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Fulltext Qualitative flow cytometric analysis of Malaysian myelodysplastic syndromes (MDS) patients

Mohadese Hashem B, Ramasamy R, Sabariah MN, Seman Z

Med J Malaysia, 2012 Feb;67(1):77-80.
PMID: 22582553 MyJurnal

Myelodysplastic syndromes (MDS) are a group of haematological malignancies categorized by ineffective hematopoiesis that result in dysplasia. Although morphological diagnosis is a traditional and standard technique that is used for the diagnosis of MDS, the heterogeneous blood and bone marrow characteristics of MDS patients can potentially obscure the right diagnosis. Thus, we have utilized flow cytometric immunophenotyping as a supportive mechanism to obtain a more accurate and faster method for detection of abnormal markers in MDS. Flow cytometry was used for analyzing bone marrow samples from newly diagnosed MDS patients to investigate the abnormal antigen expression patterns in granulocytic, monocytic, erythroid, lymphoid lineages and myeloid precursors. The results were compared with those obtained from cases that had Idiopathic Thrombocytopenic Purpura (ITP) as a control. The most common abnormality found in the granulocytic lineage was the decrease of CD10. Low expressions of CD13 were the most frequent abnormality in the monocytic lineage. The erythroid lineage was found to have low expression of CD235A+/CD71+, reduce of CD71 and decreased CD235a. In conclusion, this method is useful for confirming cases in which it is difficult to make a diagnosis by morphology.
Isolation and characterisation of mesenchymal stem cells derived from human placenta tissue

Vellasamy S, Sandrasaigaran P, Vidyadaran S, George E, Ramasamy R

World J Stem Cells, 2012 Jun 26;4(6):53-61.
PMID: 22993662

To explore the feasibility of placenta tissue as a reliable and efficient source for generating mesenchymal stem cells (MSC).
Inhibitory effects of palm α-, γ- and δ-tocotrienol on lipopolysaccharide-induced nitric oxide production in BV2 microglia

Tan SW, Ramasamy R, Abdullah M, Vidyadaran S

Cell Immunol, 2011;271(2):205-9.
PMID: 21839427 DOI: 10.1016/j.cellimm.2011.07.012

Anti-inflammatory actions of the vitamin E fragment tocotrienol have not been described for microglia. Here, we screened palm α-, γ- and δ-tocotrienol isoforms and Tocomin® 50% (contains spectrum of tocotrienols and tocopherols) for their ability to limit nitric oxide (NO) production by BV2 microglia. Microglia were treated with varying doses of tocotrienols for 24h and stimulated with 1 μg/ml lipopolysaccharide (LPS). All tocotrienol isoforms reduced NO release by LPS-stimulated microglia, with 50 μM being the most potent tocotrienol dose. Of the isoforms tested, δ-tocotrienol lowered NO levels the most, reducing NO by approximately 50% at 48 h post-LPS treatment (p
Immunomodulatory activity of polyphenols derived from Cassia auriculata flowers in aged rats

John CM, Sandrasaigaran P, Tong CK, Adam A, Ramasamy R

Cell Immunol, 2011;271(2):474-9.
PMID: 21924708 DOI: 10.1016/j.cellimm.2011.08.017

The immunomodulatory activity of Cassia auriculata (CA)-derived polyphenols was tested on aged rats. Rats (24-26 months old) were given CA polyphenols supplementation at doses of 25, 50, and 100 mg/kg for 28 days. Flow cytometry analysis of CA polyphenols-treated aged rats showed increased T and B cells percentage along with enhanced proliferation of splenocytes in both resting and LPS-stimulated cells. Increased percentage of pan T cells is further supported by an elevation of CD4+, CD8+, and CD4+CD25+ regulatory cells. In terms of innate immune cell activity, CA polyphenol supplementation reduced the oxidative burst activity of neutrophils in response to PMA and Escherichia coli activation. Our results collectively show that polyphenols derived from CA boost T cell immunity by increasing the number of T cells and its sensitivity towards stimulants and decreasing ROS production by neutrophils that could potentially harm multiple biological systems in aged individuals.
Elevated neutrophil respiratory burst activity in essential hypertensive patients

