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

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^-1）， 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^-1）， 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.

Effect of Modified Ditantang on Autophagy and Relevant Proteins in Brain Cells of Rats with Cerebral Ischemia Reperfusion Injury / 中国实验方剂学杂志

Objective:To investigate the effect of modified Ditantang on autophagy and relevant proteins in brain cells of rats with cerebral ischemia reperfusion injury. Method:The cerebral ischemic reperfusion (CIR) injury model in rats was built by reversible middle cerebral artery occlusion artery suture of middle cerebral embolism method, and randomly divided into sham group, model group, high-dose modified Ditantang group(H-dose), low-dose modified Ditantang group (L-dose, 0.384 g·kg-1) and PLXT group (0.1 g·kg-1). Sham and model groups were given normal saline by gastric perfusion, H-dose and L-dose groups were given modified Ditantang, and the PLXT group were given Piracetam tablets, intragastric volume 10 mL·kg-1. The treatment lasted for 7 d. Within 24 hours after administration, the histopathological examination was performed, the volume of cerebral infarction, neuronal apoptosis and serum levels of inflammatory factors were compared, and the expressions of autophagy related microtuble-associated protein 1 light chain 3(LC3)Ⅱ, Beclin1, B-cell lymphoma-2(Bcl-2) and p62 in brain tissues were determined. Result:Cells and blood vessel necrosis, neuron swelling and interstitial edema were observed in model group, a few neurons died, edema was reduced, swelling of nerve cells was alleviated in H-dose, L-dose and PLXT groups. The volume of cerebral infarction and neuronal apoptosis in H-dose, L-dose and PLXT groups were lower than those in model group (Pα, intedeukin (IL)-2 and IL-8 in H-dose, L-dose and PLXT groups were lower than those in model group (PPConclusion:Modified Ditantang can improve brain injury and interfere with autophagy after MCAO/R, alleviate inflammation, and regulate autophagic activity, which may be related to the down-regulation of expressions of LC3 Ⅱ, Beclin1 and the up-regulation of expression of p62.