Displaying publications 1 - 20 of 53 in total

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  1. Sharif R, Thomas P, Zalewski P, Fenech M
    Mol Nutr Food Res, 2015 Jun;59(6):1200-12.
    PMID: 25755079 DOI: 10.1002/mnfr.201400784
    An increased intake of Zinc (Zn) may reduce the risk of degenerative diseases but may prove to be toxic if taken in excess. This study aimed to investigate whether zinc carnosine supplement can improve Zn status, genome stability events, and Zn transporter gene expression in an elderly (65-85 years) South Australian cohort with low plasma Zn levels.
    Matched MeSH terms: Comet Assay
  2. Sharif R, Ghazali AR, Rajab NF, Haron H, Osman F
    Food Chem Toxicol, 2008 Jan;46(1):368-74.
    PMID: 17900779
    Malaysian locally processed raw food products are widely used as main ingredients in local cooking. Previous studies showed that these food products have a positive correlation with the incidence of cancer. The cytotoxicity effect was evaluated using MTT assay (3-(4,5-dimetil-2-thiazolil)-2,5-diphenyl-2H-tetrazolium bromide) against Chang liver cells at 2000 microg/ml following 72 h incubation. Findings showed all methanol extracts caused a tremendous drop in the percentage of cell viability at 2000 microg/ml (shrimp paste - 41.69+/-3.36%, salted fish - 37.2+/-1.06%, dried shrimp - 40.32+/-1.8%, p<0.05). To detect DNA damage in a single cell, alkaline Comet Assay was used. None of the extracts caused DNA damage to the Chang liver cells at 62.5 microg/ml following 24 h incubation, as compared to the positive control, hydrogen peroxide (tail moment - 9.50+/-1.50; tail intensity - 30.50+/-2.50). Proximate analysis which was used for the evaluation of macronutrients in food showed that shrimp paste did not comply with the protein requirement (<25%) as in Food Act 1983. Salt was found in every sample with the highest percentage being detected in shrimp paste which exceeded 20%. Following heavy metal analysis (arsenic, cadmium, lead and mercury), arsenic was found in every sample with dried shrimps showing the highest value as compared to the other samples (6.16 mg/kg). In conclusion, several food extracts showed cytotoxic effect but did not cause DNA damage against Chang liver cells. Salt was found as the main additive and arsenic was present in every sample, which could be the probable cause of the toxicity effects observed.
    Matched MeSH terms: Comet Assay
  3. Makpol S, Durani LW, Chua KH, Mohd Yusof YA, Ngah WZ
    J Biomed Biotechnol, 2011;2011:506171.
    PMID: 21541185 DOI: 10.1155/2011/506171
    This study determined the molecular mechanisms of tocotrienol-rich fraction (TRF) in preventing cellular senescence of human diploid fibroblasts (HDFs). Primary culture of HDFs at various passages were incubated with 0.5 mg/mL TRF for 24 h. Telomere shortening with decreased telomerase activity was observed in senescent HDFs while the levels of damaged DNA and number of cells in G(0)/G(1) phase were increased and S phase cells were decreased. Incubation with TRF reversed the morphology of senescent HDFs to resemble that of young cells with decreased activity of SA-β-gal, damaged DNA, and cells in G(0)/G(1) phase while cells in the S phase were increased. Elongated telomere length and restoration of telomerase activity were observed in TRF-treated senescent HDFs. These findings confirmed the ability of tocotrienol-rich fraction in preventing HDFs cellular ageing by restoring telomere length and telomerase activity, reducing damaged DNA, and reversing cell cycle arrest associated with senescence.
    Matched MeSH terms: Comet Assay
  4. Salman, M.O., Al-Wasiti, E.A., Thamir, K.A., Al-Ani, I.M., Al-Salihi, A.R.
    MyJurnal
    Introduction: We aim to investigate the effect of vasectomy on the histology of the testis as well as to evaluate DNA fragmentation in testicular tissue of male mice. Methods: Bilateral vasectomy was performed on 20 mature male mice; 10 control mice underwent sham-operation. After 6 weeks, the testes were evaluated for histological changes and DNA fragmentation by single cell gel electrophoresis (comet assay). Results: Marked alterations were observed in the testes of vasectomized mice, including degeneration of spermatids, thickened basement membrane, dilatation of the seminiferous tubules, exfoliation of germ cells, reduction in the seminiferous cell population, vacuolated appearance of the epithelium in the tubules and marked interstitial fibrosis. Single cell gel electrophoresis showed a highly significant (P
    Matched MeSH terms: Comet Assay
  5. Hussin F, Eshkoor SA, Rahmat A, Othman F, Akim A
    BMC Complement Altern Med, 2014 Jan 20;14:32.
    PMID: 24444147 DOI: 10.1186/1472-6882-14-32
    BACKGROUND: This paper is to investigate the effects of Centella asiatica on HepG2 (human hepatocellular liver carcinoma cell line). Centella asiatica is native to the Southeast Asia that is used as a traditional medicine. This study aims to determine the chemopreventive effects of the Centella asiatica juice on human HepG2 cell line.

