Current treatment options for acute renal failure are limited. The recovery from renal failure is incomplete if renal epithelial cells fail to regenerate. Since stem cells have diverse developmental potential, we propose to test the hypothesis that mouse hematopoietic stem cells (HSCs) and mouse embryonic stem cells (ES cells) can aid in the regeneration of renal tubules after acute renal failure. In our preliminary studies, we isolated HSCs from male Rosa 26 mice and injected them into female C57 BL mice that had renal ischemia/reperfusion injury. At 4 weeks after HSC transplantation, we detected Lac Z positive cells in the renal tubules of female recipient kidneys. The presence of the male specific SRY gene, and the cells with Y chromosome positive signals in female recipient kidneys confirmed that male donor HSC-derived cells had located to the kidneys. In the first aim of the proposal, we will examine whether exogenous HSCs can transdifferentiate to functional renal tubular cells by staining the kidney tissues with X-gal for the donor marker, and with antibodies to tubular transporters for renal cell markers. We will test the dose response to various numbers of HSCs injected and will examine the effect of HSCs in mice with bilateral renal ischemia/reperfusion injury. The number of donor-derived cells in the kidneys will be correlated with serum BUN and creatinine levels. We will also test whether mobilization of endogenous HSCs with stem cell factor (SCF) and G-CSF can accelerate the recovery of renal function after ischemic injury. In the second aim, we will examine if ES cells can be induced to differentiate to renal cells. Using a unique strain of transgenic mice that express GFP specifically in the epithelial cells of renal tubules and the developing GU tract, we will generate an ES cell line that will allow us to identify and isolate live tubular epithelial cells. ES cells will be induced to form embryoid bodies, and then further cultured with growth factors, conditions known to induce human ES cells to express renal cell markers. GFP positive cells will be isolated with FRCS sorting. The GFP expressing tubular epithelial cells will be injected under the capsule of the kidneys after ischemia/reperfusion injury. The kidney sections will be stained with antibodies to GFP and tubular transporters to test the functional role of transplanted ES cell-derived tubular epithelial cells. The results of this proposal will permit novel understanding of plasticity of HSCs and renal tubular development from ES cells and open up the possibility of treating renal diseases, such as acute renal failure, with stem cell replacement therapy.
