Publications by authors named "Ammu Kutty Radhakrishnan"

33 Publications

Evaluating Anticancer and Immunomodulatory Effects of (Arthrospira) and Gamma-Tocotrienol Supplementation in a Syngeneic Mouse Model of Breast Cancer.

Nutrients 2021 Jul 6;13(7). Epub 2021 Jul 6.

School of Health Sciences, International Medical University, Kuala Lumpur 57000, Malaysia.

Nutrition can modulate host immune responses as well as promote anticancer effects. In this study, two nutritional supplements, namely gamma-tocotrienol (γT3) and were evaluated for their immune-enhancing and anticancer effects in a syngeneic mouse model of breast cancer (BC). Five-week-old female BALB/c mice were fed , γT3, or a combination of and γT3 ( + γT3) for 56 days. The mice were inoculated with 4T1 cells into their mammary fat pad on day 28 to induce BC. The animals were culled on day 56 for various analyses. A significant reduction ( < 0.05) in tumor volume was only observed on day 37 and 49 in animals fed with the combination of γT3 + . There was a marked increase ( < 0.05) of CD4/CD127 T-cells and decrease ( < 0.05) of T-regulatory cells in peripheral blood from mice fed with either γT3 or The breast tissue of the combined group showed abundant areas of necrosis, but did not prevent metastasis to the liver. Although there was a significant increase ( < 0.05) of MIG-6 and Cadherin 13 expression in tumors from γT3-fed animals, there were no significant ( > 0.05) differences in the expression of MIG-6, Cadherin 13, BIRC5, and Serpine1 upon combined feeding. This showed that combined γT3 + treatment did not show any synergistic anticancer effects in this study model.
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http://dx.doi.org/10.3390/nu13072320DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8308567PMC
July 2021

Reduced infiltration of regulatory T cells in tumours from mice fed daily with gamma-tocotrienol supplementation.

Clin Exp Immunol 2021 Jul 31. Epub 2021 Jul 31.

Pathology Division, School of Medicine, International Medical University, Kuala Lumpur, Malaysia.

Gamma-tocotrienol (γT3) is an analogue of vitamin E with beneficial effects on the immune system, including immune-modulatory properties. This study reports the immune-modulatory effects of daily supplementation of γT3 on host T helper (Th) and T regulatory cell (T ) populations in a syngeneic mouse model of breast cancer. Female BALB/c mice were fed with either γT3 or vehicle (soy oil) for 2 weeks via oral gavage before they were inoculated with syngeneic 4T1 mouse mammary cancer cells (4T1 cells). Supplementation continued until the mice were euthanized. Mice (n = 6) were euthanized at specified time-points for various analysis (blood leucocyte, cytokine production and immunohistochemistry). Tumour volume was measured once every 7 days. Gene expression studies were carried out on tumour-specific T lymphocytes isolated from splenic cultures. Supplementation with γT3 increased CD4 (p < 0.05), CD8 (p < 0.05) T-cells and natural killer cells (p < 0.05) but suppressed T cells (p < 0.05) in peripheral blood when compared to animals fed with the vehicle. Higher interferon (IFN)-γ and lower transforming growth factor (TGF)-ꞵ levels were noted in the γT3 fed mice. Immunohistochemistry findings revealed higher infiltration of CD4 cells, increased expression of interleukin-12 receptor-beta-2 (IL-12ꞵ2R), interleukin (IL)-24 and reduced expression of cells that express the forkhead box P3 (FoxP3) in tumours from the γT3-fed animals. Gene expression studies showed the down-regulation of seven prominent genes in splenic CD4 T cells isolated from γT3-fed mice. Supplementation with γT3 from palm oil-induced T cell-dependent cell-mediated immune responses and suppressed T cells in the tumour microenvironment in a syngeneic mouse model of breast cancer.
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http://dx.doi.org/10.1111/cei.13650DOI Listing
July 2021

Poloxamer 188 (P188), A Potential Polymeric Protective Agent for Central Nervous System Disorders: A Systematic Review.

Curr Neuropharmacol 2021 May 28. Epub 2021 May 28.

Neuropharmacology Research Laboratory, Jeffrey Cheah School of Medicine and Health Sciences, Monash University Malaysia, Bandar Sunway 47500, Selangor, Malaysia.

Poloxamer 188 (P188) is an FDA-approved biocompatible block copolymer composed of repeating units of poly(ethylene oxide) (PEO) and poly(propylene oxide) (PPO). Due to its amphiphilic nature and high hydrophile-lipophile balance (HLB) value of 29, P188 is used as a stabilizer/emulsifier in many cosmetics and pharmaceuticals preparations. While the applications of P188 as an excipient are widely explored, the data on the pharmacological activity of P188 are scarce. Notably, the neuroprotective potential of P188 has gained a lot of interest. Therefore, this systematic review is aimed to review evidence to support the neuroprotective potential of P188 in CNS disorders. The PRISMA model was used, and five databases (Google Scholar, SCOPUS, Wiley Online Library, ScienceDirect, and PubMed) were searched with relevant keywords. The research resulted in 11 articles, which met the inclusion criteria. These articles described the protective effects of P188 on traumatic brain injury or mechanical injury in cells, neurotoxicity, Parkinson's disease, Amyotrophic lateral sclerosis (ALS), and ischemia/ reperfusion injury from stroke. All the articles were original research in experimental or pre-clinical stages using animal models or in vitro systems. The reported activities demonstrated the potential of P188 as a neuroprotective agent in improving CNS conditions such as neurodegeneration.
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http://dx.doi.org/10.2174/1570159X19666210528155801DOI Listing
May 2021

Tocotrienols Ameliorate Neurodegeneration and Motor Deficits in the 6-OHDA-Induced Rat Model of Parkinsonism: Behavioural and Immunohistochemistry Analysis.

Nutrients 2021 May 10;13(5). Epub 2021 May 10.

College of Medicine and Dentistry, James Cook University, Townsville, QLD 4811, Australia.

