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Fulltext Effect of experimental treatment on GAPDH mRNA expression as a housekeeping gene in human diploid fibroblasts

Zainuddin A, Chua KH, Abdul Rahim N, Makpol S

BMC Mol. Biol., 2010;11:59.
PMID: 20707929 DOI: 10.1186/1471-2199-11-59

Several 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 gamma-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.
γ-Tocotrienol prevents cell cycle arrest in aged human fibroblast cells through p16INK4a pathway

Zainuddin A, Chua KH, Tan JK, Jaafar F, Makpol S

J Physiol Biochem, 2017 Feb;73(1):59-65.
PMID: 27743340 DOI: 10.1007/s13105-016-0524-2

Human diploid fibroblasts (HDFs) proliferation in culture has been used as a model of aging at the cellular level. Growth arrest is one of the most important mechanisms responsible for replicative senescence. Recent researches have been focusing on the function of vitamin E in modulating cellular signaling and gene expression. Therefore, the aim of this study was to elucidate the effect of palm γ-tocotrienol (vitamin E) in modulating cellular aging through p16INK4a pathway in HDF cells. Primary culture of senescent HDFs was incubated with 70 μM of palm γ-tocotrienol for 24 hours. Silencing of p16INK4a was carried out by siRNA transfection. RNA was extracted from the different treatment groups and gene expression analysis was carried out by real-time reverse transcription polymerase chain reaction. Proteins that were regulated by p16INK4a were determined by western blot technique. The finding of this study showed that p16INK4a mRNA was overexpressed in senescent HDFs, and hypophosphorylated-pRb and cyclin D1 protein expressions were increased (p
Fulltext Antioxidant capacities and total phenolic contents increase with gamma irradiation in two types of Malaysian honey

Hussein SZ, Yusoff KM, Makpol S, Yusof YA

Molecules, 2011 Jul 27;16(8):6378-95.
PMID: 21796076 DOI: 10.3390/molecules16066378

Two types of monofloral Malaysian honey (Gelam and Nenas) were analyzed to determine their antioxidant activities and total phenolic and flavonoid contents, with and without gamma irradiation. Our results showed that both types of honey can scavenge free radicals and exhibit high antioxidant-reducing power; however, Gelam honey exhibited higher antioxidant activity (p < 0.05) than Nenas honey, which is in good correlation (r = 0.9899) with its phenolic contents. Interestingly, we also noted that both irradiated honeys have higher antioxidant activities and total phenolic and flavonoid contents compared to nonirradiated honeys by Folin-Ciocalteu and UV-spectrophotometry methods, respectively. However, HPLC analysis for phenolic compounds showed insignificant increase between irradiated and nonirradiated honeys. The phenolic compounds such as: caffeic acid, chlorogenic acid, ellagic acid, p- coumaric acid, quercetin and hesperetin as indicated by HPLC method were found to be higher in Gelam honey versus Nenas honey. In conclusion, irradiation of honey causes enhanced antioxidant activities and flavonoid compounds.
Fulltext Chlorella vulgaris Improves the Regenerative Capacity of Young and Senescent Myoblasts and Promotes Muscle Regeneration

Zainul Azlan N, Mohd Yusof YA, Alias E, Makpol S

Oxid Med Cell Longev, 2019;2019:3520789.
PMID: 31281573 DOI: 10.1155/2019/3520789

Sarcopenia is characterized by the loss of muscle mass, strength, and function with ageing. With increasing life expectancy, greater attention has been given to counteracting the effects of sarcopenia on the growing elderly population. Chlorella vulgaris, a microscopic, unicellular, green alga with the potential for various pharmaceutical uses, has been widely studied in this context. This study is aimed at determining the effects of C. vulgaris on promoting muscle regeneration by evaluating myoblast regenerative capacity in vitro. Human skeletal myoblast cells were cultured and underwent serial passaging into young and senescent phases and were then treated with C. vulgaris, followed by the induction of differentiation. The ability of C. vulgaris to promote myoblast differentiation was analysed through cellular morphology, real-time monitoring, cell proliferation, senescence-associated β-galactosidase (SA-β-gal) expression, myogenic differentiation, myogenin expression, and cell cycle profiling. The results obtained showed that senescent myoblasts exhibited an enlarged and flattened morphology, with increased SA-β-gal expression, reduced myogenic differentiation, decreased expression of myogenin, and an increased percentage of cells in the G0/G1 phase. Treatment with C. vulgaris resulted in decreased SA-β-gal expression and promotion of myogenic differentiation, as observed via an increased fusion index, maturation index, myotube size, and surface area and an increased percentage of cells that stained positive for myogenin. In conclusion, C. vulgaris improves the regenerative capacity of young and senescent myoblasts and promotes myoblast differentiation, indicating its potential to promote muscle regeneration.
Fulltext Zebrafish: A Pharmacological Model for Learning and Memory Research

