Inhibition of PI3K/Akt/mTOR pathway by apigenin induces apoptosis and autophagy in hepatocellular carcinoma cells
Graphical abstract
Introduction
Hepatocellular carcinoma is the sixth most common cancer and ranked as the third leading cause of cancer-related deaths worldwide [1,2]. Due to lacking specific clinical symptoms and signs, early diagnoses and treatments, hepatocellular carcinoma become almost impossible [3]. The treatment of liver cancer includes surgical resection, local ablation, transplantation, transarterial embolization and immunotherapy [4]. Although improved diagnosis and therapeutic methods, the overall 5-year survival rate of approximately 12% [5]. Therefore, novel agents and effective therapeutic strategies to treat liver cancer are urgently required.
Apigenin (4′,5,7-trihydroxyflavone), is a natural flavonoid that is widely distributed in fruits and vegetables, such as parsley, orange, tea, chamomile and seasonings [6]. In recent years, apigenin has been shown to possess significant anti-inflammatory, anti-oxidant and anti-tumor properties [7]. Previous studies have reported that apigenin could inhibit the growth and proliferation, promote apoptotic cell death, induce cell cycle arrest and mitochondrial membrane potential disruption of cancer cells in vitro and in vivo [[8], [9], [10], [11], [12], [13], [14], [15], [16], [17], [18], [19], [20]]. However, the precise mechanism underlying the anti-tumor effects of apigenin remains poorly understood.
Autophagy, an evolutionarily conserved pathway, plays a vital role in degradation of the superfluous proteins and organelles. Autophagy is induced by a series of conditions, such as hypoxia, cellular stress and nutrient starvation [21]. Recent studies indicate that targeted autophagy may be an effective therapeutic strategy to fight cancer [22]. However, autophagy has dual roles in regulating cell survival with regard to cancer treatment [23]. Accumulating evidence suggests that in some conditions, cancer cells tend to use autophagy as a survival mechanism to avoid chemotherapeutic agents or γ-irradiation induced apoptosis [24]. Several molecular and signaling pathways play a crucial role in regulating autophagy in many types of cancers, such as the autophagy-related gene family [25] and PI3K/Akt/mTOR signaling pathway [26,27]. The mammalian target of rapamycin (mTOR) plays a critical role in regulating the balance between cell proliferation and autophagy in response to cellular stress induced by chemotherapeutics in cancer cells [28]. Previous studies have reported that apigenin could induce autophagy and apoptosis in breast cancer cells [29]. However, the relationship between apoptosis and autophagy induced by apigenin remains unknown.
In the present study, we aimed to investigate the effect of apigenin on apoptosis and autophagy in hepatocellular carcinoma cells and xenograft mouse model. Our results showed that apigenin induced apoptosis and autophagy in vitro and in vivo. We found that inhibition of autophagy led to inhibition of cell proliferation and tumor growth. Moreover, we also demonstrated that the apigenin induced autophagy and apoptosis via inhibiting PI3K/Akt/mTOR pathway.
Section snippets
Reagents and antibodies
Apigenin was purchased from Sigma-Aldrich (St. Louis, MO, USA). Antibodies for cleaved-caspase-3(1:1500), cleaved-caspase-9(1:1000), cleaved-PARP(1:1500), Bcl-2(1:1000), Bax(1:1000), PI3K(1:1500), p-PI3K(1:1000), Akt(1:1000), p-Akt(1:800), mTOR(1:1000), p-mTOR(1:800), Atg5(1:1000), Beclin1(1:1000) and SQSTM1(1:1500) were obtained from Cell Signal Technology (Boston, USA). Antibodies for LC3B(1:1000) and actin(1:5000) were purchased from Sigma (St. Louis, MO, USA). Atg5 siRNA, GFP-LC3B plasmid,
Apigenin inhibited cell growth and induced apoptosis in hepatocellular carcinoma cells
To investigate the effect of apigenin on the viability of hepatocellular carcinoma cells, we treated HepG2 cells with apigenin We found that apigenin inhibited cell viability in a time- and dose-dependent manner (Fig. 1A and B). Moreover, Annexin V and PI staining assay showed that apigenin induced cell death in a dose-dependent manner (Fig. 1C and D). To further explore the mechanism by apigenin induced cell death in HepG2 cells, we detected the anti-apoptotic and pro-apoptotic protein
Discussion
Apigenin, one of the common natural flavonoids, has been demonstrated to inhibit tumour cell proliferation, angiogenesis and induce apoptosis in various types of tumor cells [31]. Our previous study has been demonstrated that apigenin inhibits hepatoma cell growth both in vitro and in vivo, which is associated with induction of cell cycle arrest and apoptosis [32]. However, the special mechanism underlying the anti-tumor effects of apigenin in hepatoma cell is still not well understood. In the
Conflict of interest
The authors state that they have no conflict of interests.
