Endothelial Dysfunction in Patients with Chronic Kidney Disease Results from Advanced Glycation End Products (AGE)-Mediated Inhibition of Endothelial Nitric Oxide Synthase through RAGE Activation

Background and objectives: Advanced glycation end products, known pro-inflammatory and pro-oxidative compounds that accumulate in patients with chronic kidney disease, may play a major role in their high prevalence of endothelial dysfunction and subsequent cardiovascular disease. This study examined the association of advanced glycation end product accumulation with cellular receptor for advanced glycation end product expression and endothelial dysfunction as well as the mechanisms of this association in chronic kidney disease.Design, setting, participants, & measurements: A cross-sectional study was conducted of ambulatory patients without diabetes and with different stages of chronic kidney disease (n = 51), compared with gender- and age-matched healthy subjects. Fasting blood was obtained for measurement of advanced glycation end products and mRNA receptor for advanced glycation end product expression in peripheral blood mononuclear cells. Endothelial reactivity was assessed by the microcirculatory response to local ischemia (postocclusive reactive hyperemia) and local hyperthermia (thermal hyperemia). Sera were pooled and passed through affinity columns to separate advanced glycation end product–rich fractions, which were incubated with human aortic endothelial cells, with or without blockade of receptor for advanced glycation end product, to measure their effect on endothelial nitric oxide synthase.Results: Glomerular filtration rate correlated with serum advanced glycation end product, mRNA receptor for advanced glycation end product levels, postocclusive reactive hyperemia, and thermal hyperemia. Serum advanced glycation end product correlated with receptor for advanced glycation end product and inversely with postocclusive reactive hyperemia. Advanced glycation end product–rich fractions from chronic kidney disease sera suppressed endothelial nitric oxide synthase expression of human aortic endothelial cells compared with sera from healthy subjects, an effect abrogated by receptor for advanced glycation end product blockade.Conclusions: This study demonstrates for the first time an association of excess advanced glycation end product burden with increased peripheral blood mononuclear cell mRNA receptor for advanced glycation end product and in vivo endothelial dysfunction in patients with chronic kidney disease. Endothelial dysfunction in chronic kidney disease may be partly mediated by advanced glycation end product–induced inhibition of endothelial nitric oxide synthase through receptor for advanced glycation end product activation.The epidemiologic association between chronic kidney disease (CKD) and cardiovascular disease (CVD) has been firmly established (1–3). Traditional CVD risk factors such as hypertension and hyperlipidemia do not fully explain the high prevalence of CVD in CKD (1). The recognition that inflammation, oxidative stress (OS), and endothelial dysfunction (ED) are common abnormalities in CVD has led to a detailed exploration of these pathways in search of new risk factors (4–8). Because renal failure is associated with increasing circulating levels of advanced glycation end products (AGE) (9) and these compounds are known to promote inflammation, OS, and ED, they are likely to play an important role in the pathogenesis of CVD in this population (10–12).Numerous in vitro and animal studies have implicated AGE in the pathogenesis of CVD (10–12) via several receptor-independent and receptor-dependent mechanisms. For instance, AGE have direct influence on the structural integrity of the vessel wall and the underlying basement membranes in part induced by cross-linking of subendothelial matrix molecules, such as collagen, or by disruption of matrix–matrix and matrix–cell interactions (13,14). Other actions of AGE include nitric oxide (NO) quenching (15) and endothelial NO synthase (eNOS) inhibition (16), thereby adversely affecting vascular endothelium and its protective functions, particularly vascular relaxation. The direct mechanistic links between AGE and their endothelial effects have not been clearly delineated but may include enhanced expression and activity of receptor for AGE (RAGE) (17). In fact, expression of RAGE on peripheral blood monocytes has been found to be upregulated in patients without diabetes and with CKD (18).The link among OS, inflammation, and ED as a new paradigm of atherosclerosis provides the rationale for a variety of studies, including flow-mediated vasodilation and reactive hyperemia of the microcirculation, gauging endothelial function as an index of the susceptibility of the vasculature to atherosclerosis (19–26). This noninvasive interrogation of the cutaneous microvasculature responses to both flow and thermal stimulations may offer a simple screening test for the presence of systemic ED (26). ED, defined by these noninvasive vascular tests, is a common finding in patients with CKD (22–25) and could reflect reduced NO generation (27) or impaired activity (16). Possible causes of NO deficiency are substrate (l-arginine) limitation as a result of perturbed renal biosynthesis of this amino acid or increased levels of circulating endogenous inhibitors of eNOS, such as asymmetric dimethylarginine, homocysteine, or AGE (26). Sera from patients with CKD as well as in vitro prepared AGE significantly suppress eNOS activity in cultured vascular endothelial cells (16,28), suggesting that the excess AGE in CKD may contribute to their ED.Notwithstanding all of the experimental data suggesting a major role for AGE in causing ED/CVD, the few human studies correlating serum AGE levels with cardiovascular outcome have given conflicting results. Two studies of hemodialysis patients who were followed for a variable period of time demonstrated that serum AGE levels were strong and independent predictors of mortality (29,30). Conversely, a study from Germany showed that high serum AGE levels at baseline were associated with better outcome in hemodialysis patients who were followed for 32 mo (31) and a group in Sweden showed that plasma pentosidine did not predict outcome of a group of patients who had ESRD and were identified close to the start of renal replacement therapy (32)Despite the long-term postulation that AGE excess could lead to ED in CKD, no study has specifically described the relationship between circulating AGE levels and in vivo tests of endothelial function in these patients; therefore, we performed a study to examine this relationship in a group of patients with CKD and measured two widely known circulating AGE (^ɛN-carboxymethyl-lysine [CML] and methyl-glyoxal [MG] derivatives) and the in vivo arteriolar vasodilatory response to both local ischemia and hyperthermia in a cohort of patients without diabetes and with CKD. Furthermore, we wanted to test whether the observed eNOS impairment in CKD could result from the excess of circulating AGE and whether this effect was mediated via RAGE. For this, we evaluated the effect of AGE from serum of patients with CKD on the eNOS expression of human aortic endothelial cells (HAEC) with or without blockade of RAGE expression. We also measured RAGE expression in peripheral mononuclear cells as a surrogate indicator of RAGE expression in endothelial cells. The findings suggest that CKD-related ED is due partly to excess accumulation of AGE, which activate RAGE to suppress eNOS in the vasculature.     

