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发布于:2018-11-21 17:29:45  访问:21 次 回复: 篇
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Reprogramming, genes associated with inflammation and chemotaxis were unexpectedly also part
Thus, while ER stress PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/25679764 affects ISC gene expression and proliferation in a way that is expected for a cellular stress and is in line with buy CDK8-IN-1 previous results [6], shortterm nutrient starvation appeared to trigger an apparently paradoxical pro-proliferative response.Tsalikis et al. b Overall fold induction of ISC genes compared to the total gene induction upon thapsigargin treatment and nutrient starvation. c List of ISC genes upregulated and downregulation more than 2 fold upon PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/26080418 stress. d Cell proliferation was analyzed by flow cytometry by monitoring EdU incorporation during the last 2 h of the 4 h treatment. Representative profiles of thapsigargin-treated (blue), nutrient-starved (green) and untreated control organoids (black) are shown. e Quantification of cell proliferation is presented as mean fluorescence intensity (MFI)Analysis of the AS landscape in enteroids undergoing ER stress and nutrient deprivationOne key aim of this study.Reprogramming, genes associated with inflammation and chemotaxis were unexpectedly also part of this common transcriptional signature.In primary intestinal organoids, cell proliferation is dependent on the activity of ISCs and ISC-derived transit-amplifying cells that express the marker Lgr5 and occupy an important fraction of the intestinal crypt [17]. The above results suggested that ER stress and nutrient starvation affected the expression of cell cycle and/or proliferation genes differentially in intestinal organoids (see Fig. 2 and Additional file 3: Figure S1). Therefore, to identify the overall impact of thapsigargin versus nutrient starvation on ISCs, we analyzed how these stresses affected the transcript levels of the 151 ISCenriched genes identified recently by sorting Lgr5+ intestinal epithelial cells [18]. Interestingly, the majority of ISC genes displayed reduced expression following thapsigargin treatment (fold induction <1), while the opposite result was observed in nutrient starved cells (Fig. 4a). Overall, expression of ISC genes was not upregulated (1.06 fold) by thapsigargin while this treatment upregulated gene expression 1.34 fold genome-wide (Figs. 1f and 4b). In contrast, ISC genes were significantly more upregulated (1.41 fold) than all genes (1.1 fold) by nutrient starvation (Figs. 1f and 4b). Despite these differences, we observed that a group of seven genes were similarly modulated by both stresses: Tnfrsf19, Wwtr1, Vav3, Esrrg and Notch1 were down-regulated by thapsigargin and nutrient starvation, while Car12 and Rasa3 were upregulated. These seven genes might thus represent the core group of genes involved in the cellular adaptation of ISCs to stress. Finally, we aimed to directly test the differential effect of thapsigargin and nutrient starvation on cellular proliferation in intestinal organoids by measuring EdU incorporation in organoids undergoing stress for 4 h. In agreement with the ISC gene expression data, we observed that thapsigargin stimulation resulted in potent reduction of EdU incorporation in organoids, indicative of a block in proliferation, while nutrient starvation had the opposite effect (Fig. 4d-e). Thus, while ER stress PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/25679764 affects ISC gene expression and proliferation in a way that is expected for a cellular stress and is in line with previous results [6], shortterm nutrient starvation appeared to trigger an apparently paradoxical pro-proliferative response.Tsalikis et al.
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