Guo-Ping Chen

Endothelium-independent vasorelaxant effect of sodium ferulate on rat thoracic aorta.

AIMS This study was designed to investigate the effects of sodium ferulate (SF) on rat isolated thoracic aortas and the possible mechanisms. MAIN METHODS Isometric tension was recorded in response to drugs in organ bath. Cytosolic free Ca(2+) concentration ([Ca(2+)](i)) was measured using Fluo-3 in cultured rat aortic smooth muscle cells (RASMC). KEY FINDINGS SF (0.1-30 mM) relaxed the isolated aortic rings precontracted with phenylephrine (PE) and high-K(+) in a concentration-dependent manner with respective pD(2) of 2.7+/-0.02 and 2.6+/-0.06. Mechanical removal of endothelium did not significantly modify the SF-induced relaxation. In Ca(2+)-free solution, SF noticeably inhibited extracellular Ca(2+)-induced contraction in high-K(+) and PE pre-challenged rings, and suppressed the transient contraction induced by PE and caffeine. The vasorelaxant effect of SF was unaffected by various K(+) channel blockers such as tetraethylammonium, glibenclamide, 4-aminopyridine, and barium chloride. In addition, SF concentration-dependently reduced the contraction induced by phorbol-12-myristate-13-acetate, an activator of protein kinase C (PKC), in the absence of extracellular Ca(2+), with the pD(2) of 2.9+/-0.03. In RASMC, SF had no effect on PE- or KCl-induced [Ca(2+)](i) increase either in the presence or in the absence of external Ca(2+). SIGNIFICANCE These results indicate that SF acts directly as a non-selective relaxant to vascular smooth muscle. The direct inhibition of the common pathway after [Ca(2+)](i) increase may account for the SF-induced relaxation in Ca(2+)-dependent contraction, while the blockage of the PKC-mediated contractile mechanism is likely responsible for the SF-induced relaxation in Ca(2+)-independent contraction.

Chronic inhibition of farnesyl pyrophosphate synthase attenuates cardiac hypertrophy and fibrosis in spontaneously hypertensive rats.

Farnesyl pyrophosphate synthase (FPPS), an essential enzyme in the mevalonate pathway, was reported to be upregulated in young spontaneously hypertensive rats (SHR) when compared with Wistar-Kyoto (WKY) rats, and this was accompanied by development of left ventricular hypertrophy. Five-week-old rats were daily gavaged with vehicle or an FPPS inhibitor (alendronate, 1 or 10 mg/kg) and blood pressures was monitored by the tail-cuff method every other week. Twelve weeks of alendronate treatment attenuated the left ventricular weight to body weight ratio (LVW/BW), hydroxyproline content, collagen deposition in the interstitia, and gene expression of atrial natriuretic peptide, B-type natriuretic peptide, and procollagen type I/III in the SHR left ventricle, all of which were significantly higher in SHRs than in WKY rats. Furthermore, long-term treatment with an FPPS inhibitor significantly reduced RhoA activation, ERK phosphorylation, and TGF-beta1 expression in the SHR left ventricle, all of which were upregulated more in SHRs than in WKY rats. In conclusion, chronic treatment with an FPPS inhibitor attenuates the development of cardiac hypertrophy and fibrosis, and the suppression of ERK1/2 phosphorylation and TGF-beta1 expression with inhibition of RhoA activation may be an important mechanism.

Knockdown of farnesylpyrophosphate synthase prevents angiotensin II-mediated cardiac hypertrophy

