The urinary albumin-to-creatinine ratio is a potential predictor of target lesion revascularization after percutaneous coronary intervention with coronary stents.

The association between the urinary albumin-to-creatinine ratio (UACR) and target lesion revascularization (TLR) is unknown in patients who are implanted with drug-eluting stents (DESs) or bare metal stents (BMSs) for the treatment of coronary artery disease. Of 231 Japanese patients who were implanted with DESs and/or BMSs during percutaneous coronary intervention (PCI) between July 2009 and January 2011, 118 underwent follow-up coronary angiography at 6 to 9 months after PCI; 103 were negative for qualitative tests for urine protein: 32 (31.0%)/103 patients underwent TLR for severe in-stent restenosis (ISR) and 71 did not. On the next day after admission to the hospital, first-morning-void spot urine samples were collected to calculate UACR based on urinalysis results. Pearson's product-moment correlation coefficients indicated positive associations of UACR with late loss as assessed by quantitative coronary analysis in the overall cohort, (r = +0.515, P < 0.0001), the DES subgroup (r = +0.443, P < 0.0001), and the BMS subgroup (r = +0.652, P < 0.0001). The incidence of multivessel lesions was significantly higher (P < 0.05) in the TLR group. UACR was significantly higher (P < 0.01) in the TLR group (23.88 ± 31.8 mg/gCr) than in the control group (6.29 ± 7.46 mg/gCr). Multivariate logistic regression analysis revealed UACR (odds ratio: 1.07; 95% confidence interval: 1.02-1.12; P < 0.01) to be associated with TLR. UACR was suggested to be a potential predictor of TLR required for severe ISR after PCI with coronary stents.

High-sensitivity troponin T is a prognostic marker for patients with aortic stenosis after valve replacement surgery.

BACKGROUND: Aortic stenosis (AS) is recognized as a cause of sudden cardiac death. Recently, the measurement of high-sensitivity troponin T (hs-TnT) has become possible. Several studies have clarified that hs-TnT is a marker to indicate mortality of cardiovascular diseases. OBJECTIVES: To examine whether hs-TnT can be used as a prognostic marker to predict the operative outcome of AS. METHODS: We enrolled 60 patients with AS (mean age=68.7 ± 9.6 years, male/female=30/30). Cardiac catheterization and echocardiography were performed to evaluate the severity of AS. Aortic valve replacement surgery was performed in all patients. We defined major adverse cardiac events (MACE) as composite events of heart failure, fatal arrhythmia, and all causes of death. RESULTS: We followed up the patients for 922 ± 800 days. Mean left ventricular ejection fraction was 60.0 ± 1.8%. Mean aortic valve area was 0.61 ± 0.03 cm(2). MACE occurred in 11 patients (18%), including 5 sudden cardiac deaths. We divided the patients into three groups based on the percentile of the plasma levels of hs-TnT. Kaplan-Meier curve revealed a statistically significant difference in MACE rate among the groups (log-rank test, χ(2)=13.0, p=0.002). We conducted a Cox proportional hazard analysis with a model including age, sex, estimated glomerular filtration rate, and hs-TnT tertile as explanatory variables to predict MACE. We found that hs-TnT tertile to be a significant factor to predict MACE (hazard ratio: 3.71, p=0.03). CONCLUSIONS: hs-TnT can be a prognostic marker for patients with AS after valve replacement surgery.

S100A12 as a marker to predict cardiovascular events in patients with chronic coronary artery disease.

BACKGROUND: S100A12, a calgranulin family protein released from white blood cells, is involved in inflammatory cardiovascular disease. It was hypothesized that the plasma level of S100A12 can be used to predict outcome in patients with chronic coronary artery disease (CAD). The purpose of this study was to clarify the clinical significance of S100A12 in patients with stable CAD. METHODS AND RESULTS: A total of 652 patients with stable CAD were studied. All patients underwent percutaneous coronary intervention and successful revascularization. Major adverse cardiovascular events (MACE) were defined as a composite of events of CHF, recurrence of angina pectoris, acute myocardial infarction, stroke, critical arrhythmia, intervention to peripheral arteries and cardiac death. The mean follow-up period was 973±639 days. MACE occurred in 108 patients (16.6%). Plasma S100A12 level had a significant positive correlation with high-sensitivity C-reactive protein (hs-CRP) level. On Kaplan-Meier curve analysis the incidence of MACE was significantly different among S100A12 quartiles (P=0.026). The highest S100A12 quartile (Q4) had a significantly higher MACE rate than the lowest quartile (Q1) (P=0.002). In contrast, hs-CRP was not significant for predicting MACE in the present subjects (P=0.074). A Cox proportional hazard model showed that S100A12 was an independent factor for predicting MACE in multivariate models. CONCLUSIONS: S100A12 could be a novel biomarker for predicting cardiovascular events for predicting MACE in patients with stable CAD.