Ramasamy R, Maqbool M, Mohamed AL, Noah RM

Cell Immunol, 2010;263(2):230-4.
PMID: 20471005 DOI: 10.1016/j.cellimm.2010.04.004

Neutrophils play a significant role in maintaining the integrity of innate immunity via their potent respiratory burst activity. However, the uncontrolled activation of respiratory burst in neutrophils also attributes to chronic diseases such as primary hypertension and atherosclerosis. In our study, we have investigated the activation of respiratory burst function of neutrophils harvested from essential hypertensive patients. In the presence of stimuli PMA and opsonized zymosan (OZ), hypertensive patients' neutrophils secrete significantly higher amount of superoxide anions compared to normotensive control. Although the magnitude of activation varies between both groups, yet the kinetics of activation is similar. When normotensive control's neutrophils were pre-treated with hypertensive serum, the cells failed to migrate toward fMLP which indicates the impairment of the migration property. In conclusion, the respiratory burst activity of neutrophils is affected by hypertension and their elevated superoxide anions production could be an aggravating factor in hypertension-related complication.
The immunosuppressive effects of human bone marrow-derived mesenchymal stem cells target T cell proliferation but not its effector function

Ramasamy R, Tong CK, Seow HF, Vidyadaran S, Dazzi F

Cell Immunol, 2008 Feb;251(2):131-6.
PMID: 18502411 DOI: 10.1016/j.cellimm.2008.04.009

Mesenchymal stem cells (MSC) are non-haematopoietic stem cells that are capable of differentiating into tissues of mesodermal origin. MSC play an important role in supporting the development of fetal and adult haematopoiesis. More recently, MSC have also been found to exhibit inhibitory effect on T cell responses. However, there is little information on the mechanism of this immunosuppression and our study addresses this issue by targeting T cell functions at various level of immune responses. We have generated MSC from human adult bone marrow (BM) and investigated their immunoregulatory function at different phases of T cell responses. MSC showed the ability to inhibit mitogen (CD3/CD28 microbeads)-activated T cell proliferation in a dose-dependent manner. In order to evaluate the specificity of this immunosuppression, the proliferation of CD4(+) and CD8(+) cells were measured. MSC equally inhibit CD4(+) and CD8(+) subpopulations of T cells in response to PHA stimulation. However, the antiproliferative effect of MSC is not due to the inhibition of T cell activation. The expression of early activation markers of T cells, namely CD25 and CD69 were not significantly altered by MSC at 24, 48 and 72h. Furthermore, the immunosuppressive effect of MSC mainly targets T cell proliferation rather than their effector function since cytotoxicity of T cells is not affected. This work demonstrates that the immunosuppressive effect of MSC is exclusively a consequence of an anti-proliferative activity, which targets T cells of different subpopulations. For this reason, they have the potential to be exploited in the control of unwanted immune responses such as graft versus host disease (GVHD) and autoimmunity.
Fulltext Immunophenotype and differentiation capacity of bone marrow-derived mesenchymal stem cells from CBA/Ca, ICR and Balb/c mice

Ooi YY, Rahmat Z, Jose S, Ramasamy R, Vidyadaran S

World J Stem Cells, 2013 Jan 26;5(1):34-42.
PMID: 23362438 DOI: 10.4252/wjsc.v5.i1.34

To assess the capacity to isolate and expand mesenchymal stem cells (MSC) from bone marrow of CBA/Ca, ICR and Balb/c mice.
The Bahasa Melayu version of Cornell Musculoskeletal Discomfort Questionnaire (CMDQ): Reliability and validity study in Malaysia

Shariat A, Tamrin SB, Arumugam M, Ramasamy R

Work, 2016 Mar 09;54(1):171-8.
PMID: 26967034 DOI: 10.3233/WOR-162269

BACKGROUND: The Cornell Musculoskeletal Discomfort Questionnaire (CMDQ) was developed to assess the level of musculoskeletal discomfort among office workers related to their ergonomic situation.
OBJECTIVE: The primary objective of this initial study is to analyze the validity and dependability of the Malay translation of the Cornell Musculoskeletal Discomfort Questionnaire.
METHODS: The questionnaire was self-administered two times, with an interval of two weeks in order to evaluate the accuracy of the original findings with a retest. The study involved 115 participants.
RESULTS: The range of Cronbach Alpha coefficient showed a considerable consistency of the items for each sub-scale (Cronbach's a > 0.95). The range of Kappa coefficients was between (ICC = 0.690-0.949, p
Human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSC) inhibit the proliferation of K562 (human erythromyeloblastoid leukaemic cell line)