    METHODS: Different methods including flow cytometry, comet assay and reverse transcription-polymerase chain reaction (RT-PCR) were used to show the effects of juice exposure on the level of DNA damage and the reduction of cancerous cells. MTT assay is a colorimetric method applied to measure the toxic effects of juice on cells.

    RESULTS: The Centella asiatica juice was not toxic to normal cells. It showed cytotoxic effects on tumor cells in a dose dependent manner. Apoptosis in cells was started after being exposed for 72 hr of dose dependent. It was found that the higher percentage of apoptotic cell death and DNA damage was at the concentration above 0.1%. In addition, the juice exposure caused the reduction of c-myc gene expression and the enhancement of c-fos and c-erbB2 gene expressions in tumor cells.

    CONCLUSIONS: It was concluded that the Centella asiatica juice reduced liver tumor cells. Thus, it has the potential to be used as a chemopreventive agent to prevent and treat liver cancer.

    Matched MeSH terms: Comet Assay
  6. Hussin F, Eshkoor SA, Rahmat A, Othman F, Akim A, Eshak Z
    Asian Pac J Cancer Prev, 2015;16(14):6047-53.
    PMID: 26320494
    BACKGROUND: Hepatocellular carcinoma is one of the most common cancers worldwide. Its prevalence is increasing in many countries. Plant products can be used to protect against cancer due to natural anticancer and chemopreventive constituents. Strobilanthes crispus is one of plants with potential chemopreventive ability.

    OBJECTIVE: This study aimed to evaluate the anticancer effects of Strobilanthes crispus juice on hepatocellular carcinoma cells.

    MATERIALS AND METHODS: MTT assays, flow cytometry, comet assays and the reverse transcription- polymerase chain reaction (RT-PCR) were used to determine the effects of juice on DNA damage and cancer cell numbers.

    RESULTS: This juice induced apoptosis after exposure of the HepG2 cell line for 72 h. High percentages of apoptotic cell death and DNA damage were seen at the juice concentrations above 0.1%. It was found that the juice was not toxic for normal cells. In addition, juice exposure increased the expression level of c-myc gene and reduced the expression level of c-fos and c-erbB2 genes in HepG2 cells. The cytotoxic effects of juice on abnormal cells were in dose dependent.

    CONCLUSIONS: It was concluded that the Strobilanthes crispus juice may have chemopreventive effects on hepatocellular carcinoma cells.

