Effect of salvianolic acid B on TNF-α induced cerebral microcirculatory changes in a micro-invasive mouse model.

PURPOSE: To investigate the effects of salvianolic acid B (SAB) on tumor necrosis factor a (TNF-α) induced alterations of cerebral microcirculation with a bone-abrading model. METHODS: The influences of craniotomy model and bone-abrading model on cerebral microcirculation were compared. The bone-abrading method was used to detect the effects of intracerebroventricular application of 40 µg/kg·bw TNF-α on cerebral venular leakage of fluorescein isothiocyanate (FITC)- albulmin and the rolling and adhesion of leukocytes on venules with fluorescence tracer rhodamine 6G. The therapeutical effects of SAB on TNF-α induced microcirculatory alteration were observed, with continuous intravenous injection of 5 mg/kg·h SAB starting at 20 min before or 20 min after TNF-α administration, respectively. The expressions of CD11b/CD18 and CD62L in leukocytes were measured with flow cytometry. Immunohistochemical staining was also used to detect E-selectin and ICAM-1 expression in endothelial cells. RESULTS: Compared with craniotomy method, the bone-abrading method preserved a higher erythrocyte velocity in cerebral venules and more opening capillaries. TNF-α intervention only caused responses of vascular hyperpermeability and leukocyte rolling on venular walls, without leukocyte adhesion and other hemodynamic changes. Pre- or post-SAB treatment attenuated those responses and suppressed the enhanced expressions of CD11b/CD18 and CD62L in leukocytes and E-selectin and ICAM-1 in endothelial cells induced by TNF-α. CONCLUSIONS: The pre- and post-applications of SAB during TNF-α stimulation could suppress adhesive molecular expression and subsequently attenuate the increase of cerebral vascular permeability and leukocyte rolling.

Cerebralcare Granule® attenuates blood-brain barrier disruption after middle cerebral artery occlusion in rats.

Disruption of blood-brain barrier (BBB) and subsequent edema are major contributors to the pathogenesis of ischemic stroke, for which the current clinical therapy remains unsatisfied. Cerebralcare Granule® (CG) is a compound Chinese medicine widely used in China for treatment of cerebrovascular diseases. CG has been demonstrated efficacy in attenuating the cerebral microcirculatory disturbance and hippocampal neuron injury following global cerebral ischemia. However, the effects of CG on BBB disruption following cerebral ischemia have not been investigated. In this study, we examined the therapeutic effect of CG on the BBB disruption in a focal cerebral ischemia/reperfusion (I/R) rat model. Male Sprague-Dawley rats (250 to 300 g) were subjected to 1h middle cerebral artery occlusion (MCAO). CG (0.4 g/kg or 0.8 g/kg) was administrated orally 3h after reperfusion for the first time and then once daily up to 6 days. The results showed that Evans blue extravasation, brain water content, albumin leakage, infarction volume and neurological deficits increased in MCAO model rats, and were attenuated significantly by CG treatment. T2-weighted MRI and electron microscopy further confirmed the brain edema reduction in CG-treated rats. Treatment with CG improved cerebral blood flow (CBF). Western blot analysis and confocal microscopy showed that the tight junction proteins claudin-5, JAM-1, occludin and zonula occluden-1 between endothelial cells were significantly degradated, but the protein expression of caveolin-1, the principal marker of caveolae in endothelial cells, increased after ischemia, all of which were alleviated by CG treatment. In conclusion, the post-treatment with CG significantly reduced BBB permeability and brain edema, which were correlated with preventing the degradation of the tight junction proteins and inhibiting the expression of caveolin-1 in the endothelial cells. These findings provide a novel approach to the treatment of ischemic stroke.

Protective effects of 3,4-dihydroxyphenyl lactic acid on lipopolysaccharide-induced cerebral microcirculatory disturbance in mice.

This study intended to examine the effect of 3,4-dihydroxyphenyl lactic acid (DLA), a major ingredient of Salvia miltiorrhiza, on lipopolysaccahride (LPS) -induced mouse cerebral cortical microcirculatory disturbance. Velocity of red blood cells in, and albumin leakage from venules, and the numbers of leukocytes rolling on, and adherent to the venular wall were determined by an up-right microscope after LPS (5 mg/kg/h) infusion with or without administration of DLA (5 mg/kg/h). Expression of adhesion molecules CD11b/CD18 and L-selectin on neutrophils, plasma concentration of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) were evaluated by flow cytometry. Concentration of TNF-α in supernatants of LPS-stimulated mononuclear cells was evaluated in vitro by flow cytometry as well. LPS exposure significantly increased the number of rolling and adherent leukocytes as well as albumin leakage, and decreased the velocity of red blood cells in venules. In addition, LPS stimulation apparently increased the expression of CD11b/CD18 on neutrophils, the concentration of plasma TNF-α, and the production of TNF-α from mononuclear cells. Treatment with DLA significantly ameliorated LPS-induced insults in mice, including cerebral microcirculatory disturbance, the expression of CD11b/CD18 on neutrophils, and the increased concentration of plasma TNF-α and the production of TNF-α from mononuclear cells.

The protective effect of Cerebralcare Granule® on brain edema, cerebral microcirculatory disturbance, and neuron injury in a focal cerebral ischemia rat model.