Fonseka M, Ramasamy R, Tan BC, Seow HF

Cell Biol Int, 2012 Sep;36(9):793-801.
PMID: 22335239 DOI: 10.1042/CBI20110595

hUCB-MSC (human umbilical cord blood-derived mesenchymal stem cells) offer an attractive alternative to bone marrow-derived MSC for cell-based therapy by being less invasive a source of biological material. We have evaluated the effect of hUCB-MSC on the proliferation of K562 (an erythromyeloblastoid cell line) and the cytokine secretion pattern of hUCB-MSC. Co-culturing of hUCB-MSC and K562 resulted in inhibition of proliferation of K562 in a dose-dependent manner. However, the anti-proliferative effect was reduced in transwells, suggesting the importance of direct cell-to-cell contact. hUCB-MSC inhibited proliferation of K562, arresting them in the G0 /G1 phase. NO (nitric oxide) was not involved in the hUCB-MSC-mediated tumour suppression. The presence of IL-6 (interleukin 6) and IL-8 were obvious in the hUCB-MSC conditioned media, but no significant increase was found in 29 other cytokines. Th1 cytokines, IFNα (interferon α), Th2 cytokine IL-4 and Th17 cytokine, IL-17 were not secreted by hUCB-MSC. There was an increase in the number of hUCB-MSC expressing the latent membrane-bound form of TGFβ1 co-cultured with K562. The anti-proliferative effect of hUCB-MSC was due to arrest of the growth of K562 in the G0 /G1 phase. The mechanisms underlying increased IL-6 and IL-8 secretion and LAP (latency-associated peptide; TGFβ1) by hUCB-MSC remains unknown.
Fulltext Characterization and Expression of Senescence Marker in Prolonged Passages of Rat Bone Marrow-Derived Mesenchymal Stem Cells

Ridzuan N, Al Abbar A, Yip WK, Maqbool M, Ramasamy R

Stem Cells Int, 2016;2016:8487264.
PMID: 27579045 DOI: 10.1155/2016/8487264

The present study is aimed at optimizing the in vitro culture protocol for generation of rat bone marrow- (BM-) derived mesenchymal stem cells (MSCs) and characterizing the culture-mediated cellular senescence. The initial phase of generation and characterization was conducted using the adherent cells from Sprague Dawley (SD) rat's BM via morphological analysis, growth kinetics, colony forming unit capacity, immunophenotyping, and mesodermal lineage differentiation. Mesenchymal stem cells were successfully generated and characterized as delineated by the expressions of CD90.1, CD44H, CD29, and CD71 and lack of CD11b/c and CD45 markers. Upon induction, rBM-MSCs differentiated into osteocytes and adipocytes and expressed osteocytes and adipocytes genes. However, a decline in cell growth was observed at passage 4 onwards and it was further deciphered through apoptosis, cell cycle, and senescence assays. Despite the enhanced cell viability at later passages (P4-5), the expression of senescence marker, β-galactosidase, was significantly increased at passage 5. Furthermore, the cell cycle analysis has confirmed the in vitro culture-mediated cellular senescence where cells were arrested at the G0/G1 phase of cell cycle. Although the currently optimized protocols had successfully yielded rBM-MSCs, the culture-mediated cellular senescence limits the growth of rBM-MSCs and its potential use in rat-based MSC research.
Fulltext Generation and characterisation of human umbilical cord derived mesenchymal stem cells by explant method

