Higher spermidine intake is linked to lower mortality: a prospective population-based study.

Background: Spermidine administration is linked to increased survival in several animal models. Objective: The aim of this study was to test the potential association between spermidine content in diet and mortality in humans. Design: This prospective community-based cohort study included 829 participants aged 45-84 y, 49.9% of whom were male. Diet was assessed by repeated dietitian-administered validated food-frequency questionnaires (2540 assessments) in 1995, 2000, 2005, and 2010. During follow-up between 1995 and 2015, 341 deaths occurred. Results: All-cause mortality (deaths per 1000 person-years) decreased across thirds of increasing spermidine intake from 40.5 (95% CI: 36.1, 44.7) to 23.7 (95% CI: 20.0, 27.0) and 15.1 (95% CI: 12.6, 17.8), corresponding to an age-, sex- and caloric intake-adjusted 20-y cumulative mortality incidence of 0.48 (95% CI: 0.45, 0.51), 0.41 (95% CI: 0.38, 0.45), and 0.38 (95% CI: 0.34, 0.41), respectively. The age-, sex- and caloric ratio-adjusted HR for all-cause death per 1-SD higher spermidine intake was 0.74 (95% CI: 0.66, 0.83; P < 0.001). Further adjustment for lifestyle factors, established predictors of mortality, and other dietary features yielded an HR of 0.76 (95% CI: 0.67, 0.86; P < 0.001). The association was consistent in subgroups, robust against unmeasured confounding, and independently validated in the Salzburg Atherosclerosis Prevention Program in Subjects at High Individual Risk (SAPHIR) Study (age-, sex-, and caloric ratio-adjusted HR per 1-SD higher spermidine intake: 0.71; 95% CI: 0.53, 0.95; P = 0.019). The difference in mortality risk between the top and bottom third of spermidine intakes was similar to that associated with a 5.7-y (95% CI: 3.6, 8.1 y) younger age. Conclusion: Our findings lend epidemiologic support to the concept that nutrition rich in spermidine is linked to increased survival in humans. This trial was registered at www.clinicaltrials.gov as NCT03378843.

Very-Low-Density Lipoprotein-Associated Apolipoproteins Predict Cardiovascular Events and Are Lowered by Inhibition of APOC-III.

BACKGROUND: Routine apolipoprotein (apo) measurements for cardiovascular disease (CVD) are restricted to apoA-I and apoB. Here, the authors measured an unprecedented range of apolipoproteins in a prospective, population-based study and relate their plasma levels to risk of CVD. OBJECTIVES: This study sought to measure apolipoproteins directly by mass spectrometry and compare their associations with incident CVD and to obtain a system-level understanding of the correlations of apolipoproteins with the plasma lipidome and proteome. METHODS: Associations of 13 apolipoproteins, 135 lipid species, and 211 other plasma proteins with incident CVD (91 events), defined as stroke, myocardial infarction, or sudden cardiac death, were assessed prospectively over a 10-year period in the Bruneck Study (N = 688) using multiple-reaction monitoring mass spectrometry. Changes in apolipoprotein and lipid levels following treatment with volanesorsen, a second-generation antisense drug targeting apoC-III, were determined in 2 human intervention trials, one of which was randomized. RESULTS: The apolipoproteins most significantly associated with incident CVD were apoC-II (hazard ratio per 1 SD [HR/SD]: 1.40; 95% confidence interval [CI]: 1.17 to 1.67), apoC-III (HR/SD: 1.38; 95% CI: 1.17 to 1.63), and apoE (HR/SD: 1.31; 95% CI: 1.13 to 1.52). Associations were independent of high-density lipoprotein (HDL) and non-HDL cholesterol, and extended to stroke and myocardial infarction. Lipidomic and proteomic profiles implicated these 3 very-low-density lipoprotein (VLDL)-associated apolipoproteins in de novo lipogenesis, glucose metabolism, complement activation, blood coagulation, and inflammation. Notably, apoC-II/apoC-III/apoE correlated with a pattern of lipid species previously linked to CVD risk. ApoC-III inhibition by volanesorsen reduced plasma levels of apoC-II, apoC-III, triacylglycerols, and diacylglycerols, and increased apoA-I, apoA-II, and apoM (all p < 0.05 vs. placebo) without affecting apoB-100 (p = 0.73). CONCLUSIONS: The strong associations of VLDL-associated apolipoproteins with incident CVD in the general community support the concept of targeting triacylglycerol-rich lipoproteins to reduce risk of CVD.