The Rho guanosine triphosphatases (Rho GTPases) family, including RhoA, plays an important role in angiotensin II (Ang II)-mediated cardiac hypertrophy. Farnesylpyrophosphate synthase (FPPS)-catalyzed isoprenoid intermediates are vital for activation of RhoA. The present study was designed to investigate the role of FPPS in myocardial hypertrophy mediated with Ang II. First, we demonstrated that FPPS expression was elevated both in cultured neonatal cardiomyocytes (NCMs) following Ang II treatment and in the hypertrophic myocardium of 18-week-old spontaneously hypertensive rats (SHRs). Then, the importance of FPPS was assessed by RNA interference (RNAi) against FPPS in NCMs. Successful FPPS silencing in NCMs completely inhibited the hypertrophy marker genes of β-myosin heavy chain (β-MHC) and brain natriuretic peptide (BNP), as well as cell surface area. Furthermore, FPPS knockdown prevented elevated RhoA activity compared with non-silenced controls. Similarly, increased-phosphorylation of p-38 and c-Jun N-terminal kinase (JNK) mitogen-activated protein kinases (MAPK) by Ang II was attenuated. In vivo gene transfer also attenuated hypertrophic responses as indexed by left ventricular weight/body weight (LVW/BW), heart weight/body weight (HW/BW), and echocardiography, as well as expression of β-MHC and BNP mRNA in SHRs. In conclusion, FPPS with RhoA associated p-38 and JNK MAPK signaling might play an important role in Ang II-induced cardiac hypertrophy.

Inhibition of farnesyl pyrophosphate synthase attenuates angiotensin II-induced cardiac hypertrophy and fibrosis in vivo.

Farnesyl pyrophosphate synthase (FPPS), as a key branchpoint of the mevalonate pathway, catalyzes the synthesis of isoprenoid intermediates. The isoprenoid intermediates are needed for protein isoprenylation to participate in cardiac remodeling. We have previously demonstrated that both knockdown of FPPS with small interfering RNA and inhibition of FPPS by alendronate could prevent Ang II-induced hypertrophy in cultured cardiomyocytes. In this study, we evaluated the effects of FPPS inhibition in Ang II-mediated cardiac hypertrophy and fibrosis in vivo. Wild type mice were separately treated with saline, Ang II (2.88 mg/kg per day), FPPS inhibitor alendronate (0.1 mg/kg per day), or the combination of Ang II (2.88 mg/kg per day) and alendronate (0.1 mg/kg per day) for 4 weeks. The results showed that Ang II increased FPPS expression, and the increases of Ang II-induced synthesis of the isoprenoid intermediates, FPP and GGPP, were significantly inhibited by FPPS inhibitor. In the meantime, FPPS inhibition attenuated Ang II-mediated cardiac hypertrophy and fibrosis as indexed by the heart weight to body weight ratio, echocardiographic parameters, histological examinations and expression of ANP and BNP mRNA. Furthermore, it was also found that FPPS inhibitor attenuated Ang II-induced increases of RhoA activity and p-38 MAPK phosphorylation and TGF-β1 mRNA expression. In conclusion, FPPS might play an important role in Ang II-induced cardiac hypertrophy and fibrosis in vivo, at least in part through RhoA, p-38 MAPK and TGF-β1.

Expression and Clinical Significance of Ezrin in Non–Small-Cell Lung Cancer

BACKGROUND Ezrin is known to regulate cellular survival, adhesion, migration, and invasion and has been identified as 1 of the key components of tumor progression and metastasis. The purpose of this study was to evaluate the clinicopathologic and prognostic significance of ezrin expression in non-small-cell lung cancer (NSCLC). MATERIALS AND METHODS We investigated the expression pattern of ezrin immunohistochemically in 89 paraffin samples of NSCLC between January 1998 and December 2006 and conducted survival analyses. In addition, 73 frozen specimens (including tumorous and precancerous tissues) of NSCLC and 28 frozen specimens of benign pneumonic diseases collected between January 2009 and December 2009 were analyzed by Western blot and reverse transcriptase-polymerase chain reaction (RT-PCR). RESULTS In 89 paraffin samples, ezrin was expressed in 40 cases, either in the cytoplasm or on the membrane. Ezrin-positive expression was significantly associated with increased tumor stage and lymph node (LN) metastasis. The positive rate of cytoplasm expression was significantly associated with LN metastasis. Importantly, ezrin-positive expression independently predicted inferior overall survival (OS) and disease-free survival (DFS). In 73 frozen specimens of NSCLC and 28 frozen specimens of benign pneumonic diseases, the ezrin mRNA, protein, and phospho-ezrin protein expressions in tumorous tissues were higher than they were in precancerous tissues and benign pneumonic tissues. CONCLUSION These results suggested that high-level ezrin expression contributed to NSCLC progression and that phosphorylation and subcellular translocation of ezrin might be the important mechanisms. Ezrin might be a potential prognostic marker of progression in NSCLC.

