Displaying publications 1 - 20 of 68 in total

  1. Ng MH, Kushairi A
    Molecules, 2017 Aug 29;22(9).
    PMID: 28850073 DOI: 10.3390/molecules22091424
    There are six tocol analogs present in palm oil, namely α-tocopherol (α-T), α-tocomonoenol (α-T₁), α-tocotrienol (α-T₃), γ-tocotrienol (γ-T₃), β-tocotrioenol (β-T₃) and δ-tocotrienol (δ-T₃). These analogs were difficult to separate chromatographically due to their similar structures, physical and chemical properties. This paper reports on the effect of pressure and injection solvent on the separation of the tocol analogs in palm oil. Supercritical CO₂ modified with ethanol was used as the mobile phase. Both total elution time and resolution of the tocol analogs decreased with increased pressure. Ethanol as an injection solvent resulted in peak broadening of the analogs within the entire pressure range studied. Solvents with an eluent strength of 3.4 or less were more suitable for use as injecting solvents.
  2. Ng MH, Choo YM
    J Chromatogr Sci, 2016 Apr;54(4):633-8.
    PMID: 26941414 DOI: 10.1093/chromsci/bmv241
    Palm oil is the richest source of natural carotenes, comprising 500-700 ppm in crude palm oil (CPO). Its concentration is found to be much higher in oil extracted from palm-pressed fiber, a by-product from the milling of oil palm fruits. There are 11 types of carotenes in palm oil, excluding the cis/trans isomers of some of the carotenes. Qualitative separation of these individual carotenes is particularly useful for the identification and confirmation of different types of oil as the carotenes profile is unique to each type of vegetable oil. Previous studies on HPLC separation of the individual palm carotenes reported a total analyses time of up to 100 min using C30 stationary phase. In this study, the separation was completed in <5 min. The qualitative separation was successfully carried out using a commonly used stationary phase, C18.
  3. Ng MH, Din AK
    J Sep Sci, 2020 Jan;43(1):285-291.
    PMID: 31294513 DOI: 10.1002/jssc.201900342
    Tocochromanols consisting of tocopherols and tocotrienols, is collectively known as vitamin E. Similarity in their structures, physical and chemical properties rendered the tocochromanols to be subject of chromatography interest. Supercritical fluid chromatography is a highly efficient tool for the separation and analysis of tocochromanols. Separation and analysis of tocochromanols using supercritical fluid chromatography had been carried out in the past using capillary or packed columns. Each of these techniques offer their own advantages and drawbacks. Besides being used for analysis, packed column supercritical fluid chromatography found applications as a purification and content enrichment tool. Emergence of new equipment and stationary phase technologies in recent years also helped in making supercritical fluid chromatography a highly efficient tool for the separation and analysis of tocochromanols. This paper gives an insight into the use of capillary and packed columns in supercritical fluid chromatography for the separation and/or analysis of tocochromanols. The types of stationary phase used, as well as chromatographic conditions are also discussed.
  4. Looi LM, Ng MH, Cheah PL
    Malays J Pathol, 2007 Jun;29(1):33-5.
    PMID: 19105326 MyJurnal
    The unique ability of tumour cells to proliferate indefinitely is crucial to neoplastic progression as it allows these cells to express the aggressive properties of cancer without the censure of physiological ageing. This is in contrast to normal somatic cells which are subject to a "mitotic clock," a phenomenon that has been linked to telomeric shortening after each round of cell replication, so that eventually the loss of genetic material reaches a critical stage and the cells undergo senescence and cell death. A study was conducted to investigate the role of telomerase, an RNA-containing enzyme that restores the telomere length, in the neoplastic cell immortalization and progression process. Fresh human tissue samples taken from excision specimens received by the Department of Pathology, University of Malaya Medical Centre, were investigated for telomerase activity using a commercial Telomerase PCR-ELISA kit (Boehringer Mannheim). Specimens comprised 33 breast lesions (10 infiltrating breast adenocarcinoma, 13 fibroadenoma and 10 non-neoplastic breast tissue), 27 colonic lesions (17 colonic adenocarcinoma and 10 non-neoplastic colonic mucosa) and 42 cervical lesions (20 cervical carcinoma and 22 non-neoplastic cervical tissues). Telomerase activity was found in 6 (60%) of 10 breast carcinomas, 6 (46%) of 13 fibroadenomas, none of the 10 nonneoplastic breast samples, 3 (17.6%) of 17 colon carcinomas and none of the 10 non-neoplastic colonic mucosal samples, 12 (60%) of 20 cervical carcinoma and 3 (13.6%) of 22 non-neoplastic cervical samples. 5/10 (50%) Stage I, 4/7 (57%) Stage II, 2/2 (100%) Stage III and 1/1 (100%) Stage IV cervical carcinomas showed telomerase activity. These findings support a contributory role for telomerase in tumourigenesis with activation occurring from neoplastic transformation and increasing with tumour progression.
