Indoxyl sulfate (IS), a uremic toxin, is considered as a risk factor for accelerated atherosclerosis in patients with chronic kidney disease. As uptake of oxidized low-density lipoprotein (Ox-LDL) in macrophages is a key event in the progression of atherosclerosis, the aim of this study was to determine direct effects of IS on uptake of Ox-LDL in macrophages. Flow cytometric analysis revealed that IS significantly stimulated Ox-LDL uptake by THP-1 macrophages in both dose- and time-dependent manners. A CD36 inhibitor, sulfosuccinimidyl oleate (SSO), and ERK1/2 inhibitors, PD98059 and U0126, could suppress the IS-stimulated Ox-LDL uptake. IS also stimulated CD36 expression, which was inhibited by PD98059 and U0126. Western blotting analysis showed that IS significantly activated ERK1/2 mitogen-activated protein kinase (MAPK) pathway by increasing its phosphorylation level. Further, CCK-8 assay showed that IS exerted its effects without affecting cell viability. In conclusion, IS stimulated Ox-LDL uptake through up-regulation of CD36 expression in THP-1 macrophages, partly via ERK1/2 MAPK pathway. This might be one of the mechanisms underlying the progression of atherosclerosis in patients with chronic kidney disease.

• MeSH
• CD36 Antigens physiology   MeSH
• Atherosclerosis * physiopathology   MeSH
• Renal Insufficiency, Chronic * complications   MeSH
• Extracellular Signal-Regulated MAP Kinases physiology   MeSH
• Indican * physiology   adverse effects   MeSH
• Lipoproteins, LDL physiology   MeSH
• Macrophages physiology   MeSH
• MAP Kinase Signaling System physiology   drug effects   MeSH
• Cell Line, Tumor physiology   MeSH
• Flow Cytometry MeSH
• In Vitro Techniques MeSH
• Cell Survival drug effects   MeSH
• Publication type
• Research Support, Non-U.S. Gov't MeSH

Cilostazol is a phosphodiesterase-3 inhibitor that functions as a platelet aggregation inhibitor and is used for treating peripheral artery diseases and ischemic stroke. Dendritic cells (DCs) play an active role in the immunological processes related to atherosclerosis. Cilostazol has anti-atherogenic and anti-inflammatory effects, but the effects of cilostazol on DC maturation remain unknown. The purpose of this study was to determine the effects of cilostazol on lipopolysaccharide (LPS)-induced maturation of DCs. DC2.4 cells were treated with cilostazol for 12 h and subsequently stimulated with LPS to induce maturation. Cilostazol reduced the expression of maturation-associated markers induced by LPS, such as CD40, CD86, and MHCII, improved the endocytotic function, and decreased production of the tumour necrosis factor alpha (TNF-α) and interleukin-6 (IL-6) of these cells. To further elucidate the mechanisms responsible for the inhibition of DC2.4 maturation by cilostazol, we investigated the effect of cilostazol on LPS-stimulated nuclear factor-kappa B (NF-κB) activation. Our results indicated that cilostazol treatment decreased IκBα degradation and inhibited NF-κB p65 translocation, and the inhibitory effects of cilostazol were cAMP-independent. Therefore, inhibition of NF-κB by cilostazol might result in the suppression of DC maturation. In conclusion, cilostazol suppressed LPS-stimulated DC maturation, which might contribute to its anti-atherosclerosis effect.

• MeSH
• CD40 Antigens immunology   MeSH
• B7-2 Antigen immunology   MeSH
• Atherosclerosis immunology   physiopathology   MeSH
• Immunity, Cellular immunology   drug effects   MeSH
• Dendritic Cells * chemistry   immunology   drug effects   MeSH
• Fibrinolytic Agents MeSH
• Histocompatibility Antigens Class II immunology   MeSH
• Platelet Aggregation Inhibitors MeSH
• Phosphodiesterase 3 Inhibitors MeSH
• Interleukin-6 immunology   MeSH
• Lipopolysaccharides therapeutic use   MeSH
• NF-kappa B * genetics   immunology   drug effects   MeSH
• Polymerase Chain Reaction MeSH
• Flow Cytometry MeSH
• In Vitro Techniques MeSH
• Tetrazoles * pharmacology   immunology   MeSH
• Tumor Necrosis Factor-alpha immunology   MeSH
• Vasodilator Agents MeSH
• Blotting, Western MeSH
• Publication type
• Research Support, Non-U.S. Gov't MeSH

• MeSH
• Single-Cell Gene Expression Analysis methods   instrumentation   MeSH
• Survival Analysis MeSH
• Receptors, Chimeric Antigen * immunology   therapeutic use   MeSH
• Progression-Free Survival MeSH
• Humans MeSH
• B-Cell Maturation Antigen immunology   drug effects   MeSH
• Multiple Myeloma * therapy   MeSH
• Drug-Related Side Effects and Adverse Reactions epidemiology   MeSH
• Cytokine Release Syndrome chemically induced   epidemiology   MeSH
• T-Lymphocytes immunology   MeSH
• Treatment Outcome MeSH
• Check Tag
• Humans MeSH
• Male MeSH
• Female MeSH
• Publication type
• Clinical Study MeSH
• Research Support, Non-U.S. Gov't MeSH

A novel phenylacetic acid (PAA)-induced CoA-ligase-encoding gene, designated as phlC, has been cloned from penicillin-producing fungus Penicillium chrysogenum. The open reading frame of phlC cDNA was 1671 bp and encoded a 556 amino acid residues protein with the consensus AMP binding site and a peroxisomal targeting signal 1 on its C terminus. The deduced amino acid sequence showed 37% and 38% identity with characterized P. chrysogenum Phl and PhlB protein, respectively. Functional recombinant PhlC protein was overexpressed in Escherichia coli. The purified recombinant enzyme was capable to convert PAA into its corresponding CoA ester with a specific activity of 129.5 ± 3.026 pmol/min per mg protein. Similar to Phl and PhlB, PhlC displayed broad substrate spectrum and showed higher activities to medium- and long-chain fatty acids. The catalytic properties of PhlC have been determined and compared to those of Phl and PhlB.

• MeSH
• Acetyl Coenzyme A biosynthesis   MeSH
• Escherichia coli genetics   MeSH
• Phenylacetates metabolism   MeSH
• Cloning, Molecular MeSH
• Coenzyme A Ligases chemistry   genetics   metabolism   MeSH
• Molecular Sequence Data MeSH
• Penicillium chrysogenum enzymology   genetics   MeSH
• Reverse Transcriptase Polymerase Chain Reaction MeSH
• Recombinant Proteins chemistry   genetics   metabolism   MeSH
• Amino Acid Sequence MeSH
• Sequence Alignment MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field.

• MeSH
• Autophagy * physiology   MeSH
• Autophagosomes MeSH
• Biomarkers MeSH
• Biological Assay standards   MeSH
• Humans MeSH
• Lysosomes MeSH
• Autophagy-Related Proteins metabolism   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Research Support, N.I.H., Extramural MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH
• Guideline MeSH

• MeSH
• Autophagy * physiology   MeSH
• Biological Assay methods   standards   MeSH
• Humans MeSH
• Computer Simulation MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, N.I.H., Extramural MeSH
• Guideline MeSH