Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a secreted protein that enhances degradation of the LDL receptor. While agents that inhibit PCSK9 markedly reduce atherogenic lipoproteins and ...show great promise for event reduction, it is unknown whether plasma PCSK9 levels predict incident cardiovascular events.
In a nested case-control evaluation conducted in a prospective cohort of >28 000 initially healthy American women, we measured plasma concentrations of PCSK9 at baseline among 358 participants who subsequently developed major cardiovascular events (cases) and among 358 age, smoking, and hormone replacement therapy matched participants who remained free of disease during 17 years of follow-up (controls). Proprotein convertase subtilisin/kexin type 9 level was not significantly related to smoking status, hypertension, obesity, or a family history of premature cardiovascular disease but was positively associated with apolipoprotein B-100 (r = 0.20, P< 0.001), and triglycerides (r = 0.13, P = 0.004). No associations were observed between PCSK9 and apo A1, HDLC, lipoprotein(a), or high-sensitivity C-reactive protein. Despite modest positive association with atherogenic lipids, baseline levels of PCSK9 did not predict the first cardiovascular events; the odds ratios (ORs) for future vascular events for the lowest (referent) to highest baseline quartiles of PCSK9 were 1.0, 0.94, 0.98, and 1.15 (P-trend = 0.53). In contrast, the corresponding ORs for baseline apo B levels were 1.0, 1.14, 1.34, and 1.94 (P-trend = 0.002).
In a large-scale primary prevention cohort, plasma levels of PCSK9 measured at baseline did not predict future cardiovascular events.
Purpose
The comparability between serum, plasma, and urinary measurements of estrogen metabolites via liquid chromatography–tandem mass spectrometry (LC–MS/MS) has not been largely explored, and it ...is unclear if urinary LC–MS/MS measurements are suitable surrogates of circulating levels.
Methods
Serum, plasma (EDTA and heparin), and urinary estrogen/estrogen metabolite levels were measured via LC–MS/MS in paired samples from 64 healthy volunteers (18 men, 20 premenopausal women, 26 postmenopausal women). Geometric means and Spearman correlation coefficients were used to compare individual and combined pathway levels of estrogens/estrogen metabolites across biologic matrices by sex/menopausal status.
Results
Measured concentrations of estrogens/estrogen metabolites across blood matrices were almost identical (percent differences < 4.8%). Parent estrogen concentrations measured in serum and urine were moderately correlated in postmenopausal women (estrone:
r
= 0.69, estradiol:
r
= 0.69). Correlations were similar comparing unconjugated serum estradiol to urinary estrone (
r
= 0.76) and urinary estradiol (
r
= 0.65) in postmenopausal women but were moderate to low in premenopausal women (
r
= 0.60, 0.40, respectively)/men (
r
= 0.33, 0.53, respectively). Comparing metabolite ratios, proportionally higher concentrations of 16-pathway metabolites were measured in urine versus serum across sex/menopausal status groups (e.g., postmenopausal women: 50.3% 16-pathway metabolites/total in urine versus 35.3% in serum).
Conclusions
There is strong agreement between estrogen/estrogen metabolites measurements in serum, heparin plasma, and EDTA plasma. Individual estrogen metabolite concentrations were moderately correlated between urine and serum, but were not well correlated when evaluating pathway- or relative estrogen concentrations. Differences between serum and urine are likely explained by differences in metabolism and/or excretion.
Acute kidney injury (AKI) is associated with high morbidity and mortality. The lack of sensitive and specific injury biomarkers has greatly impeded the development of therapeutic strategies to ...improve outcomes of AKI.
The unique objective of this study was to evaluate the diagnostic performance of nine urinary biomarkers of AKI—kidney injury molecule‐1 (KIM‐1), neutrophil gelatinase associated lipocalin (NGAL), interleukin‐18 (IL‐18), hepatocyte growth factor (HGF), cystatin C (Cys), N‐acetyl‐β‐D‐glucosaminidase (NAG), vascular endothelial growth factor (VEGF), chemokine interferon‐inducible protein 10 (IP‐10; CXCL10), and total protein—in a cross‐sectional comparison of 204 patients with or without AKI.
