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GSE3224
Mus musculus
9 Downloadable Samples
Affymetrix Mouse Expression 430A Array (moe430a)
Description
C2C12 myoblasts were infected with a retrovirus expressing Pax7d or with an empty virus (puro) as a control. All of the samples originated from the same common pool of parental C2C12. This pool was split into six streams. A single prep of Pax7d-puro virus was split into three volumes and used to infect three of the streams. A single prep of puro-alone virus was similarly split in three and used to infect the remaining three streams. From the point of the infection forward each stream was maintained distinct from the others. Cells were infected and grown simultaneously under identical conditions.
Publication Title
Pax7 activates myogenic genes by recruitment of a histone methyltransferase complex.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 4 Downloadable Samples
- Affymetrix Mouse Gene 2.0 ST Array (mogene20st)
Description
Besides symptoms caused by central nervous system (CNS) lesions, the majority of patients with multiple sclerosis (MS) also exhibit gastrointestinal dysfunction that has frequently been noted, but was not directly linked to the autoimmune etiology of the disease.We studied the enteric nervous system (ENS) in a murine model of MS by histology and electron microscopy. Serum IgG against enteric neurons and enteroglia was measured by ELISA and binding to the ENS was confirmed by immunohistochemistry. Target antigens were identified by mass spectrometry. Gastrointestinal dysfunction was determined by measuring dye transit time. RNA expression profiling was conducted with small intestines of MP4-immunized and control-immunized mice. Data from the mouse model were confirmed in MS patients by immunohistochemistry of the ENS in bowel resectates. In addition, ELISA was performed on plasma samples to detect antibodies against four specific target antigens as identified in the mouse model. ENS degeneration was evident already before the onset of clinical disease in the mouse model. Pathology was predominantly antibody-mediated and caused a significant decrease in gastrointestinal transit, which was associated with severe gliosis of the ENS. Unlike the dense infiltrates that developed in the perivascular compartments of the CNS of MP4-immunized mice, the infiltrates in the ENS consisted of single cells scattered throughout the tissue. RNA expression profiling could support these results, as the expression of inflammatory markers in the small intestine was similar between MP4-immunized and HEL-immunized mice. We identified four specific target antigens derived from enteric neurons and/or enteroglia. Antibodies against all four target antigens were present in MS patients. MS patients also showed gliosis and signs of ENS degeneration in the small intestine. For the first time, this study establishes a pathomechanistic link between the well-established autoimmune attack on the CNS and the ENS in MS. The presence of ENS pathology prior to CNS degeneration introduces entirely novel ways to explain MS etiology and immunopathogenesis.
Publication Title
The enteric nervous system is a potential autoimmune target in multiple sclerosis.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 27 Downloadable Samples
- Affymetrix Mouse Gene 1.0 ST Array (mogene10st)
Description
Recent studies have been successful at utilizing ectopic expression of transcription factors to generate induced cardiomyocytes (iCMs) from fibroblasts, albeit at a low frequency in vitro. This work investigates the influence of small molecules that have been previously reported to improve differentiation to cardiomyocytes as well as reprogramming to iPSCs in conjunction with ectopic expression of the transcription factors Hand2, Nkx2.5, Gata4, Mef2C, and Tbx5 on the conversion to functional iCMs. We utilized a reporter system in which the calcium indicator GCaMP is driven by the cardiac Troponin T promoter to quantify iCM yield. The TGF inhibitor, SB431542 (SB), was identified as a small molecule capable of increasing the conversion of both mouse embryonic fibroblasts and adult cardiac fibroblasts to iCMs up to ~5 fold. Further characterization revealed that inhibition of TGF by SB early in the reprogramming process led to the greatest increase in conversion of fibroblasts to iCMs in a dose-responsive manner. Global transcriptional analysis at Day 3 post-induction of the transcription factors revealed an increased expression of genes associated with the development of cardiac muscle in the presence of SB compared to the vehicle control. Incorporation of SB in the reprogramming process increases the efficiency of iCM generation, one of the major goals necessary to enable the use of iCMs for discovery-based applications and for the clinic.
Publication Title
Inhibition of TGFβ signaling increases direct conversion of fibroblasts to induced cardiomyocytes.
