Bai-He Hu

Cardiotonic pills, a compound Chinese medicine, protects ischemia-reperfusion-induced microcirculatory disturbance and myocardial damage in rats

Cardiotonic pills (CP) is a compound Chinese medicine widely used in China, as well as other countries, for the treatment of cardiovascular disease. However, limited data are available regarding the mechanism of action of CP on myocardial function during ischemia-reperfusion (I/R) injury. In this study, we examined the effect of CP on I/R-induced coronary microcirculatory disturbance and myocardial damage. Male Sprague-Dawley rats were subjected to left coronary anterior descending branch occlusion for 30 min followed by reperfusion with or without pretreatment with CP (0.1, 0.4, or 0.8 g/kg). Coronary blood flow, vascular diameter, velocity of red blood cells, and albumin leakage were evaluated in vivo after reperfusion. Neutrophil expression of CD18, malondialdehyde, inhibitor-kappaBalpha, myocardial infarction, endothelial expression of intercellular adhesion molecule 1, apoptosis-related proteins, and histological and ultrastructural evidence of myocardial damage were assessed after reperfusion. Pretreatment with CP (0.8 g/kg) significantly attenuated the I/R-induced myocardial microcirculatory disturbance, including decreased coronary blood flow and red blood cell velocity in arterioles, increased expression of CD18 on neutrophils and intercellular adhesion molecule 1 on endothelial cells, and albumin leakage from venules. In addition, the drug significantly ameliorated the I/R-induced myocardial damage and apoptosis indicated by increased malondialdehyde, infarct size, myocardial ultrastructural changes, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling-positive myocardial cells, inhibitor-kappaBalpha degradation, and expression of Bcl-2, Bax, and caspase-3 in myocardial tissues. The results provide evidence for the potential role of CP in preventing microcirculatory disturbance and myocardial damage following I/R injury.

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.

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.

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.

QiShenYiQi Pills® prevents cardiac ischemia–reperfusion injury via energy modulation☆

BACKGROUND QiShenYiQi Pills® (QSYQ) is a compound Chinese medicine used in China for alleviating cardiac function. The present study was designed to explore the effect and mechanism of QSYQ on ischemia-reperfusion (I/R)-induced disorders in myocardial structure and function, with particularly focusing on the regulation of energy metabolism. METHODS Sprague-Dawley rats, with or without QSYQ pretreatment, were subjected to 30 min occlusion of the left anterior descending coronary artery and followed by 90 min or 24h reperfusion. Myocardial blood flow (MBF) and cardiac function were evaluated at baseline, immediately after ischemia and 30, 60, 90 min, and 24h after reperfusion. Myocardial infarction, myocardial histology and ultrastructure were assessed. Double staining of alpha-cardiac actinin and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling was conducted to assess myocardial apoptosis. ATP, ADP and AMP content was determined by Enzyme-Linked Immunosorbent Assay, F-actin in myocardial cells determined by immunofluorescence microscopy and expression of ATP synthase α, ATP5D, and phosphorylated-Myosin Light Chain (P-MLC) determined by western blotting. RESULTS Pre-treatment with QSYQ protected against I/R-induced MBF decrease, myocardial infarction and apoptosis at 90 min and 24h after reperfusion. Moreover, I/R 90 min caused an impairment on cardiac function, a decrease in the ratio of ADP/ATP and AMP/ATP, accompanying with reduction of ATP 5D expression and increase in the expression of P-MLC, meanwhile, myocardium to exhibit myocardial fiber rupture, interstitial edema, and mitochondria swelling, all of which were significantly ameliorated by pre-treatment with QSYQ. CONCLUSIONS The results of the present study suggest an involvement of regulation of energy metabolism in the action of QSYQ to protect against myocardial I/R injury.

Levo-tetrahydropalmatine attenuates mouse blood-brain barrier injury induced by focal cerebral ischemia and reperfusion: Involvement of Src kinase

The restoration of blood flow following thrombolytic therapy causes ischemia and reperfusion (I/R) injury leading to blood-brain barrier (BBB) disruption and subsequent brain edema in patients of ischemic stroke. Levo-tetrahydropalmatine (l-THP) occurs in Corydalis genus and some other plants. However, whether l-THP exerts protective role on BBB disrpution following cerebral I/R remains unclear. Male C57BL/6N mice (23 to 28 g) were subjected to 90 min middle cerebral artery occlusion, followed by reperfusion for 24 h. l-THP (10, 20, 40 mg/kg) was administrated by gavage 60 min before ischemia. We found I/R evoked Evans blue extravasation, albumin leakage, brain water content increase, cerebral blood flow decrease, cerebral infarction and neurological deficits, all of which were attenuated by l-THP treatment. Meanwhile, l-THP inhibited tight junction (TJ) proteins down-expression, Src kinase phosphorylation, matrix metalloproteinases-2/9 (MMP-2/9) and caveolin-1 activation. In addition, surface plasmon resonance revealed binding of l-THP to Src kinase with high affinity. Then we found Src kinase inhibitor PP2 could attenuate Evans blue dye extravasation and inhibit the caveolin-1, MMP-9 activation, occludin down-expression after I/R, respectively. In conclusion, l-THP attenuated BBB injury and brain edema, which were correlated with inhibiting the Src kinase phosphorylation.

