GWAS meta-analysis followed by Mendelian randomization revealed potential control mechanisms for circulating α-Klotho levels.

The protein α-Klotho acts as transmembrane co-receptor for fibroblast growth factor 23 (FGF23) and is a key regulator of phosphate homeostasis. However, α-Klotho also exists in a circulating form, with pleiotropic, but incompletely understood functions and regulation. Therefore, we undertook a genome-wide association study (GWAS) meta-analysis followed by Mendelian randomization (MR) of circulating α-Klotho levels. Plasma α-Klotho levels were measured by enzyme-linked immunosorbent assay (ELISA) in the Ludwigshafen Risk and Cardiovascular Health and Avon Longitudinal Study of Parents and Children (mothers) cohorts, followed by a GWAS meta-analysis in 4376 individuals across the two cohorts. Six signals at five loci were associated with circulating α-Klotho levels at genome-wide significance (P < 5 × 10-8), namely ABO, KL, FGFR1, and two post-translational modification genes, B4GALNT3 and CHST9. Together, these loci explained >9% of the variation in circulating α-Klotho levels. MR analyses revealed no causal relationships between α-Klotho and renal function, FGF23-dependent factors such as vitamin D and phosphate levels, or bone mineral density. The screening for genetic correlations with other phenotypes followed by targeted MR suggested causal effects of liability of Crohn's disease risk [Inverse variance weighted (IVW) beta = 0.059 (95% confidence interval 0.026, 0.093)] and low-density lipoprotein cholesterol levels [-0.198 (-0.332, -0.063)] on α-Klotho. Our GWAS findings suggest that two enzymes involved in post-translational modification, B4GALNT3 and CHST9, contribute to genetic influences on α-Klotho levels, presumably by affecting protein turnover and stability. Subsequent evidence from MR analyses on α-Klotho levels suggest regulation by mechanisms besides phosphate-homeostasis and raise the possibility of cross-talk with FGF19- and FGF21-dependent pathways, respectively. Significance statement: α-Klotho as a transmembrane protein is well investigated along the endocrine FGF23-α-Klotho pathway. However, the role of the circulating form of α-Klotho, which is generated by cleavage of transmembrane α-Klotho, remains incompletely understood. Genetic analyses might help to elucidate novel regulatory and functional mechanisms. The identification of genetic factors related to circulating α-Klotho further enables MR to examine causal relationships with other factors. The findings from the first GWAS meta-analysis of circulating α-Klotho levels identified six genome-wide significant signals across five genes. Given the function of two of the genes identified, B4GALNT3 and CHST9, it is tempting to speculate that post-translational modification significantly contributes to genetic influences on α-Klotho levels, presumably by affecting protein turnover and stability.

Platelet and Monocyte Activity Markers and Mortality in Patients with End-Stage Renal Disease.

BACKGROUND: The association of platelet and monocyte activity markers with long-term mortality was assessed in hemodialysis (HD) patients. METHODS: In 41 HD patients (25 male, 16 female), surface expression of CD40L and CD62P on platelets, tissue factor (TF) binding on monocytes, and platelet-monocyte aggregates were measured by flow cytometry. Plasma levels of MCP-1, IL-6, TNFα, and soluble CD40L were analyzed by enzyme linked immunosorbent assay. Cox proportional hazard regression analyses and Kaplan-Meier curve were calculated. The predefined endpoint was all-cause mortality. RESULTS: The study follow-up was 11.54 years. Thirty-one patients (75.6%) died within the study period. Mean patient survival after study inclusion was 5.45 +/- 4.24 years. TF on monocytes above the median of the study population was significantly and independently associated with total mortality (HR (95% CI) 3.45 (1.32 - 9.07); p = 0.01). Cumulative mortality in patients with TF on monocytes above median was significantly higher compared to pa-tients with TF on monocytes below median value (log rank p < 0.01). Platelet-monocyte aggregates, the expression of CD40L and CD62P on platelets, and plasma levels of sCD40L, IL-6, MCP-1, and TNFα were not significantly correlated with mortality. CONCLUSIONS: The present study confirms a high mortality in ESRD patients. TF binding on monocytes was significantly correlated with increased mortality and may identify a subgroup of patients at higher risk.

