Sex and statin-related genetic associations at the PCSK9 gene locus: results of genome-wide association meta-analysis.

BACKGROUND: Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a key player of lipid metabolism with higher plasma levels in women throughout their life. Statin treatment affects PCSK9 levels also showing evidence of sex-differential effects. It remains unclear whether these differences can be explained by genetics. METHODS: We performed genome-wide association meta-analyses (GWAS) of PCSK9 levels stratified for sex and statin treatment in six independent studies of Europeans (8936 women/11,080 men respectively 14,825 statin-free/5191 statin-treated individuals). Loci associated in one of the strata were tested for statin- and sex-interactions considering all independent signals per locus. Independent variants at the PCSK9 gene locus were then used in a stratified Mendelian Randomization analysis (cis-MR) of PCSK9 effects on low-density lipoprotein cholesterol (LDL-C) levels to detect differences of causal effects between the subgroups. RESULTS: We identified 11 loci associated with PCSK9 in at least one stratified subgroup (p < 1.0 × 10-6), including the PCSK9 gene locus and five other lipid loci: APOB, TM6SF2, FADS1/FADS2, JMJD1C, and HP/HPR. The interaction analysis revealed eight loci with sex- and/or statin-interactions. At the PCSK9 gene locus, there were four independent signals, one with a significant sex-interaction showing stronger effects in men (rs693668). Regarding statin treatment, there were two significant interactions in PCSK9 missense mutations: rs11591147 had stronger effects in statin-free individuals, and rs11583680 had stronger effects in statin-treated individuals. Besides replicating known loci, we detected two novel genome-wide significant associations: one for statin-treated individuals at 6q11.1 (within KHDRBS2) and one for males at 12q24.22 (near KSR2/NOS1), both with significant interactions. In the MR of PCSK9 on LDL-C, we observed significant causal estimates within all subgroups, but significantly stronger causal effects in statin-free subjects compared to statin-treated individuals. CONCLUSIONS: We performed the first double-stratified GWAS of PCSK9 levels and identified multiple biologically plausible loci with genetic interaction effects. Our results indicate that the observed sexual dimorphism of PCSK9 and its statin-related interactions have a genetic basis. Significant differences in the causal relationship between PCSK9 and LDL-C suggest sex-specific dosages of PCSK9 inhibitors.

The protein "proprotein convertase subtilisin/kexin type 9" (PCSK9) regulates the levels of low-density lipoprotein cholesterol (LDL-C) in blood, and thus, contributes to the risk of cardio-vascular diseases. Women tend to have higher PCSK9 plasma levels throughout their life, although the difference is smaller in patients under LDL-C lowering medication (e.g., statins). We investigated the interplay of genetics, statin-treatment and sex, using combined data from six European studies. We detected 11 genetic regions associated with PCSK9 levels, of which one was specific for women (at SLCO1B3, a statin-transporter gene), and three were specific for men (e.g., ALOX5, encoding a protein linked to chronic inflammatory diseases such as atherosclerosis). We also tested if statin use changed the genetic effect and found five genes only associated with PCSK9 levels in untreated participants. Variants in the gene encoding PCSK9 were most strongly associated and had heterogeneous effects in dependence on statin treatment and sex: On one hand, there were genetic variants with stronger effects in men than women. Those variants are also linked to sex-differential gene expression of PCSK9. On the other hand, there were also variants with treatment-depending effects, linked to protein structure and functionality of PCSK9. This indicates that the observed sexual and treatment-related effects on PCSK9 levels have a genetic basis. In addition, we compared the causal effects of PCSK9 on LDL-C levels between men and women and found a different response to statin treatment. This highlights the need for sex-sensitive dosages of lipid-lowering medication.

Bioactive adrenomedullin and interleukin-6 in COVID-19: potential biomarkers of acute kidney injury and critical illness.

