Microdissection and SAGE as a combined tool to reveal gene expression in ductal carcinoma in situ of the breast.
The interplay between cancer cells and the normal surrounding tissue is believed to influence the biological behavior of the tumor. However, the presence of multiple cell types within the prelevated tumor specimen may attenuate changes that occur specifically in the malignant cells within their microenvironment. To study gene expression of the malignant cells in situ, we used a new microdissection method to separate ductal carcinoma in situ (DCIS) cells from the surrounding stroma, immunological infiltrates, and endothelial cells. We applied an adapted microSAGE protocol, without total mRNA amplification, to study their gene expression profile. Three thousand two hundred one different transcripts were identified in a total of 29 534 observed tags. Of these unique tags, 88.3% matched known GenBank sequences and 11.7% represented unknown transcripts. As compared to a total DCIS SAGE library, microdissection combined with SAGE revealed additional genes expressed only in normal surrounding, probably stromal, cells and not or significantly less in DCIS tumor cells. This study demonstrates that microdissection can be combined with SAGE as a tool to study transcriptomes. This approach provides important new information on differential gene expression both in tumor cells and normal surrounding tissue. Several of the observed differences indeed disappear when the total tumor mass is analyzed.
Biomedisch Onderzoeksinstituut, Limburgs Universitair Centrum, School for Life Sciences, Transnational University Limburg, Universitaire Campus, Diepenbeek, Belgium.
Ilse
I
Verlinden
Jaak
J
Janssens
Jef
J
Raus
Luc
L
Michiels
D000328:Adult; D001943:Breast Neoplasms / Q000235:genetics* / Q000473:pathology; D018270:Carcinoma, Ductal, Breast / Q000235:genetics* / Q000473:pathology; D005260:Female; D020869:Gene Expression Profiling / Q000379:methods*; D015972:Gene Expression Regulation, Neoplastic; D015723:Gene Library; D006801:Humans; D042282:Microdissection; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D012334:RNA, Neoplasm / Q000032:analysis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction
D012333:RNA, Messenger; D012334:RNA, Neoplasm