Haptoglobin 2-2 genotype is not associated with cardiovascular risk in subjects with elevated glycohemoglobin-results from the Bruneck Study.

BACKGROUND: Haptoglobin (Hp) is an abundant plasma protein with antioxidant properties. The Hp 2-2 genotype has previously been linked to coronary heart disease risk in individuals with elevated glycosylated hemoglobin (HbA1c). We investigated the association of Hp and HbA1c with cardiovascular disease (CVD) in the longitudinal, population-based Bruneck Study. METHODS AND RESULTS: Hp genotype was determined by polymerase chain reaction according to standard procedures and HbA1c concentration by a Diabetes Control and Complications Trial-aligned assay. HbA1c was measured in 1995, 2000, and 2005. Occurrence of the combined CVD endpoint of myocardial infarction or stroke was recorded between 1995 and 2010. Outcome analyses employed the Cox proportional hazards model with HbA1c category as time-varying covariate. At baseline in 1995, 806 subjects (male sex, 49.3%; age, mean ± standard deviation, 62.70 ± 11.08 years) were included. During follow-up, 123 subjects experienced at least 1 CVD event (48 suffered myocardial infarction, 68 stroke, and 7 both). Among subjects with HbA1c ≥ 6.5% (≥ 48 mmol/mol), those with the Hp 2-2 genotype did not show an elevated risk of incident CVD compared with those with other genotypes (age- and sex-adjusted hazard ratio [95% CI], 0.47 [0.19, 1.13], P=0.092) and a null association was also observed in subjects with HbA1c<6.5% (1.10 [0.75, 1.62], P=0.629) (P for interaction=0.082). CONCLUSIONS: Subjects with the Hp 2-2 genotype and elevated HbA1c compared with subjects with other Hp genotypes and elevated HbA1c did not show increased CVD risk.

Blockade of receptor activator of nuclear factor-κB (RANKL) signaling improves hepatic insulin resistance and prevents development of diabetes mellitus.

Hepatic insulin resistance is a driving force in the pathogenesis of type 2 diabetes mellitus (T2DM) and is tightly coupled with excessive storage of fat and the ensuing inflammation within the liver. There is compelling evidence that activation of the transcription factor nuclear factor-κB (NF-κB) and downstream inflammatory signaling pathways systemically and in the liver are key events in the etiology of hepatic insulin resistance and ß-cell dysfunction, although the molecular mechanisms involved are incompletely understood. We here test the hypothesis that receptor activator of NF-κB ligand (RANKL), a prototypic activator of NF-κB, contributes to this process using both an epidemiological and experimental approach. In the prospective population-based Bruneck Study, a high serum concentration of soluble RANKL emerged as a significant (P<0.001) and independent risk predictor of T2DM manifestation. In close agreement, systemic or hepatic blockage of RANKL signaling in genetic and nutritional mouse models of T2DM resulted in a marked improvement of hepatic insulin sensitivity and amelioration or even normalization of plasma glucose concentrations and glucose tolerance. Overall, this study provides evidence for a role of RANKL signaling in the pathogenesis of T2DM. If so, translation to the clinic may be feasible given current pharmacological strategies to lower RANKL activity to treat osteoporosis.

Impact of reference category and number of traits in the cluster on risk of coronary heart disease in metabolic syndrome: prospective data from the Bruneck study.

BACKGROUND: We assessed the risk of coronary heart disease (CHD) in subjects with metabolic syndrome using different reference categories and focusing on the number of traits in the cluster. METHODS: For 15 years, we followed 840 subjects from the general population living in Bruneck, northeastern Italy, aged 40-79 years, without CHD at baseline. Metabolic syndrome was diagnosed at baseline using American Heart Association/National Heart, Lung, and Blood Institute criteria. Subjects with the syndrome were compared to subjects without, as well as to subjects without any metabolic abnormality, using Cox models adjusted for sex, age, smoking, and low-density lipoprotein-cholesterol. There were 89 incident CHD cases. RESULTS: In subjects with the metabolic syndrome, the risk of CHD was1.5-fold higher when subjects without the syndrome were the reference category. CHD risk, however, was 12.5-fold higher (95% confidence interval [CI], 1.7-92.7, P=0.014) when subjects without any metabolic abnormality composed the reference category. As compared to subjects with no abnormalities (who had a trivial number of CHD events), the risk increased from subjects with one (hazard ratio 7.6, 95% CI 1.0-56.5, P=0.047) to those with 2, 3, and 4/5 abnormalities (11.6, 1.6-84.9, P=0.016; 12.9, 1.7-96.0, P=0.013; and 10.1, 1.3-79.2, P=0.028), respectively. CONCLUSIONS: When compared to the reference category of people without any metabolic abnormality, those with the metabolic syndrome had high cardiovascular risk. However, in the Bruneck population, the risk of CHD seems to be similar in subjects having two or three to five clinical features of metabolic syndrome. Therefore, the clinical utility of identifying subjects with the syndrome using current diagnostic criteria remains uncertain and might be the focus of further specific studies.

