Temporal changes in miRNA abundance in the corpus luteum during early pregnancy
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ABSTRACT: Although rescue of the corpus luteum is required for pregnancy, luteal function during maternal recognition of pregnancy remains largely unexplored. CL were collected from pregnant cattle on days 14, 17, 20, and 23, to encompass the maternal recognition of pregnancy period. Nanostring technology was used to profile miRNA. A total of 27 miRNA changed. MiRNA that increased were predicted to inhibit phosphatidylinositol signaling, while those that decreased may be negative regulators of steroidogenesis. Overall, these data indicate that there are changes in the CL of pregnancy that are important for continued luteal function.
Project description:Although rescue of the corpus luteum is required for pregnancy, luteal function during maternal recognition of pregnancy remains largely unexplored. CL were collected from pregnant cattle on days 14, 17, 20, and 23, to encompass the maternal recognition of pregnancy period. Next Generation Sequencing was used to profile mRNA abundance during this time, while tandem mass spectrometry and Nanostring technology were used to profile proteins and miRNA, respectively. A total of 1157 mRNA were differentially abundant. mRNA that increased were regulators of interferon signaling and DNA repair, while those that decreased were associated with luteolytic processes, such as calcium signaling and matrix metallopeptidase (MMP) signaling, indicating inhibition of these processes. mRNA that were maximally abundant on day 20 were primarily associated with immune processes. Overall, these data indicate that there are changes in the CL of pregnancy that are important for continued luteal function.
Project description:To evaluate functional changes of the corpus luteum (CL) during early pregnancy in cows, gene expression profiles of the CL at the time of maternal recognition were investigated. Microarray analysis, using a 15 K bovine oligo DNA microarray, demonstrated 30 and 266 differentially expressed genes in the CL on days 15 (P15) and 18 (P18) of pregnancy compared with the CL on day 15 (NP15) of non-pregnancy (n=4 for each group, >2-fold change relative to NP15; P<0.05).
Project description:The absence of luteolytic signal on non-pregnant bitches leads to the existence of a physiological pseudopregnancy maintained by a long lasting luteal function. During the diestrus, hormonal regulation of the canine CL changes. While in later stages prolactin is the main luteotropic hormone, in earlier stages the CL is independent from hypophysary hormones. At this time, prostaglandins are among the main luteotropic factors. In the present project, the aim was to further understand the effects of prostaglandins withdrawal on luteal function. In the present project, next generation sequencing (NGS), RNA-seq, was applied to explore the modulatory role of prostaglandins in the canine corpus luteum (CL) during the first half of diestrus. For this, transcriptome analysis of genes differently expressed in the CL of bitches treated in vivo with a COX2 inhibitor (Previcox, Merial) was performed. Additionally, by using just control samples, effects dependent on time were also explored and used as primary validation of results. Higher represented differentially expressed genes (DEG, p<0.01, FDR<0.1) in mature CL (days 20 and 30) referred to steroidogenesis, while in early CL (days 5 and 10) to proliferation and immune system. Then, treatment effects were investigated at each time point. No gene was concomitantly affected in all investigated groups. Thus, clearly, the effects were dioestrus stage-dependent. Higher numbers of DEG were found on day 20 (n=1741), mainly related to increased immune function, while on day 30 (n=552) they were related to decreased steroidogenesis and vascularization. Low numbers of DEG were found in early CL. Our results suggest the presence of strong compensatory effects in the early CL and multidirectional effects towards luteal gonadotropin-dependency after COX2-inhibition.