Parkinson's disease (PD) is a debilitating neurodegenerative disease, which progresses over time, causing pathological depigmentation of the substantia nigra (SN) in the midbrain due to loss of dopaminergic neurons. Emerging studies revealed the promising effects of some nutrient compounds in reducing the risk of PD. One such nutrient compound that possess neuroprotective effects and prevents neurodegeneration is tocotrienol (T3), a vitamin E family member. In the present study, a single dose intracisternal injection of 250 µg 6-hydroxydopamine (6-OHDA) was used to induce parkinsonism in male Sprague Dawley (SD) rats. Forty-eight hours post injection, the SD rats were orally supplemented with alpha (α)- and gamma (γ)-T3 for 28 days. The neuroprotective effects of α- and γ-T3 were evaluated using behavioural studies and immunohistochemistry (IHC). The findings from this study revealed that supplementation of α- and γ-T3 was able to ameliorate the motor deficits induced by 6-OHDA and improve the neuronal functions by reducing inflammation, reversing the neuronal degradation, and preventing further reduction of dopaminergic neurons in the SN and striatum (STR) fibre density.
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http://dx.doi.org/10.3390/nu13051583DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8150907PMC
May 2021

Influence of serum concentration in retinoic acid and phorbol ester induced differentiation of SH-SY5Y human neuroblastoma cell line.

Mol Biol Rep 2020 Nov 23;47(11):8775-8788. Epub 2020 Oct 23.

School of Postgraduate Studies, International Medical University, Kuala Lumpur, Malaysia.

Numerous protocols to establish dopaminergic phenotype in SH-SY5Y cells have been reported. In most of these protocols there are variations in concentration of serum used. In this paper, we compared the effects of high (10%), low (3%) and descending (2.5%/1%) serum concentration in differentiation medium containing different proportion of retinoic acid (RA) and 12-O-Tetradecanoylphorbol-13-acetate (TPA) or RA-only on the undifferentiated SH-SY5Y cells with regards to cell morphology, biochemical and gene expression alterations. Cells differentiated in culture medium containing low and descending serum concentrations showed increased number of neurite projections and reduced proliferation rates when compared to undifferentiated cells. The SH-SY5Y cells differentiated in culture medium containing 3% RA and low serum or descending (2.5%/1% RA/TPA) were found to be more susceptible to 6-hydroxydopamine (6-OHDA) induced cytotoxicity. Cells differentiated with RA/TPA or RA differentiated showed increased production of the α-synuclein (SNCA) neuroprotein and dopamine neurotransmitter compared to undifferentiated cells, regardless serum concentrations used. There was no significant difference in the expression of tyrosine hydroxylase (TH) gene between undifferentiated and differentiated SH-SY5Y cells. However, the expression of dopamine receptor D2 (DRD2) gene was markedly increased (p<0.05) in differentiated cells with 3% serum and RA only when compared to undifferentiated cells. In conclusion, to terminally differentiate SH-SY5Y cells to be used as a cell-based model to study Parkinson's disease (PD) to investigate molecular mechanisms and drug discovery, the optimal differentiation medium should contain 3% serum in RA-only.
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http://dx.doi.org/10.1007/s11033-020-05925-2DOI Listing
November 2020

Efficacy of Emu Oil Transfersomes for Local Transdermal Delivery of 4-OH Tamoxifen in the Treatment of Breast Cancer.

Pharmaceutics 2020 Aug 25;12(9). Epub 2020 Aug 25.

Jeffrey Cheah School of Medicine and Health Sciences, Monash University Malaysia, Jalan Lagoon Selatan, Bandar Sunway, Selangor Darul Ehsan 47500, Malaysia.

Oral tamoxifen used in the prevention and treatment of ductal carcinoma in situ (DCIS) (estrogen-positive) patients has limited acceptance, due to its adverse side effects. The efficacy of tamoxifen is related to its major metabolite, 4-hydroxytamoxifen. Local transdermal therapy of 4-hydroxytamoxifen to the breast might avert the toxicity of oral tamoxifen, while maintaining efficacy. We aim to study the skin irritancy, as well as to evaluate the efficacy of the developed transfersome formulations, with/without emu oil, using a syngeneic mouse model of breast cancer. We also quantified tamoxifen/4-hydroxytamoxifen concentrations in blood plasma and performed histopathology. The skin irritancy test showed that the pure emu oil and transfersome formulations with or without the emu oil did not cause skin irritancy in the animals studied. A sensitive and specific LC-MS/MS method for the quantification of tamoxifen and 4-hydroxytamoxifen was developed and validated. Studies on tumor volume and necrosis (histopathology) using the breast cancer mouse model showed that the 4-OHT transfersomal formulations, with and without emu oil, showed comparable efficacy with that of orally administered tamoxifen. However, the transfersomal formulations, with and without emu oil, resulted in significantly lower (10.24 ± 0.07 and 32.45 ± 0.48 ng/mL, respectively) plasma concentrations of 4-hydroxytamoxifen, compared to the oral tamoxifen (TAMX) group (634.42 ± 7.54 ng/mL). This study demonstrated the potential use of emu oil in a local transdermal formulation for the treatment of breast cancer and its reduced adverse effects.
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http://dx.doi.org/10.3390/pharmaceutics12090807DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7558379PMC
August 2020

A Novel Method to Identify Autoantibodies against Putative Target Proteins in Serum from beta-Thalassemia Major: A Pilot Study.

Biomedicines 2020 Apr 26;8(5). Epub 2020 Apr 26.

School of Medicine, International Medical University, Bukit Jalil, Kuala Lumpur 57000, Malaysia.