Propeptide big-endothelin, N-terminal-pro brain natriuretic peptide and mortality. The Ludwigshafen risk and cardiovascular health (LURIC) study.

CONTEXT: The endothelin system (Big-ET-1) is a key regulator in cardiovascular (CV) disease and congestive heart failure (CHF). OBJECTIVES: We have examined the incremental value of Big-ET-1 in predicting total and CV mortality next to the well-established CV risk marker N-Terminal Pro-B-Type Natriuretic Peptide (NT-proBNP). METHODS: Big-ET-1 and NT-proBNP were determined in 2829 participants referred for coronary angiography (follow-up 9.9 years). RESULTS: Big-ET-1 is an independent predictor of total, CV mortality and death due to CHF. DISCUSSION: The conjunct use of Big-ET-1 and NT-proBNP improves the risk stratification of patients with intermediate to high risk of CV death and CHF. CONCLUSIONS: Big-ET-1improves risk stratification in patients referred for coronary angiography.

Efficacy of Empagliflozin in Patients With Heart Failure Across Kidney Risk Categories.

BACKGROUND: Empagliflozin reduces the risk of major heart failure outcomes in heart failure with reduced or preserved ejection fraction. OBJECTIVES: The goal of this study was to evaluate the effect of empagliflozin across the spectrum of chronic kidney disease in a pooled analysis of EMPEROR-Reduced and EMPEROR-Preserved (Empagliflozin Outcome Trial in Patients with Chronic Heart Failure with Reduced or Preserved Ejection Fraction, respectively). METHODS: A total of 9,718 patients were grouped into Kidney Disease Improving Global Outcomes (KDIGO) categories based on estimated glomerular filtration rate (eGFR) and urine albumin-to-creatinine ratio into low-, moderate-, high-, and very-high-risk categories, comprising 32.0%, 29.1%, 21.9%, and 17.0% of the participants, respectively. RESULTS: In the placebo arm, when compared with lower risk categories, patients at higher risk experienced a slower rate of decline in eGFR, but a higher risk of a composite kidney event. Empagliflozin reduced the risk of cardiovascular death or heart failure hospitalizations similarly in all KDIGO categories (HR: 0.81; 95% CI: 0.66-1.01 for low-; HR: 0.63; 95% CI: 0.52-0.76 for moderate-; HR: 0.82; 95% CI: 0.68-0.98 for high-; and HR: 0.84; 95% CI: 0.71-1.01 for very-high-risk groups; P trend = 0.30). Empagliflozin reduced the rate of decline in eGFR whether it was estimated by chronic slope, total slope, or unconfounded slope. When compared with the unconfounded slope, the magnitude of the effect on chronic slope was larger, and the effect on total slope was smaller. In EMPEROR-Reduced, patients at lowest risk experienced the largest effect of empagliflozin on eGFR slope; this pattern was not observed in EMPEROR-Preserved. CONCLUSIONS: The benefit of empagliflozin on major heart failure events was not influenced by KDIGO categories. The magnitude of the renal effects of the drug depended on the approach used to calculate eGFR slopes.

Lowering of Circulating Sclerostin May Increase Risk of Atherosclerosis and Its Risk Factors: Evidence From a Genome-Wide Association Meta-Analysis Followed by Mendelian Randomization.