Project description:The corpus luteum (CL) is essential for maintenance of pregnancy in all mammals and luteal rescue, which occurs around day 16-19 in the cow, is necessary to maintain luteal progesterone production. Transcriptomic and proteomic profiling were performed to compare the day 17 bovine CL of the estrous cycle and pregnancy. Among mRNA and proteins measured, 140 differentially abundant mRNA and 24 differentially abundant proteins were identified. Pathway analysis was performed using four programs. Modulated pathways included T cell receptor signaling, vascular stability, cytokine signaling, and extracellular matrix remodeling. Two mRNA that were less in pregnancy were regulated by prostaglandin (PG) F2A in culture, while two mRNA that were greater in pregnancy were regulated by interferon tau (IFNT). To identify mRNA that could be critical regulators of luteal fate, the mRNA that were differentially abundant during early pregnancy were compared to mRNA that were differentially abundant during luteal regression. Eight mRNA were common to both datasets, including mRNA related to regulation of steroidogenesis and gene transcription. A subset of differentially abundant mRNA and proteins, including those associated with extracellular matrix functions, were predicted targets of differentially abundant microRNA (miRNA). Integration of miRNA and protein data, using miRPath, revealed pathways such as extracellular matrix-receptor interactions, abundance of glutathione, and cellular metabolism and energy balance. Overall, this study has provided a comprehensive profile of molecular changes in the corpus luteum during maternal recognition of pregnancy and has indicated that some of these functions may be miRNA-regulated.
Project description:Corpus luteum (CL) is an ephemeral gland whose main function is to secrete progesterone required for the establishment and maintenance of pregnancy. It is very well established that development and maintenance of CL function in primates requires action of luteinizing hormone (LH) but the extent and mechanism by which LH contributes to the maintenance of CL function through out the luteal phase is not known. To study the nuclear actions mediated by LH, we evaluated global genomic changes in CL of monkeys treated with GnRH receptor antagonist to inhibit pituitary LH secretion. Affymetrix microarray analysis was performed on RNA samples from CL obtained from VEH or CET treated monkeys. Results demonstrate that LH regulates expression of a number of genes which might be important for maintenance of CL structure and function. Keywords: CL, LH, CET, gene expression
Project description:Corpus luteum (CL) is an ephemeral gland whose main function is to secrete progesterone required for the establishment and maintenance of pregnancy. It is very well established that development and maintenance of CL function in primates requires action of luteinizing hormone (LH) but the extent and mechanism by which LH contributes to the maintenance of CL function through out the luteal phase is not known. To study the nuclear actions mediated by LH, we evaluated global genomic changes in CL of monkeys treated with GnRH receptor antagonist to inhibit pituitary LH secretion. Affymetrix microarray analysis was performed on RNA samples from CL obtained from VEH or CET treated monkeys. Results demonstrate that LH regulates expression of a number of genes which might be important for maintenance of CL structure and function. Keywords: CL, LH, CET, gene expression A single s.c. injection of GnRH receptor antagonist, Cetrorelix (CET), administered at a dose of 150 µg/kg BW has been shown to induce luteolysis. For the purpose of microarray analysis, VEH (5.25% glucose) or CET (treatment) was injected to female monkeys on day 7 of luteal phase and CL collected at 24 h post treatment (n=3) was cut into small quarters and snap frozen in liquid nitrogen. To minimize between animal variations, CL for both VEH and CET treatments were collected from the same monkeys.
Project description:To explore chorionic gonadotropin (CG)-regulated gene expression in the primate corpus luteum (CL), adult female rhesus macaques were treated with a model of simulated early pregnancy (SEP). Total RNA was isolated from individual CL and hybridized to AffymetrixM-bM-^DM-" GeneChip Rhesus Macaque Genome Arrays The level of 1192 transcripts changed expression > 2-fold (one-way ANOVA, FDR correction; P<0.05) during SEP when compared to Day 10 untreated controls, and the majority of changes occurred between Days 10 and 12 of SEP. To compare transcript levels between SEP rescued and regressing CL, previously banked rhesus GeneChip array data from the mid- to late and very late luteal phase were analyzed with time-matched intervals in SEP. Comparing RMA-normalized transcripts from the natural cycle with those from luteal rescue revealed 7677 transcripts changing in expression pattern >2 fold (one-way ANOVA, FDR correction; P<0.05) between the two groups. Clustering of samples revealed that the SEP samples possessed the most related transcript expression profiles. Regressed CL (days 18-19, around menses) were the most unlike all other CL. The most affected KEGG pathway was Steroid Biosynthesis, and most significantly absent pathways following SEP treatment includes groups of genes whose products promote cell-death. By further comparing the genome-wide changes in luteal gene expression during rescue in SEP, with those in CL during luteolysis in the natural menstrual cycle, it is possible to identify key regulatory pathways promoting fertility. Simulated early pregnancy (SEP) treatment was begun on day 9 as Duffy and Stouffer (1997) by treatment of females with recombinant human chorionic gonadotropin (hCG; NovarelM-bM-^DM-", Ferring Pharmaceuticals Inc. Parsippany, NJ, USA) in increasing dosages (15, 30,45,90,180,360,720,1440, and 2880 IU) twice daily by intramuscular injection. CL were collected by laparotomy on days 10, 12, 15, and 18, representing 1, 3, 6 and 9 days of hCG treatment (n=4 CL/day). Additionally, luteal day 10 untreated CL were collected to serve as baseline controls for SEP CL. All CL were dissected away from luteal tissue, sectioned, and snap-frozen in liquid nitrogen and stored at -80M-BM-0C until RNA and protein isolation by TRIzolM-BM-. extraction (Invitrogen, Carlsbad, CA, USA) according to manufacturerM-bM-^@M-^Ys protocols.
