High environmental temperatures induce detrimental effects on various reproductive processes in cattle. According to the predicted global warming the number of days with unfavorable ambient temperatures will further increase. The objective of this study was to investigate effects of acute heat stress during the late pre-ovulatory phase on morphological, physiological and molecular parameters of dominant follicles in cycling cows during lactation. Eight German Holstein cows in established lactation were exposed to heat stress (28C) or thermoneutral conditions (15C) with pair-feeding for four days. After synchronization growth of dominant follicles was monitored by ultrasonogrphy, and 21 hrs after an induced pre-ovulatory LH surge antral steroid hormones and granulosa cell-specific gene expression profiles were determined. The data showed that the pre-ovulatory growth of dominant follicles and the estradiol, but not the progesterone concentrations tended to be slightly affected. mRNA microarray and hierarchical cluster analysis revealed distinct expression profiles in granulosa cells derived from heat stressed compared to pair-fed animals. Among the 255 affected genes heatstress-, stress- or apoptosis associated genes were not present. But instead, we found up-regulation of genes essentially involved in G-protein coupled signaling pathways, extracellular matrix composition, and several members of the solute carrier family as well as up-regulation of FST encoding follistatin. In summary, the data of the present study show that acute pre-ovulatory heat stress can specifically alter gene expression profiles in granulosa cells, however without inducing stress related genes and pathways and suggestively can impair follicular growth due to affecting the activin-inhibin-follistatin system.
Exposure of Lactating Dairy Cows to Acute Pre-Ovulatory Heat Stress Affects Granulosa Cell-Specific Gene Expression Profiles in Dominant Follicles.
Specimen part
View SamplesThe molecular mechanisms that regulate the pivotal transformation processes observed in the follicular wall following the pre-ovulatory LH-surge, are still not established, particularly for cells of the thecal layer. To elucidate thecal and granulosa cell type-specific biological functions and signaling pathways, large dominant bovine follicles were collected before and 21 hrs after an exogenous GnRH induced LH surge. Because LH receptor density varies within the granulosa cell populations, antral granulosa (aGC; those aspirated by follicular puncture) and membrane associated granulosa (mGC; those scraped from the follicular wall) were compared to thecal cell expression profiles determined by mRNA microarrays. Thecal cell gene expression was less affected in the peri-ovulatory follicle when compared to granulosa cells, as evidenced by only 2% versus 25% of the ~11,000 genes expressed changing in response to the LH surge, respectively. The majority of the 203 LH-regulated thecal genes were also LH regulated in granulosa cells, leaving a total of 58 genes as LH-regulated theca cell specific genes. Most of the 58 genes (i.e., 74%) thecal specific genes including several known thecal markers (CYP17A1, NR5A1) were downregulated, while most genes identified are new to theca. Many of the newly identified upregulated thecal genes (e.g., PTX3, RND3, PPP4R4) were also upregulated in granulosa. Minimal expression differences were observed between aGC and mGC, however, transcripts encoding extracellular proteins (NID2) and matrix modulators (ADAMTS1, SASH1) predominated these differences. We also identified large numbers of unknown LH-regulated granulosa cell genes and discuss their putative roles in ovarian function.
Research resource: preovulatory LH surge effects on follicular theca and granulosa transcriptomes.
Specimen part
View SamplesHuman ILCs are classically categorized into five subsets; cytotoxic CD127-CD94+ NK cells and non-cytotoxic CD127+CD94-, ILC1s, ILC2s, ILC3s and LTi cells. Here, we identify a novel subset within the CD127+ ILC population, characterized by the expression of the cytotoxic marker CD94. These CD94+ ILCs strongly resemble conventional ILC3s in terms of phenotype, transcriptome and cytokine production, but are highly cytotoxic. IL-15 was unable to induce differentiation of CD94+ ILCs towards mature NK cells. Instead, CD94+ ILCs retained RORγt, CD127 and CD200R expression and produced IL-22 in response to IL-15. Culturing non-cytotoxic CD127+ ILC1s or ILC3s with IL-12 induced upregulation of CD94 and cytotoxic activity, effects that were not observed with IL-15 stimulation. Thus, human helper ILCs can acquire a cytotoxic program without differentiating into NK cells.