    Matched MeSH terms: Comet Assay
  7. Chye SM, Hseu YC, Liang SH, Chen CH, Chen SC
    Bull Environ Contam Toxicol, 2008 Jan;80(1):58-62.
    PMID: 18058049
    Para-Phenylenediamine (PPD), the main aromatic amines used in the hair dye formation, and its four derivatives (2-chloro-p-phenylenediamine, 4-chloro-o-phenylenediamine, 2-nitro-p-phenylenediamine, and 4-nitro-o-phenylenediamine) were examined for their potential to produce single strand DNA breaks in human lymphocytes using the alkaline comet assay. Results revealed that all the tested chemicals within the range of doses from 100 microM to 500 microM showed the genotoxicity in a dose-dependent manner after the incubation of lymphocytes with these chemicals for 2 h. In this study, we first reported that PPD and its four derivatives can elicit the type of single strand breaks in human lymphocytes.
    Matched MeSH terms: Comet Assay/methods*
  8. Shahar S, Shafurah S, Hasan Shaari NS, Rajikan R, Rajab NF, Golkhalkhali B, et al.
    Asian Pac J Cancer Prev, 2011;12(3):605-11.
    PMID: 21627352
    BACKGROUND: There is a paucity of information on risk factors of prostate cancer, especially those related to dietary and lifestyle among Asian populations.

    OBJECTIVE: This study aimed to determine the relationship between dietary intake (macronutrients, fruits, vegetables and lycopene), lifetime physical activity and oxidative DNA damage with prostate cancer.

    DESIGN: A case control study was carried out among 105 subjects (case n=35, control n=70), matched for age and ethnicity. Data on sociodemographic, medical, dietary intake, consumption of lycopene rich food and lifetime physical activity were obtained through an interview based questionnaire. Anthropometric measurements including weight, height and waist hip circumferences were also carried out on subjects. A total of 3 mL fasting venous blood was drawn to assess lymphocyte oxidative DNA damage using the alkaline comet assay.

    RESULTS: Cases had a significantly higher intake of fat (27.7 ± 5.5%) as compared to controls (25.1 ± 5.9%) (p < 0.05). Mean intakes of fruits and vegetables (3.11 ± 1.01 servings/d)(p < 0.05), fruits (1.23 ± 0.59 servings/d) (p<0.05) and vegetables (1.97 ± 0.94 servings/d) were higher in controls than cases (2.53 ± 1.01, 0.91 ∓ 0.69, 1.62 ± 0.82 servings/d). A total of 71% of cases did not met the recommendation of a minimum of three servings of fruits and vegetables daily, as compared to 34% of controls (p < 0.05) (adjusted OR 6.52 (95% CI 2.3-17.8)) (p < 0.05). Estimated lycopene intake among cases (2,339 ∓ 1,312 mcg/d) were lower than controls (3881 ∓ 3120 mcg/d) (p< 0.01). Estimated lycopene intake of less than 2,498 mcg/day (50th percentile) increased risk of prostate cancer by double [Adjusted OR 2.5 (95%CI 0.99-6.31)]. Intake of tomatoes, watermelon, guava, pomelo, papaya, mango, oranges, dragon fruit, carrot, tomato sauce and barbeque sauce were higher in controls compared to cases. Intake of tomato sauce of more than 2.24 g/d (25th percentile), papaya more than 22.7 g/d (50th percentile) and oranges more than 19.1g/h (50th percentile) reduced prostate cancer risk by 7.4 (Adjusted OR 7.4 (95% CI 1.17-46.8)), 2.7 (adjusted OR 2.75 (95% CI 1.03-7.39)) and 2.6 times (adjusted OR = 2.6 (95% CI=1.01-6.67)), respectively (p < 0.05 for all parameters). No oxidative damage was observed among subjects. Past history of not engaging with any physical activities at the age of 45 to 54 years old increased risk of prostate cancer by approximately three folds (Adjusted OR 2.9(95% CI = 0.8-10.8)) (p < 0.05). In conclusion, low fat diet, high intake of fruits, vegetables and lycopene rich foods and being physical active at middle age were found to be protective. Thus, it is essential for Malaysian men to consume adequate fruits and vegetables, reduce fat intake and engage in physical activity in order to reduce prostate cancer risk.