Project description:Recently, microRNAs (miRNAs) have emerged as new players in the fine tuning of some reproductive functions in mammals via posttranscriptional gene regulation mechanisms. Importantly, miRNAs have been suggested to be an important regulators of various ovarian functions. Applying custom made multispecies arrays we aimed to analyze expression profile of miRNAs in corpus luteum to answer the question whether miRNAs can be involved in maintenance of luteal function during early pregnancy in pigs.
Project description:To explore chorionic gonadotropin (CG)-regulated gene expression in the primate corpus luteum (CL), adult female rhesus macaques were treated with a model of simulated early pregnancy (SEP). Total RNA was isolated from individual CL and hybridized to Affymetrix™ GeneChip Rhesus Macaque Genome Arrays The level of 1192 transcripts changed expression > 2-fold (one-way ANOVA, FDR correction; P<0.05) during SEP when compared to Day 10 untreated controls, and the majority of changes occurred between Days 10 and 12 of SEP. To compare transcript levels between SEP rescued and regressing CL, previously banked rhesus GeneChip array data from the mid- to late and very late luteal phase were analyzed with time-matched intervals in SEP. Comparing RMA-normalized transcripts from the natural cycle with those from luteal rescue revealed 7677 transcripts changing in expression pattern >2 fold (one-way ANOVA, FDR correction; P<0.05) between the two groups. Clustering of samples revealed that the SEP samples possessed the most related transcript expression profiles. Regressed CL (days 18-19, around menses) were the most unlike all other CL. The most affected KEGG pathway was Steroid Biosynthesis, and most significantly absent pathways following SEP treatment includes groups of genes whose products promote cell-death. By further comparing the genome-wide changes in luteal gene expression during rescue in SEP, with those in CL during luteolysis in the natural menstrual cycle, it is possible to identify key regulatory pathways promoting fertility.
Project description:In monoovulatory species like bovines, a striking diversity in regulation of the corpus luteum (CL) function exists within species during different stages of the luteal phase. The function and life span of CL appears to be regulated by the interplay of both luteotrophic and luteolytic factors, whose roles and actions are varied from one species to another. Although, studies continue to expand our current understanding of the cellular and molecular actions of prostaglandin F 2alpha (PGF2α - a physiological luteolysin)-induced luteolysis, knowledge of the mechanism whereby, PGF2α mediates its luteolytic actions remains poorly understood. The mechanism of PGF2α-induced luteolysis is a complex process involving changes in expression of multiple genes encoding gene products that regulate steroidogenesis, stress related or apoptotic genes, factors involved in immune response and enzymes that stimulate PGF2α production. Thus, in the present study, efforts were made to identify the temporal changes in the global gene expression profile in the CL of buffalo cows in response to PGF2α treatment. The molecular signature of genome-wide transcriptional changes in the CL tissue subjected to PGF2α-induced luteolysis will expand our knowledge on basic mechanism/s that regulates the luteolytic process. The results obtained in this study provide insight into the mechanisms underlying the PGF2α- induced regression of CL.Keywords: CL, PGF2α, gene expression