Plasma endoglin as a marker to predict cardiovascular events in patients with chronic coronary artery diseases.

Recent clinical studies have revealed that the expression of endoglin, an accessory protein for the TGF-ß receptor, is increased in patients with atherosclerotic diseases. The plasma endoglin level is thought to represent endothelial activation, inflammation, and senescence. To clarify the significance of plasma endoglin in chronic coronary artery disease. Human umbilical vein endothelial cells (HUVECs) were cultured to examine changes in soluble endoglin (s-endoglin) levels caused by atherogenic stimulation in vitro. We studied 318 patients with stable coronary artery disease who underwent a successful percutaneous coronary intervention (PCI). Patients with acute coronary syndrome were excluded. Major adverse cardiovascular events (MACE) were congestive heart failure, acute myocardial infarction, stroke, and sudden cardiac death. All patients were followed-up to examine MACE after the procedure. We confirmed that the levels of s-endoglin was increased in the culture medium of HUVECs by senescence, tumor necrosis factor-α and hydrogen peroxide. In a clinical study, mean follow-up period was 1055 ± 612 days (49-2136 days) with 27 incidents of MACE (8.5%). We divided patients into three groups according to the plasma s-endoglin levels. Kaplan-Meier curves revealed that the highest endoglin group had a significantly higher MACE rate than the lowest endoglin group (log-rank test, p = 0.009). A Cox proportional hazards model showed that chronic kidney disease, left ventricular ejection fraction and s-endoglin level were significant factors to predict MACE. Plasma endoglin could be a marker to predict cardiovascular events in patients with chronic coronary artery disease after PCI.

Granzyme B as a novel factor involved in cardiovascular diseases.

Apoptosis plays an important role in cardiovascular diseases such as atherosclerosis, ischemic heart disease, and congestive heart failure. Previous studies have demonstrated that oxidative stress, physiological stress, and inflammatory cytokines such as tumor necrosis factor and Fas ligand are involved in apoptosis of cardiovascular system. We demonstrate that another apoptosis-related pathway, i.e. granzyme B/perforin system is involved in cardiovascular diseases. Expression of granzyme B, a member of serine protease family is increased in acute coronary syndrome, coronary artery disease with end-stage renal disease, and subacute stage of acute myocardial infarction. Although granzyme B is extensively researched in immunological disorders, the role of granzyme B/perforin system was not clear in the cardiovascular field. In addition, little is known regarding the inhibition of granzyme B system in the clinical situation. In this review we demonstrate recent findings of granzyme B in cardiovascular diseases and possible therapeutic applications of inhibiting the granzyme B/perforin system.

Functional molecular imaging of ILK-mediated Akt/PKB signaling cascades and the associated role of beta-parvin.

Visualization and quantification of the dynamics of protein-protein interactions in living cells can be used to explore the macromolecular events involved in signal transduction processes. In this study, functional molecular imaging using a luciferase-based complementation method demonstrated how the integrin-linked kinase (ILK)-mediated protein complex controls downstream signals. The luciferase complementation assay showed that Akt1 preferentially binds to beta-parvin rather than to ILK within the complex. Moreover, photon flux from the interaction between beta-parvin and Akt1 increased following serum stimulation, and the beta-parvin-Akt1 interaction was dependent on phosphoinositide 3-kinase. Intriguingly, small interfering (si)RNA-mediated beta-parvin knockdown increased photon flux from the interaction between ILK and Akt1, leading to stabilization of hypoxia-inducible factor-1alpha and increased expression of vascular endothelial growth factor-A. These data from functional molecular imaging demonstrated that beta-parvin plays a regulatory role in the ILK-mediated Akt (also called protein kinase B) signaling cascades, suggesting that beta-parvin might be a crucial modulator of cell survival.

