Upregulation of Nav1.6 expression in the rostral ventrolateral medulla of stress-induced hypertensive rats.

The rostral ventrolateral medulla (RVLM) plays a key role in mediating the development of stress-induced hypertension (SIH) by excitation and/or inhibition of sympathetic preganglionic neurons. The voltage-gated sodium channel Nav1.6 has been found to contribute to neuronal hyperexcitability. To examine the expression of Nav1.6 in the RVLM during SIH, a rat model was established by administering electric foot-shocks and noises. We found that Nav1.6 protein expression in the RVLM of SIH rats was higher than that of control rats, peaking at the tenth day of stress. Furthermore, we observed changes in blood pressure correlating with days of stress, with systolic blood pressure (SBP) found to reach a similarly timed peak at the tenth day of stress. Percentages of cells exhibiting colocalization of Nav1.6 with NeuN, a molecular marker of neurons, indicated a strong correlation between upregulation of Nav1.6 expression in NeuN-positive cells and SBP. The level of RSNA was significantly increased after 10 days of stress induction than control group. Compared with the SIHR, knockdown of Nav1.6 in RVLM of the SIHR decreased the level of SBP, heart rate (HR) and renal sympathetic nerve activity (RSNA). These results suggest that upregulated Nav1.6 expression within neurons in the RVLM of SIH rats may contribute to overactivation of the sympathetic system in response to SIH development.

Effects of CDC42 on the proliferation and invasion of gastric cancer cells.

Cell division cycle 42 (CDC42), which is a member of the Rho GTPase family, has been reported to regulate the metastasis of various human cancer cells; however, the role of CDC42 in gastric cancer (GC) remains unclear. The present study aimed to investigate the effects of CDC42 on the proliferation, migration and invasion of GC. Furthermore, the molecular mechanisms underlying the effects of CDC42 on GC were explored. The expression levels of CDC42 in the AGS and SGC7901 human GC cell lines were reduced by RNA interference. Knockdown of CDC42 significantly inhibited the proliferation of AGS and SGC7901 cells, and it was suggested that this inhibitory process may be due to cell cycle arrest at G1/S phase and downregulation of cyclin A, cyclin D1, cyclin E and proliferating cell nuclear antigen. Furthermore, knockdown of CDC42 markedly inhibited the migration and invasion of GC cells, and suppressed the expression of matrix metalloproteinase 9. These results indicated that CDC42 is a key regulator involved in regulating the proliferation, migration and invasion of GC, and it may be considered a potential therapeutic target in GC.

Protective effects of Gingko biloba extract 761 on myocardial infarction via improving the viability of implanted mesenchymal stem cells in the rat heart.

When introduced into the infarcted heart, bone marrow­derived mesenchymal stem cells (MSCs) prevent the heart from deleterious remodeling and improve its recovery. The aim of the present study was to investigate the effects of Ginkgo biloba extract (EGb) 761 on the infarcted myocardium microenvironment following MSC transplantation. The established rat myocardial infarction (MI) model, with implanted PKH­26 marked MSCs (1x105 cells), were randomly divided into two groups: The control group (injected with normal saline) and the EGb 761 treatment group (injected with 100 mg/kg/day EGb 761). The following indices for cardiac function, including the extent of inflammation, oxidative stress, MSC apoptosis and MSC differentiation were measured 1, 2 and 7 days after treatment. The anti­inflammatory effect of EGb 761 was observed by histological examination. Compared with the respective control group, the malondialdehyde content significantly decreased and the superoxide dismutase, catalase and glutathione peroxidase activity significantly increased in the EGb761­treated groups. In addition, the apoptotic index gradually decreased (P<0.05) with the extension of MI time in the EGb761-treated groups compared to the respective control groups, suggesting that EGb761 exhbits anti-oxidative effects. In addition, the level of the Fas protein was positively correlated with the implanted MSC apoptotic ratio. Following 7 days of MSC transplantation with EGb 761 treatment, the expression of cTnI in PKH26­labeled MSCs was observed in the transplanted myocardium. Cardiac function, including the ejection fraction, left ventricular end­systolic pressure and dp/dtmax significantly increased, and the left ventricular end diastolic diameters, left ventricular end­diastolic volumes and left ventricular end­diastolic pressure significantly decreased (P<0.05, vs. the control group). The results demonstrated that EGb 761 is important in improving cardiac function and the infarcted myocardium microenvironment. The present study indicated that the protective effects of EGb 761 on the infarcted myocardium may be mediated by improving the viability and the differentiation of the implanted MSCs into cardiomyocytes.

Repeated electroacupuncture attenuating of apelin expression and function in the rostral ventrolateral medulla in stress-induced hypertensive rats.

