Apolipoprotein C-III protein concentrations and gene polymorphisms in type 1 diabetes: Associations with lipoprotein subclasses

doi:10.1016/j.metabol.2004.05.004

Metabolism

Volume 53, Issue 10, October 2004, Pages 1296-1304

https://doi.org/10.1016/j.metabol.2004.05.004 Get rights and content

Abstract

Serum apolipoprotein C-III (apoCIII) concentration and apoCIII gene polymorphisms have been shown to be a risk factor for cardiovascular disease; however, the underlying mechanisms remain unclear. In addition, no studies have been performed that address these issues in type 1 diabetes. The current study investigated apoCIII protein and apoCIII gene variation in a normotriglyceridemic (82 ± 57 mg/dL) population of patients with type 1 diabetes, the Diabetes Control and Complications Trial/Epidemiology of Diabetes Intervention and Complications (DCCT/EDIC) cohort. Blood samples were obtained in 409 patients after an overnight fast. Serum apoCIII concentration was highly correlated with multiple changes in lipids and lipoproteins that resulted in an adverse cardiovascular disease risk profile. Higher apoCIII concentrations were associated (P < .0001) with increased triglycerides (r = 0.78), total (r = 0.61) and low-density lipoprotein (LDL) (r = 0.40) cholesterol, apoA-I (r = 0.26), and apoB (r = 0.50), and these relationships persisted after controlling for age, gender, body mass index (BMI), and hemoglobin A_1c (HbA_1c). Nuclear magnetic resonance (NMR) lipoprotein subclass analyses demonstrated that apoCIII was correlated with an increase in very-low-density lipoprotein (VLDL) subclasses (P = .0001). There also was a highly significant positive relationship between serum apoCIII concentration and the LDL particle concentration in both men (r = 0.49, P = .001) and women (r = 0.40, P = .001), and a highly significant negative relationship between serum apoCIII levels and average LDL particle size in both men (r = −0.37, P = .001) and women (r = −0.22, P = .001) due primarily to an augmentation in the small L1 subclass (r = 0.42, P = .0001). Neither the T⁻⁴⁵⁵→C polymorphism affecting an insulin response element in the apoCIII gene promoter nor a SacI polymorphism in the 3′UTR were associated with any alterations in circulating apoCIII concentrations, serum lipids, apolipoprotein concentrations, lipoprotein composition, or parameters measured by NMR lipoprotein subclass analyses. In summary, elevated apoCIII concentration was associated with risk factors for cardiovascular disease in normolipidemic type 1 diabetic patients through associated changes in lipoprotein subfraction distributions, which were independent of apoCIII genotype.

Section snippets

Patients and blood samples

A blood sample was collected after an overnight fast of at least 8 hours and prior to insulin administration from 409 type 1 diabetic patients consecutively appearing for their biennial evaluation of lipid levels and exam according to the DCCT/EDIC research protocol.29 Serum samples for measurement of total cholesterol, HDL cholesterol, and triglycerides concentrations were shipped on dry ice to the trial’s Central Biochemistry Laboratory, Department of Laboratory Medicine and Pathology,

Relationships between ApoCIII protein levels and clinical variables, lipids, and lipoproteins

Table 2 details the simple regressions between serum apoCIII levels with lipids and lipoproteins in the DCCT/EDIC cohort of patients with type 1 diabetes. With respect to the traditional lipid panel, apoCIII was highly correlated (P = .0001) with serum triglycerides (r = .78), total cholesterol (r = .61), and calculated LDL cholesterol (r = .40), but not with HDL cholesterol (r = 0.07; P = .265).

To more precisely study independent effects of apoCIII on these important parameters, we employed

Discussion

The results of this study show no association between apoCIII gene polymorphisms, including both the 5′ promoter (−455) and the 3′ SacI polymorphisms, and circulating levels of apoCIII protein in type 1 diabetic patients. Thus, the 5′ promoter polymorphism (−455) that disrupts an insulin response element9, 10 did not appear to affect apoCIII gene expression in type 1 diabetes. Furthermore, apoCIII gene polymorphisms were not associated with any alterations in serum triglycerides, total

Acknowledgements

The authors gratefully acknowledge Karina Moller, Yanis Bellil, Lyle Walton, and Leslie Potter for performing lipoprotein ultracentrifugation and various assays; Kirby Smith for DNA extractions; Jenny Smith and Leslie Nicholson for study coordination; and John Bercik, the General Clinical Research Center (GCRC) (M01-RR-1070) computer systems manager for computer database support. The authors also thank Dr Jim Otvos of LipoMed, Inc for performing NMR lipoprotein subclass analyses. Finally, the