Median urinary concentrations of each biomarker were significantly higher in patients with AKI than in those without AKI (p < 0.001). The area under the receiver operating characteristics curve (AUC‐ROC) for the combination of biomarkers using a logic regression model risk score of 2.93*(NGAL > 5.72 and HGF > 0.17) + 2.93*(PROTEIN > 0.22) −2*(KIM < 0.58) was greater (0.94) than individual biomarker AUC‐ROCs. Age‐adjusted levels of urinary KIM‐1, NAG, HGF, VEGF, and total protein were significantly higher in patients who died or required renal replacement therapy (RRT) when compared to those who survived and did not require RRT.
Our results demonstrate the comparative value of multiple biomarkers in the diagnosis and prognosis of AKI.
Study objective Previously, we used a proteomics approach for the discovery of new diagnostic markers of acute appendicitis and identified leucine-rich α-2-glycoprotein (LRG) that was elevated in the ...urine of children with acute appendicitis and enriched in diseased appendices. Here, we sought to evaluate the diagnostic utility of enzyme-linked immunosorbent assay (ELISA) of urine LRG in a blinded, prospective, cohort study of children being evaluated for acute abdominal pain. Methods Urine LRG concentration was measured with a commercially available LRG ELISA and selected ion monitoring mass spectrometry. Urine LRG test performance was evaluated blindly against the pathologic diagnosis and histologic grade of appendicitis. Results Urine LRG was measured in 49 patients. Mean urine LRG concentration measured with commercial LRG ELISA was significantly elevated in patients with acute appendicitis but exhibited an interference effect. Direct measurements using selected ion monitoring mass spectrometry demonstrated that LRG was elevated more than 100-fold in patients with acute appendicitis compared with those without, with the receiver operating characteristic area under the curve of 0.98 (95% confidence interval 0.96 to 1.0). Among patients with acute appendicitis, elevations of urine LRG measured with ELISA and selected ion monitoring mass spectrometry correlated with the histologic severity of appendicitis. Conclusion Urine LRG ELISA allows for discrimination between patients with and without acute appendicitis but exhibits limited accuracy because of immunoassay interference. Direct measurements of urine LRG with selected ion monitoring mass spectrometry demonstrate superior diagnostic performance. Development of a clinical-grade urine LRG assay is needed to advance the diagnostic accuracy of clinical evaluations of appendicitis.
Kawasaki disease (KD) is a systemic vasculitis of unknown etiology. Absence of definitive diagnostic markers limits the accuracy of clinical evaluations of suspected KD with significant increases in ...morbidity. In turn, incomplete understanding of its molecular pathogenesis hinders the identification of rational targets needed to improve therapy. We used high‐accuracy mass spectrometry proteomics to analyse over 2000 unique proteins in clinical urine specimens of patients with KD. We discovered that urine proteomes of patients with KD, but not those with mimicking conditions, were enriched for markers of cellular injury such as filamin and talin, immune regulators such as complement regulator CSMD3, immune pattern recognition receptor muclin, and immune cytokine protease meprin A. Significant elevations of filamin C and meprin A were detected in both the serum and urine in two independent cohorts of patients with KD, comprised of a total of 236 patients. Meprin A and filamin C exhibited superior diagnostic performance as compared to currently used markers of disease in a blinded case‐control study of 107 patients with suspected KD, with receiver operating characteristic areas under the curve of 0.98 (95% confidence intervals CI of 0.97–1 and 0.95–1, respectively). Notably, meprin A was enriched in the coronary artery lesions of a mouse model of KD. In all, urine proteome profiles revealed novel candidate molecular markers of KD, including filamin C and meprin A that exhibit excellent diagnostic performance. These disease markers may improve the diagnostic accuracy of clinical evaluations of children with suspected KD, lead to the identification of novel therapeutic targets, and allow the development of a biological classification of Kawasaki disease.
High‐accuracy mass spectrometry proteomics identifies meprin A and filamin C as superior diagnostic markers for Kawasaki disease.
