Robert H. Knopp

Shotgun proteomics implicates protease inhibition and complement activation in the antiinflammatory properties of HDL.

HDL lowers the risk for atherosclerotic cardiovascular disease by promoting cholesterol efflux from macrophage foam cells. However, other antiatherosclerotic properties of HDL are poorly understood. To test the hypothesis that the lipoprotein carries proteins that might have novel cardioprotective activities, we used shotgun proteomics to investigate the composition of HDL isolated from healthy subjects and subjects with coronary artery disease (CAD). Unexpectedly, our analytical strategy identified multiple complement-regulatory proteins and a diverse array of distinct serpins with serine-type endopeptidase inhibitor activity. Many acute-phase response proteins were also detected, supporting the proposal that HDL is of central importance in inflammation. Mass spectrometry and biochemical analyses demonstrated that HDL3 from subjects with CAD was selectively enriched in apoE, raising the possibility that HDL carries a unique cargo of proteins in humans with clinically significant cardiovascular disease. Collectively, our observations suggest that HDL plays previously unsuspected roles in regulating the complement system and protecting tissue from proteolysis and that the protein cargo of HDL contributes to its antiinflammatory and antiatherogenic properties.

Efficacy and safety of atorvastatin in the prevention of cardiovascular end points in subjects with type 2 diabetes: the Atorvastatin Study for Prevention of Coronary Heart Disease Endpoints in non-insulin-dependent diabetes mellitus (ASPEN).

OBJECTIVE—Cardiovascular disease (CVD) risk is increased in type 2 diabetes. The purpose of this study was to assess the effect of 10 mg of atorvastatin versus placebo on CVD prevention in subjects with type 2 diabetes and LDL cholesterol levels below contemporary guideline targets. RESEARCH DESIGN AND METHODS—Subjects were randomly assigned to receive 10 mg of atorvastatin or placebo in a 4-year, double-blind, parallel-group study. The composite primary end point comprised cardiovascular death, nonfatal myocardial infarction, nonfatal stroke, recanalization, coronary artery bypass surgery, resuscitated cardiac arrest, and worsening or unstable angina requiring hospitalization. RESULTS—A total of 2,410 subjects with type 2 diabetes were randomized. Mean LDL cholesterol reduction in the atorvastatin group over 4 years was 29% versus placebo (P < 0.0001). When we compared atorvastatin versus placebo, composite primary end point rates were 13.7 and 15.0%, respectively (hazard ratio 0.90 [95% CI 0.73–1.12]). In the subset of 1,905 subjects without prior myocardial infarction or interventional procedure, 10.4% of atorvastatin- and 10.8% of placebo-treated subjects experienced a primary end point (0.97 [0.74–1.28]). In the 505 subjects with prior myocardial infarction or interventional procedure, 26.2% of atorvastatin- and 30.8% of placebo-treated subjects experienced a primary end point (0.82 [0.59–1.15]). Relative risk reductions in fatal and nonfatal myocardial infarction were 27% overall (P = 0.10) and 19% (P = 0.41) and 36% (P = 0.11) for subjects without and with prior myocardial infarction or interventional procedure, respectively. CONCLUSIONS—Composite end point reductions were not statistically significant. This result may relate to the overall study design, the types of subjects recruited, the nature of the primary end point, and the protocol changes required because of changing treatment guidelines. For these reasons, the results of the Atorvastatin Study for Prevention of Coronary Heart Disease Endpoints in Non-Insulin-Dependent Diabetes Mellitus (ASPEN) did not confirm the benefit of therapy but do not detract from the imperative that the majority of diabetic patients are at risk of coronary heart disease and deserve LDL cholesterol lowering to the currently recommended targets.

Defective removal of cellular cholesterol and phospholipids by apolipoprotein A-I in Tangier Disease.

