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      Scoring criteria for the evaluation of micronuclei in oral exfoliated cells

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      , 1
      South Asian Journal of Cancer
      Medknow Publications & Media Pvt Ltd

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          Abstract

          Dear Editor, With reference to the article titled, “An in vivo cytogenetic analysis of human oral squamous cell carcinoma”[1] published in your esteemed journal, I would like to bring into notice a few important points regarding the scoring criteria used for the evaluation of micronuclei. The criteria used for the evaluation of micronuclei in the above-mentioned study are as follows:[2] The diameter of micronucleus (MN) in human lymphocytes usually varies between 1/16 and 1/3 of the mean diameter of the main nuclei which corresponds to 1/256 and 1/9 of the area of one of the main nuclei in a BN cell, respectively MN is round or oval MN is nonrefractile and it can, therefore, be readily distinguished from artifacts such as staining particles MN is not linked or connected to the main nuclei MN may touch but will not overlap the main nuclei, and the micronuclear boundary should be distinguishable from the nuclear boundary MN usually has the same staining intensity as the main nuclei but occasionally staining may be more intense. First of all, the above-said criteria are for the evaluation of micronuclei in lymphocytes and not exfoliated oral cells (as in the given study, micronuclei were evaluated in exfoliated oral cells). Second, the micronuclei showed in the images provided do not comply by the criteria used (micronuclei shown in the image are refractile and overlapping). The most accepted criterion[3] for the evaluation of micronuclei in oral exfoliated cells is as follows: Rounded smooth perimeter suggestive of a membrane Less than one-third of the diameter of the associated nucleus, but large enough to discern the shape and color Staining intensity similar to that of the nucleus Texture similar to that of the nucleus Same focal plane as the nucleus Absence of overlap with, or bridge to, the nucleus. Since there is a considerable similarity between the two criteria, using the former criterion is technically not wrong, provided it is fulfilled completely. To conclude, I request the authors to go through their images again and make sure they are compatible with the criterion used. The purpose of this letter is not to highlight an error rather to focus on the appropriate criterion to be used for future studies. Such parameters play a vital role in establishing the validity of the study. Financial support and sponsorship Nil. Conflicts of interest There are no conflicts of interest.

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          The micronucleus assay in human buccal cells as a tool for biomonitoring DNA damage: the HUMN project perspective on current status and knowledge gaps.

          The micronucleus (MN) assay in exfoliated buccal cells is a useful and minimally invasive method for monitoring genetic damage in humans. This overview has concluded that although MN assay in buccal cells has been used since the 1980s to demonstrate cytogenetic effects of environmental and occupational exposures, lifestyle factors, dietary deficiencies, and different diseases, important knowledge gaps remain about the characteristics of micronuclei and other nuclear abnormalities, the basic biology explaining the appearance of various cell types in buccal mucosa samples and effects of diverse staining procedures and scoring criteria in laboratories around the world. To address these uncertainties, the human micronucleus project (HUMN; see http://www.humn.org) has initiated a new international validation project for the buccal cell MN assay similar to that previously performed using human lymphocytes. Future research should explore sources of variability in the assay (e.g. between laboratories and scorers, as well as inter- and intra-individual differences in subjects), and resolve key technical issues, such as the method of buccal cell staining, optimal criteria for classification of normal and degenerated cells and for scoring micronuclei and other abnormalities. The harmonization and standardization of the buccal MN assay will allow more reliable comparison of the data among human populations and laboratories, evaluation of the assay's performance, and consolidation of its world-wide use for biomonitoring of DNA damage.
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            HUMN project: detailed description of the scoring criteria for the cytokinesis-block micronucleus assay using isolated human lymphocyte cultures.

            Criteria for scoring micronuclei and nucleoplasmic bridges in binucleated cells in the cytokinesis-block micronucleus assay for isolated human lymphocyte cultures are described in detail. Morphological characteristics of mononucleated cells, binucleated cells, and multinucleated cells as well as necrotic and apoptotic cells and nuclear buds are also described. These criteria are illustrated by a series of schematic diagrams as well as a comprehensive set of colour photographs that are of practical assistance during the scoring of slides. These scoring criteria, diagrams and photographs have been used in a HUman MicronNucleus (HUMN) project inter-laboratory slide-scoring exercise to evaluate the extent of variability that can be attributable to individual scorers and individual laboratories when measuring the frequency of micronuclei and nucleoplasmic bridges in binucleated cells as well as the nuclear division index. The results of the latter study are described in an accompanying paper. It is expected that these scoring criteria will assist in the development of a procedure for calibrating scorers and laboratories so that results from different laboratories for the cytokinesis-block micronucleus assay may be more comparable in the future.
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              An in vivo cytogenetic analysis of human oral squamous cell carcinoma

              Background: Oral cancer ranks in the top three of all cancers in India, which accounts for over 30% of all cancers reported in the country. The micronucleus test (MNT) is one of the most widely applied short term tests used in genetic toxicology to evaluate the mutagenicity and carcinogenicity. Aims: The present study aims at an in vivo cytogenetic analysis of human oral squamous cell carcinoma and to assess the applicability of MNT in diagnosing early detection of oral carcinoma. Materials and Methods: Exfoliated scrape smears were collected from the clinically diagnosed 136 patients suffering from oral precancerous and cancerous lesions. The wet fixed smears were stained by adopting Papanicolaou's staining protocol and counter-stained with Giemsa's solution. Results: The frequency of micronucleated cells has been observed to be in increasing order with the increase of the age-groups and from control to precancerous to cancerous cases significantly in both sexes. Conclusion: Micronucleus formation in the oral mucosa could be a biomarker of genetic damage and also a potential onco-indicator in the long run of oral carcinogenesis. Therefore, MNT can be applied for the early detection of oral carcinoma in the human being.
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                Author and article information

                Journal
                South Asian J Cancer
                South Asian J Cancer
                SAJC
                South Asian Journal of Cancer
                Medknow Publications & Media Pvt Ltd (India )
                2278-330X
                2278-4306
                Apr-Jun 2017
                : 6
                : 2
                : 89
                Affiliations
                [1]Department of Oral and Maxillofacial Pathology, Christian Dental College, CMC, Ludhiana, Punjab, India
                [1 ]Department of Oral and Maxillofacial Pathology, Bapuji Dental College and Hospital, Davangere, Karnataka, India
                Author notes
                Correspondence to: Dr. Sonal Grover, E-mail: sonalgrvr@ 123456yahoo.com
                Article
                SAJC-6-89a
                10.4103/2278-330X.208842
                5506822
                64ec33c7-4ba6-45d7-b124-b07606ebf39f
                Copyright: © 2017 The South Asian Journal of Cancer

                This is an open access article distributed under the terms of the Creative Commons Attribution-NonCommercial-ShareAlike 3.0 License, which allows others to remix, tweak, and build upon the work non-commercially, as long as the author is credited and the new creations are licensed under the identical terms.

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