Transcription profiling by array of the impact of feeding strategies on the scalable expansion of human pluripotent stem cells (hHSC_F1285T_iPS2) in single-use stirred tank bioreactors
Ontology highlight
ABSTRACT:
INSTRUMENT(S): Agilent Micro Array Scanner G2565CA
ORGANISM(S): Homo sapiens
SUBMITTER: Christina Kropp
PROVIDER: E-MTAB-4149 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:Comparison of gene expression profile of human pluripotent stem cells (clone hCBiPS2) expanded in batch-fed and perfusion stirred bioreactors after 0, 3 and 7 days.
2016-08-22 | E-MTAB-3898 | biostudies-arrayexpress
Project description:Hepatocellular carcinoma (HCC) is a frequent cancer with poor prognosis and with limited possibilities for anti-cancer treatment. Evidence has recently accumulated suggesting that multiple signaling pathways are activated in human cancer. It has been recently shown by exosome sequencing of HCC that 161 putative driver genes are associated with 11 recurrently altered pathways, suggesting that we need to inhibit these multiple pathways for HCC therapy. THOC5, a member of transcription/export (TREX) complex, that plays a role in less than 1% of mRNA processing in normal cells such as fibroblasts or macrophages, is not required for maintenance of mature hepatocytes. In this study, we have examined the role of THOC5 in human HCC. Enhanced THOC5 expression was not observed in differentiation grading 1 (G1) human HCC, but 78% of G2 and G3 exhibited increased levels of THOC5. Furthermore, the 50% depletion of THOC5 in hepatocellular carcinoma cell lines, Huh7 and HepG2 causes lipid accumulation and apoptosis. Transcriptome analysis using THOC5 depleted Huh7 and HepG2 cells revealed that 1049 and 837 genes were downregulated upon depletion of THOC5 in Huh7 and HepG2 cells, respectively. Among these genes, 396 were commonly downregulated in both cell lines. Some of these, such as tissue inhibitor of metalloproteinase 3 (TIMP3), tripartite motif containing 24 (TRIM24), ribosomal protein S6 kinase polypeptide 3 (RPS6KA3) and transmembrane emp24-like trafficking protein 10 (TMED10) are known to be fine tuners for several signal transduction pathways. The expression of these proteins was correlated with the THOC5 expression level in HCC. To inhibit multiple signaling pathways we are currently examining the effects of a combination of siRNAs against THOC5 target genes in HCC cell lines. Our data suggest that THOC5, which controls a set of genes that are involved in HCC malignancy can be a potential biomarker for HCC. In addition, THOC5 target gene, TMED10 may also be a novel biomarker of HCC. Furthermore, the suppression of the THOC5 gene per se or multiple THOC5 target genes may represent a novel strategy for cancer therapy. Huh7 and HepG2 cells were infected with lentiviral THOC5 and control shRNAs. Five days after infection, total RNAs were isolated and subjected to microarray analysis.
2015-11-02 | E-GEOD-70178 | biostudies-arrayexpress
Project description:Type I interferons (IFNs) play a key role in linking the innate and adaptive immune responses. IFNs can have both positive and negative effects on the development of memory CD8+ T cells, but their contribution to the priming of naïve T cells is unclear. Using OT-I TCR transgenic T cells, we show strong effects on TCR-induced functional response when sensitized by IFN-β prior to their stimulation with cognate antigen. These effects are only seen in the absence of co-stimulation and at suboptimal TCR stimulation conditions. Pre-stimulation of naïve T cells inhibits certain TCR-mediated functions like cell proliferation and the induction of genes such as IFN-γ and IL-2. However, type I IFN does not influence the induction of other TCR-induced functions like the expression of the T cell activation marker CD25 and a number of other genes. IFN-mediated impairment of proliferation is due to the lack of IL-2 induction since it can be compensated by exogenous IL-2. Further analysis reveals an IFN-mediated inhibition of Ca2+ flux and ERK phosphorylation. In vivo studies confirm that pre-stimulation with IFNs inhibits early T cell responses, whereas long-term responses are positively affected. Our results reveal that IFN exerts negative effects on naïve T cells in a timely defined range under sub-optimal TCR stimulation conditions. These results highlight the importance of timing of immune-regulatory events and reveal a novel role for type I IFNs by shaping the immune response, such that T cells are not able to react until optimal stimulation is provided. 8 Arrays, untreated cells as references, IFNb treatment and/or TCR stimulation at various lengths of time
2015-12-31 | E-GEOD-41381 | biostudies-arrayexpress
Project description:Human iPSC-Mac or PBMC-mac were seeded on 24-well plates (500.000cells/well) and cultured over night. On the next day, cells were washed three times with PBS. Subsequently, P. aeruginosa laboratory strain PAO1 in RPMI medium without antibiotics (MOI10) was centrifugated on the cells (600xg) and incubated at 37°C. Cells with medium only served as non-infected controls. After 1h cells were de-attached, washed and resuspended in RNA lysis buffer. RNA isolation was performed with RNAeasy micro Kit (Qiagen), according to manufactures instructions. Human iPSC samples were obtained after sorting of iPSC for TRA-1-60+ to separate iPSCs from feeder cells.
