Effect of active fraction of Eriocaulon sieboldianum on human leukemia K562 cells via proliferation inhibition, cell cycle arrest and apoptosis induction

https://doi.org/10.1016/j.etap.2015.11.001Get rights and content

Highlights

  • We identified the most anticancer fraction of E. sieboldianum.

  • E3 could effectively inhibit Aurora kinase and block cell cycle of K562 cells.

  • E3 could induce apoptosis of K562 cells via p53 dependent pathway.

  • E3 could induce apoptosis of K562 cells via mitochondria apoptosis pathway.

  • This study could raise the application value of E. sieboldianum in cancer therapy.

Abstract

Eriocaulon sieboldianum (Sieb. & Zucc. ex Steud.), a genus of Eriocaulon in the Eriocaulaceae family, is an edible and medicinal plant used in traditional Chinese medicine. It was processed into healthcare beverages for expelling wind-heat, protecting eyes, and reducing blood fat. Also, it has been used with other herbs as Traditional Chinese herbal compound to treat cancer as adjuvants in tumor therapy in China. However, the active fractions and precise cellular mechanisms of E. sieboldianum extract remain to be illustrated. The goal of this study was to investigate the effects of the active fraction of E. sieboldianum on the growth of K562 cells and understand the possible mechanisms of its action. Our findings suggested that the fraction E3 of E. sieboldianum could effectively inhibit the activity of Aurora kinase and induce apoptosis via blocking cell cycle, up-regulating the expression of proapoptotic proteins including p53 and Bax and reducing the expression of Bcl-2. The levels of Cytochrome C, cleaved caspase-9, cleaved caspase-3 and cleaved PARP were also found to be increased after treatment with fraction E3 of E. sieboldianum.

This study could improve the development of E. sieboldianum and raise its application value in cancer adjuvant therapy. Considering it is both a dietary supplement and a traditional Chinese herbal medicine which exhibits anticancer activities, it can be developed into functional food.

Introduction

Eriocaulon sieboldianum Sieb. & Zucc. ex Steud. is a genus of Eriocaulon in the Eriocaulaceae family under the name “Sai-Gu-Jing-Cao” (Siebold Pipewort) in China. So far, Ericaulon buergerianum, sold on the market, includes the whole grass of E. sieboldianum and E. buergerianum. Both they have been used in traditional Chinese medicine as an anti-inflammatory, ophthalmic and antimicrobial medicine (Chinese Pharmaceutical Association, 2006, Chinese Pharmacopoeia Commission, 2010). Often in combination with other herbs, they are also processed into healthcare beverages for expelling wind-heat, protecting eyes, and reducing blood fat (Song et al., 2008, Xu et al., 2013). Besides, E. sieboldianum extract has been used to treat cancer in conjunction with other herbs as adjuvants in tumor therapy in China (Duke and Ayensu, 1985a, Duke and Ayensu, 1985b, Graham et al., 2000). Three stigmastane-skeleton sterols have been isolated and identified from the whole plants of E. sieboldianum by Song et al., including stigmasta-7, 22-dien-3β, 4β-diol, which is a new compound, stigmasterol-3-O-β-d-glucopyranoside and stigmasta-5-en-3β-ol (β-sitosterol) (Song et al., 2008). However, the active fractions and the anticancer mechanisms of E. sieboldianum are remained to be determined.

Aurora kinase is a family of serine/threonine kinases in mammalian. In human, Aurora family comprises of Aurora kinase A (Aurora-A), Aurora kinase B (Aurora-B), and Aurora kinase C (Aurora-C). Aurora kinases play an essential role in orchestrating bipolar spindle establishment, chromosome segregation and cytokinesis. Aurora kinase was reported to be strongly associated with human cancer and was demonstrated to be overexpressed in many human cancers (Fu et al., 2007). Aurora-A regulates chromosome maturation and mitotic spindle formation; Aurora-B controls chromosomal segregation and cytokinesis (Hauf et al., 2003, Katayama and Sen, 2010); whereas, Aurora-C is involved in meiosis (Zekri et al., 2012). It is reported that Aurora-B is responsible for the mitotic phosphorylation of Ser10 residue of histone H3 (Wei et al., 1999) and this phosphorylation is thought to be necessary for chromosome condensation (Vader and Lens, 2008). Furthermore, autophosphorylation of Aurora-B within its activation segment occurs on threonine 232 in human Aurora-B and this autophosphorylation is important for the full activation of Aurora-B (Giet et al., 2005). Previous studies proposed Aurora-B as a promising drug target in cancer treatment (Salmela et al., 2012). So far, many inhibitors of Aurora-B have been reported and several inhibitors are in phase I/II clinical trial. Besides, three flavonoids, luteolin, eupatorin and S39 have been demonstrated to be inhibitors of aurora kinases (Green et al., 2011, Xie et al., 2012, Li et al., 2013). However, there is little reports about natural inhibitors of Aurora-A.

Apoptosis plays a key role in maintenance of homeostasis in organisms. Aurora kinase has been reported to play a pivotal role in cell proliferation and survival in cancer cells, and apoptosis is induced in several cancers after aurora kinase inhibition (Jung et al., 2005). P53 was found to be necessary for cellular senescence induced by alteration of genes involving mitosis and chromosome segregation, including Aurora kinase overexpression (Jung et al., 2005, Huang et al., 2008). Aurora kinase inhibition also led to changes of several apoptosis-related protein expressions including Bax, Bcl-2 and caspases (Huang et al., 2008).

In this study, the effects of E. sieboldianum extract on Aurora kinase and apoptosis were investigated. In short, the mechanism of the anti-tumor activity of the E. sieboldianum extract against K562 cells was examined.

Section snippets

Plant collection and sample preparation

E. sieboldianum was purchased from Jiangxi province of China. It was identified and authenticated by Prof. Jincai Lu, School of Traditional Chinese Materia Medica, Shenyang Pharmaceutical University, China. A voucher specimen (No. 20130301) was deposited at the Department of Pharmacy, General Hospital of Shenyang Military Area Command, Shenyang, China.

Chopped and air-dried whole plants of E. sieboldianum (1.0 kg) were extracted with 60% EtOH for three times (2 h each time) and concentrated under

NMR and HPLC analysis of E. sieboldianum extract and fraction E3

In 1H NMR spectrum, EAC and NBA extract shown a group signals of aromatic hydrogen from δ6.5 to δ8.5 (Fig. 1A). However, almost no obvious signals of aromatic protons were observed in the 1H NMR spectrum of PE extract and only few signals were found in this interval. In addition, NBA extract exhibited the signals of protons from δ3.0 to δ5.5. HPLC analysis indicated that peaks of PE extract and DCM extract were observed after 40 min when the mobile phase of EAC extract was used, suggesting that

Discussion and conclusion

In recent years, many biological activities have been studied from numerous species of plants or crops (Hu et al., 2013). Traditional Chinese medicine (TCM) demonstrates obvious advantages in anti-tumor treatment for its multiple targets, radiation resistance and immunity enhancement. Besides, Chinese medicine treatment can prolong survival of chemotherapy patients and improve their quality of life. Hence, many Chinese medicine preparations have been widely used in the prevention and treatment

Conflict of interest

The authors declare no conflicts of interest.

Acknowledgment

This study was supported by Liaoning science and technology research projects (no.2013225086).

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