Abstract: FR-PO246
Automated Detection and Quantification of Individual Collagen Fibers in a Mouse Model of Polycystic Kidney Disease
Session Information
- Genetic Diseases of the Kidneys: Cystic - II
November 04, 2022 | Location: Exhibit Hall, Orange County Convention Center‚ West Building
Abstract Time: 10:00 AM - 12:00 PM
Category: Genetic Diseases of the Kidneys
- 1101 Genetic Diseases of the Kidneys: Cystic
Authors
- Sussman, Caroline R., Mayo Foundation for Medical Education and Research, Rochester, Minnesota, United States
- Holmes, Heather L., Mayo Foundation for Medical Education and Research, Rochester, Minnesota, United States
- Harris, Peter C., Mayo Foundation for Medical Education and Research, Rochester, Minnesota, United States
- Romero, Michael F., Mayo Foundation for Medical Education and Research, Rochester, Minnesota, United States
Background
Histological assessment of fibrosis is a standard tool for evaluating PKD. The most commonly used stains are Trichrome or Picrosirius red (PSR), which are imaged using brightfield to calculate a percent positive tissue area (cystic index). These stains are limited by variability due to specimen handling and analysis. PSR emits a red fluorescent signal, which can be used to automatically detect and analyze individual fiber characteristics and determine fiber density.
Methods
Four-month-old Pkd1RC/RC mice were compared to Pkd1+/+ (WT). All mice were F1 progeny from a 129/C57BL6J cross. Following euthanasia, kidneys were fixed in 4% PFA, cryosectioned, stained with PSR, and images were obtained using brightfield and fluorescent imaging. Tissue area and fibrotic index were determined using ImageJ, and individual collagen fibers were automatically detected, and metrics determined using CT-FIRE fiber detection software (LOCI; Madison, WI). Statistical measurements were obtained using PRISM (GraphPad Software, Inc.).
Results
Standard brightfield imaging showed a significantly higher fibrotic index in RC/RC than WT. Analysis of individual fibers by CT-FIRE indicated this was most likely due to an increase in fiber density rather than fiber width. Fiber density was significantly higher in female RC/RC vs WT (4275 ± 254 vs 3423 ± 102 fibers/mm2 respectively (mean ± SEM), p=0.03 by ANOVA, n=3-9) and trended higher in males and overall (4328 ± 203 vs 3667 ± 230 fibers/mm2 respectively (mean ± SEM), p=0.05 by ANOVA, n=6-17). Frequency histograms showed broader distribution of collagen fiber width in RC/RC with a significantly greater percentage of fibers in the lowest quartile. No significant differences were seen in fiber length, straightness, or angle. Overall, fiber width had a bell-shaped distribution, with almost half of fibers having intermediate width, fiber length was skewed towards shorter lengths and straightness was skewed towards being straighter. Fiber angles were more evenly distributed from 0-180 degrees with slightly greater prevalence of angles closer to 90 degrees.
Conclusion
These data demonstrate the potential of CT-FIRE software to provide more objective, higher resolution information about collagen during PKD. Future studies will determine whether it can provide insight into collagen dynamics preceding cysts.
Funding
- NIDDK Support