ABSTRACT
BACKGROUND AND AIMS: Recent studies highlighted the role of calcification processes in the clinical progression of chronic cardiovascular diseases. In this study we investigated the relationship between the chemical composition of calcification and atherosclerotic plaque stability in carotid arteries. METHODS AND RESULTS: To this end, we characterized the calcification on 229 carotid plaques, by morphology, immunohistochemistry, transmission electron microscopy and energy dispersive X-ray microanalysis. Plaques were classified into two categories: unstable and stable. No significant differences were found in the incidence of the various risk factors between patients with and without carotid calcification, with the exception of diabetes. The energy dispersive X-ray microanalysis allowed us to identify two types of calcium salts in the atheromatous plaques, hydroxyapatite (HA) and calcium oxalate (CO). Our results showed that calcification is a common finding in carotid plaques, being present in 77.3% of cases, and the amount of calcium is not a factor of vulnerability. Noteworthy, we observed an association between HA calcification and unstable plaques. On the contrary, CO calcifications were mainly detected in stable plaques. CONCLUSIONS: The presence of different types of calcification in atheromatous plaques may open new perspectives in understanding the molecular mechanisms of atheroma formation and plaque instability.
Subject(s)
Calcium Oxalate/analysis , Carotid Arteries/chemistry , Carotid Artery Diseases/metabolism , Durapatite/analysis , Plaque, Atherosclerotic , Vascular Calcification/metabolism , Aged , Biomarkers/analysis , Biopsy , Carotid Arteries/ultrastructure , Carotid Artery Diseases/pathology , Disease Progression , Female , Humans , Immunohistochemistry , Male , Microscopy, Electron, Transmission , Middle Aged , Risk Factors , Rupture, Spontaneous , Spectrometry, X-Ray Emission , Vascular Calcification/pathologyABSTRACT
BACKGROUND/OBJECTIVES: The unresolved chronic inflammation of white adipose tissue (WAT) in obesity leads to interstitial deposition of fibrogenic proteins as reparative process. The contribution of omental adipose tissue (oWAT) fibrosis to obesity-related complications remains controversial. The aim of our study was to investigate whether oWAT fibrosis may be related to insulin resistance in severely obese population. SUBJECTS/METHODS: Forty obese subjects were studied by glucose clamp before undergoing bariatric surgery and thus stratified according to insulin resistance severity (M-value). From the first (Group B: n=13; M=1.9±0.7 mg kg(-1) min(-1)) and the highest (Group A: n=14; M=4.5±1.4 mg kg(-1) min(-1)) M-value tertiles, which were age-, waist- and body mass index-matched, oWAT samples were then obtained.Gene expression of collagen type I, III and VI, interleukin-6, profibrotic mediators (transforming growth factor (TGF)-ß1, activin A, connective tissue growth factor), hypoxia inducible factor-1α (HIF-1α) and macrophage (CD68, monocyte chemotactic protein (MCP)-1, CD86, CD206, CD150) markers were analyzed by quantitative reverse transcription PCR. Adipocyte size and total fibrosis were assessed by histomorphometry techniques. RESULTS: Fibrosis at morphological level resulted significantly greater in Group B compared with Group A, although collagens gene expression did not differ. Notably, collagen VI messenger RNA significantly correlated with collagen I, collagen III, HIF-1α, TGF-ß1, CD68, MCP-1 and CD86 transcription levels, supporting their relation with fibrosis development. CONCLUSIONS: In conclusion, we show for the first time that human oWAT fibrosis in severe obesity is consistent with a higher degree of insulin resistance measured by glucose clamp. Therefore, collagen deposition could represent a maladaptive mechanism contributing to obesity-related metabolic complications.