Tan JK, Nazar FH, Makpol S, Teoh SL

Molecules, 2022 Oct 30;27(21).
PMID: 36364200 DOI: 10.3390/molecules27217374

Learning and memory are essential to organism survival and are conserved across various species, especially vertebrates. Cognitive studies involving learning and memory require using appropriate model organisms to translate relevant findings to humans. Zebrafish are becoming increasingly popular as one of the animal models for neurodegenerative diseases due to their low maintenance cost, prolific nature and amenability to genetic manipulation. More importantly, zebrafish exhibit a repertoire of neurobehaviors comparable to humans. In this review, we discuss the forms of learning and memory abilities in zebrafish and the tests used to evaluate the neurobehaviors in this species. In addition, the pharmacological studies that used zebrafish as models to screen for the effects of neuroprotective and neurotoxic compounds on cognitive performance will be summarized here. Lastly, we discuss the challenges and perspectives in establishing zebrafish as a robust model for cognitive research involving learning and memory. Zebrafish are becoming an indispensable model in learning and memory research for screening neuroprotective agents against cognitive impairment.
Fulltext A Link between Mitochondrial Dysregulation and Idiopathic Autism Spectrum Disorder (ASD): Alterations in Mitochondrial Respiratory Capacity and Membrane Potential

Hassan H, Zakaria F, Makpol S, Karim NA

Curr Issues Mol Biol, 2021 Dec 16;43(3):2238-2252.
PMID: 34940131 DOI: 10.3390/cimb43030157

Autism spectrum disorder (ASD) is a neurological disorder triggered by various factors through complex mechanisms. Research has been done to elucidate the potential etiologic mechanisms in ASD, but no single cause has been confirmed. The involvement of oxidative stress is correlated with ASD and possibly affects mitochondrial function. This study aimed to elucidate the link between mitochondrial dysregulation and idiopathic ASD by focusing on mitochondrial respiratory capacity and membrane potential. Our findings showed that mitochondrial function in the energy metabolism pathway was significantly dysregulated in a lymphoblastoid cell line (LCL) derived from an autistic child (ALCL). Respiratory capacities of oxidative phosphorylation (OXPHOS), electron transfer of the Complex I and Complex II linked pathways, membrane potential, and Complex IV activity of the ALCL were analyzed and compared with control cell lines derived from a developmentally normal non-autistic sibling (NALCL). All experiments were performed using high-resolution respirometry. Respiratory capacities of OXPHOS, electron transfer of the Complex I- and Complex II-linked pathways, and Complex IV activity of the ALCL were significantly higher compared to healthy controls. Mitochondrial membrane potential was also significantly higher, measured in the Complex II-linked pathway during LEAK respiration and OXPHOS. These results indicate the abnormalities in mitochondrial respiratory control linking mitochondrial function with autism. Correlating mitochondrial dysfunction and autism is important for a better understanding of ASD pathogenesis in order to produce effective interventions.
Fulltext Effects of Microalgae on Metabolic Syndrome

Tamel Selvan K, Goon JA, Makpol S, Tan JK

Antioxidants (Basel), 2023 Feb 10;12(2).
PMID: 36830009 DOI: 10.3390/antiox12020449

