A 3-(4,-5-dimethylthiazo-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay is a method that used to measure cell viability. It is based on the conversion of MTT by succinic dehydrogenase enzyme into insoluble formazan. Dissolution of formazan by using proper solvent is the most important step of MTT assay to obtain valid and reliable data. In this study, we observed several solvents [isopropanol, dimethyl sulfoxide (DMSO), and sodium dodecyl sulphate (SDS)] to validate MTT assay by using MCF-7 cells. The observation was performed by MTT addition at concentration of 0.5µg/µL, 3-4h cells incubation at 37°C, dissolution of formed formazan crystal and absorbance measurement at 570nm. The result showed that formazan completely dissolved in DMSO and 10% SDS. The most advantage of using SDS was it avoided the removal of partially dissolved formazan. In this observation, we also found that pH was a very important factor in SDS solution that affected the reaction. The use of optimal condition on MTT assay by SDS-0.01M HCl and SDS-0.025M HCl as formazan solvents showed that IC 50 of curcumin were 32.3±0.78µM and 24.08±1.72µM respectively, while WST-1 assay resulted IC 50 of curcumin 80.69±5.35µM. Altogether, this study strongly indicated that SDS-0.01M HCl was the best formazan solvent for MTT assay.

The use of 5-fluorouracil (5-FU) in colon cancer as the primary chemotherapy has not been meet satisfactory effectiveness.  Therefore, the development of new chemicals as a chemopreventive agent and a combination agent (co-chemotherapeutic agent) for colon cancer is important.  Pentagamavunone-0 (2,5-bis-(4'-hydroxy-3'-methoxybenzylidine) cyclopentanone) (PGV-0), one of curcumin analogs, exhibits cytotoxic effect and apoptosis induction in various cancer cell lines, including colon cancer cell, better than curcumin.  This study aimed to investigate the cytotoxic potency of PGV-0 in combination with 5-FU and their effects, in single or in combination, on cell cycle toward WiDr colon cancer cell line.  The cells were treated with combination concentrations of PGV-0 and 5-FU, and examined by MTT cell viability assay.  The value of combination index (CI) as a parameter of cytotoxic combination assay was measured by a combination index method.  Cells were stained with propidium iodide and the cell cycle distribution was determined by flowcytometry. CI calculation showed additive effects between PGV-0 and 5-FU.  Combination of PGV-0 and 5-FU gave synergism on cell cycle.  Single treatment of PGV-0 increased apoptosis, illustrated as subG1-phase accumulation, stronger than single treatment of 5-FU.  Meanwhile, combination of PGV-0 and 5-FU demonstrated S-phase arrest.  Based on these results, it can be concluded that PGV-0 has the potential to be developed as a co-chemotherapeutic agent for colon cancer but still requires further tracking of its molecular mechanisms.Keywords: Pentagamavunone-0 (PGV-0), 5-fluorouracil (5-FU), colon cancer,combination, cell cycle

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The production of recombinant proteins for clinical applications using mammalian cell technology has become a prevalent system because of its capacity in assembling functional proteins.  One of the main problems with CHO-K1 cells is that this cell has to grow in the presence of serum. However, the presence of serum will complicate the downstream step for protein production. Thus, protein produced in media without serum, theoretically, would be easier to purify.  Technically, this type of cell can be produced by growing the CHO-K1 cells in serum-free media by using adaptation method in suspension condition. This research showed that through sequential adaptation using conditioned media, the CHO-K1 cell line that produces the human erythropoietin gene (hEPO) was able to grow in suspension culture using serum-free media.  Based on Western blot analysis, it showed that the protein (hEPO) was able to be expressed in suspension culture with molecular mass of about 47 kDa.

Muscle tissues make up about 40-50% of the human bodies. Satellite cells, which present in between the basal lamina and myofiber, are the adult muscle stem cells or myoblasts which are important for the regeneration of muscle tissues. Anticancer agents generally possess high cytotoxicity to either cancer cells or normal cells. Their effects on muscle cells generate cachexia or the deterioration of muscle tissues. Chemopreventive agents which possess lower cyctotoxic effects are expected to show higher safety in normal cells. Therefore, we investigated the effects of chemopreventive agents curcumin, naringin, and epigallocathecin-3-gallate (EGCG), which show anticancer properties in cancer cells, in C2C12 myoblast cells. We observed the C2C12 cell viability by MTT and WST assays, cell migration by wound healing scratch assay, as well as differentiation assay after treatment with the chemopreventive agents. The results indicated that curcumin showed highest cytotoxicity compared to naringin and EGCG. In addition, naringin and EGCG exhibited lower cytotoxicity. Both naringin and EGCG inhibited C2C12 cell migration at cell density 150, 000 cells/ml. Whereas, at cell density 100, 000 cells/ml, there was no significant effects of naringin as well as EGCG. Altogether, the results suggest that naringin and EGCG possess lower cytotoxic effect on C2C12 myoblast cells whereas curcumin showed stronger cytotoxicity at concentration higher than 20 µM.