Postdoctoral Fellow Johns Hopkins University School of Medicine Baltimore, Maryland, United States
Background: Cyclohexanone (CXO), an organic solvent, is ubiquitous in medicine as an adhesive for disposable PVC-based medical products such as IV bags and tubing. Infants and children are exposed in inpatient hospital settings after cardiac surgery, receiving ECMO and/or total parenteral nutrition (TPN). Previously, we reported CXO exposure is associated with worse neurodevelopmental outcomes in humans and rats, though the mechanism is unknown. Objective: To test the hypothesis that neonatal exposure to clinically relevant levels of CXO in rats is associated with fundamental developmental pathophysiology defined by increased systemic inflammation, oligodendrocyte injury, gliosis and neurodevelopmental deficits. Design/Methods: Male and female rats received intraperitoneal injections of either CXO (0.7ul/g body weight/day based on CXO concentrations in NICU TPN) or saline from postnatal day 7 (P7) to P14. Rats were euthanized from P15-P70 with brains and blood analyzed for secreted cytokine and chemokine profiles, terminal deoxynucleotidyl transferase dUTP nick end labeling (Tunel), fluorescence in situ hybridization for mitofusin 2 (mfn2), or immunohistochemistry for glial fibrillary acidic protein (GFAP), myelin basic protein (MBP), degraded MBP (dMBP) and ionized calcium-binding adaptor molecule 1 (Iba1). Images were analyzed with confocal microscopy, Imaris, and Python. Comparisons were made with a student’s t-test, Mann-Whitney test, or Welch’s t-test with p< 0.05 as statistically significant. Results: CXO increased peripheral and central inflammation, changed glial morphology, and caused significant myelin breakdown. Acutely (P15), CXO increased IFN-gamma (p < 0.05), IL-6 (p < 0.05), IL-10 (p < 0.01), and CXCL1 (p < 0.05) in the serum (n=15/group). At P28, CXO elevated serum TNF-alpha (p < 0.05, n=6/group). P28 CXO brains exhibited decreased myelin density, increased astrocytic atrophy, and decreased mfn2 (n=3/group). In CXO white matter, DNA fragmentation increased, and microglia reacted. CXO-exposed microglia (n=3/group, >1700 cells/group) have larger perimeters and areas (p < 0.01) with reduced circularities (p < 0.001).
Conclusion(s): Clinically relevant exposure to CXO for 7 days in neonatal rats increased inflammation, induced neuronal deficits, and impaired neural-immune interactions. Our analysis uncovered multiple possible mechanisms of CXO neurotoxicity, including inflammatory-mediated myelin degradation and mitochondrial failure. More studies are needed to determine the specific mechanisms and explore treatments for worsened outcomes from NICU CXO exposure.