Roles of ghrelin in left-ventricular remodelling after acute myocardial infarction.

OBJECTIVE: The purpose of this study was to elucidate the role of ghrelin after acute myocardial infarction (AMI) in left ventricular (LV) remodelling. DESIGN: Prospective observational study. SETTING: Jichi Medical University Hospital. PATIENTS: Fifty consecutive patients experiencing their first AMI. INTERVENTIONS: Ghrelin was measured on the day of admission, day 7, day 14 and 6 months after AMI. Patients were treated by percutaneous coronary intervention, and successful myocardial reperfusion was accomplished within 12 h after onset. To analyse LV remodelling, the authors performed left ventriculographies on the day of admission and 6 months after AMI. MAIN OUTCOME MEASURES: Changes in LV volume. RESULTS: Plasma ghrelin increased significantly after AMI (admission: 40.9±7.3; day 7: 89.5±11.0; day 14: 92.6±11.8 fmol/ml, p<0.0001). There was a significant correlation between ghrelin on day 14 and changes in LV volume over 6 months (r=+0.46, p<0.001). Multivariate regression analysis showed that ghrelin on day 14 is a significant predictor of LV remodelling after AMI (ß=+0.44, p=0.001). CONCLUSION: To our knowledge, this is the first report that shows a relation between circulating ghrelin after AMI and the progression of LV remodelling in the chronic phase. The elevation of ghrelin after AMI might be a compensatory mechanism to attenuate LV remodelling.

Plasma granzyme B as a predicting factor of coronary artery disease--clinical significance in patients with chronic renal failure.

OBJECTIVES: To elucidate the role of granzyme B in coronary artery disease (CAD) in patients with chronic kidney disease (CKD). We hypothesized that granzyme B plays an important role in the formation of coronary artery lesions in patients with CKD. PATIENTS AND METHODS: We studied 141 patients (116 men and 25 women; mean age, 64.2±9.6 years) and 16 control subjects. Diagnosis of CAD was confirmed by selective coronary angiography. CKD was defined as a sustained decrease in the estimated glomerular filtration (eGFR) rate less than 60 mL/min/1.73 m(2) over 3 months. We assigned patients to three groups: CAD without CKD (CAD group, n=46), CKD without CAD (CKD group, n=18), and CAD with CKD (CAD/CKD group, n=77). Plasma granzyme B was measured by enzyme-linked immunosorbent assay. Factors contributing to the severity of CAD were analyzed by multiple regression analysis in patients with CAD. RESULTS: Plasma levels of high-sensitivity CRP (hs-CRP) and granzyme B in the CAD/CKD group were significantly higher than in other groups. A significant positive correlation was observed between plasma hs-CRP and granzyme B levels. A significant negative correlation was observed between eGFR and granzyme B levels. Multiple regression analysis revealed that granzyme B and hs-CRP levels were independent predicting variables of the number of stenoses in major coronary arteries. CONCLUSIONS: These results indicate that granzyme B might be a novel risk factor for the formation of coronary atherosclerosis by inducing apoptosis of vascular tissues in patients with CKD.

Elevation of plasma granzyme B levels after acute myocardial infarction.

BACKGROUND: Apoptosis is reported to play an important role in left ventricular (LV) remodeling after acute myocardial infarction (AMI). Granzyme B is a member of the serine esterase family, which has an important role in cellular apoptosis and extracellular matrix degradation. METHODS AND RESULTS: Peripheral blood samples were obtained from 33 patients with a first-onset AMI treated by percutaneous coronary intervention (mean age: 61.4+/-8.7 years old) on days 1, 7 and 14 after onset. Plasma levels of tumor necrosis factor (TNF)-alpha, a soluble form of the Fas ligand (sFasL), and granzyme B were measured. TIMI grade 3 recanalization was accomplished in all patients within 12 h after onset. The LV end-diastolic volume index (LVEDVI) was calculated on day 1 and at 6 months after onset. Plasma levels of TNF-alpha, sFasL and granzyme B increased significantly on days 7 and 14 after onset of AMI. Stepwise multivariate regression analysis showed that the plasma granzyme B level on day 14 is a significant explanatory variable for changes in the LVEDVI. CONCLUSIONS: Plasma levels of granzyme B increased after AMI, which might be an important factor in the progression of late LV remodeling after AMI.

