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Effect of Yiqi Fuzheng Jiedu Decoction on Autophagy and Growth of A549 Cells: An Exploration Based on p53/AMPK Signaling Pathway / 中国实验方剂学杂志
Chinese Journal of Experimental Traditional Medical Formulae ; (24): 65-75, 2021.
Article in Chinese | WPRIM | ID: wpr-905928
ABSTRACT

Objective:

To investigate the effect and mechanism of Yiqi Fuzheng Jiedu decoction (YQFZJDD) on autophagy and growth of A549 cells.

Method:

A549 cells were intervened with YQFZJDD-containing serum prepared in advance. The levels of microtubule-associated protein 1 light chain 3 (LC3), homologue of yeast autophagy-related gene 6 (Beclin1), p62, p53, adenosine 5'-monophosphate-activated protein kinase (AMPK), phosphorylated AMPK (p-AMPK), mammalian target of rapamycin (mTOR), and phosphorylated mTOR (p-mTOR) were detected by Western blot assay, and microtubule-associated protein 1A/1B light chain 3B (MAP1LC3B) by immunofluorescence (IF) assay. The proliferation, invasion, and senescence of A549 cells were separately measured by 5-ethynyl-2'-deoxyuridine (EDU) staining, Transwell assay, and <italic>β</italic>-galactosidase staining. In the presence of autophagy inhibitor 3-methyladenine (3-MA, 5 mmol·L<sup>-1</sup>), the cells were divided into the 10% fetal bovine serum (blank) group, 10% control serum (control) group, low-, medium-, and high-dose 10% YQFZJDD-containing serum groups, and high-dose 10% YQFZJDD-containing serum + 3-MA group, followed by the measurement of A549 cell proliferation, invasion, and senescence. In the adoption of p53 inhibitor Pifithrin-<italic>α</italic> (PFT-<italic>α</italic>, 10 μmol·L<sup>-1</sup>), the cells were divided into the control group, PFT-<italic>α</italic> group, high-dose YQFZJDD-containing serum group, and high-dose YQFZJDD-containing serum + PFT-<italic>α</italic> group. Then the LC3-Ⅱ and Beclin1 expression, MAP1LC3B fluorescence intensity, as well as A549 cell proliferation, invasion and senescence were determined.

Result:

Compared with the blank group and control group, YQFZJDD-containing serum at the medium and high doses up-regulated the protein expression levels of LC3-Ⅱ and Beclin1 in A549 cells after 48-h intervention in a dose-dependent manner (<italic>P</italic><0.01). Besides, YQFZJDD-containing serum at the low-, medium-, and high-doses down-regulated p62 and p-mTOR/mTOR expression (<italic>P</italic><0.05, <italic>P</italic><0.01), elevated p53 and p-AMPK/AMPK (<italic>P</italic><0.01), decreased the number of proliferative and invasive cells, increased the number of senescent cells (<italic>P</italic><0.01), and enhanced the IF intensity of MAP1LC3B, with the optimal effect observed in the high-dose YQFZJDD-containing serum group (<italic>P</italic><0.01). Compared with the high-dose YQFZJDD-containing serum group, the high-dose YQFZJDD-containing serum + 3-MA group and high-dose YQFZJDD-containing serum + PFT-<italic>α</italic> group exhibited significantly increased proliferative and invasive cells but decreased senescent cells (<italic>P</italic><0.05, <italic>P<</italic>0.01). Meanwhile, the IF intensity of MAP1LC3B in the high-dose YQFZJDD-containing serum + PFT-<italic>α</italic> group was weakened and the LC3-Ⅱ and Beclin1 protein expression levels declined (<italic>P</italic><0.05, <italic>P<</italic>0.01).

Conclusion:

YQFZJDD promotes the autophagy of A549 cells through the p53/AMPK signaling pathway to inhibit their proliferation, invasion and migration but accelerate senescence, thus playing a crucial role in inhibiting the progression of lung cancer.

Full text: Available Index: WPRIM (Western Pacific) Language: Chinese Journal: Chinese Journal of Experimental Traditional Medical Formulae Year: 2021 Type: Article

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Full text: Available Index: WPRIM (Western Pacific) Language: Chinese Journal: Chinese Journal of Experimental Traditional Medical Formulae Year: 2021 Type: Article