Abstract Background The role of the m 6 A-binding protein YTHDC2 in the occurrence and development of colorectal cancer (CRC) is unclear. We aimed to explore the molecular mechanisms underlying this process and clarify the signaling pathway involved. Methods Firstly, the relationship between YTHDC2 and CRC in TCGA database was analyzed to identify relevant signaling pathways and biological processes. Then, western blot was used to analyze expression of YTHDC2 in HCT116 and Caco2 cells. After knockdown or overexpression of YTHDC2 in the above cells, RT-PCR and western blot were used to analyze p38MAPK, p-p38MAPK, and downstream apoptosis-related proteins in the MAPK signaling pathway. Flow cytometry was performed to detect changes in apoptosis. Results And the results were shown that the expression of YTHDC2 was significantly lower in tumor tissues than in normal tissues. Increased expression of YTHDC2 was associated with better overall survival among patients with CRC. Gene set enrichment analysis revealed that YTHDC2 regulates the MAPK signaling pathway. Flow cytometry revealed apoptosis was significantly reduced and enhanced in response to YTHDC2 knockdown and overexpression, respectively. There was no significant change in the expression of p38MAPK, while p-p38MAPK was significantly increased in response to overexpression and decreased in response to knockdown. Gene Expression Profiling Interactive Analysis showed apoptotic protein expression to be positively correlated with YTHDC2 expression, consistent with the results of RT-PCR and western blot. Conclusion In general, apoptosis of CRC cells is promoted by YTHDC2 via activation of the exogenous death receptor and endogenous mitochondrial apoptosis-related pathways in the p38MAPK signaling pathway.
Nitric oxide-releasing compounds were shown to activate an ADP-ribosyltransferase activity in the cytosol of Dictyostelium discoideum. The enzyme ADP-ribosylated a cytosolic protein of approximately 41 kDa, p41. Neither cGMP nor GTP and its analogues affected this ADP-ribosylation. p41 differs from other substrates ADP-ribosylated by cholera, pertussis, or diphtheria toxins. Treatment of ADP-ribosylated p41 with snake venom phosphodiesterase released adenosine 5'-monophosphate, indicating a mono-ADP-ribose-protein linkage. This linkage was stable to neutral hydroxylamine but was sensitive to mercury ions and iodomethane, suggesting an attachment to a cysteine residue. Treatment of intact cells with nitric oxide-releasing compounds appeared to stimulate the ADP-ribosylation of p41 and this modification was reversible.
Calycosin is a bioactive component of Astragali Radix, a Chinese herb for treating allergy. We have previously demonstrated that calycosin effectively inhibited allergic inflammation efficiently. The aim of this study was to explore the mechanism of calycosin on epithelial cells in allergic inflammation.An initial stage of atopic dermatitis (AD) model in which mice were just sensitized with FITC, was established in vivo and immortalized human keratinocytes (HaCaT cells) were utilized in vitro. Initiative key cytokines, TSLP and IL-33, were measured by ELISA, qPCR, immunofluorescence and Western blot. The junctions in epithelial cells were observed by electron microscopy and tight junctions (TJs) (Occludin and ZO-1) were assessed by Western blot and immunofluorescence. TLR4, MyD88, TAK1, TIRAP and NF-κB were measured by qPCR or Western blot.The results showed that TSLP and IL-33 were inhibited significantly by calycosin in the initial stage of AD model. Simultaneously, calycosin attenuated the separated gap among the epithelial cells and increased the expression of TJs. TSLP/IL-33 and TJs were similarly affected in LPS-stimulated HaCaT cells in vitro. Meanwhile, calycosin not only inhibited the expressions of TLR4, MyD88, TAK1 and TIRAP, but also reduced NF-κB activation in vitro and in vivo. An NF-κB inhibitor enhanced the expressions of TJs and reduced that of TSLP/IL-33 in LPS-stimulated HaCaT cells.These results indicated that calycosin reduced the secretion of TSLP/IL-33 and attenuated the disruption of epithelial TJs by inhibiting TLR4 mediated NF-κB signaling pathway. These findings help to understand the beneficial effects of calycosin on AD, and to develop effective preventive or therapeutic strategies to combat this disease and other epithelial barrier deletion-mediated allergic diseases.
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