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Citation Excerpt :
In-vitro studies showed that exposure of vascular endothelial cells to apoC-III resulted in inhibition of insulin-stimulated eNOS activity and NO production, suggestive of endothelial dysfunction. In type 1 diabetes, higher apoC-III levels were correlated with lipoprotein subclasses [16] and positively associated with nephropathy and retinopathy [17], suggesting that it may play a role in diabetic microvascular complications. Its role in T2D has been largely unexplored.
Diabetic peripheral neuropathy (DPN) is a common complication of Type 2 diabetes (T2D). Apart from hyperglycemia, its pathogenesis is poorly understood. Apolipoprotein-CIII (apoC-III) associated with triglyceride metabolism, is a risk factor for cardiovascular disease. Its role in DPN is not well-established. We studied the associations of apoC-III, endothelial function and DPN.
In patients with T2D, anthropometric data, fasting blood, and urine were collected for biochemistry and urine albumin/creatinine measurements (uACR). Endothelial function assessments were performed by laser Doppler flowmetry/imaging. DPN was considered present if there was an abnormal finding in monofilament (≤ 8 of 10 points) or neurothesiometer testing ≥ 25 V on either foot. Plasma apoC-III was assessed by ELISA.
Monofilament and neurothesiometer readings were measured in 1981 patients, mean age 57.4 ± 10.8 years old. DPN prevalence was 10.8% (n = 214). Patients with DPN compared to those without, were significantly older (p < 0.0001), with longer duration of T2D (p < 0.0001), had higher BMI (p = 0.006), higher glucose (p = 0.015) and HbA1c (p < 0.0001), Systolic blood pressure (SBP) (p < 0.0001), lower eGFR (p < 0.0001), higher urine ACR (p < 0.0001), poorer endothelium-dependent and endothelium-independent vasodilation (both p < 0.0001), higher VCAM-1 (p < 0.0001) and higher apoC-III [285.3 (195.2–405.6) vs 242.9(165.0–344.0) μg/ml]. After adjustment, log transformed apoC-III, remained independently associated with the presence of DPN (B = 0.965, SE = 0.397, p = 0.015).
Plasma apoC-III is higher in patients with DPN. Apart from its known association with lipids and macrovascular complications, this study suggests its association with DPN. Whether regulating apoC-III metabolism may be an important new therapeutic approach to managing dyslipidemia and microvascular complications in T2D remains to be proven in future mechanistic and clinical studies.
Disialylated apolipoprotein C-III proteoform is associated with improved lipids in prediabetes and type 2 diabetes1
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The apoC-III proteoform containing two sialic acid residues (apoC-III₂) has different in vitro effects on lipid metabolism compared with asialylated (apoC-III₀) or the most abundant monosialylated (apoC-III₁) proteoforms. Cross-sectional and longitudinal associations between plasma apoC-III proteoforms (by mass spectrometric immunoassay) and plasma lipids were tested in two randomized clinical trials: ACT NOW, a study of pioglitazone in subjects with impaired glucose tolerance (n = 531), and RACED (n = 296), a study of intensive glycemic control and atherosclerosis in type 2 diabetes patients. At baseline, higher relative apoC-III₂ and apoC-III₂/apoC-III₁ ratios were associated with lower triglycerides and total cholesterol in both cohorts, and with lower small dense LDL in the RACED. Longitudinally, changes in apoC-III₂/apoC-III₁ were inversely associated with changes in triglycerides in both cohorts, and with total and small dense LDL in the RACED. apoC-III₂/apoC-III₁ was also higher in patients treated with PPAR-γ agonists and was associated with reduced cardiovascular events in the RACED control group. Ex vivo studies of apoC-III complexes with higher apoC-III₂/apoC-III₁ showed attenuated inhibition of VLDL uptake by HepG2 cells and LPL-mediated lipolysis, providing possible functional explanations for the inverse association between a higher apoC-III₂/apoC-III₁ and hypertriglyceridemia, proatherogenic plasma lipid profiles, and cardiovascular risk.
Icosapent ethyl (eicosapentaenoic acid ethyl ester): Effects on plasma apolipoprotein C-III levels in patients from the MARINE and ANCHOR studies
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In agreement with prior research,23 patients with higher baseline ApoC-III levels also had higher TG levels, as demonstrated in comparing MARINE with ANCHOR. Furthermore, the analysis demonstrated strong correlations between baseline levels of ApoC-III and lipids/lipoproteins related to TRLs (i.e., TG, TG/HDL-C ratio, non-HDL-C).14,23–25 In contrast to TRL correlations, the analysis revealed weaker correlations between ApoC-III and ApoB: less robust correlations in ANCHOR (r = 0.31–0.36; P < .0001) and no correlations in patients with very high TG levels from the MARINE study.
Apolipoprotein C-III (ApoC-III) regulates lipoprotein and triglyceride (TG) metabolism and may have a causal role in cardiovascular disease. In the Multi-Center, Placebo-Controlled, Randomized, Double-Blind, 12-Week Study With an Open-Label Extension (MARINE) and ANCHOR studies, icosapent ethyl, a high-purity prescription eicosapentaenoic acid ethyl ester, reduced TG, and other atherogenic lipid parameters without increasing low-density lipoprotein cholesterol (LDL-C) compared with placebo.
To evaluate the effects of icosapent ethyl on plasma ApoC-III levels in patients from 2 phase 3 studies.
MARINE and ANCHOR were 12-week double-blind studies of icosapent ethyl in adult patients. Patients in MARINE had very high TG levels (≥500 and ≤2000 mg/dL) and patients in ANCHOR had high TG levels (≥200 and <500 mg/dL) despite statin control of LDL-C. This post hoc analysis of MARINE and ANCHOR assessed the median percent change from baseline in plasma ApoC-III levels vs placebo and includes subgroup analyses by statin use/efficacy and median ApoC-III levels.
