Circulating metabolomic markers in association with overall burden of microvascular complications in type 1 diabetes.

INTRODUCTION: Diabetic retinopathy (DR), diabetic kidney disease (DKD) and distal symmetric polyneuropathy (DSPN) share common pathophysiology and pose an additive risk of early mortality. RESEARCH DESIGN AND METHODS: In adults with type 1 diabetes, 49 metabolites previously associated with either DR or DKD were assessed in relation to presence of DSPN. Metabolites overlapping in significance with presence of all three complications were assessed in relation to microvascular burden severity (additive number of complications-ie, presence of DKD±DR±DSPN) using linear regression models. Subsequently, the same metabolites were assessed with progression to endpoints: soft microvascular events (progression in albuminuria grade, ≥30% estimated glomerular filtration rate (eGFR) decline, or any progression in DR grade), hard microvascular events (progression to proliferative DR, chronic kidney failure, or ≥40% eGFR decline), and hard microvascular or macrovascular events (hard microvascular events, cardiovascular events (myocardial infarction, stroke, or arterial interventions), or cardiovascular mortality), using Cox models. All models were adjusted for sex, baseline age, diabetes duration, systolic blood pressure, HbA1c, body mass index, total cholesterol, smoking, and statin treatment. RESULTS: The full cohort investigated consisted of 487 participants. Mean (SD) follow-up was 4.8 (2.9, 5.7) years. Baseline biothesiometry was available in 202 participants, comprising the cross-sectional cohort. Eight metabolites were significantly associated with presence of DR, DKD, and DSPN, and six with additive microvascular burden severity. In the full cohort longitudinal analysis, higher levels of 3,4-dihydroxybutanoic acid (DHBA), 2,4-DHBA, ribonic acid, glycine, and ribitol were associated with development of events in both crude and adjusted models. Adding 3,4-DHBA, ribonic acid, and glycine to a traditional risk factor model improved the discrimination of hard microvascular events. CONCLUSIONS: While prospective studies directly assessing the predictive ability of these markers are needed, our results strengthen the role of clinical metabolomics in relation to risk assessment of diabetic complications in chronic type 1 diabetes.

Endothelial glycocalyx and cardio-renal risk factors in type 1 diabetes.

BACKGROUND: Glycocalyx lines the inner surface of the capillary endothelium. Capillaroscopy enables visualization of the sublingual capillaries and measurement of the Perfused Boundary Region (PBR) as an estimate of the glycocalyx. Novel software enables assessment of the PBR estimated at a fixed high flow level (PBR-hf) and an overall microvascular assessment by the MicroVascular Health Score (MVHS). Damaged glycocalyx may represent microvascular damage in diabetes and assessment of its dimension might improve early cardio-renal risk stratification. AIM: To assess the associations between PBR, PBR-hf and MVHS and cardio-renal risk factors in persons with type 1 diabetes (T1D); and to compare these dimensions in persons with T1D and controls. METHODS: Cross-sectional study including 161 persons with T1D stratified according to level of albuminuria and 50 healthy controls. The PBR, PBR-hf and MVHS were assessed by the GlycoCheck device (valid measurements were available in 136 (84.5%) with T1D and in all the controls). Higher PBR and PBR-hf indicate smaller glycocalyx width. Lower MVHS represents a worse microvascular health. RESULTS: There were no associations between PBR, PBR-hf or MVHS and the cardio-renal risk factors in persons with T1D, except for higher PBR-hf and lower MVHS in females (p = 0.01 for both). There was no difference in PBR, PBR-hf or MVHS in persons with normo-, micro- or macroalbuminuria. The PBR was higher (2.20±0.30 vs. 2.03±0.18µm; p<0.001) and MVHS lower (3.15±1.25 vs. 3.53±0.86µm; p = 0.02) in persons with T1D compared to controls (p≤0.02). After adjustment for cardio-renal risk factors the difference in PBR remained significant (p = 0.001). CONCLUSIONS: The endothelial glycocalyx dimension was impaired in persons with T1D compared to controls. We found no association between the endothelial glycocalyx dimension and the level of albuminuria or cardio-renal risk factors among persons with T1D. The use of the GlycoCheck device in T1D may not contribute to cardio-renal risk stratification.