Identification of human cytotoxic ILC3s.
Specimen part, Subject
View SamplestPTEN-/- mice display a deletion of the PTEN tumor suppressor gene specifically in T cells (cross PTEN flox/flox x lck-Cre). They develop T cell lymphoma with a primary thymic tumor and invasion of most organ at late stage of the disease.
Pharmacological inhibition of carbonic anhydrase XII interferes with cell proliferation and induces cell apoptosis in T-cell lymphomas.
Specimen part, Disease, Disease stage
View SamplesAnalysis of ventricular derived mRNA from Med1fl/fl and Med1fl/fl cardiac knockout mice. Results provide insight into the molecual rmechanisms underlying dilated cardiomyopathy. Overall design: Methods: Ventricular samples (4 per group) from 21-day-old Med1fl/fl and Med1 cardiac knockout mice were used to generate polyA enriched stranded RNA libraries followed by RNAseq using the Illumina HiSeq platform. Raw sequence reads were analyzed with BaseSpace (www.illumina.com) by aligning reads to the mus musculus mm10 genome using the TopHat Alignment app. Transcripts were assembled and significant differentially expressed genes were determined with the Cufflinks Assembly and DE app using a false discovery rate <0.05. qRT–PCR validation was performed using SYBR Green assays
Cardiac Med1 deletion promotes early lethality, cardiac remodeling, and transcriptional reprogramming.
Specimen part, Subject
View SamplesThe formation of neuronal connections requires the precise guidance of developing axons towards their targets. In the Drosophila visual system, photoreceptor neurons (R cells) project from the eye into the brain. These cells are grouped into some 750 clusters comprised of eight photoreceptors or R-cells each. R cells fall into three classes, R1-R6, R7 and R8. Posterior R8 cells are the first to project axons into the brain. How these axons select a specific pathway is not known.
Robo-3--mediated repulsive interactions guide R8 axons during Drosophila visual system development.
Specimen part
View SamplesContext: Endometrium in polycystic ovary syndrome (PCOS) presents altered gene expression indicating progesterone resistance and predisposing to reduced endometrial receptivity and endometrial cancer. Objective: We hypothesized that an altered endocrine/metabolic environment in PCOS may result in an endometrial disease phenotype affecting the gene expression of different endometrial cell populations, including stem cells and their differentiated progeny. Design and setting: A prospective study conducted at an academic medical center. Patients and Main Outcome Measures: Proliferative phase endometrium was obtained from 6 overweight/obese PCOS (NIH criteria) and 6 overweight/obese controls. Microarray analysis was performed on fluorescence-activated cell sorting (FACS)-isolated endometrial epithelial cells (eEP), endothelial cells (eEN), stromal fibroblasts (eSF) and mesenchymal stem cells (eMSC). Gene expression data were validated using microfluidic Q-RT-PCR and immunohistochemistry (IHC). Results: The comparison between eEPPCOS and eEPCtrl showed dysregulation of inflammatory genes and genes with oncogenic potential (CCL2, IL-6, ORM1, TNAIFP6, SFRP4, SPARC). eSFPCOS and eSFCtrl showed upregulation of inflammatory genes (C4A/B, CCL2, ICAM1, TNFAIP3). Similarly, in eMSCPCOS vs. eMSCCtrl the most upregulated genes were related to inflammation and cancer (IL-8, ICAM1, SPRR3, LCN2). IHC scoring showed increased expression of CCL2 in eEPPCOS and eSFPCOS compared to eEPCtrl and eSFCtrl and IL-6 in eEPPCOS compared to eEPCtrl. Conclusions: Isolated endometrial cell populations in women with PCOS showed altered gene expression revealing inflammation and pro-oncogenic changes, independent of BMI, especially in eEPPCOS and eMSCPCOS, compared to controls. The study reveals an endometrial disease phenotype in women with PCOS with potential negative effects on endometrial function and long-term health.
