Effects of polysaccharides from abalone (Haliotis discus hannai Ino) on HepG2 cell proliferation
Introduction
Abalone, Haliotis discus hannai Ino, is a large, single-shelled marine mollusc of the genus Haliotis. Abalone is widely cultured in East Asia as an economically important food resource [1]. Owing to its nutritive and pharmaceutical value, abalone has received extensive attention [1], [2], [3], [4], [5]. Abalone viscera accounts for 15–25% of the total body weight of the abalone, and this visceral matter is normally discarded as industrial waste [3]. The visceral waste contains many proteins, polysaccharides and fatty acids [4]. A substantial number of studies have been published regarding the activities of abalone polysaccharides, including antitumour [4], immunoregulatory [5] and antioxidative [6] activities. However, there are few reports concerning the cytoproliferative activity of the polysaccharides from pleopods and from the viscera of H. discus hannai Ino.
There has been a rapid development in the fields of cell therapy and regenerative medicine and in the production of bio-medicine using mammalian cell culture [7]. Accelerating cell proliferation is fundamental for these applications. It has been reported that sericin, a protein derived from silkworms, is a good supplement for cell culture media to accelerate the proliferation of mammalian cells [8], [9]. Sericin was added to freezing media as an alternative to foetal bovine serum (FBS) and successfully improved the survival of various cell lines during cryopreservation [10], [11]. Cell proliferation depends on intracellular signal transduction mediated by receptors, such as enzyme-linked receptors and protein degradation-dependent receptors [12]. Cyclins are positive regulators of cell cycle progression that are produced at specific periods during the cell cycle, and their expression levels and locations are tightly controlled [13]. Cyclins are the positive regulatory subunits of a class of protein kinases termed cyclin-dependent kinases (CDKs) [14]. Earlier studies have reported that sulphated polysaccharides from the sea cucumber Stichopus iaponicus could induce the proliferation of rat astrocytes by causing the accumulation of Cyclin D1, which integrates with the extracellular signals to activate CDK4 and/or CDK6 [15]. Progesterone was also able to promote the viability of mouse embryonic stem cells through the up-regulation of cyclins and cyclin-dependent kinases [16].
In the present study, three types of polysaccharides, AAP, AVAP I, and AVAP II, were isolated from the abalone H. discus hannai Ino and purified. The polysaccharides’ compositions were subsequently identified. We investigated the cell proliferation activity of these polysaccharides, and further studies examined AVAP I and the relationship between its composition and its effects on cell proliferation. We also examined whether the cell cycle-regulating genes were involved in inducing proliferation. Our results indicated that AVAP I had a significant effect on HepG2 cell proliferation and could be used as a supplement for stimulating the proliferation of mammalian cells.
Section snippets
Materials
Fresh abalones, cultivated in Jiaonan, Shandong Province, China, were collected in the Nanshan aquatic market of Qingdao, shelled, eviscerated, vacuum freeze-dried and crushed. DEAE-cellulose anion-exchange resin was from Whatman (Brentford, England), and Sephacryl™ S-300 HR was from Pharmacia Co. (Uppsala, Sweden). A TSK G4000 PWXL column was from TOSOH Biosep (Tokyo, Japan). The human hepatoma cell line HepG2 was purchased from the Institute of Basic Medicine, Shandong Academy of Medical
Isolation, purification and chemical analysis of the polysaccharides
Three polysaccharides, AAP, AVAP I and AVAP II, were isolated and purified from the pleopods and viscera of H. discus hannai Ino. The extracted crude polysaccharide was firstly purified by ion-exchange chromatography on a DEAE-cellulose column and the main polysaccharide fraction from the pleopods was eluted with 0.42–0.60 mol/L NaCl and collected, two fractions from the viscera were respectively eluted with 0.28–0.40 mol/L NaCl and 0.44–0.56 mol/L NaCl and collected. The pooled polysaccharide
Discussion
In the present study, we isolated three polysaccharides (AAP, AVAP I and AVAP II) from the abalone H. discus hannai Ino. The composition analysis of the polysaccharides revealed that all three were sulphated, and their sulphate contents were 21.5%, 15.2% and 20.9% for AAP, AVAP I and AVAP II, respectively. The monosaccharide composition of AVAP I and AVAP II was similar and primarily composed of Rha, GlcA and Gal, but the composition of AAP was different, containing GlcA, GalN, Fuc and Gal. Wu
Acknowledgments
This study was supported by the National Marine Public Welfare Scientific Research Project of China (No. 201105029) and the Programme for Changjiang Scholars and Innovative Research Team in University (IRT1188).
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