Peripheral artery disease (PAD) is due to the blockage of the arteries supplying blood to the lower limbs usually secondary to atherosclerosis. The most severe clinical manifestation of PAD is critical limb ischemia (CLI), which is associated with a risk of limb loss and mortality due to cardiovascular events. Currently CLI is mainly treated by surgical or endovascular revascularization, with few other treatments in routine clinical practice. There are a number of problems with current PAD management strategies, such as the difficulty in selecting the appropriate treatments for individual patients. Many patients undergo repeated attempts at revascularization surgery, but ultimately require an amputation. There is great interest in developing new methods to identify patients who are unlikely to benefit from revascularization and to improve management of patients unsuitable for surgery. Circulating biomarkers that predict the progression of PAD and the response to therapies could assist in the management of patients. This review provides an overview of the pathophysiology of PAD and examines the association between circulating biomarkers and PAD presence, severity and prognosis. While some currently identified circulating markers show promise, further larger studies focused on the clinical value of the biomarkers over existing risk predictors are needed. Full article

Abdominal aortic aneurysm (AAA) is an inflammatory disease associated with marked changes in the cellular composition of the aortic wall. This study aims to identify microRNA (miRNA) expression in aneurysmal inflammatory cells isolated by laser microdissection from human tissue samples. The distribution of inflammatory cells (neutrophils, B and T lymphocytes, mast cells) was evaluated in human AAA biopsies. We observed in half of the samples that adventitial tertiary lymphoid organs (ATLOs) with a thickness from 0.5 to 2 mm were located exclusively in the adventitia. Out of the 850 miRNA that were screened by microarray in isolated ATLOs (n = 2), 164 miRNAs were detected in ATLOs. The three miRNAs (miR-15a-3p, miR-30a-5p and miR-489-3p) with the highest expression levels were chosen and their expression quantified by RT-PCR in isolated ATLOs (n = 4), M1 (n = 2) and M2 macrophages (n = 2) and entire aneurysmal biopsies (n = 3). Except for the miR-30a-5p, a similar modulation was found in ATLOs and the two subtypes of macrophages. The modulated miRNAs were then evaluated in the plasma of AAA patients for their potential as AAA biomarkers. Our data emphasize the potential of miR-15a-3p and miR-30a-5p as biomarkers of AAA but also as triggers of ATLO evolution. Further investigations will be required to evaluate their targets in order to better understand AAA pathophysiology. Full article

Abdominal aortic aneurysm (AAA) is a complex disorder that has a significant impact on the aging population. While both genetic and environmental risk factors have been implicated in AAA formation, the precise genetic markers involved and the factors influencing their expression remain an area of ongoing investigation. DNA methylation has been previously used to study gene silencing in other inflammatory disorders and since AAA has an extensive inflammatory component, we sought to examine the genome-wide DNA methylation profiles in mononuclear blood cells of AAA cases and matched non-AAA controls. To this end, we collected blood samples and isolated mononuclear cells for DNA and RNA extraction from four all male groups: AAA smokers (n = 11), AAA non-smokers (n = 9), control smokers (n = 10) and control non-smokers (n = 11). Methylation data were obtained using the Illumina 450k Human Methylation Bead Chip and analyzed using the R language and multiple Bioconductor packages. Principal component analysis and linear analysis of CpG island subsets identified four regions with significant differences in methylation with respect to AAA: kelch-like family member 35 (KLHL35), calponin 2 (CNN2), serpin peptidase inhibitor clade B (ovalbumin) member 9 (SERPINB9), and adenylate cyclase 10 pseudogene 1 (ADCY10P1). Follow-up studies included RT-PCR and immunostaining for CNN2 and SERPINB9. These findings are novel and suggest DNA methylation may play a role in AAA pathobiology. Full article