Yusoff Z, Maqbool M, George E, Hassan R, Ramasamy R

Med J Malaysia, 2016 Jun;71(3):105-10.
PMID: 27495882 MyJurnal

Mesenchymal stem cells (MSCs) derived from human umbilical cord (UC) have been considered as an important tool for treating various malignancies, tissue repair and organ regeneration. Umbilical cord-derived mesenchymal stem cells (UC-MSCs) are better alternative to MSCs that derived from bone marrow (BM-MSCs) as they are regarded as medical waste with little ethical concern for research and easily culture-expanded. In this present study, the foetal distal end of human UC was utilised to generate MSC by explant method. Upon in vitro culture, adherent cells with fibroblastic morphology were generated with rapid growth kinetics. Under the respective inductive conditions, these cells were capable of differentiating into adipocytes and osteocytes; express an array of standard MSC's surface markers CD29, CD73, CD90, CD106 and MHC-class I. Further assessment of immunosuppression activity revealed that MSCs generated from UC had profoundly inhibited the proliferation of mitogen-activated T lymphocytes in a dosedependent manner. The current laboratory findings have reinforced the application of explant method to generate UCMSCs thus, exploring an ideal platform to fulfil the increasing demand of MSCs for research and potential clinical use.
Fulltext Impaired redox environment modulates cardiogenic and ion-channel gene expression in cardiac-resident and non-resident mesenchymal stem cells

Subramani B, Subbannagounder S, Ramanathanpullai C, Palanivel S, Ramasamy R

Exp Biol Med (Maywood), 2017 03;242(6):645-656.
PMID: 28092181 DOI: 10.1177/1535370216688568

Redox homeostasis plays a crucial role in the regulation of self-renewal and differentiation of stem cells. However, the behavioral actions of mesenchymal stem cells in redox imbalance state remain elusive. In the present study, the effect of redox imbalance that was induced by either hydrogen peroxide (H2O2) or ascorbic acid on human cardiac-resident (hC-MSCs) and non-resident (umbilical cord) mesenchymal stem cells (hUC-MSCs) was evaluated. Both cells were sensitive and responsive when exposed to either H2O2 or ascorbic acid at a concentration of 400 µmol/L. Ascorbic acid pre-treated cells remarkably ameliorated the reactive oxygen species level when treated with H2O2. The endogenous antioxidative enzyme gene (Sod1, Sod2, TRXR1 and Gpx1) expressions were escalated in both MSCs in response to reactive oxygen species elevation. In contrast, ascorbic acid pre-treated hUC-MSCs attenuated considerable anti-oxidative gene (TRXR1 and Gpx1) expressions, but not the hC-MSCs. Similarly, the cardiogenic gene (Nkx 2.5, Gata4, Mlc2a and β-MHC) and ion-channel gene ( IKDR, IKCa, Ito and INa.TTX) expressions were significantly increased in both MSCs on the oxidative state. On the contrary, reduced environment could not alter the ion-channel gene expression and negatively regulated the cardiogenic gene expressions except for troponin-1 in both cells. In conclusion, redox imbalance potently alters the cardiac-resident and non-resident MSCs stemness, cardiogenic, and ion-channel gene expressions. In comparison with cardiac-resident MSC, non-resident umbilical cord-MSC has great potential to tolerate the redox imbalance and positively respond to cardiac regeneration. Impact statement Human mesenchymal stem cells (h-MSCs) are highly promising candidates for tissue repair in cardiovascular diseases. However, the retention of cells in the infarcted area has been a major challenge due to its poor viability and/or low survival rate after transplantation. The regenerative potential of mesenchymal stem cells (MSCs) repudiate and enter into premature senescence via oxidative stress. Thus, various strategies have been attempted to improve the MSC survival in 'toxic' conditions. Similarly, we investigated the response of cardiac resident MSC (hC-MSCs) and non-resident MSCs against the oxidative stress induced by H2O2. Supplementation of ascorbic acid (AA) into MSCs culture profoundly rescued the stem cells from oxidative stress induced by H2O2. Our data showed that the pre-treatment of AA is able to inhibit the cell death and thus preserving the viability and differentiation potential of MSCs.
Fulltext Musculoskeletal disorders and their relationship with physical activities among office workers: a review