Mesenchymal stem/progenitors and other endometrial cell types from women with polycystic ovary syndrome (PCOS) display inflammatory and oncogenic potential.
Sex, Specimen part
View SamplesTo investigate whether and how expression of the oncogenic transcription factor EVI1 influences gene regulation by phorbol esters and vice versa, the human myeloid cell line U937 was transduced with an EVI1 expression vector or empty vector as a control. Cells were treated with 12-Otetradecanoylphorbol 13-acetate (TPA) or its solvent ethanol as a control. RNA was extracted and subjected to gene expression microarray analysis.
The oncogene EVI1 enhances transcriptional and biological responses of human myeloid cells to all-trans retinoic acid.
Cell line
View SamplesThe product of the ecotropic virus integration site 1 (EVI1) gene, whose overexpression is associated with a poor prognosis in myeloid leukemias and some epithelial tumors, regulates gene transcription both through direct DNA binding and through modulation of the activity of other sequence specific transcription factors. Previous results from our laboratory have shown that EVI1 influenced transcription regulation in response to the myeloid differentiation inducing agent, all-trans retinoic acid (ATRA), in a dual manner: it enhanced ATRA induced transcription of the RARb gene, but repressed the ATRA induction of the EVI1 gene itself. In the present study, we asked whether EVI1 would modulate the ATRA regulation of a larger number of genes, as well as biological responses to this agent, in human myeloid cells. U937 and HL-60 cells ectopically expressing EVI1 through retroviral transduction were subjected to microarray based gene expression analysis, and to assays measuring cellular proliferation, differentiation, and apoptosis. These experiments showed that EVI1 modulated the ATRA response of several dozens of genes, and in fact reinforced it in the vast majority of cases. A particularly strong synergy between EVI1 and ATRA was observed for GDF15, which codes for a member of the TGF-b superfamily of cytokines. In line with the gene expression results, EVI1 enhanced cell cycle arrest, differentiation, and apoptosis in response to ATRA, and knockdown of GDF15 counteracted some of these effects.
The oncogene EVI1 enhances transcriptional and biological responses of human myeloid cells to all-trans retinoic acid.
Cell line
View SamplesThe product of the ecotropic virus integration site 1 (EVI1) gene, whose overexpression is associated with a poor prognosis in myeloid leukemias and some epithelial tumors, regulates gene transcription both through direct DNA binding and through modulation of the activity of other sequence specific transcription factors. Previous results from our laboratory have shown that EVI1 influenced transcription regulation in response to the myeloid differentiation inducing agent, all-trans retinoic acid (ATRA), in a dual manner: it enhanced ATRA induced transcription of the RARb gene, but repressed the ATRA induction of the EVI1 gene itself. In the present study, we asked whether EVI1 would modulate the ATRA regulation of a larger number of genes, as well as biological responses to this agent, in human myeloid cells. U937 and HL-60 cells ectopically expressing EVI1 through retroviral transduction were subjected to microarray based gene expression analysis, and to assays measuring cellular proliferation, differentiation, and apoptosis. These experiments showed that EVI1 modulated the ATRA response of several dozens of genes, and in fact reinforced it in the vast majority of cases. A particularly strong synergy between EVI1 and ATRA was observed for GDF15, which codes for a member of the TGF-b superfamily of cytokines. In line with the gene expression results, EVI1 enhanced cell cycle arrest, differentiation, and apoptosis in response to ATRA, and knockdown of GDF15 counteracted some of these effects.
The oncogene EVI1 enhances transcriptional and biological responses of human myeloid cells to all-trans retinoic acid.
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