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A thorough understanding of the circadian clock requires qualitative evaluation of circadian clock gene expression. Thus far, no simple and effective method for detecting human clock gene expression has become available. This limitation has greatly hampered our understanding of human circadian rhythm. Here we report a convenient, reliable, and less invasive method for detecting human clock gene expression using biopsy samples of hair follicle cells from the head or chin. We show that the circadian phase of clock gene expression in hair follicle cells accurately reflects that of individual behavioral rhythms, demonstrating that this strategy is appropriate for evaluating the human peripheral circadian clock. Furthermore, using this method, we indicate that rotating shift workers suffer from a serious time lag between circadian gene expression rhythms and lifestyle. Qualitative evaluation of clock gene expression in hair follicle cells, therefore, may be an effective approach for studying the human circadian clock in the clinical setting.

Statins are 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors, which are widely used to lower serum cholesterol levels in the primary and secondary prevention of cardiovascular disease. Recent experimental and clinical evidence suggests that the beneficial effects of statins may extend beyond their cholesterol-lowering effects, to include so-called pleiotropic effects. These cholesterol-independent effects include improving endothelial function, attenuating vascular and myocardial remodeling, inhibiting vascular inflammation and oxidation, and stabilizing atherosclerotic plaques. The mechanism underlying some of these pleiotropic effects is the inhibition of isoprenoid synthesis by statins, which leads to the inhibition of intracellular signaling molecules Rho, Rac and Cdc42. In particular, inhibition of Rho and one of its downstream targets, Rho kinase, may be a predominant mechanism contributing to the pleiotropic effects of statins. The aim of the present review is to provide an update on the non-cholesterol-dependent statin effects in the cardiovascular system and highlight some of the recent findings from bench to bedside to support the concept of statin pleiotropy.

Background: In addition to inhibiting cholesterol synthesis, statins (HMG-CoA reductase inhibitors) decrease the formation of isoprenoid intermediates required for the activation of key signaling pathways, including Rho/Rho kinase (ROCK). In experimental settings, statins inhibit ROCK and reverse vascular dysfunctions in atherosclerosis, independent of cholesterol reduction. It is not known whether statins inhibit ROCK activity in humans with atherosclerosis. Methods: We investigated 35 patients with stable atherosclerosis in a randomized, double-blind study comparing treatment with high-dose (80 mg/d) or low-dose (10 mg/d) atorvastatin to placebo for 28 days. Blood samples for leukocyte ROCK activity, fasting lipids, and high-sensitivity C-reactive protein (hs-CRP) were obtained on days 0, 7, 14, and 28 after randomization and change over time with the two statin treatments relative to placebo was examined. Results: Atorvastatin 80 mg/d reduced ROCK activity (p = 0.002 vs. placebo). This decline was rapid and significant within 2 weeks of treatment. The inhibition of ROCK by atorvastatin (80 mg/d) remained significant even after controlling for changes in low-density lipoprotein cholesterol (LDL-C) and triglycerides (p = 0.01). Furthermore, there was no correlation between changes in ROCK activity and changes in LDL-C (r = 0.2, p = 0.25) or triglycerides (r =0 .1,p = 0.55). There was a modest correlation between ROCK inhibition and change in hs-CRP among patients randomized to atorvastatin 80 mg/d (r = 0.6, p = 0.07). Conclusions: These first-in-man findings demonstrate that high-dose atorvastatin rapidly inhibits the proatherogenic Rho/ROCK pathway, independent of cholesterol reduction. This inhibition may contribute to the clinical benefits of statins. Rho/ROCK may provide a useful therapeutic target in patients with

Currently available modalities for the treatment of acute ischemic stroke are aimed at preserving or augmenting cerebral blood flow. Experimental evidence suggests that statins, which show 25-30% reduction of stroke incidence in clinical trials, confer stroke protection by upregulation of eNOS and increasing cerebral blood flow. The upregulation of eNOS by statins is mediated by inhibition of small GTP-binding protein RhoA. Our recent study uncovered a unique role for a Rho-family member Rac1 in stroke protection. Rac1 in endothelium does not affect cerebral blood flow. Instead, inhibition of endothelial Rac1 leads to broad upregulation of the genes relevant to neurovascular protection. Intriguingly, inhibition of endothelial Rac1 enhances neuronal cell survival through endothelium-derived neurotrophic factors, including artemin. This review discusses the emerging therapeutic opportunities to target neurovascular signaling beyond the BBB, with special emphasis on the novel role of endothelial Rac1 in stroke protection.

Wild European plum (Prunus xdomestica L.) is a rare and endangered species in China. To conserve this germplasm, a protocol for in vitro propagation of this species was developed. Nodal segments were used as the explants in our study. For the initiation of culture, B5 medium was better than Murashige and Skoog (MS) medium, which showed rapid growth of axillary bud within 3 days. Shoot organogenesis could be induced from both 6-benzylaminopurine (BAP) with α-naphthalene acetic acid (NAA) and BAP with indole-3-butyric acid (IBA) in B5 medium, but the regeneration efficiency depended on the ratio and concentration used. The higher regeneration efficiency existed in BAP:NAA=3-5:1 and BAP:IBA=1-3:1, when BAP concentration was in 0.5-1.0 mg/L. Rooting efficiency could be greatly increased with the addition of 0.4 mg/L IBA and no callus was observed on shoots in rooting media. Rooted plantlets were successfully acclimatized and grown in different transplanting medium. Plantlets in sawdust show higher survival rate than in soil and in vermiculite. This new system proved to be a practical way for germplasm conservation and for further investigation of wild European plum.