Metabolic syndrome (MetS) is a cluster of metabolic disturbances, including abdominal obesity, hypertension, hypertriglyceridemia, reduced high-density lipoprotein cholesterol (HDL-C) and hyperglycemia. Adopting a healthier lifestyle and multiple drug-based therapies are current ways to manage MetS, but they have limited efficacy, albeit the prevalence of MetS is rising. Microalgae is a part of the human diet and has also been consumed as a health supplement to improve insulin sensitivity, inflammation, and several components of MetS. These therapeutic effects of microalgae are attributed to the bioactive compounds present in them that exhibit antioxidant, anti-inflammatory, anti-obesity, antihypertensive, hepatoprotective and immunomodulatory effects. Therefore, studies investigating the potential of microalgae in alleviating MetS are becoming more popular, but a review on this topic remains scarce. In this review, we discuss the effects of microalgae, specifically on MetS, by reviewing the evidence from scientific literature covering in vitro and in vivo studies. In addition, we also discuss the underlying mechanisms that modulate the effects of microalgae on MetS, and the limitations and future perspectives of developing microalgae as a health supplement for MetS. Microalgae supplementation is becoming a viable approach in alleviating metabolic disturbances and as a unique addition to the management of MetS.
Fulltext Human adipose-derived mesenchymal stem cells promote recovery of injured HepG2 cell line and show sign of early hepatogenic differentiation

Liau LL, Makpol S, Azurah AGN, Chua KH

Cytotechnology, 2018 Aug;70(4):1221-1233.
PMID: 29549558 DOI: 10.1007/s10616-018-0214-8

Currently, orthotopic liver transplantation is the gold standard therapy for liver failure. However, it is limited by the insufficient organ donor and risk of immune rejection. Stem cell therapy is a promising alternative treatment for liver failure. One of the most ideal sources of stem cells for regenerative medicine is adipose-derived stem cells (ADSCs). In this study, primary ADSCs seeded on cell culture insert were indirectly co-cultured with injured HepG2 to elucidate the role of ADSCs in promoting the recovery of injured HepG2 in non-contact manner. HepG2 recovery was determined by the surface area covered by cells and growth factor concentration was measured to identify the factors involved in regeneration. Besides, HepG2 were collected for q-PCR analysis of injury, hepatocyte functional and regenerative markers expression. For the ADSCs, expression of hepatogenic differentiation genes was analyzed. Results showed that non-contact co-culture with ADSCs helped the recovery of injured HepG2. ELISA quantification revealed that ADSCs secreted higher amount of HGF and VEGF to help the recovery of injured HepG2. Furthermore, HepG2 co-cultured with ADSCs expressed significantly lower injury markers as well as significantly higher regenerative and functional markers compared to the control HepG2. ADSCs co-cultured with injured HepG2 expressed significantly higher hepatic related genes compared to the control ADSCs. In conclusion, ADSCs promote recovery of injured HepG2 via secretion of HGF and VEGF. In addition, co-cultured ADSCs showed early sign of hepatogenic differentiation in response to the factors released or secreted by the injured HepG2.
Fulltext Gelam Honey Inhibits the Production of Proinflammatory, Mediators NO, PGE(2), TNF-α, and IL-6 in Carrageenan-Induced Acute Paw Edema in Rats

Hussein SZ, Mohd Yusoff K, Makpol S, Mohd Yusof YA

Evid Based Complement Alternat Med, 2012;2012:109636.
PMID: 22919407 DOI: 10.1155/2012/109636

Natural honey is well known for its therapeutic value and has been used in traditional medicine of different cultures throughout the world. The aim of this study was to investigate the anti-inflammatory effect of Malaysian Gelam honey in inflammation-induced rats. Paw edema was induced by a subplantar injection of 1% carrageenan into the rat right hind paw. Rats were treated with the nonsteroidal anti-inflammatory drug (NSAID) Indomethacin (10 mg/kg, p.o.) or Gelam honey at different doses (1 or 2 g/kg, p.o.). The increase in footpad thickness was considered to be edema, which was measured using a dial caliper. Plasma and paw tissue were collected to analyze the production of inflammatory mediators, such as NO, PGE(2), TNF-α, and IL-6, as well as iNOS and COX-2. The results showed that Gelam honey could reduce edema in a dose-dependent fashion in inflamed rat paws, decrease the production of NO, PGE(2), TNF-α, and IL-6 in plasma, and suppress the expression of iNOS, COX-2, TNF-α, and IL-6 in paw tissue. Oral pretreatment of Gelam honey at 2 g/kg of body weight at two time points (1 and 7 days) showed a significantly decreased production of proinflammatory cytokines, which was similar to the effect of the anti-inflammatory drug Indomethacin (NSAID), both in plasma and tissue. Thus, our results suggest that Gelam honey has anti-inflammatory effects by reducing the rat paw edema size and inhibiting the production of proinflammatory mediators. Gelam honey is potentially useful for treating inflammatory conditions.
Fulltext Zingiber Officinale Roscoe Prevents Cellular Senescence of Myoblasts in Culture and Promotes Muscle Regeneration