    Matched MeSH terms: Comet Assay
  9. Eshkoor S, Ismail P, Rahman S, Moin S, Adon M
    Balkan J. Med. Genet., 2013 Dec;16(2):45-52.
    PMID: 24778563 DOI: 10.2478/bjmg-2013-0031
    The ageing process is influenced by many internal and external factors. The toxic substances in the environment can cause genomic damages to cells, which increase the risk of early ageing. Furthermore, the cytochrome P450 1A2 (CYP1A2) gene polymorphism is a susceptibility factor and may enhance the risk of DNA damage in cells. The current study was carried out to show whether occupational exposure could cause genotoxicity in cells carrying the CYP1A2 gene polymorphism, thus enhancing the likelihood of early ageing. This study was conducted on mechanical workshop workers and a control group by collecting buccal cells from their mouths. Restriction fragment length polymorphism-polymerase chain reaction (RFLP-PCR) was used to identify the CYP1A2 gene polymorphism in the cells. In addition, three extra methods including micronuclei (MN) test, comet assay and real-time PCR (RT-PCR) were applied to determine the effects of gene polymorphisms on DNA damage and ageing from occupational exposure. The results showed that DNA damage in the cells carrying the mutated genotype was higher than the wild genotype. In addition, the difference in MN frequency (p = 0.001) and relative telomere length (p = 0.002) between workers and controls was significant (p <0.05) in the mutated genotype. The findings indicated a possible protective effect of gene polymorphism against early ageing, which was characterized by lack of a significant influence of CYP1A2 gene polymorphism on genetic material in the subjects (p >0.05). It was concluded that the CYP1A2 gene could be a contributing factor to prevent early ageing from occupational exposure.
    Matched MeSH terms: Comet Assay
  10. Chin SF, Hamid NA, Latiff AA, Zakaria Z, Mazlan M, Yusof YA, et al.
    Nutrition, 2008 Jan;24(1):1-10.
    PMID: 17884341
    The free radical theory of aging (FRTA) suggests that free radicals are the leading cause of deteriorating physiologic function during senescence. Free radicals attack cellular structures or molecules such as DNA resulting in various modifications to the DNA structures. Accumulation of unrepaired DNA contributes to a variety of disorders associated with the aging process.
    Matched MeSH terms: Comet Assay
  11. Inayat-Hussain SH, Wong LT, Chan KM, Rajab NF, Din LB, Harun R, et al.
    Toxicol Lett, 2009 Dec 15;191(2-3):118-22.
    PMID: 19698770 DOI: 10.1016/j.toxlet.2009.08.012
    Goniothalamin, a styryllactone, has been shown to induce cytotoxicity via apoptosis in several tumor cell lines. In this study, we have examined the potential role of several genes, which were stably transfected into T-cell lines and which regulate apoptosis in different ways, on goniothalamin-induced cell death. Overexpression of full-length receptor for activated protein C-kinase 1 (RACK-1) and pc3n3, which up-regulates endogenous RACK-1, in both Jurkat and W7.2 T cells resulted in inhibition of goniothalamin-induced cell death as assessed by MTT and clonogenic assays. However, overexpression of rFau (antisense sequence to Finkel-Biskis-Reilly murine sarcoma virus-associated ubiquitously expressed gene) in W7.2 cells did not confer resistance to goniothalamin-induced cell death. Etoposide, a clinically used cytotoxic agent, was equipotent in causing cytotoxicity in all the stable transfectants. Assessment of DNA damage by Comet assay revealed goniothalamin-induced DNA strand breaks as early as 1 h in vector control but this effect was inhibited in RACK-1 and pc3n3 stably transfected W7.2 cells. This data demonstrate that RACK-1 plays a crucial role in regulating cell death signalling pathways induced by goniothalamin.
    Matched MeSH terms: Comet Assay
  12. Siew EL, Chan KM, Williams GT, Ross D, Inayat-Hussain SH
    Free Radic. Biol. Med., 2012 Oct 15;53(8):1616-24.
    PMID: 22687461 DOI: 10.1016/j.freeradbiomed.2012.05.046