  5. Shamsul BS, Aminuddin BS, Ng MH, Ruszymah BH
    Med J Malaysia, 2004 May;59 Suppl B:196-7.
    PMID: 15468885
    Bone marrow harvested by aspiration contains connective tissue progenitor cells which can be selectively isolated and induced to express bone phenotype in vitro. The osteoblastic progenitor can be estimated by counting the number of cells attach using the haemacytometer. This study was undertaken to test the hypothesis that human aging is associated with a significant change on the number of osteoblastic progenitors in the bone marrow. Bone marrow aspirates were harvested from 38 patients, 14 men (age 11-70) and 24 women (age 10-70) and cultured in F12: DMEM (1:1). In total 15 bone marrow samples have been isolated from patients above 40 years old (men/women) of age. Fourteen (93.3%) of this samples failed to proliferate. Only one (6.7%) bone marrow sample from a male patient, aged 59 years old was successfully cultured. Seventy percent (16/23) of the samples from patient below than 40 years old were successfully cultured. However, our observation on the survival rate for cells of different gender from patient below 40 years old does not indicate any significant difference. From this study, we conclude that the growth of bone marrow stromal cells possibly for bone engineering is better from bone marrow aspirates of younger patient.
  6. Boon CM, Ng MH, Choo YM, Mok SL
    PLoS One, 2013;8(2):e55908.
    PMID: 23409085 DOI: 10.1371/journal.pone.0055908
    Oleic acid has been shown to lower high blood pressure and provide cardiovascular protection. Curiosity arises as to whether super olein (SO), red palm olein (RPO) and palm olein (PO), which have high oleic acid content, are able to prevent the development of hypertension.
  7. Raviadaran R, Ng MH, Manickam S, Chandran D
    Ultrason Sonochem, 2019 Apr;52:353-363.
    PMID: 30555038 DOI: 10.1016/j.ultsonch.2018.12.012
    This study aimed to formulate a stable palm oil-based water-in-oil (W/O) nano-emulsion. Emphasis was placed on the effects of polyglycerol polyricinoleate (PGPR), medium chain triglyceride (MCT), lecithin and sodium chloride (NaCl) addition towards the stability of nano-emulsion. Among the performed analyses were mean droplet diameter (MDD), dispersity index (DI), critical micelle concentration (CMC), lipid peroxidation, viscosity, sedimentation index (SI) and surface morphology. The most stable optimized palm oil-based W/O nano-emulsion was produced using 61.25 wt% of palm oil, 26.25 wt% of MCT, 2.5 wt% of PGPR and 10 wt% of water (0.5 M of NaCl). The MDD and DI of the obtained W/O nano-emulsion were 143.1 ± 8.8 and 0.131 ± 0.094, respectively. After 2 weeks, no sedimentation was observed in W/O nano-emulsion with MDD and DI were 151.2 ± 6.5 nm and 0.156 ± 0.025 respectively. This study clearly found that polyricinoleate non-polar fatty acids of PGPR bound to non-polar fatty acids of palm oil through van der Waals intermolecular forces. While, polyglycerol polar head of PGPR interacts with water molecules through hydrogen bonding, as well as by the bound glyceride units of palm oil. The addition of NaCl further reduced MDD by 70 nm and improved the stability of nano-emulsion through electrostatic and steric repulsions attributed to the dissociation of Na+ and Cl- ions. This study aids to widen the knowledge and interest on the utilization of palm oil for the generation of W/O nano-emulsion, as well as to better understand the interaction between palm oil and PGPR/NaCl in producing nano-emulsion.