Abnormal immune reactivity in patients with beta-thalassemia (beta-thal) major can be associated with poor prognosis. Immunome protein-array analysis represents a powerful approach to identify novel biomarkers. The Sengenics Immunome Protein Array platform was used for high-throughput quantification of autoantibodies in 12 serum samples collected from nine beta-thal major patients and three non-thalassemia controls, which were run together with two pooled normal sera (Sengenics Internal QC samples). To obtain more accurate and reliable results, the evaluation of the biological relevance of the shortlisted biomarkers was analyzed using an Open Target Platform online database. Elevated autoantibodies directed against 23 autoantigens on the immunome array were identified and analyzed using a penetrance fold change-based bioinformatics method. Understanding the autoantibody profile of beta-thal major patients would help to further understand the pathogenesis of the disease. The identified autoantigens may serve as potential biomarkers for the prognosis of beta-thal major.
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http://dx.doi.org/10.3390/biomedicines8050097DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7277850PMC
April 2020

Advancing the Role of Gamma-Tocotrienol as Proteasomes Inhibitor: A Quantitative Proteomic Analysis of MDA-MB-231 Human Breast Cancer Cells.

Biomolecules 2019 12 21;10(1). Epub 2019 Dec 21.

Department of Molecular Medicine, Faculty of Medicine, University of Malaya, 50603 Kuala Lumpur, Malaysia.

Tocotrienol, an analogue of vitamin E has been known for its numerous health benefits and anti-cancer effects. Of the four isoforms of tocotrienols, gamma-tocotrienol (γT3) has been frequently reported for their superior anti-tumorigenic activity in both in vitro and in vivo studies, when compared to its counterparts. In this study, the effect of γT3 treatment in the cytoplasmic and nuclear fraction of MDA-MB-231 human breast cancer cells were assessed using the label-free quantitative proteomics analysis. The cytoplasmic proteome results revealed the ability of γT3 to inhibit a group of proteasome proteins such as PSMA, PSMB, PSMD, and PSME. The inhibition of proteasome proteins is known to induce apoptosis in cancer cells. As such, the findings from this study suggest γT3 as a potential proteasome inhibitor that can overcome deficiencies in growth-inhibitory or pro-apoptotic molecules in breast cancer cells. The nuclear proteome results revealed the involvement of important nuclear protein complexes which hardwire the anti-tumorigenesis mechanism in breast cancer following γT3 treatment. In conclusion, this study uncovered the advancing roles of γT3 as potential proteasomes inhibitor that can be used for the treatment of breast cancer.
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http://dx.doi.org/10.3390/biom10010019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7022772PMC
December 2019

Palm Tocotrienol-Adjuvanted Dendritic Cells Decrease Expression of the Gene in Murine Breast Cancer Cells and Tissues.

Vaccines (Basel) 2019 Nov 27;7(4). Epub 2019 Nov 27.

.Pathology Division, School of Medicine, International Medical University, Bukit Jalil, 57000 Kuala Lumpur, Malaysia.

The aim of this study was to evaluate the effectiveness of immunotherapy using dendritic cells (DC) pulsed with tumor lysate (a DC vaccine) in combination with daily supplementation of tocotrienol-rich fraction (TRF) to potentiate anti-tumor immune responses. We had previously reported that DC-vaccine immunotherapy together with TRF supplementation induced protective immunity to tumor challenge. Breast cancer was induced in female BALB/c mice. The mice were randomly assigned into the treatment groups. At autopsy, peripheral blood was collected in heparinized tube and the expression of cell surface molecules (CD40, CD80, CD83, and CD86) that are crucial for T-cell activation and survival were analyzed by flow cytometry. Tumor was excised from each animal and snap-frozen. Total RNA was extracted from each tumor tissue for microarray and gene expression analysis. Total protein was extracted from tumor tissue for protein expression studies using Western blotting. The results show that systemic administration of 1 mg TRF daily in combination with DC-vaccine immunotherapy (DC + TL + TRF) caused a marked reduction ( < 0.05) of tumor size and increased ( < 0.05) the survival rates of the tumor-inoculated mice. The expression of CD40, CD80, CD83, and CD86 were upregulated in peripheral blood from the DC + TL + TRF group compared to other groups. In addition, there was higher expression of FasL in tumor-excised mice from the DC + TL + TRF group compared to other groups. FasL plays an important role in maintaining immune privilege and is required for cytotoxic T-lymphocyte (CTL) activity. Microarray analysis identified several genes involved in the regulation of cancer. In this study, we focused on the special AT rich binding protein 1 gene, which was reported to have dual functions, one of which was to induce aggressive growth in breast cancer cells. Tumors from DC + TL + TRF mice showed lower ( < 0.05) expression of gene. Further study will be conducted to investigate the molecular functions of and the role of in 4T1 mammary cancer cells and DC. In conclusion, TRF supplementation can potentiate the effectiveness of DC-vaccine immunotherapy.
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http://dx.doi.org/10.3390/vaccines7040198DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6963955PMC
November 2019

Bioactive Compounds: Natural Defense Against Cancer?

Biomolecules 2019 11 21;9(12). Epub 2019 Nov 21.

Jeffrey Cheah School of Medicine and Health Sciences, Monash University Malaysia, Bandar Sunway, Selangor 47500, Malaysia.

Cancer is a devastating disease that has claimed many lives. Natural bioactive agents from plants are gaining wide attention for their anticancer activities. Several studies have found that natural plant-based bioactive compounds can enhance the efficacy of chemotherapy, and in some cases ameliorate some of the side-effects of drugs used as chemotherapeutic agents. In this paper, we have reviewed the literature on the anticancer effects of four plant-based bioactive compounds namely, curcumin, myricetin, geraniin and tocotrienols (T3) to provide an overview on some of the key findings that are related to this effect. The molecular mechanisms through which the active compounds may exert their anticancer properties in cell and animal-based studies also discussed.
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http://dx.doi.org/10.3390/biom9120758DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6995630PMC
November 2019

Investigation of the curative effects of palm vitamin E tocotrienols on autoimmune arthritis disease in vivo.

Sci Rep 2019 11 14;9(1):16793. Epub 2019 Nov 14.

Department of Biomedical Science, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia, Serdang, 43400, Malaysia.

