Xiao-Fei Lv

Downregulation of TMEM16A Calcium-Activated Chloride Channel Contributes to Cerebrovascular Remodeling During Hypertension by Promoting Basilar Smooth Muscle Cell Proliferation

Background— The Ca2+-activated chloride channel (CaCC) plays an important role in a variety of physiological functions. In vascular smooth muscle cells, CaCC is involved in the regulation of agonist-stimulated contraction and myogenic tone. The physiological functions of CaCC in blood vessels are not fully revealed because of the lack of specific channel blockers and the uncertainty concerning its molecular identity. Methods and Results— Whole-cell patch-clamp studies showed that knockdown of TMEM16A but not bestrophin-3 attenuated CaCC currents in rat basilar smooth muscle cells. The activity of CaCC in basilar smooth muscle cells isolated from 2-kidney, 2-clip renohypertensive rats was decreased, and CaCC activity was negatively correlated with blood pressure (n=25; P<0.0001) and medial cross-sectional area (n=24; P<0.0001) in basilar artery during hypertension. Both upregulation of CaMKII activity and downregulation of TMEM16A expression contributed to the reduction of CaCC in the hypertensive basilar artery. Western blot results demonstrated that angiotensin II repressed TMEM16A expression in basilar smooth muscle cells (n=6; P<0.01). Knockdown of TMEM16A facilitated and overexpression of TMEM16A inhibited angiotensin II–induced cell cycle transition and cell proliferation determined by flow cytometry and BrdU incorporation (n=6 in each group; P<0.05). TMEM16A affected cell cycle progression mainly through regulating the expression of cyclin D1 and cyclin E. Conclusions— TMEM16A CaCC is a negative regulator of cell proliferation. Downregulation of CaCC may play an important role in hypertension-induced cerebrovascular remodeling, suggesting that modification of the activity of CaCC may be a novel therapeutic strategy for hypertension-associated cardiovascular diseases such as stroke.

Downregulation of FAP suppresses cell proliferation and metastasis through PTEN/PI3K/AKT and Ras-ERK signaling in oral squamous cell carcinoma

It is largely recognized that fibroblast activation protein (FAP) is expressed in cancer-associated fibroblasts (CAFs) of many human carcinomas. Furthermore, FAP was recently also reported to be expressed in carcinoma cells of the breast, stomach, pancreatic ductal adenocarcinoma, colorectum, and uterine cervix. The carcinoma cell expression pattern of FAP has been described in several types of cancers, but the role of FAP in oral squamous cell carcinoma (OSCC) is unknown. The role of endogenous FAP in epithelium-derived tumors and molecular mechanisms has also not been reported. In this study, FAP was found to be expressed in carcinoma cells of OSCC and was upregulated in OSCC tissue samples compared with benign tissue samples using immunohistochemistry. In addition, its expression level was closely correlated with overall survival of patients with OSCC. Silencing FAP inhibited the growth and metastasis of OSCC cells in vitro and in vivo. Mechanistically, knockdown of FAP inactivated PTEN/PI3K/AKT and Ras-ERK and its downstream signaling regulating proliferation, migration, and invasion in OSCC cells, as the inhibitory effects of FAP on the proliferation and metastasis could be rescued by PTEN silencing. Our study suggests that FAP acts as an oncogene and may be a potential therapeutic target for patients with OSCC.

Involvement of Chk1-Cdc25A-cyclin A/CDK2 pathway in simvastatin induced S-phase cell cycle arrest and apoptosis in multiple myeloma cells.

Statins have been demonstrated to effectively inhibit proliferation and induce apoptosis in cancer cells by inhibition of geranylgeranylation, however its novel molecular mechanism remains to be determined. Recently simvastatin has been found to result in the synergistic induction of apoptosis with 7-hydroxystaurosporine (UCN-01) (a Chk1 inhibitor) in myeloma cells. Therefore we hypothesized that Chk1 plays a role in the anti-myeloma effect of simvastatin. Interestingly, we found that simvastatin caused a dose-dependent increase in S phase cell cycle and induced significant apoptosis. The results of western blot showed that simvastatin-induced S-phase cell cycle arrest was associated with activation of Chk1, downregulation of Cdc25A, cyclin A and CDK2 expression. Additionally, simvastatin-induced apoptosis was accompanied by diminished Bcl-2 protein expression, increased cytosolic cytochrome c level, and activation of caspase 9 and caspase 3. Further investigation revealed that silence of Chk1 expression by Chk1 specific siRNA inhibited simvastatin-induced activation of Chk1, downregulation of Cdc25A, cyclin A and CDK2 expression, and diminished S phase cell cycle arrest. Additionally, inhibition of Chk1 expression enhanced simvastatin-induced downregulation of Bcl-2, caspase 9 cleavage and subsequent apoptosis. These results suggested that the Chk1-Cdc25A-cyclin A/CDk2 pathway was involved in simvastatin-induced S-phase cell cycle arrest and apoptosis in multiple myeloma cell lines.

