Immunohistochemical localization of advanced glycosylation end products in coronary atheroma and cardiac tissue in diabetes mellitus

Advanced glycosylation end products (AGEs) accumulate on long-lived extracellular matrix proteins and have been implicated in the micro- and macrovascular complications of diabetes mellitus. Within the arterial wall, AGE-modified proteins increase vascular permeability, inactivate nitric oxide activity, and induce the release of growth-promoting cytokines. Recently developed anti-AGE antibodies were used in an immunohistochemical analysis of coronary arteries obtained from type II diabetic and nondiabetic patients. High levels of AGE reactivity were observed within the atherosclerotic plaque present in vessels from selected patients with diabetes. Considered together with the pathological effects of AGEs on vascular wall homeostasis, these data support the role of advanced glycosylation in the rapidly progressive atherosclerosis associated with diabetes mellitus.     

Cytotoxic CD4+ T cells associated with the expression of major histocompatibility complex class II antigen of mouse myeloma cells secreting interferon-gamma are cytolytic in vitro and tumoricidal in vivo

The kidney is responsible for regulation of water and electrolyte balance, filtration and absorption of plasma proteins, and control of blood volume and pressure. Homeostasis achieved by the kidney is controlled in large part by the action of hormones or proteins on specific transmembrane receptors. Conversely, many renal diseases, including that resulting from atherosclerosis, are characterised by scarring and abnormal proliferation of cellular components of the kidney, and these processes are mediated in large part by these same receptors. The G protein-coupled receptors constitute a large and diverse class of proteins, characterised by the possession of seven transmembrane-spanning domains. These receptors bind polypeptide growth factors, which function to transmit a variety of signals from the extracellular to the intracellular milieu. The receptor-associated G proteins utilised by the kidney derive their specificity not only by activating or inhibiting various second-messenger molecules, but also by their location on particular cell types. In this review, several G protein-coupled receptors will be discussed from the perspective of their importance to kidney function and to the pathogenesis of renal disease, atherosclerosis, and hypertension.     

Induction of host signal transduction pathways by Helicobacter pylori

Adherence of Helicobacter pylori to cultured gastric epithelial cells is associated with several cellular events, including the tyrosine phosphorylation of a 145-kDa host protein; the reorganization of the host cell actin and associated cellular proteins, like vasodilator-stimulated phosphoprotein, adjacent to the attached bacterial cell; and the subsequent release of the cytokine, interleukin 8 (IL-8). H. pylori isolated from patients with ulcer disease and gastric cancer contain a DNA insertion, the cag pathogenicity island (PAI), that is not present in bacteria isolated from individuals with asymptomatic infection. Mutations in a number of PAI genes abolish tyrosine phosphorylation and IL-8 synthesis but not the cytoskeletal rearrangements. Kinase inhibition studies suggest there are two distinct pathways operative in stimulating IL-8 release from host cells and one of these H. pylori pathways is independent of the tyrosine phosphorylation step.     

Morphometric analysis of cellular and vascular changes in gingival connective tissue in long-term insulin-dependent diabetes

This study examined cellular and vascular changes in gingival connective tissue samples by stereologic point-counting procedures and interactive digital analyzing systems in long-term insulin-dependent diabetes mellitus patients. Gingival connective tissue capillaries representing a clinically healthy sulcus with no evidence of periodontal disease at the site of biopsy were studied in 29 patients with diabetes. Based upon their long-term medical records, 19 were identified as having poorly controlled (PIDD) and 10 as controlled insulin-dependent diabetes mellitus (CIDD). Ten nondiabetic, age- and gender-matched individuals served as controls. Thirty-nine biopsies were processed for light microscopy, and the blood vessel area was analyzed using an interactive digital analyzing system; 9 gingival biopsies, 5 diabetic and 4 controls, were processed for morphometric electron microscopic analysis. For each individual, site-specific recordings were made for the plaque index, bleeding index, probing depth, loss of attachment, and radiographic loss of interproximal alveolar bone. No evident signs of periodontitis occurred at the biopsy sites. For each PIDD patient, respective volumetric and numeric densities of cellular components including fibroblasts, neutrophilic granulocytes, monocyte/macrophages, mast cells, lymphocytes, blast cells, and plasma cells were recorded in the inflamed connective tissue (ICT). Non-cellular components such as collagen fibers and blood vessels were also recorded. PIDD patients had elevated plasma cell levels relative to controls and they appeared also to have a decreased collagen fiber density. In addition, fibroblasts occupied less volume in the ICT of PIDD patients than in controls. PIDD patients had the largest mean area of cross-section of the blood vessels, but this difference was not statistically significant (P > or = 0.211; t-test). No specific characteristics of ICT or vascular changes were detectable in adult well-controlled long-term diabetics under similar plaque conditions. Swollen and proliferated endothelial cells were frequently found in PIDD patients and the mean distance from the lumen to the outer border of basement membrane was greater in the PIDD than in the controls (P < 0.001; t-test). Overall, our findings that cellular, vascular, and connective tissue changes indicative of increased catabolism rather than anabolism detected in gingiva are especially associated with poorly controlled long-term insulin-dependent diabetes.     

