Intraventricular dimethyl sulfoxide (DMSO) induces hydrocephalus in a dose-dependent pattern

Abstract

Introduction: Dimethyl sulfoxide (DMSO), a widely utilized solvent in the medical industry, has been associated with various adverse effects, even at low concentrations, including damage to mitochondrial integrity, altered membrane potentials, caspase activation, and apoptosis. Notably, therapeutic molecules for central nervous system treatments, such as embolic agents or some chemotherapy drugs that are dissolved in DMSO, have been associated with hydrocephalus as a secondary complication. Our study investigated the potential adverse effects of DMSO on the brain, specifically focusing on the development of hydrocephalus and the effect on astrocytes.

Methods: Varied concentrations of DMSO were intraventricularly injected into 3-day-old mice, and astrocyte cultures were exposed to similar concentrations of DMSO. After 14 days of injection, magnetic resonance imaging (MRI) was employed to quantify the brain ventricular volumes in mice. Immunofluorescence analysis was conducted to delineate DMSO-dependent effects in the brain. Additionally, astrocyte cultures were utilized to assess astrocyte viability and the effects of cellular apoptosis.

Results: Our findings revealed a dose-dependent induction of ventriculomegaly in mice with 2%, 10%, and 100% DMSO injections (p < 0.001). The ciliated cells of the ventricles were also proportionally affected by DMSO concentration (p < 0.0001). Furthermore, cultured astrocytes exhibited increased apoptosis after DMSO exposure (p < 0.001).

Conclusion: Our study establishes that intraventricular administration of DMSO induces hydrocephalus in a dose-dependent manner. This observation sheds light on a potential explanation for the occurrence of hydrocephalus as a secondary complication in intracranial treatments utilizing DMSO as a solvent.

Graphical abstract

Fig. 1

DMSO induces ventriculomegaly in a dose-dependent pattern. (A) Graphic representation of the experimental design. (B) MRI images indicating the ventricular frontal horn associated with different concentrations of DMSO and the control group. Scale bar, 1 mm. (C) Histological images of the areas of interest labeled with H&E show the lateral ventricles at the prefrontal and parietal levels. Scale bar, 1 mm (D) Ventricular volume quantification (∗ p < 0.05, ∗∗∗ p < 0.001, One-way ANOVA with posthoc Tukey test. The box represents the mean, and whiskers denote ± SD. The gray dashed line indicates the mean plus 2 standard deviations of control mice, and the red dotted line indicates the mean plus 5 standard deviations). (E) Percentages of mice with ventriculomegaly (mice with larger ventricles than mean ventricular size plus two standard deviations of control mice). (F) Percentages of mice with ventriculomegaly (mice with larger ventricles than mean ventricular size + 5 standard deviations of control mice). (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 2

DMSO-injected mice show a dose-dependent ventricular lining disruption. (A) Representative immunofluorescence of the ventricular lining with the cilia marker anti-βIV tubulin (red) and the astrocyte marker anti-GFAP (green). Scale bars 150 μm. (B) Magnifications of the dashed squares in A. Arrows indicate continuity in the ciliated cells, and asterisks indicate disrupted areas. Scale bars 50 μm. (C) Representative images labeled with H&E (top) and anti-GFAP and anti-βIV tubulin showing ventricular lining disruption (asterisk) in the frontal horn of the lateral ventricles adjacent to an area with normal ciliated cells (arrow). Scale bar, 150 μm. (D) Representative images indicating labeled with H&E (left) and anti-GFAP and anti-βIV tubulin and TUNEL, indicating apoptosis activation (arrows) in the periventricular area adjacent to the ventricular lining disruption (asterisk). Scale bars 20 μm (E) Percentage of ciliated lining disruption (∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001, One-way ANOVA with posthoc Tukey test. Box represents mean, and whiskers denote ± SD). (F) Correlation between ventricular lining disruption and ventricular volume. The gray dashed line indicates the control ventricular volume mean plus 2 standard deviations, and the red dotted line indicates the mean plus 5 standard deviations. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 3

DMSO induces a dose-dependent apoptosis in astrocyte cultures. (A) Schematic overview of the experimental design (B) Representative images indicating the total cell count labeled with DAPI (blue) and the cells in apoptosis marked with a TUNEL assay (red). Scale bar, 100 μm. (C) Percentage of apoptosis increases significantly at 10 and 100% of DMSO concentration. Quantification (∗∗ p < 0.01, ∗∗∗ p < 0.001, One-way ANOVA with posthoc Tukey test. The box represents the mean, and whiskers denote ± SD). (D) Graphic representation of the cell count associated with DMSO concentration. (∗∗∗∗ p < 0.0001, One-way ANOVA with posthoc Tukey test. The box represents the mean, and whiskers denote ± SD). (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)