Studies have revealed that apelin is a novel multifunctional peptide implicated both in blood pressure (BP) regulation and cardiac function control. Evidence shows that apelin and its receptor (APJ) in the rostral ventrolateral medulla (RVLM) may play an important role in central BP regulation; however, its role is controversial and very few reports have shown the relationship between acupuncture and apelin. Our study aims to both investigate the apelinergic system role in stress-induced hypertension (SIH) and determine whether acupuncture therapy effects on hypertension involve the apelinergic system in the RVLM. We established the stress-induced hypertensive rat (SIHR) model using electric foot-shock stressors with noise interventions. The expression of both apelin and the APJ receptor in the RVLM neurons was examined by immunohistochemical staining and Western blots. The results showed apelin expression increased remarkably in SIHR while APJ receptor expression showed no significant difference between control and SIHR groups. Microinjection of apelin-13 into the RVLM of control rats or SIHR produced pressor and tachycardic effects. Furthermore, effects induced by apelin-13 in SIHR were significantly greater than those of control rats. In addition, repetitive electroacupuncture (EA) stimulation at the Zusanli (ST-36) acupoint attenuated hypertension and apelin expression in the RVLM in SIHR; it also attenuated the pressor effect elicited by exogenous apelin-13 microinjection in SIHR. The results suggest that augmented apelin in the RVLM was part of the manifestations of SIH; the antihypertensive effects of EA might be associated with the attenuation of apelin expression and function in the RVLM, which might be a novel role for EA in SIH setting.

Nitric oxide modulates cardiovascular function in the rat by activating adenosine A2A receptors and inhibiting acetylcholine release in the rostral ventrolateral medulla.

1. Nitric oxide (NO), a gas transmitter, modulates many physiological processes, including the central regulation of cardiovascular activity. However, the mechanisms underlying the regulation of cardiovascular activity remain relatively unexplored. In the present study, we hypothesized that central NO-dependent sympathetic inhibition is mediated by activation of adenosine A(2A) receptors (A(2A)R) and inhibition of acetylcholine (ACh) release in the rostral ventrolateral medulla (RVLM). 2. L-Arginine (L-Arg; an NO donor; 100 nmol/100 nL) was microinjected into the RVLM of male Sprague-Dawley rats and heart rate variability (HRV) was assessed as an index of cardiac sympathovagal balance. Following microinjection of L-Arg, decreases were seen in mean arterial pressure (MAP), heart rate (HR) and the ratio of the low- to high-frequency components (LF/HF) of HRV. Pretreatment of rats with SCH58261 (40 pmol/60 nL into the RVLM), a competitive antagonist of the A(2A) R, attenuated these effects. 3. Western blot analysis and ELISA revealed that adenosine and A(2A)R levels increased in the RVLM following L-Arg microinjection, whereas ACh and muscarinic M(1) receptor levels decreased significantly, in parallel with the cardiovascular responses to L-Arg microinjection. The decrease in ACh levels was abolished by SCH58261 pretreatment. 4. Microinjection of N(G)-nitro-L-arginine methyl ester (a non-selective inhibitor of NO synthase; 15 nmol/100 nL) into the RVLM significantly increased MAP, HR and sympathetic activity, as evidenced by HRV (LF, HF and the LF/HF ratio were all increased). 5. The results indicate that the central NO/NO synthase system in the RVLM may modulate cardiovascular activity by activating the A(2A)R, which subsequently inhibits activation of the muscarinic M(1) receptor.

The protective effect of 17beta-estradiol postconditioning against hypoxia/reoxygenation injury in human gastric epithelial cells.

The purpose of this study was to investigate the effects and mechanisms of 17beta-estradiol pharmacological postconditioning on gastric epithelial cells hypoxia/reoxygenation injury by using an in vitro model of human gastric epithelial cells. The model of hypoxia/reoxygenation was established with human gastric epithelial cell line. The gastric epithelial cell viability was detected by 3-(4, 5-dimethylthazol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) assays. Gastric epithelial cellular apoptosis was determined by Hoechst 33258 fluorochrome staining and flow cytometric analysis. Contents of malondialdehyde (MDA) and the activity of superoxide dismutase (SOD) were measured by Colorimetry analysis. The protein expression of Bcl-2 and Bax in different groups was determined by Western blot analyses and immunocytochemistry assay. 17beta-estradiol (10(-8), 10(-7) and 10(-6)mol/l) inhibited hypoxia/reoxygenation injury and 17beta-estradiol (10(-6)mol/l) obviously attenuated hypoxia/reoxygenation injury 3h hypoxia followed by 4h reoxygenation. 17beta-estradiol promoted gastric epithelial cell viability and inhibited the gastric epithelial cell apoptosis, and meanwhile, decreased the MDA content and increased SOD activity. The level of Bcl-2 protein was restored to the normal level by 17beta-estradiol pharmacological postconditioning. In contrast, the Bax protein level was markedly reduced by 17beta-estradiol pharmacological postconditioning. These effects of 17beta-estradiol were inhibited by pretreatment with fulvestrant. These data suggested that 17beta-estradiol seems involved in regulation of gastric hypoxia/reoxygenation injury and gastroprotection, and its protective effects were strongly related to estrogen receptor.

The protective effect of capsaicin receptor-mediated genistein postconditioning on gastric ischemia-reperfusion injury in rats.