BACKGROUND: The aim of this study was to investigate whether bioactive adrenomedullin (bio-ADM) and interleukin-6 (IL-6) are related to acute kidney injury (AKI) and severe illness in COVID-19 patients. METHODS: 153 patients with COVID-19 admitted to the emergency department (ED) were included. Blood samples were collected from each patient at admission. Bio-ADM and IL-6, as well as DPP3 and routinely measured markers were evaluated regarding the endpoints AKI (22/128 hospitalized patients) and a composite endpoint of admission to intensive care unit and/or in-hospital death (n = 26/153 patients). RESULTS: Bio-ADM and IL-6 were significantly elevated in COVID-19 patients with AKI compared to COVID-19 patients without AKI (each p < 0.001). According to ROC analyses IL-6 and bio-ADM had the largest AUC (0.84 and 0.81) regarding the detection of AKI. Furthermore, bio-ADM and IL-6 were significantly elevated in COVID-19 patients reaching the composite endpoint (each p < 0.001). Regarding the composite endpoint ROC analysis showed an AUC of 0.89 for IL-6 and 0.83 for bio-ADM in COVID-19 patients. In the multivariable logistic model bio-ADM and IL-6 presented as independent significant predictors regarding both endpoints AKI and the composite endpoint in COVID-19 patients (as well as creatinine regarding the composite endpoint; each p < 0.05), opposite to leukocytes, C-reactive protein (CRP) and dipeptidyl peptidase 3 (DPP3; each p = n.s.). CONCLUSION: Elevated levels of bio-ADM and IL-6 are associated with AKI and critical illness in patients with COVID-19. Therefore, both biomarkers may be potential tools in risk stratification in COVID-19 patients at presentation in the ED.

X-chromosome and kidney function: evidence from a multi-trait genetic analysis of 908,697 individuals reveals sex-specific and sex-differential findings in genes regulated by androgen response elements.

X-chromosomal genetic variants are understudied but can yield valuable insights into sexually dimorphic human traits and diseases. We performed a sex-stratified cross-ancestry X-chromosome-wide association meta-analysis of seven kidney-related traits (n = 908,697), identifying 23 loci genome-wide significantly associated with two of the traits: 7 for uric acid and 16 for estimated glomerular filtration rate (eGFR), including four novel eGFR loci containing the functionally plausible prioritized genes ACSL4, CLDN2, TSPAN6 and the female-specific DRP2. Further, we identified five novel sex-interactions, comprising male-specific effects at FAM9B and AR/EDA2R, and three sex-differential findings with larger genetic effect sizes in males at DCAF12L1 and MST4 and larger effect sizes in females at HPRT1. All prioritized genes in loci showing significant sex-interactions were located next to androgen response elements (ARE). Five ARE genes showed sex-differential expressions. This study contributes new insights into sex-dimorphisms of kidney traits along with new prioritized gene targets for further molecular research.

Novel functions of Tribbles-homolog 1 in liver, adipocytes and atherosclerosis.

PURPOSE OF REVIEW: Human genetics studies have sparked great interest in the pseudokinase Tribbles homolog 1, as variant at the TRIB1 gene locus were robustly linked to several cardiometabolic traits, including plasma lipids and coronary artery disease. In this review, we summarize recent findings from mouse models that investigated the function of hepatic and adipocyte Trib1 in lipid metabolism and its role in atherosclerosis. RECENT FINDINGS: Studies in atherosclerosis prone low-density lipoprotein (LDL)-receptor knockout mice suggested that systemic Trib1 -deficiency promotes atherosclerotic lesion formation through the modulation of plasma lipids and inflammation. Further, investigations in mice with hepatocyte specific deletion of Trib1 identified a novel role in the catabolism of apoB-containing lipoproteins via regulation of the LDL-receptor. Moreover, recent studies on Trib1 in adipocytes uncovered critical functions in adipose tissue biology, including the regulation of plasma lipid and adiponectin levels and the response to ß3-adrenergic receptor activation. SUMMARY: Functional studies in mice have expanded our understanding of how Trib1 contributes to various aspects of cardiometabolic diseases. They support the notion that Trib1 exerts tissue-specific effects, which can result in opposing effects on cardiometabolic traits. Additional studies are required to fully elucidate the molecular mechanisms underlying the cellular and systemic effects of Trib1 .

Population-based study of the durability of humoral immunity after SARS-CoV-2 infection.

SARS-CoV-2 antibody quantity and quality are key markers of humoral immunity. However, there is substantial uncertainty about their durability. We investigated levels and temporal change of SARS-CoV-2 antibody quantity and quality. We analyzed sera (8 binding, 4 avidity assays for spike-(S-)protein and nucleocapsid-(N-)protein; neutralization) from 211 seropositive unvaccinated participants, from the population-based longitudinal TiKoCo study, at three time points within one year after infection with the ancestral SARS-CoV-2 virus. We found a significant decline of neutralization titers and binding antibody levels in most assays (linear mixed regression model, p<0.01). S-specific serum avidity increased markedly over time, in contrast to N-specific. Binding antibody levels were higher in older versus younger participants - a difference that disappeared for the asymptomatic-infected. We found stronger antibody decline in men versus women and lower binding and avidity levels in current versus never-smokers. Our comprehensive longitudinal analyses across 13 antibody assays suggest decreased neutralization-based protection and prolonged affinity maturation within one year after infection.

