This proposal describes the development of a regenerative treatment for acute kidney injury based on recent advances in reprogramming to change cell fate. Specifically, nuclear reprogramming technology will be employed to create a population of induced nephron progenitor cells. The nephron progenitor cells of the kidney are the only cell type known to be capable of differentiation into all parts of the nephron. Unfortunately, these cells are only found in the cap mesenchyme and are not normally present in the adult kidney. Our collaborators little and colleagues have identified six factors that can convert tubule cells into induced nephron progenitor cells. To capitalize on recent advances in nuclear reprogramming technology for the treatment of acute kidney injury, we will create an artificial population of induced nephron progenitors. This population will be created in situ, or in place, following the delivery of the reprogramming genes to kidney cells of mice in vivo. Others have used in situ reprogramming to successfully treat mouse models of diabetes and cardiac ischemia, suggesting that in situ reprogramming technology may also be useful for protection from ischemia reperfusion injury in the kidney. The reprogramming genes will be carried into the existing kidney cells using a newly developed gene transfer technique. An inducible piggyBac transposon will be used to integrate the genes into the genomes of renal cells and reprogramming will be initiated upon doxycycline induction. Markers of nephron progenitor cells that are not found normally in adult kidneys (Six2, Cited1) will be examined to determine the number and location of induced nephron progenitors in the treated kidneys. Following acute kidney injury, measures of kidney function such as serum creatinine and long-term fibrosis will be assayed to determine the effect of the induced nephron progenitor population on the severity and recovery from injury. Finally, we will determine how the induced nephron progenitor cells protect from acute kidney injury by performing fate- mapping studies in a double transgenic mouse model. These double transgenic mice express a tamoxifen- inducible Cre recombinase from the Cited1 promoter and contain a LacZ transgene that will express LacZ only after Cre recombination. Therefore, only cells that are progeny of the Cited1+ induced nephron progenitor cells will express LacZ. This will allow tracking of the nephron progenitor cell fate by visualization of all LacZ+ cells following acute kidney injury. We hypothesize that the induced nephron progenitors may function to ameliorate acute kidney injury by migrating to sites of damage and replacing the damaged cells, thereby restoring injured nephrons. This would represent a novel mechanism of recovery from acute kidney injury. In conclusion, in situ reprogramming to create induced nephron progenitors represents a potentially powerful treatment for acute kidney injury.
Acute kidney injury is a major cause of morbidity and mortality in the Veteran population, affecting 22% of Veterans admitted to VA intensive care units. Those who survive acute kidney injury may never regain full renal function. Many go on to develop end-stage renal disease, which is itself an expensive and deadly disease requiring dialysis or kidney transplantation for survival. Kidney regeneration is poor following acute kidney injury and there are no treatments available at this time to directly encourage regeneration. New treatments that target regeneration of the kidney are needed because the mammalian kidney is comprised of a limited set of nephrons that are present at birth and lost over time, never to be regained, so renal injuries are permanent. We seek to increase the regenerative capacity of the kidney so that it can better recover from acute kidney injury.