Subject(s)
Cardiovascular Diseases/complications , Diabetes Mellitus, Type 2/complications , Diabetes Mellitus, Type 2/diagnosis , Glycated Hemoglobin/analysis , Aged , Blood Glucose/analysis , Diabetic Retinopathy/blood , Female , Glucose Intolerance , Humans , Male , Middle Aged , Reproducibility of ResultsABSTRACT
Endothelial dysfunction and impaired autophagic activity have a crucial role in aging-related diseases such as cardiovascular dysfunction and atherosclerosis. We have identified miR-216a as a microRNA that is induced during endothelial aging and, according to the computational analysis, among its targets includes two autophagy-related genes, Beclin1 (BECN1) and ATG5. Therefore, we have evaluated the role of miR-216a as a molecular component involved in the loss of autophagic function during endothelial aging. The inverse correlation between miR-216a and autophagic genes was conserved during human umbilical vein endothelial cells (HUVECs) aging and in vivo models of human atherosclerosis and heart failure. Luciferase experiments indicated BECN1, but not ATG5 as a direct target of miR-216a. HUVECs were transfected in order to modulate miR-216a expression and stimulated with 100 µg/ml oxidized low-density lipoprotein (ox-LDL) to induce a stress repairing autophagic process. We found that in young HUVECs, miR-216a overexpression repressed BECN1 and ATG5 expression and the ox-LDL induced autophagy, as evaluated by microtubule-associated protein 1 light chain 3 (LC3B) analysis and cytofluorimetric assay. Moreover, miR-216a stimulated ox-LDL accumulation and monocyte adhesion in HUVECs. Conversely, inhibition of miR-216a in old HUVECs rescued the ability to induce a protective autophagy in response to ox-LDL stimulus. In conclusion, mir-216a controls ox-LDL induced autophagy in HUVECs by regulating intracellular levels of BECN1 and may have a relevant role in the pathogenesis of cardiovascular disorders and atherosclerosis.
Subject(s)
Atherosclerosis/metabolism , Autophagy , Heart Failure/metabolism , Human Umbilical Vein Endothelial Cells/cytology , Human Umbilical Vein Endothelial Cells/metabolism , MicroRNAs/metabolism , Apoptosis Regulatory Proteins/genetics , Apoptosis Regulatory Proteins/metabolism , Atherosclerosis/genetics , Atherosclerosis/physiopathology , Autophagy-Related Protein 5 , Beclin-1 , Heart Failure/genetics , Heart Failure/physiopathology , Humans , Lipoproteins, LDL/metabolism , Membrane Proteins/genetics , Membrane Proteins/metabolism , MicroRNAs/genetics , Microtubule-Associated Proteins/genetics , Microtubule-Associated Proteins/metabolismABSTRACT
Diabetic foot ulceration remains one of the most common and most serious consequences of diabetes. Persistently high levels of matrix metalloproteases (MMPs) contribute to wound chronicity. Our aim was to assess the concentrations of MMPs and tissue inhibitors of metalloproteinases (TIMPs) in neuropathic and ischemic diabetic foot ulcers by analyzing biopsy samples. In this study, biopsies were taken from 35 diabetic foot ulcers of type 2 diabetes mellitus patients and distinguished in neuropathic (n = 14) or ischemic (n = 21). Zymography assay was utilized for the analysis of MMP-2 and MMP-9 activity. TACE activity was evaluated by a specific fluorimetric assay. mRNA levels of MMPs as well as TIMPs were detected using quantitative real-time polymerase chain reaction. The activity of MMP9 and A Disintegrin and A MetalloProtease Domain 17/TNF-Alpha Converting Enzyme (ADAM17/TACE) was significantly increased in ischemic compared to neuropathic biopsies. No differences were detected between both groups for the mRNA levels of MMPs as well as of ADAMs. However, TIMP3 mRNA expression was decreased in ischemic samples. The combination of increased activity of MMP9 and ADAM17/TACE with decreased concentrations of TIMP-3 mRNA expression in ischemic diabetic foot ulcers compared to neuropathic samples suggests that the increased proteolytic environment may represent a causative factor in the ulcer progression. New treatment strategies for healing diabetic foot ulcers could be directed toward increasing levels of TIMP3.