Shariat A, Mohd Tamrin SB, Arumugam M, Danaee M, Ramasamy R

Malaysian Journal of Public Health Medicine, 2016;16(1):62-74.
MyJurnal

Lower back, neck and shoulder pain are the most prevalent musculoskeletal problems affecting office workers worldwide, and they have both personal and socioeconomic consequences as well. Several hypotheses regarding the underlying mechanisms and the maintenance behind office work-related musculoskeletal disorders have been presented. There is some evidence, based on epidemiological studies as well as studies upon smaller groups of subjects, that individuals who sit and work for a long time not only show cognitive impairment at the workplace, but also suffer from poorer and fragmented daytime sleep, in addition to increased risks of developing various psychological, physiological and medical impairments and musculoskeletal disorders. The related physical mechanisms behind musculoskeletal disorders are discussed in the context of new findings. The main causes, as well as varying levels in severity of musculoskeletal disorders,not to mentionthe link between such disorders in the neck, shoulder and lower back regions and physical activity among office workers are also stated. The main objective of this review paper is to conduct a systematic review to identify musculoskeletal disorders and how these disorders are
correlated with physical activity among office workers. The results of this review indicate that the musculoskeletal disorder is a critical issue among office workers and the main cause is related to the absence of physical activity as well as the subjects’ sedentary lifestyle. As a practical message, regular physical activity can be effective in the prevention and decrease of physical discomfort among office workers who suffer from musculoskeletal pain.
Comparative reliability of different instruments used to measure the severity of musculoskeletal disorders in office workers

Shariat A, Tamrin SB, Arumugam M, Danaee M, Ramasamy R

Work, 2016 Jun 8;54(3):753-8.
PMID: 27286077 DOI: 10.3233/WOR-162313

The accuracy of instruments such as questionnaires and the goniometer are critical for measuring the severity of musculoskeletal disorders among office workers.
Fulltext Electromagnetic field exposure as a plausible approach to enhance the proliferation and differentiation of mesenchymal stem cells in clinically relevant scenarios

Hamid HA, Sarmadi VH, Prasad V, Ramasamy R, Miskon A

J Zhejiang Univ Sci B, 2022 Jan 15;23(1):42-57.
PMID: 35029087 DOI: 10.1631/jzus.B2100443

Mesenchymal stem/stromal cell (MSC)‍-based therapy has been regarded as one of the most revolutionary breakthroughs in the history of modern medicine owing to its myriad of immunoregulatory and regenerative properties. With the rapid progress in the fields of osteo- and musculoskeletal therapies, the demand for MSC-based treatment modalities is becoming increasingly prominent. In this endeavor, researchers around the world have devised new and innovative techniques to support the proliferation of MSCs while minimizing the loss of hallmark features of stem cells. One such example is electromagnetic field (EMF) exposure, which is an alternative approach with promising potential. In this review, we present a critical discourse on the efficiency, practicability, and limitations of some of the relevant methods, with insurmountable evidence backing the implementation of EMF as a feasible strategy for the clinically relevant expansion of MSCs.
Package of Office Exercise Training as a New Idea for Office Workers

Shariat A, Bahri Mohd Tamrin S, Arumugam M, Ramasamy R, Danaee M

Iran J Public Health, 2016 Apr;45(4):544-5.
PMID: 27252928
Fulltext Office exercise training to reduce and prevent the occurrence of musculoskeletal disorders among office workers: a hypothesis

Shariat A, Mohd Tamrin SB, Arumugam M, Danaee M, Ramasamy R

Malays J Med Sci, 2016 Jul;23(4):54-8.
PMID: 27660545 MyJurnal DOI: 10.21315/mjms2016.23.4.7

Pain in specific areas of the body (including the lower back, neck, and shoulders) due to extended periods of sitting and inactivity is the most widespread musculoskeletal disorder worldwide and has consequences that are both socio-economic and personal. This condition is particularly prevalent in industrialised countries, affecting roughly 70% to 80% of adults at some point in their lives; approximately 1% of the U.S. population is chronically disabled by this type of pain disorder. A practical way to reduce the prevalence of musculoskeletal pain among office workers would have a significant positive impact. More work is required to develop a package of exercises designed to prevent and treat musculoskeletal pain in office workers. Such a package would be preferable to pharmacological treatments, which can have undesirable side effects. The main objective of this package would be to increase the flexibility and strength of trunk muscles in order to decrease the soreness, pain, and degree of discomfort. In this article, we introduce our proposed package of exercises, which are based on guidelines issued bythe American College of Sports Medicine.
Fulltext Nutritional Compositions and Antiproliferative Activities of Different Solvent Fractions from Ethanol Extract of Cyphomandra betacea (Tamarillo) Fruit

