A Novel Indolizine Derivative Induces Apoptosis Through the Mitochondria p53 Pathway in HepG2 Cells

Abstract

Indolizine derivatives are a class of compounds with excellent biological activity. In this study, a series of indolizine derivatives, compound 1 (C1), compound 2 (C2), compound 3 (C3), and compound 4 (C4), were synthesized. 3-(4,5-dimethylthiazole)-2,5-diphenyltetraazolium bromide (MTT) assay was used to evaluate their cytotoxicity against HepG2 (p53-wild), A549, and HeLa cell lines. HepG2 cells apoptosis induced by C3 was determined using Hoechst staining and acridine orange/ethidium bromide staining. Cells’ apoptotic ratio was measured by Annexin V–FITC/PI double staining. Changes in mitochondrial membrane potential and intracellular reactive oxygen species (ROS) in HepG2 cells after C3 treatment were determined. Immunofluorescence staining and Western blot analysis were carried out to detect p53 levels and analyze the apoptosis-associated proteins, respectively. Moreover, the cytotoxic activity of C3 was examined in two other hepatocellular carcinoma (HCC) cell lines with different p53 status including Huh-7 cells (p53-mutant) and Hep3B cells (p53-null). The results indicated that C3 showed stronger inhibition towards HepG2 cells than other cell lines. Fluorescent staining and flow cytometry analysis confirmed that C3 induced apoptosis of HepG2 cells. C3 could also increase intracellular ROS and cause a decrease in the mitochondrial membrane potential. C3 promoted p53 activation and increased p53 accumulation in nuclei. The expression of p53 and Bax was increased with the down-regulation of Bcl-2, which promoted the release of cytochrome c and caspase-3 activation. Collectively, the study demonstrated that C3 caused HepG2 cell apoptosis via the mitochondria p53 pathway. These results inspired us to further develop indolizine derivatives as potential potent inhibitors against liver cancer.

Long non‐coding RNA ANRIL enhances mitochondrial function of hepatocellular carcinoma by regulating the MiR‐199a‐5p/ARL2 axis

“…Channels are open enough to release apoptotic factors such as cytochrome c from mitochondria into the cytoplasmic matrix, causing cell death . Mitochondrial apoptotic pathway is characterized as the increase of mitochondrial membrane potential and reactive oxygen . Here, it was found that ANRIL overexpression increased D‐loop expression, resulted in a higher ATP level, enhanced mitochondrial membrane potential, and increased mitochondrial marker expression.…”

“…However, the roles of ANRIL in regulating mitochondrial function are never been revealed. The main pathways of apoptosis are as follows: the external pathway is called death receptor pathway and the internal mode is mainly mitochondrial endogenous apoptosis . The second main function of mitochondria is to participate in cell energy metabolism, which affects ATP production through respiratory chain.…”

Colon tissue-accumulating mesoporous carbon nanoparticles loaded with Musca domestica cecropin for ulcerative colitis therapy

“…Unexpectedly, MCNs also demonstrated anti-apoptotic effects. To identify the protection mechanism, we assessed ROS production and mitochondrial membrane potential changes, which are closely related to apoptosis 50 . Increased ROS production inducing apoptosis was verified in Figure 5 E-F .…”

Antiproliferative and Apoptotic Effects of Cardamonin against Hepatocellular Carcinoma HepG2 Cells

“…HIF-1 can induce apoptosis through stabilization of wild-type p53, whereas it prevents apoptosis in the mutant p53 gene. HepG2 is the only HCC cell that has wild type p53 [38]. Moreover, HIF-1A can induce apoptosis via an increase in the stability of the product of the tumor suppressor gene p53.…”