Mohd Sahardi NFN, Jaafar F, Mad Nordin MF, Makpol S

Evid Based Complement Alternat Med, 2020;2020:1787342.
PMID: 32419792 DOI: 10.1155/2020/1787342

Background: Ageing resulted in a progressive loss of muscle mass and strength. Increased oxidative stress in ageing affects the capacity of the myoblast to differentiate leading to impairment of muscle regeneration. Zingiber officinale Roscoe (ginger) has potential benefits in reversing muscle ageing due to its antioxidant property. This study aimed to determine the effect of ginger in the prevention of cellular senescence and promotion of muscle regeneration.
Methods: Myoblast cells were cultured into young and senescent state before treated with different concentrations of ginger standardised extracts containing different concentrations of 6-gingerol and 6-shogaol. Analysis on cellular morphology and myogenic purity was carried out besides determination of SA-β-galactosidase expression and cell cycle profile. Myoblast differentiation was quantitated by determining the fusion index, maturation index, and myotube size.
Results: Treatment with ginger extracts resulted in improvement of cellular morphology of senescent myoblasts which resembled the morphology of young myoblasts. Our results also showed that ginger treatment caused a significant reduction in SA-β-galactosidase expression on senescent myoblasts indicating prevention of cellular senescence, while cell cycle analysis showed a significant increase in the percentage of cells in the G0/G1 phase and reduction in the S-phase cells. Increased myoblast regenerative capacity was observed as shown by the increased number of nuclei per myotube, fusion index, and maturation index.
Conclusions: Ginger extracts exerted their potency in promoting muscle regeneration as indicated by prevention of cellular senescence and promotion of myoblast regenerative capacity.
Fulltext Chlorella vulgaris Modulates Genes and Muscle-Specific microRNAs Expression to Promote Myoblast Differentiation in Culture

Zainul Azlan N, Mohd Yusof YA, Alias E, Makpol S

Evid Based Complement Alternat Med, 2019;2019:8394648.
PMID: 31428175 DOI: 10.1155/2019/8394648

Background: Loss of skeletal muscle mass, strength, and function due to gradual decline in the regeneration of skeletal muscle fibers was observed with advancing age. This condition is known as sarcopenia. Myogenic regulatory factors (MRFs) are essential in muscle regeneration as its activation leads to the differentiation of myoblasts to myofibers. Chlorella vulgaris is a coccoid green eukaryotic microalga that contains highly nutritious substances and has been reported for its pharmaceutical effects. The aim of this study was to determine the effect of C. vulgaris on the regulation of MRFs and myomiRs expression in young and senescent myoblasts during differentiation in vitro.
Methods: Human skeletal muscle myoblast (HSMM) cells were cultured and serial passaging was carried out to obtain young and senescent cells. The cells were then treated with C. vulgaris followed by differentiation induction. The expression of Pax7, MyoD1, Myf5, MEF2C, IGF1R, MYOG, TNNT1, PTEN, and MYH2 genes and miR-133b, miR-206, and miR-486 was determined in untreated and C. vulgaris-treated myoblasts on Days 0, 1, 3, 5, and 7 of differentiation.
Results: The expression of Pax7, MyoD1, Myf5, MEF2C, IGF1R, MYOG, TNNT1, and PTEN in control senescent myoblasts was significantly decreased on Day 0 of differentiation (p<0.05). Treatment with C. vulgaris upregulated Pax7, Myf5, MEF2C, IGF1R, MYOG, and PTEN in senescent myoblasts (p<0.05) and upregulated Pax7 and MYOG in young myoblasts (p<0.05). The expression of MyoD1 and Myf5 in young myoblasts however was significantly decreased on Day 0 of differentiation (p<0.05). During differentiation, the expression of these genes was increased with C. vulgaris treatment. Further analysis on myomiRs expression showed that miR-133b, miR-206, and miR-486 were significantly downregulated in senescent myoblasts on Day 0 of differentiation which was upregulated by C. vulgaris treatment (p<0.05). During differentiation, the expression of miR-133b and miR-206 was significantly increased with C. vulgaris treatment in both young and senescent myoblasts (p<0.05). However, no significant change was observed on the expression of miR-486 with C. vulgaris treatment.
Conclusions: C. vulgaris demonstrated the modulatory effects on the expression of MRFs and myomiRs during proliferation and differentiation of myoblasts in culture. These findings may indicate the beneficial effect of C. vulgaris in muscle regeneration during ageing thus may prevent sarcopenia in the elderly.
Fulltext Effects of palm vitamin e on bone-formation-related gene expression in nicotine-treated rats

