Zhi-Han Tang

Hydrogen sulfide, the next potent preventive and therapeutic agent in aging and age-associated diseases.

ABSTRACT Hydrogen sulfide (H2S) is the third endogenous signaling gasotransmitter, following nitric oxide and carbon monoxide. It is physiologically generated by cystathionine-γ-lyase, cystathionine-β-synthase, and 3-mercaptopyruvate sulfurtransferase. H2S has been gaining increasing attention as an important endogenous signaling molecule because of its significant effects on the cardiovascular and nervous systems. Substantial evidence shows that H2S is involved in aging by inhibiting free-radical reactions, activating SIRT1, and probably interacting with the age-related gene Klotho. Moreover, H2S has been shown to have therapeutic potential in age-associated diseases. This article provides an overview of the physiological functions and effects of H2S in aging and age-associated diseases, and proposes the potential health and therapeutic benefits of H2S.

PCSK9 siRNA suppresses the inflammatory response induced by oxLDL through inhibition of NF-κB activation in THP-1-derived macrophages.

Proprotein convertase subtilisin/kexin 9 (PCSK9), a member of the protein-converting enzyme family, is highly expressed in adult hepatocytes and small intestinal enterocytes. To our knowledge, in this study, we demonstrate for the first time that PCSK9 is upregulated in a dose-dependent manner via oxidized low-density lipoprotein (oxLDL) stimulation in THP-1-derived macrophages. PCSK9 small interfering RNA (siRNA) suppresses the oxLDL-induced inflammatory cytokine expression in THP-1-derived macrophages. The exposure of macrophages to oxLDL markedly increased the expression of NF-κB protein in the nucleus. However, this effect was significantly attenuated by PCSK9 siRNA. These findings indicate that PCSK9 expression is induced by oxLDL, and that PCSK9 siRNA protects against inflammation via the inhibition of NF-κB activation in oxLDL-stimulated THP-1-derived macrophages. Our results suggest that PCSK9 may be used as a therapeutic target for the treatment of atherosclerosis since PCSK9 siRNA suppresses oxLDL-induced IκB-α degradation and NF-κB nuclear translocation into THP-1-derived macrophages.

Hydrogen sulfide prevents H₂O₂-induced senescence in human umbilical vein endothelial cells through SIRT1 activation.

The aim of the present study was to investigate the attenuation of endothelial cell senescence by H2S and to explore the mechanisms underlying the anti-aging effects of H2S. Senescence was induced in human umbilical vein endothelial cells (HUVECs) by incubation in 25 µmol/l H2O2 for 1 h. Senescence-associated β-galactosidase (SA-β-gal) activity was examined to determine the effects of H2S on senescent HUVECs. The results indicated that SA-β-gal activity in the H2O2-treated HUVECs was 11.2 ± 1.06%, which was attenuated in the NaHS group. Pretreatment with nicotinamide (NAM), a sirtuin 1 (SIRT1) inhibitor, inhibited the reduction in senescence associated with H2S. Immunoblot analyses revealed that SIRT1 levels in senescent HUVECs treated with NaHS (60 µM) were indistinguishable from controls; however, analyses of SIRT1 activity indicated that SIRT1 enzyme activity was enhanced. In addition, we found that H2S improves the function of senescent HUVECs. The present study demonstrated that H2S protects against HUVEC senescence, potentially through modulation of SIRT1 activity. Furthermore, this study establishes a novel endothelial protective effect of H2S.

SIRT1 in cardiovascular aging

Cardiovascular disease (CVD) is the leading cause of death worldwide, with aging as the key independent risk factor. Effective interventions are necessary to delay aging. Sirtuin1 (SIRT1), a NAD(+)-dependent histone deacetylase, is closely related to lifespan extension. SIRT1 exerts beneficial effects on aging and age-related diseases, such as atherosclerosis. In this review, we summarize the current knowledge on the functions of SIRT1 in cardiovascular aging, focusing on the underlying molecular mechanisms, including inhibition of oxidative stress and inflammation, and induction of autophagy. We also demonstrate that moderate up-regulation or activation of SIRT1 in cardiovascular aging and age-related CVD may confer important application values.