Plasma microRNA profiling reveals loss of endothelial miR-126 and other microRNAs in type 2 diabetes.

RATIONALE: MicroRNAs (miRNAs) have been implicated in the epigenetic regulation of key metabolic, inflammatory, and antiangiogenic pathways in type 2 diabetes (DM) and may contribute to common disease complications. OBJECTIVE: In this study, we explore plasma miRNA profiles in patients with DM. METHODS AND RESULTS: Total RNA was extracted from plasma samples of the prospective population-based Bruneck study. A total of 13 candidate miRNAs identified by microarray screening and miRNA network inference were quantified by quantitative PCR in all diabetic patients of the Bruneck study and age- and sex-matched controls (1995 evaluation, n=80 each). Quantitative PCR assessment revealed lower plasma levels of miR-20b, miR-21, miR-24, miR-15a, miR-126, miR-191, miR-197, miR-223, miR-320, and miR-486 in prevalent DM, but a modest increase of miR-28-3p. Findings emerged as robust in multivariable analysis and were independent of the standardization procedure applied. For endothelial miR-126, results were confirmed in the entire Bruneck cohort (n=822) in univariate (odds ratio [95% confidence interval], 0.38 [0.26 to 0.55]; P=2.72 × 10(-7)) and multivariate analyses (0.57 [0.37 to 0.86]; P=0.0082). Importantly, reduced miR-15a, miR-29b, miR-126, miR-223, and elevated miR-28-3p levels antedated the manifestation of disease. Most differences in miRNA levels were replicated in plasma obtained from hyperglycemic Lep(ob) mice. High glucose concentrations reduced the miR-126 content of endothelial apoptotic bodies. Similarly in patients with DM, the reduction of miR-126 was confined to circulating vesicles in plasma. CONCLUSIONS: We reveal a plasma miRNA signature for DM that includes loss of endothelial miR-126. These findings might explain the impaired peripheral angiogenic signaling in patients with DM.

Telomere length and risk of incident cancer and cancer mortality.

CONTEXT: Telomeres are essential to preserve the integrity of the genome. Critically short telomeres lead to replicative cell senescence and chromosomal instability and may thereby increase cancer risk. OBJECTIVE: To determine the association between baseline telomere length and incident cancer and cancer mortality. DESIGN, SETTING, AND PARTICIPANTS: Leukocyte telomere length was measured by quantitative polymerase chain reaction in 787 participants free of cancer at baseline in 1995 from the prospective, population-based Bruneck Study in Italy. MAIN OUTCOME MEASURES: Incident cancer and cancer mortality over a follow-up period of 10 years (1995-2005 with a follow-up rate of 100%). RESULTS: A total of 92 of 787 participants (11.7%) developed cancer (incidence rate, 13.3 per 1000 person-years). Short telomere length at baseline was associated with incident cancer independently of standard cancer risk factors (multivariable hazard ratio [HR] per 1-SD decrease in log(e)-transformed telomere length, 1.60; 95% confidence interval [CI], 1.30-1.98; P < .001). Compared with participants in the longest telomere length group, the multivariable HR for incident cancer was 2.15 (95% CI, 1.12-4.14) in the middle length group and 3.11 (95% CI, 1.65-5.84) in the shortest length group (P < .001). Incidence rates were 5.1 (95% CI, 2.9-8.7) per 1000 person-years in the longest telomere length group, 14.2 (95% CI, 10.0-20.1) per 1000 person-years in the middle length group, and 22.5 (95% CI, 16.9-29.9) per 1000 person-years in the shortest length group. The association equally applied to men and women and emerged as robust under a variety of circumstances. Furthermore, short telomere length was associated with cancer mortality (multivariable HR per 1-SD decrease in log(e)-transformed telomere length, 2.13; 95% CI, 1.58-2.86; P < .001) and individual cancer subtypes with a high fatality rate. CONCLUSION: In this study population, there was a statistically significant inverse relationship between telomere length and both cancer incidence and mortality.