Cardiovascular actions of cannabinoids and their generation during shock

Marijuana is a widely abused recreational drug well known for its psychoactive properties. Cannabinoids, the active ingredients of marijuana, elicit their neurobehavioral effects by interacting with the CB1 cannabinoid receptor subtype, expressed primarily in the brain but also present in some peripheral tissues. A second receptor subtype, the CB2 receptor, is expressed on cells of the immune system and is thought to be responsible for the immunosuppressant effects of cannabinoids. Recently, endogenous lipidlike substances have been identified, including arachidonyl ethanolamide (anandamide) and 2-arachidonyl glyceride, that bind to cannabinoid receptors and mimic many of the neurobehavioral effects of plant-derived cannabinoids. Both plant-derived cannabinoids and the endogenous ligands have been shown to elicit hypotension and bradycardia via activation of peripherally located CB1 receptors. Possible underlying mechanisms include presynaptic CB1 receptor mediated inhibition of norepinephrine release from peripheral sympathetic nerve terminals, and/or direct vasodilation via activation of vascular cannabinoid receptors. The latter may also be the target of endocannabinoids of vascular endothelial origin. Recent studies indicate that a peripheral endogenous cannabinoid system in circulating macrophages and platelets is activated in hemorrhagic and septic shock and may contribute to the hypotension associated with these conditions via activation of vascular cannabinoid receptors. The potential role of this mechanism in human shock conditions is under investigation.     

Receptor-associated protein (RAP): a specialized chaperone for endocytic receptors

A number of cellular control mechanisms have evolved that facilitate and evaluate post-translational steps in protein biosynthesis. Chaperones or escort proteins are an important part of these cellular control mechanisms. They associate with newly synthesized proteins and assure correct folding and post-translational modification including disulfide bridge formation, glycosylation and complex formation. The receptor-associated protein (RAP) is a novel type of chaperone recently identified that is especially designed to assist in the biosynthesis and intracellular transport of endocytic receptors. Experimental evidence suggests that RAP acts as a receptor antagonist and prevents association of newly synthesized receptors with their ligands during transport to the cell surface. This mechanism seems to be required in cell types that express both receptor and ligand because premature receptor-ligand interaction in the secretory pathway interferes with proper export of the receptors to the cell surface. This review describes studies that have uncovered this unique protein biosynthesis mechanism.     

Insulin resistance implications for type II diabetes mellitus and coronary heart disease

PURPOSE: To review information on the implications of insulin resistance for type II diabetes mellitus (non-insulin-dependent diabetes mellitus) and coronary heart disease, and to derive guidance from this information for the management of these conditions. DATA SOURCES: A MEDLINE search of English-language articles published between 1985 and July 1996, and review of the bibliographies of articles obtained through the MEDLINE search and textbooks. STUDY SELECTION: Primary research articles, reviews and perspectives on the epidemiology of diabetes and cardiovascular diseases and on intervention outcomes in these diseases. DATA EXTRACTION: Study design and quality were assessed, with particular attention to methods, study population size and other characteristics. Conclusions of review articles and perspectives were analyzed critically. DATA SYNTHESIS: Type II diabetes is associated with a two- to fourfold excess of coronary heart disease, compared to nondiabetic populations. In most studies, glycemia and duration of clinical diabetes were found to be only weak risk factors for coronary heart disease. Conventional coronary heart disease risk factors such as dyslipidemia and hypertension have been associated with coronary heart disease in type II diabetes subjects. Hyperinsulinemia and insulin resistance have been predictive of the development of type II diabetes and, in some studies, of coronary heart disease. CONCLUSION: Strategies to prevent the development of coronary heart disease in diabetic and possibly prediabetic subjects should emphasize a multifactorial approach, including: a) improved glycemic control; b) aggressive treatment of risk factors for coronary heart disease, including insulin resistance; c) primary prevention of NIDDM; and d) use of glucose lowering agents that improve insulin sensitivity and cardiovascular risk factors.     