Subject(s)
Diabetes Mellitus, Type 2/complications , Diabetes Mellitus, Type 2/genetics , Diabetic Foot/genetics , Diabetic Neuropathies/genetics , Ischemia/genetics , Tissue Inhibitor of Metalloproteinase-3/genetics , ADAM Proteins/metabolism , ADAM17 Protein , Aged , Aged, 80 and over , Diabetes Mellitus, Type 2/metabolism , Diabetic Foot/complications , Diabetic Foot/metabolism , Diabetic Neuropathies/complications , Diabetic Neuropathies/metabolism , Disease Progression , Down-Regulation , Female , Humans , Ischemia/complications , Ischemia/metabolism , Male , Middle Aged , Tissue Inhibitor of Metalloproteinase-3/metabolismABSTRACT
AIMS/HYPOTHESIS: Overexpression of the gene encoding phosphoprotein enriched in astrocytes 15 (PEA15), also known as phosphoprotein enriched in diabetes (PED), causes insulin resistance and diabetes in transgenic mice and has been observed in type 2 diabetic individuals. The aim of this study was to investigate whether PEA15 overexpression occurs in individuals at high risk of diabetes and whether it is associated with specific type 2 diabetes subphenotypes. SUBJECTS AND METHODS: We analysed PEA15 expression in euglycaemic first-degree relatives (FDR) of type 2 diabetic subjects. RESULTS: The expression of PEA15 in peripheral blood leucocytes (PBLs) paralleled that in fat and skeletal muscle tissues. In PBLs from the FDR, PEA15 expression was two-fold higher than in euglycaemic individuals with no family history of diabetes (control subjects), both at the protein and the mRNA level (p < 0.001). The expression of PEA15 was comparable in FDR and type 2 diabetic subjects and in each group close to one-third of the subjects expressed PEA15 levels more than 2 SD higher than the mean of control subjects. Subjects with IFG with at least one type 2 diabetes-affected FDR also overexpressed PEA15 (p < 0.05). In all the groups analysed, PEA15 expression was independent of sex and unrelated to age, BMI, waist circumference, systolic and diastolic BP, and fasting cholesterol, triacylglycerol and glucose levels. However, in euglycaemic FDR of type 2 diabetic subjects, PEA15 expression was inversely correlated with insulin sensitivity (r = -557, p = 0.01). CONCLUSIONS/INTERPRETATION: We conclude that PEA15 overexpression represents a common defect in FDR of patients with type 2 diabetes and is correlated with reduced insulin sensitivity in these individuals.
Subject(s)
Diabetes Mellitus, Type 2/genetics , Gene Expression Regulation , Insulin Resistance/genetics , Intracellular Signaling Peptides and Proteins/genetics , Phosphoproteins/genetics , Adult , Apoptosis Regulatory Proteins , Blood Glucose/metabolism , DNA Primers , Diabetes Mellitus, Type 2/physiopathology , Family , Female , Humans , Male , Phosphoproteins/metabolism , RNA/genetics , RNA/isolation & purification , Reverse Transcriptase Polymerase Chain ReactionABSTRACT
The N-n-propyl-N-(2-phenylethyl)-2-(3-hydroxyphenyl)ethylamine (1, RU 24213) had been previously identified as selective agonist of DA D2 receptor subtype. In this paper we describe the synthesis and in vitro binding affinities of several derivatives of 1 substituted with fluorine, chlorine and methyl or hydroxy groups on the phenyl ring of the N-2-phenylethyl moiety. The results obtained indicate that these substitutions do not improve the D2 binding affinity. The introduction on the phenyl ring of two fluorine or chlorine atoms decreases with D1 affinity, and the dichloro derivatives are highly selective for the D2 receptor. Preliminary behavioural tests confirm that the dichloro derivatives behave as D2 selective agonists.
Subject(s)
Ethylamines/chemical synthesis , Receptors, Dopamine/drug effects , Animals , Benzazepines/pharmacology , Dopamine Agents/pharmacology , Ethylamines/pharmacology , In Vitro Techniques , Ligands , Metoclopramide/pharmacology , Motor Activity/drug effects , Neostriatum/drug effects , Neostriatum/metabolism , Phenethylamines/pharmacology , Rats , Receptors, Dopamine D1/drug effects , Receptors, Dopamine D2/drug effects , Spiperone/pharmacology , Stereotyped Behavior/drug effectsABSTRACT
The synthesis of 2-(4-fluoro-3-hydroxyphenyl)ethylamine (26) and of some N,N-dialkyl derivatives (27-30) starting from 4-fluoro-3-hydroxytoluene and their in vitro binding affinities for dopamine (DA) receptor are reported. The amine 26 can be regarded as a molecular modification of DA in which the para hydroxyl group is replaced by fluorine. The new compounds 26-30 were evaluated for their affinity at D-1 and D-2 DA receptor subtypes by displacement of [3H]SCH 23390 (D-1 selective) and [3H]spiperone (D-2 selective). The amine 26 had about 2-fold less affinity for D-1 and D-2 binding sites than DA. The substitution of the amino group with ethyl, n-propyl, and 2-phenylethyl groups decreased the affinity for D-1 binding sites but greatly enhanced the effectiveness on D-2 binding sites. The N-ethyl- (28) and N-n-propyl-N-(2-phenylethyl)-2-(4-fluoro-3- hydroxyphenyl)ethylamine (30) were the most potent members of the series with high selectivity for D-2 binding sites. A similar effect was observed with isomeric N-n-propyl-N-(2-phenylethyl)-2-(3-fluoro-4-hydroxyphenyl)ethylamine (31) which was approximately 65 times more selective for D-2 sites vs D-1 sites. The introduction of a 2-phenylethyl group on the nitrogen atom induce the highest effect, perhaps as a consequence of an increased liposolubility or of binding to a complementary lipophilic site on the receptor.
