Carol Forsblom

Diabetic nephropathy is associated with low-grade inflammation in Type 1 diabetic patients.

Aims/hypothesisIncreased concentrations of C-reactive protein and interleukin-6, a finding suggestive of the presence of inflammation, have been observed in Type 2 diabetes. In such patients, C-reactive protein was predictive of diabetic nephropathy. Studies on low-grade inflammatory markers and nephropathy in Type 1 diabetic patients have shown conflicting results. Therefore we studied whether low-grade inflammation is associated with diabetic nephropathy in Type 1 diabetic patients.MethodsWe divided 194 Type 1 diabetic patients into three groups from the Finnish Diabetic Nephropathy Study based upon their albumin excretion rate. Patients with normoalbuminuria (n=67) had no antihypertensive medication or signs of cardiovascular disease, while patients with microalbuminuria (n=64) or macroalbuminuria (n=63) were all treated with an angiotensin-converting enzyme inhibitor, a drug that could attenuate low-grade inflammation. As a measure of insulin sensitivity we used estimated glucose disposal rate. C-reactive protein was measured by radioimmunoassay and interleukin-6 by high sensitivity enzyme immunoassay.ResultsC-reactive protein was higher in micro- and macroalbuminuric patients compared to normoalbuminuric patients (normoalbuminuria 2.0±1.7, microalbuminuria 2.6±1.7, macroalbuminuria 2.9±2.5xa0mg/l; p=0.016), while interleukin-6 increased in parallel with the severity of the renal disease (1.9±1.5, 2.9±3.3, 3.6±3.1xa0ng/l; p<0.0001). In multiple regression analysis albumin excretion rate was the only variable independently associated with C-reactive protein (p=0.03), whereas albumin excretion rate (p=0.0003), HDL-cholesterol (p=0.0135) and duration of diabetes (p=0.0176) were independently associated with interleukin-6.Conclusions/interpretationLow-grade inflammatory markers are associated with diabetic nephropathy in Type 1 diabetic patients. The predictive value needs to be assessed.

Metabolomics Reveals Signature of Mitochondrial Dysfunction in Diabetic Kidney Disease

Diabetic kidney disease is the leading cause of ESRD, but few biomarkers of diabetic kidney disease are available. This study used gas chromatography-mass spectrometry to quantify 94 urine metabolites in screening and validation cohorts of patients with diabetes mellitus (DM) and CKD(DM+CKD), in patients with DM without CKD (DM-CKD), and in healthy controls. Compared with levels in healthy controls, 13 metabolites were significantly reduced in the DM+CKD cohorts (P≤0.001), and 12 of the 13 remained significant when compared with the DM-CKD cohort. Many of the differentially expressed metabolites were water-soluble organic anions. Notably, organic anion transporter-1 (OAT1) knockout mice expressed a similar pattern of reduced levels of urinary organic acids, and human kidney tissue from patients with diabetic nephropathy demonstrated lower gene expression of OAT1 and OAT3. Analysis of bioinformatics data indicated that 12 of the 13 differentially expressed metabolites are linked to mitochondrial metabolism and suggested global suppression of mitochondrial activity in diabetic kidney disease. Supporting this analysis, human diabetic kidney sections expressed less mitochondrial protein, urine exosomes from patients with diabetes and CKD had less mitochondrial DNA, and kidney tissues from patients with diabetic kidney disease had lower gene expression of PGC1α (a master regulator of mitochondrial biogenesis). We conclude that urine metabolomics is a reliable source for biomarkers of diabetic complications, and our data suggest that renal organic ion transport and mitochondrial function are dysregulated in diabetic kidney disease.