OBJECTIVE: In this study, we aimed to establish the causal effects of lowering sclerostin, target of the antiosteoporosis drug romosozumab, on atherosclerosis and its risk factors. METHODS: A genome-wide association study meta-analysis was performed of circulating sclerostin levels in 33,961 European individuals. Mendelian randomization (MR) was used to predict the causal effects of sclerostin lowering on 15 atherosclerosis-related diseases and risk factors. RESULTS: We found that 18 conditionally independent variants were associated with circulating sclerostin. Of these, 1 cis signal in SOST and 3 trans signals in B4GALNT3, RIN3, and SERPINA1 regions showed directionally opposite signals for sclerostin levels and estimated bone mineral density. Variants with these 4 regions were selected as genetic instruments. MR using 5 correlated cis-SNPs suggested that lower sclerostin increased the risk of type 2 diabetes mellitus (DM) (odds ratio [OR] 1.32 [95% confidence interval (95% CI) 1.03-1.69]) and myocardial infarction (MI) (OR 1.35 [95% CI 1.01-1.79]); sclerostin lowering was also suggested to increase the extent of coronary artery calcification (CAC) (ß = 0.24 [95% CI 0.02-0.45]). MR using both cis and trans instruments suggested that lower sclerostin increased hypertension risk (OR 1.09 [95% CI 1.04-1.15]), but otherwise had attenuated effects. CONCLUSION: This study provides genetic evidence to suggest that lower levels of sclerostin may increase the risk of hypertension, type 2 DM, MI, and the extent of CAC. Taken together, these findings underscore the requirement for strategies to mitigate potential adverse effects of romosozumab treatment on atherosclerosis and its related risk factors.

Circulating Sclerostin Levels Are Positively Related to Coronary Artery Disease Severity and Related Risk Factors.

Romosozumab is a newly available treatment for osteoporosis acting by sclerostin inhibition. Its cardiovascular safety has been questioned after finding excess cardiovascular disease (CVD)-related events in a pivotal phase 3 trial. Previous studies of relationships between circulating sclerostin levels and CVD and associated risk factors have yielded conflicting findings, likely reflecting small numbers and selected patient groups. We aimed to characterize relationships between sclerostin and CVD and related risk factors in more detail by examining these in two large cohorts, Ludwigshafen Risk and Cardiovascular Health study (LURIC; 34% female, mean age 63.0 years) and Avon Longitudinal Study of Parents and Children study (ALSPAC) mothers (mean age 48.1 years). Together these provided 5069 participants with complete data. Relationships between sclerostin and CVD risk factors were meta-analyzed, adjusted for age, sex (LURIC), body mass index, smoking, social deprivation, and ethnicity (ALSPAC). Higher sclerostin levels were associated with higher risk of diabetes mellitus (DM) (odds ratio [OR] = 1.25; 95% confidence interval [CI] 1.12, 1.37), risk of elevated fasting glucose (OR 1.15; CI 1.04, 1.26), and triglyceride levels (ß 0.03; CI 0.00, 0.06). Conversely, higher sclerostin was associated with lower estimated glomerular filtration rate (eGFR) (ß -0.20; CI -0.38, -0.02), HDL cholesterol (ß -0.05; CI -0.10, -0.01), and apolipoprotein A-I (ß -0.05; CI -0.08, -0.02) (difference in mean SD per SD increase in sclerostin, with 95% CI). In LURIC, higher sclerostin was associated with an increased risk of death from cardiac disease during follow-up (hazard ratio [HR] = 1.13; 1.03, 1.23) and with severity of coronary artery disease on angiogram as reflected by Friesinger score (0.05; 0.01, 0.09). Associations with cardiac mortality and coronary artery severity were partially attenuated after adjustment for risk factors potentially related to sclerostin, namely LDL and HDL cholesterol, log triglycerides, DM, hypertension, eGFR, and apolipoprotein A-I. Contrary to trial evidence suggesting sclerostin inhibition leads to an increased risk of CVD, sclerostin levels appear to be positively associated with coronary artery disease severity and mortality, partly explained by a relationship between higher sclerostin levels and major CVD risk factors. © 2021 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).

Author Correction: Cardiovascular risk algorithms in primary care: Results from the DETECT study.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Renal function, N-terminal Pro-B-Type natriuretic peptide, propeptide big-endothelin and patients with heart failure and preserved ejection fraction.

