Kenneth K. Wu

Impact of weight loss on plasminogen activator inhibitor (PAI-1), factor VII, and other hemostatic factors in moderately overweight adults.

Based on previous cross-sectional findings, we hypothesized that weight loss could improve several hemostatic factors associated with cardiovascular disease. In a randomized controlled trial, moderately overweight men and women were assigned to one of four weight loss treatment groups or to a control group. Measurements of plasminogen activator inhibitor-1 (PAI-1) antigen, tissue-type plasminogen activator (t-PA) antigen, D-dimer antigen, factor VII activity, fibrinogen, and protein C antigens were made at baseline and after 6 months in 90 men and 88 women. Net treatment weight loss was 9.4 kg in men and 7.4 kg in women. There was no net change (p > 0.05) in D-dimer, fibrinogen, or protein C with weight loss. Significant (p < 0.05) decreases were observed in the combined treatment groups compared with the control group for mean PAI-1 (31% decline), t-PA antigen (24% decline), and factor VII (11% decline). Decreases in these hemostatic variables were correlated with the amount of weight lost and the degree that plasma triglycerides declined; these correlations were stronger in men than women. These findings suggest that weight loss can improve abnormalities in hemostatic factors associated with obesity.

Association of hemostatic variables with prevalent cardiovascular disease and asymptomatic carotid artery atherosclerosis. The Atherosclerosis Risk in Communities (ARIC) Study Investigators.

The relation of hemostatic factor levels to the occurrence of cardiovascular disease is incompletely established. The Atherosclerosis Risk in Communities Study measured fibrinogen, factor VII, factor VIII, von Willebrand factor, antithrombin III, protein C, activated partial thromboplastin time, and other cardiovascular risk factors in nearly 15,000 men and women aged 45 to 64. This analysis assessed the relations of these hemostatic factors with prevalent cardiovascular disease and asymptomatic carotid artery intimal-medial thickness measured by B-mode ultrasound. Compared with participants without cardiovascular disease, those with cardiovascular disease had higher levels of fibrinogen, factor VIII, and von Willebrand factor in both sexes. The other hemostatic factors were less consistently associated with prevalent cardiovascular disease. Only fibrinogen was associated with carotid intimal-medial thickness. Adjusted for age, race, and field center, the odds ratio for carotid wall thickness in the 90th percentile or greater, compared with < 50th percentile, for each SD higher fibrinogen concentration (65 mg/dL) was 1.42 (95% confidence interval, 1.25, 1.62) in men and 1.43 (1.25, 1.64) in women. This population-based study provides further evidence that fibrinogen and possibly factor VIII and von Willebrand factor are risk factors for cardiovascular disease.

Associations of fish intake and dietary n-3 polyunsaturated fatty acids with a hypocoagulable profile. The Atherosclerosis Risk in Communities (ARIC) Study.

Recent epidemiological evidence indicates that the hemostatic profile is an important predictor of cardiovascular disease, yet its dietary determinants are not well established. An important question is whether dietary fatty acid intake influences blood levels of coagulation proteins. We examined potential dietary determinants of six hemostatic factors--fibrinogen, factor VII, factor (vWF), protein C, and antithrombin III--in four population-based samples totaling over 15,000 participants, blacks and whites, in the Atherosclerosis Risk in Communities (ARIC) Study. Usual dietary intake was assessed by a food frequency questionnaire. Cross-sectional associations were explored using multiple linear regression analysis, adjusting for gender, race, age, body mass index, smoking status, alcohol use, diabetes, and field center. Dietary intake of n-3 polyunsaturated fatty acids (PUFAs) showed negative associations with fibrinogen, factor VIII, and vWF (blacks and whites) and a positive association with protein C (whites only). Fish intake, the major source of dietary n-3 PUFAs, was similarly related to the hemostatic profile: a 1 serving per day greater fish intake was associated with the following predicted differences (95% confidence interval): fibrinogen, -2.9 mg/dL (-6.3, 0.5); factor VIII, -3.3% (-5.4, -1.3); vWF, -2.7% (-5.2, -0.1) (blacks and whites); and protein C, +0.07 microgram/mL (0.03, 0.11) (whites only). Other nutrients or foods were variably associated with the hemostatic factors. These population-based associations, although cross-sectional, suggest that increases in n-3 PUFA intake from fish may modify the blood levels of several coagulation factors.