Dipeptidyl-peptidase 3 and IL-6: potential biomarkers for diagnostics in COVID-19 and association with pulmonary infiltrates.

Coronavirus SARS-CoV-2 spread worldwide, causing a respiratory disease known as COVID-19. The aim of the present study was to examine whether Dipeptidyl-peptidase 3 (DPP3) and the inflammatory biomarkers IL-6, CRP, and leucocytes are associated with COVID-19 and able to predict the severity of pulmonary infiltrates in COVID-19 patients versus non-COVID-19 patients. 114 COVID-19 patients and 35 patients with respiratory infections other than SARS-CoV-2 were included in our prospective observational study. Blood samples were collected at presentation to the emergency department. 102 COVID-19 patients and 28 non-COVID-19 patients received CT imaging (19 outpatients did not receive CT imaging). If CT imaging was available, artificial intelligence software (CT Pneumonia Analysis) was used to quantify pulmonary infiltrates. According to the median of infiltrate (14.45%), patients who obtained quantitative CT analysis were divided into two groups (> median: 55 COVID-19 and nine non-COVID-19, ≤ median: 47 COVID-19 and 19 non-COVID-19). DPP3 was significantly elevated in COVID-19 patients (median 20.85 ng/ml, 95% CI 18.34-24.40 ng/ml), as opposed to those without SARS-CoV-2 (median 13.80 ng/ml, 95% CI 11.30-17.65 ng/ml; p < 0.001, AUC = 0.72), opposite to IL-6, CRP (each p = n.s.) and leucocytes (p < 0.05, but lower levels in COVID-19 patients). Regarding binary logistic regression analysis, higher DPP3 concentrations (OR = 1.12, p < 0.001) and lower leucocytes counts (OR = 0.76, p < 0.001) were identified as significant and independent predictors of SARS-CoV-2 infection, as opposed to IL-6 and CRP (each p = n.s.). IL-6 was significantly increased in patients with infiltrate above the median compared to infiltrate below the median both in COVID-19 (p < 0.001, AUC = 0.78) and in non-COVID-19 (p < 0.05, AUC = 0.81). CRP, DPP3, and leucocytes were increased in COVID-19 patients with infiltrate above median (each p < 0.05, AUC: CRP 0.82, DPP3 0.70, leucocytes 0.67) compared to infiltrate below median, opposite to non-COVID-19 (each p = n.s.). Regarding multiple linear regression analysis in COVID-19, CRP, IL-6, and leucocytes (each p < 0.05) were associated with the degree of pulmonary infiltrates, as opposed to DPP3 (p = n.s.). DPP3 showed the potential to be a COVID-19-specific biomarker. IL-6 might serve as a prognostic marker to assess the extent of pulmonary infiltrates in respiratory patients.

Feasibility of peripheral blood stem cell collection from sickle cell trait donors with an intensified G-CSF regimen.

OBJECTIVES: Allogeneic hematopoietic stem cell transplantation (HSCT) is the only curative treatment for SCD and bone marrow from an HLA-matched sibling is currently the standard of care. Haploidentical HSCT from a family donor with a TCR αß/CD19 depleted graft (T-haplo) is an increasingly successful alternative, which requires the generation of G-CSF stimulated peripheral stem cell (PBSC) from haploidentical relatives. These sickle cell trait (SCT) donors reported to develop SCD-related complications in conditions of severe stress. METHODS: In this retrospective analysis, we compared the safety and efficacy of PBSC mobilization with a G-CSF intensified mobilization regimen in SCT donors with a conventional G-CSF mobilization regimen in healthy donors. RESULTS: The reported adverse events were similar during intensified G-CSF mobilization, apheresis, and shortly after stem cell apheresis in SCT and control donors. In SCT and control donors, we were able to mobilize high yields of CD34+ stem cells and the harvested CD34+ cell count was comparable with control donors. CONCLUSIONS: Peripheral stem cell mobilization using an intensified G-CSF regimen is safe, and well tolerated among SCT donors. SCT donors are a valid alternative for collection of peripheral CD34+ stem cells for T-cell-depleted haploidentical stem cell transplantation.