Background— Notch1 regulates binary cell fate determination and is critical for angiogenesis and cardiovascular development. However, the pathophysiological role of Notch1 in the postnatal period is not known. We hypothesize that Notch1 signaling in vascular smooth muscle cells (SMCs) may contribute to neointimal formation after vascular injury. Methods and Results— We performed carotid artery ligation in wild-type, control (SMC-specific Cre recombinase transgenic [smCre-Tg]), general Notch1 heterozygous deficient (N1 +/− ), SMC-specific Notch1 heterozygous deficient (smN1 +/− ), and general Notch3 homozygous deficient (N3 −/− ) mice. Compared with wild-type or control mice, N1 +/− and smN1 +/− mice showed a 70% decrease in neointimal formation after carotid artery ligation. However, neointimal formation was similar between wild-type and N3 −/− mice. Indeed, SMCs derived from explanted aortas of either N1 +/− - or smN1 +/− mice showed decreased chemotaxis and proliferation and increased apoptosis compared with control or N3 −/− mice. This correlated with decreased staining of proliferating cell nuclear antigen–positive cells and increased staining of cleaved caspase-3 in the intima of N1 +/− - or smN1 +/− mice. In SMCs derived from CHF1/Hey2 −/− mice, activation of Notch signaling did not lead to increased SMC proliferation or migration. Conclusions— These findings indicate that Notch1, rather than Notch3, mediates SMC proliferation and neointimal formation after vascular injury through CHF1/Hey2 and suggest that therapies that target Notch1/CHF1/Hey2 in SMCs may be beneficial in preventing vascular proliferative diseases.

Background Rho-kinase (ROCK) is a downstream effector of Rho GTPase that is known to regulate various pathological processes important to the development of ischemic stroke, such as thrombus formation, inflammation, and vasospasm. Inhibition of ROCK leads to decreased infarct size in animal models of ischemic stroke. This study tests the hypothesis that ROCK activity increases during the acute phase of ischemic stroke. Methods Serial blood samples were drawn from 10 patients with acute ischemic stroke presenting within 24 h of symptom onset and with NIHSS scores ≥ 4. Samples were taken at 24, 48, and 72 h. Leukocyte ROCK activity was determined by immunoblotting leukocyte lysates with antibodies to the phosphorylated form of myosin-binding subunit (P-MBS) of myosin light chain phosphatase (MLCP). MBS and P-MBS contents were normalized to α-tubulin, and ROCK activity was expressed as the ratio of P-MBS to MBS. ROCK activities in these 10 patients were compared to baseline ROCK activities in 10 control subjects without acute illness and matched for sex, age, and number of vascular risk factors using a two-tailed Student's t -test. Results The mean NIHSS score in patients with stroke was 15.4. ROCK activity was significantly increased at 24 and 48 h in patients after acute ischemic stroke when compared to control values, with peak elevations at 48 h after stroke onset. There was no apparent correlation between ROCK activity and stroke severity based on NIHSS. Conclusions Leukocyte ROCK activity is increased in patients after acute ischemic stroke with maximal activity occurring about 48 h after stroke onset. These findings suggest that activation of ROCK may play a role in the pathogenesis of ischemic stroke in humans.

Background— Alteration of the circadian rhythm and increased vascular senescence are linked to cardiovascular disease. Per2 , a circadian gene, is known to regulate endothelium-dependent vasomotion. However, the mechanism by which Per2 affects endothelial function is unknown. We hypothesize that endothelial dysfunction in Per2 mutant (Per2 m/m ) mice is mediated in part by increased vascular senescence and impaired endothelial progenitor cell (EPC) function. Methods and Results— Endothelial cells from Per2 m/m mice exhibit increased protein kinase Akt signaling, greater senescence, and impaired vascular network formation and proliferation. Indeed, Per2 m/m mice have impaired blood flow recovery and developed autoamputation of the distal limb when subjected to hind-limb ischemia. Furthermore, matrigel implantation into Per2 m/m mice resulted in less neovascularization. Because EPCs contribute to angiogenesis, we studied the role of Per2 in these cells using bone marrow transplantation. Basal EPC levels were similar between wild-type and Per2 m/m mice. However, compared with wild-type bone marrow transplantation mice, EPC mobilization was impaired in Per2 m/m bone marrow transplantation mice in response to ischemia or VEGF stimulation. Bone marrow transplantation or infusion of wild-type EPC restored blood flow recovery and prevented autoamputation in Per2 m/m mice. Conclusion— These findings indicate that mutation of Per2 causes Akt-dependent senescence and impairs ischemia-induced revascularization through the alteration of EPC function.

Bradykinin 2 receptor (B2R) deficiency predisposes to cardiac hypertrophy and hypertension. The pathways mediating these effects are not known. Two-month-old B2R knockout (KO) and wild-type (WT) mice were assigned to 4 treatment groups (n = 12–14/group): control (vehicle); nitro-l-arginine methyl ester (l-NAME) an NO synthase inhibitor; simvastatin (SIM), an NO synthase activator; and SIM+l-NAME. Serial echocardiography was performed and blood pressure (BP) at 6 weeks was recorded using a micromanometer. Myocardial eNOS and mitogen-activated protein kinase (MAPK, including ERK, p38, and JNK) protein expression were measured. Results showed that (i) B2RKO mice had significantly lower ejection fraction than did WT mice (61% ± 1% vs. 73% ± 1%), lower myocardial eNOS and phospho-eNOS, normal systolic BP, and higher LV mass, phospho-p38, and JNK; (ii) l-NAME increased systolic BP in KO mice (117 ± 19 mm Hg) but not in WT mice and exacerbated LV hypertrophy and dysfunction; and (iii) in KO mice, SIM decreased hypertrophy, p38, and JNK, improved function, increased capillary eNOS and phospho-eNOS, and prevented l-NAME-induced LV hypertrophy without lowering BP. We conclude that disruption of the B2R causes maladaptive cardiac hypertrophy with myocardial eNOS downregulation and MAPK upregulation. SIM reverses these abnormalities and prevents the development of primary cardiac hypertrophy as well as hypertrophy secondary to l-NAME-induced hypertension.

The anaplastic lymphoma kinase (ALK) is an oncogene product involved in hematopoietic and non-hematopoietic malignancies. Recent studies have demonstrated that nucleophosmin (NPM)-ALK, originated from the fusion of NPM and ALK genes, causes cell transformation through diverse mechanisms. Here, we show a novel mechanism by which NPM-ALK transforms lymphoid tumor cells to become resistant to glucocorticoid (GC) or dexamethasone (Dex) treatment. Transformed BaF3 cells by NPM-ALK were much more resistant to Dex compared with their parental cells, and concurrently had a constitutive activation of mammalian target of rapamycin (mTOR) signaling, as evidenced by hyperphosphorylation of its downstream effectors, p70 S6 kinase (p70S6K) and eukaryotic initiation factor 4E-binding protein 1 (4E-BP1). The mTOR inhibitor rapamycin suppressed activation of p70S6K in BaF3/NPM-ALK cells and reversed GC resistance by synergistically inhibiting mTOR signaling pathway, enhancing cell cycle arrest at G(1) phase and promoting apoptotic cell death. In conclusion, our data indicate that the ALK fusion kinase, NPM-ALK, induces GC resistance by activating mTOR signaling, and addition of mTOR inhibitors to the chemotherapeutic regimen of ALK+ lymphomas may improve the prognosis.

