Kanserli hücre hatları, pasaj sayısı arttıkça genomik organizasyonunu ve karyotipini değiştirir: sitogenetik bir çalışma

Year 2018, Volume: 43 Issue: 4, 923 - 930, 29.12.2018

M. Bertan Yılmaz Erdal Tunç , N. Seda Ilgaz , Hale Öksüz Ezgi Öztecik Lütfiye Özpak İşıl Öcal , Ayfer Pazarbaşı , Osman Demirhan

https://doi.org/10.17826/cumj.375325

Abstract

Amaç: İnsan sağlığına ilişkin bilimsel araştırmalarda
memelilerin sınırlı kullanımı yeni araştırma stratejilerinin geliştirilmesine
yol açmıştır. Bunlardan birisi memeli hücre kültürü tekniğidir. Piyasada
bulunan, sitogenetik ve biyokimyasal belirteçlerle iyi karakterize edilmiş
kanser hücre hatları farklı laboratuvarlar arasındaki sonuçların
karşılaştırılmasına olanak sağlar. Bununla birlikte, bu hücre dizileri,
kültürde uzun bir süre muhafaza edildiklerinden, önceki pasajlarda tanımlanmış
veya saptanmamış, hücre hatlarının özelliklerini ve ajanlara karşı tepkilerini
değiştirebilecek mutasyonlar meydana gelebilir. Burada sitogenetik olarak, altı
farklı hücre hattının tekrarlanan hücre kültürlerinde yeni kromozomal
düzenlenmeleri araştırdık.

Metod: MCF7, HCT116, A549, SHSY5Y, HEPG2 ve NIH3T3 hücre
hatları, %10 FBS ve %1 penisilin-streptomisin içeren DMEM besiyerinde kültüre
edildi. Metafaz kromozomlarının analizi için GTG bantlama prosedürü kullanıldı
ve en az 20 metafaz analiz edildi.

Bulgular: İncelenen hücre kültürlerinde pasaj sayısı
artışıyla kromozom sayı varyasyonları ve yapısal değişiklikler tespit ettik.

Sonuç: İlaçları test etmek, moleküler mekanizmaları
tanımlamak, çevresel etkileri anlamak için hücre hatları uzun süredir
araştırmalarda kullanılmaktadır. Bir hücre dizisinin en önemli özelliği, konak
organizmasıyla olan genotip ve karyotip benzerlikleridir. Kanser Hücresi
soyları, genomik/kromozomal dengesizliğe sahiptir ve bu da her pasajda fenotip
değişimi ile birlikte karyotip değişimine neden olabilmaktedir. Bu nedenle, hücre
kültürlerinin kullanılacağı bir araştırma projesine başlamadan muhakkak
kullanılan hücre hattının karyotipini doğrulamak gerekmektedir.

Keywords

Kanserli hücre hattı, sitogenetik analiz, MCF7, HCT116, A549, SHSY5Y, HEPG2, NIH3T3

Cancerous cell lines alter their genomic organization and karyotype with increased passage number: a cytogenetic study

Abstract

Purpose: The limited use of mammals in human health related
scientific research has led to the development of new research strategies like
cell culture techniques. Commercially available cancerous cell lines that
are well characterized by cytogenetics and biochemical markers allow comparison
of results among different laboratories. However, as these cell lines tend to
be maintained in culture over long periods of time, mutations can occur that
may change characteristics and responses of cell lines that have initially been
identified or non-existed at earlier passages. Here we cytogenetically
investigated the chromosomal rearrangements in repeated cultures of six
different cell lines over continuous passages.

Method: MCF7, HCT116,
A549, SHSY5Y, HEPG2, and NIH3T3 cell lines were cultured in DMEM containing 10%
FBS and 1% penicillin-streptomycin.  GTG
banding procedure was used for the analysis of metaphase chromosomes, at least
20 metaphases were analyzed per cell line.

Results: We found chromosome
number variations and structural changes in the all examined cell cultures as
the passage numbers increase.

Conclusion: Cell
lines have long been used in research to test drugs, to delineate molecular
mechanisms, to understand the environmental effects and so on. The most
important feature of a cell line is its genotype and karyotype similarities
with their host organism. Cancer Cell lines, possess genomic/chromosomal instability that also lead them to change
their phenotype along with their karyotype from one passage to next. Therefore,
it is always best to verify karyotype before employing a specific cell line in
a research project.

