BackgroundSeveral genes have been used as housekeeping genes and choosing an appropriate reference gene is important for accurate quantitative RNA expression in real time RT-PCR technique. The expression levels of reference genes should remain constant between the cells of different tissues and under different experimental conditions. The purpose of this study was to determine the effect of different experimental treatments on the expression of glyceraldehyde 3-phosphate dehydrogenase (GAPDH) mRNA so that the reliability of GAPDH as reference gene for quantitative real time RT-PCR in human diploid fibroblasts (HDFs) can be validated. HDFs in 4 different treatment groups viz; young (passage 4), senescent (passage 30), H2O2-induced oxidative stress and γ-tocotrienol (GTT)-treated groups were harvested for total RNA extraction. Total RNA concentration and purity were determined prior to GAPDH mRNA quantification. Standard curve of GAPDH expression in serial diluted total RNA, melting curve analysis and agarose gel electrophoresis were used to determine the reliability of GAPDH as reference gene.ResultsHDFs with different experimental treatments exhibited diverse cell morphology with different expression of senescence-associated β-galactosidase (SA β-gal) activity. However the expression level of GAPDH was consistent in all treatment groups.ConclusionThe study demonstrated that GAPDH is reliable as reference gene for quantitative gene expression analysis in HDFs. Therefore it can be used as housekeeping gene for quantitative real time RT-PCR technique in human diploid fibroblasts particularly in studying cellular senescence.

In this study, we determined the molecular mechanism ofγ-tocotrienol (GTT) in preventing cellular aging by focusing on its anti-apoptotic effect in stress-induced premature senescence (SIPS) model of human diploid fibroblasts (HDFs). Results obtained showed that SIPS exhibited senescent-phenotypic characteristic, increased expression of senescence-associatedβ-galactosidase (SAβ-gal) and promoted G0/G1cell cycle arrest accompanied by shortening of telomere length with decreased telomerase activity. Both SIPS and senescent HDFs shared similar apoptotic changes such as increased Annexin V-FITC positive cells, increased cytochromecrelease and increased activation of caspase-9 and caspase-3 (P<0.05). GTT treatment resulted in a significant reduction of Annexin V-FITC positive cells, inhibited cytochromecrelease and decreased activation of caspase-9 and caspase-3 (P<0.05). Gene expression analysis showed that GTT treatment down regulated BAX mRNA, up-regulated BCL2A1 mRNA and decreased the ratio of Bax/Bcl-2 protein expression (P<0.05) in SIPS. These findings suggested that GTT inhibits apoptosis by modulating the upstream apoptosis cascade, causing the inhibition of cytochromecrelease from the mitochondria with concomitant suppression of caspase-9 and caspase-3 activation. In conclusion, GTT delays cellular senescence of human diploid fibroblasts through the inhibition of intrinsic mitochondria-mediated pathway which involved the regulation of pro- and anti-apoptotic genes and proteins.

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Initial moisture content of ground powder is very important factor in maintaining the physical quality of the product during storage. This research was conducted to investigate the effect of initial moisture content of Orthosiphon stamineus ground powder on physical properties such as moisture content, water activity and colour properties during short term storage period. In this research, the experimental treatments were arranged in a completed randomized design (CRD) that consisted of two main factors which were initial moisture content (8, 13 and 20 %) and storage period (0, 2, 4, 6, 8 and 10 weeks). The changes of moisture content, water activity and colour properties were analyzed using moisture analyzer, water activity meter and chroma meter, respectively. From the results, it was revealed that the best initial moisture content to maintain the quality of O. stamineus ground powder was 13 %. At early of storage period, moisture content and water activity were slightly reduced and they started to stable at week 8 to 10. The consistency of moisture content and water activity is expected to last for the next 6 months under same storage condition. However, it could be observed that the colour of O. staminus ground powder was slightly affected during short term storage.

In recent years, the consumption of antioxidants in athletes has markedly increased. Exercise and endurance activities require a large energy as well as cause a high turnover of antioxidant capacity through sweat losses, musculoskeletal repair process and metabolism in the body. Antioxidant system in the body becomes depleted during exercise which causes cells and tissues are more susceptible to reactive oxygen species (ROS). Antioxidants includes vitamin E have received attention predominantly as a nutritional strategy to maximize sport performance and preventing detrimental effects of ROS, which continuously generated during strenuous exercise. Vitamin E is a nonenzymatic and potent lipid-soluble antioxidants that able to scavenge and neutralize the ROS. Vitamin E has the ability to protect against oxidative stress in human and animal, as well as enhances physical endurance. Therefore, the aim of this review is to evaluate the potential effects of vitamin E in modulating oxidative stress in order to enhance physical performance in athletes. To date there is still poor evidence that dietary supplementation with vitamin E will improve sport performance. However it can be suggested that antioxidants supplementation rich in vitamin E may act as alternative strategy to improve physical performance.