ABSTRACT
BACKGROUND AND AIMS: Tsukushi (TSK) is a recently identified hepatokine, and we aimed to investigate the association between systemic TSK and the severity of nonalcoholic fatty liver disease (NAFLD) in subjects with and without type 2 diabetes mellitus (DM). METHODS: Three hundred ninety-three DM and 289 without DM individuals were recruited for transient elastography assessment to determine liver steatosis and fibrosis. Serum TSK was measured by ELISA. The presence of NAFLD was defined as controlled attenuation parameter ≥ 248â dB/m. RESULTS: NAFLD was present in 276 (70.2%) and 129 (44.6%) subjects with and without DM respectively, and they had higher serum TSK levels than those without NAFLD [DM group: 91.0â ng/mL (61.7-133.8) vs 82.5 (60.9-118.5), P < .01 respectively; without DM group: 97.1â ng/mL (69.3-148.6) vs 80.8 (53.4-111.6) respectively, P < .01]. Univariate analysis showed that serum TSK significantly correlated with the degree of steatosis and fibrosis both in subjects with and without DM. On multivariable regression analysis, only liver stiffness and estimated glomerular filtration rate were significant determinants of TSK level, and the relationship was independent of diabetes and serum adiponectin. Out of 405 subjects with NAFLD, 49 had either advanced fibrosis or cirrhosis. The area under receiver operating characteristic curve of serum TSK to indicate advanced fibrosis or cirrhosis was 0.70 (95% CI .62-.77), which was significantly better than that of fibrosis-4 index, 0.64 (95% CI .55-.72), P < .05. CONCLUSION: Serum TSK levels were increased in subjects with NAFLD and reflected the severity of liver fibrosis.
Subject(s)
Diabetes Mellitus, Type 2 , Elasticity Imaging Techniques , Non-alcoholic Fatty Liver Disease , Humans , Non-alcoholic Fatty Liver Disease/pathology , Liver/pathology , Diabetes Mellitus, Type 2/complications , Diabetes Mellitus, Type 2/pathology , Liver Cirrhosis/pathologyABSTRACT
OBJECTIVE: Carbamylation is part of the aging process and causes adverse changes in the structure and function of proteins. Lipoproteins are subjected to carbamylation. We investigated the usefulness of carbamylated HDL as a prognostic indicator of survival in patients with type 2 diabetes and the association with mortality outcomes. RESEARCH DESIGN AND METHODS: Baseline plasma carbamylated HDL was measured by ELISA in a cohort of 1,517 patients with type 2 diabetes. The primary outcome was all-cause mortality, and the secondary outcomes were cause-specific deaths, including cardiovascular, renal, infection, and cancer related. RESULTS: Over a median follow-up of 14 years, 292 patients died, and the mortality rate was 14.5 per 1,000 person-years. Plasma carbamylated HDL level was higher in those with a fatal outcome (46.1 ± 17.8 µg/mL vs. 32.9 ± 10.7; P < 0.01). Patients in the third (hazard ratio [HR] 2.11; 95% CI 1.40-3.17; P < 0.001) and fourth quartiles (HR 6.55; 95% CI 4.67-9.77; P < 0.001) of carbamylated HDL had increased mortality risk. After adjustment for conventional risk factors, elevated carbamylated HDL was independently associated with all-cause mortality (HR 1.39; 95% CI 1.28-1.52; P < 0.001) as well as with all the cause-specific mortalities. Adding plasma carbamylated HDL level improved the power of the multivariable models for predicting all-cause mortality, with significant increments in C index (from 0.78 to 0.80; P < 0.001), net reclassification index, and integrated discrimination improvement. CONCLUSIONS: Carbamylation of HDL renders HDL dysfunctional, and carbamylated HDL is independently associated with mortality outcomes in patients with type 2 diabetes.
Subject(s)
Cardiovascular Diseases , Diabetes Mellitus, Type 2 , Cholesterol, HDL , Humans , Kidney , Proportional Hazards Models , Risk FactorsABSTRACT
BACKGROUND AND OBJECTIVES: Protein carbamylation is a consequence of uremia and carbamylated lipoproteins contribute to atherogenesis in CKD. Proteins can also be carbamylated by a urea-independent mechanism, and whether carbamylated lipoproteins contribute to the progression of CKD has not been investigated. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: A case-control study was performed to determine whether there were changes in plasma levels of carbamylated lipoproteins in individuals with type 2 diabetes with eGFR >60 ml/min per 1.73 m2 compared with a group of age- and sex-matched healthy controls. A cohort of 1320 patients with type 2 diabetes with baseline eGFR ≥30 ml/min per 1.73 m2 was longitudinally followed up to evaluate the association between carbamylated lipoproteins and progression of CKD. The primary kidney outcome was defined as doubling of serum creatinine and/or initiation of KRT during follow-up. Plasma carbamylated LDLs and HDLs was measured by ELISA. RESULTS: In individuals with diabetes with eGFR >60 ml/min per 1.73 m2, both plasma carbamylated LDL and HDL levels were higher compared with healthy controls (P<0.001). After a mean follow-up of 9 years of the diabetic cohort, individuals in the top quartile of carbamylated LDL (hazard ratio, 2.21; 95% confidence interval, 1.42 to 3.46; P<0.001) and carbamylated HDL (hazard ratio, 4.53; 95% confidence interval, 2.87 to 7.13; P<0.001) had higher risk of deterioration of kidney function compared with those in the lowest quartile. On multivariable Cox regression analysis, plasma carbamylated LDL was no longer associated with kidney outcome after adjusting for baseline eGFR and potential confounding factors. However, the association between plasma carbamylated HDL and kidney outcome remained significant and was independent of HDL cholesterol. CONCLUSIONS: Plasma carbamylated HDL but not carbamylated LDL was independently associated with progression of CKD in patients with type 2 diabetes.