Adeno-associated virus vector-mediated systemic interleukin-10 expression ameliorates hypertensive organ damage in Dahl salt-sensitive rats.

BACKGROUND: Inflammation plays an important role in the pathogenesis of hypertension and hypertensive organ damage. Interleukin (IL)-10, a pleiotropic anti-inflammatory cytokine, exerts vasculoprotective effects in many animal models. In the present study, we examined the preventive effects of adeno-associated virus (AAV) vector-mediated sustained IL-10 expression against hypertensive heart disease and renal dysfunction in Dahl salt-sensitive rats. METHODS: We injected the rats intramuscularly with an AAV type 1-based vector encoding rat IL-10 or enhanced green fluorescent protein (EGFP) at 5 weeks of age; subsequently, the rats were fed a high-sodium diet from 6 weeks of age. RESULTS: Sustained IL-10 expression significantly improved survival rate of Dahl salt-sensitive rats compared with EGFP expression (62.5% versus 0%, p < 0.001); it also caused 26.0% reduction in systolic blood pressure at 15 weeks (p < 0.0001). Echocardiography exhibited a 22.0% reduction in hypertrophy (p < 0.0001) and a 26.3% improvement in fractional shortening (p < 0.0001) of the rat left ventricle in the IL-10 group compared to the EGFP group. IL-10 expression also caused a 21.7% decrease in the heart weight/body weight index and cardiac atrial natriuretic peptide levels. Histopathological studies revealed that IL-10 decreased inflammatory cell infiltration, fibrosis, and transforming growth factor-beta(1) levels in the failing heart. Furthermore, IL-10 expression significantly reduced urine protein excretion with increased glomerular filtration rates. CONCLUSIONS: This is the first study to demonstrate that the anti-inflammatory cytokine IL-10 has a significant anti-hypertensive effect. AAV vector-mediated IL-10 expression potentially prevents the progression of refractory hypertension and hypertensive organ damage in humans.

Redox imbalance in patients with coronary artery disease showing progression of atherosclerotic lesions.

OBJECTIVES: To clarify the relationship between changes in redox balance and the development of new coronary lesions in patients with coronary artery disease (CAD). METHODS: We studied 82 CAD patients (70 males and 12 females, mean age 61.8 +/- 9.2 years) who underwent repeated coronary angiography within 1 year after percutaneous coronary intervention. Levels of serum lipid peroxide, erythrocyte glutathione peroxidase activity, and the redox state of erythrocyte (ratio of reduced to oxidized glutathione, the GSH/GSSG ratio) were measured at the time of follow-up coronary angiography. According to the development of significant stenotic lesions, we divided the patients into two groups: 57 patients without the development of new stenotic lesions (group A) and 25 patients showing new significant stenotic lesions within 1 year (group B). RESULTS: The serum lipid peroxide level in group B was significantly higher than those of group A (2.61 +/- 0.32 vs 1.74 +/- 0.16 nmol/ml, p < 0.01). Erythrocyte glutathione peroxidase activity did not differ significantly between two groups. The erythrocyte GSH/GSSG ratio in group B was significantly lower than that of group A (83 +/- 9.6 vs 126 +/- 7.3, p < 0.01). The sensitivity and specificity of GSH/GSSG ratio to detect CAD patients with the development of significant coronary stenosis were 80.0% and 61.4%, respectively. CONCLUSIONS: CAD patients who showed development of new coronary lesions within 1 year have increased oxidative stress and imbalanced erythrocyte redox state. The GSH/GSSG ratio, an indicator for redox balance, could be a useful marker to identify high-risk CAD patients.

Determinants of exaggerated difference in morning and evening blood pressure measured by self-measured blood pressure monitoring in medicated hypertensive patients: Jichi Morning Hypertension Research (J-MORE) Study.