Although Rho-associated kinase (ROCK) activity has been implicated in cardiovascular diseases, the tissue- and isoform-specific roles of ROCKs in the vascular response to injury are not known. To address the role of ROCKs in this process, we generated haploinsufficient Rock1 (Rock1(+/-)) and Rock2 (Rock2(+/-)) mice and performed carotid artery ligations. Following this intervention, we found reduced neointima formation in Rock1(+/-) mice compared with that of WT or Rock2(+/-) mice. This correlated with decreased vascular smooth muscle cell proliferation and survival, decreased levels proinflammatory adhesion molecule expression, and reduced leukocyte infiltration. In addition, thioglycollate-induced peritoneal leukocyte recruitment and accumulation were substantially reduced in Rock1(+/-) mice compared with those of WT and Rock2(+/-) mice. To determine the role of leukocyte-derived ROCK1 in neointima formation, we performed reciprocal bone marrow transplantation (BMT) in WT and Rock1(+/-) mice. Rock1(+/-) to WT BMT led to reduced neointima formation and leukocyte infiltration following carotid ligation compared with those of WT to WT BMT. In contrast, WT to Rock1(+/-) BMT resulted in increased neointima formation. These findings indicate that ROCK1 in BM-derived cells mediates neointima formation following vascular injury and suggest that ROCK1 may represent a promising therapeutic target in vascular inflammatory diseases.

The high-operating-temperature (HOT) midwave infrared (MWIR) n-on-p detector has been pursued using the high-density vertically integrated photodiode (HDVIP®) architecture. In this device, arsenic-doped HgCdTe grown by liquid-phase epitaxy (LPE) is used, passivated on both surfaces with interdiffused CdTe. Dark current densities on these diodes as low as 2.5 mA/cm2 normalized to a 5 μm cutoff at 250 K have been demonstrated. 1/f noise at 1 Hz, measured at 250 K, is found to be 6 × 10−11 A/rHz-cm measured on a cutoff of 4.43 μm. These results agree with the theoretical predictions for the devices made.

Imide-containing siloxane-urethane copolymers (I-PU copolymers) were synthesized from 4,4'-diphenylmethane diisocyanate (MDI), OH-terminated polydimethylsiloxane (PDMS) and pyromellitic dianhydride (PMDA). The hard segment consisted of urethane and imide segments and the soft segment was PDMS. The DSC of I-PU copolymers suggests that they had a high degree of phase separation in their low hard segment contents and phase mixing in high hard segment contents. DMA indicates similar phase behaviour and shows that incorporating imide segments into the copolymers can maintain the high modulus at high temperature. The progress of degradation of I-PU copolymers was determined by TGA and standard kinetics methods and there were three stages at around 250~425 °C, 425~560 °C and 560~700 °C, respectively. The first stage was dominated by the hard-segment structure and content. Incorporating imide segments into PU raises the characteristic temperatures of degradation in the first stage. The second stage resulted from the degradation of the PDMS soft segment to form oligomers by the interchange reaction. The degradation of the imide segment leads to the third stages because the maximum rate of degradation in DTG curves increases with the imide content.

Alpha-actinins are critical components of the actin cytoskeleton. Here we show that alpha-actinins serve another important biological function by binding to and competitively inhibiting calcium-dependent activation of endothelial NOS (eNOS). Alpha-actinin-2 was found to associate with eNOS in a yeast two-hybrid screen. In vascular endothelial cells, which only express alpha-actinin-1 and -4, alpha-actinin-4 interacted and colocalized with eNOS. Addition of alpha-actinin-4 directly inhibited eNOS recombinant protein, and overexpression of alpha-actinin-4 inhibited eNOS activity in eNOS-transfected COS-7 cells and bovine aortic endothelial cells (BAECs). In contrast, knockdown of alpha-actinin-4 by siRNA increased eNOS activity in BAECs. The alpha-actinin-4-binding site on eNOS was mapped to a central region comprising the calmodulin-binding domain, and the eNOS-binding site on alpha-actinin-4 was mapped to the fourth spectrin-like rod domain, R4. Treatment of endothelial cells with a calcium ionophore, A23187, decreased alpha-actinin-4-eNOS interaction, leading to translocation of alpha-actinin-4 from plasma membrane to cytoplasm. Indeed, addition of calmodulin displaced alpha-actinin-4 binding to eNOS and increased eNOS activity. These findings indicate that eNOS activity in vascular endothelial cells is tonically and dynamically regulated by competitive interaction with alpha-actinin-4 and calmodulin.

Photoacoustic (PA) imaging has been used mainly for anatomical and functional imaging. Although functionalized nanoparticles also have been developed for PA molecular imaging, only single targeting has been demonstrated. In this study, PA imaging of multiple targets using gold nanorods is demonstrated experimentally using HER2 and CXCR4 as target molecules. The two corresponding monoclonal antibodies were conjugated to two types of gold nanorod with different aspect ratios. Gold nanorods with mean aspect ratios of 5.9 and 3.7 exhibited peak optical absorptions at 1000 and 785 nm, respectively. Appropriate selection of laser irradiation wavelength enhances PA signals by 7-12 dB and allows signals from gold nanorods corresponding to specific bindings to be distinguished. This approach potentially allows the expression levels of different oncogenes of cancer cells to be revealed simultaneously.

The Rho kinase (ROCK) isoforms, ROCK1 and ROCK2, were initially discovered as downstream targets of the small GTP-binding protein Rho. Because ROCKs mediate various important cellular functions such as cell shape, motility, secretion, proliferation, and gene expression, it is likely that this pathway will intersect with other signaling pathways known to contribute to cardiovascular disease. Indeed, ROCKs have already been implicated in the regulation of vascular tone, proliferation, inflammation, and oxidative stress. However, it is not entirely clear how ROCKs are regulated, what some of their downstream targets are, and whether ROCK1 and ROCK2 mediate different cellular functions. Clinically, inhibition of ROCK pathway is believed to contribute to some of the cardiovascular benefits of statin therapy that are independent of lipid lowering (ie, pleiotropic effects). To what extent ROCK activity is inhibited in patients on statin therapy is not known, but it may have important clinical implications. Indeed, several pharmaceutical companies are already actively engaged in the development of ROCK inhibitors as the next generation of therapeutic agents for cardiovascular disease because evidence from animal studies suggests the potential involvement of ROCK in hypertension and atherosclerosis.