Cathepsin L stimulates autophagy and inhibits apoptosis of ox-LDL-induced endothelial cells: Potential role in atherosclerosis

The activation of endothelial cells by oxidized low-density lipoprotein (ox-LDL) with subsequent increases in endothelial permeability occurs in the early stage of atherosclerosis. Cathepsin L (CATL) is one of the cysteine proteases and has been implicated in advanced atherosclerotic lesions and plaque instability. This study aimed to explore the role of CATL in ox-LDL-induced early atherosclerotic events and to delineate the underlying mechanism. Results showed that ox-LDL upregulated CATL protein levels and activation in human umbilical vein endothelial cells (ECs) in a concentration-dependent manner and stimulated EC autophagy and apoptosis and increased EC monolayer permeability. Concomitantly, VE-cadherin expression was decreased. When ECs were pretreated with a CATL inhibitor, ox-LDL-induced autophagy was inhibited while apoptosis was further increased. In addition, the VE-cadherin protein level was increased, and the EC monolayer permeability was reduced. Taken together, the present study showed that the upregulated expression and activation of CATL induced by ox-LDL, increased EC autophagy and antagonized EC apoptosis, which partly neutralized the effect of increased EC monolayer permeability mediated by the downregulation of VE-cadherin. Thus, the proatherogenic effect of CATL was partly neutralized by inducing autophagy and inhibiting apoptosis in early stages of atherosclerosis.

New role of PCSK9 in atherosclerotic inflammation promotion involving the TLR4/NF-κB pathway

BACKGROUND AND AIMS Proprotein convertase subtilisin/kexin 9 (PCSK9) has emerged as a popular target in the development of new cholesterol-lowering drugs and therapeutic interventions for atherosclerosis. PCSK9 could accelerate atherosclerosis through mechanisms beyond the degradation of the hepatic low-density lipoprotein receptor. Several clinical studies suggested that PCSK9 is involved in atherosclerotic inflammation. Accordingly, this study aimed to explore the role of PCSK9 in vascular inflammation that promotes atherosclerotic progression. METHODS We examined whether PCSK9 silencing via transduction with the lentivirus-mediated PCSK9 shRNA (LV-PCSK9 shRNA) vector affects the formation of vascular lesions in hyperlipidemia-induced atherosclerosis in apolipoprotein E knockout (apoE KO) mice. In vitro, the effects of PCSK9 on oxLDL-induced macrophages inflammation were investigate using LV-PCSK9 and LV-PCSK9 shRNA for PCSK9 overexpression and PCSK9 silencing. RESULTS Immunohistochemical analysis showed that PCSK9 expression increased within atherosclerotic plaques in apoE KO mice. These in vivo results showed that the LV-PCSK9 shRNA group of mice developed less aortic atherosclerotic plaques compared with the control group. These lesions also had the reduced number of macrophages and decreased expression of vascular inflammation regulators, such as tumor necrosis factor-α, interleukin 1 beta, monocyte chemoattractant protein-1, toll-like receptor 4 and nuclear factor kappa B (NF-κB). We further showed that PCSK9 overexpression in macrophages in vitro increased the secretion of oxLDL-induced proinflammatory cytokines. PCSK9 overexpression upregulated TLR4 expression and increased p-IκBα levels, IkBα degradation, and NF-κB nuclear translocation in macrophages, but PCSK9 knockdown had the opposite effects in oxLDL-treated macrophages. CONCLUSIONS PCSK9 gene interference could suppress atherosclerosis directly through decreasing vascular inflammation and inhibiting the TLR4/NF-κB signaling pathway without affecting plasma cholesterol level in high-fat diet-fed apoE KO mice. PCSK9 may be an inflammatory mediator in the pathogenesis of atherosclerosis.

Janus-like role of fibroblast growth factor 2 in arteriosclerotic coronary artery disease: atherogenesis and angiogenesis.

Angiogenic stimulation is a promising new strategy for treating patients with arteriosclerotic coronary artery disease. This strategy aims to ameliorate cardiac function by improving myocardial perfusion and lowering the risk of myocardial infarction. However, angiogenesis may contribute to the growth of atherosclerotic lesions. Atherogenesis is also a potential side effect of angiogenic therapy. Early clinical trials were performed using fibroblast growth factor 2 (FGF2) protein, which enhances the formation of new collateral vessels to reduce ischaemic symptoms. Conversely, angiogenic stimulation by FGF2 is a dilemma because it could cause negative angiogenic effects, such as atherosclerosis. Thus far, clinical trials in patients with recombinant FGF2 protein therapy have not yet yielded undisputable beneficial effects. Future trials should determine whether an improvement can be obtained in patients with coronary artery disease using a combination of FGF2 and other growth factors or a combination of the FGF2 gene and stem cell therapy. This review summarises the multiple roles of FGF2 in the progression of atherosclerosis, its effect on pro-angiogenesis and improvement of cardiac function in coronary artery disease, and the potentially unfavourable effect of angiogenesis on the prevention and treatment of atherogenesis.