  8. Raviadaran R, Ng MH, Manickam S, Chandran D
    Ultrason Sonochem, 2020 Jun;64:104995.
    PMID: 32106064 DOI: 10.1016/j.ultsonch.2020.104995
    In this work, the effects of thickeners and tonicity towards producing stable palm oil-based water-in-oil-in-water (W/O/W) multiple nanoemulsion using ultrasound and microfluidizer were investigated. Palm oil, Sucragel, polyglycerol polyricinoleate, Tween 80, Xanthan gum, and NaCl were used. W/O/W was formed under the optimized conditions of ultrasound at 40% amplitude and for 180 s of irradiation time, whereas for the microfluidizer, the optimized conditions were 350 bar and 8 cycles. This is the first work that successfully utilized Sucragel (oil-based thickener) in imparting enhanced stability in W/O/W. W/O/W with isotonic stabilization produced the lowest change in the mean droplet diameter (MDD), NaCl concentration, and water content by 1.5%, 2.6%, and 0.4%, respectively, due to reduced water movement. The final optimized W/O/W possessed MDD and dispersity index of 175.5 ± 9.8 and 0.232 ± 0.012, respectively. The future direction of formulating stable W/O/W would be by employing oil phase thickeners and isotonicity. The observed ~12 times lesser energy consumed by ultrasound than microfluidizer to generate a comparable droplet size of ~235 nm, further confirms its potential in generating the droplets energy-efficiently.
  9. Looi LM, Azura WW, Cheah PL, Ng MH
    Pathology, 2001 Aug;33(3):283-6.
    PMID: 11523925
    This investigation was carried out to gain insight into the prevalence of pS2 expression in invasive ductal breast carcinoma in the Malaysian population and its correlation with oestrogen receptor (ER) protein expression and tumour aggressiveness. Seventy consecutive infiltrating ductal breast carcinomas treated with mastectomy and axillary lymph node clearance were investigated, using the standard avidin-biotin complex immunoperoxidase method with microwave antigen retrieval and commercial monoclonal antibodies (Dako), for expression of pS2 and human ER. This was correlated against histological grade (modified Bloom and Richardson) and the presence of axillary lymph node metastasis of these carcinomas. Four (5.7%) were grade 1, 40 (57.1%) grade 2 and 26 (37.1%) grade 3 tumours. A total of 45 (64%) showed histological evidence of axillary lymph node metastasis. Forty (57%) were ER-positive, while 31 (44%) were pS2-positive. There was a statistically significant correlation between pS2 and ER expressions (chi2-test with Yates correction: P<0.005). There was no correlation between pS2 expression and histological grade (P>0.1) and the presence of lymph node metastasis (P>0.1). Our findings support the views that pS2 may be a co-marker of endocrine responsiveness in invasive breast cancer and that it does not influence breast cancer biology in terms of potential for metastatic spread.
  10. Lim WL, Chowdhury SR, Ng MH, Law JX
    PMID: 33947053 DOI: 10.3390/ijerph18094764
    Tissue-engineered substitutes have shown great promise as a potential replacement for current tissue grafts to treat tendon/ligament injury. Herein, we have fabricated aligned polycaprolactone (PCL) and gelatin (GT) nanofibers and further evaluated their physicochemical properties and biocompatibility. PCL and GT were mixed at a ratio of 100:0, 70:30, 50:50, 30:70, 0:100, and electrospun to generate aligned nanofibers. The PCL/GT nanofibers were assessed to determine the diameter, alignment, water contact angle, degradation, and surface chemical analysis. The effects on cells were evaluated through Wharton's jelly-derived mesenchymal stem cell (WJ-MSC) viability, alignment and tenogenic differentiation. The PCL/GT nanofibers were aligned and had a mean fiber diameter within 200-800 nm. Increasing the GT concentration reduced the water contact angle of the nanofibers. GT nanofibers alone degraded fastest, observed only within 2 days. Chemical composition analysis confirmed the presence of PCL and GT in the nanofibers. The WJ-MSCs were aligned and remained viable after 7 days with the PCL/GT nanofibers. Additionally, the PCL/GT nanofibers supported tenogenic differentiation of WJ-MSCs. The fabricated PCL/GT nanofibers have a diameter that closely resembles the native tissue's collagen fibrils and have good biocompatibility. Thus, our study demonstrated the suitability of PCL/GT nanofibers for tendon/ligament tissue engineering applications.