Faecal biomarkers in type 1 diabetes with and without diabetic nephropathy.

Gastrointestinal dysbiosis is common among persons with type 1 diabetes (T1D), but its potential impact on diabetic nephropathy (DN) remains obscure. We examined whether faecal biomarkers, previously associated with low-grade gastrointestinal inflammation, differ between healthy controls and T1D subjects with and without DN. Faecal samples were analyzed for levels of calprotectin, intestinal alkaline phosphatase (IAP), short-chain fatty acids (SCFA) and immunoglobulins in subjects with T1D (n = 159) and healthy controls (NDC; n = 50). The subjects with T1D were stratified based on albuminuria: normoalbuminuria (< 30 mg/g; n = 49), microalbuminuria (30-299 mg/g; n = 50) and macroalbuminuria (≥ 300 mg/g; n = 60). aecal calprotectin, IAP and immunoglobulin levels did not differ between the T1D albuminuria groups. However, when subjects were stratified based on faecal calprotectin cut-off level (50 µg/g), macroalbuminuric T1D subjects exceeded the threshold more frequently than NDC (p = 0.02). Concentrations of faecal propionate and butyrate were lower in T1D subjects compared with NDC (p = 0.04 and p = 0.03, respectively). Among T1D subjects, levels of branched SCFA (BCFA) correlated positively with current albuminuria level (isobutyrate, p = 0.03; isovalerate, p = 0.005). In our study cohort, fatty acid metabolism seemed to be altered among T1D subjects and those with albuminuria compared to NDC. This may reflect gastrointestinal imbalances associated with T1D and renal complications.

Plasma trimethylamine N-oxide and its metabolic precursors and risk of mortality, cardiovascular and renal disease in individuals with type 2-diabetes and albuminuria.

AIMS: The trimethylamine N-oxide (TMAO) pathway is related to intestinal microbiota and has been associated to risk of cardiovascular disease (CVD). We investigated associations between four plasma metabolites in the TMAO pathway and risk of all-cause mortality, CVD and deterioration in renal function in individuals with type 2-diabetes (T2D) and albuminuria. MATERIALS AND METHODS: Plasma concentrations of TMAO, choline, carnitine, and betaine were measured by liquid chromatography-tandem mass spectrometry at baseline in 311 individuals with T2D and albuminuria. Information on all-cause mortality and fatal/non-fatal CVD during follow-up was obtained from registries. The association of each metabolite, and a weighted sum score of all four metabolites, with the endpoints were examined. Serum creatinine was measured at follow-up visits and the renal endpoint was defined as eGFR-decline of ≥30%. Associations were analysed using proportional hazards models adjusted for traditional risk factors. RESULTS: Baseline mean(SD) age was 57.2(8.2) years and 75% were males. Follow-up was up to 21.9 years (median (IQR) follow-up 6.8 (6.1-15.5) years for mortality and 6.5 (5.5-8.1) years for CVD events). The individual metabolites and the weighted sum score were not associated with all-cause mortality (n = 106) or CVD (n = 116) (adjusted p≥0.09). Higher choline, carnitine and the weighted sum score of the four metabolites were associated with higher risk of decline in eGFR (n = 106) (adjusted p = 0.001, p = 0.03 and p<0.001, respectively). CONCLUSIONS: In individuals with T2D and albuminuria, higher choline, carnitine and a weighted sum of four metabolites from the TMAO pathway were risk markers for deterioration in renal function during long-term follow-up. Metabolites from the TMAO pathway were not independently related to risk of all-cause mortality or CVD.

Visit-to-visit variability of clinical risk markers in relation to long-term complications in type 1 diabetes.

