culture. There was no significant difference between cells cultured with OCS or CS 3.5 and no difference between cells obtained from the ipsi- or contralateral oviduct. Estradiol-17β (E2, 10 pg/mL) or progesterone (P4, 10 ng/mL) stimulation showed that the cultured BOEC are able to respond to hormonal signals in a manner similar to their reaction in vivo (Ulbrich et al. 2003 J. Steroid. Biochem. Mol. Biol. 84, 279–89). Progesterone receptor mRNA was up-regulated by E2 and estrogen receptor β mRNA was up-regulated by P4. The culture system for bovine oviduct epithelial cells thus provides an adequate tool to investigate mechanisms of the embryo-maternal communication in cattle. This work was supported by the Deutsche Forschungsgemeinschaft (Research Unit “Mechanisms of Embryo-Maternal Communication”; FOR 478/1).

278: Gene Expression Profiling of Prostatic Epithelial Cells in Two-Dimensional Versus Three-Dimensional Cultures

2007 ◽  
Vol 177 (4S) ◽  
pp. 93-93
Author(s):  
Toshiyuki Tsunoda ◽  
Junichi Inocuchi ◽  
Darren Tyson ◽  
Seiji Naito ◽  
David K. Ornstein
Keyword(s):  
Gene Expression ◽  
Epithelial Cells ◽  
Gene Expression Profiling ◽  
Expression Profiling ◽  
Three Dimensional ◽  
Two Dimensional
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