The tocotrienol-rich fraction (TRF) from palm oil contains vitamin E, which possesses potent antioxidant and anti-inflammatory activities. Rheumatoid arthritis (RA) is a chronic joint inflammatory disease characterised by severe joint pain, cartilage destruction, and bone erosion owing to the effects of various pro-inflammatory mediators and cytokines. Here, we investigated the therapeutic effects of TRF in a rat model of collagen-induced arthritis (CIA). Arthritis was induced by a single intradermal injection of collagen type II in Dark Agouti (DA) rats. Rats were then treated with or without TRF by oral gavage from day 28 after the first collagen injection. Arthritic rats supplemented with TRF showed decreased articular index scores, ankle circumferences, paw volumes, and radiographic scores when compared with untreated rats. The untreated arthritic rats showed higher plasma C-reactive protein levels (p < 0.05) and production of pro-inflammatory cytokines than arthritic rats fed TRF. Moreover, there was a marked reduction in the severity of histopathological changes observed in arthritic rats treated with TRF compared with that in untreated arthritic rats. Overall, the results show that TRF had beneficial effects in this rat model of RA.
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http://dx.doi.org/10.1038/s41598-019-53424-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6856359PMC
November 2019

Medical students' orientation toward lifelong learning in an outcome-based curriculum and the lessons learnt.

Med Teach 2021 Jul 13;43(sup1):S6-S11. Epub 2019 Aug 13.

Faculty of Medicine and Health, International Medical University, Kuala Lumpur, Malaysia.

Background: Lifelong learning (LL) is an important outcome of medical training. The objective of this study is to measure the orientation of medical students toward LL and to determine the types of self-directed learning (SDL) activities that contribute toward LL skills.

Methods: The Jefferson Scale of Physician Lifelong Learning for medical student (JeffSPLL-MS) questionnaire was used. Factor analysis was performed, Cronbach's alpha and effect size were calculated. The types of learning activities that contribute to LL skills were identified.

Results: Three-factor structure emerged from the factor analysis and were identified as learning beliefs and motivation, skills in seeking information and attention to learning opportunities. A significant increase ( < .05; ES = 0.27) in orientation toward LL with academic progression was observed. Clinical students improved significantly in the domains of 'skills in seeking information' ( < .001; ES = 0.48) and 'attention to learning opportunities' ( < .001; ES = 0.55). Problem-based learning, flipped classroom, guided reading, projects and experiential learning activities are perceived to be effective for promoting LL.

Conclusions: Medical students' LL skills develop progressively from preclinical to clinical years. Self-directed learning activities are perceived to be effective in promoting LL skills.
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http://dx.doi.org/10.1080/0142159X.2019.1646894DOI Listing
July 2021

Tocotrienols Modulate Breast Cancer Secretomes and Affect Cancer-Signaling Pathways in MDA-MB-231 Cells: A Label-Free Quantitative Proteomic Analysis.

Nutr Cancer 2019 14;71(8):1263-1271. Epub 2019 May 14.

Department of Molecular Medicine, Faculty of Medicine, University of Malaya , Kuala Lumpur , Malaysia.

Tocotrienols (T3), a family of vitamin E, are reported to possess potent anti-cancer effects but the molecular mechanisms behind these effects still remain unclear. The aim of this study was to investigate how T3 exert anti-cancer effects on MDA-MB-231 human breast cancer cells. The MDA-MB-231 cells were chosen for this study as they are triple-negative and highly metastatic cells, which form aggressive tumors in experimental models. The MDA-MB-231 cells were treated with varying concentrations (0-20 µg mL) of gamma (γ) or delta (δ) T3 and the secretome profiles of these cells treated with half maximal inhibitory concentration (IC) of γT3 (5.8 µg mL) or δT3 (4.0 µg mL) were determined using label-free quantitative proteomic strategy. A total of 103, 174 and 141 proteins were identified with ProteinLynx Global Server (PLGS) score of more than 200 and above 25% sequence coverage in the untreated control and T3-treated cell culture supernatant respectively. A total of 18 proteins were dysregulated between untreated control and T3 (δT3 or γT3) treated conditions. The results showed that T3 treatment downregulated the exogenous Cathepsin D and Serpine1 proteins but upregulated Profilin-1 protein, which play a key role in breast cancer in the MDA-MB-231 cells. These findings strongly suggest that T3 may induce differential expression of secreted proteins involved in the cytoskeletal regulation of RHO GTPase signaling pathway.
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http://dx.doi.org/10.1080/01635581.2019.1607407DOI Listing
July 2020

Protective Mechanisms of Flavonoids in Parkinson's Disease.

Oxid Med Cell Longev 2015 20;2015:314560. Epub 2015 Oct 20.

Discipline of Biomedicine, College of Public Health, Medical and Veterinary Sciences, James Cook University, Townsville, QLD 4811, Australia.

Parkinson's disease is a chronic, debilitating neurodegenerative movement disorder characterized by progressive degeneration of dopaminergic neurons in the substantia nigra pars compacta region in human midbrain. To date, oxidative stress is the well accepted concept in the etiology and progression of Parkinson's disease. Hence, the therapeutic agent is targeted against suppressing and alleviating the oxidative stress-induced cellular damage. Within the past decades, an explosion of research discoveries has reported on the protective mechanisms of flavonoids, which are plant-based polyphenols, in the treatment of neurodegenerative disease using both in vitro and in vivo models. In this paper, we have reviewed the literature on the neuroprotective mechanisms of flavonoids in protecting the dopaminergic neurons hence reducing the symptoms of this movement disorder. The mechanism reviewed includes effect of flavonoids in activation of endogenous antioxidant enzymes, suppressing the lipid peroxidation, inhibition of inflammatory mediators, flavonoids as a mitochondrial target therapy, and modulation of gene expression in neuronal cells.
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http://dx.doi.org/10.1155/2015/314560DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4630416PMC
July 2016

Modulatory effects of mesenchymal stem cells on leucocytes and leukemic cells: A double-edged sword?