Sphingosine-1-phosphate (S1P) is a lipid mediator that exerts multiple cellular functions through activation of a subfamily of G-protein-coupled receptors. Although there is evidence that S1P plays a role in the developing and adult CNS, little is known about the ability of brain parenchyma to synthesize this lipid. We have therefore analyzed the brain distribution of the enzymatic activity of the S1P synthesizing enzyme, sphingosine kinase (SPHK) [EC:2.7.1.91], as well as mRNA distribution for one of the two isoforms of this enzyme, sphingosine kinase 2. SPHK activity, measured by the conversion of [(3)H]sphingosine to [(3)H]S1P, is highest in cerebellum, followed by cortex and brainstem. Lowest activities were found in striatum and hippocampus. Sensitivity to 0.1% Triton-X suggests that this activity is accounted for by SPHK2. RT-PCR and in situ hybridization studies show that mRNA for this isoform has a distribution similar to that of SPHK activity. In vivo and in vitro ischemia increase SPHK activity and SPHK2 mRNA levels. These results indicate that SPHK2 is the predominant S1P-synthesizing isoform in normal brain parenchyma. Its heterogeneous distribution, in particular laminar distribution in cortex, suggests a neuronal localization and a possible role in cortical and cerebellar functions, in normal as well as ischemic brain.

Hg1−x Cd x Te samples of x ~ 0.3 (in the midwave infrared, or MWIR, spectral band) were prepared by molecular beam epitaxy (MBE) for fabrication into 30-μm-pitch, 256 × 256, front-side-illuminated, high-density vertically-integrated photodiode (HDVIP) focal plane arrays (FPAs). These MBE Hg1−x Cd x Te samples were grown on CdZnTe(211) substrates prepared in this laboratory; they were ~10-μm thick and were doped with indium to ~5 × 1014 cm−3. Standard HDVIP process flow was employed for array fabrication. Excellent array performance data were obtained from these MWIR arrays with MBE HgCdTe material. The noise-equivalent differential flux (NEΔΦ) operability of the best array is 99.76%, comparable to the best array obtained from liquid-phase epitaxy (LPE) material prepared in this laboratory.

Metabolic syndrome (MetS) is a cluster of atherosclerotic cardiovascular disease risk factors and is closely associated with insulin resistance and obesity (1,2). The precise diagnosis of MetS, however, is still somewhat controversial, and no serological markers are available for its detection. Nevertheless, according to the criteria of National Cholesterol Educational Program Adult Treatment Panel (ATP)-III (3), the prevalence of MetS in U.S. adults who are older than 20 years of age is approximately 24% (4). Metabolic syndrome is a growing problem worldwide, particularly in the Asian population (5). The World Health Organization Expert Consultation recommended a lower cut-off point for observed risk in Asian populations than that for non-Asian populations, varying from 22 to 25 kg/m2 (6). Some studies on migrants from Asian countries to Western countries also indicated that Asian populations might be more susceptible to the Western diet and lifestyle than white ones in developing the MetS (7,8). Patients with MetS have twice the incidence of developing new-onset diabetes compared with those without MetS (9). Metabolic syndrome, therefore, is associated with increasing risk of developing diabetes and cardiovascular disease. Several inflammatory mediators and adipocytokines, including tumor necrosis factor (TNF)-alpha (10), interleukin (IL)-6 (11), high-sensitivity C-reactive protein (hs-CRP) (12,13), and adiponectin (14) are associated with the development of insulin resistance and MetS (15,16). Despite these associations, the pathogenesis of MetS remains unknown. We recently reported that Rho kinase (ROCK) mediates the expression of plasminogen activator inhibitor-1 under hyperglycemic conditions (17). Furthermore, ROCK is upregulated under inflammatory conditions (18) and may be involved in adipocyte differentiation (19). Thus, growing evidence suggest that ROCK may contribute to the pathogenesis of MetS. However, clinical studies are lacking, which definitively link ROCK activity with MetS. Rho kinase is a serine/threonine kinase that mediates the downstream signaling of the small guanosine triphosphate-binding protein, Rho, on the actin cytoskeleton (20). ROCK consists of 2 isoforms, ROCK1 and ROCK2. In mostly animal models, the inhibition of ROCK ameliorates many cardiovascular conditions, including hypertension (21), atherosclerosis (22), myocardial fibrosis (23), and stroke (24). Furthermore, ROCK also could regulate insulin signaling and glucose metabolism through direct phosphorylation of the insulin receptor substrate (IRS)-1 (25). Thus, it is likely that ROCK plays an important role in the pathogenesis of MetS and diabetes. However, evidence is lacking showing that ROCK activity is increased in human subjects with MetS. In the current study, we measured ROCK activity in Taiwanese population with MetS and determined whether ROCK activity is an independent marker of MetS and whether it correlates with other components and risk markers of MetS.

Background—Recurrent cerebrovascular events constitute an estimated 200 000 of the 700 000 strokes reported annually in the United States, which makes secondary stroke prevention an important goal in the management of disease among patients who have experienced stroke or transient ischemic attack.Methods and Results—Various pharmacological approaches have been advocated, but the relative efficacy and safety of these regimens has remained the subject of much debate. The results of recent clinical trials on the use of antiplatelet therapy suggest that patients with a history of stroke or transient ischemic attack may constitute a population distinct from patients with coronary or peripheral vascular disease. This may be caused, in part, by the differing etiologies of stroke and the increased vulnerability of cerebral vessels to bleeding. Indeed, dual antiplatelet therapy, which has been found to be beneficial for the treatment of acute coronary syndromes and percutaneous coronary interventions, does not confer secondary stroke protection. The emerging paradigm is that some level of platelet inhibition is required for secondary stroke protection; a level beyond which increased risk of bleeding arises.Conclusion—Because the vast majority of patients with ischemic stroke have recurrent stroke or transient ischemic attack, rather than myocardial infarction, as their next event, antiplatelet therapies for these patients should be administered according to what has been shown to be efficacious for secondary stroke protection rather than for myocardial protection. Combination therapies, which provide optimal platelet inhibition as well as vascular protection, may offer the best strategy for secondary stroke protection.

