Epithelial cell-targeted transgene expression enables isolation of cyan fluorescent protein (CFP)-expressing prostate stem/progenitor cells

Lankenau Institute for Medical Research, 100 Lancaster Avenue, Wynnewood, PA 19096, USA.
Transgenic Research (Impact Factor: 2.32). 10/2011; 20(5):1073-86. DOI: 10.1007/s11248-010-9478-2
Source: PubMed


To establish a method for efficient and relatively easy isolation of a cell population containing epithelial prostate stem cells, we developed two transgenic mouse models, K5/CFP and K18/RFP. In these models, promoters of the cytokeratin 5 (Krt5) and the cytokeratin 18 (Krt18) genes regulate cyan and red fluorescent proteins (CFP and RFP), respectively. CFP and RFP reporter protein fluorescence allows for visualization of K5(+) and K18(+) epithelial cells within the cellular spatial context of the prostate gland and for their direct isolation by FACS. Using these models, it is possible to test directly the stem cell properties of prostate epithelial cell populations that are positively selected based on expression of cytoplasmic proteins, K5 and K18. After validating appropriate expression of the K5/CFP and K18/RFP transgenes in the developing and adult prostate, we demonstrate that a subset of CFP-expressing prostate cells exhibits stem cell proliferation potential and differentiation capabilities. Then, using prostate cells sorted from double transgenic mice (K5/CFP + K18/RFP), we compare RNA microarrays of sorted K5(+)K18(+) basal and K5(-)K18(+) luminal epithelial cells, and identify genes that are differentially expressed. Several genes that are over-expressed in K5(+) cells have previously been identified as potential stem cell markers. These results suggest that FACS isolation of prostate cells from these mice based on combining reporter gene fluorescence with expression of potential stem cell surface marker proteins will yield populations of cells enriched for stem cells to a degree that has not been attained by using cell surface markers alone.

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    • "B6.129S2-Trp53tm1Tyj/J mice carrying a deletion of exons 2–6 of Trp53 were obtained from The Jackson Laboratory and maintained in a standard special pathogen free (SPF) facility at the University of Helsinki. The reporter mice were from Prof. Janet Sawicki laboratory from the Lankenau Institute for Medical Research41. All animal experiments were approved by the National Animal Experiment Board of Finland (Eläinkoelautakunta, ELLA) in compliance with the Finnish Act on Animal Experimentation (62/2006). "
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