Blood Cells Mol Dis 2015 Dec 1;55(4):351-7. Epub 2015 Aug 1.

Division of Pathology, Faculty of Medicine and Health Sciences, International Medical University, Malaysia. Electronic address:

Mesenchymal stem cells (MSCs) have drawn much attention amongst stem cell researchers in the past few decades. The ability of the MSC to differentiate into cells of mesodermal and non-mesodermal origins has made them an attractive approach for cell-based therapy and regenerative medicine. The MSCs have immunosuppressive activities that may have considerable therapeutic values in autoimmune diseases. However, despite the many beneficial effects reported, there is a growing body of evidence, which suggests that MSCs could be a culprit of enhanced tumour growth, metastasis and drug resistance in leukaemia, via some modulatory effects. Many controversies regarding the interactions between MSCs and leukaemia still exist. Furthermore, the role of MSCs in leukemogenesis and its progression remain largely unknown. Hence it is important to understand how the MSCs modulate leukaemia before these cells could be safely used in the treatment of leukaemia patients.
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http://dx.doi.org/10.1016/j.bcmd.2015.07.017DOI Listing
December 2015

toxicity evaluation of a standardized extract of leaf.

Toxicol Rep 2014 21;1:718-725. Epub 2014 Sep 21.

School of Medicine and Health Sciences, Monash University, Sunway Campus, 46150 Bandar Sunway, Malaysia.

In this study, the acute and subchronic toxicity effect of the leaf extract (SA) was evaluated. For the acute toxicity study, a single dose of 2000 mg/kg of the SA was given by oral-gavage to male Sprague-Dawley (SD) rats. The rats were observed for mortality and toxicity signs for 14 days. In the subchronic toxicity study the SA was administered orally at doses of 50, 100, and 200 mg/kg per day for 28 days to male SD rats. The animals were sacrificed at the end of the experiment. The parameters measured including food and water intake, body weight, absolute and relative organ weight, blood biochemical tests and histopathology observation. In both the acute and subchronic toxicity studies, SA did not show any visible signs of toxicity. There were also no significant differences between the control and SA treated rats in terms of their food and water intake, body weight, absolute and relative organ weight, biochemical parameters or gross and microscopic appearance of the organs. There were no acute or subchronic toxicity observed and our results indicate that this extract could be devoid of any toxic risk. This is the first study reported the safety and toxicity of SA.
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http://dx.doi.org/10.1016/j.toxrep.2014.09.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5598474PMC
September 2014

Tocotrienol-adjuvanted dendritic cells inhibit tumor growth and metastasis: a murine model of breast cancer.

PLoS One 2013 19;8(9):e74753. Epub 2013 Sep 19.

Pathology Division, Faculty of Medicine and Health, International Medical University, Bukit Jalil, Kuala Lumpur, Malaysia ; Nutrition Unit, Malaysian Palm Oil Board, Bandar Baru Bangi, Selangor, Malaysia.

Tocotrienol-rich fraction (TRF) from palm oil is reported to possess anti-cancer and immune-enhancing effects. In this study, TRF supplementation was used as an adjuvant to enhance the anti-cancer effects of dendritic cells (DC)-based cancer vaccine in a syngeneic mouse model of breast cancer. Female BALB/c mice were inoculated with 4T1 cells in mammary pad to induce tumor. When the tumor was palpable, the mice in the experimental groups were injected subcutaneously with DC-pulsed with tumor lysate (TL) from 4T1 cells (DC+TL) once a week for three weeks and fed daily with 1 mg TRF or vehicle. Control mice received unpulsed DC and were fed with vehicle. The combined therapy of using DC+TL injections and TRF supplementation (DC+TL+TRF) inhibited (p<0.05) tumor growth and metastasis. Splenocytes from the DC+TL+TRF group cultured with mitomycin-C (MMC)-treated 4T1 cells produced higher (p<0.05) levels of IFN-γ and IL-12. The cytotoxic T-lymphocyte (CTL) assay also showed enhanced tumor-specific killing (p<0.05) by CD8(+) T-lymphocytes isolated from mice in the DC+TL+TRF group. This study shows that TRF has the potential to be used as an adjuvant to enhance effectiveness of DC-based vaccines.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0074753PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3777960PMC
June 2014

Supplementation with natural forms of vitamin E augments antigen-specific TH1-type immune response to tetanus toxoid.

Biomed Res Int 2013 7;2013:782067. Epub 2013 Jul 7.

Pathology Division, Faculty of Medicine and Health, International Medical University, Bukit Jalil, Kuala Lumpur, Wilayah Persekutuan, Malaysia.

This study compared the ability of three forms of vitamin E [tocotrienol-rich fraction (TRF), alpha-tocopherol (α-T), and delta-tocotrienol (δ-T3)] to enhance immune response to tetanus toxoid (TT) immunisation in a mouse model. Twenty BALB/c mice were divided into four groups of five mice each. The mice were fed with the different forms of vitamin E (1 mg) or vehicle daily for two weeks before they were given the TT vaccine [4 Lf] intramuscularly (i.m.). Booster vaccinations were given on days 28 and 42. Serum was collected (days 0, 28, and 56) to quantify anti-TT levels. At autopsy, splenocytes harvested were cultured with TT or mitogens. The production of anti-TT antibodies was augmented (P < 0.05) in mice that were fed with δ-T3 or TRF compared to controls. The production of IFN-γ and IL-4 by splenocytes from the vitamin E treated mice was significantly (P < 0.05) higher than that from controls. The IFN-γ production was the highest in animals supplemented with δ-T3 followed by TRF and finally α-T. Production of TNF-α was suppressed in the vitamin E treated group compared to vehicle-supplemented controls. Supplementation with δ-T3 or TRF can enhance immune response to TT immunisation and production of cytokines that promote cell-mediated (TH1) immune response.
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http://dx.doi.org/10.1155/2013/782067DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3722853PMC
March 2014

Effect of Spirulina (Arthrospira) supplementation on the immune response to tetanus toxoid vaccination in a mouse model.