Department of Cardiovascular & Renal Medicine, Saga University Faculty of Medicine, Saga, Japan Periodontitis is a chronic inflammatory disease associated with degradation of periodontal tissues and ishighly prevalent in late middle age. Statins, such as simvastatin, are pharmacologic 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) re-ductase inhibitors that inhibit cholesterol synthesis, which is important in development of arteriosclerosis.Many cardiovascular studies have suggested that statins also have anti-inflammatory effects which areindependent of cholesterol lowering. As a chronic inflammatory disease, periodontitis shares some mecha-nisms with atherosclerosis. Since oral epithelial cells are implicated in periodontal inflammation, we mea-sured simvastatin effects on cytokine [interleukin (IL)-1β, IL-6, IL-8, IL-10, IL-12p70, and tumor necrosisfactor α (TNFα)] production by cultured human epithelioid cell line KB cells in response to IL-1 α. Simvastatindecreased production, an effect reversed by adding mevalonate or geranylgeranyl pyrophosphate (GGPP),but not farnesyl pyrophosphate (FPP). Simvastatin was found to reduce NF-κB and AP-1 promoter activityin KB cells. Our results support an anti-inflammatory effect of simvastatin on human oral epithelial cells.Rec.9/29/2006, Acc.12/7/2006, pp107-111

Arteriovenous malformations are common in patients with Continuous-Flow Left Ventricular Assist Devices (LVAD) and may reflect deregulation of angiogenesis. Angiopoietin-2 (Ang-2) is a stimulator of angiogenesis produced by endothelial cells in response to thrombin. In this study, we evaluate Ang-2 expression in patients with LVAD compared with stable heart failure (HF) or orthotopic heart transplant (OHT) and test the hypothesis that elevated Ang-2 in patients with LVAD leads to increased angiogenesis. Blood samples were taken from 79 patients (n= 21 HF, 41 LVAD, 17 OHT) during routine heart catheterization. Ang-2 was measured by ELISA and thrombin was measured by Western Blot. Cultured human umbilical vein endothelial cells (HUVECs) were incubated with plasma from each patient and Ang-2 expression was measured by RT-PCR. HUVECs were then incubated on Matrigel with serum from each patient in the presence or absence of an Ang-2 blocking antibody and tube formation was measured by microscopy. Also, vena caval endothelial cells were isolated from discarded guide wires used during catheterization and Ang-2 expression was measured by quantitative immunofluorescence. In patients with LVAD vs. HF or OHT, Ang-2 was elevated in both serum (12.3±9.6, 4.5±2.1, and 4.9±2.0 ng/mL respectively, p10 ng/mL was associated with a higher 3 month risk of GI bleeding vs. Ang-2 in vitro . Therefore, Ang-2 overexpression may contribute to AVM formation in patients with LVAD.

Rationale: Severe myocardial contractile dysfunction following resuscitation from cardiac arrest (CA) is a major contributor to CA mortality. The pathophysiology and etiology of this dysfunction is not known and there are no pharmacological therapies known to improve outcomes. Previously, we demonstrated that Dynamin related protein 1 (Drp1) is activated and recruited to the mitochondria during CA and that the Drp1 inhibitor Mdivi-1 improves post CA survival. Objective: To determine the effects of CA length on myocardial and mitochondrial function. We also sought to determine the effects of Mdivi-1 on post CA outcomes. Methods and Results: Asystolic cardiac arrest (CA) was induced in mice by IV injection of 0.08 mg/g KCL. CPR begun at 4, 8, 12, and 16 minutes post-cardiac arrest had rates of return of spontaneous circulation (ROSC) of 100%(12/12), 93%(14/15), 71%(10/14), and 44% (4/9) and 2-hour survival of 100%(12/12), 67%(10/15), 50%(7/14), and 11%(1/9). Transthoracic echocardiography 15 min post-resuscitation demonstrated percent fractional shortening of 36±4% (Sham,n=6), 30±4% (4 minCA,n=11), 24±5% (8minCA,n=10), 15±2% (12minCA,n=12). In surviving animals, myocardial dysfunction persisted for 2 hours post-resuscitation, but slowly recovered to baseline by 72 hours. No evidence of myocardial necrosis, inflammation, or apoptosis was noted following resuscitation. Progressive increases in mitochondrial derived reactive oxygen species (ROS) during CA was observed by MitoSOX red myocardial tissue staining. Mitochondria isolated from 12 min CA hearts demonstrated decreased substrate coupled and uncoupled respiration. Mdivi-1, a mitochondrial inhibitor of division (fission), improved survival and neurological scores in mice following an 8 min cardiac arrest compared to controls. Conclusions: Severe, time dependent myocardial stunning (contractile dysfunction in the absence of irreversible injury) was observed following asystolic cardiac arrest. This myocardial stunning was associated with mitochondrial injury and improved by an inhibitor of Drp1. Strategies targeting ischemia/reperfusion-induced changes in mitochondrial dynamics hold promise for improving myocardial function and survival following cardiac arrest.

An innovative cooling technology, multiple magnetic vibrating fins system with only one piezoelectric actuator, is investigated for the thermal management in this study. The vibrating driving force is generated by a piezoelectric actuator and then transfer to multiple magnetic fins by the magnetic forces. The multiple magnetic vibrating fins system in this study uses only one piezoelectric actuator to drive ten fins to vibrate simultaneously which means that it not only decrease the cost but also distinctly increase the application value. In this study, the resonance frequency and the vibration amplitude of the multiple magnetic vibrating fins is investigated to optimize its thermal performance. Compare to traditional fixed fins, the experiment results show that the improved thermal performance of the vibrating fins cooling system (η) can be improved up to 34.7%, 38.5%, 37.4% and 25.9% under the pitch of vibrating fins of 6 mm, 8 mm, 10 mm, and 12 mm, respectively, with the power consumption of 0.2 W.