Cellular aging reflected by leukocyte telomere length predicts advanced atherosclerosis and cardiovascular disease risk.

OBJECTIVE: To determine the association between leukocyte telomere length (TL) and atherosclerosis and its clinical sequelae stroke and myocardial infarction. METHODS AND RESULTS: Within the scope of the prospective population-based Bruneck Study, leukocyte TL was measured by quantitative polymerase chain reaction in 800 women and men aged 45 to 84 years (in 1995). The manifestation of cardiovascular disease (CVD) (1995-2005) and the progression of atherosclerosis (1995-2000) were carefully assessed. The TL was shorter in men than in women (age-adjusted mean [95% CI], 1.41 [1.33 to 1.49] versus 1.55 [1.47 to 1.62]; P=0.02) and inversely correlated to age (r=-0.22, P<0.001) and family history of CVD (P=0.03). Participants with CVD events during follow-up (n=88) had significantly shorter telomeres (age- and sex-adjusted mean [95% CI], 1.25 [1.08 to 1.42] versus 1.51 [1.45 to 1.57]; P<0.001). In multivariable Cox models, baseline TL emerged as a significant and independent risk predictor for the composite CVD end point and its individual components (myocardial infarction and stroke); however, this was not the case for de novo stable angina and intermittent claudication. Subjects in the top and bottom TL tertile group differed in their CVD risk by a factor of 2.72 (95% CI, 1.41 to 5.28), which is the risk ratio attributable to a 13.9-year difference in chronological age. Remarkably, in our atherosclerosis progression model, TL was strongly associated with advanced, but not early, atherogenesis. All findings were consistent in women and men. CONCLUSIONS: Our findings indicate a differential role of telomere shortening in the various stages of atherosclerosis, with preferential involvement in advanced vessel pathology and acute vascular syndromes.

Single nucleotide polymorphism on chromosome 9p21 and endothelial progenitor cells in a general population cohort.

OBJECTIVE: Recent studies have revealed that sequence variation on chromosome 9p21 is a major genetic determinant for coronary heart disease and stroke, however, the underlying mechanism remains unknown. This genomic region contains the gene encoding cyclin-dependent kinase 2A, a regulator of proliferation and differentiation of endothelial progenitor cell (EPC) which has been implicated in vascular repair and protection against cardiovascular disease. We investigated whether carriers and non-carriers of the chromosome 9p21 variation differed in their number and function of EPCs. METHODS: We genotyped a cohort of 769 individuals (who participated in the Bruneck Study) for the single nucleotide polymorphism rs1333049 on chromosome 9p21 and determined circulating EPC numbers and EPC colony formation units (n=538 and 512, respectively). RESULTS: There was no evidence of reduced EPC numbers or colony formation units in carriers of the chromosome 9p21 risk variant. On the contrary, circulating EPC numbers and colony formation units tended to be higher in carriers of the variant (p=0.189 for EPC numbers and p=0.190 for EPC colony formation units). CONCLUSION: These results indicate that the chromosome 9p21 variant does not influence the risk of coronary heart disease and stroke through an effect on circulating EPCs.

Vascular cell adhesion molecule 1 as a predictor of severe osteoarthritis of the hip and knee joints.

OBJECTIVE: Osteoarthritis (OA) is a leading cause of pain and physical disability in middle-aged and older individuals. We undertook this study to determine predictors of the development of severe OA, apart from age and overweight. METHODS: Joint replacement surgery due to severe hip or knee OA was recorded over a 15-year period in the prospective Bruneck cohort study. Demographic characteristics and lifestyle and biochemical variables, including the level of soluble vascular cell adhesion molecule 1 (VCAM-1), were assessed at the 1990 baseline visit and tested as predictors of joint replacement surgery. RESULTS: Between 1990 and 2005, hip or knee joint replacement due to OA was performed in 60 subjects. VCAM-1 level emerged as a highly significant predictor of the risk of joint replacement surgery. Intervention rates were 1.9, 4.2, and 10.1 per 1,000 person-years in the first, second, and third tertiles, of the VCAM-1 level, respectively. In multivariable logistic regression analysis, the adjusted relative risk of joint replacement surgery in the highest versus the lowest tertile group of VCAM-1 level was 3.9 (95% confidence interval 1.7-8.7) (P<0.001). Findings were robust in various sensitivity analyses and were consistent in subgroups. Addition of the VCAM-1 level to a risk model already including age, sex, and body mass index resulted in significant gains in model discrimination (C statistic) and calibration and in more accurate risk classification of individual participants. CONCLUSION: The level of soluble VCAM-1 emerged as a strong and independent predictor of the risk of hip and knee joint replacement due to severe OA. If our findings can be reproduced in other epidemiologic cohorts, they will assist in routine risk classification and will contribute to a better understanding of the etiology of OA.

