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Turkish Journal of Medical Sciences

Abstract

Background/aim: 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.

Materials and methods: 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.

Results: 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.

Conclusion: 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.

Author ORCID Identifier

ABULİMİTİABULIMITI ABULAİTİABULAITI: 0009-0004-5203-308X

XİAOHONG SUN: 0009-0000-2217-3066

WARESIJIANG YIBULAYIN: 0009-0007-4560-2573

DAN HE: 0009-0003-6118-6734

KEMİNGKEMING XU: 0009-0003-7077-7683

XİAYİMAİERDAN YİBULAYİN: 0009-0005-9092-5668

DOI

10.55730/1300-0144.6163

Keywords

apoptosis, mitochondrial autophagy, Oleanolic acid-chitosan compound, PTEN/AKT signaling pathway

First Page

292

Last Page

301

Publisher

The Scientific and Technological Research Council of Türkiye (TÜBİTAK)

Creative Commons License

Creative Commons Attribution 4.0 International License
This work is licensed under a Creative Commons Attribution 4.0 International License.

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