J Diet Suppl 2013 Sep 9;10(3):229-40. Epub 2013 Aug 9.

School of Postgraduate Studies and Research, Faculty of Medicine and Health, International Medical University, Kuala Lumpur, Malaysia.

The aim of this study was to investigate whether Spirulina (Arthrospira) supplementation could enhance the immune response to tetanus toxoid (TT) vaccine in a mouse model. Vaccination of TT was performed on day 7 and 21 in mice fed daily with Spirulina (50 and 150 mg/kg body weight). Both Spirulina supplementation and TT vaccination did not significantly affect body weight gain of the mice. Supplementation of Spirulina significantly enhanced IgG level (p = .01) after the first but not after the second TT vaccination. The anti-TT IgG levels of the groups that received low dose and high dose of Spirulina were not significantly different. Spirulina supplementation did not show significant effects on in vitro splenocyte proliferation and cytokine (IFN-γ and IL-4) production induced by Con A and TT. This study showed that Spirulina supplementation could enhance primary immune response in terms of antibody production, but not secondary immune response following TT vaccination in a mouse model.
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http://dx.doi.org/10.3109/19390211.2013.822452DOI Listing
September 2013

Single nucleotide polymorphism in the promoter of the human interleukin-13 gene is associated with asthma in Malaysian adults.

Biomed Res Int 2013 25;2013:981012. Epub 2013 Jun 25.

Pathology Division, Faculty of Medicine and Health, International Medical University, 126, Jalan Jalil Perkasa 19, Bukit Jalil, 57000 Kuala Lumpur, Malaysia.

Asthma susceptibility genes are mapped to a region on human chromosome 5q31-q33, which contains a cluster of proinflammatory cytokine genes such as interleukin-13 (IL-13), which is associated with asthma. This study investigated the allele frequencies of two single nucleotide polymorphisms (SNPs) (-1111C>T and 4257C>A) in the IL-13 gene between asthmatics and healthy volunteers as well as the relationship between these SNPs and IL-13 production. DNA extracted from buffy coat of asthmatic and control subjects was genotyped using the PCR-RFLP method. Amount of IL-13 produced by mitogen-stimulated peripheral blood leucocytes PBLs (PBLs) was determined by ELISA. The frequencies of the -1111C and 4257G wild-type alleles were 0.52 and 0.55 in asthmatics and were 0.67 and 0.56 in controls. A significant (P < 0.05) association was found between genotype and allele frequencies of SNP at position -1111C>T between asthmatic and control groups (OR, 1.810; 95% CI = 1.184 to 2.767; P < 0.05). The mitogen-stimulated PBLs from asthmatics produced higher amounts of IL-13 production (P < 0.001). The 4257GA heterozygous and 4257AA homozygous mutant alleles were associated with higher IL-13 production in asthmatics (P < 0.05). Our results show that the -1111T mutant allele are associated with asthma and the 4257A mutant alleles are associated with elevated IL-13 production.
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http://dx.doi.org/10.1155/2013/981012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3707285PMC
February 2014

Comparing the ability of freshly generated and cryopreserved dendritic cell vaccines to inhibit growth of breast cancer in a mouse model.

Biores Open Access 2012 Oct;1(5):239-46

Pathology Division, Faculty of Medicine and Health, International Medical University , Kuala Lumpur, Malaysia .

Repetitive vaccinations with dendritic cell (DC)-based vaccines over long periods of time can break pre-existing tolerance to tumors and achieve clinically relevant immune response. This requires a large number of DCs to be generated under good manufacturing protocol, which is time- and cost intensive. Thus, producing a large numbers of DCs at one time point and cryopreserving these cells in ready-for-use aliquots for clinical application may overcome this constraint. This could also reduce batch-to-batch variations. In this study, we generated DCs from bone marrow obtained from BALB/c mice. Some of the generated DCs were cryopreserved before conducting various tests. There were no significant differences in the morphology and phenotype between cryopreserved and freshly generated DCs. Both types of DCs pulsed with tumor lysate (TL) from 4T1 murine mammary cancer cells (DC+TL) possessed a similar capacity to stimulate the proliferation of T-cells. In addition, cryopreserved and fresh DC pulsed with TL showed similar tumor growth inhibition patterns. Both DCs induced initial retardation of tumor growth (p<0.05) and prolonged the survival (p<0.05) of tumor-bearing mice treated with DC+TL as compared with nontreated control mice. Cryopreserved DCs shared similar therapeutic efficacy to fresh DCs, and this finding lends supports the routine use of cryopreserved DCs in future clinical trials.
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http://dx.doi.org/10.1089/biores.2012.0229DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3559207PMC
October 2012

Amelioration of collagen-induced arthritis in female dark agouti rats by glucosamine treatment.

ISRN Pharmacol 2013 14;2013:562905. Epub 2013 Feb 14.

School of Veterinary and Biomedical Sciences, James Cook University, Townsville, QLD 4811, Australia.

The present study assessed the therapeutic efficacy of glucosamine hydrochloride against collagen-induced arthritis in female Dark Agouti rats (DA). Arthritis was induced by intradermaly injecting a collagen and complete Freund's adjuvant suspension at multiple sites in the rat at a dose of 4 mg/kg of body weight and thereafter followed by two more boosters of the same dose, after the 1st week and 2nd week of primary immunization. After 21 days from the day of primary immunization, the arthritic group rats were given oral supplementation of glucosamine hydrochloride at a dose of 300 mg/kg of body weight until day 45. The arthritic group treated with glucosamine hydrochloride from day 21 to day 45 showed significant reduction in arthritic histopathological changes of the joints, reduction in paw thickness and also a significant decrease in C-reactive protein and TNF-alpha in the serum. Treatment with 300 mg/kg of glucosamine hydrochloride was able to reverse the arthritic changes, hence suggesting that glucosamine has a therapeutic effect against collagen-induced arthritis.
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http://dx.doi.org/10.1155/2013/562905DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3586518PMC
March 2013

Colostrum supplementation protects against exercise-induced oxidative stress in skeletal muscle in mice.