Sample Metadata Fields
Specimen part, Treatment
View Samples- Homo sapiens
- 9 Downloadable Samples
- Affymetrix Human Genome U133A 2.0 Array (hgu133a2)
Description
Transdifferentiation has been recently described as a novel method for converting human fibroblasts into induced cardiomyocyte-like cells. Such an approach can produce differentiated cells to study physiology or pathophysiology, examine drug interactions or toxicities, and engineer tissues. Here we describe the transdifferentiation of human dermal fibroblasts towards the cardiac cell lineage via the induced expression of transcription factors (TFs) GATA4, TBX5, MEF2C, MYOCD, NKX2-5, and delivery of microRNAs miR-1 and miR-133a. Cells undergoing transdifferentiation expressed ACTN2 and TNNT2 and partially organized their cytoskeleton in a cross-striated manner. The conversion process was associated with significant upregulation of a cohort of cardiac-specific genes, activation of pathways associated with muscle contraction and physiology, and downregulation of fibroblastic markers. We used a genetically encoded calcium indicator and readily detected active calcium transients although no spontaneous contractions were observed in transdifferentiated cells. Finally, we determined that inhibition of Janus kinase 1, inhibition of glycogen synthase kinase 3, or addition of NRG1 significantly enhanced the efficiency of transdifferentiation. Overall, we describe a method for achieving transdifferentiation of human dermal fibroblasts into induced cardiomyocyte-like cells via transcription factor overexpression, microRNA delivery, and molecular pathway manipulation.
Publication Title
Core Transcription Factors, MicroRNAs, and Small Molecules Drive Transdifferentiation of Human Fibroblasts Towards The Cardiac Cell Lineage.
Sample Metadata Fields
Specimen part, Treatment, Time
View Samples- Mus musculus
- 8 Downloadable Samples
- Affymetrix Mouse Gene 1.0 ST Array (mogene10st)
Description
Moderate alcohol exposure during pregnancy can result in a heterogeneous range of neurobehavioural and cognitive effects, termed fetal alcohol spectrum disorders (FASD). We have developed a mouse model of FASD that involves moderate ethanol exposure throughout gestation achieved by voluntary maternal consumption. This model results in phenotypes relevant to FASD. Since ethanol is known to directly affect the expression of genes in the developing brain leading to abnormal cell death, changes to cell proliferation, migration, and differentiation, and potential changes to epigenetic patterning, we hypothesize that this leaves a long-term footprint on the adult brain. However, the long-term effects of prenatal ethanol exposure on brain gene expression, when behavioural phenotypes are apparent, are unclear.
Publication Title
Long-lasting alterations to DNA methylation and ncRNAs could underlie the effects of fetal alcohol exposure in mice.
Sample Metadata Fields
Sex, Specimen part, Treatment
View Samples- Mus musculus
- 4 Downloadable Samples
- Affymetrix Mouse Gene 1.0 ST Array (mogene10st)
Description
The developing brain is particularly sensitive to ethanol during the brain growth spurt or synaptogenesis (third human trimester equivalent). This has been shown to lead to abnormal brain development and behavioural changes in the adult mouse that are relevant to those seen in humans with fetal alcohol spectrum disorders (FASD). We evaluated the long-term (postnatal day 60 young adult) gene expression changes that occur in the brain due to ethanol exposure during synaptogenesis.
Publication Title
Long-lasting alterations to DNA methylation and ncRNAs could underlie the effects of fetal alcohol exposure in mice.
Sample Metadata Fields
Treatment
View Samples- Mus musculus
- 12 Downloadable Samples
- Affymetrix Mouse Genome 430A 2.0 Array (mouse430a2)
Description
Transient over-expression of defined combinations of master regulator genes can effectively induce cellular reprogramming: the acquisition of an alternative predicted phenotype from a differentiated cell lineage. This can be of particular importance in cardiac regenerative medicine wherein the heart lacks the capacity to heal itself, but simultaneously contains a large pool of fibroblasts. In this study we determined the cardio-inducing capacity of ten transcription factors to actuate cellular reprogramming of mouse embryonic fibroblasts into cardiomyocyte-like cells. Over-expression of transcription factors MYOCD and SRF alone or in conjunction with Mesp1 and SMARCD3 significantly enhanced the basal but necessary cardio-inducing effect of the previously reported GATA4, TBX5, and MEF2C. In particular, combinations of five or seven transcription factors significantly enhanced the activation of cardiac reporter vectors, and induced an upregulation of cardiac-specific genes. Global gene expression analysis also demonstrated a significantly greater cardio-inducing effect when the transcription factors MYOCD and SRF were used. Detection of cross-striated cells was highly dependent on the cell culture conditions and was enhanced by the addition of valproic acid and JAK inhibitor. Although we detected Ca2+ transient oscillations in the reprogrammed cells, we did not detect significant changes in resting membrane potential or spontaneously contracting cells. This study further elucidates the cardio-inducing effect of the transcriptional networks involved in cardiac cellular reprogramming, contributing to the ongoing rational design of a robust protocol required for cardiac regenerative therapies.
Publication Title
Transcription factors MYOCD, SRF, Mesp1 and SMARCD3 enhance the cardio-inducing effect of GATA4, TBX5, and MEF2C during direct cellular reprogramming.