SDF1 gene variation is associated with circulating SDF1alpha level and endothelial progenitor cell number: the Bruneck Study.

BACKGROUND: Stromal cell-derived factor-1 (SDF1) and its receptor CXC chemokine receptor 4 (CXCR4) play a critical role in progenitor cell homing, mobilization and differentiation. It would be interesting to assess the predictive value of SDF-1alpha level for EPC number, and to ascertain whether there is a relationship between SDF1 gene variation, plasma SDF-1alpha level, and the number and function of circulating EPCs. We also tested whether EPC number and function was related to CXCR4 gene variation. METHODOLOGY AND PRINCIPAL FINDINGS: We genotyped a cohort of individuals who participated in the Bruneck Study for single nucleotide polymorphisms (SNPs) in the SDF1 and CXCR4 genes, and measured blood SDF1alpha level as well as EPC number and function. SDF1alpha levels were correlated with age, gender, alcohol consumption, circulating reticulocyte numbers, and concentrations of matrix metalloproteinase-9, C-reactive protein, cystatin C, fibrinogen and homocytein. In blood samples taken in 2005, EPC number was inversely associated with SDF1alpha level (p<0.001). EPC number in 2005 was also inversely associated with SDF1alpha level in 2000 (p = 0.009), suggesting a predictive value of plasma SDF1alpha level for EPC number. There was an association between the SDF1 gene rs2297630 SNP A/A genotype, increased SDF1alpha level (p = 0.002) and lower EPC number (p = 0.006). CONCLUSIONS: Our data indicate that a SDF1 gene variation (rs2297630) has an influence on SDF1alpha level and circulating EPC number, and that plasma SDF1alpha level is a predictor of EPC number.

Endothelial progenitor cells, cardiovascular risk factors, cytokine levels and atherosclerosis--results from a large population-based study.

BACKGROUND: EPC number and functionality are assumed to reflect the endogenous vascular repair capacity with the EPC pool declining in higher ages and being exhausted by unfavorable life-style and risk factors. This intriguing and clinically highly relevant concept, however, has so far been derived from small case-control studies and patient series. METHODOLOGY AND PRINCIPLE FINDINGS: In the population-based Bruneck Study EPC number and EPC-colony forming units (EPC-CFU) were assessed as part of the fourth follow-up evaluation (2005) in 571 and 542 subjects, respectively. EPC number declined with age (p = 0.013), was significantly lower in women (p = 0.006) and higher in subjects on statin, hormone replacement or ACE inhibitor/angiotensin-receptor blockers, and correlated positively with moderate alcohol consumption. Unexpectedly, a positive relation between EPC number and several vascular risk factors emerged. In a step forward multivariate linear regression analysis EPC number was independently related with SDF1alpha, MMP-9, triglycerides, alcohol consumption, and Hba1c. EPC-CFU in turn was related to SDF1alpha and diastolic blood pressure. Moreover, EPC number showed a significant positive association with the Framingham risk score (P = 0.001). Finally, there was an inverse association between EPC number and common carotid artery intima-media thickness (p = 0.02) and the carotid artery atherosclerosis score (p = 0.059). CONCLUSIONS: Our population-based data confirm the decline of EPC number with advancing age and lend first epidemiological support to a role of SDF-1alpha and MMP9 in EPC differentiation, mobilization and homing, but are conflict with the view that EPC number is unfavorably affected by cardiovascular risk factors. EPC number increases with the cardiovascular risk estimated by the Framingham risk score (FRS), which in the absence of similar changes for EPC-CFU. Finally, we demonstrate a significant inverse association between EPC number and extent of carotid atherosclerosis even though this association was only of moderate strength and not entirely consistent in other vascular territories.

Insulin resistance as estimated by homeostasis model assessment predicts incident symptomatic cardiovascular disease in caucasian subjects from the general population: the Bruneck study.