  11. Liau LL, Ruszymah BHI, Ng MH, Law JX
    Curr Res Transl Med, 2020 01;68(1):5-16.
    PMID: 31543433 DOI: 10.1016/j.retram.2019.09.001
    Mesenchymal stromal cells (MSCs) are widely used in the clinic because they involve fewer ethical issues and safety concerns compared to other stem cells such as embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs). MSCs derived from umbilical cord Wharton's jelly (WJ-MSCs) have excellent proliferative potential and a faster growth rate and can retain their multipotency for more passages in vitro compared to adult MSCs from bone marrow or adipose tissue. WJ-MSCs are used clinically for repairing tissue injuries of the spinal cord, liver and heart with the aim of regenerating tissue. On the other hand, WJ-MSCs are also used clinically to ameliorate immune-mediated diseases based on their ability to modulate immune responses. In the field of tissue engineering, WJ-MSCs capable of differentiating into multiple cell lineages have been used to produce a variety of engineered tissues in vitro that can then be transplanted in vivo. This review discusses the characteristics of WJ-MSCs, the differences between WJ-MSCs and adult MSCs, clinical studies involving WJ-MSCs and future perspectives of WJ-MSC research and clinical applications. To summarize, WJ-MSCs have shown promise in treating a variety of diseases clinically. However, most clinical trials/studies reported thus far are relatively smaller in scale. The collected evidence is insufficient to support the routine use of WJ-MSC therapy in the clinic. Thus, rigorous clinical trials are needed in the future to obtain more information on WJ-MSC therapy safety and efficacy.
  12. Yeo GEC, Ng MH, Nordin FB, Law JX
    Int J Mol Sci, 2021 May 27;22(11).
    PMID: 34072224 DOI: 10.3390/ijms22115749
    Rapid growth of the geriatric population has been made possible with advancements in pharmaceutical and health sciences. Hence, age-associated diseases are becoming more common. Aging encompasses deterioration of the immune system, known as immunosenescence. Dysregulation of the immune cell production, differentiation, and functioning lead to a chronic subclinical inflammatory state termed inflammaging. The hallmarks of the aging immune system are decreased naïve cells, increased memory cells, and increased serum levels of pro-inflammatory cytokines. Mesenchymal stem cell (MSC) transplantation is a promising solution to halt immunosenescence as the cells have excellent immunomodulatory functions and low immunogenicity. This review compiles the present knowledge of the causes and changes of the aging immune system and the potential of MSC transplantation as a regenerative therapy for immunosenescence.
  13. Raviadaran R, Ng MH, Chandran D, Ooi KK, Manickam S
    Mater Sci Eng C Mater Biol Appl, 2021 Feb;121:111808.
    PMID: 33579452 DOI: 10.1016/j.msec.2020.111808
    This work aimed to evaluate the effects of encapsulated tocotrienols (TRF) and caffeic acid (CA) in water-in-oil-in-water (W/O/W) multiple nanoemulsion with cisplatin towards cancer cells. This work is important considering the limited efficacy of cisplatin due to tumour resistance, as well as its severe side effects. A549 and HEP G2 cancer cell lines were utilised for evaluating the efficacy of the encapsulated W/O/W while HEK 293 normal cell line was used for evaluating the toxicity. TRF, CA and CIS synergistically improved apoptosis in the late apoptotic phase in A549 and HEP G2 by 23.1% and 24.9%, respectively. The generation of ROS was enhanced using TRF:CA:CIS by 16.9% and 30.2% for A549 and HEP G2, respectively. Cell cycle analysis showed an enhanced cell arrest in the G0/G1 phase for both A549 and HEP G2. TRF, CA and CIS led to cell death in A549 and HEP G2. For HEK 293, ~33% cell viability was found when only CIS was used while >95% cell viability was observed when TRF, CA and CIS were used. This study demonstrates that the encapsulated TRF and CA in W/O/W with CIS synergistically improved therapeutic efficacy towards cancer cells, as well as lowered the toxicity effects towards normal cells.
