J Neurocrit Care.  2023 Jun;16(1):18-27. 10.18700/jnc.220052.

Neuroprotective effects of chloroquine on neurological scores, blood-brain barrier permeability, and brain edema after traumatic brain injury in male rats

• Karimi M
• Nazari S
• Shirani F
• Alivirdiloo V
• Siahposht-Khachaki A
• Edalatkhah S
• Khanian R

• KMID: 2543388
• DOI: http://doi.org/10.18700/jnc.220052

Abstract

Background
Traumatic brain injury (TBI) is one of leading causes of death among young people worldwide. Chloroquine, an antimalarial drug, has been shown to easily cross the blood-brain barrier (BBB) and inhibit autophagy in a variety of disorders, including Alzheimer disease and brain ischemia. We investigated the effects of chloroquine on neuronal protection after induction of brain trauma in male rats.
Methods
A total of 120 male Wistar rats were treated with chloroquine at doses of 1.5, 3, and 6 mg/kg intraperitoneally after induction of diffuse TBIs. The veterinary coma scale was used to assess short-term neurological deficits. BBB disruption was evaluated using the Evans Blue dye method 6-hour post-injury. Vestibulomotor function was evaluated using the beam walk and beam balance methods. Histopathological changes in the brain tissue in different groups were evaluated using light microscopy and hematoxylin-eosin staining. Brain water and cerebrospinal fluid (CSF) contents of matrix metalloproteinase 9 (MMP-9) were assessed using the wet/dry method and enzyme-linked immunosorbent assay, respectively.
Results
The results showed that injecting chloroquine (3 and 6 mg/kg) 30 minutes after TBI significantly reduced brain edema and BBB disruption, and recovered neurological deficits post-TBI (P<0.01). Furthermore, CSF MMP-9 was significantly reduced after administration of 1.5 mg/kg chloroquine (P<0.01).
Conclusion
Chloroquine has neuroprotective effects in the brain, and thus, has the potential to mitigate the effects of brain trauma. It is possible that the anti-inflammatory and neurogenic effects of chloroquine are due to a decrease in MMP secretion in the CSF.

Keyword

Chloroquine; Brain trauma; Neural protection

Figure

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  Fig. 1. Beam-walk and beam-balance task scores in three consecutive days after traumatic brain injury (TBI) injury (n=8 per group). (A) Beam-walk task transverse time in seconds in three consecutive days after TBI. Data were analyzed using repeated measure two-way analysis of variance (ANOVA) with Greenhouse-Geisser correction. Tukey’s HSD test was used as the post-hoc test. (B) Area under the curve (AUC) calculated separately for the beam walk during experimental TBI in male rats in different days (before trauma [pre], on trauma day [D0], first [D1], second [D2] and third [D3] days). (C) Beam-balance task transverse time in seconds in three consecutive days after TBI. Data are analyzed using repeated measure two-way ANOVA with Greenhouse-Geisser correction. Tukey’s HSD test was used as the post-hoc test. (D) AUC calculated separately for the beam balance during experimental TBI in male rats in different days (pre, D0, D1, D2 and D3 days). CQ, chloroquine. a)P<0.05; b)P<0.01; c)P<0.001.

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  Fig. 2. The effects of chloroquine (CQ) on brain water content (%) 24 hours after traumatic brain injury (TBI). Each bar shows mean±standard error of mean for eight rats. Data were analyzed using one-way analysis of variance test. Newman-Keuls test was used as the post-hoc test. Compared with saline and TBI groups: a)P<0.05; b)P<0.01.

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  Fig. 3. The effects of chloroquine (CQ) on brain tissue Evans blue dye content 6 hours after traumatic brain injury (TBI). Each bar shows mean±standard error of mean for eight rats. Data were analyzed using one-way analysis of variance test. Newman-Keuls test was used as the post-hoc test. Compared with saline and TBI groups: a)P<0.05; b)P<0.01.

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  Fig. 4. Cerebrospinal fluid (CSF) content of matrix metalloproteinase 9 (MMP-9) measured by enzyme-linked immunosorbent assay 24 hours after traumatic brain injury (TBI) injury (n=8 in each group). Each bar shows mean±standard error of mean for eight rats. Data were analyzed using one-way analysis of variance test. Newman-Keuls test was used as the post hoc test. CQ, chloroquine. Compared with saline and TBI groups: a)P<0.05; b)P<0.01.

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  Fig. 5. The effects of chloroquine (CQ) on histopathology changes in 24 hours post-traumatic brain injury (TBI; H&E, ×400). (A) TBI group, (B) saline, (C) CQ 1.5 mg/kg, (D) CQ 3 mg/kg, and (E) CQ 6 mg/kg in male rat’s brain. , swollen astrocyte; ★, blood vessel; *, degenerated neuron; →, edematous neuron; ↗, endothelial cell; , Normal neuron.

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