We investigated transcriptional control of gene expression in human abdominal aortic aneurysm (AAA). We previously identified 3274 differentially expressed genes in human AAA tissue compared to non-aneurysmal controls. Four expressed transcription factors (ELF1, ETS2, STAT5 and RUNX1) were selected for genome-wide chromatin immunoprecipitation. Transcription factor binding was enriched in 4760 distinct genes (FDR < 0.05), of which 713 were differentially expressed in AAA. Functional classification using Gene Ontology (GO), KEGG, and Network Analysis revealed enrichment in several biological processes including “leukocyte migration” (FDR = 3.09 × 10⁻⁰⁵) and “intracellular protein kinase cascade” (FDR = 6.48 × 10⁻⁰⁵). In the control aorta, the most significant GO categories differed from those in the AAA samples and included “cytoskeleton organization” (FDR = 1.24 × 10⁻⁰⁶) and “small GTPase mediated signal transduction” (FDR = 1.24 × 10⁻⁰⁶). Genes up-regulated in AAA tissue showed a highly significant enrichment for GO categories “leukocyte migration” (FDR = 1.62 × 10⁻¹¹), “activation of immune response” (FDR = 8.44 × 10⁻¹¹), “T cell activation” (FDR = 4.14 × 10⁻¹⁰) and “regulation of lymphocyte activation” (FDR = 2.45 × 10⁻⁰⁹), whereas the down-regulated genes were enriched in GO categories “cytoskeleton organization” (FDR = 7.84 × 10⁻⁰⁵), “muscle cell development” (FDR = 1.00 × 10⁻⁰⁴), and “organ morphogenesis” (FDR = 3.00 × 10⁻⁰⁴). Quantitative PCR assays confirmed a sub-set of the transcription factor binding sites including those in MTMR11, DUSP10, ITGAM, MARCH1, HDAC8, MMP14, MAGI1, THBD and SPOCK1. Full article

HMG-CoA (3-hydroxy-3-methylglutaryl-coenzyme A) reductase inhibitors (statins) have been suggested to attenuate abdominal aortic aneurysm (AAA) growth. However, the effects of statins in human AAA tissues are not fully elucidated. The aim of this study was to investigate the direct effects of statins on proinflammatory molecules in human AAA walls in ex vivo culture. Simvastatin strongly inhibited the activation of nuclear factor (NF)-κB induced by tumor necrosis factor (TNF)-α in human AAA walls, but showed little effect on c-jun N-terminal kinase (JNK) activation. Simvastatin, as well as pitavastatin significantly reduced the secretion of matrix metalloproteinase (MMP)-9, monocyte chemoattractant protein (MCP)-2 and epithelial neutrophil-activating peptide (CXCL5) under both basal and TNF-α-stimulated conditions. Similar to statins, the Rac1 inhibitor NSC23766 significantly inhibited the activation of NF-κB, accompanied by a decreased secretion of MMP-9, MCP-2 and CXCL5. Moreover, the effect of simvastatin and the JNK inhibitor SP600125 was additive in inhibiting the secretion of MMP-9, MCP-2 and CXCL5. These findings indicate that statins preferentially inhibit the Rac1/NF-κB pathway to suppress MMP-9 and chemokine secretion in human AAA, suggesting a mechanism for the potential effect of statins in attenuating AAA progression. Full article

Our previous analysis using genome-wide microarray expression data revealed extreme overrepresentation of immune related genes belonging the Natural Killer (NK) Cell Mediated Cytotoxicity pathway (hsa04650) in human abdominal aortic aneurysm (AAA). We followed up the microarray studies by immunohistochemical analyses using antibodies against nine members of the NK pathway (VAV1, VAV3, PLCG1, PLCG2, HCST, TYROBP, PTK2B, TNFA, and GZMB) and aortic tissue samples from AAA repair operations (n = 6) and control aortae (n = 8) from age-, sex- and ethnicity-matched donors from autopsies. The results confirmed the microarray results. Two different members of the NK pathway, HCST and GRZB, which act at different steps in the NK-pathway, were actively transcribed and translated into proteins in the same cells in the AAA tissue demonstrated by double staining. Furthermore, double staining with antibodies against CD68 or CD8 together with HCST, TYROBP, PTK2B or PLCG2 revealed that CD68 and CD8 positive cells expressed proteins of the NK-pathway but were not the only inflammatory cells involved in the NK-pathway in the AAA tissue. The results provide strong evidence that the NK Cell Mediated Cytotoxicity Pathway is activated in human AAA and valuable insight for future studies to dissect the pathogenesis of human AAA. Full article

Most reported studies with animal models of abdominal aortic aneurysm (AAA) and several studies with patients have suggested that doxycycline favourably modifies AAA; however, a recent large long-term clinical trial found that doxycycline did not limit aneurysm growth. Thus, there is currently no convincing evidence that doxycycline reduces AAA expansion. Here, we critically review the available experimental and clinical information about the effects of doxycycline when used as a pharmacological treatment for AAA. The view that AAA can be considered an autoimmune disease and the observation that AAA tissue shows clonal expansion of T cells is placed in the light of the well-known inhibition of mitochondrial protein synthesis by doxycycline. In T cell leukaemia animal models, this inhibitory effect of the antibiotic has been shown to impede T cell proliferation, resulting in complete tumour eradication. We suggest that the available evidence of doxycycline action on AAA is erroneously ascribed to its inhibition of matrix metalloproteinases (MMPs) by competitive binding of the zinc ion co-factor. Although competitive binding may explain the inhibition of proteolytic activity, it does not explain the observed decreases of MMP mRNA levels. We propose that the observed effects of doxycycline are secondary to inhibition of mitochondrial protein synthesis. Provided that serum doxycycline levels are kept at adequate levels, the inhibition will result in a proliferation arrest, especially of clonally expanding T cells. This, in turn, leads to the decrease of proinflammatory cytokines that are normally generated by these cells. The drastic change in cell type composition may explain the changes in MMP mRNA and protein levels in the tissue samples. Full article