Abukhadir SS, Mohamed N, Makpol S, Muhammad N

Evid Based Complement Alternat Med, 2012;2012:656025.
PMID: 23049610 DOI: 10.1155/2012/656025

The study determines the effects of palm vitamin E on the gene expression of bone-formation-related genes in nicotine-treated rats. Male rats were divided into three groups: normal saline olive oil (NSO), nicotine olive oil (NO), and nicotine palm vitamin E (NE). The treatment was carried out in 2 phases. During the first 2 months, the NSO group received normal saline while the NO and NE groups received nicotine 7 mg/kg, 6 days a week, intraperitoneally. The following 2 months, normal saline and nicotine administration was stopped and was replaced with oral supplementation of olive oil for the NSO and NO groups and oral supplementation of palm vitamin E (60 mg/kg) for the NE group. Both femurs were harvested to determine the gene expression of bone morphogenetic protein-2 (BMP-2), Osterix (OSX), and Runt-related transcription factor 2 (RUNX2). Nicotine significantly downregulated the gene expression. This effect was reversed by palm vitamin E treatment. In conclusion, palm vitamin E may play a role in osteoblast differentiation and can be considered as an anabolic agent to treat nicotine-induced osteoporosis.
Fulltext Modulation of Cell Cycle Profile by Chlorella vulgaris Prevents Replicative Senescence of Human Diploid Fibroblasts

Saberbaghi T, Abbasian F, Mohd Yusof YA, Makpol S

Evid Based Complement Alternat Med, 2013;2013:780504.
PMID: 23573154 DOI: 10.1155/2013/780504

In this study, the effects of Chlorella vulgaris (CV) on replicative senescence of human diploid fibroblasts (HDFs) were investigated. Hot water extract of CV was used to treat HDFs at passages 6, 15, and 30 which represent young, presenescence, and senescence ages, respectively. The level of DNA damage was determined by comet assay while apoptosis and cell cycle profile were determined using FACSCalibur flow cytometer. Our results showed direct correlation between increased levels of damaged DNA and apoptosis with senescence in untreated HDFs (P < 0.05). Cell cycle profile showed increased population of untreated senescent cells that enter G0/G1 phase while the cell population in S phase decreased significantly (P < 0.05). Treatment with CV however caused a significant reduction in the level of damaged DNA and apoptosis in all age groups of HDFs (P < 0.05). Cell cycle analysis showed that treatment with CV increased significantly the percentage of senescent HDFs in S phase and G2/M phases but decreased the population of cells in G0/G1 phase (P < 0.05). In conclusion, hot water extract of Chlorella vulgaris effectively decreased the biomarkers of ageing, indicating its potential as an antiageing compound.
Fulltext Tocotrienol-rich fraction, [6]-gingerol and epigallocatechin gallate inhibit proliferation and induce apoptosis of glioma cancer cells

Rahman AA, Makpol S, Jamal R, Harun R, Mokhtar N, Ngah WZ

Molecules, 2014 Sep 12;19(9):14528-41.
PMID: 25221872 DOI: 10.3390/molecules190914528

