Cilostazol reduces blood brain barrier dysfunction, white matter lesion formation and motor deficits following chronic cerebral hypoperfusion.

Cerebral small vessel disease (CSVD) is a pathological process leading to lacunar infarcts, leukoaraiosis and cerebral microbleeds. Dysfunction of the blood brain barrier (BBB) has been proposed as a mechanism in the progression cerebral small vessel disease. A rodent model commonly used to study some aspects of CSVD is bilateral common carotid artery occlusion (BCCAO) in the rat. In the present study it was determined that gait impairment, as determined by a tapered beam test, and BBB permeability increased following BCCAO. Cilostazol, a type III phosphodiesterase inhibitor, has been shown to have anti-apoptotic effects and prevent white matter vacuolation and rarefaction induced by BCCAO in rats. In this study the protective effect of cilostazol administration on the increase BBB permeability following BCCAO was determined as well as the effect on plasma levels of circulating microparticles (MPs), cerebral white matter rarefaction, glial activation and gait disturbance. The effect of cilostazol on in vitro endothelial barriers was also evaluated. Cilostazol treatment improved BBB permeability and reduced gait disturbance, visual impairment and microglial activation in optic tract following BCCAO in vivo. It also reduced the degree of cell death and the reduction in trans-endothelial electrical resistance (TEER) in artificial endothelial barriers in vitro induced by MP treatment of in vitro barriers.

Microparticles generated during chronic cerebral ischemia increase the permeability of microvascular endothelial barriers in vitro.

Numbers of circulating microparticles (MPs) are elevated in a variety of cardiovascular disorders, and recent studies indicate that they are involved in inflammatory intercellular signaling. In the present study the signaling properties of MPs were assessed in an in vitro model of the blood brain barrier. MPs isolated from the plasma of rats exposed to chronic cerebral ischemia caused a significant reduction in the transendothelial electrical resistance (TEER) when applied to in vitro endothelial barriers, while MPs isolated from an equal volume of plasma from unoperated or sham operated rats did not. The reduction in TEER was attenuated by treating endothelial barriers prior to exposure to MPs with the caspase 3 inhibitor AC-DEVD-CHO, the TNF-α inhibitor SPD304, the tumor necrosis factor alpha-converting enzyme (TACE, ADAM 17) inhibitor TAPI-0-1 and the Rho kinase (ROCK) inhibitor Y-27632, and by treating the MPs themselves with these inhibitors prior to applying them to cultured cells. This observation indicates that MPs generated during cerebral ischemia contain pro-TNF-α, active TACE and active ROCK. ROCK and Ras homolog gene family member A (RhoA) were detected in MPs by western blot. The growth factor VEGF stimulated transcellular transport in endothelial barriers while exposure to MPs did not. We conclude that the increase in permeability of artificial barriers induced by MPs is primarily due to enhanced apoptosis induced by activation of the TNF-α pathway and activated caspase 3 and Rho kinases delivered to endothelial cells by MPs.

Microparticles generated during chronic cerebral ischemia deliver proapoptotic signals to cultured endothelial cells.

Circulating microparticles (MPs) are involved in many physiological processes and numbers are increased in a variety of cardiovascular disorders. The present aims were to characterize levels of MPs in a rodent model of chronic cerebral hypoperfusion (CCH) and to determine their signaling properties. MPs were isolated from the plasma of rats exposed to CCH and quantified by flow cytometry. When MPs were added to cultured endothelial cells or normal rat kidney cells they induced cell death in a time and dose dependent manner. Analysis of pellets by electron microscopy indicates that cell death signals are carried by particles in the range of 400 nm in diameter or less. Cell death involved the activation of caspase 3 and was not a consequence of oxidative stress. Inhibition of the Fas/FasL signaling pathway also did not improve cell survival. MPs were found to contain caspase 3 and treating the MPs with a caspase 3 inhibitor significantly reduced cell death. A TNF-α receptor blocker and a TRAIL neutralizing antibody also significantly reduced cell death. Levels of circulating MPs are elevated in a rodent model of chronic cerebral ischemia. MPs with a diameter of 400 nm or less activate the TNF-α and TRAIL signaling pathways and may deliver caspase 3 to cultured cells.