Intestinal IgA-positive plasma cells are highly sensitive indicators of alloreaction early after allogeneic transplantation and associate with both graft-versus-host disease and relapse-related mortality.

Intestinal immunoglobulin A (IgA) is strongly involved in microbiota homeostasis. Since microbiota disruption is a major risk factor of acute graft-versus-host disease (GvHD), we addressed the kinetics of intestinal IgA-positive (IgA+) plasma cells by immunohistology in a series of 430 intestinal biopsies obtained at a median of 1,5 months after allogeneic stem cell transplantation (allo-SCT) from 115 patients (pts) at our center. IgA+ plasma cells were located in the subepithelial lamina propria and suppressed in the presence of histological aGvHD (GvHD Lerner stage 0: 131+/-8 IgA+ plasma cells/mm2; stage 1-2: 108+/-8 IgA+ plasma cells/mm2; stage 3-4: 89+/-16 IgA+ plasma cells/mm2; P=0.004). Overall, pts with IgA+ plasma cells below median had an increased treatment related mortality (P=0.04). Time courses suggested a gradual recovery of IgA+ plasma cells after day 100 in the absence but not in the presence of GvHD. Vice versa IgA+ plasma cells above median early after allo-SCT were predictive of relapse and relapse-related mortality (RRM): pts with low IgA+ cells had a 15% RRM at 2 and at 5 years, while pts with high IgA+ cells had a 31% RRM at 2 years and more than 46% at 5 years; multivariate analysis indicated high IgA+ plasma cells in biopsies (hazard ratio =2.7; 95% confidence interval: 1.04-7.00) as independent predictors of RRM, whereas Lerner stage and disease stage themselves did not affect RRM. In contrast, IgA serum levels at the time of biopsy were not predictive for RRM. In summary, our data indicate that IgA+ cells are highly sensitive indicators of alloreaction early after allo-SCT showing association with TRM but also allowing prediction of relapse independently from the presence of overt GvHD.

Urinary N-Terminal Pro-Brain Natriuretic Peptide Predicts Acute Kidney Injury and Severe Disease in COVID-19.

INTRODUCTION: The ongoing COVID-19 pandemic is placing an extraordinary burden on our health care system with its limited resources. Accurate triage of patients is necessary to ensure medical care for those most severely affected. In this regard, biomarkers could contribute to risk evaluation. The aim of this prospective observational clinical study was to assess the relationship between urinary N-terminal pro-brain natriuretic peptide (NT-proBNP) and acute kidney injury (AKI) as well as severe disease in patients with COVID-19. METHODS: 125 patients treated with an acute respiratory infection in the emergency department of the University Hospital Regensburg were analyzed. These patients were divided into a COVID-19 cohort (n = 91) and a cohort with infections not caused by severe acute respiratory syndrome-coronavirus-2 (n = 34). NT-proBNP was determined from serum and fresh urine samples collected in the emergency department. Clinical endpoints were the development of AKI and a composite one consisting of AKI, intensive care unit admission, and in-hospital death. RESULTS: 11 (12.1%) COVID-19 patients developed AKI during hospitalization, whereas 15 (16.5%) reached the composite endpoint. Urinary NT-proBNP was significantly elevated in COVID-19 patients who suffered AKI or reached the composite endpoint (each p < 0.005). In a multivariate regression analysis adjusted for age, chronic kidney disease, chronic heart failure, and arterial hypertension, urinary NT-proBNP was identified as independent predictor of AKI (p = 0.017, OR = 3.91 [CI: 1.28-11.97] per standard deviation [SD]), as well as of the composite endpoint (p = 0.026, OR 2.66 [CI: 1.13-6.28] per SD). CONCLUSION: Urinary NT-proBNP might help identify patients at risk for AKI and severe disease progression in COVID-19.

Overexpressing high levels of human vaspin limits high fat diet-induced obesity and enhances energy expenditure in a transgenic mouse.

