Mn death certificate

Adiponectin, predominantly synthesized in the adipose tissue, seems to play an important role in carbohydrate and lipid metabolism and vascular biology (1). It has been found to be a major modulator of insulin action and resistance (2) and to predict the development of type 2 diabetes (3-8). Furthermore, it seems to have substantial anti-inflammatory properties (1). Adiponectin is also related to lipid metabolism, particularly higher levels of HDL cholesterol and lower levels of triglycerides (9). These data suggest that high adiponectin levels may be related to lower risk for coronary heart disease (CHD), and we demonstrated recently that adiponectin levels are associated with a lower risk for myocardial infarction among healthy men in the Health Professionals Follow-up Study (10). Lifestyle and pharmaceutical approaches that increase adiponectin levels therefore might be valuable in decreasing atherosclerotic risk, particularly in individuals with type 2 diabetes, who are at high risk. However, it remains unclear whether adiponectin levels predict CHD risk among individuals with type 2 diabetes, in whom a complex array of metabolic abnormalities most likely contributes to the elevated risk. Glycemia, blood lipids, and inflammatory markers seem to be independently associated with adiponectin levels (11), but it also remains unresolved which pathways may mediate the potential association between adiponectin and CHD risk among diabetic individuals. We therefore evaluated whether adiponectin levels predict CHD events among diabetic men and whether inflammatory markers, cholesterol levels, or Hb[A.sub.1c] mediates this association.

RESEARCH DESIGN AND METHODS

The Health Professionals Follow-up Study is a prospective cohort study of 51,529 U.S. male health professionals (dentists, veterinarians, pharmacists, optometrists, osteopathic physicians, and podiatrists) who were aged 40-75 years at study initiation in 1986. This cohort is followed through biennial mailed questionnaires that focus on various lifestyle factors and health outcomes. In addition, between 1993 and 1994, 18,159 study participants provided blood samples by overnight courier. Among participants who returned blood samples, 1,000 had a confirmed diagnosis of type 2 diabetes (as reported on a supplementary questionnaire sent to all men who reported a diagnosis of diabetes) at baseline or during follow-up through 1998. The present study included 745 men who did not report a diagnosis of angina pectoris, myocardial infarction, coronary bypass surgery or coronary angioplasty, or stroke on any of the biennial questionnaires before blood collection.

Diabetes and cardiovascular end point definitions. The supplementary diabetes questionnaire ascertained the type of diabetes diagnosed (type 1 or type 2), results of diagnostic blood sugars, and information about symptoms at time of diagnosis and use of hypoglycemic medication. In accordance with the criteria of the National Diabetes Data Group (12), confirmation of type 2 diabetes required at least one of the following self-reports on the supplementary questionnaire: 1) an elevated plasma glucose concentration (fasting plasma glucose [greater than or equal to] 7.8 mmol/l, random plasma glucose [greater than or equal to] 11.1 mmol/l, and/or plasma glucose [greater than or equal to] 11.1 mmol/l after [greater than or equal to] 2 h during an oral glucose tolerance test) plus at least one classic symptom (excessive thirst, polyuria, weight loss, or hunger); 2) no symptoms but at least two elevated plasma glucose concentrations (by the above criteria) on different occasions; or 3) treatment with hypoglycemic medication (insulin or oral hypoglycemic agent). We used the National Diabetes Data Group criteria to define diabetes because the majority of our participants had their diabetes diagnosed before the release of the American Diabetes Association criteria in 1997 (13). The validity of self-reported diabetes using the supplementary questionnaire has been documented in a subsample of 71 men from the Health Professionals Follow-up Study cohort. Of these, 12 had incomplete records, whereas the diagnosis of type 2 diabetes was confirmed in 57 (97%) of the remaining 59 (14).