Peripheral artery disease (PAD) is a broad disorder encompassing multiple forms of arterial disease outside of the heart. As such, PAD development is a multifactorial process with a variety of manifestations. For example, aneurysms are pathological expansions of an artery that can lead to rupture, while ischemic atherosclerosis reduces blood flow, increasing the risk of claudication, poor wound healing, limb amputation, and stroke. Current PAD treatment is often ineffective or associated with serious risks, largely because these disorders are commonly undiagnosed or misdiagnosed. Active areas of research are focused on detecting and characterizing deleterious arterial changes at early stages using non-invasive imaging strategies, such as ultrasound, as well as emerging technologies like photoacoustic imaging. Earlier disease detection and characterization could improve interventional strategies, leading to better prognosis in PAD patients. While rodents are being used to investigate PAD pathophysiology, imaging of these animal models has been underutilized. This review focuses on structural and molecular information and disease progression revealed by recent imaging efforts of aortic, cerebral, and peripheral vascular disease models in mice, rats, and rabbits. Effective translation to humans involves better understanding of underlying PAD pathophysiology to develop novel therapeutics and apply non-invasive imaging techniques in the clinic. Full article

Peripheral artery disease (PAD) is a broad term encompassing a range of atherosclerotic and aneurysmal conditions of the extra-coronary arteries [1]. [...] Full article

The acute phase protein serum amyloid A (SAA), a marker of inflammation, induces expression of pro-inflammatory and pro-thrombotic mediators including ICAM-1, VCAM-1, IL-6, IL-8, MCP-1 and tissue factor (TF) in both monocytes/macrophages and endothelial cells, and induces endothelial dysfunction—a precursor to atherosclerosis. In this study, we determined the effect of pharmacological inhibition of known SAA receptors on pro-inflammatory and pro-thrombotic activities of SAA in human carotid artery endothelial cells (HCtAEC). HCtAEC were pre-treated with inhibitors of formyl peptide receptor-like-1 (FPRL-1), WRW4; receptor for advanced glycation-endproducts (RAGE), (endogenous secretory RAGE; esRAGE) and toll-like receptors-2/4 (TLR2/4) (OxPapC), before stimulation by added SAA. Inhibitor activity was also compared to high-density lipoprotein (HDL), a known inhibitor of SAA-induced effects on endothelial cells. SAA significantly increased gene expression of TF, NFκB and TNF and protein levels of TF and VEGF in HCtAEC. These effects were inhibited to variable extents by WRW4, esRAGE and OxPapC either alone or in combination, suggesting involvement of endothelial cell SAA receptors in pro-atherogenic gene expression. In contrast, HDL consistently showed the greatest inhibitory action, and often abrogated SAA-mediated responses. Increasing HDL levels relative to circulating free SAA may prevent SAA-mediated endothelial dysfunction and ameliorate atherogenesis. Full article

Doxorubicin (DOX) is an effective chemotherapeutic agent; however; its use is limited by some side effects; such as cardiotoxicity and thrombocytopenia. DOX-induced cardiotoxicity has been intensively investigated; however; DOX-induced thrombocytopenia has not been clearly elucidated. Here we show that DOX-induced mitochondria-mediated intrinsic apoptosis and glycoprotein (GP)Ibα shedding in platelets. DOX did not induce platelet activation; whereas; DOX obviously reduced adenosine diphosphate (ADP)- and thrombin-induced platelet aggregation; and impaired platelet adhesion on the von Willebrand factor (vWF) surface. In addition; we also show that DOX induced intracellular reactive oxygen species (ROS) production and mitochondrial ROS generation in a dose-dependent manner. The mitochondria-targeted ROS scavenger Mito-TEMPO blocked intracellular ROS and mitochondrial ROS generation. Furthermore; Mito-TEMPO reduced DOX-induced platelet apoptosis and GPIbα shedding. These data indicate that DOX induces platelet apoptosis; and impairs platelet function. Mitochondrial ROS play a pivotal role in DOX-induced platelet apoptosis and GPIbα shedding. Therefore; DOX-induced platelet apoptosis might contribute to DOX-triggered thrombocytopenia; and mitochondria-targeted ROS scavenger would have potential clinical utility in platelet-associated disorders involving mitochondrial oxidative damage. Full article

