Downregulation of CX<Subscript>3</Subscript>CR1 ameliorates experimental colitis: evidence for CX<Subscript>3</Subscript>CL1-CX<Subscript>3</Subscript>CR1-mediated immune cell recruitment

Purpose

Inflammatory conditions like inflammatory bowel diseases (IBD) are characterized by increased immune cell infiltration. The chemokine ligand CX₃CL1 and its receptor CX₃CR1 have been shown to be involved in leukocyte adhesion, transendothelial recruitment, and chemotaxis. Therefore, the objective of this study was to describe CX3CL1-CX3CR1-mediated signaling in the induction of immune cell recruitment during experimental murine colitis.

Methods

Acute colitis was induced by dextran sodium sulfate (DSS), and sepsis was induced by injection of lipopolysaccharide (LPS). Serum concentrations of CX₃CR1 and CX₃CL1 were measured by ELISA. Wild-type and CX₃CR1^-/- mice were challenged with DSS, and on day 6, intravital microscopy was performed to monitor colonic leukocyte and platelet recruitment. Intestinal inflammation was assessed by disease activity, histopathology, and neutrophil infiltration.

Results

CX₃CR1 was upregulated in DSS colitis and LPS-induced sepsis. CX₃CR1^-/- mice were protected from disease severity and intestinal injury in DSS colitis, and CX₃CR1 deficiency resulted in reduced rolling of leukocytes and platelets.

Conclusions

In the present study, we provide evidence for a crucial role of CX₃CL1-CX₃CR1 in experimental colitis, in particular for intestinal leukocyte recruitment during murine colitis. Our findings suggest that CX₃CR1 blockade represents a potential therapeutic strategy for treatment of IBD.

     

Associations between physical fitness and HbA<Subscript>1c</Subscript> in type 2 diabetes mellitus

Aim/hypothesis  

In people with type 2 diabetes, exercise improves glucose control (as reflected in HbA_1c) and physical fitness, but it is not clear to what extent these exercise-induced improvements are correlated with one another. We hypothesised that reductions in HbA_1c would be related: (1) to increases in aerobic fitness and strength respectively in patients performing aerobic training or resistance training; and (2) to changes in strength and aerobic fitness in patients performing aerobic and resistance training.     

Haemoglobin A<Subscript>1c</Subscript> levels and subsequent cardiovascular disease in persons without diabetes: a meta-analysis of prospective cohorts

Aims/hypothesis  

The aim of this meta-analysis was to determine the relationship between HbA_1c levels and subsequent cardiovascular outcomes in individuals without diabetes.     

HbA<Subscript>1c</Subscript> is associated with altered expression in blood of cell cycle- and immune response-related genes

Aims/hypothesis

Individuals with type 2 diabetes are heterogeneous in their glycaemic control as tracked by blood HbA_1c levels. Here, we investigated the extent to which gene expression levels in blood reflect current and future HbA_1c levels.

Methods

HbA_1c levels at baseline and 1 and 2 year follow-up were compared with gene expression levels in 391 individuals with type 2 diabetes from the Hoorn Diabetes Care System Cohort (15,564 genes, RNA sequencing). The functions of associated baseline genes were investigated further using pathway enrichment analysis. Using publicly available data, we investigated whether the genes identified are also associated with HbA_1c in the target tissues, muscle and pancreas.

Results

At baseline, 220 genes (1.4%) were associated with baseline HbA_1c. Identified genes were enriched for cell cycle and complement system activation pathways. The association of 15 genes extended to the target tissues, muscle (n = 113) and pancreatic islets (n = 115). At follow-up, expression of 25 genes (0.16%) associated with 1 year HbA_1c and nine genes (0.06%) with 2 year HbA_1c. Five genes overlapped across all time points, and 18 additional genes between baseline and 1 year follow-up. After adjustment for baseline HbA_1c, the number of significant genes at 1 and 2 years markedly decreased, suggesting that gene expression levels in whole blood reflect the current glycaemic state and but not necessarily the future glycaemic state.

Conclusions/interpretation

HbA_1c levels in individuals with type 2 diabetes are associated with expression levels of genes that link to the cell cycle and complement system activation.