New susceptibility loci associated with kidney disease in Type 1 diabetes

Diabetic kidney disease, or diabetic nephropathy (DN), is a major complication of diabetes and the leading cause of end-stage renal disease (ESRD) that requires dialysis treatment or kidney transplantation. In addition to the decrease in the quality of life, DN accounts for a large proportion of the excess mortality associated with type 1 diabetes (T1D). Whereas the degree of glycemia plays a pivotal role in DN, a subset of individuals with poorly controlled T1D do not develop DN. Furthermore, strong familial aggregation supports genetic susceptibility to DN. However, the genes and the molecular mechanisms behind the disease remain poorly understood, and current therapeutic strategies rarely result in reversal of DN. In the GEnetics of Nephropathy: an International Effort (GENIE) consortium, we have undertaken a meta-analysis of genome-wide association studies (GWAS) of T1D DN comprising ∼2.4 million single nucleotide polymorphisms (SNPs) imputed in 6,691 individuals. After additional genotyping of 41 top ranked SNPs representing 24 independent signals in 5,873 individuals, combined meta-analysis revealed association of two SNPs with ESRD: rs7583877 in the AFF3 gene (Pu200a=u200a1.2×10−8) and an intergenic SNP on chromosome 15q26 between the genes RGMA and MCTP2, rs12437854 (Pu200a=u200a2.0×10−9). Functional data suggest that AFF3 influences renal tubule fibrosis via the transforming growth factor-beta (TGF-β1) pathway. The strongest association with DN as a primary phenotype was seen for an intronic SNP in the ERBB4 gene (rs7588550, Pu200a=u200a2.1×10−7), a gene with type 2 diabetes DN differential expression and in the same intron as a variant with cis-eQTL expression of ERBB4. All these detected associations represent new signals in the pathogenesis of DN.

Discovery of early-stage biomarkers for diabetic kidney disease using ms-based metabolomics (FinnDiane study)

Diabetic kidney disease (DKD) is a devastating complication that affects an estimated third of patients with type 1 diabetes mellitus (DM). There is no cure once the disease is diagnosed, but early treatment at a sub-clinical stage can prevent or at least halt the progression. DKD is clinically diagnosed as abnormally high urinary albumin excretion rate (AER). We hypothesize that subtle changes in the urine metabolome precede the clinically significant rise in AER. To test this, 52 type 1 diabetic patients were recruited by the FinnDiane study that had normal AER (normoalbuminuric). After an average of 5.5xa0years of follow-up half of the subjects (26) progressed from normal AER to microalbuminuria or DKD (macroalbuminuria), the other half remained normoalbuminuric. The objective of this study is to discover urinary biomarkers that differentiate the progressive form of albuminuria from non-progressive form of albuminuria in humans. Metabolite profiles of baseline 24xa0h urine samples were obtained by gas chromatography–mass spectrometry (GC–MS) and liquid chromatography–mass spectrometry (LC–MS) to detect potential early indicators of pathological changes. Multivariate logistic regression modeling of the metabolomics data resulted in a profile of metabolites that separated those patients that progressed from normoalbuminuric AER to microalbuminuric AER from those patients that maintained normoalbuminuric AER with an accuracy of 75% and a precision of 73%. As this data and samples are from an actual patient population and as such, gathered within a less controlled environment it is striking to see that within this profile a number of metabolites (identified as early indicators) have been associated with DKD already in literature, but also that new candidate biomarkers were found. The discriminating metabolites included acyl-carnitines, acyl-glycines and metabolites related to tryptophan metabolism. We found candidate biomarkers that were univariately significant different. This study demonstrates the potential of multivariate data analysis and metabolomics in the field of diabetic complications, and suggests several metabolic pathways relevant for further biological studies.

Association Testing of Previously Reported Variants in a Large Case–Control Meta-Analysis of Diabetic Nephropathy

We formed the GEnetics of Nephropathy–an International Effort (GENIE) consortium to examine previously reported genetic associations with diabetic nephropathy (DN) in type 1 diabetes. GENIE consists of 6,366 similarly ascertained participants of European ancestry with type 1 diabetes, with and without DN, from the All Ireland-Warren 3-Genetics of Kidneys in Diabetes U.K. and Republic of Ireland (U.K.-R.O.I.) collection and the Finnish Diabetic Nephropathy Study (FinnDiane), combined with reanalyzed data from the Genetics of Kidneys in Diabetes U.S. Study (U.S. GoKinD). We found little evidence for the association of the EPO promoter polymorphism, rs161740, with the combined phenotype of proliferative retinopathy and end-stage renal disease in U.K.-R.O.I. (odds ratio [OR] 1.14, P = 0.19) or FinnDiane (OR 1.06, P = 0.60). However, a fixed-effects meta-analysis that included the previously reported cohorts retained a genome-wide significant association with that phenotype (OR 1.31, P = 2 × 10−9). An expanded investigation of the ELMO1 locus and genetic regions reported to be associated with DN in the U.S. GoKinD yielded only nominal statistical significance for these loci. Finally, top candidates identified in a recent meta-analysis failed to reach genome-wide significance. In conclusion, we were unable to replicate most of the previously reported genetic associations for DN, and significance for the EPO promoter association was attenuated.