Cardioprotection by polysaccharide sulfate against ischemia/ reperfusion injury in isolated rat hearts

AbstractAim:Polysaccharide sulfate (PSS) is a new type of heparinoid synthesized with alginic acid as the basic material and then by chemical introduction of effective groups. Although PSS is successfully applied in ischemic cardio-cerebrovascular disease, its effect on cardiac function after ischemia/reperfusion (I/R) injury has previously not been investigated. The aim of the present study was to investigate whether PSS can protect the heart from I/R injury and the underlying mechanism of protection.Methods:Isolated rat hearts were perfused (Langendorff) and subjected to 20 min global ischemia followed by 60 min reperfusion with Krebs Henseleit solution or PSS (0.3–100 mg/L). Myocardial contractile function was continuously recorded. Creatine kinase (CK) and lactate dehydrogenase (LDH) leakage were measured. Tumor necrosis factor-α (TNF-α) expression in cardiomyocytes was investigated. Western blot analysis for extracellular regulated kinases (ERKs), c-jun amino-terminal kinase (JNKs) and p38 mitogen-activated protein kinase (MAPK) activity was performed.Results:After I/R, cardiac contractility decreased, CK and LDH levels increased in the coronary effluent, and TNF-α expression increased in cardiomyocytes. PSS administration at concentrations of 1–30 mg/L improved cardiac contractility, reduced CK and LDH release and inhibited TNF-α production. Phosphorylated-p38MAPK (p-p38MAPK) and p-p54/p46-JNK increased in I/R rat hearts but diminished in PSS (1–30 mg/L) treated hearts. P-p44/p42-ERK levels were unchanged. In contrast, high concentrations of PSS (100 mg/L) had adverse effects that caused a worsening of heart function.Conclusion:PSS has dose-dependent cardioprotective effects on the rat heart after I/R injury. The beneficial effects may be mediated through normalization of the activity of p38 MAPK and JNK pathways as well as controlling the level of TNF-α expression.

Tissue-specific effects of atorvastatin on 3-hydroxy-3-methylglutaryl-coenzyme A reductase expression and activity in spontaneously hypertensive rats

AbstractAim:Cardiovascular remodeling is closely associated with cholesterol and is attenuated by statins. The spontaneously hypertensive rat (SHR) has a low serum cholesterol level and evident cardiovascular remodeling. The aims of the present study were to characterize the effects of atorvastatin on tissue cholesterol content and 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase expression and activity in four tissues from SHR: liver, heart, aorta and kidney.Methods:SHR and normotensive Wistar-Kyoto rats (WKY) were treated daily with atorvastatin (50 mg/kg) for 8 weeks. Cholesterol levels of serum and tissues (liver, heart, aorta and kidney) were determined by commercial enzymatic methods. Western blot analysis and high performance liquid chromatogram (HPLC) were used to assay the expression and activity of enzyme respectively.Results:Treatment with atorvastatin decreased cholesterol content and HMG-CoA reductase expression and activity in all four tissues of SHR. However, in WKY, atorvastatin only altered HMG-CoA reductase in liver, where the protein expression was upregulated but the enzyme activity was decreased.Conclusion:The present study demonstrates that the effects of atorvastatin on tissue cholesterol content and HMG-CoA reductase are strain- and tissue-specific.