2018-10-23 | E-MTAB-5436 | biostudies-arrayexpress
Project description:Human pluripotent stem cells were aggregated in mTeSR medium for 4 days and subjected to an optimzed small molecule-based cardiac differentiation protocol (modified from Kempf et. al., Stem Cell Reports, 2014) in agitated Erlenmeyer flasks. Differentiation was induced using 7.5µM CHIR99021 (day 0). Cardiac mesoderm specification was carried out using 5µM IWP2 either directly after CHIR99021 exposure after 24h (day 1; optimized protocol) or after 72h hours (day 3; standard procedure). RNA samples were harvested at undifferentiated state (day 0), after differentiation induction (day 1) and after 3 days with and without IWP2 treatment.
2019-01-13 | E-MTAB-6828 | biostudies-arrayexpress
Project description:Background: Evidence has recently accumulated suggesting that mature hepatocytes have the unique capacity to autonomously decide their replication fate. The molecular mechanism by which hepatocytes switch from an essentially quiescent state to rapidly proliferation after liver injury is not fully understood. To provide molecular insights into the self-renewal of mature hepatocytes, we have applied an ex vivo liver slice culture system. Results: After liver injury, activation of p38 and Erk1/2 began in mature hepatocytes within 5 min. Erk1/2 was activated at the edge of the cut as well as on the surface of liver slices. The number of hepatocytes that contain activated Erk1/2 increased within 1 h and then decreased. Concomitantly, immediate early genes (IEGs), such as Jun, early gene response 1 (Egr1) and Myc were induced and Ki67 positive hepatocytes appeared 24 h after liver injury in the slices. In agreement with in vivo studies, transcriptome analysis using 2 h slice culture reveals that 63% of upregulated genes were downstream of lipopolysaccharide (LPS) stimulation which induces interleukin 6 (IL 6) or tumor necrosis factor (TNF) alpha pathways. Furthermore, circadian clock regulator, such as members of the basic helix-loop-helix family, Bhlhe40 and Bhlhe41, were highly upregulated for 4 h after injury. Although upon injury Jun or Egr1 were induced in liver slices from proliferation-defective liver, no circadian clock regulator genes were upregulated. Moreover, using an in vitro cell culture system we show that Bhlhe40 is required for the G0-G1 transition. Conclusion: Our data suggest that the circadian clock regulator, Bhlhe40, is involved in the G0-G1 transition. An ex vivo system using normal and proliferation defective KO liver is a useful tool for identification of genes that trigger cell proliferation shortly after liver injury. This method may also be applied for measurement of the liver regeneration potential of individual livers at the priming phase. In one dual-color microarray hybridization, mRNA expression changes after 2h ex vivo incubation of liver slices were examined.
2015-12-31 | E-GEOD-65995 | biostudies-arrayexpress
Project description:1-day-old C57BL/6 mice were left untreated (co) or orally infected with 10E2 CFU wildtype (wt) or delta invC SPI1 mutant Salmonella Typhimurium (ATCC14028). Four biological replicates obtained from individual animals were exmained; each group contained animals from at least 2 different litters. On day 4 p.i., animals were sacrificed and intestinal epithelial cells were isolated from total small intestine (protocol according to: Lotz et al., J. Exp. Med. 2006). Total RNA was isolated by TriZol and its purity was examined using a Bioanalyzer. We used microarrays to detail the global gene expression in primary total isolated intestinal epithelial cells. Four biological replicates of isolated intestinal epithelial cells obtained 4 daysp.i. from neonate mice infected at the age of 1 day (each group with animals from at least two litteres). Single color array.
2014-09-01 | E-GEOD-51160 | biostudies-arrayexpress
Project description:Intestinal epithelial cells of 28-day-old female mice were isolated according to a recently published protocol (Lotz et al., J Exp. Med. 2006). Isolated intestinal epithelial cells of four individual animals (four biological replicates) were examined. Single color array.
2014-08-23 | E-GEOD-51269 | biostudies-arrayexpress
Project description:Transcription of immediate early genes (IEGs) in response to extrinsic and intrinsic signals is tightly regulated at multiple stages. THOC5, a member of the TREX (transcription/export) complex plays a role in expression of only a subset of constitutively active genes. In this report, we show that THOC5 plays a role in the 3M-BM-4end-processing of more than 90% of IEGs induced by serum stimulation, however THOC5 depletion does not influence the expression of the most rapidly induced IEGs, such as Fos or Jun. One group of THOC5 target genes, including Id1, Id3 and Wnt11 transcripts were not released from chromatin in THOC5 depleted cells. Another group of genes, such as Myc, and Smad7 transcripts, were released with shortening of 3M-BM-4UTR by alternative cleavage, and were spliced but were exported to a lesser extent in THOC5 depleted cells. Furthermore, upon stimulation with serum THOC5 forms a complex with polyadenylation specific factor 100 (CPSF100). THOC5 is required for recruitment of CPSF100 to 3M-BM-4UTR of THOC5 target genes, but not for THOC5 independent genes. These data suggest the presence of a novel mechanism for the control of immediate early genes response via 3M-BM-4end-processing. In 2 dual-color microarray hybridizations, mRNA expression changes after 1h of FCS treatment were examined. MEF(Rosa26ERT2 Cre: THOC5 (flox/flox)) cells in which a THOC5 knockdown is inducible upon Tamoxifen treatment were analyzed. M4040 represents cells without Taxoxifen pre-treatment (wildtype reference) whereas M4041 represents the same celltype pre-treated for 2 days with Tamoxifen to induce a THOC5 knockdown.
2014-11-03 | E-GEOD-59561 | biostudies-arrayexpress
Project description:comparison of human pluripotent stem cell aggregates from Batch (BA) and Cyclic-Perfusion (CPAs) feeding generated in stirred bioreactors.
2014-11-30 | E-MTAB-2814 | biostudies-arrayexpress