BACKGROUND: Clinical characteristics such as HbA1c , systolic blood pressure (SBP), albuminuria and estimated glomerular filtration rate (eGFR) are important when treating type 1 diabetes. We investigated the variability in these measures as risk markers for micro- and macrovascular complications. METHODS: This prospective study included 1062 individuals with type 1 diabetes. Visit-to-visit variability of HbA1c , SBP, albuminuria and eGFR was calculated as the SD of the residuals in individual linear regression models using all available measures in a specified period of 3 years (VV). Endpoints included were as follows: cardiovascular events (CVE) defined as myocardial infarction, non-fatal stroke, or coronary or peripheral arterial intervention; end-stage kidney disease (ESKD) defined as eGFR <15 ml/min/1.73 m2 , chronic dialysis or kidney transplantation; eGFR decline ≥30%; and mortality. Adjustment included age, sex, cholesterol, HbA1c , SBP, body mass index, smoking, albuminuria, eGFR, and mean, intercept, slope of respective exposure variables and regression models. RESULTS: SBP VV was significantly associated with CVE (adjusted hazard ratio per 50% increase, (CI 95%); p: 1.21 [1.05-1.39]; p = 0.008), ESKD (1.51 [1.16-1.96]; p = 0.002) and mortality (1.25 [1.09-1.44]; p = 0.002). HbA1c VV was significantly associated with mortality (1.51 [1.30-1.75]; p < 0.001); albuminuria VV with eGFR decline (1.14 [1.08-1.20]; p = 0.024) and ESKD (1.14 [1.02-1.27]; p < 0.001), but neither CVE nor mortality. Adjusted eGFR VV was not associated with endpoints. CONCLUSION: In type 1 diabetes, higher variability of basic clinical risk markers adds important risk stratification information for the development of micro- and macrovascular complications.

Assessment of the sublingual microcirculation with the GlycoCheck system: Reproducibility and examination conditions.

BACKGROUND: The glycocalyx is an extracellular layer lining the lumen of the vascular endothelium, protecting the endothelium from shear stress and atherosclerosis and contributes to coagulation, immune response and microvascular perfusion. The GlycoCheck system estimates glycocalyx' thickness in vessels under the tongue from perfused boundary region (PBR) and microvascular perfusion (red blood cell (RBC) filling) via a camera and dedicated software. OBJECTIVES: Evaluating reproducibility and influence of examination conditions on measurements with the GlycoCheck system. METHODS: Open, randomised, controlled study including 42 healthy smokers investigating day-to-day, side-of-tongue, inter-investigator variance, intraclass-correlation (ICC) and influence of examination conditions at intervals from 0-180 minutes on PBR and RBC filling. RESULTS: Mean (SD) age was 24.9 (6.1) years, 52% were male. There was no significant intra- or inter-investigator variation for PBR or RBC filling nor for PBR for side-of-tongue. A small day-to-day variance was found for PBR (0.012µm, p = 0.007) and RBC filling (0.003%, p = 0.005) and side-of-tongue, RBC filling (0.025%, p = 0.009). ICC was modest but highly improved by increasing measurements. Small significant influence of cigarette smoking (from 40-180 minutes), high calorie meal intake and coffee consumption was found. The latter two peaking immediately and tapering off but remained significant up to 180 minutes, highest PBR changes for the three being 0.042µm (p<0.05), 0.183µm (p<0.001) and 0.160µm (p<0.05) respectively. CONCLUSIONS: Measurements with the GlycoCheck system have a moderate reproducibility, but highly increases with multiple measurements and a small day-to-day variability. Smoking, meal and coffee intake had effects up to 180 minutes, abstinence is recommended at least 180 minutes before GlycoCheck measurements. Future studies should standardise conditions during measurements.

Carotid-Femoral Pulse Wave Velocity as a Risk Marker for Development of Complications in Type 1 Diabetes Mellitus.