     

Translating HbA<Subscript>1c</Subscript> measurements into estimated average glucose values in pregnant women with diabetes

Aims/hypothesis

This study aimed to examine the relationship between average glucose levels, assessed by continuous glucose monitoring (CGM), and HbA_1c levels in pregnant women with diabetes to determine whether calculations of standard estimated average glucose (eAG) levels from HbA_1c measurements are applicable to pregnant women with diabetes.

Methods

CGM data from 117 pregnant women (89 women with type 1 diabetes; 28 women with type 2 diabetes) were analysed. Average glucose levels were calculated from 5–7 day CGM profiles (mean 1275 glucose values per profile) and paired with a corresponding (±1 week) HbA_1c measure. In total, 688 average glucose–HbA_1c pairs were obtained across pregnancy (mean six pairs per participant). Average glucose level was used as the dependent variable in a regression model. Covariates were gestational week, study centre and HbA_1c.

Results

There was a strong association between HbA_1c and average glucose values in pregnancy (coefficient 0.67 [95% CI 0.57, 0.78]), i.e. a 1% (11 mmol/mol) difference in HbA_1c corresponded to a 0.67 mmol/l difference in average glucose. The random effects model that included gestational week as a curvilinear (quadratic) covariate fitted best, allowing calculation of a pregnancy-specific eAG (PeAG). This showed that an HbA_1c of 8.0% (64 mmol/mol) gave a PeAG of 7.4–7.7 mmol/l (depending on gestational week), compared with a standard eAG of 10.2 mmol/l. The PeAG associated with maintaining an HbA_1c level of 6.0% (42 mmol/mol) during pregnancy was between 6.4 and 6.7 mmol/l, depending on gestational week.

Conclusions/interpretation

The HbA_1c–average glucose relationship is altered by pregnancy. Routinely generated standard eAG values do not account for this difference between pregnant and non-pregnant individuals and, thus, should not be used during pregnancy. Instead, the PeAG values deduced in the current study are recommended for antenatal clinical care.

     

Synergistic growth inhibition in HL-60 cells by the combination of acyclic retinoid and vitamin K<Subscript>2</Subscript>

Purpose  

The aim of this study was to assess the effects of acyclic retinoid (ACR) and vitamin K₂ (VK₂) in HL-60 cells.     

Effect of simultaneous vaccination with H1N1 and GAD-alum on GAD<Subscript>65</Subscript>-induced immune response

Aims/hypothesis

A European Phase III trial of GAD formulated with aluminium hydroxide (GAD-alum) failed to reach its primary endpoint (preservation of stimulated C-peptide secretion from baseline to 15 months in type 1 diabetes patients), but subgroup analysis showed a clinical effect when participants from Nordic countries were excluded, raising concern as to whether the mass vaccination of the Swedish and Finnish populations with the Pandemrix influenza vaccine could have influenced the study outcomes. In the current study, we aimed to assess whether Pandemrix vaccination affects the specific immune responses induced by GAD-alum and the C-peptide response.

Methods

In this secondary analysis, we analysed data acquired from the Swedish participants in the Phase III GAD-alum trial who received subcutaneous GAD-alum vaccination (two doses, n = 43; four doses, n = 46) or placebo (n = 48). GAD autoantibodies (GADA) and H1N1 autoantibodies, GAD₆₅-induced cytokine secretion and change in fasting and stimulated C-peptide levels from baseline to 15 months were analysed with respect to the relative time between H1N1 vaccination and the first injection of GAD-alum.

Results

GADA levels at 15 months were associated with the relative time between GAD-alum and Pandemrix administration in participants who received two doses of the GAD-alum vaccine (p = 0.015, r = 0.4). Both in participants treated with two doses and four doses of GAD-alum, GADA levels were higher when the relative time between vaccines was ≥210 days (p < 0.05). In the group that received two doses of GAD-alum, levels of several GAD₆₅-induced cytokines were higher in participants who received the H1N1 vaccination and the first GAD-alum injection at least 150 days apart, and the change in fasting and stimulated C-peptide at 15 months was associated with the relative time between vaccines. Neither of these effects were observed in individuals who received four doses of GAD-alum.

Conclusions/interpretation

In individuals who received two doses of GAD-alum, receiving the Pandemrix vaccine closer to the first GAD-alum injection, i.e. <150 days, seemed to affect both GAD₆₅-induced immune response and C-peptide preservation.

Trial registration:

ClinicalTrials.gov NCT00723411.