Renal dysfunction may limit the clinical application of NT-proBNP in the diagnosis of heart failure. In general practice, where echocardiography is not readily available, a biomarker for the diagnosis of a heart failure with preserved ejection fraction (HFpEF) would be useful. Since cardiac diseases frequently coincide with renal disease, there is a high need of valid risk stratification methods in patients affected with both. We therefore examined NT-proBNP and another biomarker, Big-Endothelin-1, as a marker of HFpEF in patients with CKD. NT-proBNP and Big-ET-1 were determined in 439 patients with HFpEF in the Ludwigshafen Risk and Cardiovascular Health (LURIC) study. NT-proBNP plasma level has shown an exponential increase with declining GFR, while Big-ET-1 plasma level increased only in a moderate and linear fashion. In patients without CKD, a NT-proBNP cut-off point at 250 pg/mL was suitable for the discrimination between HFpEF and patients without HF. When the GFR was less than 60 mL/min/1.73m2, the NT-proBNP cut-off point should be raised to 750 pg/mL. At a cutoff point at 0.85 fmol/L, Big-ET-1 allowed to distinguish patients with HFpEF from persons without HF, independently of GFR. In general, NT-proBNP is a good indicator of suspected heart failure. While for NT-proBNP different cut-off points have to be considered in the diagnosis of HFpEF, a single cut-off point of Big-ET-1 was appropriate in the diagnosis of HFpEF, regardless of the presence or absence of CKD. An additional measurement of Big-ET-1 improves the diagnosis of HFpEF in patients with chronic kidney disease.

Association of soluble CD40L with short-term and long-term cardiovascular and all-cause mortality: The Ludwigshafen Risk and Cardiovascular Health (LURIC) study.

BACKGROUND AND AIMS: The CD40-CD40 Ligand (CD40L) system has an important role in vascular inflammation. For this reason, we assessed the association of soluble CD40L with cardiovascular and all-cause mortality in the Ludwigshafen Risk and Cardiovascular Health (LURIC) study. METHODS: Plasma levels of sCD40L were determined in 2759 persons using an enzyme immunoassay. Cox proportional hazard regressions were performed to evaluate the association between plasma concentration of sCD40 ligand and short-term (12 months) and long-term (10 years) mortality. Subpopulation analyses were conducted in seven different risk groups. Cox regression models were adjusted for traditional risk factors. RESULTS: The present study did not reveal significant association between sCD40L plasma levels and all-cause mortality, as well as cardiovascular mortality at one-year follow-up. In selected subgroups only, significant association between elevated sCD40L plasma levels and short-term all-cause and cardiovascular mortality could be observed. With regard to long-term all-cause and cardiovascular mortality analyses, no significant correlation with increased plasma levels of sCD40L could be detected, neither overall nor in any subgroup. CONCLUSIONS: Soluble sCD40L is not associated with cardiovascular and all-cause mortality in this large cohort. Only in selected patient subgroups elevated levels of sCD40L correlate with short-term mortality but this correlation disappears in long-term analysis.

Cardiovascular risk algorithms in primary care: Results from the DETECT study.

Guidelines for prevention of cardiovascular diseases use risk scores to guide the intensity of treatment. A comparison of these scores in a German population has not been performed. We have evaluated the correlation, discrimination and calibration of ten commonly used risk equations in primary care in 4044 participants of the DETECT (Diabetes and Cardiovascular Risk Evaluation: Targets and Essential Data for Commitment of Treatment) study. The risk equations correlate well with each other. All risk equations have a similar discriminatory power. Absolute risks differ widely, in part due to the components of clinical endpoints predicted: The risk equations produced median risks between 8.4% and 2.0%. With three out of 10 risk scores calculated and observed risks well coincided. At a risk threshold of 10 percent in 10 years, the ACC/AHA atherosclerotic cardiovascular disease (ASCVD) equation has a sensitivity to identify future CVD events of approximately 80%, with the highest specificity (69%) and positive predictive value (17%) among all the equations. Due to the most precise calibration over a wide range of risks, the large age range covered and the combined endpoint including non-fatal and fatal events, the ASCVD equation provides valid risk prediction for primary prevention in Germany.