Attenuating effect of post-treatment with QiShen YiQi Pills on myocardial fibrosis in rat cardiac hypertrophy

QiShen YiQi Pills(®) (QSYQ) is a compound Chinese medicine used for treatment of cardiovascular diseases. However, the potential of QSYQ to inhibit cardiac fibrosis in left ventricle hypertrophy is not explored to date. We investigated the effects of post-treatment with QSYQ on rat myocardial fibrosis in left ventricle hypertrophy induced by pressure over-load through ascending aortic stenosis. QSYQ was administrated 4 weeks after the surgery, at a dose of 0.8 g/kg/day over the next 4 weeks, while echocardiography was performed 4 and 8 weeks, respectively, after the surgery. Eight weeks after the surgery, myocardial blood flow was determined by Laser-Doppler Perfusion Imager and the ratio of heart weight to body weight (HW/BW) was estimated, in concurrent evaluation of myocardial histology and ultrastructure, as well as collagen content by sirius red staining, and immunohistochemistry staining for CD68 and transforming growth factor beta 1. Post-treatment with QSYQ significantly alleviated left ventricular posterior wall end diastolic thickness and the HW/BW, increased left ventricle ejection fraction and left ventricle fractional shortening. QSYQ also decreased myocardial fibrosis size. The expression of CD68 and transforming growth factor beta 1 were obviously suppressed after QSYQ treatment. The results suggest that post-treatment with QSYQ attenuates pressure over-load-induced cardiac hypertrophy and myocardial fibrosis through interfering in inflammatory process.

ROCK-dependent ATP5D modulation contributes to the protection of notoginsenoside NR1 against ischemia-reperfusion-induced myocardial injury

Cardiac ischemia-reperfusion (I/R) injury remains a challenge for clinicians, which initiates with energy metabolism disorder. The present study was designed to investigate the protective effect of notoginsenoside R1 (NR1) on I/R-induced cardiac injury and underlying mechanism. Male Sprague-Dawley rats were subjected to 30-min occlusion of the left coronary anterior descending artery followed by reperfusion with or without NR1 pretreatment (5 mg·kg(-1)·h(-1)). In vitro, H9c2 cells were cultured under oxygen and glucose deprivation/reoxygenation conditions after NR1 (0.1 mM), Rho kinase (ROCK) inhibitor Y-27632 (10 μM), or RhoA/ROCK activator U-46619 (10 nM) administration. Myocardial infarct size, myocardial histology, and cardiac function were evaluated. Myofibril and mitochondria morphology were observed by transmission electron microscopy. F-actin and apoptosis were determined by immunofluorescence and TUNEL staining. ATP and AMP content were assessed by ELISA. Phosphorylated-AMP-activated protein kinase, ATP synthase subunits, apoptosis-related molecules, and the level and activity of ROCK were determined by Western blot analysis. We found that NR1 pretreatment ameliorated myocardial infarction, histological injury, and cardiac function induced by I/R. Furthermore, similar to the effect of Y-27632, NR1 improved H9c2 cell viability, maintained actin skeleton and mitochondria morphology, and attenuated apoptosis induced by oxygen and glucose deprivation/reoxygenation. Importantly, NR1 prevented energy abnormity, inhibited the expression and activation of ROCK, and restored the expression of the mitochondrial ATP synthase δ-subunit both in vivo and in vitro, whereas U-46619 suppressed the effect of NR1. These results prove NR1 as an agent able to prevent I/R-induced energy metabolism disorder via inhibiting ROCK and enhancing mitochondrial ATP synthase δ-subunits, which at least partially contributes to its protection against cardiac I/R injury.

Treatment with cardiotonic pills(®) after ischemia-reperfusion ameliorates myocardial fibrosis in rats.

The present study was designed to evaluate whether CP was beneficial in alleviating myocardial fibrosis following I/R injury.

Ginsenoside Rb1 ameliorates lipopolysaccharide-induced albumin leakage from rat mesenteric venules by intervening in both trans- and paracellular pathway

Lipopolysaccharide (LPS) is one of the common pathogens that causes mesentery hyperpermeability- and intestinal edema-related diseases. This study evaluated whether ginsenoside Rb1 (Rb1), an ingredient of a Chinese medicine Panax ginseng, has beneficial effects on mesentery microvascular hyperpermeability induced by LPS and the underlying mechanisms. Male Wistar rats were continuously infused with LPS (5 mg · kg(-1) · h(-1)) via the left jugular vein for 90 min. In some rats, Rb1 (5 mg · kg(-1) · h(-1)) was administrated through the left jugular vein 30 min after LPS infusion. The dynamics of fluorescein isothiocynate-labeled albumin leakage from mesentery venules was assessed by intravital microscopy. Intestinal tissue edema was evaluated by hematoxylin and eosin staining. The number of caveolae in endothelial cells of microvessels was examined by electron microscopy. Confocal microscopy and Western blotting were applied to detect caveolin-1 (Cav-1) expression and phosphorylation, junction-related proteins, and concerning signaling proteins in intestinal tissues and human umbilical vein endothelial cells. LPS infusion evoked an increased albumin leakage from mesentery venules that was significantly ameliorated by Rb1 posttreatment. Mortality and intestinal edema around microvessels were also reduced by Rb1. Rb1 decreased caveolae number in endothelial cells of microvessels. Cav-1 expression and phosphorylation, VE-Cadherin phosphorylation, ZO-1 degradation, nuclear factor-κB (NF-κB) activation, and Src kinase phosphorylation were inhibited by Rb1. Rb1 ameliorated microvascular hyperpermeability after the onset of endotoxemia and improved intestinal edema through inhibiting caveolae formation and junction disruption, which was correlated to suppression of NF-κB and Src activation.