Sample Metadata Fields
Specimen part
View Samples- Mus musculus
- 28 Downloadable Samples
- Affymetrix Mouse Gene 1.0 ST Array (mogene10st)
Description
Direct conversion of fibroblasts to induced cardiomyocytes (iCMs) has great potential for regenerative medicine. Recent publications have reported significant progress, but the evaluation of reprogramming has relied upon non-functional measures such as flow cytometry for cardiomyocyte markers or GFP expression driven by a cardiomyocyte-specific promoter. The issue is one of practicality: the most stringent measures - electrophysiology to detect cell excitation and the presence of spontaneously contracting myocytes - are not readily quantifiable in the large numbers of cells screened in reprogramming experiments. However, excitation and contraction are linked by a third functional characteristic of cardiomyocytes: the rhythmic oscillation of intracellular calcium levels. We set out to optimize direct conversion of fibroblasts to iCMs with a quantifiable calcium reporter to rapidly assess functional transdifferentiation. We constructed a reporter system in which the calcium indicator GCaMP is driven by the cardiomyocyte-specific Troponin T promoter. Using calcium activity as our primary outcome measure, we compared several published combinations of transcription factors along with novel combinations in mouse embryonic fibroblasts. The most effective combination consisted of Hand2, Nkx2.5, Gata4, Mef2c, and Tbx5 (HNGMT). This combination is >50-fold more efficient than GMT alone and produces iCMs with cardiomyocyte marker expression, robust calcium oscillation, and spontaneous beating that persists for weeks following inactivation of reprogramming factors. HNGMT is also significantly more effective than previously published factor combinations for the transdifferentiation of adult mouse cardiac fibroblasts to iCMs. Quantification of calcium function is a convenient and effective means for the identification and evaluation of cardiomyocytes generated by direct reprogramming. Using this stringent outcome measure, we conclude that HNGMT produces iCMs more efficiently than previously published methods.
Publication Title
Optimization of direct fibroblast reprogramming to cardiomyocytes using calcium activity as a functional measure of success.
Sample Metadata Fields
No sample metadata fields
View Samples- Homo sapiens
- 6 Downloadable Samples
- Affymetrix Human Exon 1.0 ST Array [transcript (gene) version (huex10st)
Description
Grainyhead genes are involved in wound healing and developmental neural tube closure. In light of the high degree of similarity between the epithelial-mesenchymal transitions (EMTs) occurring in wound healing processes and the cancer stem cell-like compartment of tumors, including TGF--dependence, we investigated the role of a Grainyhead gene (GRHL2) in oncogenic EMT. Grainyhead was specifically down-regulated in the claudin-low subclass of mammary tumors and in the basal-B subclass of breast cancer cell lines. Functionally, GRHL2 suppressed TGF--induced, Twist-induced or spontaneous EMT, enhanced anoikis-sensitivity, and suppressed mammosphere generation in mammary epithelial cells. These effects were mediated, in part, by its suppression of ZEB1 expression, through direct repression of the ZEB1 promoter. GRHL2 also inhibited Smad-mediated transcription, and up-regulated mir200b/c as well as the TGF- receptor antagonist, BMP2. The expression of GRHL2 in the breast cancer cell line MDA-MB-231 triggered a mesenchymal-to-epithelial transition and sensitized the cells to anoikis. These results indicate that GRHL2 is a suppressor of the oncogenic EMT.
Publication Title
Suppression of the epithelial-mesenchymal transition by Grainyhead-like-2.
Sample Metadata Fields
Specimen part
View Samples- Homo sapiens
- 7 Downloadable Samples
- (ffymetrixhumanexon1.0starray[cdf:huex10stv2,corer3,a20071112,ep)
Description
KAP1 (TRIM28) is a transcriptional regulator in embryonic development that controls stem cell self-renewal, chromatin organization and the DNA damage response, acting as an essential co-repressor for KRAB family zinc finger proteins (KRAB-ZNF). To gain insight into the function of this large gene family, we developed an antibody that recognizes the conserved zinc fingers linker region (ZnFL) in multiple KRAB-ZNF. Here we report that the expression of many KRAB-ZNF along with active SUMOlyated KAP1 is elevated widely in human breast cancers. KAP1 silencing in breast cancer cells reduced proliferation and inhibited the growth and metastasis of tumor xenografts. Conversely, KAP1 overexpression stimulated cell proliferation and tumor growth. In cells where KAP1 was silenced, we identified multiple downregulated genes linked to tumor progression and metastasis, including EREG/epiregulin, PTGS2/COX2, MMP1, MMP2 and CD44, along with downregulation of multiple KRAB-ZNF proteins. KAP1-dependent stabilization of KRAB-ZNF required direct interactions with KAP1. Together, our results show that KAP1-mediated stimulation of multiple KRAB-ZNF contributes to the growth and metastasis of breast cancer.
Publication Title
KAP1 promotes proliferation and metastatic progression of breast cancer cells.
Sample Metadata Fields
Cell line
View Samples