OBJECTIVE: The purpose of this study was to evaluate whether insulin resistance is associated to cardiovascular disease (CVD) and to understand whether this association can be explained by traditional and novel CVD risk factors associated with this metabolic disorder. RESEARCH DESIGN AND METHODS: We examined a sample representative of the population of Bruneck, Italy (n = 919; aged 40-79 years). Insulin-resistant subjects were those with a score in the top quartile of the homeostasis model assessment (HOMA) for insulin resistance (HOMA-IR). Risk factors correlated with insulin resistance included BMI, A1C, HDL cholesterol, triglycerides, blood pressure, high-sensitivity C-reactive protein (hsCRP), fibrinogen, oxidized LDL, vascular cell adhesion molecule-1 (VCAM-1), and adiponectin. Subjects without CVD at baseline were followed up for 15 years for incident CVD, a composite end point including fatal and nonfatal myocardial infarction and stroke, transient ischemic attack, and any revascularization procedure. RESULTS: During follow-up, 118 subjects experienced a first symptomatic CVD event. Levels of HOMA-IR were higher at baseline among subjects who developed CVD (2.8) compared with those remaining free of CVD (2.5) (P < 0.05). Levels of HOMA-IR also were significantly correlated (P < 0.05) with most CVD risk factors we evaluated. In Cox proportional hazard models, insulin-resistant subjects had an age-, sex-, and smoking-adjusted 2.1-fold increased risk (95% CI 1.3-3.1) of incident symptomatic CVD relative to non-insulin-resistant subjects. After sequential adjustment for physical activity and classic risk factors (A1C, LDL cholesterol, and hypertension) as well as BMI, HDL cholesterol, triglycerides, and novel risk factors, including fibrinogen, oxidized LDL, hsCRP, VCAM-1, and adiponectin, the association between HOMA-IR and incident CVD remained significant and virtually unchanged (hazard ratio 2.2 [95% CI 1.4-3.6], P < 0.001). CONCLUSIONS: HOMA-estimated insulin resistance is associated with subsequent symptomatic CVD in the general population independently of all classic and several nontraditional risk factors. These data suggest that insulin resistance may be an important target to reduce CVD risk.

High-sensitivity C-reactive protein and risk of nontraumatic fractures in the Bruneck study.

BACKGROUND: Chronic inflammatory diseases are associated with bone loss and an enhanced fracture risk. It is unknown, however, whether low-grade inflammation in healthy individuals, as estimated by the high-sensitivity C-reactive protein (hs-CRP) level, interferes with bone metabolism and affects the risk of nontraumatic fractures. METHODS: Lifetime bone fractures were carefully recorded in the cohort of the population-based Bruneck Study (n = 919) along with information on the date of occurrence and associated circumstances. The serum level of hs-CRP was measured from blood samples collected during the 1990 baseline examination and the 1995, 2000, and 2005 follow-up examinations. In addition, lifestyle and demographic characteristics, bone ultrasonographic data at the heel, and variables of bone metabolism were assessed. RESULTS: Between September 1, 1990, and August 31, 2005, 69 subjects experienced nontraumatic hip or vertebral fractures. The incidence of nontraumatic fractures was 1.3, 3.8, and 13.9 per 1000 person-years in the tertile groups for hs-CRP. In multivariate pooled logistic regression analysis, the adjusted relative risk (95% confidence interval) of nontraumatic fracture in the highest vs lowest tertile group for hs-CRP was 9.4 (3.6-24.8) (P < .001). The exclusion of subjects with cardiovascular disease, dementia, malignancies, and chronic inflammatory disease had little effect on the results obtained. The hs-CRP level was unrelated to ultrasonographic measures of bone density, but showed an inverse relation to laboratory markers of bone turnover, like beta-crosslaps and osteocalcin concentration (P < .001). CONCLUSIONS: The hs-CRP level is a significant and independent risk predictor of nontraumatic fracture. This finding is consistent with the hypothesis of a tight interplay between low-grade inflammation and bone turnover.

Oxidized phospholipids predict the presence and progression of carotid and femoral atherosclerosis and symptomatic cardiovascular disease: five-year prospective results from the Bruneck study.