Cancer cells preferentially utilize glycolysis, instead of oxidative phosphorylation, for metabolism even in the presence of oxygen. This phenomenon of aerobic glycolysis, referred to as the “Warburg effect”, commonly exists in a variety of tumors. Recent studies further demonstrate that both genetic factors such as oncogenes and tumor suppressors and microenvironmental factors such as spatial hypoxia and acidosis can regulate the glycolytic metabolism of cancer cells. Reciprocally, altered cancer cell metabolism can modulate the tumor microenvironment which plays important roles in cancer cell somatic evolution, metastasis, and therapeutic response. In this article, we review the progression of current understandings on the molecular interaction between cancer cell metabolism and the tumor microenvironment. In addition, we discuss the implications of these interactions in cancer therapy and chemoprevention. Full article

Two bidentate NS ligands were synthesized by the condensation reaction of S-2-methylbenzyldithiocarbazate (S2MBDTC) with 2-methoxybenzaldehyde (2MB) and 3-methoxybenzaldehyde (3MB). The ligands were reacted separately with acetates of Cu(II), Ni(II) and Zn(II) yielding 1:2 (metal:ligand) complexes. The metal complexes formed were expected to have a general formula of [M(NS)₂] where M = Cu²⁺, Ni²⁺, and Zn²⁺. These compounds were characterized by elemental analysis, molar conductivity, magnetic susceptibility and various spectroscopic techniques. The magnetic susceptibility measurements and spectral results supported the predicted coordination geometry in which the Schiff bases behaved as bidentate NS donor ligands coordinating via the azomethine nitrogen and thiolate sulfur. The molecular structures of the isomeric S2M2MBH (1) and S2M3MBH (2) were established by X-ray crystallography to have very similar l-shaped structures. The Schiff bases and their metal complexes were evaluated for their biological activities against estrogen receptor-positive (MCF-7) and estrogen receptor-negative (MDA-MB-231) breast cancer cell lines. Only the Cu(II) complexes showed marked cytotoxicity against the cancer cell lines. Both Schiff bases and other metal complexes were found to be inactive. In concordance with the cytotoxicity studies, the DNA binding studies indicated that Cu(II) complexes have a strong DNA binding affinity. Full article

Systemic lupus erythematosus (SLE) is an autoimmune disease involving multiple organs and the presence of anti-nuclear antibodies. The pathogenesis of SLE has been intensively studied but remains far from clear. B and T lymphocyte abnormalities, dysregulation of apoptosis, defects in the clearance of apoptotic materials, and various genetic and epigenetic factors are attributed to the development of SLE. The latest research findings point to the association between abnormal epigenetic regulation and SLE, which has attracted considerable interest worldwide. It is the purpose of this review to present and discuss the relationship between aberrant epigenetic regulation and SLE, including DNA methylation, histone modifications and microRNAs in patients with SLE, the possible mechanisms of immune dysfunction caused by epigenetic changes, and to better understand the roles of aberrant epigenetic regulation in the initiation and development of SLE and to provide an insight into the related therapeutic options in SLE. Full article

Mannose receptor (MR) is a member of pattern-recognition receptors (PRRs), which plays a significant role in immunity responses. Much work on MR has been done in mammals and birds while little in fish. In this report, a MR gene (designated as zfMR) was cloned from zebra fish (Danio rerio), which is an attractive model for the studies of animal diseases. The full-length cDNA of zfMR contains 6248 bp encoding a putative protein of 1428 amino acids. The predicted amino acid sequences showed that zfMR contained a cysteine-rich domain, a single fibronectin type II (FN II) domain, eight C-type lectin-like domains (CTLDs), a transmembrane domain and a short C-terminal cytoplasmic domain, sharing highly conserved structures with MRs from the other species. The MR mRNA could be detected in all examined tissues with highest level in kidney. The temporal expression patterns of MR, IL-1β and TNF-α mRNAs were analyzed in the liver, spleen, kidney and intestine post of infection with Aeromonas sobria. By immunohistochemistry assay, slight enhancement of MR protein was also observed in the spleen and intestine of the infected zebra fish. The established zebra fish-A. sobria infection model will be valuable for elucidating the role of MR in fish immune responses to infection. Full article