Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2015 Jul 17;16(7):16401-13.
doi: 10.3390/ijms160716401.

A New Ligustrazine Derivative-Selective Cytotoxicity by Suppression of NF-κB/p65 and COX-2 Expression on Human Hepatoma Cells. Part 3

Chenze Zhang  1 , Wenqiang Yan  2 , Bi Li  3 , Bing Xu  4 , Yan Gong  5 , Fuhao Chu  6 , Yuzhong Zhang  7 , Qiuli Yao  8 , Penglong Wang  9 , Haimin Lei  10
Affiliations

A New Ligustrazine Derivative-Selective Cytotoxicity by Suppression of NF-κB/p65 and COX-2 Expression on Human Hepatoma Cells. Part 3

Chenze Zhang et al. Int J Mol Sci. .

Abstract

A new anticancer ligustrazine derivative, 3β-hydroxyolea-12-en-28-oic acid- 3,5,6-trimethylpyrazin-2-methylester (T-OA, C38H58O3N2), was previously reported. It was synthesized via conjugating hepatoprotective and anticancer ingredients of traditional Chinese medicine. We found that T-OA exerted its anticancer activity by preventing the expression of nuclear transcription factor NF-κB/p65 and COX-2 in S180 mice. However, the selective cytotoxicity of T-OA on various kinds of cell lines has not been studied sufficiently. In the present study, compared with Cisplatin, T-OA was more toxic to human hepatoma cell line Bel-7402 (IC50 = 6.36 ± 1.56 µM) than other three cancer cell lines (HeLa, HT-29, BGC-823), and no toxicity was observed toward Madin-Darby canine kidney cell line MDCK (IC50 > 150 µM). The morphological changes of Bel-7402 cells demonstrated that T-OA had an apoptosis-inducing effect which had been substantiated using 4',6-diamidino-2-phenylindole (DAPI) staining, acridine orange (AO)/ethidium bromide (EB) staining, flow cytometry and mitochondrial membrane potential assay. Combining the immumohistochemical staining, we found T-OA could prevent the expression of NF-κB/p65 and COX-2 in Bel-7402 cells. Both of the proteins have been known to play roles in apoptosis and are mainly located in the nuclei. Moreover subcellular localization was performed to reveal that T-OA exerts in nuclei of Bel-7402 cells. The result was in accordance with the effects of down-regulating the expression of NF-κB/p65 and COX-2.

Keywords: NF-κB/p65 and COX-2; hepatoma; ligustrazine derivative; selective cytotoxicity.

PubMed Disclaimer

Figures

Figure 1
Figure 1
Structure of T-OA.
Figure 2
Figure 2
Selective cytotoxicity of T-OA on Bel-7402 and MDCK cell lines.
Figure 3
Figure 3
Morphological changes of Bel-7402 cells assessed by Giemsa staining. (a) Control group without T-OA; (b) Treated with 5 µM T-OA; (c) Treated with 10 µM T-OA; and (d) Treated with 20 µM T-OA. (×400).
Figure 4
Figure 4
DAPI (I) and acridine orange/ethidium bromide (AO/EB) (II) staining of the Bel-7402 cells treated by T-OA. (Ia) Control group without T-OA; (Ib) Treated with 5 µM T-OA; (Ic) Treated with 10 µM T-OA; (Id) Treated with 20 µM T-OA; (IIa) Control group without T-OA; (IIb) Treated with 5 µM T-OA; (IIc) Treated with 10 µM T-OA; (IId) Treated with 20 µM T-OA. (×200).
Figure 5
Figure 5
Apoptosis ratio detection by Annexin V/PI assay on the Bel-7402 cells treated by T-OA. (a) Control group without T-OA; (b) Treated with 5 µM T-OA; (c) Treated with 10 µM T-OA; (d) Treated with 20 µM T-OA.
Figure 6
Figure 6
Mitochondrial membrane potential detection by Rhodamine 123 staining on the Bel-7402 cells treated by T-OA. (a) Control group without T-OA; (b) Treated with 5 µM T-OA; (c) Treated with 10 µM T-OA; and (d) Treated with 20 µM T-OA.
Figure 7
Figure 7
Immunohistochemical analysis of NF-κB/p65 (I) and COX-2 (II) in control and T-OA treated groups of Bel-7402 cells. (Ia) Control group without T-OA; (Ib) Treated with 5 µM T-OA; (Ic) Treated with 10 µM T-OA; (Id) Treated with 20 µM T-OA; (IIa) Control group without T-OA; (IIb) Treated with 5 µM T-OA; (IIc) Treated with 10 µM T-OA; and (IId) Treated with 20 µM T-OA. (×400).
Figure 8
Figure 8
Subcellular localization of T-OA in Bel-7402 cells. (a) Control group at 488 nm; (b) Control group at 514 nm; (c) Merged image of control group; (d) T-OA fluorescent analogue treated group at 488 nm; (e) T-OA fluorescent analogue treated group at 514 nm; and (f) Merged image of T-OA fluorescent analogue treated group. (×600).
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

References

    1. Zhang J.L., Wang H., Chen C., Pi H.F., Ruan H.L., Zhang P., Wu J.Z. Addictive evaluation of cholic acid-verticinone ester, a potential cough therapeutic agent with agonist action of opioid receptor. Acta Pharmacol. Sin. 2009;30:559–566. doi: 10.1038/aps.2009.43. - DOI - PMC - PubMed
    1. Zhang J.L., Wang H., Pi H.F., Ruan H.L., Zhang P., Wu J.Z. Structural analysis and antitussive evaluation of five novel esters of verticinone and bile acids. Steroids. 2009;74:424–434. doi: 10.1016/j.steroids.2008.12.007. - DOI - PubMed
    1. Hao Y.Z., Wang P.L., Hong Y., Lei H.M. Synthesis and structure identification of tetramethylpyrazine-protocatechuic acid and effects on hypoxic-ischemic brain damage. Pharmacol. Clin. Chin. Mater. Med. 2010;26:41–45.
    1. Wang P.L., Cheng Y.T., Xu K., An Y.W., Wang W., Li Q.S., Han Q.J., Li Q., Zhang H.G., Lei H.M. Synthesis and anti-tumor evaluation of one novel tetramethylpyrazine-rhein derivative. Asian J. Chem. 2013;25:4885–4888.
    1. Chu F.H., Xu X., Li G., Gu S., Xu K., Gong Y., Xu B., Wang M.N., Zhang W.Z., Zhang Y.Z., et al. Amino acid derivatives of ligustrazine-oleanolic acid as new cytotoxic agents. Molecules. 2014;19:18215–18231. doi: 10.3390/molecules191118215. - DOI - PMC - PubMed

Publication types

LinkOut - more resources