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bioRxiv2025

Void spot assay visualization optimization for use of Void Whizzard in rats (Rattus norvegicus)

Void spot assay (VSA) noninvasively evaluates urination. VSA is often not performed in rats due to difficulty analyzing larger papers compared with mouse. This study optimizes VSA for rats by comparing post-assay visualization techniques: bright field light (BF), ultraviolet light (UV), and ninhydrin spray (N). Male rats were placed in cages lined by filter paper for 4 hours. Water and food was provided ad lib. After the assay all papers were dried overnight. BF images were photographed (digital camera). UV images were captured after cutting papers in half using a Darkroom ultraviolet imaging cabinet. Papers were sprayed with ninhydrin and photographed (digital camera). All images were converted to black and white for analysis with Void Whizzard. UV vs. BF showed significant differences in area. All three groups had significant differences in overall spot count and in the smallest sized spots (0-0.1 cm2). UV vs. N and UV vs. BF showed significant differences in 0.1-0.25 cm2 spots, UV vs. N in 0.25-0.5 cm2, and N vs. BF in spots 0.5-1 cm2. Overall BF visualization proved difficult. N provided an ideal way to highlight urine and image with a digital camera. Human fingerprints from pre-assay handling of paper interfered with analysis of the smallest sized spots however there were no differences in detection of larger spots, spot distribution, or overall spot area. This study contributes to the development of a standardized VSA protocol for assessing bladder function in both mouse and rat models.

physiology↗

bioRxiv2020

An immunohistochemical prostate cell identification key indicates that aging shifts procollagen 1A1 production from myofibroblasts to fibroblasts in dogs prone to prostate-related urinary dysfunction

Background The identity and spatial distribution of prostatic cell types has been determined in humans but not in dogs, even though aging- and prostate-related voiding disorders are common in both species and mechanistic factors, such as prostatic collagen accumulation, appear to be shared between species. In this publication we characterize the regional distribution of prostatic cell types in the young intact dog to enable comparisons with human and mice and we examine how the cellular source of procollagen 1A1 changes with age in intact male dogs.Methods A multichotomous decision tree involving sequential immunohistochemical stains was validated for use in dog and used to identify specific prostatic cell types and determine their distribution in the capsule, peripheral, periurethral and urethral regions of the young intact canine prostate. Prostatic cells identified using this technique include perivascular smooth muscle cells, pericytes, endothelial cells, luminal, intermediate, and basal epithelial cells, neuroendocrine cells, myofibroblasts, fibroblasts, fibrocytes, and other hematolymphoid cells. Corresponding images were made freely accessible through the GUDMAP database at https://doi.org/10.25548/16-WMM4.Results The prostatic peripheral region harbors the largest proportion of epithelial cells. Aging does not change the density of hematolymphoid cells, fibroblasts, and myofibroblasts in the peripheral region or in the fibromuscular capsule, regions where we previously observed aging- and androgen-mediated increases in prostatic collagen abundance Instead, we observed aging-related changes the procollagen 1A1 positive prostatic cell identity from a myofibroblast to a fibroblast.Conclusions Hematolymphoid cells and myofibroblasts are often identified as sources of collagen in tissues prone to aging-related fibrosis. We show that these are not the likely sources of pathological collagen synthesis in older intact male dogs. Instead, we identify an aging-related shift in the prostatic cell type producing procollagen 1A1 that will help direct development of cell type and prostate appropriate therapeutics for collagen accumulation.View Full Text

pathology↗