Plant bioactives [6]-gingerol (GING), epigallocatechin gallate (EGCG) and asiaticoside (AS) and vitamin E, such as tocotrienol-rich fraction (TRF), have been reported to possess anticancer activity. In this study, we investigated the apoptotic properties of these bioactive compounds alone or in combination on glioma cancer cells. TRF, GING, EGCG and AS were tested for cytotoxicity on glioma cell lines 1321N1 (Grade II), SW1783 (Grade III) and LN18 (Grade IV) in culture by the (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxy-phenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt) (MTS) assay. With the exception of AS, combinations of two compounds were tested, and the interactions of each combination were evaluated by the combination index (CI) using an isobologram. Different grades of glioma cancer cells showed different cytotoxic responses to the compounds, where in 1321N1 and LN18 cells, the combination of EGCG + GING exhibited a synergistic effect with CI = 0.77 and CI = 0.55, respectively. In contrast, all combinations tested (TRF + GING, TRF + EGCG and EGCG + GING) were found to be antagonistic on SW1783 with CI values of 1.29, 1.39 and 1.39, respectively. Combined EGCG + GING induced apoptosis in both 1321N1 and LN18 cells, as evidenced by Annexin-V FITC/PI staining and increased active caspase-3. Our current data suggests that the combination of EGCG + GING synergistically induced apoptosis and inhibits the proliferation 1321N1 and LN18 cells, but not SW1783 cells, which may be due to their different genetic profiles.
Fulltext Vitamin E in sarcopenia: current evidences on its role in prevention and treatment

Khor SC, Abdul Karim N, Ngah WZ, Yusof YA, Makpol S

Oxid Med Cell Longev, 2014;2014:914853.
PMID: 25097722 DOI: 10.1155/2014/914853

Sarcopenia is a geriatric syndrome that is characterized by gradual loss of muscle mass and strength with increasing age. Although the underlying mechanism is still unknown, the contribution of increased oxidative stress in advanced age has been recognized as one of the risk factors of sarcopenia. Thus, eliminating reactive oxygen species (ROS) can be a strategy to combat sarcopenia. In this review, we discuss the potential role of vitamin E in the prevention and treatment of sarcopenia. Vitamin E is a lipid soluble vitamin, with potent antioxidant properties and current evidence suggesting a role in the modulation of signaling pathways. Previous studies have shown its possible beneficial effects on aging and age-related diseases. Although there are evidences suggesting an association between vitamin E and muscle health, they are still inconclusive compared to other more extensively studied chronic diseases such as neurodegenerative diseases and cardiovascular diseases. Therefore, we reviewed the role of vitamin E and its potential protective mechanisms on muscle health based on previous and current in vitro and in vivo studies.
Fulltext Gelam honey attenuated radiation-induced cell death in human diploid fibroblasts by promoting cell cycle progression and inhibiting apoptosis

Tengku Ahmad TA, Jaafar F, Jubri Z, Abdul Rahim K, Rajab NF, Makpol S

BMC Complement Altern Med, 2014;14:108.
PMID: 24655584 DOI: 10.1186/1472-6882-14-108

The interaction between ionizing radiation and substances in cells will induce the production of free radicals. These free radicals inflict damage to important biomolecules such as chromosomes, proteins and lipids which consequently trigger the expression of genes which are involved in protecting the cells or repair the oxidative damages. Honey has been known for its antioxidant properties and was used in medical and cosmetic products. Currently, research on honey is ongoing and diversifying. The aim of this study was to elucidate the role of Gelam honey as a radioprotector in human diploid fibroblast (HDFs) which were exposed to gamma-rays by determining the expression of genes and proteins involved in cell cycle regulation and cell death.
Fulltext Piper betle induces phase I & II genes through Nrf2/ARE signaling pathway in mouse embryonic fibroblasts derived from wild type and Nrf2 knockout cells

Wan Hasan WN, Kwak MK, Makpol S, Wan Ngah WZ, Mohd Yusof YA

BMC Complement Altern Med, 2014;14:72.
PMID: 24559113 DOI: 10.1186/1472-6882-14-72

Nuclear factor-erythroid 2 p45 related factor 2 (Nrf2) is a primary transcription factor, protecting cells from oxidative stress by regulating a number of antioxidants and phase II detoxifying enzymes. Dietary components such as sulforaphane in broccoli and quercetin in onions have been shown to be inducers of Nrf2. Piper betle (PB) grows well in tropical climate and the leaves are used in a number of traditional remedies for the treatment of stomach ailments and infections among Asians. The aim of this study was to elucidate the effect of Piper betle (PB) leaves extract in Nrf2 signaling pathway by using 2 types of cells; mouse embryonic fibroblasts (MEFs) derived from wild-type (WT) and Nrf2 knockout (N0) mice.
Fulltext Comparative effects of biodynes, tocotrienol-rich fraction, and tocopherol in enhancing collagen synthesis and inhibiting collagen degradation in stress-induced premature senescence model of human diploid fibroblasts