Tangier disease is a rare genetic disorder characterized by extremely low plasma levels of HDL and apo A-I, deposition of cholesteryl esters in tissues, and a high prevalence of cardiovascular disease. We examined the possibility that HDL apolipoprotein-mediated removal of cellular lipids may be defective in Tangier disease. With fibroblasts from normal subjects, purified apo A-I cleared cells of cholesteryl esters, depleted cellular free cholesterol pools available for esterification, and stimulated efflux of radiolabeled cholesterol, phosphatidylcholine, and sphingomyelin. With fibroblasts from two unrelated Tangier patients, however, apo A-I had little or no effect on any of these lipid transport processes. Intact HDL also was unable to clear cholesteryl esters from Tangier cells even though it promoted radiolabeled cholesterol efflux to levels 50-70% normal. Passive desorption of radiolabeled cholesterol or phospholipids into medium containing albumin or trypsinized HDL was normal for Tangier cells. Binding studies showed that the interaction of apo A-I with high-affinity binding sites on Tangier fibroblasts was abnormal. These results indicate that apo A-I has an impaired ability to remove cholesterol and phospholipid from Tangier fibroblasts, possibly because of a defective interaction of apo A-I with cell-surface binding sites. Failure of apo A-I to acquire cellular lipids may account for the rapid catabolism of nascent HDL particles and the low plasma HDL levels in Tangier disease.

Effect of ritonavir on lipids and post-heparin lipase activities in normal subjects

BackgroundIntensive therapy of HIV infection with highly active antiretroviral therapy (HAART) dramatically reduces viral loads and improves immune status. Abnormalities of lipid levels, body fat distribution, and insulin resistance have been commonly reported after starting HAART. Whether the lipid abnormalities result from changes in metabolism after an improvement in HIV status or are partly attributable to the effects of protease inhibitor use is unknown. MethodsTwenty-one healthy volunteers participated in a 2 week double-blind, placebo-controlled study on the effect of the protease inhibitor ritonavir on total lipids, apolipoproteins, and post-heparin plasma lipase activities. ResultsThose taking ritonavir (n = 11) had significantly higher levels of plasma triglyceride, VLDL cholesterol, IDL cholesterol, apolipoprotein B, and lipoprotein (a) compared with placebo (n = 8). HDL cholesterol was lower with therapy as a result of a reduction in HDL3 cholesterol. Post-heparin lipoprotein lipase (LpL) activity did not change but hepatic lipase activity decreased 20% (P < 0.01) in those taking ritonavirrcompared with placebo. Although all lipoprotein subfractions became triglyceride enriched, most of the increase in triglyceride was in VLDL and not in IDL particles. ConclusionTreatment with ritonavir in the absence of HIV infection or changes in body composition results in hypertriglyceridemia that is apparently not mediated by impaired LpL activity or the defective removal of remnant lipoproteins, but could be caused by enhanced formation of VLDL. Long-term studies of patients with HIV infection receiving HAART will be necessary to determine the impact of these drugs and associated dyslipidemia on the risk of coronary artery disease.

Effect of Estrogen/Progestin Potency on Lipid/Lipoprotein Cholesterol

We studied 374 women taking oral contraceptives, 284 women taking estrogen preparations after menopause, and 1086 women taking no hormones, to determine the relation of plasma lipids and lipoprotein cholesterol concentrations to various types of estrogen/progestin formulations. Premenopausal women, using oral contraceptives containing a relatively low dose of estrogen combined with a medium or high dose of progestin (Norlestrin, Ovral, or Demulen) had a 24 per cent higher median concentration of low-density-lipoprotein cholesterol than did those not using hormones (P less than 0.05). Women using oral contraceptives that are high in estrogen and low in progestin (Enovid or Oracon) had significantly higher concentrations of high-density-lipoprotein cholesterol than did nonusers; those using Ovral, a low-estrogen and high-progestin formulation, had significantly lower levels of high-density-lipoprotein cholesterol. In postmenopausal women the use of estrogen was associated with concentrations of low-density-lipoprotein cholesterol that were 11 to 19 per cent below the levels in postmenopausal women who did not use hormones. The effects of estrogen-progestin balance on low-density and high-density lipoproteins may underlie the increased incidence of stroke and myocardial infarction in women of childbearing age who take oral contraceptives.

