Irbesartan treatment does not influence plasma levels of the dicarbonyls methylglyoxal, glyoxal and 3-deoxyglucosone in participants with type 2 diabetes and microalbuminuria: An IRMA2 sub-study.

AIM: Angiotensin receptor blockers (ARBs) reduce vascular complications in diabetes independently of blood pressure. Experimental studies suggested that ARBs may restore the detoxifying enzyme glyoxalase 1, thereby lowering dicarbonyls such as methylglyoxal. Human data on the effects of ARBs on plasma dicarbonyl levels are lacking. We investigated, in individuals with type 2 diabetes, whether irbesartan lowered plasma levels of the dicarbonyls methylglyoxal, glyoxal, 3-deoxyglucosone and their derived advanced glycation end products (AGEs), and increased d-lactate, reflecting greater methylglyoxal flux. METHODS: We analysed a subset of the Irbesartan in Patients with T2D and Microalbuminuria (IRMA2) study. We measured plasma dicarbonyls methylglyoxal, glyoxal and 3-deoxyglucosone, free AGEs and d-lactate using ultra-performance liquid chromatography tandem mass-spectrometry (UPLC-MS/MS) in the treatment arm receiving 300 mg irbesartan (n = 121) and a placebo group (n = 101) at baseline and after 1 and 2 years. Effect of treatment was analysed with repeated measurements ANOVA. RESULTS: There was a slight, but significant difference in baseline median methylglyoxal levels [placebo 1119 (907-1509) nmol/l vs. irbesartan 300 mg 1053 (820-1427) nmol/l], but no significant changes were observed in any of the plasma dicarbonyls over time in either group and there was no effect of irbesartan treatment on plasma free AGEs or d-lactate levels at either 1 or 2 years. CONCLUSION: Irbesartan treatment does not change plasma levels of the dicarbonyls methylglyoxal, glyoxal and 3-deoxyglucosone, free AGEs or d-lactate in type 2 diabetes. This indicates that increased dicarbonyls in type 2 diabetes are not targetable by ARBs, and other approaches to lower systemic dicarbonyls are needed in type 2 diabetes. (Clinical Trial Registry No: #NCT00317915).

Fasting and post-oral-glucose-load levels of methylglyoxal are associated with microvascular, but not macrovascular, disease in individuals with and without (pre)diabetes: The Maastricht Study.

AIMS: Reactive dicarbonyl compounds, such as methylglyoxal (MGO), rise during an oral glucose tolerance test (OGTT), particularly in (pre)diabetes. Fasting MGO levels are associated with chronic kidney disease (CKD) and cardiovascular disease (CVD) in patients with poorly controlled type 2 diabetes mellitus (T2DM). Yet, whether fasting or post-OGTT plasma MGO levels are associated with vascular disease in people with (pre)diabetes is unknown. METHODS: Subjects with normal glucose metabolism (n=1796; age: 57.9±8.2 years; 43.3% men), prediabetes (n=478; age: 61.6±7.6 years; 54.0% men) and T2DM (n=669; age: 63.0±7.5 years; 67.0% men) from the Maastricht Study underwent OGTTs. Plasma MGO levels were measured at baseline and 2h after OGTT by mass spectrometry. Prior CVD was established via questionnaire. CKD was reflected by estimated glomerular filtration rate (eGFR) and albuminuria; retinopathy was assessed using retinal photographs. Data were analyzed using logistic regression adjusted for gender, age, smoking, systolic blood pressure, total-to-HDL cholesterol ratio, triglycerides, HbA1c, BMI and medication use. Odd ratios (ORs) were expressed per standard deviation of LN-transformed MGO. RESULTS: Fasting and post-OGTT MGO levels were associated with higher ORs for albuminuria ≥30mg/24h [fasting: 1.12 (95% CI: 0.97-1.29); post-OGTT: 1.19 (1.01-1.41)], eGFR<60mL/min/1.73 m2 [fasting: 1.58 (95% CI: 1.38-1.82), post-OGTT: 1.57 (1.34-1.83)] and retinopathy [fasting: 1.59 (95% CI: 1.01-2.53), post-OGTT: 1.38 (0.77-2.48)]. No associations with prior CVD were found. CONCLUSION: Fasting and post-OGTT MGO levels were associated with microvascular disease, but not prior CVD. Thus, therapeutic strategies directed at lowering MGO levels may prevent microvascular disease.

NO-mediated cGMP synthesis in cultured cholinergic neurons from the basal forebrain of the fetal rat.