Rosiglitazone and PPAR-γ Overexpression Protect Mitochondrial Membrane Potential and Prevent Apoptosis by Upregulating Anti-Apoptotic Bcl-2 Family Proteins

To determine the involvement of peroxisome proliferator‐activated receptor‐γ (PPAR‐γ) in cytoprotection, we subjected N2‐A cells to oxygen–glucose deprivation followed by reoxygenation (H‐R). Following H‐R insults, H2O2 production was increased while cell viability declined, which was accompanied by loss of mitochondrial membrane potential (MMP), cytochrome c release, caspases 9 and 3 activation, poly(ADP‐ribose)polymerase (PARP) cleavage and apoptosis. Rosiglitazone up to 5 µM protected cell viability, normalized MMP, and prevented apoptotic signals. The protective effect of rosiglitazone was abrogated by GW9662, a PPAR‐γ antagonist, or a specific PPAR‐γ small interference RNA (siRNA) but not a control scRNA. PPAR‐γ overexpression alone was effective in maintaining MMP and preventing apoptosis and its protective effect was also abrogated by PPAR‐γ siRNA or GW9662. To elucidate the mechanism by which PPAR‐γ protects MMP and prevents apoptosis, we analyzed Bcl‐2, Bcl‐xl, and phosphorylated Bad (p‐Bad). H‐R suppressed them. Rosiglitazone or PPAR‐γ overexpression restored them via PPAR‐γ. Rosiglitazone or PPAR‐γ overexpression preserved phosphorylated Akt and 3‐phosphoinositide‐dependent kinase‐1 (PDK‐1) in a PPAR‐γ dependent manner. These results indicate that ligand‐activated PPAR‐γ protects N2‐A cells against H‐R damage by enhancing Bcl‐2/Bcl‐xl and maintaining p‐Bad via preservation of p‐Akt. J. Cell. Physiol. 220: 58–71, 2009.

Protein C, Antithrombin, and Venous Thromboembolism Incidence. A Prospective Population-Based Study

Although deficiencies of protein C and antithrombin, 2 natural plasma anticoagulants, are known risk factors for venous thrombosis, population-based prospective incidence data on these associations are lacking. Venous thromboembolic events have been identified in adults in 2 longitudinal cohort studies, the Atherosclerosis Risk in Communities (ARIC) Study and the Cardiovascular Health Study (CHS). Incidence was examined in relation to prediagnostic plasma levels of protein C (ARIC Study only) and antithrombin. Over a mean of 8.1 years of follow-up, there were 130 incident venous thromboembolic events that were not due to cancer in the ARIC Study. The age-adjusted incidence was elevated 3.36-fold (95% CI 1.24 to 9.11) in the 1.1% of subjects with protein C values <2.0 mg/L compared with subjects with higher values. In contrast, in the ARIC Study and the CHS, there was no association between low plasma antithrombin and venous thromboembolism. In conclusion, in this population-based study, a low protein C, but not antithrombin, level has been determined to be associated with an increased incidence of venous thromboembolism. Attributable risk estimates suggest that low protein C levels account for ≈2.5% of venous thromboembolic events in the ARIC population.

Analysis of Protein-DNA Binding by Streptavidin-Agarose Pulldown

Binding of nuclear transactivators to sequence-specific regulatory elements on the promoter regions is of fundamental importance in gene expression and regulation. DNA-bound transactivators recruit transcription coactivators or repressors and an array of associated proteins that interact with the basal transcription factors, thereby activating the transcription machinery. Analysis of the large complex of proteins that bind to DNA is an important step in elucidating the mechanisms by which gene expressions are regulated. Commonly used techniques to determine DNA-protein binding such as the electrophoretic mobility shift assay (EMSA) have limited value for analyzing simultaneously a large number of proteins in the complex. We describe here a streptavidin-agarose pulldown assay that is capable of analyzing quantitatively binding of an array of proteins to DNA probes. The assay is easy to perform and does not require radiolabeled probes. It involves incubation of nuclear extract proteins with 5biotinylated double-stranded DNA probes and streptavidin-agarose beads. The complex is pulled down, and proteins in the complex are dissociated and analyzed by Western blotting. This method has been shown to be useful in determining the regulation of binding of transactivators, p300/CBP, and associated proteins to the cyclooxygenase-2 (COX-2) promoter.

β-Fibrinogen Gene −455G/A Polymorphism and Coronary Heart Disease Incidence: The Atherosclerosis Risk in Communities (ARIC) Study

Abstract PURPOSE: The −455G/A (HaeIII) polymorphism of the β-fibrinogen gene influences levels of plasma fibrinogen. We determined whether it influences risk of coronary heart disease. METHODS: We conducted a case-cohort study nested within a prospective investigation, the Atherosclerosis Risk in Communities Study. We accumulated 398 incident coronary heart disease cases over a median of 5.3 years of follow-up and compared their −455G/A status with a random sample of the cohort n = 498 . RESULTS: Plasma fibrinogen was higher p = 0.04 in AA homozygous participants (341 mg/dL) than in persons carrying the G allele: GA (290 mg/dL), GG (298 mg/dL). However, there was no significant association between −455G/A and incident CHD. CONCLUSIONS: Although a small effect cannot be excluded, −455G/A does not appear to be an important genetic determinant of CHD.