Background and Objectives: Assay cost, quality, and availability pose challenges for vitamin D surveys in limited resource settings. This study aimed to validate an inexpensive vitamin D assay ...(ELISA) under real-world conditions in Mongolia, the northernmost developing country, to characterize the assay's usefulness and inform the design of epidemiologic studies in similar regions.
Methods and Study Design: We collected paired summer and winter serum samples from 120 men and women (aged 20-57 years) in urban and rural Mongolia, analyzed each sample for 25(OH)D concentration using both Immunodiagnostic Systems ELISA and DiaSorin LIAISON 25(OH)D TOTAL, and compared the assays using multiple statistics. LIAISON was itself validated by participation in the DEQAS program.
Results: Correlation and agreement between assays were higher in summer (Pearson's correlation=0.60, Spearman's rank correlation=0.67, Lin's concordance correlation=0.56) than winter (rP=0.37, rS=0.43, rC=0.33), although ELISA less accurately assigned subjects to sufficiency categories in summer (percent agreement=44%) than winter (58%), during the latter of which most subjects were deficient (25(OH)D categories used: >75 nmol/L (optimal), 50-75 nmol/L (adequate), 25-50 nmol/L (inadequate), <25 nmol/L (deficient)). Compared with LIAISON, ELISA tended to indicate higher vitamin D status in both seasons (mean paired difference: 7.0 nmol/L (95% CI: 3.5-10.5) in summer, 5.2 nmol/L (95% CI: 2.9-7.5) in winter).
Conclusions: ELISA proved useful for measuring and ranking subjects' vitamin D status in Mongolia during summer, but levels were too low in winter to sensitively discriminate between subjects, and ELISA overestimated status in both seasons. These findings have implications for the timing and interpretation, respectively, of vitamin D surveys in highly deficient populations.
Background and Objectives: Assay cost, quality, and availability pose challenges for vitamin D surveys in limited resource settings. This study aimed to validate an inexpensive vitamin D assay ...(ELISA) under real-world conditions in Mongolia, the northernmost developing country, to characterize the assay's usefulness and inform the design of epidemiologic studies in similar regions.
Methods and Study Design: We collected paired summer and winter serum samples from 120 men and women (aged 20-57 years) in urban and rural Mongolia, analyzed each sample for 25(OH)D concentration using both Immunodiagnostic Systems ELISA and DiaSorin LIAISON 25(OH)D TOTAL, and compared the assays using multiple statistics. LIAISON was itself validated by participation in the DEQAS program.
Results: Correlation and agreement between assays were higher in summer (Pearson's correlation=0.60, Spearman's rank correlation=0.67, Lin's concordance correlation=0.56) than winter (rP=0.37, rS=0.43, rC=0.33), although ELISA less accurately assigned subjects to sufficiency categories in summer (percent agreement=44%) than winter (58%), during the latter of which most subjects were deficient (25(OH)D categories used: >75 nmol/L (optimal), 50-75 nmol/L (adequate), 25-50 nmol/L (inadequate), <25 nmol/L (deficient)). Compared with LIAISON, ELISA tended to indicate higher vitamin D status in both seasons (mean paired difference: 7.0 nmol/L (95% CI: 3.5-10.5) in summer, 5.2 nmol/L (95% CI: 2.9-7.5) in winter).
Conclusions: ELISA proved useful for measuring and ranking subjects' vitamin D status in Mongolia during summer, but levels were too low in winter to sensitively discriminate between subjects, and ELISA overestimated status in both seasons. These findings have implications for the timing and interpretation, respectively, of vitamin D surveys in highly deficient populations.
Polychlorinated biphenyls (PCBs), known endocrine disruptors, were banned in 1979 but persist in the environment. Previous studies are inconsistent regarding prenatal exposure to PCBs and pregnancy ...outcomes. We investigated associations between prenatal exposure to PCBs and gestational length and birth weight.
In a sample of 600 infants (born between 1960 and 1963) randomly selected from Child Health and Development Studies participants followed through adolescence we measured 11 PCB congeners in maternal post partum sera (within three days of delivery). Length of gestation was computed from the reported first day of the last menstrual period (LMP) and delivery date. Linear regression was used to estimate associations between PCB exposure and gestational age and birth weight, adjusting for potential confounders. PCBs were grouped according to hypothesized biological action (1b (sum of weak phenobarbital inducers), 2b (sum of limited dioxin activity), and 3 (sum of CYP1A and CYP2b inducers)) or degree of ortho- substitution (mono, di, tri). Secondary analyses examined associations between total PCB exposure and exposure to individual congeners.
