Fibroblast-Specific PAI-1 Depletion Increases Macrophages but Not Kidney Fibrosis after Unilateral Ureteral Obstruction
Laura S Peterson, Amir M Khan, Haichun Yang, Agnes B Fogo. Vanderbilt University Medical Center, Nashville, TN; Chicago Medical School at Rosalind Franklin University of Medicine and Science, Chicago, IL
Background: Plasminogen activator inhibitor 1 (PAI-1) is a serine protease implicated in pro-fibrotic kidney disease. PAI-1 inhibits matrix breakdown and plays important roles in chemotaxis and proliferation of inflammatory and wound-healing cells. Systemic PAI-1 knockout mice have decreased interstitial renal scarring in response to ureteral obstruction. A multitude of cell types in the kidney express PAI-1 under stress. This study was designed to determine the effect of fibroblast-specific PAI-1 depletion in mice after ureteral obstruction.
Design: We created mice with floxed PAI-1 and bred to tenascin C Cre mice to knock down PAI-1 in fibroblasts after administration of tamoxifen. Unilateral ureteral obstruction (UUO) was performed in PAI-1loxP/loxP/Cre (n=4) and control mice (Cre or PAI-1loxP/lox, n=8), and both kidneys were harvested after 10 days. The efficiency of the inducible fibroblast specific PAI-1 knockout was measured by double staining for PAI-1 and GFP, which binds to tenascin C. Interstitial fibrosis was measured by Sirius red staining. PAI-1, collagen I and III mRNA was determined by real time PCR. Fibroblast and macrophage infiltration were assessed by FSP-1 and F4/80 staining.
Results: 76-87% of GFP positive cells, i.e. fibroblasts, were PAI-1 negative in PAI-1loxP/loxP/ Cre mice while control mice showed PAI-1 positivity in all interstitial cells, indicating effective knockdown of PAI-1 in this model. PAI-1loxP/loxP/Cre also had less PAI-1 mRNA in obstructed kidneys, 54.7% compared to control mice. F4/80 positive cells were increased in PAI-1loxP/loxP/Cre vs. control (43.7 ± 2.6 vs. 23.1 ± 3.7/HPF, p<0.05). However, fibroblast cell number was not different between the two groups (10.7 ± 7.6 vs. 18.6 ± 11.0/HPF, p NS). Further, fibrosis, assessed by Sirius red positive area, collagen I and III mRNA, did not differ between the two groups.
Conclusions: Our studies demonstrate efficacy of fibroblast-specific PAI-1 depletion in mice. Macrophage accumulation in obstructed kidneys is markedly increased in the absence of PAI-1 signaling from fibroblasts. Interestingly, this increase in macrophages is not accompanied by change in fibrosis or number of fibroblasts. Whether the phenotype of macrophages is altered when fibroblast PAI-1 is diminished, or whether PAI-1 in cells other than fibroblasts crucially modify ultimate fibrosis, awaits further study.
Category: Kidney (does not include tumors)
Monday, March 4, 2013 1:00 PM
Poster Session II # 255, Monday Afternoon