Makpol S, Jam FA, Khor SC, Ismail Z, Mohd Yusof YA, Ngah WZ

Oxid Med Cell Longev, 2013;2013:298574.
PMID: 24396567 DOI: 10.1155/2013/298574

Biodynes, tocotrienol-rich fraction (TRF), and tocopherol have shown antiaging properties. However, the combined effects of these compounds on skin aging are yet to be investigated. This study aimed to elucidate the skin aging effects of biodynes, TRF, and tocopherol on stress-induced premature senescence (SIPS) model of human diploid fibroblasts (HDFs) by determining the expression of collagen and MMPs at gene and protein levels. Primary HDFs were treated with biodynes, TRF, and tocopherol prior to hydrogen peroxide (H2O2) exposure. The expression of COL1A1, COL3A1, MMP1, MMP2, MMP3, and MMP9 genes was determined by qRT-PCR. Type I and type III procollagen proteins were measured by Western blotting while the activities of MMPs were quantified by fluorometric Sensolyte MMP Kit. Our results showed that biodynes, TRF, and tocopherol upregulated collagen genes and downregulated MMP genes (P < 0.05). Type I procollagen and type III procollagen protein levels were significantly increased in response to biodynes, TRF, and tocopherol treatment (P < 0.05) with reduction in MMP-1, MMP-2, MMP-3, and MMP-9 activities (P < 0.05). These findings indicated that biodynes, TRF, and tocopherol effectively enhanced collagen synthesis and inhibited collagen degradation and therefore may protect the skin from aging.
Fulltext Comparative effect of Piper betle, Chlorella vulgaris and tocotrienol-rich fraction on antioxidant enzymes activity in cellular ageing of human diploid fibroblasts

Makpol S, Yeoh TW, Ruslam FA, Arifin KT, Yusof YA

BMC Complement Altern Med, 2013;13:210.
PMID: 23948056 DOI: 10.1186/1472-6882-13-210

Human diploid fibroblasts (HDFs) undergo a limited number of cellular divisions in culture and progressively reach a state of irreversible growth arrest, a process termed cellular ageing. Even though beneficial effects of Piper betle, Chlorella vulgaris and tocotrienol-rich fraction (TRF) have been reported, ongoing studies in relation to ageing is of interest to determine possible protective effects that may reverse the effect of ageing. The aim of this study was to evaluate the effect of P. betle, C. vulgaris and TRF in preventing cellular ageing of HDFs by determining the activity of antioxidant enzymes viz.; catalase, superoxide dismutase (SOD) and glutathione peroxidase.
Differences in protein changes between stress-induced premature senescence and replicative senescence states

Aan GJ, Hairi HA, Makpol S, Rahman MA, Karsani SA

Electrophoresis, 2013 Aug;34(15):2209-17.
PMID: 23712505 DOI: 10.1002/elps.201300086

Replicative senescence and stress-induced premature senescence (SIPS) cells are known to share certain traits. However, whether these cells are different at the protein level is unclear. Thus, this study has utilized proteomics to identify differences in the proteomes of replicative senescence and SIPS cells compared to normal cells. Replicative senescence was induced by serial passage of normal cells in culture. SIPS was established by exposure to H2 O2 at a subcytotoxic concentration of 20 μM for two weeks. Following 2DE, protein profiles were compared and protein spots that changed in abundance were identified by MALDI-TOF MS. Quantitative real-time RT-PCR was then performed to evaluate the transcript expression of selected altered proteins. A total of 24 and 10 proteins were found to have changed in abundance in replicative senescence and SIPS cells, respectively, when compared to young cells. Quantitative RT-PCR revealed that nine genes showed the same direction of change as observed in the proteomics analysis. Very little overlap was observed between proteins that changed in replicative senescence and SIPS cells, suggesting that although both SIPS and replicative senescence cells share hallmarks of cellular senescence, they were different in terms of proteins that changed in abundance.

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