GENO-SİTOTOKSİSİTE ÇALIŞMALARINA SİTOM YAKLAŞIMI

Yıl 2019, , 261 - 269, 31.07.2019

Ceren Börçek Kasurka

https://doi.org/10.18036/estubtdc.599209

Öz

ÖZET

 

DNA hasarının
kromozom seviyesinde çalışılması, genetik toksikoloji araştırmalarının temel
dayanaklarındandır. Mikronükleus yöntemi, kromozom hasarını değerlendirmek
amacıyla geliştirilen yöntemlerden biridir. Mikronükleuslar; sentromeri olmayan
kromozom veya kromatid fragmentlerinden ya da anafazda geç kalıp kutuplara göç
edemeyen kromozom veya kromatidlerden oluşan ve telofazda hücre zarı ile
kuşatılarak kardeş hücre çekirdeklerine dahil olmadan sitoplazmada kalan,
çekirdek dışı küçük cisimciklerdir. Bu yöntemde mikronükleusların yanısıra
nükleoplazmik köprüler (NPK) ve nükleer tomurcuklar (NT) değerlendirilebilmekte
ve hücre ölümü ile nükleer bölünme indeksi de ölçülebilmektedir. Disentrik
kromozomların sentromerlerinin farklı kutuplara göç etmesinden orjinlenen NPK,
DNA’nın yanlış onarımının; kromozom rearanjmanının, telomer uç birleşmesinin; NT
ise gen amplifikasyonu ve/veya değişmiş gen dozajının göstergesi olarak
sayılmaktadır. Ayrıca her hücrenin sahip olduğu çekirdek sayısının
değerlendirilerek nükleer bölünme indeksinin hesaplanmasının yanısıra nekrotik
ve apoptotik hücrelerin de belirlenebilmesi mitotik aktivite ve
sitotoksisitenin tayinine olanak sağlamaktadır. Analiz edilebilen tüm bu
parametreler mikronükleus yöntemini genetik kusurların, beslenme
yetersizliklerinin veya ekzojen kaynaklı genotoksinlerin sebep olduğu kromozomal
instabilite fenotipinin ve değişmiş hücresel canlılığın kapsamlı olarak
ölçülebildiği “sitom” yöntemi haline getirmiştir. Bu derlemede mikronükleus
yönteminin genotoksisite çalışmalarındaki yeri ve önemi ile sitom yaklaşımının
bu yönteme kattığı yeniliklere değinilmiştir.

Anahtar Kelimeler

Mikronükleus, Sitom, Genotoksisite, Sitotoksisite

CYTOM APPROACH TO GENO-CYTOTOXICITY STUDIES

Öz

ABSTRACT

 

Working at the chromosomal level of DNA
damage is the mainstay of the genetic toxicology researches. The micronucleus assay is one of the developed methods
to evaluate chromosomal damage. Micronuclei
are non-centromeric chromosomes/chromatids fragments, or chromosomes/chromatids
which lagging in the anaphase and cannot migrate to the poles, located in the
cytoplasm without being included in the sister cell nuclei and surrounded by
cell membranes in telophase. In this method, besides micronuclei, nucleoplasmic
bridges (NPBs) and nuclear buds (NBUDs) can be evaluated, cell death and
nuclear division index (NDI) can be measured. NPBs originated from the
centromeres of dicentric chromosomes
migrating to different poles, are indicative
of DNA mis-repair, chromosome
rearrangement, telomere end-fusions. NBUDs are considered as markers of gene amplification and/or altered
gene dosage. In addition to NDI, establishing to necrotic and apoptotic cells
may allow the determination of mitotic activity and cytotoxicity. All these
analysed parameters have transformed micronucleus method into "cytome" assay by which chromosomal
instability phenotypes and altered cellular viability caused by genetic
defects, nutritional deficiencies, genotoxins from exogenous sources can be
measured comprehensively.
In this review, the place and importance of micronucleus method in genotoxicity
studies and the innovations caused by the cytome approach are mentioned.

