Robotic Ultrasound and Novel Collagen Analyses for Polycystic Kidney Disease Research Using Mice

Abstract

Background: 3D imaging and histology are critical tools for assessing polycystic kidney disease (PKD) in patients and animal models. Magnetic resonance (MR) imaging provides micron resolution, but is time consuming, expensive, and access to equipment and expertise is limiting. Robotic ultrasound (US) imaging has lower spatial resolution but is faster, more cost effective, and accessible. Similarly, Picrosirius red (PSR) staining and brightfield microscopy is commonly used to assess fibrosis; however, alternative methods have been shown in non-kidney tissues to provide greater sensitivity and more detailed structural characterization. Methods: In this study, we evaluated the utility of robotic US and alternative methods of quantifying PSR staining for PKD research. We compared longitudinal total kidney volume (TKV) measurements using US and MR. We additionally compared PSR imaging and quantification using standard brightfield with that by circularly polarized light with hue analysis, and fluorescence imaging analyzed using CT-FIRE software for automatic detection of individual collagen fibers. Results: Increased TKV was detected by US in Pkd1 RC/RC vs wild type (WT) at timepoints spanning early to established disease. US inter-observer variability was greater but allowed scanning in 2-5 minutes/mouse while MR required 20-30 minutes/mouse. While no change in fibrotic index was detected in this cohort of relatively mild disease using brightfield, polarized light showed fibers skewed thinner in Pkd1 RC/RC vs WT. Fluorescence imaging showed a higher density of collagen fibers in Pkd1 RC/RC vs WT, and fibers were thinner and curvier with no change in length. Additionally, fiber density was higher in both glomeruli and tubules in Pkd1 RC/RC , and glomeruli had a higher fiber density than tubules in Pkd1 RC/RC , and trended higher in WT. Conclusions: These studies show robotic ultrasound is a rigorous imaging tool for pre-clinical PKD research. Additionally, they demonstrate the increased sensitivity of polarized and fluorescence analysis of PSR-stained collagen.