Adipose tissue inflammation and insulin resistance are hallmarks in the development of metabolic diseases resulting from overweight and obesity, such as type 2 diabetes and non-alcoholic fatty liver disease. In obesity, adipocytes predominantly secrete proinflammatory adipokines that further promote adipose tissue dysfunction with negative effects on local and systemic insulin sensitivity. Expression of the serpin vaspin (SERPINA12) is also increased in obesity and type 2 diabetes, but exhibits compensatory roles in inflammation and insulin resistance. This has in part been demonstrated using vaspin-transgenic mice. We here report a new mouse line (h-vaspinTG) with transgenic expression of human vaspin in adipose tissue that reaches vaspin concentrations three orders of magnitude higher than wild type controls (>200 ng/ml). Phenotyping under chow and high-fat diet conditions included glucose-tolerance tests, measurements of energy expenditure and circulating parameters, adipose tissue and liver histology. Also, ex vivo glucose uptake in isolated adipocytes and skeletal muscle was analyzed in h-vaspinTG and littermate controls. The results confirmed previous findings, revealing a strong reduction in diet-induced weight gain, fat mass, hyperinsulinemia, -glycemia and -cholesterolemia as well as fatty liver. Insulin sensitivity in adipose tissue and muscle was not altered. The h-vaspinTG mice showed increased energy expenditure under high fat diet conditions, that may explain reduced weight gain and overall metabolic improvements. In conclusion, this novel human vaspin-transgenic mouse line will be a valuable research tool to delineate whole-body, tissue- and cell-specific effects of vaspin in health and disease.

Polygenic scores for estimated glomerular filtration rate in a population of general adults and elderly - comparative results from the KORA and AugUR study.

BACKGROUND: Polygenic scores (PGSs) combining genetic variants found to be associated with creatinine-based estimated glomerular filtration rate (eGFRcrea) have been applied in various study populations with different age ranges. This has shown that PGS explain less eGFRcrea variance in the elderly. Our aim was to understand how differences in eGFR variance and the percentage explained by PGS varies between population of general adults and elderly. RESULTS: We derived a PGS for cystatin-based eGFR (eGFRcys) from published genome-wide association studies. We used the 634 variants known for eGFRcrea and the 204 variants identified for eGFRcys to calculate the PGS in two comparable studies capturing a general adult and an elderly population, KORA S4 (n = 2,900; age 24-69 years) and AugUR (n = 2,272, age ≥ 70 years). To identify potential factors determining age-dependent differences on the PGS-explained variance, we evaluated the PGS variance, the eGFR variance, and the beta estimates of PGS association on eGFR. Specifically, we compared frequencies of eGFR-lowering alleles between general adult and elderly individuals and analyzed the influence of comorbidities and medication intake. The PGS for eGFRcrea explained almost twice as much (R2 = 9.6%) of age-/sex adjusted eGFR variance in the general adults compared to the elderly (4.6%). This difference was less pronounced for the PGS for eGFRcys (4.7% or 3.6%, respectively). The beta-estimate of the PGS on eGFRcrea was higher in the general adults compared to the elderly, but similar for the PGS on eGFRcys. The eGFR variance in the elderly was reduced by accounting for comorbidities and medication intake, but this did not explain the difference in R2-values. Allele frequencies between general adult and elderly individuals showed no significant differences except for one variant near APOE (rs429358). We found no enrichment of eGFR-protective alleles in the elderly compared to general adults. CONCLUSIONS: We concluded that the difference in explained variance by PGS was due to the higher age- and sex-adjusted eGFR variance in the elderly and, for eGFRcrea, also by a lower PGS association beta-estimate. Our results provide little evidence for survival or selection bias.

Quantification of bulk lipid species in human platelets and their thrombin-induced release.

Lipids play a central role in platelet physiology. Changes in the lipidome have already been described for basal and activated platelets. However, quantitative lipidomic data of platelet activation, including the released complex lipids, are unavailable. Here we describe an easy-to-use protocol based on flow-injection mass spectrometry for the quantitative analysis of bulk lipid species in basal and activated human platelets and their lipid release after thrombin activation. We provide lipid species concentrations of 12 healthy human donors, including cholesteryl ester (CE), ceramide (Cer), free cholesterol (FC), hexosylceramide (HexCer), lysophosphatidylcholine (LPC), lysophosphatidylethanolamine (LPE), phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylinositol (PI), phosphatidylserine (PS), sphingomyelin (SM) and triglycerides (TG). The assay exhibited good technical repeatability (CVs < 5% for major lipid species in platelets). Except for CE and TG, the inter-donor variability of the majority of lipid species concentrations in platelets was < 30% CV. Balancing of concentrations revealed the generation of LPC and loss of TG. Changes in lipid species concentrations indicate phospholipase-mediated release of arachidonic acid mainly from PC, PI, and PE but not from PS. Thrombin induced lipid release was mainly composed of FC, PS, PC, LPC, CE, and TG. The similarity of the released lipidome with that of plasma implicates that lipid release may originate from the open-canalicular system (OCS). The repository of lipid species concentrations determined with this standardized platelet release assay contribute to elucidating the physiological role of platelet lipids and provide a basis for investigating the platelet lipidome in patients with hemorrhagic or thrombotic disorders.