Myocardial, neural and vascular aspects of ischemic preconditioning

Ischemic preconditioning can be obtained with brief coronary occlusions. It has been studied in different animal species including dogs, pigs, rabbits and rats. The suggested duration of the occlusions ranges from four periods of 5 min, separated from each other by 5 min of reperfusion, to one period of 2.5 min. In addition to the reduction of the size of a subsequent infarction, preconditioning is responsible for the attenuation of the ischemia-reperfusion injury. The protection has a short duration and does not exceed two hours. Myocardial, neural and endothelial factors are involved in preconditioning. The myocardial component includes an increased release of adenosine with activation of A1 adenosine receptors, the activation of a protein-kinase C and possibly of antioxidant enzymes. The neural component includes a reduction in the release of noradrenaline from the postganglionic sympathetic fibers and a reduced myocardial sensitivity to noradrenaline. The increased myocardial release of adenosine, together with the reduced adrenergic activity, is consistent with the reduction in myocardial metabolism which has been observed after preconditioning. The coronary vascular endothelium is concerned in an increased release of nitric oxide which seems to be responsible for a prevention of reperfusion arrhythmias. In addition to the protective effect exerted on the myocardium, ischemic preconditioning seems to be responsible for a change in the coronary responsiveness to short periods of occlusion followed by release. This change in responsiveness is mainly represented by a greater velocity of the increase in flow occurring in the coronary reactive hyperemia.     

Diabetic periodontitis: possible lipid-induced defect in tissue repair through alteration of macrophage phenotype and function

BACKGROUND: Diabetes mellitus is a major health problem in the United States affecting approximately 13 million people. The five 'classic' complications which have historically been associated with the condition are microangiopathy, neuropathy, nephropathy, microvascular disease, and delayed wound healing. Recently, periodontal disease (PD) has been declared the 'sixth' major complication of diabetes as diabetics demonstrate an increased incidence and severity of PD. The cellular and molecular basis for diabetic PD is unknown. HYPOTHESIS: Recent evidence suggests that PD and delayed dermal wound healing may be manifestations of the same general systemic deficit in diabetes involving impairment of the cellular and molecular signal of wounding via alterations in macrophage phenotype. Diabetes-induced hyperlipidemia may interfere with the normal cellular and molecular signal of wounding by alteration of macrophage function and subsequent dysregulation of cytokines at the wound site. RESULTS: Preliminary data in both animal models and humans suggests that hyperglycemia, in combination with elevations of serum low density lipoproteins and triglycerides, leads to formation of advanced glycation end products (AGEs) which may alter macrophage phenotype. This may be responsible for dysregulation of macrophage cytokine production and increased inflammatory tissue destruction and alveolar bone loss. IMPLICATIONS: Future investigations will consider diabetic PD in the context of a generalized systemic wound healing deficit that manifests as PD in the face of constant pathologic wounding of the gingiva (bacterial plaque) or delayed dermal wound healing in instances of periodic traumatic wounding to other parts of the body. These types of studies will provide information concerning defective tissue repair in diabetics that will have clinical relevance for the understanding of PD and delayed dermal healing as well as applications of appropriate and specific therapies.     