PI3K/Akt/FoxO3a signaling mediates cardioprotection of FGF-2 against hydrogen peroxide-induced apoptosis in H9c2 cells

Cardiovascular disease is a growing major global public health problem. Oxidative stress is regarded as one of the key regulators of pathological physiology, which eventually leads to cardiovascular disease. However, mechanisms by which FGF-2 rescues cells from oxidative stress damage in cardiovascular disease is not fully elucidated. Herein this study was designed to investigate the protective effects of FGF-2 in H2O2-induced apoptosis of H9c2 cardiomyocytes, as well as the possible signaling pathway involved. Apoptosis of H9c2 cardiomyocytes was induced by H2O2 and assessed using methyl thiazolyl tetrazolium assay, Hoechst, and TUNEL staining. Cells were pretreated with PI3K/Akt inhibitor LY294002 to investigate the possible PI3K/Akt pathways involved in the protection of FGF-2. The levels of p-Akt, p-FoxO3a, and Bim were detected by immunoblotting. Stimulation with H2O2 decreased the phosphorylation of Akt and FoxO3a, and induced nuclear localization of FoxO3a and apoptosis of H9c2 cells. These effects of H2O2 were abrogated by pretreatment with FGF-2. Furthermore, the protective effects of FGF-2 were abolished by PI3K/Akt inhibitor LY294002. In conclusion, our data suggest that FGF-2 protects against H2O2-induced apoptosis of H9c2 cardiomyocytes via activation of the PI3K/Akt/FoxO3a pathway.

Tet methylcytosine dioxygenase 2 inhibits atherosclerosis via upregulation of autophagy in ApoE −/− mice

Tet methylcytosine dioxygenase 2 (TET2) mediates the conversion of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC). The loss of TET2 is associated with advanced atherosclerotic lesions. Our previous study showed that TET2 improves endothelial cell function by enhancing endothelial cell autophagy. Accordingly, this study determined the role of TET2 in atherosclerosis and potential mechanisms. In ApoE−/− mice fed high-fat diet, TET2 overexpression markedly decreased atherosclerotic lesions with uniformly increased level of 5hmC and decreased level of 5mC in genomic DNA. TET2 overexpression also promoted autophagy and downregulated inflammation factors, such as vascular cell adhesion molecule 1, intercellular adhesion molecule 1, monocyte chemotactic protein 1, and interleukin-1. Consistently, TET2 knockdown with small hairpin RNA (shRNA) in ApoE−/− mice decreased 5hmC and increased 5mC levels in atherosclerotic lesions. Meanwhile, autophagy was inhibited and atherosclerotic lesions progressed with an unstable lesion phenotype characterized by large lipid core, macrophage accumulation, and upregulated inflammation factor expression. Experiments with the cultured endothelial cells revealed that oxidized low-density lipoprotein (ox-LDL) inhibited endothelial cell autophagy. TET2 shRNA strengthened impaired autophagy and autophagic flux in the ox-LDL-treated endothelial cells. TET2 overexpression reversed these effects by decreasing the methylation level of the Beclin 1 promoter, which contributed to the downregulation of inflammation factors. Overall, we identified that TET2 was downregulated during the pathogenesis of atherosclerosis. The downregulation of TET2 promotes the methylation of the Beclin 1 promoter, leading to endothelial cell autophagy, impaired autophagic flux, and inflammatory factor upregulation. Upregulation of TET2 may be a novel therapeutic strategy for treating atherosclerosis.

Lowering serum lipids via PCSK9-targeting drugs: current advances and future perspectives

Proprotein convertase subtilisin/kexin type 9 (PCSK9), also known as neural apoptosis regulated convertase (NARC1), is a key modulator of cholesterol metabolism. PCSK9 increases the serum concentration of low-density lipoprotein cholesterol by escorting low-density lipoprotein receptors (LDLRs) from the membrane of hepatic cells into lysosomes, where the LDLRs are degraded. Owing to the importance of PCSK9 in lipid metabolism, considerable effort has been made over the past decade in developing drugs targeting PCSK9 to lower serum lipid levels. Nevertheless, some problems and challenges remain. In this review we first describes the structure and function of PCSK9 and its gene polymorphisms. We then discuss the various designs of pharmacological targets of PCSK9, including those that block the binding of PCSK9 to hepatic LDLRs (mimetic peptides, adnectins, and monoclonal antibodies), inhibit PCSK9 expression (the clustered regularly interspaced short palindromic repeats/Cas9 platform, small molecules, antisense oligonucleotides, and small interfering RNAs), and interfere with PCSK9 secretion. Finally, this review highlights future challenges in this field, including safety concerns associated with PCSK9 monoclonal antibodies, the limited utility of PCSK9 inhibitors in the central nervous system, and the cost-effectiveness of PCSK9 inhibitors.