OBJECTIVES: The purpose of this work was to determine the predictive value of oxidized phospholipids (OxPLs) present on apolipoprotein B-100 particles (apoB) in carotid and femoral atherosclerosis. BACKGROUND: The OxPLs are pro-inflammatory and pro-atherogenic and may be detected using the antibody E06 (OxPL/apoB). METHODS: The Bruneck study is a prospective population-based survey of 40- to 79-year-old men and women initiated in 1990. Plasma levels of OxPL/apoB and lipoprotein (a) [Lp(a)] were measured in 765 of 826 (92.6%) and 671 of 684 (98.1%) subjects alive in 1995 and 2000, respectively, and correlated with ultrasound measures of carotid and femoral atherosclerosis. RESULTS: The distribution of the OxPL/apoB levels was skewed to lower levels and nearly identical to Lp(a) levels. The OxPL/apoB and Lp(a) levels were highly correlated (r = 0.87, p < 0.001), and displayed long-term stability and lacked correlations with most cardiovascular risk factors and lifestyle variables. The number of apolipoprotein (a) kringle IV-2 repeats was inversely related to Lp(a) mass (r = -0.48, p < 0.001) and OxPL/apoB levels (r = -0.46, p < 0.001). In multivariable analysis, OxPL/apoB levels were strongly and significantly associated with the presence, extent, and development (1995 to 2000) of carotid and femoral atherosclerosis and predicted the presence of symptomatic cardiovascular disease. Both OxPL/apoB and Lp(a) levels showed similar associations with atherosclerosis severity and progression, suggesting a common biological influence on atherogenesis. CONCLUSIONS: This study suggests that pro-inflammatory oxidized phospholipids, present primarily on Lp(a), are significant predictors of the presence and extent of carotid and femoral atherosclerosis, development of new lesions, and increased risk of cardiovascular events. The OxPL biomarkers may provide valuable insights into diagnosing and monitoring cardiovascular disease.

Association of serum-soluble heat shock protein 60 with carotid atherosclerosis: clinical significance determined in a follow-up study.

BACKGROUND AND PURPOSE: Previous work has shown that soluble heat shock protein 60 (HSP60; sHSP60), present in circulating blood, is associated with carotid atherosclerosis. In the current evaluation, we tested the hypothesis that sHSP60 levels are associated with the progression of carotid arteriosclerosis, prospectively. METHODS: The association of sHSP60 with early atherogenesis (5-year development and progression of nonstenotic carotid plaques) was investigated as part of the population-based prospective Bruneck Study. The current study focused on the follow-up period between 1995 and 2000 and, thus, included 684 subjects. RESULTS: sHSP60 levels measured in 1995 and 2000 were highly correlated (r=0.40; P<0.001), indicating consistency over a 5-year period. Circulating HSP60 levels were significantly correlated with antilipopolysaccharide and anti-HSP60 antibodies. It was also elevated in subjects with chronic infection (top quintile group of HSP60, among subjects with and without chronic infection: 23.8% versus 17.0%; P=0.003 after adjustment for age and sex). HSP60 levels were significantly associated with early atherogenesis, both in the entire population (multivariate odds ratio, for a comparison between quintile group V versus I+II: 2.0 [1.2 to 3.5] and the subgroup free of atherosclerosis at the 1995 baseline: 3.8 [1.6 to 8.9]). The risk of early atherogenesis was additionally amplified when high-sHSP60 and chronic infection were present together. CONCLUSIONS: Our study provides the first prospective data confirming an association between high levels of sHSP60 and early carotid atherosclerosis. This possibly indicates an involvement of sHSP60 in activating proinflammatory processes associated with early vessel pathology.

Urinary albumin excretion is independently associated with carotid and femoral artery atherosclerosis in the general population.

AIMS: In diabetic patients, increased urinary albumin excretion (UAE), termed microalbuminuria when in the range between 30 and 300 mg/dL per day, is associated with a higher risk of atherosclerosis and its complications. Whether or not this notion applies to the general population is a matter of ongoing controversy because none of the few previous investigations among non-diabetics strictly represent the general community. METHODS AND RESULTS: Urinary albumin-to-creatinine ratio (uACR), a measure of UAE, was assessed from overnight spot urine samples in a population-based cohort of 684 individuals. The ratio was significantly related to age, gender, blood pressure, diabetes, markers of systemic inflammation, liver enzymes, and parathyroid hormone levels (P<0.001 each). Moreover, uACR emerged as a highly significant risk predictor of carotid and femoral artery atherosclerosis in the general community and the non-diabetic subpopulation alike (age/sex-adjusted P<0.001 each). In multivariable logistic regression analyses, odds ratios (95% CI) of carotid and femoral atherosclerosis amounted to 1.28 (1.01-1.61) and 1.44 (1.15-1.81) for a one unit increase in log(e)-transformed uACR (P=0.040 and 0.002). Corresponding odds ratios in non-diabetic subjects were 1.41 (1.09-1.84) and 1.54 (1.19-1.99) (P=0.010 and 0.001). Multivariable linear regression analyses yielded significant, or near significant, relations with carotid and femoral artery intima-media thickness and atherosclerosis scores (P=0.058-0.001). CONCLUSION: The uACR is significantly and independently associated with the presence and severity of atherosclerosis in the general population. The relation obtained was of a dose-response type and extended to levels far below what is termed microalbuminuria. The novel aspects of our study are its focus on various vascular territories and representivity of the general healthy population.