Prevention of atherosclerosis in diabetics

The high prevalence of atherosclerotic (macrovascular) complications in diabetes (1.5-6x higher than in non-diabetics) stimulated evaluation of new pathogenetic findings which could have an impact on prevention. In type 1 diabetics the development of nephropathy is a factor hastening the development of macroangiopathy. In type 2 diabetics on whom attention is concentrated at the moment interaction of various metabolic deviations is involved which include changes of lipoproteins (drop of HDL, changes in the size and composition of LDL), insulin resistance and glycosylation of proteins. There is an enhanced tendency to lipoprotein oxidation (LDL) which promote the development of cholesterol rich plaques in the arterial walls. Their rupture may cause occlusion and ensuing risks for life. Possibilities of prevention are not adequately made use of. This is due to a tendency to underrate the serious character of type 2 diabetes and also the high percentage of diabetics where the disease was diagnosed late. The metabolic syndrome which develops as a result of insulin resistance precedes for years manifestations of diabetes. Its detection makes it possible to screen subjects at risk, some of whom develop diabetes. At the same time it is also a pathogenetic factor of macrovascular complications. It leads to the cumulation of a number of risk conditions. More effective prevention can be implemented by intervention of all associated risk factors (smoking, hypertension), in the application of lifestyle provisions of energy reduction by promoting physical activity and by a rational diet (diabetes, obesity, hyperlipidaemia). The justification of pharmacotherapy for the high risk groups of diabetics with hyperlipidaemia is supported by results of recently published investigations where statins were used. For the sub-population of subjects at risk the perspective should be screening of risk factors, early diagnosis of diabetes, education, continuous primary care, comprehensive prevention using lifestyle provisions as well as advances in modern pharmacotherapy.     

Recent progress in clinical aspects of receptor research

Clinical receptology encompasses broad areas, including receptor or postreceptor defects due to mutations of receptor or other genes, abnormalities due to receptor antibodies and secondary changes of receptors under various pathological conditions. Recent progress in molecular biology has succeeded in cloning genes of receptors, G-proteins and other cellular proteins that are involved in the signal transduction and clarified their germ-line and somatic mutations. It is of importance that mutations of receptors and G-proteins do not necessarily cause loss of function but sometimes cause gain of function of receptors or G-proteins, thus leading to hyperfunction. Molecular basis that causes either loss or gain of function has been studied but is not completely understood. Some examples of gain of function mutatious of G-protein coupled receptors, tyrosin kinase-type receptors and G alpha protein are shown. Another important aspect in receptor research is that mutation of a single receptor gene sometimes result in different phenotypes and even different modes of inheritance. For example, mutations of rhodopsin (a G-protein coupled receptor) gene cause retinitis pigmentosa of autosomal dominant type and autosomal recessive type and also cause congenital stationary night blindness. Exact mechanisms responsible for such differences are not completely understood. There are polymorphisms in some genes that may be involved in some diseases. An example is a polymorphism in beta 3-adrenergic receptor that is claimed but not clearly demonstrated to be a cause of obesity or type II diabetes. Such polymorphism is possibly a gene in polygenic diseases. Receptology is important for elucidating pathogenesis of complex diseases.     

Lens glutathione, lens protein glycation and electrophoretic patterns of lens proteins in STZ induced diabetic rats

As diabetes is a very complex disease, with the pathological symptoms varying with age, diabetic type and means of control, it still warrants many in vivo and in vitro studies. During hyperglycaemia, increases in the sorbitol pathway, nonenzymatic glycosylation of lens proteins and damage to antioxidant systems have been reported to cause opacification of the lens leading to cataract formation. In this study, intracapsular extracts of lenses from STZ induced diabetic female rats were examined. Total protein, glutathione and nonenzymatic glycosylation were determined by the Lowry, Ellman reagent and thiobarbituric acid methods respectively. Laemmli protein electrophoresis was also carried out on the lens homogenates. After a period of as short as 5 weeks, a decrease in lens glutathione, and an increase in nonenzymatic glycosylation of lens proteins were found. The electrophoresis showed an increase in proteins of high molecular weight.     