Regional homogeneity changes in heroin-dependent individuals: resting-state functional MR imaging study.

PURPOSE To identify heroin-related modulations of neural activity in the resting state in heroin-dependent individuals (HDIs) by using resting-state functional magnetic resonance (MR) imaging and a regional homogeneity method and to investigate whether these changes of neural activity can be related to duration of heroin use and to decision-making deficits in HDIs. MATERIALS AND METHODS This prospective study was approved by the appropriate ethics committee, and written informed consent was obtained from each participant. Thirty-one HDIs receiving methadone-maintained treatment and 24 control subjects participated. Resting-state functional MR imaging was performed by using a gradient-echo echo-planar imaging sequence. Regional homogeneity was calculated by using software. Voxel-based analysis of the regional homogeneity maps between control and HDI groups was performed with two-sample t tests by using software. Statistical maps were set at P less than .05 and were corrected for multiple comparisons. The Iowa gambling task (IGT) was used to assess participant decision making during uncertainty. Abnormal clusters revealed by group comparison were extracted and correlated with behavioral performance at the IGT and with duration of heroin use. RESULTS Regional homogeneity was diminished in the bilateral medial orbitofrontal cortex (OFC), bilateral dorsal medial thalamus, bilateral cuneus, and lingual gyrus in HDIs compared with control subjects. There were negative correlations between mean regional homogeneity in the medial OFC, bilateral cuneus, and lingual gyrus and duration of heroin use. There was a positive correlation between mean regional homogeneity in the medial OFC and performance level at the IGT. CONCLUSION The present study reveals resting-state abnormalities in HDIs that may lead to further improvement of the understanding of the neural substrates of cognitive impairment in HDIs.

Decrease of Intracellular Chloride Concentration Promotes Endothelial Cell Inflammation by Activating Nuclear Factor-κB Pathway

Recent evidence suggested that ClC-3 channel/antiporter is involved in regulation of nuclear factor (NF)-&kgr;B activation. However, the mechanism explaining how ClC-3 modulates NF-&kgr;B signaling is not well understood. We hypothesized that ClC-3-dependent alteration of intracellular chloride concentration ([Cl−]i) underlies the effect of ClC-3 on NF-&kgr;B activity in endothelial cells. Here, we found that reduction of [Cl−]i increased tumor necrosis factor-&agr; (TNF&agr;)-induced expression of intercellular adhesion molecule 1 and vascular cell adhesion molecule 1 and adhesion of monocytes to endothelial cells (P<0.05; n=6). In Cl− reduced solutions, TNF&agr;-evoked I&kgr;B kinase complex &bgr; and inhibitors of &kgr;B&agr; phosphorylation, inhibitors of &kgr;B&agr; degradation, and NF-&kgr;B nuclear translocation were enhanced. In addition, TNF&agr; and interleukin 1&bgr; could activate an outward rectifying Cl− current in human umbilical vein endothelial cells and mouse aortic endothelial cells. Knockdown or genetic deletion of ClC-3 inhibited or abolished this Cl− conductance. Moreover, Cl− channel blockers, ClC-3 knockdown or knockout remarkably reduced TNF&agr;-induced intercellular adhesion molecule 1 and vascular cell adhesion molecule 1expression, monocytes to endothelial cell adhesion, and NF-&kgr;B activation (P<0.01; n=6). Furthermore, TNF&agr;-induced vascular inflammation and neutrophil infiltration into the lung and liver were obviously attenuated in ClC-3 knockout mice (P<0.01; n=7). Our results demonstrated that decrease of [Cl−]i induced by ClC-3-dependent Cl− efflux promotes NF-&kgr;B activation and thus potentiates TNF&agr;-induced vascular inflammation, suggesting that inhibition of ClC-3-dependent Cl− current or modification of intracellular Cl− content may be a novel therapeutic approach for inflammatory diseases.