BACKGROUND: No published study has addressed the effect of genistein postconditioning on gastric ischemia-reperfusion (GI-R) injury in rats. AIM: To examine whether capsaicin receptor-mediated genistein postconditioning protects against gastric ischemia-reperfusion injury via the PI3K/Akt signal pathway. METHODS AND RESULTS: Chloraldurat-anesthetized rats underwent occlusion of the celiac artery for 30 min, followed by 60 min of reperfusion. Based on this animal model of gastric ischemia-reperfusion injury, genistein at doses of 100, 500 or 1,000 µg/kg was administered via peripheral vein 5 min before reperfusion. The dose of 500 µg/kg was optimal for postconditioning, at which the severity of I-R-induced gastric injury significantly decreased. Immunohistochemistry also showed that gastric mucosal cell apoptosis decreased. Capsazepine (CPZ), a specific antagonist for the capsaicin receptor, was administered (1,000 µg/kg, i.v.) just before ischemia. Capsaicin (50 mg/kg, s.c.) once a day for 4 days reversed the protective effects of genistein. Reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting showed increased calcitonin gene-related peptide (CGRP) expression in genistein group but not in capsazepine or capsaicin group. CGRP inhibitor CGRP8-37 also prevented the effects of genistein in decreasing gastric mucosal injury index. In addition, PI3K inhibitor LY294002 (1.5 mg/kg) reversed the protective effect of genistein. Compared with genistein group, Western blots also demonstrated decreased Akt phosphorylation in LY294002 group. CONCLUSION: Our data suggest that capsaicin receptors mediated the protective effects of genistein postconditioning. CGRP secreted by activated capsaicin-sensitive neurons played an important role in the protective effects of genistein. PI3K/Akt pathway was also involved in the protective effects of genistein.

[The protective mechanism of N-acetylcysteine against ischemia/reperfusion induced gastric injury in rats].

The present study aimed to investigate the protective mechanism of N-acetylcysteine (NAC) against gastric ischemia /reperfusion (GI/R) injury in rats. After intravenous injection (IV) of NAC (150 mg/kg) into femoral vein, the rats were subjected to 30 min of ischemia induced by clamping the celiac artery followed by 60 min of reperfusion. After the gastric mucosal damage index (GMDI) had been calculated, gastric mucosal cell in situ apoptosis was detected by TUNEL method. The protein expression of p-ERK, p-JNK and NF-kappaB, and mRNA expression of TNF-alpha and Caspase-3 in gastric mucosa were evaluated by using Western-blot or RT-PCR, respectively. The results showed that NAC not only attenuated the GI-R injury, but also decreased gastric mucosal cellular apoptosis. Furthermore, NAC increased the protein expression of p-ERK, while inhibited protein expression of p-JNK, NF-kappaB in gastric mucosa. NAC also decreased the expression of TNF-alpha mRNA and Caspase-3 mRNA in gastric mucosa. Capsazepine (CPZ) (400 mg/kg, IV) reversed the protective effect of NAC against GI/R injury in rats. These results suggest that NAC can protect rats against GI/R injury. This protective effect is possibly mediated by the up-regulation of p-ERK and down-regulation of p-JNK and NF-kappaB. In addition, vanilloid receptor subtype 1 may be involved in the protective mechanism of NAC against GI/R injury.

[The neuroregulatory effect of cerebellar fastigial nucleus stimulation on gastric ischemia-reperfusion injury in rats].

In the present study, rat model of gastric ischemia-reperfusion (GI-R) injury was established by clamping the celiac artery for 30 min followed by 1 h of reperfusion. Subsequently, the regulatory effect of electrical stimulation of cerebellar fastigial nucleus (FN) on GI-R injury and its neural mechanisms were investigated in Sprague-Dawley rats. The results are as follows. Electrical stimulation of the cerebellar FN not only obviously attenuated the GI-R injury in an intensity-dependent manner, but also decreased the apoptosis rate of gastric mucosal cells. Chemical lesion of FN eliminated the protective effect of electrical stimulation of FN on GI-R injury. Electrical stimulation of cerebellar FN decreased both the frequency and amplitude of the discharges of greater splanchnic nerve, but it could not change the discharge of greater splanchnic nerve following the lesion of the lateral hypothalamic area (LHA). After bilateral section of the greater splanchnic nerves, electrical stimulation of the FN also attenuated the GI-R injury. Chemical lesion of the LHA reversed the protective effect of electrical stimulation of FN on GI-R injury. Electrical stimulation of FN increased the activity of superoxide dismutase (SOD), but decreased the content of malondialdehyde (MDA) in gastric mucosa under GI-R. These results indicate that the cerebellar FN may regulate GI-R injury. Therefore, the cerebellar FN is an important brain site protecting the stomach against GI-R. The LHA and greater splanchnic nerves participate in the regulatory effects of cerebellar FN stimulation on GI-R injury. In addition, antioxidation may also be involved in the protection mechanism of cerebellar FN stimulation.