Gene expression analysis on small numbers of invasive cells collected by chemotaxis from primary mammary tumors of the mouse.
BACKGROUND
cDNA microarrays have the potential to identify the genes involved in invasion and metastasis. However, when used with whole tumor tissue, the results average the expression patterns of different cell types. We have combined chemotaxis-based cell collection of the invasive subpopulation of cells within the primary tumor with array-based gene expression analysis to identify the genes necessary for the process of carcinoma cell invasion.


RESULTS
Invasive cells were collected from live primary tumors using microneedles containing chemotactic growth factors to mimic chemotactic signals thought to be present in the primary tumor. When used with mammary tumors of rats and mice, carcinoma cells and macrophages constitute the invasive cell population. Microbeads conjugated with monoclonal anti-CD11b (Mac-1alpha) antibodies were used to separate macrophages from carcinoma cells. We utilized PCR-based cDNA amplification from small number of cells and compared it to the quality and complexity of conventionally generated cDNA to determine if amplified cDNA could be used with fidelity for array analysis of this cell population. These techniques showed a very high level of correlation indicating that the PCR based amplification technique yields a cDNA population that resembles, with high fidelity, the original template population present in the small number of cells used to prepare the cDNA for use with the chip.


CONCLUSIONS
The specific collection of invasive cells from a primary tumor and the analysis of gene expression in these cells are is now possible. By further comparing the gene expression patterns of cells collected by invasion into microneedles with that of carcinoma cells obtained from the whole primary tumor, the blood, and whole metastatic tumors, genes that contribute to the invasive process in carcinoma cells may be identified.
Wang
Weigang
W
Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, Bronx, New York 10461, USA. wgwang@aecom.yu.edu
Wyckoff
Jeffrey B
JB
Wang
Yarong
Y
Bottinger
Erwin P
EP
Segall
Jeffrey E
JE
Condeelis
John S
JS
D000818:Animals; D002277:Carcinoma / Q000235:genetics*; D002469:Cell Separation / Q000379:methods; D002633:Chemotaxis / Q000235:genetics*; D018076:DNA, Complementary / Q000032:analysis; D005260:Female; D020869:Gene Expression Profiling / Q000379:methods*; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics*; D008264:Macrophages / Q000737:chemistry / Q000378:metabolism / Q000473:pathology; D015674:Mammary Neoplasms, Animal / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D051379:Mice; D009361:Neoplasm Invasiveness / Q000235:genetics; D021141:Nucleic Acid Amplification Techniques; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D016133:Polymerase Chain Reaction; D012334:RNA, Neoplasm / Q000235:genetics; D051381:Rats
D018076:DNA, Complementary; D012334:RNA, Neoplasm