Keywords

Cancerous Cell line, cytogenetic analysis, MCF7, HCT116, A549, SHSY5Y, HEPG2, NIH3T3

References

• 1. Kytölä S, Rummukainen J, Nordgren A, Karhu R, Farnebo F, Isola J, et al. Chromosomal alterations in 15 breast cancer cell lines by comparative genomic hybridization and spectral karyotyping. Genes Chromosomes Cancer. 2000 Jul;28(3):308-17.
• 2. Macleod RA, Kaufmann M, Drexler HG. Cytogenetic analysis of cancer cell lines. Methods Mol Biol. 2011;731:57-78
• 3. Wenger SL, Senft JR, Sargent LM, Bamezai R, Bairwa N, Grant SG. Comparison of Established Cell Lines at Different Passages by Karyotype and Comparative Genomic Hybridization. Biosci Rep. 2004 Dec;24(6):631-9.
• 4. https://www.phe-culturecollections.org.uk/media/130237/mcf7-cell-line-profile.pdf
• 5. https://www.lgcstandards-atcc.org/Products/All/HTB-22.aspx?geo_country=tr#characteristics.
• 6. Rondón-Lagos M, Verdun Di Cantogno L, Marchiò C, Rangel N, Payan-Gomez C, Gugliotta P, et al. Differences and homologies of chromosomal alterations within and between breast cancer cell lines: a clustering analysis. Mol Cytogenet. 2014 Jan 23;7(1):8.
• 7. https://www.lgcstandards-atcc.org/products/all/CCL-247.aspx?geo_country=tr#general information
• 8. Knutsen T, Padilla-Nash HM, Wangsa D, Barenboim-Stapleton L, Camps J, McNeil N, et al. Definitive molecular cytogenetic characterization of 15 colorectal cancer cell lines. Genes Chromosomes Cancer. 2010 Mar;49(3):204-23.
• 9. https://www.lgcstandards-atcc.org/Products/All/CCL-185.aspx?geo_country=tr#characteristics
• 10. Peng KJ, Wang JH, Su WT, Wang XC, Yang FT, Nie WH. Characterization of two human lung adenocarcinoma cell lines by reciprocal chromosome painting. Dongwuxue Yanjiu. 2010 Apr;31(2):113-21.
• 11. Luk C, Tsao MS, Bayani J, Shepherd F, Squire JA. Molecular cytogenetic analysis of non-small cell lung carcinoma by spectral karyotyping and comparative genomic hybridization. Cancer Genet Cytogenet. 2001 Mar;125(2):87-99.
• 12. https://www.mskcc.org/research-advantage/support/technology/tangible-material/human-neuroblastoma-cell-line-sh-sy5y.
• 13. https://www.lgcstandards-atcc.org/Products/Cells_and_Microorganisms/By_Tissue/Bone_ Marrow/CRL-2266.aspx?geo_country=tr#characteristics
• 14. Kim GJ, Park SY, Kim H, Chun YH, Park SH. Chromosomal aberrations in neuroblastoma cell lines identified by cross species color banding and chromosome painting. Cancer Genet Cytogenet. 2001 Aug;129(1):10-6.
• 15. https://www.lgcstandards-atcc.org/products/all/HB-8065.aspx?geo_country=tr# characteristics
• 16. http://www.hepg2.com/
• 17. Leibiger C, Kosyakova N, Mkrtchyan H, Glei M, Trifonov V, Liehr T First molecular cytogenetic high resolution characterization of the NIH 3T3 cell line by murine multicolor banding.. J Histochem Cytochem. 2013 Apr;61(4):306-12.
• 18. http://www.nih3t3.com/
• 19. https://www.ncbi.nlm.nih.gov/pubmed/26026906
• 20. Heinrich F, Contioso VB, Stein VM, Carlson R, Tipold A, Ulrich R, et al. Passage-dependent morphological and phenotypical changes of a canine histiocytic sarcoma cell line (DH82 cells). Vet Immunol Immunopathol. 2015 Jan 15;163(1-2):86-92.
• 21. Pelliccia F, Ubertini V, Bosco N. The importance of molecular cytogenetic analysis prior to using cell lines in research: The case of the KG-1a leukemia cell line. Oncol Lett. 2012 Aug;4(2):237-240.
• 22. Horbach SPJM, Halffman W. The ghosts of HeLa: How cell line misidentification contaminates the scientific literature. PLoS One. 2017 Oct 12;12(10):e0186281.