Simvastatin Ameliorates Rat Cerebrovascular Remodeling During Hypertension via Inhibition of Volume-Regulated Chloride Channel

Statins have pleiotropic actions against the development of vascular remodeling and the incidence of ischemic stroke. Although previous studies have suggested that posttranslational modification of several proteins, such as Rho by mevalonate-derived isoprene groups, geranylgeranyl pyrophosphate or farnesyl pyrophosphate, underlie the pleiotropic effects of statins, the detailed mechanisms remain elusive. Recent growing evidence demonstrated that ClC-3 volume-regulated chloride channel plays an important role in cell proliferation, and the activity of this channel is increased in basilar smooth muscle cells from a hypertensive rat. We hypothesized that inhibition of volume-regulated chloride channel may contribute to the beneficial effects of statins on cerebrovascular remodeling during hypertension. Our study here demonstrated that simvastatin ameliorated hypertension-caused cerebrovascular remodeling. In rat basilar smooth muscle cells, simvastatin inhibited cell proliferation and activation of volume-regulated chloride channel, and these effects of simvastatin were abolished by pretreatment with mevalonate or geranylgeranyl pyrophosphate. In addition, Rho A inhibitor C3 exoenzyme and Rho kinase inhibitor Y-27632 both reduced cell proliferation and activation of volume-regulated chloride channel. Moreover, ClC-3 overexpression decreased the suppressive effect of simvastatin on cell proliferation and increased estimated IC50 of simvastatin on endothelin 1- and hypo-osmolarity-induced cell proliferation from 3.40±0.08 and 3.50±0.10 &mgr;mol/L to 5.30±0.70 and 5.60±0.70 &mgr;mol/L, respectively (P<0.01; n=6). Furthermore, the expression of ClC-3 was increased in basilar artery during hypertension, and simvastatin normalized the upregulation of ClC-3. Our data suggested that simvastatin ameliorates cerebrovascular remodeling in the hypertensive rat through inhibition of vascular smooth muscle cell proliferation by suppression of volume-regulated chloride channel.

Relationship between lipid profiles and plasma total homocysteine, cysteine and the risk of coronary artery disease in coronary angiographic subjects

BackgroundHomocysteine and cysteine are considered as risk factors of cardiovascular disease. Homocysteine influences the liver expression of ApoA-I and decreases its blood level and HDL in genetic mice model. We aimed therefore to evaluate whether homocysteine and cysteine are associated with lipid parameters, and the joint effects of them on the risk of coronary artery disease (CAD). Plasma total homocysteine (tHcy), cysteine (tCys) and lipid markers were measured in 2058 consecutive coronary artery angiographic patients.ResultsPlasma tHcy but not tCys correlated negatively with ApoA-I (r = -0.153, P < 0.001) and with HDL cholesterol (r = -0.148, P < 0.001), and correlated positively with the risk of CAD (OR: 1.61; 95% confidence interval; 1.26 to 2.05). Combination of high tHcy and high tCys levels was associated with decreased ApoA-I and HDL cholesterol levels, and with increased risk of CAD (OR: 1.696, 95% CI (1.301-2.211)). Furthermore, low HDL cholesterol combined with low tHcy or high tHcy all had increased risk for CAD (OR: 1.254, 95% CI (1.114-1.565); OR: 1.332, 95% CI (1.093-1.624); respectively) whereas high HDL cholesterol counteracted the harmful effect of high tHcy on the risk of CAD. However, only the combination of high tHcy and high ApoA-I had an increased risk for CAD (OR: 1.438, 95% CI (1.170-1.768)).ConclusionsThe association of homocysteine and cysteine, ApoA-I or HDL cholesterol and their joint effects provide new insights on its role on CAD.

Progressive white matter microstructure damage in male chronic heroin dependent individuals: a DTI and TBSS study.