BACKGROUND: Morning blood pressure (BP) surge in ambulatory BP monitoring was a risk factor for stroke in our previous study. We studied the determinants of the morning minus evening systolic BP difference (ME difference) in self-measured BP monitoring, as a possible risk factor for stroke in medicated hypertensive patients. METHODS: Nine hundred sixty-nine hypertensive outpatients receiving stable antihypertensive drug treatment were studied using self-measured BP monitoring in the morning and evening. RESULTS: The ME difference ranged from -37.3 to 53.3 mm Hg (mean 7.9 mm Hg). The highest quartile (Q4) of the ME difference group (>15.0 mm Hg) had older age (68.0+/-9.8 years v 66.2+/-10.3 years, P=.01) and higher prevalence of men (48.3% v 39.9%, P=.02), regular alcohol drinkers (34.7% v 26.0%, P=.01) and beta-blocker use (26.9% v 19.9%, P=.03) than the other quartile groups (Q1 to Q3), whereas there was no significant difference in the average of morning and evening (ME average) BP. In logistic regression analysis controlling for ME average and other confounding factors, independent risks for Q4 of ME difference were older age (10 years older: odds ratio [OR] 1.21, P=.01, 95% confidence interval (CI) 1.04-1.42), regular alcohol drinker (OR 1.51, P=.04, 95% CI 1.01-2.26), and beta-blocker use (OR 1.50, P=.02, 95% CI 1.06-2.12). CONCLUSIONS: Older age, beta-blocker use, and regular alcohol drinking were significant determinants of the exaggerated ME difference in medicated hypertensive patients.

Collagen metabolism in extracellular matrix may be involved in arterial stiffness in older hypertensive patients with left ventricular hypertrophy.

Collagen metabolism in the extracellular matrix (ECM) is related to the pathogenesis of cardiovascular stiffness and remodeling in hypertension. We evaluated the association between collagen metabolism markers and the newly developed parameter, brachial-ankle pulse wave velocity (baPWV), in older hypertensive patients with left ventricular hypertrophy (LVH). We performed echocardiography and baPWV measurement using a new device, form PWV/ABI (Colin Medical Technology, Komaki, Japan), and measured plasma levels of markers of collagen metabolism such as procollagen type I C-terminal propeptide (PICP: a marker of collagen synthesis), collagen type I pyridinoline cross-linked C-terminal telopeptide (ICTP: a marker of collagen type I degradation), matrix metalloproteinase-1 (MMP-1: a marker of collagen degradation) and tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) in 46 hypertensive patients with LVH. BaPWV was correlated with the plasma level of PICP (r=0.33, p=0.03) and ICTP (r=0.29, p=0.05) and the total TIMP-1/MMP-1 ratio (an index of collagen turnover; r=0.30, p=0.04). BaPWV was negatively correlated with the E/A ratio of left ventricular inflow (r=-0.36, p<0.05), while baPWV was not correlated with left ventricular mass index (LVMI; r=-0.175, p=0.25) or deceleration time of the mitral E wave (DCT; r=0.15, p=0.31). The measures of hypertensive heart disease, such as the E/A ratio, DCT or LVMI were not correlated with any collagen markers in this study. In multiple regression analysis adjusted for confounding factors such as age, sex, pulse pressure, mean blood pressure, pulse rate, LVMI, E/A ratio and DCT, the positive correlation between baPWV and total TIMP-1/MMP-1 ratio remained significant (p<0.05). In conclusion, arterial stiffness in high-risk older hypertensive patients may involve ECM collagen metabolism.

[Relationship between redox state of whole arterial blood glutathione and left ventricular function after acute myocardial infarction].

OBJECTIVES: To investigate the role of oxidative stress in left ventricular function after acute myocardial infarction. METHODS: We studied 41 patients with acute myocardial infarction (30 men and 11 women, mean age 61.7 +/- 11.6 years) with Thrombolysis in Myocardial Infarction grade 3 recanalization of occluded coronary arteries within 12 hr after onset. Cardiac catheterization was performed at the time of admission and before discharge. Three markers for oxidative stress were measured: plasma lipid hydroperoxide, plasma creatol and whole arterial blood glutathione at the time of admission. RESULTS: Mean time from onset to recanalization was 5.2 +/- 0.6 hr. The patients were divided into two groups according to the changes in left ventricular wall motion (LVWM); patients who showed improvement in LVWM and those without improvement. There were no significant differences in age, sex, coronary risk factors, severity of coronary artery disease, time from onset to recanalization or ejection fraction between two groups. Maximum creatine kinase and C-reactive protein levels in patients without LVWM improvement were significantly higher than in patients with improvement. Plasma levels of lipid hydroperoxide and creatol did not differ significantly between two groups. On the other hand, reduced glutathione/oxidized glutathione ratio in arterial blood in patients without LVWM improvement was significantly lower than in patients with LVWM improvement (69.8 +/- 3.4 vs 85.5 +/- 2.9, p < 0.05). CONCLUSIONS: Our results suggest that whole arterial blood glutathione is more oxidized in acute myocardial infarction patients without LVWM improvement than in patients with improvement. Redox state of arterial blood can be a predicting factor for left ventricular function after acute myocardial infarction.