Subject(s)
Dopamine Agents/chemical synthesis , Phenethylamines/chemical synthesis , Animals , Benzazepines/metabolism , Chemical Phenomena , Chemistry , Dopamine Agents/pharmacology , Dopamine Antagonists , Phenethylamines/pharmacology , Rats , Rats, Inbred Strains , Receptors, Dopamine/drug effects , Structure-Activity RelationshipABSTRACT
The 2-(3-fluoro-4-hydroxyphenyl)ethylamine and its N,N-dialkyl derivatives were synthesized. The affinity of new compounds for dopamine binding sites was measured in a test involving displacement of [3H]SCH 23390 (D-1 selective) and [3H]spiperone (D-2 selective) from homogenized rat striatal tissue. No compound proved effective in displacing [3H]SCH 23390. Two derivatives are selective displacers of [3H]spiperone.
Subject(s)
Dopamine/analogs & derivatives , Receptors, Dopamine/drug effects , Animals , Antipsychotic Agents/pharmacology , Benzazepines/pharmacology , Chemical Phenomena , Chemistry , Corpus Striatum/drug effects , Corpus Striatum/metabolism , Dopamine/chemical synthesis , Dopamine/metabolism , Dopamine/pharmacology , In Vitro Techniques , Radioligand Assay , Rats , Receptors, Dopamine/metabolism , Spiperone/pharmacologyABSTRACT
Some N-alkyl derivatives of 2-(3-hydroxybenzyl)piperidine and of 2-(3,4-dihydroxybezyl)piperidine were synthesized and evaluated pharmacologically in vitro by competition with [3H]spiperone for binding to a homogenized rat striatal tissue, and for ability to stimulate adenylate cyclase. The N-methyl-2-(3,4-dihydroxybenzyl)piperidine shows a fairly good affinity for the D-2 dopamine receptor. The N-alkyl 2-(3,4-dihydroxybenzyl)piperidines produce a faible stimulation of adenylate cyclase activity.
Subject(s)
Piperidines/pharmacology , Receptors, Dopamine/drug effects , Adenylyl Cyclase Inhibitors , Animals , Benzyl Compounds/chemical synthesis , Benzyl Compounds/pharmacology , Binding, Competitive/drug effects , Chemical Phenomena , Chemistry , Corpus Striatum/metabolism , In Vitro Techniques , Male , Piperidines/chemical synthesis , Rats , Rats, Inbred Strains , Spiperone/metabolism , Synaptosomes/enzymologyABSTRACT
The syntheses and a preliminary pharmacological evaluation of some aminopropylindolizines and aminopropyltetrahydroindolizines are reported. All compounds showed anti-5-hydroxytryptamine, anti-histamine, and antiacetylcholine activities. Some also exhibited weak CNS activity.
Subject(s)
Indolizines/chemical synthesis , Acetylcholine/antagonists & inhibitors , Animals , Behavior, Animal/drug effects , Female , Guinea Pigs , Histamine H1 Antagonists/chemical synthesis , In Vitro Techniques , Indolizines/pharmacology , Male , Mice , Muscle Contraction/drug effects , Muscle, Smooth/drug effects , Propylamines/chemical synthesis , Propylamines/pharmacology , Rats , Serotonin Antagonists/chemical synthesis , Uterine Contraction/drug effectsABSTRACT
The synthesis and antimonoamine oxidase activity of some N'-substituted hydrazides of indolizine-2-carboxylic acid are described. They all inhibit monoamine oxidase and are more active than iproniazid. The structure-activity relationships also are discussed.
Subject(s)
Indolizines/chemical synthesis , Monoamine Oxidase Inhibitors/chemical synthesis , Carboxylic Acids/chemical synthesis , Carboxylic Acids/pharmacology , In Vitro Techniques , Indolizines/pharmacology , Iproniazid/pharmacology , Structure-Activity RelationshipABSTRACT
Several 5-monosubstituted and 5,5-disubstituted 5,6-dihydroimidazo[1,2-c]quinazolines were synthesized in high yields by the condensation of 2-(o-aminophenyl)imidazole with the required aliphatic or aromatic aldehydes or ketones. Some of these compounds caused inhibition of ADP-induced platelet aggregation in vitro at concentration of 10(-5) to 10(-4) M. The structure-activity relationships of these inhibitors are discussed.