Background The value of carotid-femoral pulse wave velocity (cfPWV) as risk factor for development of complications in type 1 diabetes mellitus remains to be determined. We investigated associations between cfPWV and renal outcomes, cardiovascular events, and all-cause mortality in people with type 1 diabetes mellitus. Methods and Results cfPWV was measured with SphygmoCor in 633 people with type 1 diabetes mellitus. Median (interquartile range) follow-up was 6.2 (5.8-6.7) years. End points included progression in albuminuria group, decline in estimated glomerular filtration rate (eGFR) ≥30%, end-stage kidney disease, cardiovascular event, mortality, and a composite renal end point. Hazard ratios (HRs) were calculated per 1-SD increase in cfPWV. Adjustments included age, sex, hemoglobin A1c, mean arterial pressure, body mass index, low-density lipoprotein cholesterol, smoking, urine albumin excretion rate, and eGFR. The cohort included 45% women, mean (SD) age was 54 (13) years, mean (SD) eGFR was 83.2 (27.9) mL/min per 1.73 m2, and mean (SD) cfPWV was 10.4 (3.3) m/s. Median (interquartile range) albumin excretion rate was 17 (17-63) mg/24 h. After adjustment, higher cfPWV was associated with increased hazard of progression in albuminuria (HR, 1.59; 95% CI, 1.10-2.32); decline in eGFR ≥30% (HR, 1.38; 95% CI, 1.06-1.79); cardiovascular event (HR, 1.31; 95% CI, 1.01-1.70); mortality (HR, 1.36; 95% CI, 1.00-1.85); and the composite renal end point (HR, 1.30; 95% CI, 1.04-1.63), but not with end-stage kidney disease (HR, 1.18; 95% CI, 0.62-2.26). Higher cfPWV was associated with steeper yearly increase in albumin excretion and steeper yearly decline in eGFR after adjustment (P=0.002 and P=0.01, respectively). Conclusions cfPWV was associated with increased hazard of renal outcomes, cardiovascular event, and mortality. cfPWV may be suited for risk stratification in type 1 diabetes mellitus.

Metabolomic Assessment Reveals Alteration in Polyols and Branched Chain Amino Acids Associated With Present and Future Renal Impairment in a Discovery Cohort of 637 Persons With Type 1 Diabetes.

Background: Improved understanding of the pathophysiology causing diabetic kidney disease (DKD) is imperative. The aim of this study was to uncover associations between serum metabolites and renal outcomes. Methods: Non-targeted serum metabolomics analyses were performed in samples from 637 persons with type 1 diabetes using two-dimensional gas chromatography coupled to time-of-flight mass-spectrometry. Longitudinal data at follow-up (median 5.5 years) on renal events were obtained from national Danish health registries. A composite renal endpoint (n = 123) consisted of estimated glomerular filtration rate (eGFR) decline from baseline (≥30%), progression to end-stage renal disease and all-cause mortality. Metabolites with significant associations (p < 0.05) in any of the cross-sectional analyses with eGFR and albuminuria were analyzed for specific and composite endpoints. Adjustments included traditional cardiovascular risk factors and correction for multiple testing. Results: A data-driven partial correlation analysis revealed a dense fabric of co-regulated metabolites and clinical variables dominated by eGFR. Ribonic acid and myo-inositol were inversely associated with eGFR, positively associated with macroalbuminuria (p < 0.02) and longitudinally associated with higher risk of eGFR decline ≥30% (HR 2.2-2.7, CI [1.3-4.3], p < 0.001). Ribonic acid was associated with a combined renal endpoint (HR 1.8, CI [1.3-2.3], p = 0.001). The hydroxy butyrate 3,4-dihydroxybutanoic acid was cross-sectionally associated with micro- and macroalbuminuria, urinary albumin excretion rate and inversely associated with eGFR (p < 0.04) while branched chain amino acids were associated with eGFR and lower risk of the combined renal endpoint (p < 0.02). Conclusions: Alterations in serum metabolites, particularly polyols and amino acids, were associated with renal endpoints in type 1 diabetes highlighting molecular pathways associated with progression of kidney disease. External validation is needed to further assess their roles and potentials as future therapeutic targets.

Markers of Collagen Formation and Degradation Reflect Renal Function and Predict Adverse Outcomes in Patients With Type 1 Diabetes.