Each unit increase in mono-ortho substituted PCBs was associated with a 0.30 week decrease (95% confidence interval (CI) -0.59, -0.016), corresponding to a 2.1 (95% CI -4.13, -0.11) day decrease in length of gestation. Similar associations were estimated for di-ortho substituted PCBs, (1.4 day decrease; (95% CI -2.9, 0.1)) and group 3 PCBs (0.84 day decrease; (95% CI -1.8, 0.11). We found similar associations in congener specific analyses and for the sum of congeners.
Our study provides new evidence that PCB exposure shortens length of gestation in humans. This may have public health implications for population exposures.
Soluble Tumor Necrosis Factor Receptor II (sTNFR2) is used as a biomarker to study cardiovascular disease (CVD) in diverse populations. TNF inhibitors (TNFi's) are a common treatment for inflammatory ...conditions. The objective of this study was to examine whether TNFi use impacts measured sTNFR2 levels.
We studied blood samples from a cohort of RA patients with clinical data and high sensitivity-C-reactive protein (hsCRP) measurements. To assess for interference, we tested the entire cohort for the expected positive correlation between sTNFR2 and TNFi using Pearson correlations. We then performed Pearson correlations between sTNFR2 and TNFi and sequentially removed subjects on adalimumab, etanercept, and infliximab; if interference was occurring, no correlation would be observed between hsCRP and sTNFR2, and correlation would be restored by removing subjects on the treatment causing the interference.
We studied 190 subjects, 84.2% female, 73.4% anti-CCP positive. All subjects with sTNFR2 level exceeding measurable level were on etanercept. The expected positive correlation between hsCRP and sTNFR2 was not observed when assessing the entire cohort, r = 0.05, p = 0.51. However, the expected correlation was restored only after excluding subjects on etanercept, r = 0.46, p < 0.0001, and not adalimumab or infliximab. ELISA for sTNFR2 was performed using etanercept only and demonstrated direct binding to sTNFR2.
Our data identified interference between etanercept and the TNFR2 assay. Of the TNFi's, only etanercept has a TNF-binding domain modeled after TNFR2. These data should be considered when designing studies using sTNFR2 in populations where etanercept is a treatment option.
To address whether umbilical cord blood concentrations of sex steroid hormones and the insulin-like growth factor (IGF) axis differ between African-American and White male neonates.
In 2004 and 2005, ...venous cord blood samples were collected from 75 African-American and 38 White male full-term uncomplicated births along with birth weight, placental weight, mother's age and parity, and time of birth. Testosterone, androstanediol glucuronide, estradiol, and sex hormone binding globulin (SHBG) were measured by immunoassay, and IGF-I, IGF-2, and IGF binding protein (BP)-3 by ELISA. Crude and multivariable-adjusted geometric mean concentrations were computed.
Androstanediol glucuronide, estradiol, and SHBG concentrations did not differ by race; however, the molar ratio of testosterone to SHBG was higher in African-American than White male babies after adjustment (P = 0.01). Both before and after adjustment, Whites had higher concentrations of IGF-I (adjusted; White, African-American, 93.1, 71.9 ng/mL), IGF-2 (537.3-474.8 ng/mL), and IGFBP-3 (1,673-1,482 ng/mL) than African-Americans (P < 0.05), although the molar ratio of IGF-I plus IGF-2 to IGFBP-3 did not differ by race.
The higher cord blood testosterone to SHBG ratio in African-American compared with White male babies after taking into account maternal and birth factors is compatible with the hypothesis that differences in androgen levels in utero contribute to their higher prostate cancer risk, although we would have expected crude differences as well. Lower cord blood IGF-I and IGF-2 levels in African-American compared with White male babies are not consistent with the hypothesis that differences in growth factor levels contribute to their higher prostate cancer risk.
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