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On the top of the serious array of hepatocellular carcinoma (HCC) ranks standstill among the most common cancers and causes massive death in the world population . It is mainly due to lack of potential
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drugs as well as adverse side effects by available therapeutics. One of the new strategy to treat this deadly liver cancer [2,3] is identifying natural bioactive compounds available in the dietary supplement that rekindles the direction of research against cancer diseases . Brown seaweeds are one of the major widespread groups of marine
macroalgae that prevail vast resorts of novel and rich bioactive com-pounds in a large extend. More predominately, the major cell wall component of brown seaweeds such as polysaccharide, alginate and mainly included fucoidan which is so called sulfated polysaccharide with fucose as a major principle sugar along with other monosugar like glucuronic acid, xylose and galactose. In addition, brown seaweeds have contained other elements such as minerals, tannins, polyphenols, vitamins along with proteins, lipids and carotene pigments [5–9]. These sorts of brown seaweed are being broadly used in East Asian countries as a supplement in terms of functional foods, healthcare, medicinal foods and pharmaceuticals [9,10]. There are viable in-vitro studies de-monstrated that the fucoidan act against various cancers including hepatocarcinoma and melanoma [10–12]. Furthermore, in-vitro and in-vivo studies that proved the fucoidan displaying wide range of biolo-gical activities such as anticoagulant, antithrombotic, antivirus, anti-inflammatory, antioxidant, anti-complementary, pro-survival mechan-isms and immunomodulatory activities and so on [13–15]. And also, recent and recent past reports had substantiated the evidence relay on anticancer effects of fucoidan by activating through apoptosis, sup-pression of metastasis and angiogenesis in different cancer cell types [16,17] and quite interestingly the molecular mechanisms of actions have not been fully clarified in a greater extend . However, antic-ancer properties of the fucoidan were meticulously documented in the following cancer cells such as lung, breast, liver, colon, prostate and Methylpiperidino pyrazole . A study suggested that fucoidan supplement has been improved and promoted in the deep-seated area of the fecal microbiota composition and repaired intensively the intestinal barrier function that could probably be used as an intestinal flora modulator for preventing further cancer burgeoning . While compared to medications in terms of food supplement, fucoidan can be utilized as an underlying complementary alternative therapeutics without being intolerable side effects for treating cancer [20,21].
Recent proven study in mainstream of MCF-7 breast cancer cells has been implied that the fucoidan can be a promising candidate for cancer therapy in combination of the cisplatin, doxorubicin and taxol . Further, fucoidan induced apoptosis in PC-3 human prostate cancer cells has also been well documented . Many studies demonstrated that the fucoidan has imperatively suppressed the cancer tumor and comprehensively enhanced the overall survival rate in cancer patients . In the present pragmatic investigation aimed at the anticancer effect of fucoidan in a hepatoblastoma-derived (HepG2) cell line that was thoroughly analyzed by the typical techniques such as cell viability, colony formation, cell migration, cell cycle progression, genetic damage and apoptosis along with their nuclear morphology and mitochondrial membrane potential.
2. Materials and methods
Bioassay kits such as Trevigen’s comet assay kit, annexin V-FITC assay kit, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bro-mide (MTT), Hoechst (33,342) and 5,5′6,6′-tetrachloro-1,1′3,3′tetra-ethylbenzimi-dazolycarbocyanine iodide (JC-1) staining solution, fu-coidan and propidium iodide (PI) were procured from Sigma-Aldrich. The RPMI-1640 medium, fetal bovine serum (FBS) and phosphate-buffered saline (PBS) were procured from Hi-Media (Mumbai, India). All the solvents and chemical were of analytical grade.
2.2. Cell viability assay
By the protocol of MTT assay was performed with MTT dye (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and whereby the MTT was converted into MTT-formazan in mitochondria of HepG2 cells. Based on the experimental process, the cells (1 × 105 cells/well) were seeded in 96-well plates and kept 6 h for adhering. Subsequently, Toxicology Reports 6 (2019) 556–563
the cells were constituted with fucoidan/quercetin standard at nuance of concentration such as 0, 50, 100 & 200 μg/ml (filtered by 0.2 μ Millipore filter) for 48 h. And then 100 μl of MTT dye (5 mg in 10 ml of serum free medium) was added into each well and kept at 5% CO2 incubator up to 4 h at 37◦C in the dark. Later on, the superficial media were removed and thereby precipitated formazan dissolved in 100 μl of 20% SDS (in 50% dimethyl formamide) and after construed in an ELISA reader at 540 nm . The percentage of inhibition (I%) was calculated using the following equation: I% = (Control-Treated)/Control×100%.