  14. Cheah PL, Looi LM, Ng MH, Sivanesaratnam V
    J Clin Pathol, 2002 Jan;55(1):22-6.
    PMID: 11825919
    AIM: Telomerase activity was studied in invasive uterine cervical carcinoma to assess whether it was activated during cervical malignant transformation and to look for a possible association with human papillomavirus (HPV) infection in a set of Malaysian patients.

    METHODS: Histologically confirmed invasive cervical carcinoma and benign cervices were assayed for telomerase activity using a commercial telomerase polymerase chain reaction (PCR) enzyme linked immunosorbent assay kit. The same cases were subjected to PCR detection of HPV using type specific (HPV types 6b, 11, 16, and 18) followed by L1 open reading frame (ORF) consensus primers.

    RESULTS: HPV was detected in 18 (13 HPV-16, one HPV-6b, four only L1 ORF) of 20 invasive cervical carcinoma and one (only L1 ORF) of 19 benign cervices. Raised telomerase activity (A(450 nm) > 0.215) was detected in 11 cervical carcinomas, with A(450 nm) ranging between 0.238 and 21.790 (mean, 3.952) in positive squamous carcinomas, whereas A(450 nm) was only 0.222 in the one positive adenosquamous carcinoma. Five of 11 cervical carcinomas in stage I, three of six in stage II, both in stage III, and the only case in stage IV showed telomerase activation. Increased telomerase activity was noted in five of the 12 lymph node negative, five of the seven lymph node status unknown cases, and the one case with presumed lymph node metastasis. Ten of 18 HPV positive and one of two HPV negative cervical carcinomas showed telomerase upregulation.

    CONCLUSIONS: Telomerase is activated in invasive cervical carcinoma. Although larger studies are needed, there seems to be no clear association between telomerase upregulation and HPV status, although there is a suggestion of increased telomerase activity in squamous carcinomas and late stage disease.

  15. Mahindran E, Law JX, Ng MH, Nordin F
    Int J Mol Sci, 2021 Sep 29;22(19).
    PMID: 34638883 DOI: 10.3390/ijms221910542
    Projected life expectancy continues to grow worldwide owing to the advancement of new treatments and technologies leading to rapid growth of geriatric population. Thus, age-associated diseases especially in the musculoskeletal system are becoming more common. Loss of bone (osteoporosis) and muscle (sarcopenia) mass are conditions whose prevalence is increasing because of the change in population distribution in the world towards an older mean age. The deterioration in the bone and muscle functions can cause severe disability and seriously affects the patients' quality of life. Currently, there is no treatment to prevent and reverse age-related musculoskeletal frailty. Existing interventions are mainly to slow down and control the signs and symptoms. Mesenchymal stem cell (MSC) transplantation is a promising approach to attenuate age-related musculoskeletal frailty. This review compiles the present knowledge of the causes and changes of the musculoskeletal frailty and the potential of MSC transplantation as a regenerative therapy for age-related musculoskeletal frailty.
  16. Mirzasadeghi A, Narayanan SS, Ng MH, Sanaei R, Cheng CH, Bajuri MY, et al.
    Biomed Mater Eng, 2014;24(6):2177-86.