Adrenomedullin expression in coronary circulation after stent implantation as a prognostic factor for restenosis.

The authors determined the changes in adrenomedullin (AM) expression in the coronary circulation of patients with ischemic heart disease who underwent coronary stent implantation. Blood samples were drawn from the coronary sinus before, immediately after and four hours after coronary stent implantation, and plasma levels of AM were measured. AM levels in the coronary sinus blood were significantly increased four hours after stent implantation. On the other hand, in the femoral arterial blood, no significant changes in AM levels were observed. A significant positive correlation was found between AM level in the coronary sinus blood four hours after stent implantation and late loss index six months after the procedure. These results suggest that inflammation after coronary stent implantation induces AM expression, which might play an important role in restenosis after stenting.

GIT1 mediates Src-dependent activation of phospholipase Cgamma by angiotensin II and epidermal growth factor.

Critical events for vasoconstrictor and growth factor signal transduction include stimulation of phospholipase Cgamma (PLCgamma) and elevation of intracellular calcium. c-Src has been proposed as a common mediator for these signals activated by both G protein-coupled receptors (GPCRs) and tyrosine kinase-coupled receptors (TKRs). Here we show that the GPCR kinase-interacting protein-1 (GIT1) is a substrate for c-Src that undergoes tyrosine phosphorylation in response to angiotensin II (AngII) and EGF in vascular smooth muscle and 293 cells. GIT1 associates with PLCgamma via the PLCgamma Src homology 2 and 3 domains constitutively, and the interaction is unaltered by AngII and EGF. GIT1 interaction with PLCgamma is required for PLCgamma activation based on inhibition of tyrosine phosphorylation and calcium mobilization after GIT1 knockdown with antisense GIT1 oligonucleotides. GIT1 interacts with PLCgamma via a novel Spa homology domain (SHD) and a coiled-coil domain. Deletion mutation analysis showed that GIT1(SHD) is required for AngII- and EGF-mediated PLCgamma activation (measured by phosphorylation of Tyr783 and inositol 1,4,5-trisphosphate formation). We propose that GIT1 is a novel regulator of PLCgamma function that mediates PLCgamma activation by c-Src and integrates signal transduction by GPCRs and TKRs.

Fluid shear stress attenuates hydrogen peroxide-induced c-Jun NH2-terminal kinase activation via a glutathione reductase-mediated mechanism.

c-Jun NH2-terminal kinase (JNK) is activated by a number of cellular stimuli including reactive oxygen species (ROS). Previous studies have demonstrated that fluid shear stress (flow) inhibits cytokine-induced JNK activation in endothelial cells (ECs). In the present study, we show JNK activation by ROS in ECs and hypothesized that flow inhibits ROS-induced JNK activation in ECs via modulation of cellular protection systems against ROS. JNK was activated by 300 micro mol/L hydrogen peroxide (H2O2) in bovine lung microvascular ECs (BLMVECs) with a peak at 60 minutes after stimulation (6.3+/-1.2-fold increase). Preexposure of BLMVECs to physiological steady laminar flow (shear stress=12 dyne/cm2) for 10 minutes significantly decreased H2O2-induced JNK activation. Thioredoxin and glutathione are cellular antioxidants that protect cells against ROS. Flow induced a significant increase in the ratio of reduced glutathione to oxidized glutathione consistent with a 1.6-fold increase in glutathione reductase (GR) activity. Preincubation of BLMVECs with the GR inhibitor, 1,3 bis-(2 chloroethyl)-1-nitrosourea, abolished the inhibitory effect of flow. In contrast, preincubation of BLMVECs with azelaic acid, a specific inhibitor for thioredoxin reductase, did not alter the effect of flow on H2O2-induced JNK activation. Overexpression of GR mimicked the effect of flow to inhibit JNK activation. These results suggest that flow activates GR, an important regulator of the intracellular redox state of glutathione, and exerts a protective mechanism against oxidative stress in endothelial cells.

