Monitoring of buccal epithelial cells by alkaline comet assay (single cell gel electrophoresis technique) in cytogenetic evaluation of chlorhexidine

Eren, Kaya; Özmeriç, Nurdan; Şardaş, Semra

doi:10.1007/s00784-002-0168-1

Cited by 66 publications

(46 citation statements)

References 30 publications

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“…Our results have demonstrated that Periogard® containing chlorhexidine at 0.12% was able to induce cytogenetic damage as depicted by the increase of micronucleus frequency after continuous exposure. By comparison, a level of DNA damage in peripheral blood lymphocytes and buccal mucosa cells was evidenced in individuals exposed to chlorhexidine digluconate at 0.12% for 2 weeks by single-cell gel (comet) assay [19]. In particular, a previous study conducted by our research groups has consistently demonstrated that Periogard® induced genetic damage in peripheral lymphocytes and buccal mucosa cells of rats [28].…”

Section: Discussionmentioning

confidence: 96%

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Cytogenetic damage induced by mouthrinses formulations in vivo and in vitro

et al. 2011

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The aim of the present study was to comparatively evaluate DNA damage and cellular death in cells exposed to various commercially available mouthrinses: Listerine Cepacol, Plax alcohol free, Periogard, and Plax Whitening. A total of 75 volunteers were included in the search distributed into five groups containing 15 people each for in vivo study. Exfoliated buccal mucosa cells were collected immediately before mouthrinse exposure and after 2 weeks. Furthermore, blood samples were obtained from three healthy donors for in vitro study. The micronucleus test was used to evaluate mutagenicity and cytotoxicity in vivo. The single-cell gel (comet) assay was used to determine DNA damage in vitro. After 2 weeks exposure, Periogard showed 1.8% of micronucleated cells with significant statistical differences (p < 0.05) compared to before exposure (0.27%). Plax Whitening presented high tail moment value (4.5) when compared to negative control (0.6). The addition of all mouthrinses to cells incubated with methyl methanesulfonate did not alter the number of strand breaks in the genetic material. Listerine was able to reduce genetic damage induced by hydrogen peroxide because a decrease of tail moment was noticed. The results of the present study suggest that Periogard and Plax Whitening can induce genetic damage, whereas Listerine is an antioxidant agent. Since DNA damage is considered to be prime mechanism during chemical carcinogenesis, these data may be relevant in risk assessment for protecting human health and preventing carcinogenesis.

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Section: Discussionmentioning

confidence: 96%

“…Test materials, manufacturers, and ingredients are listed in Table 1. All volunteers were requested to rinse their mouths with 15 ml of mouthrinse twice a day for 2 weeks as described elsewhere [19]. The rinsing procedures took approximately 30 s each time.…”

Section: Micronucleus Test In Buccal Mucosa Cellsmentioning

confidence: 99%

Cytogenetic damage induced by mouthrinses formulations in vivo and in vitro

et al. 2011

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“…In this context, aside from lymphocytes, the assay has been carried out using whole blood [ 31 ], buccal epithelial cells [ 32 ], tear duct epithelial cells [ 33 ], and sperm [ 34 ]. The selection of the cell population to use must be based on exposure conditions, persistence of DNA damage, and rates of cell turnover.…”

Section: Sample Type Collection and Processingmentioning

confidence: 99%

The Comet Assay In Vivo in Humans

Costa

Teixeira

2014

Genotoxicity and DNA Repair

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In the last decades, the comet assay has been used as a molecular biomarker to detect DNA damage in several human biomonitoring studies. Nevertheless, many of these studies are often inconclusive or offer poor quality data due to study design constraints. This chapter describes not only the experimental protocol to carry the alkaline version of the comet assay in human cells but also considers many of the obstacles that the researcher faces while carrying a human biomonitoring study, such as sample size, sample type, sample collection, statistics, and result communication. Understanding assay limitations, both experimental and biological, is essential to improve both data quality and data relevance obtained and to guarantee that comet assay continues to provide enhanced reliability as a biomarker in human biomonitoring studies.Key words Comet assay , Human biomonitoring , Exposure , Lymphocytes , Whole blood Comet Assay for Human BiomonitoringComet assay (also designated of single cell gel electrophoresis) is a rapid, simple, visual, and sensitive method to detect DNA breakage in eukaryotic cells [ 1 ].Briefl y, cells are embedded in agarose on a microscope slide, lysed with detergent to remove cell and nuclear membranes, and treated with a high salt solution. Nucleoids are formed, containing non-nucleosomal but still supercoiled DNA. Any breaks present in the DNA cause the supercoiling to relax locally and loops of DNA are then free to extend toward the anode during electrophoresis. At the end of this procedure, damaged cells will resemble comets (hence the name comet assay ) while non-damaged cells maintain their round appearance. Comets are viewed by fl uorescence microscopy after staining with a suitable fl uorescent DNA-binding dye [ 2 ].The pH of unwinding solution is responsible for differences in method sensitivity, as the conversion of alkali-labile sites to strand breaks is a process dependent on the pH and time of alkaline treatment. Under alkaline (pH > 13) conditions, the assay potentially

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“…Although SCGE studies have also been conducted on BM cells [Rojas et al, 1996;Valverde et al, 1997;Eren et al, 2002;Dhillon et al, 2004], low cell viability, saprophyte bacterial contamination, and their specialized cellular membranes, which make cell lysis difficult, contribute to making BM cells a more complicated cell to analyze compared with leukocytes. In this study, we evaluated the properties of BM cells in the SCGE assay and developed methodology for performing adequate cell lysis and enriching for viable cells in exfoliated BM cell samples.…”

Section: Introductionmentioning

confidence: 99%

Viable human buccal mucosa cells do not yield typical nucleoids: Impacts on the single-cell gel electrophoresis/comet assay

Pinhal

Gontijo

Reyes

et al. 2006

Environ. Mol. Mutagen.

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Buccal mucosa (BM) cells have been used in human biomonitoring studies for detecting DNA adducts and chromosomal damage in an epithelial cell population. In the present study, we have investigated if human BM cells are suitable for use in the single-cell gel electrophoresis (SCGE)/Comet assay as an approach for estimating the exposure of epithelial cells to DNA-damaging agents. Our results indicate that only a few cells from BM cell samples yield comets that can be analyzed by current methods, and that the yield of cells with comets is independent of the percentage of viable BM cells in the sample. Data generated after enzymatic enrichment of viable cells and immunomagnetic separation of epithelial cells suggest that most of the BM cells that do form comets are probably leukocytes. Moreover, by reevaluating specific cells after running the Comet assay, we found that viable epithelial BM cells give rise to atypical comets that are not included in the analysis. Comparing DNA migration patterns between small groups of smokers and nonsmokers indicated that long-term smoking had no effect on the subpopulation of cells that yield typical comets. Our results indicate that the SCGE assay, as it is commonly performed, may not be useful for genotoxicity monitoring in human epithelial BM cells.

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Monitoring of buccal epithelial cells by alkaline comet assay (single cell gel electrophoresis technique) in cytogenetic evaluation of chlorhexidine

Cited by 66 publications

References 30 publications

Cytogenetic damage induced by mouthrinses formulations in vivo and in vitro

Cytogenetic damage induced by mouthrinses formulations in vivo and in vitro

The Comet Assay In Vivo in Humans

Viable human buccal mucosa cells do not yield typical nucleoids: Impacts on the single-cell gel electrophoresis/comet assay

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