0336

Positive Influence of Hyperbaric Oxygenation on Recovery from Brain Trauma

Sanja Pekovic ¹, Toma Jovanovic², Irena Lavrnja¹, Ana Parabucki¹, Predrag Brkic², Ivana Bjelobaba¹, Sanja Dacic³, Danijela Stojkov¹, Ljubisa Rakic⁴, Mirjana Stojiljkovic^1
1University of Belgrade, Institute for Biological Research “Sinisa Stankovic”, Department of Neurobiology, Serbia, ²Institute of Medical Physiology “Richard Burian”, School of Medicine, University of Belgrade, Serbia, ³Faculty of Biology, University of Belgrade, Serbia, ⁴Serbian Academy of Sciences and Arts, Serbia

Objectives

Hyperbaric oxygen (HBO) has been used as a primary or adjunctive therapy over the last 50 years with controversial results, both in experimental and clinical studies, and definitive established mechanisms of action are still lacking. It is not unlikely that some of the effects of HBO on neuronal survival are mediated indirectly by glial cells, which are robustly activated after brain injury and are known to play important roles in neuroprotection and neurodegeneration. Previously, we have shown that cortical lesions cause massive activation of glial cells primarily in injured cortex. Since the effects of repetitive HBO treatment on glial and immune response after stab cortical injury have not been addressed so far, the aim of this study was to compare these responses in treated vs. untreated rats.

Method

Experiments were conducted on the male Wistar rats, 10 weeks old. Surgery: The coordinates of left sensorimotor cortex stab lesion were: 2 mm posterior to the bregma, 2 mm from the midline, 2 mm deep. Sham controls (SC, SC+HBO) passed the same operation protocol but without skull injury. Two groups of animals were left intact (C, C+HBO) and served as physiological controls. HBO treatment: one hour following the surgical procedure, animals were subjected to the HBO protocol for 60 minutes (compression/decompression lasted 10 minutes), pressure applied 2 - 2.5 ATA. The treatment was performed once a day for 10 days. Posttraumatic processes in the brain were evaluated using immunohistochemical method (antibodies used: GFAP, ED1, vimentin, CD40, CD40L, ICAM-1) and Western blot analysis.

Results

GFAP and vimentin immunoreactivity in the peri-lesioned region of injured cortex was significantly lowered in HBO-treated group, due to reduction of reactive astrogliosis and prevention of glial scar formation. Additionally, the macrophage and microglial activities were reduced as well. CD40 was expressed on activated microglia/macrophages after the lesion, being not detected in physiological and HBO conditions. Expression of CD40L was not found in physiological conditions, but after the injury was profoundly expressed on reactive astrocytes and T-lymphocytes around the lesion site. Double CD40L/GFAP labeling revealed their co-localization in reactive astrocytes confirming astrocytes as predominant source of CD40L in the brain. HBO treatment reduced the number of CD40L+ reactive astrocytes in the lesioned area, shifting their morphology towards resting form, whereas CD40L+ T-lymphocytes were not observed. ICAM-1 labeling of blood vessels was intensive after the lesion, while under HBO and in physiological conditions was negligible. All results were confirmed by Western blotting.

Conclusions

Results presented point to HBO-induced suppression of astrogliosis and glial scarring prevention. Additionally, considering CD40/CD40L and ICAM-1 involvement in amplification of immune response after brain injury, observed reduction of inflammatory responses after HBO, recommend HBO treatment as an attractive therapeutic tool for improving recovery from head injury.