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

Key Points:

1. Robotic ultrasound performed favorably compared with magnetic resonance imaging for evaluating total kidney volume.

2. Collagen evaluation by two novel methods of picrosirius red imaging were more informative than the standard method by brightfield imaging.

3. Findings can improve research by increasing speed and access to total kidney volume determination and sensitivity of collagen assessment.

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 and expensive, and access to equipment and expertise is limited. Robotic ultrasound (US) imaging has lower spatial resolution but is faster, more cost-effective, and accessible. Similarly, picrosirius red (PSR) staining and brightfield microscopy are 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 measurements using US and MR imaging. We additionally compared PSR imaging and quantification using standard brightfield microscopy with that by circularly polarized light with hue analysis and fluorescence imaging analyzed using curvelet transform fiber extraction software for automatic detection of individual collagen fibers.

Results: Increased total kidney volume was detected by US in Pkd1^RC/RC versus wild-type (WT) at time points spanning from early to established disease. US interobserver variability was greater but allowed scanning in 2–5 minutes/mouse, whereas MR imaging required 20–30 minutes/mouse. While no change in fibrotic index was detected in this cohort of relatively mild disease using brightfield microscopy, polarized light showed fibers skewed thinner in Pkd1^RC/RC versus WT. Fluorescence imaging showed a higher density of collagen fibers in Pkd1^RC/RC versus WT, and fibers were thinner and curvier with no change in length. In addition, 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 US is a rigorous imaging tool for preclinical PKD research. In addition, they demonstrate the increased sensitivity of polarized and fluorescence analysis of PSR-stained collagen.