Mendelian Randomization Analysis Reveals a Causal Influence of Circulating Sclerostin Levels on Bone Mineral Density and Fractures.

In bone, sclerostin is mainly osteocyte-derived and plays an important local role in adaptive responses to mechanical loading. Whether circulating levels of sclerostin also play a functional role is currently unclear, which we aimed to examine by two-sample Mendelian randomization (MR). A genetic instrument for circulating sclerostin, derived from a genomewide association study (GWAS) meta-analysis of serum sclerostin in 10,584 European-descent individuals, was examined in relation to femoral neck bone mineral density (BMD; n = 32,744) in GEFOS and estimated bone mineral density (eBMD) by heel ultrasound (n = 426,824) and fracture risk (n = 426,795) in UK Biobank. Our GWAS identified two novel serum sclerostin loci, B4GALNT3 (standard deviation [SD]) change in sclerostin per A allele (ß = 0.20, p = 4.6 × 10-49 ) and GALNT1 (ß = 0.11 per G allele, p = 4.4 × 10-11 ). B4GALNT3 is an N-acetyl-galactosaminyltransferase, adding a terminal LacdiNAc disaccharide to target glycocoproteins, found to be predominantly expressed in kidney, whereas GALNT1 is an enzyme causing mucin-type O-linked glycosylation. Using these two single-nucleotide polymorphisms (SNPs) as genetic instruments, MR revealed an inverse causal relationship between serum sclerostin and femoral neck BMD (ß = -0.12, 95% confidence interval [CI] -0.20 to -0.05) and eBMD (ß = -0.12, 95% CI -0.14 to -0.10), and a positive relationship with fracture risk (ß = 0.11, 95% CI 0.01 to 0.21). Colocalization analysis demonstrated common genetic signals within the B4GALNT3 locus for higher sclerostin, lower eBMD, and greater B4GALNT3 expression in arterial tissue (probability >99%). Our findings suggest that higher sclerostin levels are causally related to lower BMD and greater fracture risk. Hence, strategies for reducing circulating sclerostin, for example by targeting glycosylation enzymes as suggested by our GWAS results, may prove valuable in treating osteoporosis. © 2019 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals, Inc.

Familial hypercholesterolemia in primary care in Germany. Diabetes and cardiovascular risk evaluation: Targets and Essential Data for Commitment of Treatment (DETECT) study.

BACKGROUND AND AIMS: Familial hypercholesterolemia (FH) is an inherited disorder of lipoprotein metabolism characterised by impaired removal of low-density lipoproteins (LDL) from the circulation, which leads to an increased risk of cardiovascular disease (CVD). This risk can be significantly lowered by early diagnosis and treatment. In Germany, reliable estimates of the prevalence of FH are lacking. We therefore examined the prevalence rate of FH in Germany in a primary care based cohort. METHOD: We utilized records of 4722 participants in the DETECT study, in whom complete data on blood lipids and medical history were available. Prevalence rates were assessed using the Dutch Lipid Clinics Network (DLCN) and the US-MEDPED criteria. We stratified for gender and age. Group differences were analyzed using Chi2 and ANOVA tests. RESULTS: Using the DLCN (probable or definite FH) and the US.MEDPED criteria yielded prevalence rates of 1:278 and 1:295, respectively. The established diagnostic scores used in this analysis identify different patients. In women below 50 years of age, the LDL-C concentration is lower than in men, leading to the possibility of under-diagnosing FH in this group because women under the age of 50 are less likely to reach a higher DLCN-Score. CONCLUSIONS: FH has a higher than expected prevalence in Germany. Clinical diagnostic algorithms may not be concordant.