cAMP-induced actin cytoskeleton remodelling inhibits MKL1-dependent expression of the chemotactic and pro-proliferative factor, CCN1

AbstractElevation of intracellular cAMP concentration has numerous vascular protective effects that are in part mediated via actin cytoskeleton-remodelling and subsequent regulation of gene expression. However, the mechanisms are incompletely understood. Here we investigated whether cAMP-induced actin-cytoskeleton remodelling modulates VSMC behaviour by inhibiting expression of CCN1. In cultured rat VSMC, CCN1-silencing significantly inhibited BrdU incorporation and migration in a wound healing assay. Recombinant CCN1 enhanced chemotaxis in a Boyden chamber. Adding db-cAMP, or elevating cAMP using forskolin, significantly inhibited CCN1 mRNA and protein expression in vitro; transcriptional regulation was demonstrated by measuring pre-spliced CCN1 mRNA and CCN1-promoter activity. Forskolin also inhibited CCN1 expression in balloon injured rat carotid arteries in vivo. Inhibiting RhoA activity, which regulates actin-polymerisation, by cAMP-elevation or pharmacologically with C3-transferase, or inhibiting its downstream kinase, ROCK, with Y27632, significantly inhibited CCN1 expression. Conversely, expression of constitutively active RhoA reversed the inhibitory effects of forskolin on CCN1 mRNA. Furthermore, CCN1 mRNA levels were significantly decreased by inhibiting actin-polymerisation with latrunculin B or increased by stimulating actin-polymerisation with Jasplakinolide. We next tested the role of the actin-dependent SRF co-factor, MKL1, in CCN1 expression. Forskolin inhibited nuclear translocation of MKL1 and binding of MKL1 to the CCN1 promoter. Constitutively-active MKL1 enhanced basal promoter activity of wild-type but not SRE-mutated CCN1; and prevented forskolin inhibition. Furthermore, pharmacological MKL-inhibition with CCG-1423 significantly inhibited CCN1 promoter activity as well as mRNA and protein expression. Our data demonstrates that cAMP-induced actin-cytoskeleton remodelling regulates expression of CCN1 through MKL1: it highlights a novel cAMP-dependent mechanism controlling VSMC behaviour

The Severity of Fatty Liver Disease Relating to Metabolic Abnormalities Independently Predicts Coronary Calcification

Background. Nonalcoholic fatty liver disease (NAFLD) is one of the metabolic disorders presented in liver. The relationship between severity of NAFLD and coronary atherosclerotic burden remains largely unknown. Methods and Materials. We analyzed subjects undergoing coronary calcium score evaluation by computed tomography (MDCT) and fatty liver assessment using abdominal ultrasonography. Framingham risk score (FRS) and metabolic risk score (MRS) were obtained in all subjects. A graded, semiquantitative score was established to quantify the severity of NAFLD. Multivariate logistic regression analysis was used to depict the association between NAFLD and calcium score. Results. Of all, 342 participants (female: 22.5%, mean age: 48.7 ± 7.0 years) met the sufficient information rendering detailed analysis. The severity of NAFLD was positively associated with MRS (X2 = 6.12, trend P < 0.001) and FRS (X2 = 5.88, trend P < 0.001). After multivariable adjustment for clinical variables and life styles, the existence of moderate to severe NAFLD was independently associated with abnormal calcium score (P < 0.05). Conclusion. The severity of NAFLD correlated well with metabolic abnormality and was independently predict coronary calcification beyond clinical factors. Our data suggests that NAFLD based on ultrasonogram could positively reflect the burden of coronary calcification

Second-Hand Smoke–Induced Cardiac Fibrosis Is Related to the Fas Death Receptor Apoptotic Pathway without Mitochondria-Dependent Pathway Involvement in Rats