    PMID: 25226916 DOI: 10.3233/BME-141029
    The application of bone substitutes and cements has a long standing history in augmenting fractures as a complement to routine fracture fixation techniques. Nevertheless, such use is almost always in conjunction with definite means of fracture fixation such as intramedullary pins or bone plates. The idea of using biomaterials as the primary fixation bears the possibility of simultaneous fixation and bone enhancement. Intramedullary recruitment of bone cements is suggested in this study to achieve this goal. However, as the method needs primary testings in animal models before human implementation, and since the degree of ambulation is not predictable in animals, this pilot study only evaluates the outcomes regarding the feasibility and safety of this method in the presence of primary bone fixators. A number of two sheep were used in this study. Tibial transverse osteotomies were performed in both animals followed by external skeletal fixation. The medullary canals, which have already been prepared by removing the marrow through proximal and distal drill holes, were then injected with calcium phosphate cement (CPC). The outcomes were evaluated postoperatively by standard survey radiographs, morphology, histology and biomechanical testings. Healing processes appeared uncomplicated until week four where one bone fracture recurred due to external fixator failure. The results showed 56% and 48% cortical thickening, compared to the opposite site, in the fracture site and proximal and distal diaphyses respectively. This bone augmentative effect resulted in 264% increase in bending strength of the fracture site and 148% increase of the same value in the adjacent areas of diaphyses. In conclusion, IMCO, using CPC in tibia of sheep, is safe and biocompatible with bone physiology and healing. It possibly can carry the osteopromotive effect of the CPCs to provide a sustained source of bone augmentation throughout the diaphysis. Although the results must be considered preliminary, this method has possible advantages over conventional methods of bone fixation at least in bones with compromised quality (i.e. osteoporosis and bone cysts), where rigid metal implants may jeopardize eggshell cortices.
  17. Chowdhury SR, Ng MH, Hassan NS, Aminuddin BS, Ruszymah BH
    Hum. Cell, 2012 Sep;25(3):69-77.
    PMID: 22968953
    This study was undertaken in order to identify the best culture strategy to expand and osteogenic differentiation of human bone marrow stem cells (hBMSCs) for subsequent bone tissue engineering. In this regard, the experiment was designed to evaluate whether it is feasible to bypass the expansion phase during hBMSCs differentiation towards osteogenic lineages by early induction, if not identification of suitable culture media for enhancement of hBMSCs expansion and osteogenic differentiation. It was found that introduction of osteogenic factors in alpha-minimum essential medium (αMEM) during expansion phase resulted in significant reduction of hBMSCs growth rate and osteogenic gene expressions. In an approach to identify suitable culture media, the growth and differentiation potential of hBMSCs were evaluated in αMEM, F12:DMEM (1:1; FD), and FD with growth factors. It was found that αMEM favors the expansion and osteogenic differentiation of hBMSCs compared to that in FD. However, supplementation of growth factors in FD, only during expansion phase, enhances the hBMSCs growth rate and significantly up-regulates the expression of CBFA-1 (the early markers of osteogenic differentiation) during expansion, and, other osteogenic genes at the end of induction compared to the cells in αMEM and FD. These results suggested that the expansion and differentiation phase of the hBMSCs should be separately and carefully timed. For bone tissue engineering, supplementation of growth factors in FD only during the expansion phase was sufficient to promote hBMSCs expansion and differentiation, and preferably the most efficient culture condition.
  18. Ude CC, Shamsul BS, Ng MH, Chen HC, Norhamdan MY, Aminuddin BS, et al.
    Tissue Cell, 2012 Jun;44(3):156-63.
    PMID: 22402173 DOI: 10.1016/j.tice.2012.02.001
    Tracking of transplanted cells has become an important procedure in cell therapy. We studied the in vitro dye retention, survival and in vivo tracking of stem cells with PKH26 dye. Sheep BMSCs and ADSCs were labeled with 2, 4 and 8 μmol of PKH26 and monitored for six passages. Labeled BMSCs and ADSCs acquired mean cumulative population doubling of 12.7±0.4 and 14.6±0.5; unlabeled samples had 13.8±0.5 and 15.4±0.6 respectively. Upon staining with 2, 4 and 8 μmol PKH26, BMSCs had retentions of 40.0±5.8, 60.0±2.9 and 95.0±2.9%, while ADSCs had 92.0±1.2, 95.0±1.2 and 98.0±1.2%. ADSCs retentions were significantly higher at 2 and 4 μmol. On dye retention comparison at 8 μmol and 4 μmol for BMSCs and ADSCs; ADSCs were significantly higher at passages 2 and 3. The viability of BMSCs reduced from 94.0±1.2% to 90.0±0.6% and ADSCs from 94.0±1.2% to 52.0±1.2% (p<0.05) after 24h. BMSCs had significant up regulation of the cartilage genes for both the labeled and the unlabeled samples compared to ADSCs (p<0.05). PKH26 fluorescence was detected on the resected portions of the regenerated neo-cartilage. The recommended concentration of PKH26 for ADSCs is 2 μmol and BMSCs is 8 μmol, and they can be tracked up to 49 days.
