Oleanolic acid–chitosan compound inhibits mitochondrial autophagy and malignant transformation of lung cancer through the PTEN/AKT pathway

TL;DR

A compound made of oleanolic acid and chitosan fights lung cancer by affecting cell survival and autophagy through a key signaling pathway.

Contribution

The study reveals that OAC inhibits lung cancer by modulating the PTEN/AKT pathway to induce apoptosis and autophagy.

Findings

OAC significantly reduced cell viability and proliferation in A549 lung cancer cells.

OAC induced apoptosis and autophagy, marked by changes in key protein expressions and mitochondrial dysfunction.

PTEN overexpression enhanced the effects of OAC on apoptosis and autophagy.

Abstract

Oleanolic acid–chitosan compound (OAC) shows potent antinonsmall cell lung cancer (NSCLC) activity, but its mechanisms remain unclear. This study elucidates how OAC modulates autophagy and apoptosis via the Phosphatase and Tensin Homolog/Protein Kinase B (PTEN/AKT) signaling pathway.

Oleanolic acid was coupled with chitosan to synthesize OAC. A549 and MRC-5 cells were cultured in Dulbecco’s Modified Eagle Medium and Minimum Essential Medium, respectively, supplemented with 10% fetal bovine serum and penicillin–streptomycin. Cells were pretreated with chloroquine before OAC treatment. Cell viability was assessed using the CCK-8 assay, while 5-bromo-2′-deoxyuridine (BrdU) and colony formation tests were employed to evaluate cell proliferation. Apoptosis was measured by flow cytometry using Annexin V-FITC/PI (fluorescein isothiocyanate/propidium iodide) staining. Autophagy was monitored through western blot analysis of LC3, SQSTM1, Atg5, Beclin1, and LAMP1, and confirmed by immunofluorescence and transmission electron microscopy (TEM). Mitochondrial membrane potential (MMP) was determined by JC-1 staining, and acridine orange staining was used to detect acidic vesicular organelles (AVOs). Additionally, PTEN overexpression was induced by transfecting cells with the hemagglutinin tag (HA)-PTEN plasmid.

OAC treatment significantly inhibited cell viability and proliferation in A549 cells, as evidenced by decreased CCK-8 absorbance, reduced BrdU incorporation, and fewer colony formations. Flow cytometry revealed a marked increase in apoptotic cells following OAC treatment. Western blot analysis demonstrated altered expression levels of apoptosis-related proteins (Bcl-2, Bax, Cytochrome c, caspase-9, caspase-3, PARP) and autophagy markers (LC3, SQSTM1, Atg5, Beclin1, LAMP1). Immunofluorescence and TEM further confirmed the induction of autophagy. AO staining showed increased AVOs, while JC-1 staining indicated a reduction in MMP. PTEN overexpression enhanced OAC-induced apoptosis and autophagy.

OAC effectively inhibits cell proliferation and induces apoptosis and autophagy in A549 cells, with PTEN playing a regulatory role in these processes. These findings suggest that OAC may serve as a potential therapeutic agent for cancer treatment.

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