Exposure to environmental tobacco smoke has been epidemiologically linked to heart disease among nonsmokers. However, the molecular mechanism behind the pathogenesis of cardiac disease is unknown. In this study, we found that Wistar rats, exposed to tobacco cigarette smoke at doses of 5, 10, or 15 cigarettes for 30 min twice a day for 1 month, had a dose-dependently reduced heart weight to body weight ratio and enhanced interstitial fibrosis as identified by histopathologic analysis. The mRNA and activity of matrix metalloprotease-2 (MMP-2), representing the progress of cardiac remodeling, were also elevated in the heart. In addition, we used reverse-transcriptase polymerase chain reaction and Western blotting to demonstrate significantly increased levels of the apoptotic effecter caspase-3 in treated animal hearts. Dose-dependently elevated mRNA and protein levels of Fas, and promoted apoptotic initiator caspase-8 (active form), a molecule of a death-receptor–dependent pathway, coupled with unaltered or decreased levels of cytosolic cytochrome c and the apoptotic initiator caspase-9 (active form), molecules of mitochondria-dependent pathways, may be indicative of cardiac apoptosis, which is Fas death-receptor apoptotic-signaling dependent, but not mitochondria pathway dependent in rats exposed to second-hand smoke (SHS). With regard to the regulation of survival pathway, using dot blotting, we found cardiac insulin-like growth factor-1 (IGF-1) and IGF-1 receptor mRNA levels to be significantly increased, indicating that compensative effects of IGF-1 survival signaling could occur. In conclusion, we found that the effects of SHS on cardiomyocyte are mediated by the Fas death-receptor–dependent apoptotic pathway and might be related to the epidemiologic incidence of cardiac disease of SHS-exposed non-smokers

Riociguat treatment in patients with chronic thromboembolic pulmonary hypertension: Final safety data from the EXPERT registry

Objective: The soluble guanylate cyclase stimulator riociguat is approved for the treatment of adult patients with pulmonary arterial hypertension (PAH) and inoperable or persistent/recurrent chronic thromboembolic pulmonary hypertension (CTEPH) following Phase

The role of S-phase kinase-associated protein 2 in regulation of vascular smooth muscle cell proliferation 'in vitro' and 'in vivo'

EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Agrostaleyrodes arcanus, a New Genus and Species of Whitefly (Homoptera: Aleyrodidae) from Taiwan

Agrostaleyrodes arcanus, a new genus and species of whitefly (Homoptera: Aleyrodidae) from Taiwan. Zoological Studies 40(2): 177-186. The adult male, female, immature stages, and egg of a new genus and species of whitefly, Agrostaleyrodes arcanus Ko gen. et sp. nov. are described from Taiwan, where the species is widely distributed on Miscanthus sinensis (Gramineae). Scanning electron micrographs and figures are included with descriptions of immature stages and adults

Tilapia Piscidin 4 (TP4) Stimulates Cell Proliferation and Wound Closure in MRSA-Infected Wounds in Mice

Antimicrobial peptides (AMPs) are endogenous antibiotics that directly affect microorganisms, and also have a variety of receptor-mediated functions. One such AMP, Tilapia piscidin 4 (TP4), was isolated from Nile tilapia (Oreochromis niloticus); TP4 has antibacterial effects and regulates the innate immune system. The aim of the present study was to characterize the role of TP4 in the regulation of wound closure in mice and proliferation of a keratinocyte cell line (HaCaT) and fibroblast cell line (Hs-68). In vitro, TP4 stimulated cell proliferation and activated collagen I, collagen III, and keratinocyte growth factor (KGF) gene expression in Hs-68 cells, which induces keratin production by HaCaT cells. This effect was detectable at TP4 concentrations of 6.25 µg/mL in both cell lines. In vivo, TP4 was found to be highly effective at combating peritonitis and wound infection caused by MRSA in mouse models, without inducing adverse behavioral effects or liver or kidney toxicity. Taken together, our results indicate that TP4 enhances the survival rate of mice infected with the bacterial pathogen MRSA through both antimicrobial and wound closure activities mediated by epidermal growth factor (EGF), transforming growth factor (TGF), and vascular endothelial growth factor (VEGF). The peptide is likely involved in antibacterial processes and regulation of tissue homeostasis in infected wounds in mice. Overall, these results suggest that TP4 may be suitable for development as a novel topical agent for wound dressing

Rho GTPase, Rac1, regulates Skp2 levels, vascular smooth muscle cell proliferation, and intima formation in vitro and in vivo