Serum adiponectin concentration is a positive predictor of all‐cause and cardiovascular mortality in type 1 diabetes

Abstract.u2002 Forsblom C, Thomas MC, Moran J, Saraheimo M, Thorn L, Wadén J, Gordin D, Frystyk J, Flyvbjerg A, Groop P‐H, on behalf of the FinnDiane Study Group (Folkhälsan Institute of Genetics, Helsinki; Department of Medicine Helsinki University Central Hospital, Helsinki, Finland; The Baker IDI Heart and Diabetes Institute, Melbourne, Vic.; The Queen Elizabeth Hospital, Woodville, SA, Australia; and Aarhus University Hospital, Aarhus C., Denmark). Serum adiponectin concentration is a positive predictor of all‐cause and cardiovascular mortality in type 1 diabetes. J Intern Med 2011; 270: 346–355.

Progression to microalbuminuria in type 1 diabetes: development and validation of a prediction rule

Aims/hypothesisMicroalbuminuria is common in type 1 diabetes and is associated with an increased risk of renal and cardiovascular disease. We aimed to develop and validate a clinical prediction rule that estimates the absolute risk of microalbuminuria.MethodsData from the European Diabetes Prospective Complications Study (nu2009=u20091115) were used to develop the prediction rule (development set). Multivariable logistic regression analysis was used to assess the association between potential predictors and progression to microalbuminuria within 7xa0years. The performance of the prediction rule was assessed with calibration and discrimination (concordance statistic [c-statistic]) measures. The rule was validated in three other diabetes studies (Pittsburgh Epidemiology of Diabetes Complications [EDC] study, Finnish Diabetic Nephropathy [FinnDiane] study and Coronary Artery Calcification in Type 1 Diabetes [CACTI] study).ResultsOf patients in the development set, 13% were microalbuminuric after 7xa0years. Glycosylated haemoglobin, AER, WHR, BMI and ever smoking were found to be the most important predictors. A high-risk group (nu2009=u200987 [8%]) was identified with a risk of progression to microalbuminuria of 32%. Predictions showed reasonable discriminative ability, with c-statistic of 0.71. The rule showed good calibration and discrimination in EDC, FinnDiane and CACTI (c-statistic 0.71, 0.79 and 0.79, respectively).Conclusions/interpretationWe developed and validated a clinical prediction rule that uses relatively easily obtainable patient characteristics to predict microalbuminuria in patients with type 1 diabetes. This rule can help clinicians to decide on more frequent check-ups for patients at high risk of microalbuminuria in order to prevent long-term chronic complications.

Circulating ACE2 activity is increased in patients with type 1 diabetes and vascular complications.

Objective: Angiotensin-converting enzyme 2 (ACE2) is a homolog of ACE that counterbalances the actions of angiotensin (AT)II and promotes vasodilatation. Circulating ACE2 activity is increased in diabetes in experimental models. The role of ACE2 in human pathophysiology is unknown. We examined whether ACE2 activity is altered in patients with type 1 diabetes (T1D), with and without diabetic nephropathy. Methods: Quantitative ACE2 activity in serum was measured by a fluorometric assay in 859 patients with T1D in the Finnish Diabetic Nephropathy (FinnDiane) study and in 204 healthy controls. Pulse-wave analysis with augmentation index (AIx) measurement was performed in 319 patients with T1D and 114 controls. Results: ACE2 activity was increased in men with T1D and microalbuminuria (30.2u200a±u200a1.5u200angE/ml) when compared to patients without albuminuria (27.0u200a±u200a0.5u200angE/ml, Pu200a<u200a0.05) or controls (25.6u200a±u200a0.8u200angE/ml, Pu200a<u200a0.05). ACE2 activity was increased in male and female patients who were on ACE inhibitor (ACEi) treatment, also independently of albuminuria. Male and female patients with coronary heart disease (CHD) had significantly increased ACE2 activity (35.5u200a±u200a2.5 vs. 27.0u200a±u200a0.5u200angE/ml, Pu200a<u200a0.001 among male T1D patients vs. male controls). ACE2 activity correlated positively with systolic blood pressure (rsu200a=u200a0.175, Pu200a<u200a0.001), AIx (rsu200a=u200a0.191, Pu200a=u200a0.010) and diabetes duration (rsu200a=u200a0.198, Pu200a<u200a0.001), and negatively with estimated glomerular filtration rate (rsu200a=u200a−0.109, Pu200a=u200a0.016) among male T1D patients. Conclusions: ACE2 activity increases with increasing vascular tone and when the patient with T1D has microvascular or macrovascular disease, indicating that ACE2 may participate as a compensatory mechanism in the regulation of vascular and renal function in patients with T1D.