OBJECTIVE: Patients with type 1 diabetes (T1D) have a higher risk of developing chronic kidney disease, cardiovascular events (CVEs), and mortality than the general population. We hypothesized that two previously published biomarkers, namely PRO-C6, a biomarker of collagen type VI formation, and C3M, a biomarker of collagen type III degradation, may be associated with impaired renal function and have prognostic value for adverse renal, CVE, and mortality in patients with T1D. RESEARCH DESIGN AND METHODS: PRO-C6 and C3M in serum (sPRO-C6, sC3M) and urine (uPRO-C6, uC3M) were measured by ELISA in 663 patients with T1D ranging from normoalbuminuric to macroalbuminuric. Association of the biomarkers with mortality, CVEs, heart failure, decline in estimated glomerular filtration rate (eGFR) ≥30%, and end-stage renal disease (ESRD) were tested in Cox proportional hazards models after log2 transformation and adjusted for relevant clinical characteristics. Hazard ratios (HRs) were reported per doubling of biomarker levels. RESULTS: High levels of sPRO-C6 were independently associated with a higher risk of all-cause mortality (HR 2.26 [95% CI 1.31-3.87], P < 0.0031). There was an association with higher risk of CVEs (n = 94) and heart failure (n = 28) but not after adjustment (P ≥ 0.58). In relation to renal outcomes, adjusted sPRO-C6 was associated with a higher risk of eGFR decline ≥30% in T1D, with eGFR >45 and >30 mL/min/1.73 m2, and with a higher risk of ESRD (all P ≤ 0.03). Higher uPRO-C6 was associated with a lower risk of decline in eGFR. CONCLUSIONS: In patients with T1D, higher sPRO-C6 was an independent predictor of both decline in eGFR and development of ESRD and of all-cause mortality. Higher uPRO-C6 was also associated with a lower risk of decline in eGFR.

Uric Acid Is an Independent Risk Factor for Decline in Kidney Function, Cardiovascular Events, and Mortality in Patients With Type 1 Diabetes.

OBJECTIVE: Previous studies have provided inconclusive results on the role of uric acid (UA) in risk prediction. Here we aimed to improve the power and precision of the predictive value of UA for the risk of decline in kidney function, cardiovascular events (CVEs), and mortality in patients with type 1 diabetes (T1D). RESEARCH DESIGN AND METHODS: Plasma UA was measured in 670 patients with T1D and various degrees of albuminuria, ranging from normoalbuminuria to macroalbuminuria. Associations of UA with an estimated glomerular filtration rate (eGFR) decline of ≥30%, CVEs, and mortality were analyzed. The median follow-up time was 5.3 years [interquartile range (IQR) 2.7-6.2 years] for a decline in eGFR of ≥30%, 5.8 years (2.5-6.4 years) for progression in albuminuria status, 5.1 years (4.7-5.6 years) for CVE, and 6.2 years (5.8-6.7 years) for mortality. Both univariable and multivariable associations of UA with relevant outcomes and variables were reported. Hazard ratios (HRs) were calculated per doubling of the UA level. RESULTS: A doubling in UA level was associated with a higher risk of decline in eGFR of ≥30% (n = 89) (HR 3.18 [IQR 1.71-5.93]; P < 0.001), CVE (n = 94) (HR 2.25 [IQR 1.20-4.21]; P = 0.011), and mortality (n = 58) (HR 2.58 [IQR 1.12-5.90]; P = 0.025) in adjusted analyses. Adding UA to the adjusted model including conventional risk factors improved the relative integrated discrimination index by 12.6% for a decline in eGFR of ≥30% (P < 0.001), 6.5% for CVE (P = 0.010), and 11.8% (P = 0.003) for mortality. A doubling in UA level was also associated with a steeper decline in eGFR (P < 0.0026) and a steeper increase in urine albumin-to-creatinine ratio (P < 0.0027) in adjusted analysis. CONCLUSIONS: In individuals with T1D, a higher UA level is associated with a higher risk of decline in kidney function, CVE, and mortality, independently of other risk factors. Our results suggest that UA has a promising role in risk stratification among individuals with T1D.

Trimethylamine N-oxide (TMAO) as a New Potential Therapeutic Target for Insulin Resistance and Cancer.

