Associations of carotid intima media thickness with gene expression in whole blood and genetically predicted gene expression across 48 tissues.

Carotid intima media thickness (cIMT) is a biomarker of subclinical atherosclerosis and a predictor of future cardiovascular events. Identifying associations between gene expression levels and cIMT may provide insight to atherosclerosis etiology. Here, we use two approaches to identify associations between mRNA levels and cIMT: differential gene expression analysis in whole blood and S-PrediXcan. We used microarrays to measure genome-wide whole blood mRNA levels of 5647 European individuals from four studies. We examined the association of mRNA levels with cIMT adjusted for various potential confounders. Significant associations were tested for replication in three studies totaling 3943 participants. Next, we applied S-PrediXcan to summary statistics from a cIMT genome-wide association study (GWAS) of 71 128 individuals to estimate the association between genetically determined mRNA levels and cIMT and replicated these analyses using S-PrediXcan on an independent GWAS on cIMT that included 22 179 individuals from the UK Biobank. mRNA levels of TNFAIP3, CEBPD and METRNL were inversely associated with cIMT, but these associations were not significant in the replication analysis. S-PrediXcan identified associations between cIMT and genetically determined mRNA levels for 36 genes, of which six were significant in the replication analysis, including TLN2, which had not been previously reported for cIMT. There was weak correlation between our results using differential gene expression analysis and S-PrediXcan. Differential expression analysis and S-PrediXcan represent complementary approaches for the discovery of associations between phenotypes and gene expression. Using these approaches, we prioritize TNFAIP3, CEBPD, METRNL and TLN2 as new candidate genes whose differential expression might modulate cIMT.

Association of plasma proteomics with mortality in individuals with and without type 2 diabetes: Results from two population-based KORA cohort studies.

BACKGROUND: Protein biomarkers may contribute to the identification of vulnerable subgroups for premature mortality. This study aimed to investigate the association of plasma proteins with all-cause and cause-specific mortality among individuals with and without baseline type 2 diabetes (T2D) and evaluate their impact on the prediction of all-cause mortality in two prospective Cooperative Health Research in the Region of Augsburg (KORA) studies. METHODS: The discovery cohort comprised 1545 participants (median follow-up 15.6 years; 244 with T2D: 116 total, 62 cardiovascular, 31 cancer-related and 23 other-cause deaths; 1301 without T2D: 321 total, 114 cardiovascular, 120 cancer-related and 87 other-cause deaths). The validation cohort comprised 1031 participants (median follow-up 6.9 years; 203 with T2D: 76 total, 45 cardiovascular, 19 cancer-related and 12 other-cause deaths; 828 without T2D: 169 total, 74 cardiovascular, 39 cancer-related and 56 other-cause deaths). We used Cox regression to examine associations of 233 plasma proteins with all-cause and cause-specific mortality and Lasso regression to construct prediction models for all-cause mortality stratifying by baseline T2D. C-index, category-free net reclassification index (cfNRI), and integrated discrimination improvement (IDI) were conducted to evaluate the predictive performance of built prediction models. RESULTS: Thirty-five and 62 proteins, with 29 overlapping, were positively associated with all-cause mortality in the group with and without T2D, respectively. Out of these, in the group with T2D, 35, eight, and 26 were positively associated with cardiovascular, cancer-related, and other-cause mortality, while in the group without T2D, 55, 41, and 47 were positively associated with respective cause-specific outcomes in the pooled analysis of both cohorts. Regulation of insulin-like growth factor (IGF) transport and uptake by IGF-binding proteins emerged as a unique pathway enriched for all-cause and cardiovascular mortality in individuals with T2D. The combined model containing the selected proteins (five and 12 proteins, with four overlapping, in the group with and without T2D, respectively) and clinical risk factors improved the prediction of all-cause mortality by C-index, cfNRI, and IDI. CONCLUSIONS: This study uncovered shared and unique mortality-related proteins in persons with and without T2D and emphasized the role of proteins in improving the prediction of mortality in different T2D subgroups.