We assessed ApoC-III levels in 148 and 612 patients in the MARINE and ANCHOR studies, respectively. In MARINE, the approved prescription dose of icosapent ethyl (4 g/day) significantly reduced ApoC-III levels by 25.1% (P < .0001) vs placebo. In ANCHOR, icosapent ethyl 4 g/day significantly reduced ApoC-III levels by 19.2% (P < .0001) vs placebo; subanalysis by statin efficacy revealed significant reductions vs placebo in the higher-efficacy and medium-efficacy groups (24.6% and 17.2%, respectively; both P < .0001).
Icosapent ethyl 4 g/day significantly reduced plasma ApoC-III levels in patients with elevated TGs from the MARINE and ANCHOR studies.
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In their report, NMR lipoprotein parameters remained significantly different between controls and T2DM patients after adjustment for established risk factors. Other studies have been made on the effects of atorvastatin on NMR-derived lipoprotein fraction concentrations [156], the relationships between apolipoprotein C-III concentrations and lipoprotein subclasses [157], the interrelations between poor glycemia and lipoprotein metabolism [158], the association between retinopathy and lipoprotein subclasses in T1DM [159,160] and T2DM [160] diabetic patients, and the interrelations between diabetic nephropathy and lipoprotein metabolism [159]. It should be pointed out that the new associations found between retinopathy and lipoprotein subclasses were previously unknown and could not be detected using conventional lipid profiles.
Association of a DNA polymorphism of the apolipoprotein AI-CIII-AIV gene cluster with myocardial infarction in a Tunisian population
2011, European Journal of Internal Medicine
Citation Excerpt :
In our study, there was no significant difference among genotypes for any lipid, or lipoprotein traits. These findings are in agreement with some earlier studies [6,27,28,33], but not all [14,20,34–40]. The discrepancies between the studies may be explained by genetic difference between the Tunisian population used in our study and populations where this association has been detected.
Apolipoproteins AI–CIII–AIV play important roles in the metabolism of triglycerides and high-density lipoprotein cholesterol. However, whether genetic variations in the ApoAI–CIII–AIV gene cluster are associated with the risk of myocardial infarction (MI) remains uncertain. In the present study, we examined a possible association of the ApoCIII SacI polymorphism in the ApoAI–CIII–AIV gene cluster with lipid parameters and MI in a sample of the Tunisian population.
A total of 326 Tunisian patients with MI and 361 controls were included in the study. Genotypes were determined by polymerase chain reaction — restriction fragment length polymorphism (PCR-RFLP) analysis.
A significant difference in genotype distribution and allele frequency was observed between patients and controls. At the multivariate analysis after adjustment for traditional vascular risk factors, the ApoCIII SacI polymorphism was significantly associated with MI, according to co-dominant and dominant models (co-dominant model odds ratio [OR]: 1.53, 95% confidence interval [CI]: 1.0–2.35, p = 0.04; dominant model OR: 2.02, 95% CI: 1.11–3.67, p = 0.02). The MI patient group showed a significant higher frequency of the S2 allele compared to the controls (10.2% vs. 6.5%; OR: 1.64, 95% CI: 1.10–2.47, p = 0.01). There was no statistically significant association between ApoAI–CIII–AIV cluster gene polymorphism and lipid, lipoprotein, and apolipoprotein levels in both MI patients and controls.
In the current study, a significant association between the ApoCIII SacI polymorphism (presence of S2 allele) and MI in the Tunisian population was found.
LDL-containing immune complexes in the DCCT/EDIC cohort: Associations with lipoprotein subclasses
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Clinical characteristics reported in Table 1 for these 845 patients did not differ significantly from those in the 1301 patients who comprise the overall DCCT/EDIC cohort (data not shown). Additional clinical characteristics of these patients have been reported in our previous publications (Jenkins et al., 2003a, 2003b; Jenkins, Lyons, et al., 2004; Klein et al., 2004; Lopes-Virella et al., 2007; Lyons et al., 2006). Immune complexes (IC) were precipitated from serum as described previously (Lopes-Virella et al., 2007; Mironova et al., 1997).
Immune complexes containing modified LDL (LDL-IC) and NMR-determined Total LDL particle concentrations are significantly associated with intima-media thickness (IMT). We analyzed the associations between concentrations of NMR-determined lipoprotein subclasses and LDL-IC in the DCCT/EDIC cohort. LDL-IC concentrations in women and men of the DCCT/EDIC cohort did not differ significantly and were positively associated with Total LDL particle concentrations in men and women (r=0.34, r=0.32, respectively; P<.01) and with Small LDL concentration (r=0.22, r=0.13, respectively; P<.01). In women, Large LDL concentrations were also associated with LDL-IC (r=0.20, P<.01) while in men, the association was more modest (r=0.11, P<.05). Thus, both Small and Large LDL are associated with LDL-IC formation. Based on the results from statistical mediation analyses, we concluded that plasma concentrations of LDL-IC may provide a physiological link between the statistically significant association of Total LDL particle concentration with carotid artery IMT in subjects with Type 1 diabetes. Furthermore, after adjusting for conventional risk factors, there was a decrease in LDL-IC concentration even in the presence of high Total LDL particle concentrations in those women with high concentrations of Large HDL, but the association was not evident in men. This suggests that the associations between Large HDL and Total LDL particle concentrations, and their associations with LDL-IC levels, differ by gender and suggest that LDL-IC partially mediate the contribution of Total LDL particle concentration to increased carotid IMT in diabetic men.