Trib1 Deficiency Promotes Hyperlipidemia, Inflammation, and Atherosclerosis in LDL Receptor Knockout Mice.

BACKGROUND: Genetic variants at the TRIB1 gene locus are strongly associated with plasma lipid traits and the risk of coronary artery disease in humans. Here, we analyzed the consequences of Trib1 deficiency on lipid metabolism and atherosclerotic lesion formation in atherosclerosis-susceptible Ldlr-/- mice. METHODS: Trib1-/- mice were crossed onto the Ldlr-/- background to generate double-knockout mice (Trib1-/-Ldlr-/-) and fed a semisynthetic, modified AIN76 diet (0.02% cholesterol and 4.3% fat) until 20 weeks of age. RESULTS: Trib1-/-Ldlr-/- mice had profoundly larger (5.8-fold) and more advanced atherosclerotic lesions at the aortic root as compared with Trib1+/+Ldlr-/- controls. Further, we observed significantly elevated plasma total cholesterol and triglyceride levels in Trib1-/-Ldlr-/- mice, resulting from higher VLDL (very-low-density lipoprotein) secretion. Lipidomics analysis revealed that loss of Trib1 altered hepatic lipid composition, including the accumulation of cholesterol and proinflammatory ceramide species, which was accompanied by signs of hepatic inflammation and injury. Concomitantly, we detected higher plasma levels of IL (interleukin)-6 and LCN2 (lipocalin 2), suggesting increased systemic inflammation in Trib1-/-Ldlr-/- mice. Hepatic transcriptome analysis demonstrated significant upregulation of key genes controlling lipid metabolism and inflammation in Trib1-/-Ldlr-/- mice. Further experiments suggested that these effects may be mediated through pathways involving a C/EPB (CCAAT/enhancer binding protein)-PPARγ (peroxisome proliferator-activated receptor γ) axis and JNK (c-Jun N-terminal kinase) signaling. CONCLUSIONS: We provide experimental evidence that Trib1 deficiency promotes atherosclerotic lesion formation in a complex manner that includes the modulation of lipid metabolism and inflammation.

Differences in white matter hyperintensities in socioeconomically deprived groups: results of the population-based LIFE Adult Study.

OBJECTIVE: Previous studies have shown that socioeconomically deprived groups exhibit higher lesion load of the white matter (WM) in aging. The aim of this study was to (i) investigate to what extent education and income may contribute to differences in white matter hyperintensities (WMHs) and (ii) identify risk profiles related to a higher prevalence of age-associated WMH. DESIGN AND SETTING: Population-based adult study of the Leipzig Research Centre for Civilization Diseases (LIFE) in Leipzig, Germany. PARTICIPANTS: Dementia-free sample aged 40-80 years (n = 1,185) derived from the population registry. MEASUREMENTS: Information was obtained in standardized interviews. WMH (including the derived Fazekas scores) were assessed using automated segmentation of high-resolution T1-weighted anatomical and fluid-attenuated inversion recovery (FLAIR) MRI acquired at 3T. RESULTS: Despite a significant association between income and WMH in univariate analyses, results from adjusted models (age, gender, arterial hypertension, heart disease, and APOE e4 allele) indicated no association between income and WMH. Education was associated with Fazekas scores, but not with WMH and not after Bonferroni correction. Prevalence of some health-related risk factors was significantly higher among low-income/education groups. After combining risk factors in a factor analysis, results from adjusted models indicated significant associations between higher distress and more WMH as well as between obesity and more deep WMH. CONCLUSIONS: Previously observed differences in WMH between socioeconomically deprived groups might stem from differences in health-related risk factors. These risk factors should be targeted in prevention programs tailored to socioeconomically deprived individuals.

Cardiovascular Risk Factor Control in 70- to 95-Year-Old Individuals: Cross-Sectional Results from the Population-Based AugUR Study.