Previously, using brain slices, we reported NO-mediated cGMP synthesis in all cholinergic fibers in the rat neocortex. In order to answer the question whether this property of cholinergic fibers was present before or developed after birth, we investigated properties of NO-responsiveness of cultured cholinergic forebrain neurons. Basal forebrain neurons of E16 fetal rat were cultured. Under the conditions chosen and after one day of culturing, all cells had attained a cholinergic phenotype using choline acetyltransferase or the vesicular acetylcholine transporter molecule as markers. Between 95-99% of the cells also expressed neuronal NOS. In the presence of 1 mM IBMX, a non-selective phosphodiesterase (PDE) inhibitor, 10 microM of the NO donor diethylamine-NONOate (DEANO) increased cGMP synthesis in 80% of the cells. cGMP levels in the cultured forebrain neurons were also increased when cells were stimulated with DEANO in the presence of the selective PDE inhibitors BAY 60-7550 (PDE2), sildenafil (PDE5), or the mixed type inhibitor papaverine (PDE2,5,10). Subpopulations of cells from the basal forebrain expressed mRNA for PDE2, PDE5, and PDE9. Atropine increased cGMP levels in an NO-dependent manner in a small population of cultured forebrain cells in the presence of IBMX. In conclusion, cultured cholinergic basal forebrain neurons present a heterogeneous cell population in the magnitude of their response to NO. NO-responsiveness of the cultured cholinergic neurons is already detectable after one day of culturing and indicates that NO-sensitivity of the cholinergic neurons of the rat basal forebrain is present well before birth.

Nitric oxide-induced cGMP synthesis in the cholinergic system during the development and aging of the rat brain.

cGMP synthesis in cholinergic neurons of the basal forebrain, the caudate putamen, and the tegmento-pedunculopontine nucleus of the rat was studied during development after birth at P1, P4, P10, and P21, in the adult, and during aging. NO-mediated cGMP synthesis in these neurons was studied using the approach of in vitro incubation of brain slices in combination with cGMP-immunocytochemistry. The percentage of NO-responsive, cGMP-synthesizing cholinergic cells in the septum and diagonal band of Broca decreased from 75% to 6% in adult animals and to 2% in aged ones. In the caudate putamen, this decrease was from 81% to 21% in adult and 11% in aged animals. Cholinergic cells of the tegmento-pedunculopontine nucleus were unresponsive to NO and never showed cGMP-immunoreactivity. In addition, it was observed that the amount of NO-responsive, cGMP-synthesizing cholinergic fibers in the hippocampus declined in parallel with the maturation of the septal-hippocampal cholinergic pathway, whereas in the caudate putamen, this colocalization became complete 2 weeks after birth. It is concluded that the property of NO-mediated cGMP synthesis in the cholinergic nuclei of the forebrain is developmentally regulated after birth and that NO-cGMP signal transduction has a role in establishing cholinergic neuronal connections in the hippocampus and caudate putamen.

Levels of basic fibroblast growth factor, glutamine synthetase, and interleukin-6 in subretinal fluid from patients with retinal detachment.

PURPOSE: To investigate the presence of basic fibroblast growth factor, glutamine synthetase activity, and interleukin-6 in subretinal fluid from patients with retinal detachment. METHODS: In a prospective study we measured basic fibroblast growth factor, glutamine synthetase activity, interleukin-6, and total protein in subretinal fluid samples from 96 eyes from 94 consecutive patients with a retinal detachment corrected by a conventional scleral buckling operation in our clinical practice. As controls, vitreous fluid samples from eyes with a macular hole (n = 6) or pucker (n = 11) were used. Laboratory data of the patient group were compared with the control group and correlated with various clinical data. RESULTS: Levels (median, range) of basic fibroblast growth factor, glutamine synthetase activity, interleukin-6, and total protein were significantly higher in patients than in controls (P <.0001). An increased level of glutamine synthetase and total protein correlated with a longer duration of the retinal detachment (r =.4, P =.002, and r =.34, P =.001, respectively). Interleukin-6 and basic fibroblast growth factor levels did not correlate with the duration of the detachment. After multivariate logistic regression analysis, no significant relation was found between any of the tested subretinal proteins and a low visual outcome or redetachment. CONCLUSIONS: We found increased levels of basic fibroblast growth factor and glutamine synthetase in subretinal fluid from patients with retinal detachment. Basic fibroblast growth factor and glutamine synthetase may play a role in the pathogenesis and recovery after retinal detachment. The questions of whether the increased levels of basic fibroblast growth factor and glutamine synthetase result from leakage of dying glia cells (including Müller cells) and neurons and if basic fibroblast growth factor is actively produced to protect the photoreceptor cells need further research.