    The Fau gene (Finkel-Biskis-Reilly murine sarcoma virus (FBR-MuSV)-associated ubiquitously expressed gene) was identified as a potential tumor suppressor gene using a forward genetics approach. Downregulation of Fau by overexpression of its reverse sequence has been shown to inhibit apoptosis induced by DNA-damaging agents. To address a potential role of Fau in benzene toxicity, we investigated the apoptotic effects of hydroquinone (HQ), a major benzene metabolite, in W7.2 mouse thymoma cells transfected with either a plasmid construct expressing the antisense sequence of Fau (rfau) or the empty vector (pcDNA3.1) as a control. HQ induced apoptosis via increased production of reactive oxygen species and DNA damage, measured using dihydroethidine (HE) staining and alkaline Comet assay, respectively, in W7.2 pcDNA3.1 cells. In contrast, when Fau was downregulated by the antisense sequence in W7.2 rfau cells, HQ treatment did not cause DNA damage and oxidative stress and these cells were markedly more resistant to HQ-induced apoptosis. Further investigation revealed that there was an upregulation of NAD(P)H: quinone oxidoreductase 1 (NQO1), a detoxification enzyme for benzene-derived quinones, in W7.2 rfau cells. Compromising cellular NQO1 by use of a specific mechanism-based inhibitor (MAC 220) and NQO1 siRNA resensitized W7.2 rfau cells to HQ-induced apoptosis. Silencing of Fau in W7.2 wild-type cells resulted in increased levels of NQO1, confirming that downregulation of Fau results in NQO1 upregulation which protects against HQ-induced apoptosis.
    Matched MeSH terms: Comet Assay
  13. Tew LS, Cai MT, Lo LW, Khung YL, Chen NT
    Materials (Basel), 2018 Jul 09;11(7).
    PMID: 29987236 DOI: 10.3390/ma11071170
    Photodynamic therapy (PDT) is a cancer treatment that employs the production of cytotoxic reactive oxygen species (ROS), subsequently triggering tumor apoptosis and tumor size reduction. However, this approach suffers from insufficient light penetration depth. In order to mitigate this issue, pollen-structured gold clusters (PSGCs) were designed for mediating X-ray-induced PDT for radiotherapy enhancement. The structure of PSGCs provides a large surface area that is able to generate ROS upon X-ray irradiation. The synthesized PSGCs were exposed to different X-ray doses and the generated ROS was then quantified by dihydroethidium (DHE) assay. Furthermore, at the cellular level, the PDT efficacy of PSGCs was evaluated via immunofluorescence staining with γ-H2AX and comet assay. The results demonstrated that PSGCs possess a significantly high ROS-generating capacity and a remarkable PDT efficacy in the treatment of breast cancer cells, thus showing potential clinical uses in deep-tissue cancer treatment.
    Matched MeSH terms: Comet Assay
  14. Muhammad H, Omar MH, Rasid ENI, Suhaimi SN, Mohkiar FH, Siu LM, et al.
    Plants (Basel), 2021 Feb 11;10(2).
    PMID: 33670296 DOI: 10.3390/plants10020343
    The present study was carried out to assess the genotoxicity potential of Ficus deltoidea var. kunstleri aqueous extract (FDAE) using standard in vitro assays. The DNA damage of V79B cells was measured using the alkaline comet assay treated at 0.1 mg/mL (IC10) and 0.3 mg/mL (IC25) of FDAE together with positive and negative controls. For in vitro micronucleus assay, the V79B cells were treated with FDAE at five different concentrations (5, 2.5, 1.25, 0.625, and 0.3125 mg/mL) with and without S9 mixture. The bacteria reverse mutation assay of FDAE was performed on Salmonella typhimurium strains TA98, 100, 1535, 1537, and Escherichia coli strain WP2uvrA using pre-incubation method in the presence or in the absence of an extrinsic metabolic system (S9 mixture). FDAE at 0.1 and 0.3 mg/mL significantly increased DNA damage in both comet tail and tail moment (p < 0.05). No significant changes were detected in the number of micronucleated cell when compared to control. Tested at the doses up to 5000 µg/plate, the FDAE did not increase the number of revertant colonies for all strains. In conclusion, further investigation needs to be conducted in animal model to confirm the non-genotoxicity activities of FDAE.