Managing preexisting diabetes for pregnancy: Summary of evidence and consensus recommendations for care

This document presents consensus panel recommendations for the medical care of pregnant women with preexisting diabetes, including type 1 and type 2 diabetes. The intent is to help clinicians deal with the broad spectrum of problems that arise in management of diabetes before and during pregnancy, and to prepare diabetic women for treatment that may reduce complications in the years after pregnancy. A thorough discussion of the evidence supporting the recommendations is presented in the book, Management of Preexisting Diabetes and Pregnancy , authored by the consensus panel and published by the American Diabetes Association (ADA) in 2008 (1). A consensus statement on obstetrical and postpartum management will appear separately. The recommendations are diagnostic and therapeutic actions that are known or believed to favorably affect maternal and perinatal outcomes in pregnancies complicated by diabetes. The grading system adapted by the ADA was used to clarify and codify the evidence that forms the basis for the recommendations (2). Unfortunately there is a paucity of randomized controlled trials (RCTs) of the different aspects of management of diabetes and pregnancy. Therefore our recommendations are often based on trials conducted in nonpregnant diabetic women or nondiabetic pregnant women, as well as on peer-reviewed experience before and during pregnancy in women with preexisting diabetes (3–4). We also reviewed and adapted existing diabetes and pregnancy guidelines (5–10) and guidelines on diabetes complications and comorbidities (2,3,11–14). ### A. Organization of preconception and pregnancy care #### Recommendations

Equivalent efficacy of a time-release form of niacin (Niaspan) given once-a-night versus plain niacin in the management of hyperlipidemia

This study compared the efficacy and safety of a once-a-night, time-release niacin formulation, Niaspan (Kos Pharmaceuticals, Miami Lakes, FL), with plain niacin and placebo for the treatment of primary hypercholesterolemia. The study was conducted in nine academic lipid research clinics in a randomized, double-blind design. Niaspan 1.5 g at bedtime was compared with plain niacin 1.5 g/d after 8 weeks and 3.0 g/d after 16 weeks in divided doses and with placebo. A total of 223 hypercholesterolemic adult men and women participated. Compared with placebo at 8 weeks, Niaspan versus plain niacin at 1.5 g/d showed comparable efficacy, comparably lowering total cholesterol (C) (8%/8%), triglycerides (16%/18%), low-density lipoprotein (LDL)-C (12%/12%), apolipoprotein (apo B) (12%/12%), apo E (9%/7%), and lipoprotein(a) [Lp(a)] (15%/11%), and raising high-density lipoprotein (HDL)-C (20%/17%), HDL2-C (37%/33%), HDL3-C (17%/16%), and apo A-I (8%/6%) (P < or = .05 in all instances). After 16 weeks, the Niaspan effect on LDL-C and triglyceride was unchanged while the plain niacin effect approximately doubled. At equal doses of 1.5 g/d of Niapan versus plain niacin, respectively, AST increased 5.0% versus 4.8% (difference not significant [NS]), fasting plasma glucose increased 4.8% versus 4.5% (NS), and uric acid concentrations increased less, 6% versus 16% (P=.0001). Flushing events were more frequent with plain niacin versus Niaspan (1,905 v 576, P < .001). Flushing severity was slightly greater with Niaspan, but still well tolerated. In conclusion, Niaspan 1.5 g hour of sleep (hs) has comparable efficacy, a lower incidence of flushing, a lesser uric acid rise, and an equivalent hepatic enzyme effect than 500 mg thrice-daily plain niacin in hyperlipidemic subjects. Niaspan may be an equivalent or better alternative to plain niacin at moderate doses in the management of hyperlipidemia.