Previous studies proposed the multiple fans system with a piezoelectric actuator (MFPA), which combined the piezoelectric effect, magnetic effect and resonance effect to drive the passive fans vibrating simultaneously. In this study, the thermal performance of a modularized MFPA was investigated, in which the housing design was applied. Performance of the MFPA system (η) is defined to describe the improvement of the thermal performance. The results indicate that the geometry of the housing can significantly influence the thermal performance of the modularized MFPA. The results show that can be improved from 30.26 % to 37.55 % when dimensionless width number (Z*) and dimensionless length number (Y*) are decreased from 1.5 to 1.25 and from 0.92 to 0.42, separately. Moreover, in the investigation the nozzle angle (θ), the results indicate that η can be further improved to 46.61 % when θ is increased from 0 to 63.43 .

Investigations from basic biology suggest that activation of the Rho/Rho kinase pathway reduces the bioavailability of nitric oxide (NO) and thereby promotes atherosclerosis and its clinical complications. Yet, little information is available about the relationship of the Rho/Rho kinase pathway to NO bioavailability in humans with atherosclerosis. Accordingly, we determined whether inhibition of Rho kinase augments NO bioavailability and improves endothelial function in human subjects with coronary artery disease (CAD). Thirteen CAD subjects and 16 age- and sex-matched healthy controls were randomly assigned to receive the Rho kinase inhibitor, fasudil, or placebo for 1 month each in a double-blind crossover trial. Flow-mediated, endothelium-dependent and nitroglycerin-induced, endothelium-independent vasodilation were assessed by brachial artery ultrasonography. Rho kinase activity was measured in peripheral leukocytes. Fasudil increased endothelium-dependent vasodilation in CAD subjects from 9.4±1.9% to 13.4±1.9% ( P =0.001) but not in healthy controls (from 11.3±1.4% to 7.7±1.1%; P =0.07). Endothelium-independent vasodilation was not affected by fasudil in either CAD or healthy subjects. Fasudil reduced Rho kinase activity by 59±18% in CAD subjects ( P =0.001) but not in healthy subjects (by 3±6%; P =0.60). The change in endothelium-dependent vasodilation achieved with fasudil relative to placebo was inversely proportional to Rho kinase inhibition (ie, greater Rho kinase inhibition was associated with larger improvement in endothelium-dependent vasodilation) ( r =−0.48; P =0.01). These findings suggest that Rho/Rho kinase activation promotes endothelial dysfunction in humans with atherosclerosis. Inhibition of the Rho/Rho kinase pathway should provide a useful strategy to restore NO bioavailability in humans with atherosclerosis.

The research reported here integrates computational, visual, and cartographic methods to develop a geovisual analytic approach for exploring and understanding spatio-temporal and multivariate patterns. The developed methodology and tools can help analysts investigate complex patterns across multivariate, spatial, and temporal dimensions via clustering, sorting, and visualization. Specifically, the approach involves a self-organizing map, a parallel coordinate plot, several forms of reorderable matrices (including several ordering methods), a geographic small multiple display, and a 2-dimensional cartographic color design method. The coupling among these methods leverages their independent strengths and facilitates a visual exploration of patterns that are difficult to discover otherwise. The visualization system we developed supports overview of complex patterns and, through a variety of interactions, enables users to focus on specific patterns and examine detailed views. We demonstrate the system with an application to the IEEE InfoVis 2005 Contest data set, which contains time-varying, geographically referenced, and multivariate data for technology companies in the US.

Rho-kinase is a serine threonine kinase that increases vasomotor tone via its effects on both endothelium and smooth muscle. Rho-kinase inhibition reduces cerebral infarct size in wild type, but not endothelial nitric oxide synthase deficient (eNOS−/–) mice. The mechanism may be related to Rho-kinase activation under hypoxic/ischemic conditions and impaired vasodilation because of downregulation of eNOS activity. To further implicate Rho-kinase in impaired vascular relaxation during hypoxia/ischemia, we exposed isolated vessels from rat and mouse to 60 mins of hypoxia, and showed that hypoxia reversibly abolished acetylcholine-induced eNOS-dependent relaxation, and that Rho-kinase inhibitor hydroxyfasudil partially preserved this relaxation during hypoxia. We, therefore, hypothesized that if hypoxia-induced Rho-kinase activation acutely impairs vasodilation in ischemic cortex, in vivo, then Rho-kinase inhibitors would acutely augment cerebral blood flow (CBF) as a mechanism by which they reduce infarct size. To test this, we studied the acute cerebral hemodynamic effects of Rho-kinase inhibitors in ischemic core and penumbra during distal middle cerebral artery occlusion (dMCAO) in wild-type and eNOS−/– mice using laser speckle flowmetry. When administered 60 mins before or immediately after dMCAO, Rho-kinase inhibitors hydroxyfasudil and Y-27632 reduced the area of severely ischemic cortex. However, hydroxyfasudil did not reduce the area of CBF deficit in eNOS−/– mice, suggesting that its effect on CBF within the ischemic cortex is primarily endothelium-dependent, and not mediated by its direct vasodilator effect on vascular smooth muscle. Our results suggest that Rho-kinase negatively regulates eNOS activity in acutely ischemic brain, thereby worsening the CBF deficit. Therefore, rapid nontranscriptional upregulation of eNOS activity by small molecule inhibitors of Rho-kinase may be a viable therapeutic approach in acute stroke.

Many cardiovascular studies have suggested that 3-hydroxy-3-methylglutaryl co-enzyme A reductase inhibitors (statins) have anti-inflammatory effects independent of cholesterol lowering. As a chronic inflammatory disease, periodontitis shares some mechanisms with atherosclerosis. Since oral epithelial cells participate importantly in periodontal inflammation, we measured simvastatin effects on interleukin-6 and interleukin-8 production by cultured human epithelial cell line (KB cells) in response to interleukin-1α. Simvastatin decreased production, an effect reversed by adding mevalonate or geranylgeranyl pyrophosphate, but not farnesyl pyrophosphate. Simvastatin was found to reduce NF-κB and AP-1 promoter activity in KB cells. Dominant-negative Rac1 severely inhibited interleukin-1α-induced NF-κB and AP-1 promoter activity. Our results may indicate an anti-inflammatory effect of simvastatin on human oral epithelial cells, apparently involving Rac1 GTPase inhibition.