    Matched MeSH terms: Comet Assay
  15. Saberbaghi T, Abbasian F, Mohd Yusof YA, Makpol S
    PMID: 23573154 DOI: 10.1155/2013/780504
    In this study, the effects of Chlorella vulgaris (CV) on replicative senescence of human diploid fibroblasts (HDFs) were investigated. Hot water extract of CV was used to treat HDFs at passages 6, 15, and 30 which represent young, presenescence, and senescence ages, respectively. The level of DNA damage was determined by comet assay while apoptosis and cell cycle profile were determined using FACSCalibur flow cytometer. Our results showed direct correlation between increased levels of damaged DNA and apoptosis with senescence in untreated HDFs (P < 0.05). Cell cycle profile showed increased population of untreated senescent cells that enter G0/G1 phase while the cell population in S phase decreased significantly (P < 0.05). Treatment with CV however caused a significant reduction in the level of damaged DNA and apoptosis in all age groups of HDFs (P < 0.05). Cell cycle analysis showed that treatment with CV increased significantly the percentage of senescent HDFs in S phase and G2/M phases but decreased the population of cells in G0/G1 phase (P < 0.05). In conclusion, hot water extract of Chlorella vulgaris effectively decreased the biomarkers of ageing, indicating its potential as an antiageing compound.
    Matched MeSH terms: Comet Assay
  16. Jubri Z, Rahim NB, Aan GJ
    Clinics (Sao Paulo), 2013 Nov;68(11):1446-54.
    PMID: 24270958 DOI: 10.6061/clinics/2013(11)11
    This study aimed to determine the effect of manuka honey on the oxidative status of middle-aged rats.
    Matched MeSH terms: Comet Assay
  17. Lim CK, Yaacob NS, Ismail Z, Halim AS
    Toxicol In Vitro, 2010 Apr;24(3):721-7.
    PMID: 20079826 DOI: 10.1016/j.tiv.2010.01.006
    Biopolymer chitosan (beta-1,4-d-glucosamine) comprises the copolymer mixture of N-acetylglucosamine and glucosamine. The natural biocompatibility and biodegradability of chitosan have recently highlighted its potential use for applications in wound management. Chemical and physical modifications of chitosan influence its biocompatibility and biodegradability, but it is unknown as to what degree. Hence, the biocompatibility of the chitosan porous skin regenerating templates (PSRT 82, 87 and 108) was determined using an in vitro toxicology model at the cellular and molecular level on primary normal human epidermal keratinocytes (pNHEK). Cytocompatibility was accessed by using a 3-[4,5-dimethyl-2-thiazolyl]-2,5-diphenyl tetrazolium bromide (MTT) assay from 24 to 72h. To assess the genotoxicity of the PSRTs, DNA damage to the pNHEK was evaluated by using the Comet assay following direct contact with the various PSRTs. Furthermore, the skin pro-inflammatory cytokines TNF-alpha and IL-8 were examined to evaluate the tendency of the PSRTs to provoke inflammatory responses. All PSRTs were found to be cytocompatible, but only PSRT 108 was capable of stimulating cell proliferation. While all of the PSRTs showed some DNA damage, PSRT 108 showed the least DNA damage followed by PSRT 87 and 82. PSRT 87 and 82 induced a higher secretion of TNF-alpha and IL-8 in the pNHEK cultures than did PSRT 108. Hence, based on our experiments, PSRT 108 is the most biocompatible wound dressing of the three tested.
    Matched MeSH terms: Comet Assay
  18. Siew EL, Rajab NF, Osman AB, Sudesh K, Inayat-Hussain SH
    J Biomed Mater Res A, 2007 May;81(2):317-25.
    PMID: 17120221
    Among the various biomaterials available for tissue engineering and therapeutic applications, microbial polyhydroxyalkanoates offer the most diverse range of thermal and mechanical properties. In this study, the biocompatibility of poly(3-hydroxybutyrate-co-4-hydroxybutyrate) [P(3HB-co-4HB); containing 50 mol % of 4-hydroxybutyrate] copolymer produced by Delftia acidovorans was evaluated. The cytotoxicity, mode of cell death, and genotoxicity of P(3HB-co-4HB) extract against V79 and L929 fibroblast cells were assessed using MTT assay, acridine orange/propidium iodide staining, and alkaline comet assay, respectively. Our