Chronic inhibition of farnesyl pyrophosphate synthase improves endothelial function in spontaneously hypertensive rats

Farnesyl pyrophosphate synthase (FPPS, EC 2.5.1.10), an essential enzyme in the mevalonate pathway, catalyzes the synthesis of isoprenoid intermediates. The isoprenoid intermediates are needed for protein isoprenylation of RhoA for its function on regulation of endothelial nitric oxide synthase (eNOS). We previously reported that FPPS were upregulated in spontaneously hypertensive rats (SHR) when compared with Wistar-Kyoto rats (WKY), and this was accompanied by development of endothelial dysfunction. Five-week-old male rats were daily gavaged with vehicle or an FPPS inhibitor (alendronate, 1 or 10mg/kg). After 12-week administration of alendronate, endothelium-dependent and -independent vasorelaxation were measured in isolated aortic rings. Twelve-week of alendronate (10mg/kg/day) treatment restored the impaired endothelium-dependent vasodilation in SHR. Furthermore, long-term treatment with an FPPS inhibitor significantly suppressed RhoA activation and increased phospho-eNOS/eNOS ratio. In conclusion, chronic treatment with an FPPS inhibitor improves the endothelial function in SHR, and the upregulation of phospho-eNOS/eNOS ratio with inhibition of RhoA activation may be an important mechanism.

Effects of atorvastatin on calcium-regulating proteins: a possible mechanism to repair cardiac dysfunction in spontaneously hypertensive rats.

Previous clinical and experimental studies have demonstrated that statins, the inhibitors of 3-hydroxy-3-methylglutaryl-coenzyme A reductase, can improve left ventricular function in damaged hearts. Also, the normal expression of Ca2+ regulatory proteins is critical for efficient myocardial function. However, it is still unclear whether the beneficial effect of statins on cardiac function is associated with alterations of Ca2+ regulatory proteins. In this study, we investigated the effect of atorvastatin on cardiac function in spontaneously hypertensive rats (SHRs), focusing in particular on its impact on the expression of sarcoplasmic reticulum Ca2+-adenosine triphosphatase (SERCA2a), phospholamban (PLB) and its phosphorylated form (phosphorylated PLB), all of which are Ca2+ regulatory proteins in myocardium. SHRs showed decreases in gene expression of SERCA2a and phosphorylated PLB, and reduction in SERCA activity in the left ventricular myocardium, as well as reduced cardiac function, compared to age-matched Wistar Kyoto rats (WKYs). Furthermore, we showed that in SHRs atorvastatin preserved cardiac dysfunction accompanied by positive alterations in calcium regulatory proteins, with up-regulation in expression of SERCA2a and phosphorylated PLB, and with improvement of SERCA activity. Thus, atorvastatin has positive effects on calcium regulatory proteins, which may be one of the mechanisms of the beneficial effect of statins on cardiac function in spontaneously hypertensive rats.

Alteration of mevalonate pathway in proliferated vascular smooth muscle from diabetic mice: possible role in high-glucose-induced atherogenic process.

The proliferation of vascular smooth muscle cells (VSMCs) is one of the main features of atherosclerosis induced by high glucose. Mevalonate pathway is an important metabolic pathway that plays a key role in multiple cellular processes. The aim of this study was to define whether the enzyme expression in mevalonate pathway is changed in proliferated VSMCs during atherogenic process in diabetic mice. Diabetes was induced in BALB/c mice with streptozotocin (STZ, 50 mg/kg/day for 5 days). Induction of diabetes with STZ was associated with an increase of lesion area and media thickness after 8 and 16 weeks of diabetes. In aorta, there were overexpressions of some enzymes, including 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR), farnesyl pyrophosphate synthase (FPPS), geranylgeranyl pyrophosphate synthase (GGPPS), farnesyltransferase (FNT), and geranylgeranyltransferase-1 (GGT-1), and unchanged expression of squalene synthase (SQS) and phosphor-3-hydroxy-3-methylglutaryl-coenzyme A reductase (P-HMGR) in 8 and 16 weeks of diabetes. In vitro, VSMCs were cultured and treated with different glucose concentrations for 48 h. High glucose (22.2 mM) induced VSMC proliferation and upregulation of HMGR, FPPS, GGPPS, FNT, and GGT-1 but did not change the expressions of SQS and P-HMGR. In conclusion, altered expression of several key enzymes in the mevalonate pathway may play a potential pathophysiological role in atherogenic process of diabetes macrovascular complication.