Association of plasma proteomics with incident coronary heart disease in individuals with and without type 2 diabetes: results from the population-based KORA study.

BACKGROUND: Coronary heart disease (CHD) is a major global health concern, especially among individuals with type 2 diabetes (T2D). Given the crucial role of proteins in various biological processes, this study aimed to elucidate the aetiological role and predictive performance of protein biomarkers on incident CHD in individuals with and without T2D. METHODS: The discovery cohort included 1492 participants from the Cooperative Health Research in the Region of Augsburg (KORA) S4 study with 147 incident CHD cases (45 vs. 102 cases in the group with T2D and without T2D, respectively) during 15.6 years of follow-up. The validation cohort included 888 participants from the KORA-Age1 study with 70 incident CHD cases (19 vs. 51 cases in the group with T2D and without T2D, respectively) during 6.9 years of follow-up. We measured 233 plasma proteins related to cardiovascular disease and inflammation using proximity extension assay technology. Associations of proteins with incident CHD were assessed using Cox regression and Mendelian randomization (MR) analysis. Predictive models were developed using priority-Lasso and were evaluated on top of Framingham risk score variables using the C-index, category-free net reclassification index (cfNRI), and relative integrated discrimination improvement (IDI). RESULTS: We identified two proteins associated with incident CHD in individuals with and 29 in those without baseline T2D, respectively. Six of these proteins are novel candidates for incident CHD. MR suggested a potential causal role for hepatocyte growth factor in CHD development. The developed four-protein-enriched model for individuals with baseline T2D (ΔC-index: 0.017; cfNRI: 0.253; IDI: 0.051) and the 12-protein-enriched model for individuals without baseline T2D (ΔC-index: 0.054; cfNRI: 0.462; IDI: 0.024) consistently improved CHD prediction in the discovery cohort, while in the validation cohort, significant improvements were only observed for selected performance measures (with T2D: cfNRI: 0.633; without T2D: ΔC-index: 0.038; cfNRI: 0.465). CONCLUSIONS: This study identified novel protein biomarkers associated with incident CHD in individuals with and without T2D and reaffirmed previously reported protein candidates. These findings enhance our understanding of CHD pathophysiology and provide potential targets for prevention and treatment.

Plasma proteome association with coronary heart disease and carotid intima media thickness: results from the KORA F4 study.

BACKGROUND AND AIMS: Atherosclerosis is the main cause of stroke and coronary heart disease (CHD), both leading mortality causes worldwide. Proteomics, as a high-throughput method, could provide helpful insights into the pathological mechanisms underlying atherosclerosis. In this study, we characterized the associations of plasma protein levels with CHD and with carotid intima-media thickness (CIMT), as a surrogate measure of atherosclerosis. METHODS: The discovery phase included 1000 participants from the KORA F4 study, whose plasma protein levels were quantified using the aptamer-based SOMAscan proteomics platform. We evaluated the associations of plasma protein levels with CHD using logistic regression, and with CIMT using linear regression. For both outcomes we applied two models: an age-sex adjusted model, and a model additionally adjusted for body mass index, smoking status, physical activity, diabetes status, hypertension status, low density lipoprotein, high density lipoprotein, and triglyceride levels (fully-adjusted model). The replication phase included a matched case-control sample from the independent KORA F3 study, using ELISA-based measurements of galectin-4. Pathway analysis was performed with nominally associated proteins (p-value < 0.05) from the fully-adjusted model. RESULTS: In the KORA F4 sample, after Bonferroni correction, we found CHD to be associated with five proteins using the age-sex adjusted model: galectin-4 (LGALS4), renin (REN), cathepsin H (CTSH), and coagulation factors X and Xa (F10). The fully-adjusted model yielded only the positive association of galectin-4 (OR = 1.58, 95% CI = 1.30-1.93), which was successfully replicated in the KORA F3 sample (OR = 1.40, 95% CI = 1.09-1.88). For CIMT, we found four proteins to be associated using the age-sex adjusted model namely: cytoplasmic protein NCK1 (NCK1), insulin-like growth factor-binding protein 2 (IGFBP2), growth hormone receptor (GHR), and GDNF family receptor alpha-1 (GFRA1). After assessing the fully-adjusted model, only NCK1 remained significant (ß = 0.017, p-value = 1.39e-06). Upstream regulators of galectin-4 and NCK1 identified from pathway analysis were predicted to be involved in inflammation pathways. CONCLUSIONS: Our proteome-wide association study identified galectin-4 to be associated with CHD and NCK1 to be associated with CIMT. Inflammatory pathways underlying the identified associations highlight the importance of inflammation in the development and progression of CHD.