Background To investigate the WM microstructure deficits in heroin dependent individuals (HDIs) with different length of heroin dependence, and to investigate whether these WM deficits can be related to the duration of heroin use and to decision-making deficits in HDIs. Methodology/Principal Findings Thirty-six HDIs [including eighteen sHDIs (duration of heroin dependent is less than 10 years) and eighteen lHDIs (duration of dependent is between 10∼20 years)] and sixteen healthy controls participated in this study. Whole brain voxel-wise analysis of fractional anisotropy (FA), mean diffusivity (MD), axial diffusivity (Da) and radial diffusivity (Dr) were performed by tract-based spatial statistics (TBSS) to localize abnormal WM regions among groups. TBSS demonstrated that sHDIs had significantly lower FA than controls in right orbito-frontal WM, bilateral temporal WM and right parietal WM. The lHDIs had significantly lower FA throughout the brain compared with the controls and sHDIs. The lHDIs had significantly lower Da than controls in bilateral inferior frontaloccipital fasciculus, bilateral splenium of corpus callosum, left inferior longitudinal fasciculus, and had significantly higher Dr than controls in bilateral uncinatus fasciculus, bilateral inferior frontaloccipital fasciculus and bilateral cortical spinal fasciculus. Volume-of-interest (VOI) analyses detect the changes of diffusivity indices in the regions with FA abnormalities revealed by control vs sHDIs. In most VOIs, FA reductions were caused by the increase in Dr as well as the decrease in Da. Correlation analysis was used to assess the relationship between FA and behavioral measures in HDIs and controls available. Significantly positively correlations were found between the FA values in the right orbital-frontal WM, right parietal WM and IGT performance. Conclusions The extent and severity of WM integrity deficits in HDIs was associated with the length of heroin dependent. Furthermore, abnormal WM microstructure may correlate with decision-making impairments in HDIs.

ClC-3 chloride channel prevents apoptosis induced by hydrogen peroxide in basilar artery smooth muscle cells through mitochondria dependent pathway

ClC-3 Cl− channel plays an important role in cell volume regulation and cell cycle. In vascular smooth muscle cells, we have found that ClC-3 was involved in ET-1 induced cell proliferation. The present study was designed to further investigate the role of ClC-3 Cl− channel in H2O2-induced apoptosis and its underlying mechanisms in rat basilar arterial smooth muscle cell (BASMCs). By using ClC-3 cDNA and small interference RNA (siRNA) transfection strategy, it was found that overexpression of ClC-3 significantly decreased the apoptotic rate of H2O2-treated BASMCs and increased the cell viability, whereas silencing of ClC-3 with siRNA produced opposite effects and increased the apoptotic rate. ClC-3 overexpression decreased cytochrome C release and caspase-3 activation, and increased both the stability of mitochondrial membrane potential and the ratio of Bcl-2/Bax, whereas silencing of ClC-3 produced opposite effect. Furthermore, we demonstrated that overexpression of ClC-3 attenuated, whereas silencing of ClC-3 facilitated, the degradation of LaminA, one of the structural matrix proteins, in BASMCs. Our data suggest that ClC-3 Cl− channel can modulate H2O2-induced apoptosis in BASMCs via the intrinsic, mitochondrial pathway.

The impulsivity behavior is correlated with prefrontal cortex gray matter volume reduction in heroin-dependent individuals

PURPOSE Chronic exposure to heroin induced cerebral structural abnormalities may underlie heroin-related behaviors. The aim of this study was to: (1) identify cerebral structural abnormalities in heroin-dependent individuals (HDIs) by an automated and unbiased morphometric technique. (2) Define the correlation between these cerebral structural abnormalities and the impulsivity characteristic in HDIs. METHODS 24 HDIs and 24 control subjects were completed with (1) high resolution structural magnetic resonance imaging scanning and analysis of gray matter volume using voxel-based morphometry implemented in Statistical Parametric Mapping and (2) a Chinese translation Barratt Impulsiveness Scale-11 questionnaire survey. Differences in regional gray matter volume were tested using an analysis of covariance model, co-varying for global gray matter and age. Statistical maps were set at p<0.05, corrected for multiple comparisons. The abnormal brain regions were correlated with the duration of heroin use and impulsivity scores. RESULTS After adjusting for effects of age and total gray matter volume, cortical gray matter volume in the bilateral medial prefrontal cortex, bilateral dorsal lateral prefrontal cortex, and right fusiform cortex were significantly reduced in HDIs. Moreover, the gray matter volume in prefrontal cortex that showed group differences was negatively correlated with the duration of heroin use and negatively correlated with the impulsivity characteristic in HDIs. CONCLUSION These findings reveal the prefrontal cortex was impaired in HDIs, meanwhile, indicate the changes in gray matter volume are relating to the duration of heroin use and the impulsivity characteristic of the HDIs.