Cardiovascular risk factors such as high glucose, LDL-cholesterol, blood pressure, and impaired kidney function are particularly frequent in old-aged individuals. However, population-based data on the extent of cardiovascular risk factor control in the old-aged population is limited. AugUR is a cohort of the mobile "70+"-year-old population of/near Regensburg, recruited via population registries. We conducted cross-sectional analyses assessing the proportion of AugUR participants with LDL-cholesterol, HbA1c, or blood pressure beyond recommended levels and their association with impaired creatinine- and cystatin-based estimated glomerular filtration rate (eGFR, <60 mL/min/1.73 m2) or urine albumin-creatinine ratio (UACR, ≥30 mg/g). Among 2215 AugUR participants, 74.7% were taking lipid-, glucose-, blood-pressure-lowering, or diuretic medication. High LDL-cholesterol at ≥116 mg/dL was observed for 76.1% (51.1% among those with prior cardiovascular events). We found HbA1c ≥ 7.0% for 6.3%, and high or low systolic blood pressure for 6.8% or 26.5%, respectively (≥160, <120 mmHg). Logistic regression revealed (i) high HbA1c levels associated with increased risk for impaired kidney function among those untreated, (ii) high blood pressure with increased UACR, and (iii) low blood pressure with impaired eGFR, which was confined to individuals taking diuretics. Our results provide important insights into cardiovascular risk factor control in individuals aged 70-95 years, which are understudied in most population-based studies.

Granulocyte concentrate splitting does not affect phenotype and function.

BACKGROUND: More granulocyte concentrates (GCs) could be produced for more patients from the same donor if apheresis bags were split and stored for longer periods of time. Hence, we tested the hypothesis that splitting and extension of storage of GCs do not impair granulocyte function or viability. STUDY DESIGN AND METHODS: Granulocyte apheresis concentrates were produced using modified fluid gelatin as a separation enhancer, split into two portions, and stored for 24 and 48 h. Granulocyte function, represented by cell migration, reactive oxygen species (ROS) production, and neutrophil extracellular trap formation (NETosis), was measured by live-cell imaging. ROS production, adhesive surface protein expression, and viability were measured by flow cytometry. RESULTS: Splitting had no effect on any of the tested parameters. After 24 h of storage, live-cell imaging showed no significant difference in migration, time to maximum ROS production, time to half-maximum NETosis, viability, or CD11b expression, but ROS production induced by phorbol 12-myristate 13-acetate (PMA) decreased from an initial median fluorescence intensity of 1775-590 artificial units. After 48 h, PMA-induced ROS production, viability, and migration declined, as reflected by decreases in median total distance (119 vs. 63.5 µm) and median Euclidean distance (30.75 vs. 14.3 µm). CONCLUSION: Splitting GC products has no effect on granulocyte viability or function, but extended storage >24 h does compromise granulocyte function. The findings confirm that GCs should be transfused within 24 h of collection. Longer storage cannot be recommended.

Higher Infection Risk among Health Care Workers and Lower Risk among Smokers Persistent across SARS-CoV-2 Waves-Longitudinal Results from the Population-Based TiKoCo Seroprevalence Study.

SARS-CoV-2 seroprevalence was reported as substantially increased in medical personnel and decreased in smokers after the first wave in spring 2020, including in our population-based Tirschenreuth Study (TiKoCo). However, it is unclear whether these associations were limited to the early pandemic and whether the decrease in smokers was due to reduced infection or antibody response. We evaluated the association of occupation and smoking with period-specific seropositivity: for the first wave until July 2020 (baseline, BL), the low infection period in summer (follow-up 1, FU1, November 2020), and the second/third wave (FU2, April 2021). We measured binding antibodies directed to SARS-CoV-2 nucleoprotein (N), viral spike protein (S), and neutralizing antibodies at BL, FU1, and FU2. Previous infection, vaccination, smoking, and occupation were assessed by questionnaires. The 4181 participants (3513/3374 at FU1/FU2) included 6.5% medical personnel and 20.4% current smokers. At all three timepoints, new seropositivity was higher in medical personnel with ORs = 1.99 (95%-CI = 1.36-2.93), 1.41 (0.29-6.80), and 3.17 (1.92-5.24) at BL, FU1, and FU2, respectively, and nearly halved among current smokers with ORs = 0.47 (95%-CI = 0.33-0.66), 0.40 (0.09-1.81), and 0.56 (0.33-0.94). Current smokers compared to never-smokers had similar antibody levels after infection or vaccination and reduced odds of a positive SARS-CoV-2 result among tested. Our data suggest that decreased seroprevalence among smokers results from fewer infections rather than reduced antibody response. The persistently higher infection risk of medical staff across infection waves, despite improved means of protection over time, underscores the burden for health care personnel.