Mammalian cells respond to bacterial lipopolysaccharide (LPS) through a cognate receptor: Toll-like receptor 4 (TLR4). The signaling pathways, which link TLR4 to the proinflammatory transcription factor nuclear factor κB (NF-κB), occur through the intracellular docking proteins MyD88 and Trif. We hypothesize that unlike antigen-presenting cells, vascular endothelial cells (ECs) lack the Trif protein TRAM and are therefore incapable of eliciting Trif-dependent immune responses to LPS. Stimulation of wild-type mice with LPS leads to the activation of NF-κB in ECs and macrophages in vitro and in vivo. In contrast to macrophages, LPS did not activate endothelial NF-κB or NF-κB–dependent genes in MyD88 −/− mice, suggesting the absence of a functional Trif pathway in vascular ECs. Indeed, the Trif-dependent gene cxcl10 was not expressed in ECs after LPS stimulation. This correlated with diminished expression of the Trif accessory TIR protein TRAM in ECs. Overexpression of TRAM cDNA in ECs reconstituted LPS-induced Trif-dependent NF-κB activation and cxcl10 promoter activity. The functional absence of TRAM in vascular ECs restricts TLR4 signaling to MyD88-dependent pathway. This is in contrast to macrophages, which respond to LPS via both Trif- and MyD88-dependent pathways. These findings indicate that vascular ECs do not express the Trif-dependent gene subset. This implies that these genes may be dispensable for the endothelial response to bacterial infection and play no role in the endothelial contribution to the development of atherosclerosis.

Rho-associated kinases (ROCKs), the immediate downstream targets of RhoA, are ubiquitously expressed serine-threonine protein kinases that are involved in diverse cellular functions, including smooth muscle contraction, actin cytoskeleton organization, cell adhesion and motility, and gene expression. Recent studies have shown that ROCKs may play a pivotal role in cardiovascular diseases such as vasospastic angina, ischemic stroke, and heart failure. Indeed, inhibition of ROCKs by statins or other selective inhibitors leads to the upregulation and activation of endothelial nitric oxide synthase (eNOS) and reduction of vascular inflammation and atherosclerosis. Thus inhibition of ROCKs may contribute to some of the cholesterol-independent beneficial effects of statin therapy. Currently, two ROCK isoforms have been identified, ROCK1 and ROCK2. Because ROCK inhibitors are nonselective with respect to ROCK1 and ROCK2 and also, in some cases, may be nonspecific with respect to other ROCK-related kinases such as myristolated alanine-rich C kinase substrate (MARCKS), protein kinase A, and protein kinase C, the precise role of ROCKs in cardiovascular disease remains unknown. However, with the recent development of ROCK1- and ROCK2-knockout mice, further dissection of ROCK signaling pathways is now possible. Herein we review what is known about the physiological role of ROCKs in the cardiovascular system and speculate about how inhibition of ROCKs could provide cardiovascular benefits.

We aimed to clarify responsiveness to angiotensin (Ang) II in the porcine basilar artery and the role of Ang II receptor subtypes by functional, radioligand binding, and cell culture studies. Ang II induced more potent contractions in the proximal part than in the distal part of isolated porcine basilar arteries. The contraction induced by Ang II was inhibited by the Ang II type 1 (AT1) receptor antagonist losartan, but the Ang II type 2 (AT2) receptor antagonist PD123319 enhanced it. After removal of the endothelium, the effect of losartan remained but the effect of PD123319 was abolished. The specific binding site of [3H]Ang II on the smooth muscle membrane was inhibited by losartan, but not by PD123319. Stimulation of angiotensin II increased nitric oxide (NO) production in cultured basilar arterial endothelial cells. This production was inhibited by PD123319 and the NO synthase inhibitor L–NG-nitroarginine. These results suggest that the contraction induced by Ang II might be mediated via the activation of AT1 receptors on the basilar arterial smooth muscle cells and be modulated via the activation of AT2 receptors on the endothelial cells, followed by NO production.

Background— Rho GTPase and its downstream target, Rho-associated kinase (ROCK), have been implicated in diverse cardiovascular diseases such as cardiac hypertrophy. However, pharmacological inhibitors of ROCK are not entirely specific, nor can they discriminate between the ROCK isoforms ROCK1 and ROCK2. To determine the specific role of ROCK1 in the development of cardiac hypertrophy, we generated ROCK1 +/− haploinsufficient mice and determined whether cardiac hypertrophy and remodeling are decreased in these mice. Methods and Results— Litters of ROCK1 −/− mice on C57Bl/6 background were markedly underrepresented, suggesting lethality in utero or postnatally. ROCK1 +/− mice, however, are viable and fertile with no obvious phenotypic abnormalities. Basal blood pressure, heart rate, and cardiac dimension and function in ROCK1 +/− mice were similar to those in wild-type (WT) littermates. Infusion of angiotensin II (400 ng · kg −1 · min −1 for 28 days) or treatment with N G -nitro- l -arginine methyl ester (1 mg/mL in drinking water for 28 days) caused similar increases in systolic blood pressure, left ventricular wall thickness, left ventricular mass, ratio of heart weight to tibial length, and cardiomyocyte size in ROCK1 +/− mice and WT littermates. In contrast, perivascular fibrosis in hearts was increased to a lesser extent in ROCK1 +/− mice compared with WT littermates. This was associated with decreased expression of transforming growth factor-β, connective tissue growth factor, and type III collagen. In addition, perivascular fibrosis induced by transaortic constriction or myocardial infarction was decreased in ROCK1 +/− mice compared with WT littermates. Conclusions— These findings indicate ROCK1 is critical for the development of cardiac fibrosis, but not hypertrophy, in response to various pathological conditions and suggest that signaling pathways leading to the hypertrophic and profibrotic response of the heart are distinct.

We examined the effects of the retention of colour-changed flowers on long- and short-distance attractiveness of bumblebees and the likelihood of successive flower visits by bumblebees in Pedicularis monbeigiana. The lower lip changed colour with age from white to purple. Hand geitonogamous pollination significantly reduced seed production. No pollen limitation occurred in this species. Purple-phase flowers contributed minimally to pollinator attractiveness at long distance. The combination of less reproductive flowers with a lower amount of reward and floral colour change enabled plants to direct pollinators to reproductive, highly rewarding white flowers at close range. A high percentage of purple-phase flowers in an inflorescence was associated with a marked reduction in the frequency of successive flower visits to individual plants. We suggest floral colour change in P. monbeigiana may serve as a mechanism for enhancing inter-individual pollen transfer and reducing intra-individual pollen transfer.