Sex Differences in the Effect of Diabetes Mellitus on Lipoprotein Triglyceride and Cholesterol Concentrations

We studied sex differences in the serum lipid abnormalities associated with diabetes mellitus in 111 patients with insulin-dependent diabetes and 270 patients with non-insulin-dependent diabetes, who were compared with 586 nondiabetic controls. Relative to control levels, the increases in triglycerides were 17 to 34 mg per deciliter greater in diabetic women than in diabetic men. The median low-density-lipoprotein cholesterol concentration in non-insulin-dependent diabetics was 1 to 4 mg per deciliter lower than the control level in women and 16 to 22 mg per deciliter lower in men, and was 30 mg per deciliter higher than control in insulin-dependent diabetic women and similar to control in insulin-dependent diabetic men. The decrease in median high-density-lipoprotein cholesterol in non-insulin-dependent diabetics was 2 to 7 mg per deciliter greater in women than in men, and the increase in high-density-lipoprotein cholesterol in insulin-dependent diabetics was 3 mg per deciliter less in women than in men. We conclude that diabetes has a greater adverse effect on triglyceride and lipoprotein cholesterol concentrations in diabetic women than in diabetic men, and that this may explain the greater increase in risk of arteriosclerosis in diabetic women.

Contrasting effects of unmodified and time-release forms of niacin on lipoproteins in hyperlipidemic subjects: Clues to mechanism of action of niacin

To minimize the cutaneous flushing symptoms associated with niacin use, a time-release capsule form of niacin has been formulated. Thus study compares the effects of time-release niacin with those of unmodified niacin on lipoprotein lipids, including HDL2 and HDL3, apoproteins A-I and A-II, clinical chemistries, symptomatic side effects, and adherence to the medication regimen. Seventy-one primarily hypercholesterolemic subjects were randomized to either unmodified niacin or time-release niacin ad took medication for a six-month period. The two groups were closely matched on anthropomorphic and lipid variables. Adherence to the therapeutic regimen at a dose of 1.5 g/d in the first month of treatment was similar in the two groups. Thereafter, at a dose of 3.0 g/d, adherence was in excess of 90% among subjects taking unmodified niacin but only 64% among those taking time-release niacin, chiefly because of aggravated gastrointestinal symptoms; cutaneous flushing side effects, however, were slightly less common with time-release niacin. At these levels of adherence, LDL cholesterol (C) was reduced 21% by unmodified niacin and 13% by the time release form. Plasma total triglyceride was reduced more with unmodified niacin (27%) than with time-release niacin (8% maximum), and HDL-C and HDL2-C were increased significantly with unmodified niacin (26% and 36%) and were not significantly changed by time-release niacin. Increased to a similar degree on both regimens were HDL3-C (approximately 35%) and apoA-I (approximately 12%). ApoA-II was not affected by either drug regimen.(ABSTRACT TRUNCATED AT 250 WORDS)

Metabolic Control and Progression of Retinopathy: The Diabetes in Early Pregnancy Study

OBJECTIVE To evaluate the role of metabolic control in the progression of diabetic retinopathy during pregnancy. RESEARCH DESIGN AND METHODS We conducted a prospective cohort study of 155 diabetic women in the Diabetes in Early Pregnancy Study followed from the periconceptional period to 1 month postpartum. Fundus photographs were obtained shortly after conception (95% within 5 weeks of conception) and within 1 month postpartum. Glycosylated hemoglobin was measured weekly during the 1st trimester and monthly thereafter. RESULTS In the 140 patients who did not have proliferative retinopathy at baseline, progression of retinopathy was seen in 10.3, 21.1, 18.8, and 54.8% of patients with no retinopathy, microaneurysms only, mild nonproliferative retinopathy, and moderate-to-severe nonproliferative retinopathy at baseline, respectively. Proliferative retinopathy developed in 6.3% with mild and 29% with moderate-to-severe baseline retinopathy. Elevated glycosylated hemoglobin at baseline and the magnitude of improvement of glucose control through week 14 were associated with a higher risk of progression of retinopathy (adjusted odds ratio for progression in those with glycohe-moglobin ≥ 6 SD above the control mean versus those within 2 SD was 2.7; 95% confidence interval was 1.1-7.2; P = 0.039). CONCLUSIONS The risk for progression of diabetic retinopathy was increased by initial glycosylated hemoglobin elevations as low as 6 SD above the control mean. This increased risk maybe due to suboptimal control itself or to the rapid improvement in metabolic control that occurred in early pregnancy. Excellent metabolic control before conception may be required to avoid this increase in risk. Those with moderate-to-severe retinopathy at conception need more careful ophthalmic monitoring, particularly if their diabetes was suboptimally controlled at conception.