The Effect of Short-Term Exposure to Cadmium on the Expression of Vascular Endothelial Barrier Antigen in the Developing Rat Forebrain and Cerebellum: A Computerized Quantitative Immunofluorescent Study

Michael O Ibiwoye; Emily A Snyder; James Lyons; Audrey A Vasauskas; Mark J Hernandez; Arthur R Summerlin; James D Foster

doi:10.7759/cureus.23848

The Effect of Short-Term Exposure to Cadmium on the Expression of Vascular Endothelial Barrier Antigen in the Developing Rat Forebrain and Cerebellum: A Computerized Quantitative Immunofluorescent Study

Cureus. 2022 Apr 5;14(4):e23848. doi: 10.7759/cureus.23848. eCollection 2022 Apr.

Authors

Michael O Ibiwoye¹, Emily A Snyder², James Lyons³, Audrey A Vasauskas⁴, Mark J Hernandez¹, Arthur R Summerlin⁵, James D Foster¹

Affiliations

¹ Anatomy and Molecular Medicine, Alabama College of Osteopathic Medicine, Dothan, USA.
² Research, Alabama College of Osteopathic Medicine, Dothan, USA.
³ Department Clinical Sciences, Alabama College of Osteopathic Medicine, Dothan, USA.
⁴ Institutional Effectiveness, Alabama College of Osteopathic Medicine, Dothan, USA.
⁵ Pathology, Southeast Health Medical Center, Dothan, USA.

Abstract

Clinical and laboratory studies have shown that environmental exposure to cadmium produces damage to several organs, including bones, lungs, and kidneys. The involvement of cadmium in central nervous system (CNS) disorders has also been widely reported, but the precise pathophysiological mechanism is not yet fully understood. Children who were exposed to cadmium during pregnancy are known to suffer from developmental delays, learning difficulties, attention deficit hyperactivity disorder (ADHD), and other cognitive and neurobehavioral deficits. Results from numerous studies suggest that dysfunction of the blood-brain barrier (BBB) structures is an important step in the neurotoxicity of cadmium. A rat-specific BBB marker protein, the endothelial barrier antigen (EBA), has been previously isolated and classified by Sternberger and others. The mouse IgG1 clone, anti-endothelial barrier antigen (anti-EBA), detects a protein triplet (23.5kDa, 25 kDa, and 30kDa) localized to the luminal surface of central and peripheral nervous system (CNS and PNS) vascular endothelial cells with selective permeability barrier functions. This marker has been widely used for characterizing BBB alterations under demyelinating, inflammatory, and other CNS pathologies. Many studies have been published using the rat model system for studying the neurotoxic effect of acute and chronic exposure to cadmium. We applied the indirect immunofluorescent techniques using the anti-EBA antibody in conjunction with the Olympus cellSens computerized image analysis to detect and quantify the surface areas of BBB-competent microvessel profiles in paraformaldehyde-fixed, paraffin-embedded brains of term-delivered young rats after intraperitoneal injection of a single dose of cadmium chloride. We detected a statistically significant reduction in EBA-positive microvessel surface areas in the forebrain (t = 5.86, df = 1789, p-value < 0.001) and cerebellum (t=73.40, df=1337, p < 0.001) of cadmium-treated rats compared to the normal controls. Thus, this study supports the hypothesis that the EBA is a sensitive and measurable indicator for quantitative assessment of the impact of cadmium exposure in the developing rat brain.

Keywords: automated digital image analysis; blood–brain barrier; cadmium neurotoxicity; cavalieri principle; endothelial barrier antigen; rat; stereology.