BMC Res Notes 2012 Nov 22;5:649. Epub 2012 Nov 22.

Faculty of Sports Science & Recreation, University Technology Mara, Shah Alam 40450, Kuala Lumpur, Malaysia.

Background: This study examined the effects of bovine colostrum on exercise -induced modulation of antioxidant parameters in skeletal muscle in mice. Adult male BALB/c mice were randomly divided into four groups (control, colostrum alone, exercise and exercise with colostrum) and each group had three subgroups (day 0, 21 and 42). Colostrum groups of mice were given a daily oral supplement of 50 mg/kg body weight of bovine colostrum and the exercise group of mice were made to exercise on the treadmill for 30 minutes per day. Total antioxidants, lipid hydroperoxides, xanthine oxidase and super oxide dismutase level was assayed from the homogenate of hind limb skeletal muscle.

Results: Exercise-induced a significant oxidative stress in skeletal muscles as evidenced by the elevated lipid hydroperoxides and xanthine oxidase levels. There was a significant decrease in skeletal muscle total antioxidants and superoxide dismutase levels. Daily colostrum supplement significantly reduced the lipid hydroperoxides and xanthine oxidase enzyme level and increased the total antioxidant levels in the leg muscle.

Conclusion: Thus, the findings of this study showed that daily bovine colostrum supplementation was beneficial to skeletal muscle to reduce the oxidant-induced damage during muscular exercise.
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http://dx.doi.org/10.1186/1756-0500-5-649DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3571863PMC
November 2012

An effective ostrich oil bleaching technique using peroxide value as an indicator.

Molecules 2011 Jul 5;16(7):5709-19. Epub 2011 Jul 5.

School of Medicine and Health Sciences, Monash University, Sunway Campus, 46150, Malaysia.

Ostrich oil has been used extensively in the cosmetic and pharmaceutical industries. However, rancidity causes undesirable chemical changes in flavour, colour, odour and nutritional value. Bleaching is an important process in refining ostrich oil. Bleaching refers to the removal of certain minor constituents (colour pigments, free fatty acid, peroxides, odour and non-fatty materials) from crude fats and oils to yield purified glycerides. There is a need to optimize the bleaching process of crude ostrich oil prior to its use for therapeutic purposes. The objective of our study was to establish an effective method to bleach ostrich oil using peroxide value as an indicator of refinement. In our study, we showed that natural earth clay was better than bentonite and acid-activated clay to bleach ostrich oil. It was also found that 1 hour incubation at a 150 °C was suitable to lower peroxide value by 90%. In addition, the nitrogen trap technique in the bleaching process was as effective as the continuous nitrogen flow technique and as such would be the recommended technique due to its cost effectiveness.
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http://dx.doi.org/10.3390/molecules16075709DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6264162PMC
July 2011

Tocotrienol-treated MCF-7 human breast cancer cells show down-regulation of API5 and up-regulation of MIG6 genes.

Cancer Genomics Proteomics 2011 Jan-Feb;8(1):19-31

Department of Human Biology, Faculty of Medicine and Health, International Medical University, Bukit Jalil, 57000 Kuala Lumpur, Malaysia.

Background: Tocotrienols belong to the vitamin E family and have multiple anticancer effects, such as antiproliferative, antioxidant, pro-apoptosis and antimetastatic. This study aimed to identify the genes that are regulated in human breast cancer cells following exposure to various isomers of vitamin E as these may be potential targets for the treatment of breast cancer.

Materials And Methods: Gene expression profiling was performed with MCF-7 cells at inhibitory conditions of IC(50) using Illumina's Sentrix Array Human-6 BeadChips. The expression levels of selected differentially expressed genes were verified by quantitative real-time-PCR (qRT-PCR).

Results: The treatment with tocotrienol-rich palm oil fraction (TRF), α-tocopherol and isomers of tocotrienols (α, γ, and δ) altered the expression of several genes that code for proteins involved in the regulation of immune response, tumour growth and metastatic suppression, apoptotic signalling, transcription, protein biosynthesis regulation and many others.

Conclusion: Treatment of human MCF-7 cells with tocotrienol isomers causes the down-regulation of the API5 gene and up-regulation of the MIG6 gene and the differential expression of other genes reported to play a key role in breast cancer biology.
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April 2011

Protective effect of aqueous extract from Spirulina platensis against cell death induced by free radicals.

BMC Complement Altern Med 2010 Sep 21;10:53. Epub 2010 Sep 21.

International Medical University, No, 126 Jalan 19/155B, Bukit Jalil, 57000 Kuala Lumpur, Malaysia.

Background: Spirulina is a commercial alga well known to contain various antioxidants, especially phycocyanin. Apart from being sold as a nutraceutical, Spirulina is incorporated as a functional ingredient in food products and beverages. Most of the previous reports on antioxidant activity of Spirulina were based on chemical rather than cell-based assays. The primary objective of this study was to assess the antioxidant activity of aqueous extract from Spirulina based on its protective effect against cell death induced by free radicals.

Methods: The antioxidant activity of the cold water extract from food-grade Spirulina platensis was assessed using both chemical and cell-based assays. In the cell-based assay, mouse fibroblast cells (3T3) cells were incubated for 1 h in medium containing aqueous extract of Spirulina or vitamin C (positive control) at 25, 125 and 250 μg/mL before the addition of 50 μM 1,1-diphenyl-2-picrylhydrazyl (DPPH) or 3-ethylbenzothiazoline-6-sulfonic acid (ABTS). The cells were incubated for another 24 h before being assessed for cell death due to apoptosis using the Cell Death Detection ELISA Kit. Spectrophotometric assays based on DPPH and ABTS were also used to assess the antioxidant activity of the extract compared to vitamin C and vitamin E (positive controls).

