A strategy combining flow sorting and comparative genomic hybridization for studying genetic aberrations at different stages of colorectal tumorigenesis in ulcerative colitis.
BACKGROUND
DNA aneuploidy has been shown to increase the risk of developing dysplasia in ulcerative colitis (UC) and is related to tumorigenesis in the colorectum. Therefore, it is of particular interest to study genetic aberrations behind DNA aneuploidization during colorectal carcinogenesis. We wanted to elucidate further the relationship between mucosal morphology and DNA aberrations in UC.


METHODS
DNA flow cytometry was applied to multiple lesions including regenerative, dysplastic, and carcinomatous mucosa from the colectomy specimen of a male patient with long-standing UC. The lesions harbored multiple DNA aneuploid stemlines that were subjected to flow sorting. We analyzed gene alterations by degenerate oligonucleotide primer (DOP; universal primers) polymerase chain reaction (PCR)-based comparative genomic hybridization (CGH) and fluorescent in situ hybridization (FISH) in diploid and aneuploid sorted cells.


RESULTS
DOP-PCR-based CGH shows gains and losses that can be verified by FISH. We show that with this approach one can study genetic evolution of distinct DNA diploid and aberrant subpopulations through defined stages of colorectal tumorigenesis. This includes getting information related to tumor heterogeneity that cannot be obtained by CGH with DNA extracted from nonsorted cell populations. Genetic imbalance was also detected in diploid nondysplastic flow-sorted mucosal cells from the same bowel.


CONCLUSIONS
Similar gains and losses were found in aneuploid dysplasias and carcinomas at widely separated locations in the same bowel, indicating a common selection pressure in different areas of the same bowel. The common aberrations may be of importance for progression from dysplasia to carcinoma.
Clausen
O P
OP
Institute of Pathology, The National Hospital, University of Oslo, Oslo, Norway. ole.petter.clausen@rikshospitalet.no
Andersen
S N
SN
Strøomkjaer
H
H
Nielsen
V
V
Rognum
T O
TO
Bolund
L
L
Køolvraa
S
S
D000368:Aged; D000782:Aneuploidy; D002869:Chromosome Aberrations / Q000235:genetics*; D003093:Colitis, Ulcerative / Q000150:complications* / Q000235:genetics* / Q000473:pathology; D015179:Colorectal Neoplasms / Q000150:complications* / Q000235:genetics* / Q000473:pathology; D004247:DNA / Q000032:analysis / Q000235:genetics; D004171:Diploidy; D018450:Disease Progression; D005434:Flow Cytometry / Q000379:methods*; D016678:Genome; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D007413:Intestinal Mucosa / Q000166:cytology / Q000473:pathology; D007621:Karyotyping; D008297:Male; D009693:Nucleic Acid Hybridization / Q000379:methods*; D016133:Polymerase Chain Reaction
D004247:DNA