Anahtar Kelimeler

Micronucleus, Cytome, Genotoxicity, Cytotoxicity

Kaynakça

• [1] Riss TL, Moravec RA, Niles AL. Cytotoxicity testing: measuring viable cells, dead cells, and detecting mechanism of cell death. Methods Mol Biol. 2011; 740:103-14.
• [2] Atlı Şekeroğlu Z, Şekeroğlu V. Genetik toksisite testleri. TÜBAV Bilim Dergisi, 2011;4(3):221-9.
• [3] Eastmond DA, Hartwig A, Anderson D, Anwar WA, Cimino MC, Dobrev I, et al. Mutagenicity testing for chemical risk assessment: update of the WHO/IPCS Harmonized Scheme. Mutagenesis, 2009; 24(4):341-9.
• [4] Tokur O, Aksoy A. In Vitro Sitotoksisite Testleri, 2017; 112-8 p.
• [5] Jena GB, Kaul CL, Ramarao P. Genotoxicity Testing, A Regulatory Requirement For Drug Discovery And Development: Impact Of Ich Guidelines. Indian Journal of Pharmacology, 2002; 34:86-99.
• [6] Mark H, Naram R, Pham T, Shah K, Cousens LP, Wiersch C, et al. A practical cytogenetic protocol for in vitro cytotoxicity and genotoxicity testing. Annals of Clinical & Laboratory Science, 1994; 24(5):387-95.
• [7] Fenech M. The cytokinesis-block micronucleus technique: A detailed description of the method and its application to genotoxicity studies in human populations. Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis, 1993; 285(1):35-44.
• [8] Cho NY, Kim KW, Kim KK. Genomic health status assessed by a cytokinesis-block micronucleus cytome assay in a healthy middle-aged Korean population. Mutat Res. 2017; 814:7-13.
• [9] Knasmüller S, Nersesyan A, Mišík M, Gerner C, Mikulits W, Ehrlich V, et al. Use of conventional and -omics based methods for health claims of dietary antioxidants: a critical overview. British Journal of Nutrition, 2008; 99(E-S1):ES3-ES52.
• [10] Kocaman AY, Rencüzoğulları E, Topaktaş M. In vitro investigation of the genotoxic and cytotoxic effects of thiacloprid in cultured human peripheral blood lymphocytes. Environmental toxicology, 2014; 29(6):631-41.
• [11] Tucker JD, Preston RJ. Chromosome aberrations, micronuclei, aneuploidy, sister chromatid exchanges, and cancer risk assessment. Mutation Research/Reviews in Genetic Toxicology, 1996; 365(1-3):147-59.
• [12] Mateuca R, Lombaert N, Aka PV, Decordier I, Kirsch-Volders M. Chromosomal changes: induction, detection methods and applicability in human biomonitoring. Biochimie, 2006; 88(11):1515-31.
• [13] Evans HJ, Neary GJ, Williamson FS. The Relative Biological Efficiency of Single Doses of Fast Neutrons and Gamma-rays on Vicia Faba Roots and the Effect of Oxygen. International Journal of Radiation Biology and Related Studies in Physics, Chemistry and Medicine, 1959; 1(3):216-29.
• [14] Fenech M. The in vitro micronucleus technique. Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis. 2000; 455(1–2):81-95.
• [15] Thoday J. The Effect of Ionizing Radiations on the Broad Bean Root—Part IX. The British journal of radiology, 1951; 24(286):572-6.
• [16] Schmid W. The micronucleus test. Mutation Research/Environmental Mutagenesis and Related Subjects, 1975; 31(1):9-15.
• [17] Heddle JA. A rapid in vivo test for chromosomal damage. Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis, 1973; 18(2):187-90.
• [18] Fenech M, Kirsch-Volders M, Natarajan AT, Surralles J, Crott JW, Parry J, et al. Molecular mechanisms of micronucleus, nucleoplasmic bridge and nuclear bud formation in mammalian and human cells. Mutagenesis, 2011; 26(1):125-32.
• [19] Coskun M, Cayir A, Coskun M, Tok H. Evaluation of background DNA damage in a Turkish population measured by means of the cytokinesis-block micronucleus cytome assay. Mutat Res. 2013; 757(1):23-7.
• [20] Fenech M, Holland N, Chang WP, Zeiger E, Bonassi S. The HUman MicroNucleus Project—An international collaborative study on the use of the micronucleus technique for measuring DNA damage in humans. Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis. 1999; 428(1–2):271-83.
• [21] Duan H, Leng S, Pan Z, Dai Y, Niu Y, Huang C, et al. Biomarkers measured by cytokinesis-block micronucleus cytome assay for evaluating genetic damages induced by polycyclic aromatic hydrocarbons. Mutation Research/Genetic Toxicology and Environmental Mutagenesis. 2009; 677(1-2):93-9.
• [22] Fenech M, Morley AA. Measurement of micronuclei in lymphocytes. Mutation Research/Environmental Mutagenesis and Related Subjects. 1985; 147(1):29-36.
• [23] Fenech M. Cytokinesis-block micronucleus assay evolves into a "cytome" assay of chromosomal instability, mitotic dysfunction and cell death. Mutat Res. 2006; 600(1-2):58-66.
• [24] Donmez-Altuntas H, Bitgen N. Evaluation of the genotoxicity and cytotoxicity in the general population in Turkey by use of the cytokinesis-block micronucleus cytome assay. Mutat Res. 2012;748(1-2):1-7.
• [25] Üstüner D. Kromozom kırıkları ve mikronükleus-apoptoz. TÜBAV Bilim Dergisi, 2011; 4(1).
• [26] OECD. Test No. 487: In Vitro Mammalian Cell Micronucleus Test. Paris: OECD Publishing; 2016.