Results: Spirulina extract did not cause cytotoxic effect on 3T3 cells within the range of concentrations tested (0 - 250 μg/mL). The extract reduced significantly (p < 0.05) apoptotic cell death due to DPPH and ABTS by 4 to 5-fold although the activity was less than vitamin C. Based on the DPPH assay, the radical scavenging activity of the extract was higher than phycocyanin and was at least 50% of vitamin C and vitamin E. Based on the ABTS assay, the antioxidant activity of the extract at 50 μmug/mL was as good as vitamin C and vitamin E.

Conclusions: The results showed that aqueous extract of Spirulina has a protective effect against apoptotic cell death due to free radicals. The potential application of incorporating Spirulina into food products and beverages to enhance their antioxidant capacity is worth exploring.
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http://dx.doi.org/10.1186/1472-6882-10-53DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2954939PMC
September 2010

Is there a genetic variation association in the IL-10 and TNF alpha promoter gene with gestational diabetes mellitus?

Hereditas 2010 Apr;147(2):94-102

Department of Obstetrics and Gynecology, Faculty of Medicine and Health, International Medical University (IMU), Bukit Jalil, Kuala Lumpur, Malaysia.

Gestational diabetes mellitus (GDM), defined as carbohydrate intolerance diagnosed for the first time during pregnancy, affects both maternal and fetal health. Possession of a specific genetic polymorphism can be a predisposing factor for susceptibility to some diseases. The aim of this study was to investigate the association between single nucleotide polymorphisms (SNP) in the promoter gene of interleukin-10 (IL-10) as well as tumor necrosis factor-alpha (TNF alpha) with the development of GDM. Two hundred and twelve consecutive series of eligible normal pregnant women (controls) and gestational diabetes mellitus women were selected based on the study's inclusion and exclusion criteria. DNA was extracted from blood and genotyped for IL-10 at three positions and TNF alpha for gene polymorphism using the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. Plasma levels of IL-10 and TNF alpha at different gestational periods as well as postpartum were quantified using enzyme linked immunosorbent assay (ELISA). The results of the study showed that the difference in the frequency of SNP at position -597 in the promoter of the human IL-10 gene between the control and GDM groups was statistically significant (p < 0.05). In contrast, there was no significant difference in the frequency of SNP at the other two sites in the promoter region of the human IL-10 gene (-824 and -1082) as well as position -308 in the promoter of the human TNF-alpha (p > 0.05). In addition, there was no significant difference between the two groups in terms of plasma levels of IL-10 as well as TNF alpha in different stages of pregnancy. SNP at position -597 was significantly associated with the development of GDM and shows potential for use as a predictive marker for GDM.
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http://dx.doi.org/10.1111/j.1601-5223.2009.02134.xDOI Listing
April 2010

Assessment of antioxidant capacity and cytotoxicity of selected Malaysian plants.

Molecules 2010 Mar 25;15(4):2139-51. Epub 2010 Mar 25.

Department of Physiology, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia.

Thirteen Malaysian plants; Artocarpus champeden, Azadirachta indica, Fragaria x ananassa, Garcinia mangostana, Lawsonia inermis, Mangifera indica, Nephelium lappaceum, Nephelium mutobile, Peltophorum pterocarpum, Psidium guajava and Syzygium aqueum, selected for their use in traditional medicine, were subjected to a variety of assays. Antioxidant capability, total phenolic content, elemental composition, as well as it cytotoxity to several cell lines of the aqueous and ethanolic extracts from different parts of these selected Malaysian plants were determined. In general, the ethanolic extracts were better free radical scavengers than the aqueous extracts and some of the tested extracts were even more potent than a commercial grape seed preparation. Similar results were seen in the lipid peroxidation inhibition studies. Our findings also showed a strong correlation of antioxidant activity with the total phenolic content. These extracts when tested for its heavy metals content, were found to be below permissible value for nutraceutical application. In addition, most of the extracts were found not cytotoxic to 3T3 and 4T1 cells at concentrations as high as 100 microg/mL. We conclude that although traditionally these plants are used in the aqueous form, its commercial preparation could be achieved using ethanol since a high total phenolic content and antioxidant activity is associated with this method of preparation.
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http://dx.doi.org/10.3390/molecules15042139DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6257288PMC
March 2010

Association between polymorphisms in human tumor necrosis factor-alpha (--308) and -beta (252) genes and development of gestational diabetes mellitus.

Diabetes Res Clin Pract 2010 May 26;88(2):139-45. Epub 2010 Feb 26.

Department of Obstetrics and Gynaecology, Faculty of Medicine and Health, International Medical University, 126, Jalan 19/155B, Bukit Jalil, 57000 Kuala Lumpur, Malaysia.

Objective: The aim of this study is to investigate if an association exists between single nucleotide polymorphism (SNP) in the tumor necrosis factor-alpha (TNF-alpha) and TNF-beta genes.

Methods: The DNA was extracted and SNP in the human TNF-alpha and TNF-beta genes at positions -308 (G/A) and 252 (A/G), respectively, was analyzed using the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. Plasma levels of TNF-alpha in different stages of pregnancy were quantified using enzyme linked immunosorbent assay (ELISA).

Results: There was no significant difference in genotype and allele frequency of SNP at position -308 (G/A) in the promoter region of the human TNF-alpha gene as well as the SNP at position 252 (A/G) in the human TNF-beta gene between the GDM and control subjects. Using the logistic regression model, it was found that the SNP in the TNF-alpha as well as TNF-beta were not associated with development of GDM. In addition, the TNF-alpha levels in the plasma of GDM and control mothers were not significantly different.

Conclusions: In the population studied, the SNP in position -308 (G/A) of the human TNF-alpha or in position 252 (A/G) of the human TNF-beta gene is not an independent risk factor or a predictor for GDM.
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http://dx.doi.org/10.1016/j.diabres.2010.01.028DOI Listing
May 2010
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