Protein kinase C-alpha and protein kinase C-epsilon are required for Grb2-associated binder-1 tyrosine phosphorylation in response to platelet-derived growth factor.

Grb2-associated binder-1 (Gab1) is an adapter protein related to the insulin receptor substrate family. It is a substrate for the insulin receptor as well as the epidermal growth factor (EGF) receptor and other receptor-tyrosine kinases. To investigate the role of Gab1 in signaling pathways downstream of growth factor receptors, we stimulated rat aortic vascular smooth muscle cells (VSMC) with EGF and platelet-derived growth factor (PDGF). Gab1 was tyrosine-phosphorylated by EGF and PDGF within 1 min. AG1478 (an EGF receptor kinase-specific inhibitor) failed to block PDGF-induced Gab1 tyrosine phosphorylation, suggesting that transactivated EGF receptor is not responsible for this signaling event. Because Gab1 associates with phospholipase Cgamma (PLCgamma), we studied the role of the PLCgamma pathway in Gab1 tyrosine phosphorylation. Gab1 tyrosine phosphorylation by PDGF was impaired in Chinese hamster ovary cells expressing mutant PDGFbeta receptor (Y977F/Y989F: lacking the binding site for PLCgamma). Pretreatment of VSMC with (a specific PLCgamma inhibitor) inhibited Gab1 tyrosine phosphorylation as well, indicating the importance of the PLCgamma pathway. Gab1 was tyrosine-phosphorylated by phorbol ester to the same extent as PDGF stimulation. Studies using antisense protein kinase C (PKC) oligonucleotides and specific inhibitors showed that PKCalpha and PKCepsilon are required for Gab1 tyrosine phosphorylation. Binding of Gab1 to the protein-tyrosine phosphatase SHP2 and phosphatidylinositol 3-kinase was significantly decreased by PLCgamma and/or PKC inhibition, suggesting the importance of the PLCgamma/PKC-dependent Gab1 tyrosine phosphorylation for the interaction with other signaling molecules. Because PDGF-mediated ERK activation is enhanced in Chinese hamster ovary cells that overexpress Gab1, Gab1 serves as an important link between PKC and ERK activation by PDGFbeta receptors in VSMC.

Matrix metalloproteinase expression in the coronary circulation induced by coronary angioplasty.

Recent studies have revealed that matrix metalloproteinases (MMPs) play an important role in cardiovascular remodeling by degrading the extracellular matrix. We investigated changes in the expression of MMPs due to percutaneous transluminal coronary angioplasty (PTCA). We studied 47 patients with ischemic heart disease who underwent elective PTCA on isolated stenotic lesion of left coronary arteries. Twelve patients received conventional balloon angioplasty, 14 percutaneous transluminal rotational atherectomy and 21 stent implantation. Blood samples were drawn from the coronary sinus immediately before and after, as well as 4 and 24 h, after PTCA. Plasma levels of MMP-1, MMP-2, tissue inhibitor of MMP (TIMP)-1 and TIMP-2 were measured by enzyme-linked immunosorbent assay. Plasma MMP-2 activity was determined with the digestion of a specific chromogenic peptide substrate. We could observe serial changes in plasma MMP-1 levels in the coronary circulation only in one patient, because MMP-1 levels were lower than the limit of detection in other patients. On the other hand, plasma MMP-2 levels in the coronary sinus were detectable in all subjects and increased significantly 4 and 24 h after PTCA. Plasma TIMP-1 levels also showed significant increases 4 and 24 h after PTCA, whereas TIMP-2 did not show significant changes. Plasma MMP-2/TIMP-2 ratio and MMP-2 activity in the coronary sinus showed significant increases 4 and 24 h after PTCA. A positive correlation was observed between MMP-2 levels in the coronary sinus 4 h after PTCA and late loss index 6 months after PTCA. MMP-2 levels in the coronary sinus blood were significantly higher in patients with late restenosis than in those without restenosis. PTCA induces increases in plasma MMP-2 levels and activity in the coronary circulation, which may contribute to vascular remodeling and late restenosis after PTCA.