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Comparisons of Stereological and Other Approaches for Quantifying Macrophage Aggregates in Piscine Spleens
Mark A Matsche1, Vicki S Blazer2, Patricia M Mazik3
1Maryland Department of Natural Resources, Cooperative Oxford Laboratory, 904 South Morris Street, Oxford, Maryland, 21654, USA.
Abstract:
Macrophage aggregates (MAs) are focal accumulations of pigmented macrophages in the spleen and other tissues of fish. A central role of MAs is the clearance and destruction of degenerating cells and recycling of some cellular components. Macrophage aggregates also respond to chemical contaminants and infectious agents and may play a role in the adaptive immune response. Tissue damage or physiological stress can result in increased MA accumulation. As a result, MAs may be sensitive biomarkers of environmental stress in fish. Abundance of MAs in tissues has been reported in a variety of ways-most commonly as density, mean size, and relative area-but the utility of these estimates has not been compared. In this study, four different types of splenic MA abundance estimates (abundance score, density, relative area, and total volume) were compared in two fish populations (Striped Bass Morone saxatilis and White Perch M. americana) with a wide range in ages. Stereological estimates of total volume indicated an increase in MA abundance with spleen volume, which generally corresponded to fish age, and with splenic infections (mycobacteria or trematode parasites). Abundance scores were generally limited in the ability to detect changes in MA abundance by these factors, whereas density estimates were greatly influenced by changes in spleen volume. In some instances, densities declined while the total volume of MAs and spleen volume increased. Experimentally induced acute stress resulted in a decrease in spleen volume and an increase in MA density, although the total volume of MAs remained unchanged. Relative area estimates accounted for the size and number of MAs but not for changes in organ volume. Total volume is an absolute measure of MA abundance irrespective of changes in organ volume or patterns of accumulation and may provide an improved means of quantifying MAs in the spleens of fish.
Insights
Macrophage aggregates (MAs) in fish spleens are sensitive biomarkers of environmental stress. Total MA volume is the most reliable measure for quantifying abundance, unaffected by organ size changes.
Area of Science:
- Aquatic toxicology
- Fish immunology
- Histopathology
Background:
- Macrophage aggregates (MAs) are focal accumulations of pigmented macrophages in fish tissues, crucial for cellular debris clearance and immune responses.
- MAs can increase due to tissue damage, physiological stress, chemical contaminants, or infections, suggesting their potential as biomarkers of environmental stress.
- Current methods for estimating MA abundance (density, mean size, relative area) lack standardized comparison, limiting their utility.
Purpose of the Study:
- To compare the efficacy of four different methods for quantifying splenic macrophage aggregate (MA) abundance in fish.
- To determine the most reliable method for assessing MA abundance as a biomarker of environmental stress in fish populations.
Main Methods:
- Four methods were used to estimate splenic MA abundance: abundance score, density, relative area, and total volume.
- Stereological techniques were employed to estimate total MA volume.
- Two fish populations, Striped Bass (Morone saxatilis) and White Perch (M. americana), with varying ages and conditions, were studied.
- Experimentally induced acute stress was applied to assess MA responses.
Main Results:
- Total MA volume increased with spleen volume (correlated with fish age) and splenic infections (mycobacteria, trematode parasites).
- Abundance scores showed limited ability to detect changes, while density estimates were significantly influenced by spleen volume fluctuations.
- Acute stress decreased spleen volume but increased MA density, without altering total MA volume.
- Relative area estimates did not account for changes in organ volume.
Conclusions:
- Total MA volume provides an absolute and reliable measure of MA abundance, independent of spleen size or accumulation patterns.
- MA total volume is a superior metric for quantifying MAs in fish spleens compared to abundance scores, density, or relative area.
- This study highlights the importance of selecting appropriate quantification methods for using MAs as sensitive biomarkers in environmental monitoring programs.