References (47)
- et al.
Chemopreventive strategies in hepatocellular carcinoma
Nat. Rev. Gastro Hepat.
(2014) - et al.
4SC-202 activates ASK1-dependent mitochondrial apoptosis pathway to inhibit hepatocellular carcinoma cells
Biochem. Biophys. Res. Commun.
(2016) - et al.
Lancet Lancet
(2012) - et al.
Implication of intracellular ROS formation, caspase-3 activation and Egr-1 induction in platycodon d-induced apoptosis of U937 human leukemia cells
Biomed. Pharmacother.
(2009) - et al.
Rottlerin induces autophagy which leads to apoptotic cell death through inhibition of PI3K/Akt/mTOR pathway in human pancreatic cancer stem cells
Biochem. Pharmacol.
(2012) - et al.
Inactivated Sendai virus induces apoptosis and autophagy via the PI3K/Akt/mTOR/p70S6K pathway in human non-small cell lung cancer cells
Biochem. Biophys. Res. Commun.
(2015) - et al.
Current treatment strategies for inhibiting mTOR in cancer
Trends Pharmacol. Sci.
(2015) - et al.
Apigenin inhibits hepatoma cell growth through alteration of gene expression patterns
Phytomedicine
(2011) - et al.
Autophagy and metastasis: another double-edged sword
Curr. Opin. Cell Biol
(2010) - et al.
Defining biomarkers to predict sensitivity to PI3K/Akt/mTOR pathway inhibitors in breast cancer
Cancer Treat Rev.
(2013)
Development of PI3K/AKT/mTOR pathway inhibitors and their application in personalized therapy for non–small-cell lung cancer
J. Thorac. Oncol.
Effect of evodiagenine mediates photocytotoxicity on human breast cancer cells MDA-MB-231 through inhibition of PI3K/AKT/mTOR and activation of p38 pathways
Fitoterapia
A novel protoapigenone analog RY10-4 induces breast cancer MCF-7 cell death through autophagy via the Akt/mTOR pathway
Toxicol. Appl. Pharma
Hepatocellular carcinoma
N Engl. J. Med. New. Engl. J. Med.
Synergistic anticancer effects of curcumin and resveratrol in Hepa1-6 hepatocellular carcinoma cells
Oncol. Rep.
Apigenin and cancer chemoprevention: progress, potential and promise (review)
Int. J. Oncol.
Apigenin induced apoptosis in esophageal carcinoma cells by destruction membrane structures
Scanning
5-Fluorouracil combined with apigenin enhances anticancer activity through mitochondrial membrane potential (ΔΨm)-mediated apoptosis in hepatocellular carcinoma
Clin. Exp. Pharmacol. Physiol.
Apigenin induces the apoptosis and regulates MAPK signaling pathways in mouse macrophage ANA-1 cells
Plos One
Apigenin, a dietary flavonoid, inhibits proliferation of human bladder cancer T-24 cells via blocking cell cycle progression and inducing apoptosis
Cancer Cell Int.
Apigenin inhibits prostate cancer progression in TRAMP mice via targeting PI3K/Akt/FoxO pathway
Carcinogenesis
Apigenin induces ROS dependent apoptosis and ER stress in human endometriosis cells
J. Cell Physiol.
Apigenin inhibits tumor angiogenesis through decreasing HIF-1α and VEGF expression
Carcinogenesis
Cited by (271)
Manganese induces neuronal apoptosis by activating mTOR signaling pathway in vitro and in vivo
2024, Food and Chemical ToxicologyEffects of apigenin on gastric cancer cells
2024, Biomedicine and PharmacotherapySmall molecule agents for triple negative breast cancer: Current status and future prospects
2024, Translational Oncology