Cardiovascular end points consisted of fatal CHD, nonfatal myocardial infarction, and coronary bypass surgery/coronary angioplasty. The end point did not include angina pectoris. Nonfatal myocardial infarction was confirmed by reviewing medical records using the criteria of the World Health Organization of symptoms plus either typical electrocardiographic changes or elevated cardiac enzyme levels (15). Cardiovascular deaths were confirmed by review of medical records or autopsy reports with the permission of the next of kin. The cause listed on the death certificate was not sufficient alone to confirm a coronary death. Sudden deaths (i.e., death within 1 h of symptom onset in a man without known disease that could explain death) were considered cardiovascular deaths. Physicians who reviewed the records had no knowledge of the self-reported risk factor status. Deaths were reported by next of kin, the postal system, and through records of the National Death Index. Using all sources combined, it is estimated that follow-up for deaths was >98% complete (16).

Blood collection and processing. Each interested participant was sent a blood collection kit that contained instructions and needed supplies (blood tubes, tourniquet, gauze, band-aid, and needles). The participants made arrangements for the blood to be drawn. Blood samples were collected in three 10-ml liquid EDTA blood tubes, placed on ice packs stored in Styrofoam containers, and returned to our laboratory via overnight courier; >95% of the samples arrived within 24 h. After receipt, the chilled blood was centrifuged; aliquotted into plasma, erythrocytes, and buffy coat; and stored in continuously monitored nitrogen freezers at a temperature no higher than -130[degrees]C. We requested information on the date and the time of the blood sample drawing and the time elapsed since the preceding meal to identify nonfasting (>8 h) participants.

All biomarker assays were carried out on a Hitachi 911 analyzer (Roche Diagnostics, Indianapolis, IN). Adiponectin was measured by a competitive radioimmunoassay using a commercial reagent set from Linco Research (St. Louis, MO) with a day-to-day variability at adiponectin concentrations of 3, 6, and 15 ng/ml of 9.2, 6.9, and 9.2%, respectively. Hb[A.sub.1c] determination was based on turbidimetric immunoinhibition using hemolyzed whole blood or packed red cells. The day-to-day variability at Hb[A.sub.1c] concentrations of 5.5 and 9.1% was 1.9 and 3.0%, respectively. The determination of total cholesterol, triglycerides, and HDL cholesterol concentrations was simultaneously performed using reagents and calibrators from Roche Diagnostics; coefficients of variation for these measurements were <1.8%. LDL cholesterol was measured by a homogeneous direct method from Genzyme (Cambridge, MA). The day-to-day variability at LDL cholesterol concentrations of 2.32, 2.74, and 3.34 mmol/l was <3.1%. Measurement of apolipoprotein B-100 (apo[B.sub.100]) was based on the immunonephelometric assay using reagents and calibrators from Wako Chemicals USA (Richmond, VA) with a day-to-day variability of <5%. We calculated non-HDL cholesterol as the difference between total and HDL cholesterol. C-reactive protein (CRP) was measured via an immunoturbidimetric assay using reagents and calibrators from Denka Seiken (Niigata, Japan). The day-to-day variability of the assay at concentrations of 0.91, 3.07, and 13.38 mg/l was 2.8, 1.6, and 1.1%, respectively. Fibrinogen was measured with an immunoturbidimetric assay using reagents and calibrators from Kamiya Biomedical (Seattle, WA). The day-to-day variability of the assay at concentrations of 4.92, 9.51, and 16.29 [micro]g/l was 0.9, 1.1, and 1.5%, respectively. Plasma-soluble fractions of tumor necrosis factor-a receptor 2 (sTNFR2) were measured by ELISA from R & D Systems (Minneapolis, MN). The day-to-day variability of the sTNFR2 assay at concentrations of 89.9, 197, and 444 pg/ml was 5.1, 3.5, and 3.6%, respectively. Creatinine was measured by a rate-blanked method that is based on the Jaffe reaction using Roche Diagnostics reagents with a day-to-day variability of 5.0 and 2.2% at concentrations of 106.1 and 565.8 [micro]mol/l.