Texture Analysis as a Discriminating Tool: Unmasking Rodlet Cell Degranulation in Response to a Contaminant of Emerging Concern

Background: Contaminants of emerging concern (CECs) have garnered significant attention due to their potential impacts on ecology, wildlife, and human health. The interest in these contaminants arises from their inadequate regulation or lack of routine monitoring in natural environments. Among them, per- and polyfluoroalkyl substances (PFAS) are of particular concern due to their notable propensity to accumulate within the kidney, significantly influencing the excretion of these pollutants. Rodlet cells (RCs) have emerged as promising indicators of immunotoxicity in response to chemical stressors. A prior comprehensive study extensively detailed the effects of subchronic exposure to perfluorooctanoic acid (PFOA), a well-known PFAS compound, on RCs located in the hematopoietic tissue of the common carp kidney. Even at concentrations commonly found in the environment, PFOA exhibited a significant impact on the distribution patterns of RCs, concurrently enhancing exocytosis activity. Methods: The assessment of PFOA-induced RC degranulation employed texture analysis combined with linear discriminant analysis (LDA) to differentiate between various experimental exposure groups. The investigation encompassed three fish groups: an unexposed group, a group exposed to an environmentally relevant PFOA concentration (200 ng L-1), and a group exposed to a higher PFOA concentration (2 mg L-1). Texture analysis was conducted on highresolution color (RGB) images obtained from light microscopy of ultrathin sections from five fish per experimental group, stained with toluidine blue. Results: This analysis facilitated the quantification of potential cytoplasmic alterations associated with degranulation, encompassing all three RGB channels. The data subjected to LDA enabled the identification of the most distinctive texture characteristics, providing a reliable, objective, and reproducible method to differentiate between experimental groups. Remarkably, 98.0% of both the original and cross-validated cases were correctly classified. However, only one unexposed case was misclassified as a fish exposed to a 200 ng L-1 PFOA concentration, constituting the single false positive in the analysis. Conclusions: Utilizing texture analysis and LDA to quantify RC degranulation offers a dependable approach for assessing immunotoxicity within experimental models of toxicological and environmental pathology. This underscores the scientific significance of employing a morphological approach in such investigations.