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Tong C.K.,University Putra Malaysia | Vellasamy S.,University Putra Malaysia | Tan B.C.,Britannia Women and Children Specialist Center | Abdullah M.,University Putra Malaysia | And 3 more authors.
Cell Biology International

MSCs (mesenchymal stem cells) promise a great potential for regenerative medicine due to their unique properties of selfrenewal, high plasticity, modulation of immune response and the flexibility for genetic modification. Therefore, the increasing demand for cellular therapy necessitates a larger-scale production of MSC; however, the technical and ethical issues had put a halt on it. To date, studies have shown that MSC could be derived from human UC (umbilical cord), which is once considered as clinical waste. We have compared the two conventional methods which are classic enzymatic digestion and explant method with our newly tailored enzymatic-mechanical disassociation method to generate UC-MSC. The generated UC-MSCs from the methods above were characterized based on their immunophenotyping, early embryonic transcription factors expression and mesodermal differentiation ability. Our results show that enzymatic-mechanical disassociation method increase the initial nucleated cell yield greatly (approximately 160-fold) and maximized the successful rate of UC-MSC generation. Enzymatic-mechanical disassociation-derived UC-MSC exhibited fibroblastic morphology and surface markers expression of CD105, CD73, CD29, CD90 and MHC class I. Furthermore, these cells constitutively express early embryonic transcription factors (Nanog, Oct-4, Sox-2 and Rex-1), as confirmed by RT-PCR, indicating their multipotency and high self-renewal capacity. They are also capable of differentiating into osteoblasts and adipocytes when given an appropriate induction. The present study demonstrates a new and efficient approach in generating MSC from UC, hence serving as ideal alternative source of mesenchymal stem cell for clinical and research use. © The Author(s) Journal compilation. © 2011 Portland Press Limited. Source

Fonseka M.,University Putra Malaysia | Ramasamy R.,University Putra Malaysia | Tan B.C.,Britannia Women and Children Specialist Center | Seow H.F.,University Putra Malaysia
Cell Biology International

hUCB-MSC (human umbilical cord blood-derived mesenchymal stem cells) offer an attractive alternative to bone marrowderived 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 hUCBMSC. 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. hUCBMSC inhibited proliferation of K562, arresting them in the G0/G1 phase. NO (nitric oxide) was not involved in the hUCB-MSCmediated 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, IFNγ (interferon γ), 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-cul tured with K562. The ant i-proliferative ef fect 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. © The Author(s) Journal Compilation © 2012 International Federation for Cell Biology. Source

Chong P.P.,University Putra Malaysia | Ng C.K.,Kajang Hospital | Hamilton W.H.W.,Serdang Hospital | Tan B.C.,Britannia Women and Children Specialist Center | Noraihan N.,Putrajaya Hospital
Asian Pacific Journal of Cancer Prevention

Persistent high-risk human papillomavirus (HPV) infection is known to play an important role in the genesis of cervical cancer. Since new screening and prevention strategies, namely improved HPV testing and HPV vaccination have been aggressively promoted recently, it is crucial to investigate the HPV distribution in Malaysia in order to maximize their cost-effectiveness. This study was therefore conducted to assess the HPV type distribution in the most populous region, the state of Selangor. A total of 200 cervical swab samples were collected in two health-screening campaigns, and also from women attending obstetrics and gynecology clinics in several hospitals in Selangor. DNA extraction was performed and HPV DNA was detected via nested PCR using MY09/MY11 as outer primers and GP5+/GP6+ as inner primers which target the L1 gene of the viral genome. The purified PCR products were subjected to automated DNA sequencing to determine the HPV genotype. Out of 180 β-globin positive samples, 84 (46.7%) were positive for HPV DNA. The most common HPV type found was high-risk oncogenic type 16 (40%), followed by HPV type 18 (3.3%), HPV 33 (1.7%), HPV 31 (0.6%), and low-risk HPV 87 (0.6%). Our study confirmed that nested PCR method is highly sensitive in detecting HPV DNA even in low risk patients. Since a relatively high prevalence rate of HPV infection was found in this population, prompt healthcare policy changes to bring about implementation of early HPV vaccination program is desirable to prevent a high incidence of cervical cancer. Source

Ramasamy R.,University Putra Malaysia | Tong C.K.,University Putra Malaysia | Yip W.K.,University Putra Malaysia | Vellasamy S.,University Putra Malaysia | And 2 more authors.
Cell Proliferation

Background: Mesenchymal stem cells (MSC) have great potential in regenerative medicine, immunotherapy and gene therapy due to their unique properties of self-renewal, high plasticity, immune modulation and ease for genetic modification. However, production of MSC at sufficient clinical scale remains an issue as in vitro generation of MSC inadequately fulfils the demand with respect to patients. Objectives: This study has aimed to establish optimum conditions to generate and characterize MSC from human umbilical cord (UC-MSC). Materials and methods: To optimize MSC population growth, basic fibroblast growth factor (bFGF) was utilized in culture media. Effects of bFGF on expansion kinetics, cell cycle, survival of UC-MSC, cytokine secretion, expression of early stem-cell markers and immunomodulation were investigated. Results: bFGF supplementation profoundly enhanced UC-MSC proliferation by reducing population doubling time without altering immunophenotype and immunomodulatory function of UC-MSC. However, cell cycle studies revealed that bFGF drove the cells into the cell cycle, as a higher proportion of cells resided in S phase and progressed into M phase. Consistent with this, bFGF was shown to promote expression of cyclin D proteins and their relevant kinases to drive UC-MSC to transverse cell cycle check points, thus, committing the cells to DNA synthesis. Furthermore, supplementation with bFGF changed the cytokine profiles of the cells and reduced their apoptotic level. Conclusion: Our study showed that bFGF supplementation of UC-MSC culture enhanced the cells' growth kinetics without compromising their nature. © 2012 Blackwell Publishing Ltd. Source

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