Bidirectional modulation of TCA cycle metabolites and anaplerosis by metformin and its combination with SGLT2i.

BACKGROUND: Metformin and sodium-glucose-cotransporter-2 inhibitors (SGLT2i) are cornerstone therapies for managing hyperglycemia in diabetes. However, their detailed impacts on metabolic processes, particularly within the citric acid (TCA) cycle and its anaplerotic pathways, remain unclear. This study investigates the tissue-specific metabolic effects of metformin, both as a monotherapy and in combination with SGLT2i, on the TCA cycle and associated anaplerotic reactions in both mice and humans. METHODS: Metformin-specific metabolic changes were initially identified by comparing metformin-treated diabetic mice (MET) with vehicle-treated db/db mice (VG). These findings were then assessed in two human cohorts (KORA and QBB) and a longitudinal KORA study of metformin-naïve patients with Type 2 Diabetes (T2D). We also compared MET with db/db mice on combination therapy (SGLT2i + MET). Metabolic profiling analyzed 716 metabolites from plasma, liver, and kidney tissues post-treatment, using linear regression and Bonferroni correction for statistical analysis, complemented by pathway analyses to explore the pathophysiological implications. RESULTS: Metformin monotherapy significantly upregulated TCA cycle intermediates such as malate, fumarate, and α-ketoglutarate (α-KG) in plasma, and anaplerotic substrates including hepatic glutamate and renal 2-hydroxyglutarate (2-HG) in diabetic mice. Downregulated hepatic taurine was also observed. The addition of SGLT2i, however, reversed these effects, such as downregulating circulating malate and α-KG, and hepatic glutamate and renal 2-HG, but upregulated hepatic taurine. In human T2D patients on metformin therapy, significant systemic alterations in metabolites were observed, including increased malate but decreased citrulline. The bidirectional modulation of TCA cycle intermediates in mice influenced key anaplerotic pathways linked to glutaminolysis, tumorigenesis, immune regulation, and antioxidative responses. CONCLUSION: This study elucidates the specific metabolic consequences of metformin and SGLT2i on the TCA cycle, reflecting potential impacts on the immune system. Metformin shows promise for its anti-inflammatory properties, while the addition of SGLT2i may provide liver protection in conditions like metabolic dysfunction-associated steatotic liver disease (MASLD). These observations underscore the importance of personalized treatment strategies.

Stool and blood metabolomics in the metabolic syndrome: a cross-sectional study.

INTRODUCTION/OBJECTIVES: Changes in the stool metabolome have been poorly studied in the metabolic syndrome (MetS). Moreover, few studies have explored the relationship of stool metabolites with circulating metabolites. Here, we investigated the associations between stool and blood metabolites, the MetS and systemic inflammation. METHODS: We analyzed data from 1,370 participants of the KORA FF4 study (Germany). Metabolites were measured by Metabolon, Inc. (untargeted) in stool, and using the AbsoluteIDQ® p180 kit (targeted) in blood. Multiple linear regression models, adjusted for dietary pattern, age, sex, physical activity, smoking status and alcohol intake, were used to estimate the associations of metabolites with the MetS, its components and high-sensitivity C-reactive protein (hsCRP) levels. Partial correlation and Multi-Omics Factor Analysis (MOFA) were used to investigate the relationship between stool and blood metabolites. RESULTS: The MetS was significantly associated with 170 stool and 82 blood metabolites. The MetS components with the highest number of associations were triglyceride levels (stool) and HDL levels (blood). Additionally, 107 and 27 MetS-associated metabolites (in stool and blood, respectively) showed significant associations with hsCRP levels. We found low partial correlation coefficients between stool and blood metabolites. MOFA did not detect shared variation across the two datasets. CONCLUSIONS: The MetS, particularly dyslipidemia, is associated with multiple stool and blood metabolites that are also associated with systemic inflammation. Further studies are necessary to validate our findings and to characterize metabolic alterations in the MetS. Although our analyses point to weak correlations between stool and blood metabolites, additional studies using integrative approaches are warranted.