     

HbA<Subscript>1c</Subscript>, diabetes and cognitive decline: the English Longitudinal Study of Ageing

Aims/hypothesis

The aim of the study was to evaluate longitudinal associations between HbA_1c levels, diabetes status and subsequent cognitive decline over a 10 year follow-up period.

Methods

Data from wave 2 (2004–2005) to wave 7 (2014–2015) of the English Longitudinal Study of Ageing (ELSA) were analysed. Cognitive function was assessed at baseline (wave 2) and reassessed every 2 years at waves 3–7. Linear mixed models were used to evaluate longitudinal associations.

Results

The study comprised 5189 participants (55.1% women, mean age 65.6?±?9.4 years) with baseline HbA_1c levels ranging from 15.9 to 126.3 mmol/mol (3.6–13.7%). The mean follow-up duration was 8.1?±?2.8 years and the mean number of cognitive assessments was 4.9?±?1.5. A 1 mmol/mol increment in HbA_1c was significantly associated with an increased rate of decline in global cognitive z scores (?0.0009 SD/year, 95% CI ?0.0014, ?0.0003), memory z scores (?0.0005 SD/year, 95% CI ?0.0009, ?0.0001) and executive function z scores (?0.0008 SD/year, 95% CI ?0.0013, ?0.0004) after adjustment for baseline age, sex, total cholesterol, HDL-cholesterol, triacylglycerol, high-sensitivity C-reactive protein, BMI, education, marital status, depressive symptoms, current smoking, alcohol consumption, hypertension, CHD, stroke, chronic lung disease and cancer. Compared with participants with normoglycaemia, the multivariable-adjusted rate of global cognitive decline associated with prediabetes and diabetes was increased by ?0.012 SD/year (95% CI ?0.022, ?0.002) and ?0.031 SD/year (95% CI ?0.046, ?0.015), respectively (p for trend <0.001). Similarly, memory, executive function and orientation z scores showed an increased rate of cognitive decline with diabetes.

Conclusions/interpretation

Significant longitudinal associations between HbA_1c levels, diabetes status and long-term cognitive decline were observed in this study. Future studies are required to determine the effects of maintaining optimal glucose control on the rate of cognitive decline in people with diabetes.

     

Bone Marrow Mononuclear Cell Transplantation Improves Survival and Induces Hepatocyte Proliferation in Rats after CCl<Subscript>4</Subscript> Acute Liver Damage

Aim  

The aim of this research was to evaluate the effects of bone marrow mononuclear cell (BMC) transplantation in rats with toxic acute liver damage induced by carbon tetrachloride (CCl₄).     

Lafutidine,a Protective H<Subscript>2</Subscript> Receptor Antagonist,Enhances Mucosal Defense in Rat Esophagus

Background  

Luminal acid or CO₂ induces a hyperemic response in the esophagus, via activation of acid sensors on capsaicin-sensitive afferent nerves (CSAN). Since disruption of the hyperemic response to luminal CO₂ acidifies the interstitium of the esophageal mucosa, the hyperemic response may maintain interstitial pH (pH_int). We hypothesized that acid-related hyperemia maintains pH_int, preventing acid-induced injury in the esophageal mucosa.     

Effects of Ischemia and Reperfusion on P2X<Subscript>2</Subscript> Receptor Expressing Neurons of the Rat Ileum Enteric Nervous System

Purpose  

We investigated the effects of ischemia/reperfusion in the intestine (I/R-i) on purine receptor P2X₂-immunoreactive (IR) neurons of the rat ileum.     

Dimerization of AT<Subscript>2</Subscript> and Mas Receptors in Control of Blood Pressure

Purpose of Review

Angiotensin type 2 receptor (AT₂R) and receptor Mas (MasR) are part of the “protective arm” of the renin angiotensin system. Gene and pharmacological manipulation studies reveal that AT₂R and MasR are involved in natriuretic, vasodilatory, and anti-inflammatory responses and in lowering blood pressure in various animal models under normal and pathological conditions such as salt-sensitive hypertension, obesity, and diabetes. The scope of this review is to discuss co-localization and heterodimerization as potential molecular mechanisms of AT₂R- and MasR-mediated functions including antihypertensive activities.

Recent Findings

Accumulating evidences show that AT₂R and MasR are co-localized, make a heterodimer, and are functionally interdependent in producing their physiological responses. Moreover, ang-(1-7) preferably may be an AT₁R-biased agonist while acting as a MasR agonist.