  19. Looi LM, Cheah PL, Ng MH, Yip CH, Mun KS, Rahman NA
    Asian Pac J Cancer Prev, 2010;11(3):713-6.
    PMID: 21039041
    A study was initiated to explore possible differences in handling telomere attrition in the most common lignant and benign tumours of the breast in Malaysian women. Infiltrating ductal carcinoma (IDC) and fibroadenoma (FA) represented the malignant and benign prototypes respectively. 29 IDC, 28 FA and 22 benign non-lesional control (BNL) breast tissue samples were analysed for telomerase activation using a Telomerase PCR ELISA kit (Boehringer Mannheim). In addition, 23 IDC, 12 FA and 14 BNL were subjected to telomere length determination with a TeloTAGGG Telomere Length Assay Kit (Roche Diagnostic GmbH, Germany), following digestion of genomic DNA by frequently cutting restriction enzymes RsaI and HinfI. Mean telomerase activity in IDC (A450nm=0.3338), but not FA (A450nm=0.0003) was significantly raised (p<0.05) compared with BNL (A450nm=0.0031). Similarly IDC (1.2 kb), but not FA (2.2 kb), showed significant telomere shortening (p<0.05) relative to BNL (2.9 kb). The findings imply that telomere attrition and telomerase activation differ between malignant and benign tumours of the breast and may be important for targeted therapy.
  20. Ng MH, Aminuddin BS, Hamizah S, Lynette C, Mazlyzam AL, Ruszymah BH
    J Tissue Viability, 2009 Nov;18(4):109-16.
    PMID: 19632116 DOI: 10.1016/j.jtv.2009.06.003
    Previous studies suggested telomerase activity as a determinant of cell replicative capacity by delaying cell senescence. This study aimed to evaluate the feasibility of adopting telomerase activity as a selection criterion for in vitro expanded skin cells before autologous transplantation. Fibroblasts and keratinoctyes were derived from the same consenting patients aged 9-69 years, and cultured separately in serum-supplemented and serum-free media, respectively. Telomerase activity of fresh and cultured cells were measured and correlated with cell growth rate, donor age and passage number. The results showed that telomerase activity and cell growth were independent of donor age for both cell types. Telomerase was expressed in freshly digested epidermis and dermis and continued expressing in vitro. Keratinocytes consistently showed 3-12 folds greater telomerase activity than fibroblast both in vivo and in vitro. Conversely, growth rate for fibroblast exceeded that of keratinocyte. Telomerase activity decreased markedly at Passage 6 for keratinocytes and ceased by Passage 3 for fibroblasts. The decrease or cessation of telomerase activity coincided with senescence for keratinocyte but not for fibroblast, implying a telomerase-regulated cell senescence for the former and hence a predictor of replicative capacity for this cell type. Relative telomerase activity for fibroblasts from the younger age group was significantly higher than that from the older age group; 69.7% higher for fresh isolates and 31.1% higher at P0 (p<0.05). No detectable telomerase activity was to be found at later subcultures for both age groups. Similarly for keratinocytes, telomerase activity in the younger age group was significantly higher (p<0.05) compared to that in the older age group; 507.7% at P0, 36.8% at P3 and the difference was no longer significant at P6. In conclusion, the study provided evidence that telomerase sustained the proliferation of keratinocytes but not fibroblasts. Telomerase activity is an important criterion for continued survival and replication of keratinocytes, hence its positive detection before transplantation is desirable. Inferring from our results, the use of keratinocytes from Passage 3 or lesser for construction of skin substitute or cell-based therapy is recommended owing to their sustained telomerase expression.
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