Opposing mitogenic and anti-mitogenic actions of parathyroid hormone-related protein in vascular smooth muscle cells: a critical role for nuclear targeting

Parathyroid hormone-related protein (PTHrP) is a prohormone that is posttranslationally processed to a family of mature secretory forms, each of which has its own cognate receptor(s) on the cell surface that mediate the actions of PTHrP. In addition to being secreted via the classical secretory pathway and interacting with cell surface receptors in a paracrine/autocrine fashion, PTHrP appears to be able to enter the nucleus directly following translation and influence cellular events in an "intracrine" fashion. In this report, we demonstrate that PTHrP can be targeted to the nucleus in vascular smooth muscle cells, that this nuclear targeting is associated with a striking increase in mitogenesis, that this nuclear effect on proliferation is the diametric opposite of the effects of PTHrP resulting from interaction with cell surface receptors on vascular smooth muscle cells, and that the regions of the PTHrP sequence responsible for this nuclear targeting represent a classical bipartite nuclear localization signal. This report describes the activation of the cell cycle in association with nuclear localization of PTHrP in any cell type. These findings have important implications for the normal physiology of PTHrP in the many tissues which produce it, and suggest that gene delivery of PTHrP or modified variants may be useful in the management of atherosclerotic vascular disease.     

Involvement of regional lymph nodes after penetration of Schistosoma mansoni cercariae in naive and infected mice

The very low density lipoprotein receptor (VLDL-r) is a cell-surface molecule specialized for the internalization of multiple diverse ligands, including apolipoprotein E (apoE)-containing lipoprotein particles, via clathrin-coated pits. Its structure is similar to the low-density lipoprotein receptor (LDL-r), although the two have substantially different systemic distributions and regulatory pathways. The present work examines the distribution of VLDL-r in the central nervous system (CNS) and in relation to senile plaques in Alzheimer disease (AD). VLDL-r is present on resting and activated microglia, particularly those associated with senile plaques (SPs). VLDL-r immunoreactivity is also found in cortical neurons. Two exons of VLDL-r mRNA are differentially spliced in the mature receptor mRNA. One set of splice forms gives rise to receptors containing (or lacking) an extracellular O-linked glycosylation domain near the transmembrane portion of the molecule. The other set of splice forms appears to be brain-specific, and is responsible for the presence or absence of one of the cysteine-rich repeat regions in the binding region of the molecule. Ratios of the receptor variants generated from these splice forms do not differ substantially across different cortical areas or in AD. We hypothesize that VLDL-r might contribute to metabolism of apoE and apoE/A beta complexes in the brain. Further characterizations of apoE receptors in Alzheimer brain may help lay the groundwork for understanding the role of apoE in the CNS and in the pathophysiology of AD.     

Small GTP-binding proteins in the brain-corpus cardiacum-corpus allatum complex of the silkworm, Bombyx mori: involvement in the secretion of prothoracicotropic hormone

BACKGROUND: Proteolysis, modulated in part by intramural fibrinolytic system proteins and their inhibitors, appears to influence vascular smooth muscle cell (SMC) migration and proliferation and remodeling of extracellular matrix (ECM). Alterations of fibrinolysis in circulating blood and of proteolysis within vessel walls in experimental animals and patients with diabetes have been associated with accelerated vascular disease. Hyaluronan, a prominent component of ECM in normal vessels, is increased in the tunica media of macroscopically normal arterial vessels from patients with type 2 diabetes. OBJECTIVE: To determine whether hyaluronan alters the expression of the fibrinolytic system protein, plasminogen activator inhibitor type-1 (PAI-1), in human vascular SMCs, thereby potentially accelerating vascular disease in patients with type 2 diabetes. METHODS: Urokinase-type and tissue-type plasminogen activators (uPA and tPA) and PAI-1 were assayed in vascular SMC conditioned media and in cell lysates, using enzyme-linked immunosorbent assay and western blotting. RESULTS: Hyaluronan increased the 24-h release of PAI-1 into conditioned media in a concentration-dependent and time-dependent manner (1.8-fold compared with control with 1 mg/ml hyaluronan; n = 9, P < 0.01). Although the accumulation of uPA in conditioned media tended to increase also, uPA content was reduced in cell lysates (64% of control with 0.1 mg/ml hyaluronan at 24 h; n = 9, P < 0.01) without any change in PAI-1. Concentrations of tPA in conditioned media and cell lysates were unchanged. Digestion of hyaluronan with hyaluronidase (50 turbidity reducing units (TRU)/ml) or exposure of the smooth muscle cells to antihuman CD44 antibody (1 microgram/ml) that binds to the hyaluronan cell surface receptor obviated the effects of hyaluronan. CONCLUSION: Our results indicate that increases in hyaluronan increase vascular SMC expression of PAI-1, a phenomenon that may alter the balance between proteolysis and its inhibition in vessels of patients with type 2 diabetes, thereby contributing to the acceleration of macroangiopathy.     