Mutalib MA, Rahmat A, Ali F, Othman F, Ramasamy R

Malays J Med Sci, 2017 Oct;24(5):19-32.
PMID: 29386969 MyJurnal DOI: 10.21315/mjms2017.24.5.3

BACKGROUND: This study aims to examine various solvent extracts of Cyphomandra betacea (tamarillo) also known as the tree tomato, for their bioactive constituents and antioxidant activity. The study also aims to examine its effect on cancer cell death using two types of cancer cell lines (liver and breast cancer cell).
METHODS: The first part of the study evaluates the nutritional composition of tamarillo. Then, phytochemical profiling using GC-MS analysis in ethanolic tamarillo extract was conducted. Different fractions of n-butanol, ethyl acetate and aqueous fractions were obtained from the ethanolic extract of tamarillo. Then, the fractions were subjected to the quantification of total phenol (TPC) and flavonoid contents (TFC), free radical scavenging activity (SA) and also antioxidant activity (AOX) assayed by beta-carotene bleaching (BCB) assay. Finally, the capability of the ethanolic extract of tamarillo and different fractions were evaluated for their anticancer properties.
RESULTS: Findings from this study revealed that the nutritional composition (ash, protein, carbohydrate and total dietary fiber), and mineral levels (calcium, magnesium, potassium and iron) of tamarillo were moderate. The crude ethanol extract of tamarillo contained the highest phenolic and total flavonoid content. FT-IR analysis revealed the presence of alkanes, carboxylic acid, phenol, alkanes, carboxylic acids, aromatics and nitro compounds. Twelve bioactive constituents in tamarillo have been identified through GC-MS analysis. Cytotoxic activity suggests the potential of ethanolic extracts of tamarillo having a chemopreventive effect on breast and liver cancer cells.
CONCLUSION: This study reveals that tamarillo has substantial antioxidant activity as well as anticancer properties.
Evaluation of metabolic and immunological changes in streptozotocin-nicotinamide induced diabetic rats

Mojani MS, Sarmadi VH, Vellasamy S, Sandrasaigaran P, Rahmat A, Peng LS, et al.

Cell Immunol, 2014 May-Jun;289(1-2):145-9.
PMID: 24791700 DOI: 10.1016/j.cellimm.2014.04.004

Type 2 diabetes is a chronic disease with growing public health concern globally. Finding remedies to assist this health issue requires recruiting appropriate animal model for experimental studies. This study was designated to evaluate metabolic and immunologic changes in streptozotocin-nicotinamide induced diabetic rats as a model of type 2 diabetes. Male rats were induced diabetes using nicotinamide (110 mg/kg) and streptozotocin (65 mg/kg). Following 42 days, biochemical and immunological tests showed that diabetic rats had higher levels of blood glucose, WBC, certain abnormalities in lipid profile and insufficient mitogenic responses of lymphocytes (p<0.05). However, the status of the total antioxidant, inflammatory biomarkers and other parameters of full blood count (except HCT) were not significantly altered. Phenotyping assay indicated insignificant lymphocyte subtype imbalances excluding a significant rise in the level of CD4+CD25+ marker (p<0.05). This model of diabetic animals may represent some but not all symptoms of human type 2 diabetes.
Gender effect on in vitro lymphocyte subset levels of healthy individuals

Abdullah M, Chai PS, Chong MY, Tohit ER, Ramasamy R, Pei CP, et al.

Cell Immunol, 2012;272(2):214-9.
PMID: 22078320 DOI: 10.1016/j.cellimm.2011.10.009

Differences in gender immune response have resulted in differences in immune protection and susceptibility to inflammatory diseases. Cultured peripheral blood mononuclear cells (PBMC) are widely used in immunomodulation studies, yet the influence of gender is usually not considered. We examined the effect of in vitro culture and phytohaemagglutinin (PHA) stimulation on PBMC lymphocyte subsets using flowcytometry. Full blood counts of whole blood showed higher levels of lymphocyte in male subjects. Lymphocyte subsets enumeration revealed higher NK cell counts in males and higher B cells in females. Cultured PBMC resulted in significant increases in B and total T cell percentages among females and NK cells among males. PHA stimulated significantly increased percentages of NK and total T cells in males and total activated T cells (CD69+) in females. Our results showed significant gender differences in lymphocyte subsets in cultured conditions. This may affect experimental outcome.

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