Genomic instability of micronucleated cells revealed by single-cell comparative genomic hybridization.
Nuclear variation in size and shape and genomic instability are hallmarks of dedifferentiated cancer cells. Although micronuclei are a typical long-term consequence of DNA damage, their contribution to chromosomal instability and clonal diversity in cancer disease is unclear. We isolated cancer cells with or without micronuclei to perform genomic analysis. Cell suspensions of HT1080 fibrosarcoma cells from either 2D culture or after isolation from 3D collagen matrix culture were stained with Hoechst 33342 and after classification for presence or absence of a micronucleus via bright-field and epifluorescence microscopy, cells were individually aspirated with a micropipette. Subsequently, whole-genome amplification and single-cell comparative genomic hybridization (CGH) were applied to detect genomic aberrations. The data show a high-fidelity isolation and genome amplification that lacks adverse effects by prior Hoechst 33342 staining. HT1080 cells showed a high degree of divergent amplifications, but neither location nor frequency of aberrations was dependent on 2D or 3D culture conditions or micronucleation. Thus, single-cell selection of defined nuclear states is amenable to single-cell CGH and here provides first insight into the aberration drift and genomic diversity in cancer cells with and without micronuclei.
Imle
Andrea
A
Rudolf Virchow Center for Experimental Biomedicine, Department of Dermatology, Universtiy Würzburg, Germany.
Polzer
Bernhard
B
Alexander
Stephanie
S
Klein
Christoph A
CA
Friedl
Peter
P
D045744:Cell Line, Tumor; D055028:Comparative Genomic Hybridization / Q000379:methods*; D005354:Fibrosarcoma / Q000235:genetics* / Q000473:pathology; D042822:Genomic Instability / Q000235:genetics / Q000502:physiology*; D006801:Humans; D007621:Karyotyping; D048629:Micronuclei, Chromosome-Defective; D020411:Oligonucleotide Array Sequence Analysis