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Oridonin induces apoptosis and autophagy in murine fibrosarcoma L929 cells partly via NO-ERK-p53 positive-feedback loop signaling pathway.


ABSTRACT:

Aim

To investigate the role of nitric oxide (NO) in oridonin-induced apoptosis and autophagy in murine fibrosarcoma L929 cells and the underlying molecular mechanisms.

Methods

Cell viability was measured using MTT assay. Intracellular NO level, SubG(1) cell ratio and autophagy cell ratios were analyzed with flow cytometry after diaminofluorescein-2 diacetate (DAF-2DA), propidium iodide (PI) and monodansylcadaverine (MDC) staining, respectively. Protein expression was examined using Western blot analysis.

Results

Exposure of L929 cells to oridonin (50 ?mol/L) for 24 h led to intracellular NO production. Pretreatment with NOS inhibitor 1400w or L-NAME inhibited oridonin-induced apoptosis and autophagy in L929 cells. The pretreatment decreased the apoptosis-related protein Bax translocation and cytochrome c release, increased Bcl-2 level, reversed the autophagy-associated protein Beclin 1 increase and conversion of LC3 I to LC3 II. Furthermore, pretreatment with NO scavenger DTT completely inhibited oridonin-induced apoptosis and autophagy in L929 cells. In addition, oridonin (50 ?mol/L) activated ERK and p53 in L929 cells, and the interruption of ERK and p53 activation by PD 98059, pifithrin-?, or ERK siRNA decreased oridonin-induced apoptosis and autophagy. The inhibition of NO production reduced oridonin-induced ERK and p53 activation, and NO production was down-regulated by blocking ERK and p53 activation.

Conclusion

NO played a pivotal role in oridonin-induced apoptosis and autophagy in L929 cells. Taken together with our previous finding that ERK contributes to p53 activation, it appears that NO, ERK, and p53 form a positive feedback loop. Consequently, we suggest that oridonin-induced apoptosis and autophagy are modulated by the NO-ERK-p53 molecular signaling mechanism in L929 cells.

SUBMITTER: Ye YC 

PROVIDER: S-EPMC4085664 | biostudies-literature | 2012 Aug

REPOSITORIES: biostudies-literature

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Publications

Oridonin induces apoptosis and autophagy in murine fibrosarcoma L929 cells partly via NO-ERK-p53 positive-feedback loop signaling pathway.

Ye Yuan-chao YC   Wang Hong-ju HJ   Xu Lei L   Liu Wei-wei WW   Liu Bin-bin BB   Tashiro Shin-Ichi S   Onodera Satoshi S   Ikejima Takashi T  

Acta pharmacologica Sinica 20120730 8


<h4>Aim</h4>To investigate the role of nitric oxide (NO) in oridonin-induced apoptosis and autophagy in murine fibrosarcoma L929 cells and the underlying molecular mechanisms.<h4>Methods</h4>Cell viability was measured using MTT assay. Intracellular NO level, SubG(1) cell ratio and autophagy cell ratios were analyzed with flow cytometry after diaminofluorescein-2 diacetate (DAF-2DA), propidium iodide (PI) and monodansylcadaverine (MDC) staining, respectively. Protein expression was examined usin  ...[more]

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