BACKGROUND: The intake of animal products in food has been associated with both the development of insulin resistance and gastrointestinal cancers (GIC). Through the digestion of animal protein and other constituents of animal products, the commensal bacteria in the gut (the gut microbiota) forms metabolites that can contribute to the development of both insulin resistance and cancer. Trimethylamine-N-Oxide (TMAO) is such a molecule and has recently drawn a lot of attention as it may be a risk factor for - and a link between - the gut microbiota and cardiovascular and renal disease. Further, TMAO is anticipated to have significance as a biomarker of - or even an independent risk factor for - other undesirable conditions, including insulin resistance and GIC. TMAO originates from a precursor, trimethylamine (TMA) that is a metabolite of various precursors; mainly choline and carnitine from ingested foods. METHODS: We review the literature on TMAO as a shared risk factor and/or pathway between insulin resistance and GIC risk and take the reader through the literature of interventions that could reduce formation of TMAO and thereby the risk of insulin resistance and GIC. The purpose of the work is to generate a hypothesis to be tested in preclinical and clinical studies. RESULTS: TMAO seems to be associated with both insulin resistance and GIC risk and also with atherosclerotic cardiovascular disease. One shared pathway is the formation of N-Nitroso compounds, a group of metabolites that can cause DNA-damage and epigenetic changes. Levels of TMAO can be reduced by limiting the dietary intake of certain foods, most importantly animal products. Further, certain drugs, namely Meldonium and 3,3-dimethyl- 1-butanol, may inhibit the formation of TMAO by inhibiting bacterial enzymes. CONCLUSIONS: The TMAO pathway and its metabolites are possibly involved in the development of two major health problems: insulin resistance and cancer. Within these pathways novel therapeutic targets may be identified. Further research is needed in order to verify existing or develop new pharmacological agents that modify these pathways and reduce the risk of insulin resistance and GIC.

Psoriasis and new-onset diabetes: a Danish nationwide cohort study.

OBJECTIVE: Psoriasis is associated with increased risk of cardiovascular events and increased prevalence of cardiovascular risk factors. Diabetes mellitus (DM) is a major contributor to cardiovascular morbidity and mortality that may be associated with psoriasis, but conflicting results have been presented and nationwide data on the risk of new-onset DM in patients with psoriasis have not been reported. RESEARCH DESIGN AND METHODS: The study comprised a Danish population ≥ 10 years of age on 1 January 1997 who were followed until new-onset DM, death, or 31 December 2009. Information on comorbidity, concomitant medication, and socioeconomic status was linked on an individual level. The primary study end point was DM requiring pharmacotherapy. Incidence rates for the development of DM events per 1,000 observational years were calculated and adjusted. Incidence rate ratios (IRRs) were estimated by Poisson regression. RESULTS: A total of 4,614,807 subjects were eligible for analysis, with a maximum follow-up of 13 years. In the study period, 52,613 patients with psoriasis, including 6,784 patients with severe psoriasis, were identified. The overall incidence rates for new-onset DM were 3.67 (CI 3.65-3.69), 6.93 (6.63-7.25), and 9.65 (8.68-10.73) for the reference population, mild psoriasis, and severe psoriasis, respectively. Compared with the reference population, the IRR of new-onset DM was increased in all patients with psoriasis, i.e., IRR 1.49 (CI 1.43-1.56) and 2.13 (1.91-2.37) for those with mild and severe psoriasis. CONCLUSIONS: In this nationwide cohort, psoriasis was associated with increased incidence rates of new-onset DM. The association remained statistically significant after adjustment for confounding factors.

[The effect of aldosterone A on renal potassium excretion]. / Aldosterons effekt på nyrernes kaliumbalance.

Recent studies have shown expression of the following regulatory WNK kinases in the kidney: the full-length WNK1 (L-WNK1), the shorter kidney specific WNK1 transcript (KS-WNK1), formed by alternative splicing, and WNK4. Aldosterone activates expression of KS-WNK1 and inhibits WNK4 via SGK1 - both leading to stimulation of ENaC and activation of ROMK, and increased potassium excretion. Thus, further characterization of the WNK system may lead to elucidation of the dual anti-natriuretic and kaliuretic effects of aldosterone, in situations where only activation of one of these effects is needed.