Ethnicity and cardiovascular risk: variations in people of African ancestry and South Asian origin

Mortality from coronary heart disease (CHD), stroke and end-stage renal failure are high in South Asian migrants in the UK. This is associated with high prevalence of diabetes and hypertension. These seem to be manifestations of a metabolic syndrome with insulin resistance (hyperinsulinaemia) and central obesity (based on high waist-to-hip ratio rather than on conventional measures of body mass index). This is associated with sedentary lifestyle, high serum triglycerides and low HDL-cholesterol. Mortality from stroke and end-stage renal failure are high in black migrants to the UK (both Caribbeans and West Africans). However, CHD mortality is low in this group. This pattern of mortality is associated with high prevalence of hypertension and diabetes. This group tends to be obese (particularly women) according to conventional measures of body mass index and to have hyperinsulinaemia, low serum triglycerides and high HDL-cholesterol. Conventional risk factors such as cigarette smoking and hypercholesterolaemia are less prevalent in ethnic minority populations in the United Kingdom and unlikely to explain the differences seen between groups, although each risk factor is likely to contribute to the variation in vascular disease within each group. There is difficulty in reconciling the results of migration studies (eg, from rural to urban environments) pointing to major environmental influences on the changes in cardiovascular risk factors with the consistent pattern of disease of ethnic groups across the world and in subsequent generations, suggesting a certain degree of genetic susceptibility. Important environment-gene interplays might be underlying some of these processes. The detection and management of hypertension and diabetes are still unsatisfactory in inner city areas and show variations by ethnic origin. Strategies for the control of CHD and stroke adopted in European countries directed mostly to white populations may be inappropriate for ethnic minority populations.     

Targeted overexpression of protein kinase C beta2 isoform in myocardium causes cardiomyopathy

Increased cardiovascular mortality occurs in diabetic patients with or without coronary artery disease and is attributed to the presence of diabetic cardiomyopathy. One potential mechanism is hyperglycemia that has been reported to activate protein kinase C (PKC), preferentially the beta isoform, which has been associated with the development of micro- and macrovascular pathologies in diabetes mellitus. To establish that the activation of the PKCbeta isoform can cause cardiac dysfunctions, we have established lines of transgenic mice with the specific overexpression of PKCbeta2 isoform in the myocardium. These mice overexpressed the PKCbeta2 isoform transgene by 2- to 10-fold as measured by mRNA, and proteins exhibited left ventricular hypertrophy, cardiac myocyte necrosis, multifocal fibrosis, and decreased left ventricular performance without vascular lesions. The severity of the phenotypes exhibited gene dose-dependence. Up-regulation of mRNAs for fetal type myosin heavy chain, atrial natriuretic factor, c-fos, transforming growth factor, and collagens was also observed. Moreover, treatment with a PKCbeta-specific inhibitor resulted in functional and histological improvement. These findings have firmly established that the activation of the PKCbeta2 isoform can cause specific cardiac cellular and functional changes leading to cardiomyopathy of diabetic or nondiabetic etiology.     

Targeting of proteins to the Golgi apparatus

The proteins that reside in the Golgi carry out functions associated with post-translational modifications, including glycosylation and proteolytic processing, membrane transport, recycling of endoplasmic reticulum proteins and maintenance of the structural organisation of the organelle itself. The latter includes Golgi stacking, interconnections between stacks and the microtubule-dependent positioning of the organelle within the cell. There are a number of distinct groups of Golgi membrane proteins, including glycosyltransferases, recycling trans-Golgi network (TGN) proteins, peripheral membrane proteins and receptors. Considerable effort has been directed at understanding the basis of the localisation of Golgi glycosyltransferases and recycling TGN proteins; in both cases there is increasing evidence that multiple signals may be involved in their specific localisation. A number of models for the Golgi retention of glycosyltransferases have been proposed including oligomerisation, lipid-mediated sorting and intra-Golgi retrograde transport. More information is required to determine the contribution of each of these potential mechanisms in the targeting of different glycosyltransferases. Future work is also likely to focus on the relationship between the localisation of resident Golgi proteins and the maintenance of Golgi structure.