Population-based incidence rates and risk factors for type 2 diabetes in white individuals: the Bruneck study.

Incidence rates and risk factors for type 2 diabetes in low-risk populations are not well documented. We investigated these in white individuals who were aged 40-79 years and from the population of Bruneck, Italy. Of an age- and sex-stratified random sample of 1,000 individuals who were identified in 1990, 919 underwent an oral glucose tolerance test (OGTT) and an assessment of physiological risk factors for diabetes, including insulin resistance (homeostasis model assessment, HOMA-IR), and postchallenge insulin response (Sluiter's Index). Diabetes at baseline by fasting or 2-h OGTT plasma glucose (World Health Organization criteria, n = 82) was excluded, leaving 837 individuals who were followed for 10 years. Incident cases of diabetes were ascertained by confirmed diabetes treatment or a fasting glucose >or=7.0 mmol/l. At follow-up, 64 individuals had developed diabetes, corresponding to a population-standardized incidence rate of 7.6 per 1,000 person-years. Sex- and age-adjusted incidence rates were elevated 11-fold in individuals with impaired fasting glucose at baseline, 4-fold in those with impaired glucose tolerance, 3-fold in overweight individuals, 10-fold in obese individuals, and approximately 2-fold in individuals with dyslipidemia or hypertension. Incidence rates increased with increasing HOMA-IR and decreasing Sluiter's Index. As compared with normal insulin sensitivity and normal insulin response, individuals with low insulin sensitivity and low insulin response had a sevenfold higher risk of diabetes. Baseline impaired fasting glucose, BMI, HOMA-IR, and Sluiter's Index were the only independent predictors of incident diabetes in multivariate analyses. We conclude that approximately 1% of European white individuals aged 40-79 years develop type 2 diabetes annually and that "subdiabetic" hyperglycemia, obesity, insulin resistance, and impaired insulin response to glucose are independent predictors of diabetes.

Soluble RANKL and risk of nontraumatic fracture.

CONTEXT: The receptor activator of nuclear factor kappaB ligand (RANKL) is essential for osteoclast and, possibly, osteoblast activation and may represent a key link between bone formation and resorption. OBJECTIVE: To determine the relationship between serum level of RANKL and the risk of nontraumatic fracture. DESIGN, SETTING, AND PARTICIPANTS: As part of a prospective population-based study conducted in Bruneck, Italy, we recorded all fractures that occurred between 1990 and 2000 in 906 participants and classified them as traumatic (n = 115) or nontraumatic (n = 31). Serum levels of RANKL and osteoprotegerin and characteristics of bone metabolism and lifestyle were assessed in 1990 and at follow-up in 1995 and 2000. MAIN OUTCOME MEASURE: Incident nontraumatic fracture by levels of RANKL. RESULTS: Levels of RANKL did not differ between sexes and were not related to age, menopausal status, lifestyle characteristics, or data from bone ultrasound at the heel. However, RANKL emerged as a significant predictor of nontraumatic fracture. In pooled logistic regression analysis, the relative risks of nontraumatic fracture in the lowest and middle vs highest tertile for RANKL were 10.0 (95% confidence interval [CI], 2.3-43.1) and 3.9 (95% CI, 0.8-19.0) (P<.001 for trend), respectively. Patients in the highest-tertile group had a low risk of fracture even in the presence of other predisposing factors, whereas women aged 60 years or older in the lowest tertile had a 5-year rate of nontraumatic fracture greater than 7%. CONCLUSIONS: A low level of RANKL is an independent predictor of nontraumatic fracture. This finding is consistent with the hypothesis of an important role of RANKL in human bone turnover and if confirmed in future investigations may gain relevance for assessment of fracture risk.