Rho-Kinase inhibition decreases focal cerebral ischemia-induced glial activation in rats.

Abstract

Background: Rho-kinase inhibition in a rat middle cerebral artery occlusion (MCAO) model is reported to improve neurological functions and decrease infarction size.

Objective: The objective of this study is to investigate the underlying mechanisms of such improvement by evaluating the effects of Rho-kinase inhibition on astrocytes and microglial accumulation and activation in this condition.

Methods: Adult male Sprague-Dawley (SD) rats were used to generate the MCAO model, which received an I.P injection of a chemical Rho-kinase inhibitor (Fasudil- 5 mg/kg/day) or vehicle (PBS) for 2 and 4 days.

Results: Fasudil treatment significantly decreased the stroke volumes and water content in the lesion areas, as revealed by MRI. Immunostaining and Western blotting results demonstrated that Fasudil significantly decreased the levels of Aquaporin-4, a water channel protein. The number of GFAP⁺ astrocytes and Iba-1⁺ macrophage/microglia was decreased in the lesion areas. Proinflammatory transcription factor NF-κB protein levels were decreased in the Fasudil group 2 days after MCAO. Also, proinflammatory mediators including TNF-α, IL-1β, and iNOS levels were decreased. In vitro migration study using a human microglial cell line (HMO6) confirmed the inhibitory effects of Fasudil on the process. Fasudil also decreased combined IL-1β and IFNγ-induced NF-κB nuclear translocation in HMO6. Moreover, Fasudil transiently decreased combined IL-1β and IFNγ-induced iNOS, TNFα, and IL-1β mRNA levels in HMO6.

Conclusion: Our study demonstrates the inhibitory effects of Rho-kinase on NF-κB-mediated glial activation and cerebral edema, which might be a promising therapeutic target in acute cerebral ischemia conditions.