Summary

The physical interactions of AT₂R and MasR appear to be an important mechanism by which these receptors are involved in blood pressure regulation and antihypertensive activity. Whether heteromers of these receptors influence affinity or efficacy of endogenous or synthetic agonists remains a question to be considered.

     

Lipoprotein-associated phospholipase A<Subscript>2</Subscript> and future risk of subclinical disease and cardiovascular events in individuals with type 2 diabetes: the Cardiovascular Health Study

Aims/hypothesis  

Type 2 diabetes is an established risk factor for cardiovascular disease (CVD). This increased risk may be due in part to the increased levels of inflammatory factors associated with diabetes. Lipoprotein-associated phospholipase A₂ (Lp-PLA₂) is a risk marker for CVD and has pro-inflammatory effects in atherosclerotic plaques. We therefore sought to determine whether Lp-PLA₂ levels partially explain the greater prevalence of subclinical CVD and greater incidence of CVD outcomes associated with type 2 diabetes in the Cardiovascular Health Study.     

Effets cardiovasculaires des auto-anticorps anti-récepteurs béta1 et béta3-adrénergiques chez le rat Lewis

Purpose

To analyze vascular reactivity changes in response to immunization protocols with antigens corresponding to the second extracellular loop of –β₃ and –β_1 and 3 adrenergic receptors (AR).

Methods

Lewis rats were immunized for 3 months with peptidic sequences corresponding to the second extracellular loop of β₃–AR or β_1 and 3–AR. Specific β₃–AR antibodies were characterized by Elisa and purified using “Proteus Protein G” kit. Their functionality were tested in rabbit isolated ventricular cardiomyocytes. Aortic and mesenteric artery rings isolated from control or immunized rats were mounted in organ baths and precontracted with phenylephrine. Then, relaxant curves were established.

Results

SR58611A (10 nM), a preferential β₃–AR agonist and purified β₃–AR antibodies (25 μg/mL) induced a decrease of cell shortening (−39.56 ± 4.4% [n = 11] and −18.45 ± 3.9% [n = 10] respectively) in isolated cardiomyocytes. This decrease was significantly inhibited when the cardiomyocytes were pre-incubated with the L–748337 (1 μM), a selective β₃–AR antagonist (P < 0.05). In contrast with what was observed in rats immunized against the β₁–AR, vasorelaxations induced by acetylcholine and SR58611A in both aorta and mesenteric arteries were unaltered in rats immunized against the β₃–AR and β_1 and 3–AR.

Conclusion

These results show, for the first time, that β₃–AR antibodies induced a β₃–AR agonist-like activity. They would not have a vascular pathogenic action but would offset the endothelial dysfunction caused by β₁–AR antibodies.     

Size and shape of the associations of glucose,HbA<Subscript>1c</Subscript>, insulin and HOMA-IR with incident type 2 diabetes: the Hoorn Study

Aims/hypothesis

Glycaemic markers and fasting insulin are frequently measured outcomes of intervention studies. To extrapolate accurately the impact of interventions on the risk of diabetes incidence, we investigated the size and shape of the associations of fasting plasma glucose (FPG), 2 h post-load glucose (2hPG), HbA_1c, fasting insulin and HOMA-IR with incident type 2 diabetes mellitus.

Methods

The study population included 1349 participants aged 50–75 years without diabetes at baseline (1989) from a population-based cohort in Hoorn, the Netherlands. Incident type 2 diabetes was defined by the WHO 2011 criteria or known diabetes at follow-up. Logistic regression models were used to determine the associations of the glycaemic markers, fasting insulin and HOMA-IR with incident type 2 diabetes. Restricted cubic spline logistic regressions were conducted to investigate the shape of the associations.

Results

After a mean follow-up duration of 6.4 (SD 0.5) years, 152 participants developed diabetes (11.3%); the majority were screen detected by high FPG. In multivariate adjusted models, ORs (95% CI) for incident type 2 diabetes for the highest quintile in comparison with the lowest quintile were 9.0 (4.4, 18.5) for FPG, 6.1 (2.9, 12.7) for 2hPG, 3.8 (2.0, 7.2) for HbA_1c, 1.9 (0.9, 3.6) for fasting insulin and 2.8 (1.4, 5.6) for HOMA-IR. The associations of FPG and HbA_1c with incident diabetes were non-linear, rising more steeply at higher values.