results demonstrate that P(3HB-co-4HB) treated on both cell lines were comparable with clinically-used Polyglactin 910, where more than 60% of viable cells were observed following 72-h treatment at 200 mg/mL. Further morphological investigation on the mode of cell death showed an increase in apoptotic cells in a time-dependent manner in both cell lines. On the other hand, P(3HB-co-4HB) at 200 mg/mL showed no genotoxic effects as determined by alkaline comet assay following 72-h treatment. In conclusion, our study indicated that P(3HB-co-4HB) compounds showed good biocompatibility in fibroblast cells suggesting that it has potential to be used for future medical applications.
    Matched MeSH terms: Comet Assay
  19. Inayat-Hussain SH, Chan KM, Rajab NF, Din LB, Chow SC, Kizilors A, et al.
    Toxicol Lett, 2010 Mar 1;193(1):108-14.
    PMID: 20026395 DOI: 10.1016/j.toxlet.2009.12.010
    Goniothalamin (GTN) isolated from Goniothalamus sp. has been demonstrated to induce apoptosis in a variety of cancer cell lines including Jurkat T leukemia cells. However, the mechanism of GTN-induced apoptosis upstream of mitochondria is still poorly defined. In this study, GTN caused a decrease in GSH with an elevation of reactive oxygen species as early as 30 min and DNA damage as assessed by Comet assay. Analysis using topoisomerase II processing of supercoiled pBR 322 DNA showed that GTN caused DNA damage via a topoisomerase II-independent pathway suggesting that cellular oxidative stress may contribute to genotoxicity. A 12-fold increase of caspase-2 activity was observed in GTN-treated Jurkat cells after 4h treatment and this was confirmed using Western blotting. Although the caspase-2 inhibitor Z-VDVAD-FMK inhibited the proteolytic activity of caspase-2, apoptosis ensued confirming that caspase-2 activity was not crucial for GTN-induced apoptosis. However, GTN-induced apoptosis was completely abrogated by N-acetylcysteine further confirming the role of oxidative stress. Since cytochrome c release was observed as early as 1h without any appreciable change in Bcl-2 protein expression, we further investigated whether overexpression of Bcl-2 confers resistance in GTN-induced cytotoxicity. Using a panel of Jurkat Bcl-2 transfectants, GTN cytotoxicity was not abrogated in these cells. In conclusion, GTN induces DNA damage and oxidative stress resulting in apoptosis which is independent of both caspase-2 and Bcl-2.
    Matched MeSH terms: Comet Assay
  20. Chan KM, Rajab NF, Ishak MH, Ali AM, Yusoff K, Din LB, et al.
    Chem Biol Interact, 2006 Feb 1;159(2):129-40.
    PMID: 16297902
    Restenosis represents a major impediment to the success of coronary angioplasty. Abnormal proliferation of vascular smooth muscle cells (VSMCs) has been shown to be an important process in the pathogenesis of restenosis. A number of agents, particularly rapamycin and paclitaxel, have been shown to impact on this process. This study was carried out to determine the mechanisms of cytotoxicity of goniothalamin (GN) on VSMCs. Results from MTT cytotoxicity assay showed that the IC(50) for GN was 4.4 microg/ml (22 microM), which was lower compared to the clinically used rapamycin (IC(50) of 25 microg/ml [27.346 microM]). This was achieved primarily via apoptosis where up to 25.83 +/- 0.44% of apoptotic cells were detected after 72 h treatment with GN. In addition, GN demonstrated similar effects as rapamycin in inhibiting VSMCs proliferation using bromodeoxyuridine (BrdU) cell proliferation assay after 72 h treatment at IC(50) concentration (p > 0.05). In order to understand the mechanisms of GN, DNA damage detection using comet assay was determined at 2h post-treatment with GN. Our results showed that there was a concentration-dependent increase in DNA damage in VSMCs prior to cytotoxicity. Moreover, GN effects were comparable to rapamycin. In conclusion, our data show that GN initially induces DNA damage which subsequently leads to cytotoxicity primarily via apoptosis in VSMCs.
    Matched MeSH terms: Comet Assay
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