| Gene name: | JAG1 |
| Aging type: | Prevent |
| Aging characteristic: |
| Tissue type: | -- |
| Cell name: | MSC |
| Experiment: | SA-β-gal activity assay//Flow cytometry |
| Description: | As we expected,activation of Notch signaling by JAG1 led to a reduced cell senescence in both 1- and 5-day sheet cultures, as demonstrated by decreased frequencies of SA-β-gal positive cells (8.25±1.5% and 20.5±4.2%, respectively) when compared with MSCs from lgG control groups .By comparison, the G0 phase cells in JAG1 groups was 45.2±2.8% and 57.5±8.2% in the 1-day and 5-day cultures respectively, and significantly lower than those in the lgG group. |
| Target gene: | P16//CCND1 |
| Official symbol(s): | P16//CCND1 |
| R-AG-Target gene: | Downregulation//Upregulation |
| Subcategory: | Unclear |
| Target gene experiment: | RT-PCR//Western blot |
| Target gene description: | In lgG control group,relative quantification by RT -PCR revealed an ~ 5-fold up-regulation of p16 RNA and a 2.7-fold up-regulation of p21 RNA in 5-day cultures compared with 1-day cultures, which suggests that these two factors act synergistically to induce MSC senescence. Finally, protein expression of p16 in western blot confirmed our PCR results by showing thinner band in JAG1 treated cells and thicker band in Hes1 deficient cells, and no significant change was observed in protein expression of p21 .Consistent with our previous finding, CCND1 expression was induced by JAG1 in 1-day cultures and no significant changes were observed in 5-day cultures compared with lgG controls. |
| Regulatory pathway: | JAG1-NOTCH-HES1 |
| R-AG-Pathway: | Activation |
| Official symbol(s): | JAG1-NOTCH1-HES1 |
| Pathway experiment: | RT-PCR//Western blot//SA-β-gal activity assay |
| Pathway description: | Therefore, we further measured Notch target Hes1 expression in 1- and 5-day sheet cultures. RT -PCR analysis of total RNA revealed a decreased expression of Hes1from 1-day to 5-day cultures in control lgG-coated plates, while Hes1 expression at both time points was significantly increased by JAG1.Western blot analysis using protein from 5-day sheet cultures further confirmed these knockdown results by showing a significantly reduced Hes1 protein expression in shRNA lentivirus-infected MSCs with or without JAG1 treatment.More importantly, although the cellular senescence in 5-day cultures was significantly inhibited by JAG1-mediated Notch activ ation, this inhibitory effect was almost completely abrogated after knocking down Hes1 expression in these MSCs. |
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