Background— Elevated levels of plasminogen activator inhibitor-1 (PAI-1) are associated with myocardial infarction and stroke, especially in patients with diabetes. The induction of PAI-1 expression by hyperglycemia involves oxidative stress and protein kinase C (PKC). However, the mechanism by which hyperglycemia increases PAI-1 expression is unknown. Methods and Results— Compared with normoglycemia, exposure of human endothelial cells to hyperglycemia, but not mannitol, increased Rho-kinase activity in a time- and concentration-dependent manner. This increase was inhibited by a PKC inhibitor, GF109203X, and antioxidants N -acetylcysteine (NAC) and reduced form of glutathione (GSH). This correlated with inhibition of hyperglycemia-induced PAI-1 expression by GF109203X, NAC, and GSH. Hyperglycemia-increased PAI-1 mRNA and protein levels were inhibited by Rho-kinase inhibitors hydroxyfasudil and Y27632 and by a dominant-negative mutant of Rho-kinase. The mechanism for this inhibition occurs at the level of gene transcription because Rho-kinase inhibitors repress hyperglycemia-stimulated PAI-1 promoter activity without affecting mRNA stability. Hyperglycemia failed to stimulate Rho-kinase activity and PAI-1 expression in heterozygous ROCK I–knockout murine endothelial cells. Conclusions— Hyperglycemia stimulates Rho-kinase activity via PKC- and oxidative stress–dependent pathways, leading to increased PAI-1 gene transcription. These results suggest that inhibition of ROCK I may be a novel therapeutic target for preventing thromboembolic complications of diabetes and cardiovascular disease.

High operating temperature (HOT) detector concepts using midwave infrared (MWIR) (x∼0.3) p-type HgCdTe operating at temperatures within the thermoelectric cooler range are of significant interest at the present time. However, it is apparent that much work remains to be done in the areas of material, diode passivation, and diode formation technologies before the “holy grail” of photon detection at room temperature for all infrared wavelengths is achieved. Over the years, at DRS, we have developed a technology base for both n- and p-type HgCdTe materials parameters that are relevant to photodiode design and fabrication. This paper will discuss data that we have taken recently on minority carrier lifetime in MWIR and long wave infrared (LWIR) HgCdTe, particularly p type, and how it compares to current theories of Auger 7, radiative, and Shockley-Read recombination in this material. Extrinsic group IB (Cu, Au) and group V (arsenic) p-type dopants were used, together with group III (In) for n-type. The impact of the data on future HOT detector work is discussed.

Transmission spectra of liquid-phase epitaxy (LPE) Hg1-xCdxTe with Cd mole fractions in the range of 0.23

There is growing evidence that Rho-kinases (ROCKs), the immediate downstream targets of the small guanosine triphosphate-binding protein Rho, may contribute to cardiovascular disease. ROCKs play a central role in diverse cellular functions such as smooth muscle contraction, stress fiber formation and cell migration and proliferation. Overactivity of ROCKs is observed in cerebral ischemia, coronary vasospasm, hypertension, vascular inflammation, arteriosclerosis and atherosclerosis. ROCKs, therefore, may be an important and still relatively unexplored therapeutic target in cardiovascular disease. Recent experimental and clinical studies using ROCK inhibitors such as Y-27632 and fasudil have revealed a critical role of ROCKs in embryonic development, inflammation and oncogenesis. This review will focus on the potential role of ROCKs in cellular functions and discuss the prospects of ROCK inhibitors as emerging therapy for cardiovascular diseases.

Background— Notch signaling influences binary cell fate decisions in a variety of tissues. The Notch1 receptor is widely expressed during embryogenesis and is essential for embryonic development. Loss of global Notch1 function results in early embryonic lethality, but the cell type responsible for this defect is not known. Here, we identify the endothelium as the primary target tissue affected by Notch1 signaling. Methods and Results— We generated an endothelium-specific deletion of Notch1 using Tie2Cre and conditional Notch1 flox/flox mice. Mutant embryos lacking endothelial Notch1 died at approximately embryonic day 10.5 with profound vascular defects in placenta, yolk sac, and embryo proper, whereas heterozygous deletion had no effect. In yolk sacs of mutant embryos, endothelial cells formed a primary vascular plexus indicative of intact vasculogenesis but failed to induce the secondary vascular remodeling required to form a mature network of well-organized large and small blood vessels, which demonstrates a defect in angiogenesis. These vascular defects were also evident in the placenta, where blood vessels failed to invade the placental labyrinth, and in the embryo proper, where defective blood vessel maturation led to pericardial and intersomitic hemorrhage. Enhanced activation of caspase-3 was detected in endothelial and neural cells of mutant mice, which resulted in enhanced apoptotic degeneration of somites and the neural tube. Conclusions— These findings recapitulate the vascular phenotype of global Notch1 −/− mutants and indicate an essential cell-autonomous role of Notch1 signaling in the endothelium during vascular development. These results may have important clinical implications with regard to Notch1 signaling in adult angiogenesis.

Two series of siloxane-urethane copolymers were prepared from polydimethylsiloxane (PDMS) with a molecular weight of 1000 or 1800 which was used as a soft segment, 4,4′-diphenylmethane diisocyanate (MDI) and 1,4-butanediol (1,4-BD). Differential scanning calorimetry (DSC) demonstrated that the position (Tgs) and breadth (ΔB) of soft-segment glass transition of copolymers remained constant as the hard-segment content increased. Heat capacities at soft-segment glass transition of the copolymer (ΔCp) were 0.195∼0.411 J/g○C and heat capacities of pure PDMS (ΔCp0) were 0.571∼0.647 J/g○C, leading to the various ΔCp/ΔCp0 ratios. The ΔCp/ΔCp0 ratios decreased as the increasing of hard-segment content, showing poor phase separation. The FTIR spectrum confirmed the occurrence of hydrogen bonding in ether end-group of pure PDMS. The ether group of the soft segment led to interfacial mixing between soft and hard segments. The tan δ of the soft segment determined by dynamic mechanical testing (DMA) also identified the mixing of soft and hard segments. The mechanical properties of the copolymer were directly related to either the soft and hard segment contents or the chain lengths of soft and hard segments. The hard segment that reinforced the soft segment and interfacial thickness between soft and hard segment dominated the mechanical properties.