Conclusions/interpretation

FPG was most strongly associated with incident diabetes, followed by 2hPG, HbA_1c, HOMA-IR and fasting insulin. The strong association with FPG is probably because FPG is the most frequent marker for diabetes diagnosis. Non-linearity of associations between glycaemic markers and incident type 2 diabetes should be taken into account when estimating future risk of type 2 diabetes based on glycaemic markers.

     

CO<Subscript>2</Subscript> embolism can complicate transanal total mesorectal excision

Background

Carbon dioxide (CO₂) embolism is a rare but potentially devastating complication of minimally invasive abdominal and retroperitoneal surgery. Characterized by a decrease in end-tidal CO₂ (ETCO₂) and oxygen saturation (SpO₂), CO₂ emboli can cause rapid intraoperative hypotension and cardiovascular collapse. Transanal total mesorectal excision (taTME) is a novel surgical approach for rectal resection, which requires high flow CO₂ insufflation in a low volume operative field. In this setting, the incidence of CO₂ embolism is unknown; we evaluate three cases of intraoperative CO₂ embolism that occurred during the transanal portion of the TME dissection.

Methods

All taTME cases from December 2014 to March 2018 at a single institution were reviewed. Cases of CO₂ embolism were identified intraoperatively and characterized using the operative reports and anesthesia records. The transanal/pelvic insufflation included a targeted pressure of 15 mm Hg, high flow and high smoke evacuation. Physiologic derangements and management of these instances were analyzed. The postoperative course was evaluated and any complications were noted.

Results

A total of 80 taTME were performed for benign and malignant disease. Three patients (4%) developed intraoperative evidence of CO₂ embolism. Each instance occurred during the transanal portion of the dissection. Physiologic changes were marked by abrupt decrease in end-tidal ETCO₂, SpO₂, and blood pressure (BP). Management included immediate release of pneumopelvis, hemodynamic support with crystalloid or vasopressors, and placement of the patient in the Trendelenburg position with left side down. Within 10 min of the acute event, all patients had return of ETCO₂, SpO₂, and BP to pre-event levels. There were no intraoperative or postoperative sequelae including arrhythmia, myocardial infarction, stroke or death. No cases required conversion to open.

Conclusions

During taTME, rare CO₂ emboli may occur in the setting of venous bleeding during pneumopelvis, causing sudden, transient cardiovascular instability. Immediate recognition of rapid decrease in ETCO₂, SpO₂, and BP should be followed by desufflation of pneumopelvis, patient positioning in Trendelenburg and left lateral decubitus, and hemodynamic support. Increased awareness of this potential complication and maintaining a high index of suspicion will lead to preparedness of the anesthesia and surgery teams.

     

Associations of HbA1c and educational level with risk of cardiovascular events in 32 871 drug‐treated patients with Type 2 diabetes: a cohort study in primary care

Aims

To explore the association of HbA_1c and educational level with risk of cardiovascular events and mortality in patients with Type 2 diabetes.

Methods

A cohort of 32 871 patients with Type 2 diabetes aged 35 years and older identified by extracting data from electronic patient records for all patients who had a diagnosis of Type 2 diabetes and had glucose‐lowering agents prescribed between 1999 and 2009 at 84 primary care centres in Sweden. Associations of mean HbA_1c levels and educational level with risks of cardiovascular events and all‐cause mortality were analysed.

Results

The associations of HbA_1c with risk of all‐cause and cardiovascular mortality were J‐shaped, with the lowest risk observed for cardiovascular mortality at an HbA_1c level of 51 mmol/mol (6.8%) for subjects on oral agents and 56 mmol/mol (7.3%) in insulin‐treated patients. The lowest risk observed for all‐cause mortality was at an HbA_1c level of 51 mmol/mol (6.8%) for subjects on oral agents and 56 mmol/mol (7.3%) in insulin‐treated patients. There was an increased risk for cardiovascular death [hazard ratio 1.6 (1.2–2.1), P = 0.0008] at the lowest HbA_1c decile for subjects in the low education category. For subjects with higher education there was no evident J curve for cardiovascular death [hazard ratio 1.2 (0.8–1.6), P = 0.3873].