Type 2 diabetes incidence in patients initiating denosumab or alendronate treatment: a primary care cohort study.

Denosumab initiation is related to a lower risk of type 2 diabetes than alendronate in anti-osteoporotic treatment-naïve users in primary care practices. PURPOSE: Links have been suggested between bone metabolism and glucose tolerance. Downregulation of the receptor activator of nuclear factor κ B ligand (RANKL) signaling improves glucose metabolism. Denosumab, a human monoclonal antibody against RANKL, may be associated with a lower risk of type 2 diabetes (T2D). The aim was to compare incidence rates of T2DM in primary care patients initiating denosumab or alendronate, which is a first-line therapy of osteoporosis. Alendronate as comparator enhances comparability of the two cohorts. METHOD: The IQVIA Disease Analyzer comprises a representative panel of general and specialist practices (Germany). A new-user comparative study was conducted among patients with denosumab or alendronate treatment (2010-2021) without history of diabetes and age ≥ 45 years. Incidence rates (per 1,000 person-years) and Cox proportional hazard ratios (HR; 95%CI) for T2DM were estimated. RESULTS: The cohorts consisted of 3,354 denosumab (age: 75 years; women: 87%) and 27,068 alendronate (76 years; 86%) users. Overall, 1,038 persons developed T2D during 54,916 person-years. T2DM incidence rates per 1,000 person-years were 11.9 (9.5-14.4) for denosumab and 20.1 (18.8-21.3) for alendronate users, respectively. Denosumab was associated with a reduced risk of T2DM compared to alendronate, adjusting for age, sex, index year, visits, obesity, comorbidities and statins (HR: 0.73; 0.58-0.89). CONCLUSION: In this comparative study of older patients seen in routine practices, denosumab was associated with a lower risk of developing T2DM than alendronate.

Associations between multiple neurological biomarkers and distal sensorimotor polyneuropathy: KORA F4/FF4 study.

AIMS: The aim of this study was to assess associations between neurological biomarkers and distal sensorimotor polyneuropathy (DSPN). MATERIALS AND METHODS: Cross-sectional analyses were based on 1032 participants aged 61-82 years from the population-based KORA F4 survey, 177 of whom had DSPN at baseline. The prevalence of type 2 diabetes was 20%. Prospective analyses used data from 505 participants without DSPN at baseline, of whom 125 had developed DSPN until the KORA FF4 survey. DSPN was defined based on the examination part of the Michigan Neuropathy Screening Instrument. Serum levels of neurological biomarkers were measured using proximity extension assay technology. Associations between 88 biomarkers and prevalent or incident DSPN were estimated using Poisson regression with robust error variance and are expressed as risk ratios (RR) and 95% CI per 1-SD increase. Results were adjusted for multiple confounders and multiple testing using the Benjamini-Hochberg procedure. RESULTS: Higher serum levels of CTSC (cathepsin C; RR [95% CI] 1.23 (1.08; 1.39), pB-H = 0.044) and PDGFRα (platelet-derived growth factor receptor A; RR [95% CI] 1.21 (1.08; 1.35), pB-H = 0.044) were associated with prevalent DSPN in the total study sample. CDH3, JAM-B, LAYN, RGMA and SCARA5 were positively associated with DSPN in the diabetes subgroup, whereas GCP5 was positively associated with DSPN in people without diabetes (all pB-H for interaction <0.05). None of the biomarkers showed an association with incident DSPN (all pB-H>0.05). CONCLUSIONS: This study identified multiple novel associations between neurological biomarkers and prevalent DSPN, which may be attributable to functions of these proteins in neuroinflammation, neural development and myelination.