Mother-child relationship—FIRST score*

This paper describes the use of the MCR FIRST score. The score has evolved as a method of measuring certain aspects of a mother's caring for her newborn baby. The factors included are feeding, interest, response, speech and touch, and these have been combined into a numerical score. The results for 100 mothers who were assessed on days 1, 3 and 5 postpartum are presented. The MCR FIRST score is distinguished by its simplicity of use in clinical work and may be helpful in recognizing potential impairment of mother-child relationships during the early postpartum period.     

Polymorphism on leflunomide: stability and crystal structures

Two polymorphs of Leflunomide were found and studied (form I and II). Both of them were characterized by X-ray powder diffraction and thermal analysis. Single crystals were obtained and both structures were solved. Forms I and II crystallize in the space group P2(1)/c with two and one independent molecules per asymmetric unit, respectively. Thermodynamic stability of the two forms is assessed by differential scanning calorimetry. The cohesion in the crystal of form I (the more stable) is provided by both by H bonding as well as pi...pi interactions, while in form II it is given only by the former. The independent molecules in form I adopt different conformations thus allowing for a larger number of intermolecular interactions.     

Implicit operant learning of pain sensitization

Operant conditioning mechanisms have been demonstrated to be important in the development of chronic pain behavior, but it is not clear whether and how this extends to pain perception itself. The fear-avoidance theory suggests that hypersensitivity may be induced by anticipatory pain avoidance learned through negative reinforcement by acute reductions of pain and fear. But the precise mechanism of the assumed 'sensory decalibration' has not been specified. The present study with healthy subjects investigated whether operant learning of enhanced short-term sensitization may provide the 'proximal' mechanism and whether gradual learning of hypersensitivity can take place without subjects' awareness. We used an experimental model of implicit learning based on a behavioral adjustment method of sensitization measurement developed and validated previously, combining it with standard methods of operant response shaping of increased sensitization or habituation. Results indicated that operant discrimination training with reinforcement of short-term sensitization in the seconds range can produce gross up or down changes in sensitivity within an hour without subjects' awareness of reinforcement contingencies. Consequently, implicit learning of enhanced pain sensitization may be a suitable model to investigate operant plasticity of pain perception in addition to basic sensory and neuronal mechanisms and to link these with the clinical construct of pain-fear avoidance.     

Comparison of medium, temperature, and length of incubation for detection of vancomycin-resistant Enterococcus

Campylobacter (Campy; BD Diagnostics, Sparks, MD), Spectra VRE (Remel, Lenexa, KS), and bile-esculin-azide-vancomycin (BEAV; Remel) agars were compared for their ability to detect vancomycin-resistant enterococci (VRE) in 750 stool specimens. The media were compared at 24 h and 48 h of incubation at 35°C and 42°C. When incubated for 24 h at 35°C, Campy was the most sensitive (97.8%) and specific (99.9%) but was comparable to Spectra, which has a sensitivity of 95.6% and a specificity of 99.1%, whereas BEAV was significantly less sensitive (90%) and specific (96.1%). Incubation at 42°C or extended incubation at 35°C for 48 h yielded no advantage over incubation at 35°C for 24 h.