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^☆: Supported by a program project grant (Principal Investigator: W.T. Garvey) cofunded by the NIH/NHLBI (PO1-HL55782) and the Juvenile Diabetes Foundation International; Juvenile Diabetes Foundation International Research Grant No. 197028 (A.J.J.); American Heart Association Southeast Consortium (9850200V) for analyses of serum apoCIII concentration (W.V.B.); American Heart Association Established Investigator Award 9640278N (N.-A.L.); and by a research contract from the Division of Diabetes, Endocrinology and metabolic diseases of the NIDDK/NIH (DCCT/EDIC).

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Apolipoprotein C-III protein concentrations and gene polymorphisms in type 1 diabetes: Associations with lipoprotein subclasses☆

Abstract

Section snippets

Patients and blood samples

Relationships between ApoCIII protein levels and clinical variables, lipids, and lipoproteins

Discussion

Acknowledgements

Metabolism

Atherosclerosis

Atherosclerosis

Atherosclerosis

J Lipid Res

J Lipid Res

Arteriosclerosis

Lancet

Metabolism

Atherosclerosis

Atherosclerosis

Am J Cardiol

Am J Cardiol

J Lipid Res

J Lipid Res

J Biol Chem

Apolipoprotein C-III gene variation and dyslipidemia

Curr Opin Lipidol

Haplotypes identified by DNA restriction-fragment-length polymorphisms in A-I/C-III/A-IV gene region and hypertriglyceridemia

Am J Hum Genet

Apolipoprotein A-I gene polymorphism associated with premature coronary artery disease and familial hypoalphalipoproteinemia

N Engl J Med

Apolipoprotein A-I and B levels and the risk of ischemic heart disease during a five-year follow-up of men in the Quebec Cardiovascular Study

Circulation

An apolipoprotein CIII haplotype protective against hypertriglyceridemia is specified by promoter and 3′ untranslated region polymorphisms

Proc Natl Acad Sci USA

Common genetic variation in the promoter of the human apo CIII gene abolishes regulation by insulin and may contribute to hypertriglyceridemia

J Clin Invest

Mechanism of hypertriglyceridemia in human apolipoprotein (apo) CIII transgenic miceDiminished very low density lipoprotein fractional catabolic rate associated with increased apo CIII and reduced apo E on the particles

J Clin Invest

Chylomicronemia due to apolipoprotein CIII overexpression in apolipoprotein E null miceApolipoprotein CIII-induced hypertriglyceridemia is not mediated by effects on apolipoprotein E

J Clin Invest

Common genomic variation in the APOC3 promoter associated with variation in plasma lipoproteins

Arterioscler Thromb Vasc Biol

Hypertriglyceridemia and the apolipoprotein CIII gene locusLack of association with the variant insulin response element in Italian school children

Hum Genet

Polymorphic markers in apolipoprotein C-III gene flanking regions and hypertriglyceridemia

Arterioscler Thromb Vasc Biol

Increased risk for endogenous hypertriglyceridaemia is associated with an apolipoprotein C3 haplotype specified by the SstI polymorphism

Eur J Clin Invest