OBJECTIVE: The purpose of the present study was to explore the protective effects of CG on rat cerebral injury after focal cerebral I /R. METHODS: Male Sprague-Dawley rats were subjected to right middle cerebral artery occlusion for 60 minutes followed by reperfusion for 60 minutes or 24 hours. CG (0.4 or 0.8 g/kg) was administrated 90 minutes before ischemia. Brian edema was evaluated by Evan's blue dye extravasations and brain water content, leukocyte adhesion, and albumin leakage were determined with an upright fluorescence microscope, and neuron damage was assessed by 2,3,5-triphenyltetrazolium chloride staining, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling, and immunohistochemistry of caspase-3, p53, p53 upregulated modulator of apoptosis. RESULTS: Focal cerebral I/R elicited a prominent brain edema, an increase in leukocyte adhesion, and albumin leakage, as well as neuron damage. All the insults after focal cerebral I/R were significantly attenuated by pretreatment with CG. CONCLUSIONS: Pretreatment with CG significantly reduced focal cerebral I/R-induced brain edema, cerebral microcirculatory disturbance, and neuron damage, suggesting the potential of CG as a prophylactic strategy for patients in danger of stroke.

Cerebralcare Granule, a Chinese herb compound preparation, improves cerebral microcirculatory disorder and hippocampal CA1 neuron injury in gerbils after ischemia-reperfusion.

AIM OF THE STUDY: Cerebralcare Granule (CG) is a Chinese herb compound preparation that has been used for treatment of cerebrovascular related diseases. However, the effect of post-treatment with CG on ischemia and reperfusion (I/R) induced cerebral injury is so far unclear. MATERIALS AND METHODS: In present study, cerebral global I/R was induced in Mongolian gerbils by clamping bilateral carotid arteries for 30 min followed by reperfusion for 5 days, and CG (0.4 g/kg or 0.8 g/kg) was administrated 3h after the initiation of reperfusion. RESULTS: Post-treatment with CG for 5 days attenuated the I/R-induced production of hydrogen peroxide in, leukocyte adhesion to, and albumin leakage from cerebral microvessels, and, meanwhile, protected neuron from death, reduced the number of caspase-3- and Bax-positive cells, and increased Bcl-2-positive cells in hippocampal CA1 region. CONCLUSION: The results suggest that CG given after initiation of reperfusion is able to ameliorate cerebral microvascular dysfunction and hippocampal CA1 neuron damage caused by I/R.

The antioxidant Cerebralcare Granule attenuates cerebral microcirculatory disturbance during ischemia-reperfusion injury.

Cerebralcare Granule (CG) is a compound Chinese medicine used for treatment of headache and dizziness associated with cerebrovascular diseases. To clarify the mechanism underlying the clinical outcome of CG, this study investigated the effects of CG on the structure and function of cerebral microvasculature during I/R injury. A total of 138 Mongolian gerbils were included and divided into four groups, each composed of 36 or 30 animals, for evaluating various parameters of concern. A skull window was prepared for microcirculatory observation in animals, which were subjected to I/R with or without pretreatment with CG (0.4 or 0.8 g/kg). The velocity of red blood cells in the venules was observed by a high-speed video camera system, along with intravital confocal microscopic measurements of microvascular diameters, adherent leukocytes, and albumin leakage in the brain cortex. Changes in the fluorescence intensity of dihydrorhodamine 123 in cerebral microvessels and malondialdehyde level in the cortex were measured. The ultrastructure of the microvessels in the cerebral cortex was analyzed using both transmission and scanning electron microscopy. In addition, cerebral blood flow was monitored using the laser Doppler imaging technique. Pretreatment with CG (0.4 or 0.8 g/kg) significantly alleviated I/R injury-induced disorders in cerebral microvasculature, as evidenced by the data observed at 60 min of reperfusion wherein the values in CG (0.4 g/kg) pretreatment group, CG (0.8 g/kg) pretreatment group, and I/R group were 2.43 +/- 0.24, 2.28 +/- 0.18, and 6.00 +/- 0.35 for leukocyte adhesion, 2.51 +/- 0.40, 2.33 +/- 0.29, and 4.77 +/- 0.24 for albumin leakage, 7.06 +/- 0.81, 5.93 +/- 0.42, and 28.38 +/- 2.70 for dihydrorhodamine 123 fluorescence intensity in cerebral microvessels, 16.35 +/- 0.52, 14.34 +/- 0.68, and 21.46 +/- 0.71 for malondialdehyde level in the cortex, and 0.43 +/- 0.07, 0.46 +/- 0.02, and 0.17 +/- 0.08 for cerebral blood flow, respectively. I/R injury-elicited ultrastructural alterations in microvessels in cerebral cortex were also mitigated impressively by CG administration, manifested as attenuation of the reduced number of opening capillaries and the altered fine structures in endothelium, which were characterized by rough inner surface, increased intracellular vesicles, hypertrophy of digitations of intercellular contact, and swollen perivascular astroglial processes. Cerebralcare Granule is able to attenuate I/R injury-induced functional and structural changes in microvessels in the cerebral cortex of gerbils, an ability that is most likely correlated with its antioxidant potential.