Health-related quality of life in patients with type 1diabetes—association with diabetic complications (the FinnDiane Study)

BACKGROUNDnThe daily treatment of type 1 diabetes with frequent monitoring of blood glucose levels and nuisance caused by insulin administration may affect patients health-related quality of life (HRQoL). Type 1 diabetes is further burdened with an increased risk of complications which may additionally reduce a patients HRQoL. We aimed to assess HRQoL and its association with diabetic complications in a large sample of patients with type 1 diabetes.nnnMETHODSnAltogether, 1070 patients with type 1 diabetes (46% men, mean age 46 +/- 12 years, diabetes duration 29 +/- 13 years) from the Finnish Diabetic Nephropathy Study (FinnDiane Study) participated in this cross-sectional study. Data on HRQoL were obtained from 1023 patients using the 15D instrument. When studying nephropathy, patients were divided into groups based on their albumin excretion rate. Laser-treated patients were classified as having proliferative retinopathy.nnnRESULTSnThe mean 15D score was 0.899 +/- 0.095 with no differences between men and women. HRQoL decreased with increasing age among patients with and without diabetic complications. In the Tobit regression model, macroalbuminuria (-0.036), dialysis (-0.082), renal transplant (-0.053), poor glycemic control (-0.006), ageing (-0.002) and longer diabetes duration (-0.001) explained the reduction in HRQoL. In a corresponding model, the presence of proliferative retinopathy did not have a significant negative influence on HRQoL. Of individual dimensions of the 15D instrument, nephropathy affected all but five dimensions, while retinopathy affected vision, mobility, eating and usual activities.nnnCONCLUSIONSnThe 15D scores decreased with increasing age. The presence of nephropathy, but not retinopathy, reduced the subjective HRQoL in patients with type 1 diabetes.

Urinary Liver-Type Fatty Acid-Binding Protein and Progression of Diabetic Nephropathy in Type 1 Diabetes

OBJECTIVE Diabetic nephropathy (DN) has mainly been considered a glomerular disease, although tubular dysfunction may also play a role. This study assessed the predictive value for progression of a tubular marker, urinary liver-type fatty acid–binding protein (L-FABP), at all stages of DN. RESEARCH DESIGN AND METHODS At baseline, 1,549 patients with type 1 diabetes had an albumin excretion rate (AER) within normal reference ranges, 334 had microalbuminuria, and 363 had macroalbuminuria. Patients were monitored for a median of 5.8 years (95% CI 5.7–5.9). In addition, 208 nondiabetic subjects were studied. L-FABP was measured by ELISA and normalized with urinary creatinine. Different Cox proportional hazard models for the progression at every stage of DN were used to evaluate the predictive value of L-FABP. The potential benefit of using L-FABP alone or together with AER was assessed by receiver operating characteristic curve analyses. RESULTS L-FABP was an independent predictor of progression at all stages of DN. As would be expected, receiver operating characteristic curves for the prediction of progression were significantly larger for AER than for L-FABP, except for patients with baseline macroalbuminuria, in whom the areas were similar. Adding L-FABP to AER in the models did not significantly improve risk prediction of progression in favor of the combination of L-FABP plus AER compared with AER alone. CONCLUSIONS L-FABP is an independent predictor of progression of DN irrespective of disease stage. L-FABP used alone or together with AER may not improve the risk prediction of DN progression in patients with type 1 diabetes, but further studies are needed in this regard.