Journal
KIDNEY INTERNATIONAL
Volume 77, Issue 5, Pages 407-416Publisher
ELSEVIER SCIENCE INC
DOI: 10.1038/ki.2009.482
Keywords
cell survival; kidney development; renal cell biology; renal development; tubular epithelium
Categories
Funding
- European Framework 6 KIDSTEM training network
- EU
- NC3Rs
- National Centre for the Replacement, Refinement and Reduction of Animals in Research (NC3Rs) [G0700480/1] Funding Source: researchfish
Ask authors/readers for more resources
Here we describe a novel method in which embryonic kidneys are dissociated into single-cell suspensions and then reaggregated to form organotypic renal structures. Kidney cell reaggregates were transiently cultured with small-molecule Rho kinase inhibitors, which caused ureteric bud structures to form and induced formation of nephrons. These structures displayed normal morphology, expressed appropriate differentiation markers, and were connected at their distal ends to the ureteric buds, thus forming artificial tissues very similar to those found in normal embryonic kidneys. Using this culture method, it was straightforward to make fine-grained chimeras by mixing different cell types or by mixing cells transfected with different constructs before reaggregation. Chimeric renal cultures were formed using mixtures of unmarked normal host embryonic kidney cells and CellTracker-marked WT1 siRNA-carrying cells to test the hypothesis that WT1 is important to a cell's ability to contribute to nephron formation. We found a significant reduction in the ability of WT1 knockdown cells to contribute to nephron formation. This dissociation and reaggregation procedure can also be applied to embryonic lungs and to form coarse-grained hybrid tissues from mixtures of lung and kidney cells. Overall, our protocol allows very simple mixing of cells from different sources or cells subjected to different pretreatments to make fine-grained, highly dispersed chimera tissues. Kidney International (2010) 77, 407-416; doi:10.1038/ki.2009.482; published online 16 December 2009
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available