Aims Vascular smooth muscle cell (VSMC) proliferation contributes to intima formation after angioplasty or venous by-pass grafting, and during atherosclerosis. VSMC proliferation requires degradation of p27Kip1 promoted by S-phase kinase-associated protein-2 (Skp2), an F-box protein component of the Skp-Cullin-F-boxSkp2 ubiquitin-ligase. We investigated the role of Rac1 in the regulation of Skp2 in rat VSMC. Methods and results Rat carotid balloon injury increased Rac1 activity. Rho GTPase inhibition with Clostridium difficile Toxin B or specific Rac1 inhibition with adenovirus-mediated expression of domi-nant-negative Rac1 reduced Skp2 levels, and VSMC proliferation in vitro and intima formation in vivo following carotid balloon injury. Inhibition of Skp2 expression and proliferation by dominant-negative Rac1 was reversed by exogenous Skp2. Elevation of endogenous adenosine 30,50-cyclic monophosphate (cAMP) with forskolin-inhibited Rac1 activity, reduced Skp2, increased p27 Kip1 and inhibited VSMC pro-liferation, effects that were reversed by constitutively active Rac1. These effects were independent of Rac1 Cdc42/Rac interactive binding (CRIB)-domain effector proteins but associated with Rac1-dependent actin polymerization. Conclusion Rac1 activity regulates VSMC proliferation by controlling Skp2 levels. Activation of Rac1 induced by balloon injury in vivo increases Skp2 levels, which promotes VSMC proliferation and intima formation. Inhibition of this novel pathway underlies the negative effects of cAMP on VSMC proliferation

Enhanced Control of Bladder-Associated Tumors Using Shrimp Anti-Lipopolysaccharide Factor (SALF) Antimicrobial Peptide as a Cancer Vaccine Adjuvant in Mice

Shrimp anti-lipopolysaccharide factor (SALF) is an antimicrobial peptide with reported anticancer activities, such as suppression of tumor progression. In this study, we prepared a potential cancer vaccine comprised of SALF in conjunction with the cell lysate of inactivated murine bladder carcinoma cells (MBT-2), and evaluated its efficacy in a mouse tumor model. Our study shows that SALF added to cell culture media inhibits growth progression of MBT-2, and that SALF together with inactivated MBT-2 lysate elevates the level of inflammasome activity, and modulates the levels of IL-1β, MCP-1, IL-6, IL-12, and TNF-α in mouse macrophages. Immunization of 7, 14, and 21 day-old mice with the vaccine prevented growth of MBT-2 cell-mediated tumors. The vaccine was found to enhance expression of T-cell, cytotoxic T cells, and NK cells in the immunized mice groups. Recruitment of macrophages, T-helper cells, and NK cells was enhanced, but levels of VEGF were decreased in immunized mice. This report provides empirical evidence that our SALF as vaccine adjuvant enhances antitumor immunity in mice

Beyond Oncogenesis: The Role of S-Phase Kinase-Associated Protein-2 (SKP2) In Vascular Restenosis

The clinical benefits of percutaneous coronary intervention, the most prevalent procedure nowadays for the treatment of symptomatic coronary artery disease, are frequently offset by the occurrence of vascular restenosis. Although the introduction of drug-eluting stents has significantly reduced restenotic rates, the rare, but potentially fatal, delayed thrombosis remains a clinical threat. Further refinement of the drug-eluting stent based on a better understanding of cell cycle regulation between the vascular smooth muscle cell (VSMC) and endothelial cell (EC) is required. In this review, we discuss the role of S-phase kinase-associated protein-2 (Skp2), previously known as an oncoprotein, in the regulation of VSMC proliferation and its signaling axis. The currently available evidence suggests that the Rac1-Skp2-p27Kip1 signaling axis acts as a common final pathway for many factors that regulate VSMC proliferation, such as growth factors, extracellular matrices and cyclic nucleotides. Importantly, although EC proliferation is also shown to be regulated by the same axis, cAMP seems to regulate this axis differentially between VSMC and EC, rendering the underlying mechanism of this differential regulation a promising target for the development of a new generation of drug-eluting stent