Association of transketolase polymorphisms with diabetic polyneuropathy in the general population: The KORA F4 study.

AIMS: We recently reported that genetic variability in the TKT gene encoding transketolase, a key enzyme in the pentose phosphate pathway, is associated with measures of diabetic sensorimotor polyneuropathy (DSPN) in recent-onset diabetes. Here, we aimed to substantiate these findings in a population-based KORA F4 study. MATERIALS AND METHODS: In this cross-sectional study, we assessed seven single nucleotide polymorphisms (SNPs) in the transketolase gene in 952 participants from the KORA F4 study with normal glucose tolerance (NGT; n = 394), prediabetes (n = 411), and type 2 diabetes (n = 147). DSPN was defined by the examination part of the Michigan Neuropathy Screening Instrument (MNSI) using the original MNSI > 2 cut-off and two alternative versions extended by touch/pressure perception (TPP) (MNSI > 3) and by TPP plus cold perception (MNSI > 4). RESULTS: After adjustment for sex, age, BMI, and HbA1c, in type 2 diabetes participants, four out of seven transketolase SNPs were associated with DSPN for all three MNSI versions (all p ≤ 0.004). The odds ratios of these associations increased with extending the MNSI score, for example, OR (95% CI) for SNP rs62255988 with MNSI > 2: 1.99 (1.16-3.41), MNSI > 3: 2.27 (1.26-4.09), and MNSI > 4: 4.78 (2.22-10.26); SNP rs9284890 with MNSI > 2: 2.43 (1.42-4.16), MNSI > 3: 3.46 (1.82-6.59), and MNSI > 4: 4.75 (2.15-10.51). In contrast, no associations were found between transketolase SNPs and the three MNSI versions in the NGT and prediabetes groups. CONCLUSIONS: The link of genetic variation in transketolase enzyme to diabetic polyneuropathy corroborated at the population level strengthens the concept suggesting an important role of pathways metabolising glycolytic intermediates in the evolution of diabetic polyneuropathy.

Sex hormone-binding globulin may explain sex differences for glucose homeostasis and incidence of type 2 diabetes: the KORA study.

Research has indicated that sex hormone-binding globulin (SHBG) is associated with glucose homeostasis and may play a role in the etiology of type 2 diabetes (T2D). While it is unclear whether SHBG may mediate sex differences in glucose control and subsequently, incidence of T2D. We used observational data from the German population-based KORA F4 study (n = 1937, mean age: 54 years, 41% women) and its follow-up examination KORA FF4 (median follow-up 6.5 years, n = 1387). T2D was initially assessed by self-report and validated by contacting the physicians and/ or reviewing the medical charts. Mediation analyses were performed to assess the role of SHBG in mediating the association between sex (women vs. men) and glucose- and insulin-related traits (cross-sectional analysis) and incidence of T2D (longitudinal analysis). After adjustment for confounders, (model 1: adjusted for age; model 2: model 1 + smoking + alcohol consumption + physical activity), women had lower fasting glucose levels compared to men (ß = -4.94 (mg/dl), 95% CI: -5.77, -4.11). SHBG levels were significantly higher in women than in men (ß = 0.47 (nmol/l), 95% CI:0.42, 0.51). Serum SHBG may mediate the association between sex and fasting glucose levels with a proportion mediated (PM) of 30% (CI: 22-41%). Also, a potential mediatory role of SHBG was observed for sex differences in incidence of T2D (PM = 95% and 63% in models 1 and 2, respectively). Our novel findings suggest that SHBG may partially explain sex-differences in glucose control and T2D incidence.