Conclusions

Our results lend support to the recent American Diabetes Association/ European Association for the Study of Diabetes position statement that emphasizes the importance of additional factors, including the propensity for hypoglycaemia, which should influence HbA_1c targets and treatment choices for individual patients. (Clinical Trials Registry No; NCT 01121315)     

Defects in beta cell Ca<Superscript>2+</Superscript> signalling,glucose metabolism and insulin secretion in a murine model of K<Subscript>ATP</Subscript> channel-induced neonatal diabetes mellitus

Aims/hypothesis  

Mutations that render ATP-sensitive potassium (K_ATP) channels insensitive to ATP inhibition cause neonatal diabetes mellitus. In mice, these mutations cause insulin secretion to be lost initially and, as the disease progresses, beta cell mass and insulin content also disappear. We investigated whether defects in calcium signalling alone are sufficient to explain short-term and long-term islet dysfunction.     

Preconditioning by Levosimendan is Mediated by Activation of Mitochondrial Ca<Superscript>2+</Superscript>-Sensitive Potassium (mBK<Subscript>Ca</Subscript>) Channels

Purpose

Activation of mitochondrial large-conductance Ca²⁺-sensitive potassium (mBK_Ca)-channels is a crucial step for cardioprotection by preconditioning. Whether activation of these channels is involved in levosimendan-induced preconditioning is unknown. We investigated if cardioprotection by levosimendan requires activation of mBK_Ca-channels in the rat heart in vitro.

Methods

In a prospective blinded experimental laboratory investigation, hearts of male Wistar rats were randomized and placed on a Langendorff system, perfused with Krebs-Henseleit buffer at a constant pressure of 80 mmHg. All hearts were subjected to 33 min of global ischemia and 60 min of reperfusion. Before ischemia, hearts were perfused with different concentrations of levosimendan (0.03–1 μM) for determination of a dose-effect curve. In a second set of experiments, 0.3 μM levosimendan was administered in combination with the mBK_Ca-channel inhibitor paxilline (1 μM). Infarct size was determined by TTC staining.

Results

In control, animal’s infarct size was 58?±?7%. Levosimendan at a concentration of 0.3 μM reduced infarct size to 30?±?7% (P?<?0.05 vs. control). Higher concentrations with 1 μM levosimendan did not confer stronger protection. Paxilline completely blocked levosimendan-induced cardioprotection while paxilline alone had no effect on infarct size.

Conclusions

This study shows that activation of mBK_Ca-channels plays a pivotal role in levosimendan-induced preconditioning.

     

Lp-PLA<Subscript>2</Subscript> activity is associated with increased risk of diabetic retinopathy: a longitudinal disease progression study

Aims/hypothesis

The aim of the study was to examine the association between lipoprotein-associated phospholipase A₂ (Lp-PLA₂) activity levels and incident diabetic retinopathy and change in retinopathy grade.

Methods

This was a cohort study of diabetic participants with serum collected at baseline and routinely collected diabetic retinal screening data. Participants with type 2 diabetes from the GoDARTS (Genetics of Diabetes Audit and Research in Tayside Scotland) cohort were used. This cohort is composed of individuals of white Scottish ancestry from the Tayside region of Scotland. Survival analysis accounting for informative censoring by modelling death as a competing risk was performed for the development of incident diabetic retinopathy from a disease-free state in a 3 year follow-up period (n?=?1364) by stratified Lp-PLA₂ activity levels (in quartiles). The same analysis was performed for transitions to more severe grades.

Results

The hazard of developing incident diabetic retinopathy was 2.08 times higher (95% CI 1.64, 2.63) for the highest quartile of Lp-PLA₂ activity compared with the lowest. Higher Lp-PLA₂ activity levels were associated with a significantly increased risk for transitions to all grades. The hazards of developing observable (or more severe) and referable (or more severe) retinopathy were 2.82 (95% CI 1.71, 4.65) and 1.87 (95% CI 1.26, 2.77) times higher for the highest quartile of Lp-PLA₂ activity compared with the lowest, respectively.

Conclusions/interpretation

Higher Lp-PLA₂ levels are associated with increased risk of death and the development of incident diabetic retinopathy, as well as transitions to more severe grades of diabetic retinopathy. These associations are independent of calculated LDL-cholesterol and other traditional risk factors. Further, this biomarker study shows that the association is temporally sensitive to the proximity of the event to measurement of Lp-PLA_2.