Association between Adverse Reactions and Humoral Immune Response No Longer Detectable after BNT162b2 Booster Vaccination.

In a previous study, we described a highly significant association between reactogenicity and SARS-CoV-2 RBD IgG titers and wild-type neutralization capacity in males after basic vaccination with BNT162b2. The objective of this study was to assess whether this benefit was long lasting and also evident after BNT162b2 booster vaccination. Reactogenicity was classified into three groups: no or minor injection site symptoms, moderate (not further classified) and severe adverse reactions (defined as any symptom(s) resulting in sick leave). We initially compared 76 non-immunocompromised individuals who reported either no or minor injection site symptoms or severe adverse reactions after second vaccination. In total, 65 of them took part in another blood sampling and 47 were evaluated after booster vaccination. 26 weeks after second vaccination, men who reported severe adverse reactions after second vaccination had 1.7-fold higher SARS-CoV-2 RBD IgG titers (p = 0.025) and a 2.5-fold better neutralization capacity (p = 0.006) than men with no or only minor injection site symptoms. Again, no association was found in women. Reactogenicity of BNT162b2 booster vaccination was different from second vaccination according to our classification and was no longer associated with SARS-CoV-2 RBD IgG titers or wild-type neutralization capacity. To conclude, after BNT162b2 basic vaccination, the association between reactogenicity and humoral immune response in men persisted over time but was no longer detectable after BNT162b2 booster vaccination.

External validation of biomarkers for immune-related adverse events after immune checkpoint inhibition.

Immune checkpoint inhibitors have revolutionized treatment of advanced melanoma, but commonly cause serious immune-mediated complications. The clinical ambition of reserving more aggressive therapies for patients least likely to experience immune-related adverse events (irAE) has driven an extensive search for predictive biomarkers. Here, we externally validate the performance of 59 previously reported markers of irAE risk in a new cohort of 110 patients receiving Nivolumab (anti-PD1) and Ipilimumab (anti-CTLA-4) therapy. Alone or combined, the discriminatory value of these routine clinical parameters and flow cytometry biomarkers was poor. Unsupervised clustering of flow cytometry data returned four T cell subsets with higher discriminatory capacity for colitis than previously reported populations, but they cannot be considered as reliable classifiers. Although mechanisms predisposing some patients to particular irAEs have been described, we are presently unable to capture adequate information from pre-therapy flow cytometry and clinical data to reliably predict risk of irAE in most cases.

Benchmarking One-Phase Lipid Extractions for Plasma Lipidomics.

A key element of successful lipidomics analysis is a sufficient extraction of lipid molecules typically by two-phase systems such as chloroform-based Bligh and Dyer (B&D). However, numerous metabolomics and lipidomics studies today apply easy to use one-phase extractions. In this work, quantitative flow injection analysis high-resolution mass spectrometry was applied to benchmark the lipid recovery of popular one-phase extraction methods for human plasma samples. The following organic solvents were investigated: methanol (MeOH), ethanol (EtOH), 2-propanol (IPA), 1-butanol (BuOH), acetonitrile (ACN) and the solvent mixtures BuOH/MeOH (3:1) and MeOH/ACN (1:1). The recovery of polar lysophospholipids was sufficient for all tested solvents. However, nonpolar lipid classes such as triglycerides (TG) and cholesteryl esters (CE) revealed extraction efficiencies less than 5% due to precipitation in polar solvents EtOH, MeOH, MeOH/ACN, and ACN. Sample pellets also contained a substantial amount of phospholipids, for example, more than 75% of total phosphatidylcholine and sphingomyelin for ACN. The loss of lipids by precipitation was directly related to the polarity of solvents and lipid classes. Although, lipid recovery increased with the volume of organic solvent, recovery in polar MeOH remains incomplete also for less polar lipid classes such as ceramides. Addition of stable isotope-labeled internal standards prior to lipid extraction could compensate for insufficient lipid recovery for polar lipid classes including lysolipids and phospholipids but not for nonpolar CE and TG. In summary, application of one-phase extractions should be limited to polar lipid classes unless sufficient recovery/solubility of nonpolar lipids has been demonstrated. The presented data reveal that appropriate lipid extraction efficiency is fundamental to achieve accurate lipid quantification.