Subject(s)
Diabetic Nephropathies/blood , Lipoproteins, HDL/blood , Protein Carbamylation , Adult , Aged , Biomarkers/blood , Case-Control Studies , Creatinine/blood , Diabetes Mellitus, Type 2/blood , Diabetes Mellitus, Type 2/complications , Diabetic Nephropathies/diagnosis , Diabetic Nephropathies/etiology , Diabetic Nephropathies/physiopathology , Disease Progression , Female , Glomerular Filtration Rate , Humans , Kidney/physiopathology , Lipoproteins, LDL/blood , Longitudinal Studies , Male , Middle Aged , Prospective StudiesABSTRACT
AIMS: Recent clinical studies have shown that galectin-3 is a prognostic indicator in patients with coronary heart disease and in patients with heart failure. Experimental data suggest that galectin-3 may play a role in atherogenesis. We have evaluated whether serum galectin-3 level is associated with cardiovascular outcome in type 2 diabetes. MATERIALS AND METHODS: Galectin-3 was measured in baseline samples in 1495 persons with type 2 diabetes. The primary cardiovascular outcome, incident cardiovascular events, was defined as first non-fatal myocardial infarction, non-fatal stroke, coronary revascularization, or death from cardiovascular cause. The secondary outcome was all-cause mortality. RESULTS: At baseline, 12% of the subjects had prevalent cardiovascular disease. Serum galectin-3 was increased in the group with incident cardiovascular events compared with those who remained free of events during follow up (9.03 ± 2.98 ng/mL vs 8.15 ± 2.76, P < 0.01). Serum galectin-3 was also significantly increased in those subjects with a fatal outcome. The hazard ratios (HR) for cardiovascular events and all-cause mortality for individuals in the top quartile were 2.50 (95% CI 1.87, 3.36, P < 0.001) and 3.92 (95%CI 2.55, 6.01, P < 0.001), respectively. In a multivariate Cox regression analysis including traditional risk factors, log (eGFR), baseline albuminuria, and cardiovascular disease status, the HR per standard deviation change in galectin-3 was 1.13 (95% CI 1.02, 1.26, P = 0.02) for cardiovascular events and 1.17 (95% CI 1.01, 1.35, P = 0.04) for all-cause mortality. CONCLUSIONS: Serum galectin-3 is associated with adverse cardiovascular outcomes in persons with type 2 diabetes independent of traditional risk factors.
Subject(s)
Biomarkers/blood , Cardiovascular Diseases/diagnosis , Cardiovascular Diseases/mortality , Diabetes Mellitus, Type 2/mortality , Galectin 3/blood , Blood Proteins , Cardiovascular Diseases/blood , Cardiovascular Diseases/etiology , Diabetes Mellitus, Type 2/blood , Diabetes Mellitus, Type 2/complications , Female , Follow-Up Studies , Galectins , Humans , Male , Middle Aged , Prognosis , Risk Factors , Survival RateABSTRACT
AIMS/HYPOTHESIS: Galectin-3 has been implicated in cardiac and renal fibrosis and serves as a prognostic clinical indicator in heart failure. The aim of the present study was to evaluate whether serum galectin-3 level is associated with progressive kidney disease in type 2 diabetes. METHODS: Galectin-3 was measured in baseline samples by ELISA in 1320 participants with type 2 diabetes with eGFR ≥30 ml min-1 1.73 m-2. The primary outcome was defined as doubling of serum creatinine and/or initiation of renal replacement therapy during follow-up. The secondary outcome was progression to macroalbuminuria in individuals with normo- or microalbuminuria at baseline. RESULTS: Serum galectin-3 levels were significantly increased in a random subgroup of 270 type 2 diabetic individuals with eGFR >60 ml min-1 1.73 m-2 compared with an age- and sex-matched non-diabetic control group (7.58 ± 2.29 ng/ml vs 6.10 ± 1.91 ng/ml, respectively, p < 0.01). In the whole diabetic cohort, after a mean follow-up of 9 years, galectin-3 was independently associated with doubling of serum creatinine (HR 1.19; 95% CI 1.14, 1.24, p < 0.001) and incident macroalbuminuria (HR 1.20; 95% CI 1.12, 1.30, p < 0.001), even after adjusting for traditional risk factors, baseline eGFR and albuminuria status. Individuals with galectin-3 levels in the highest quartile had a fourfold risk of renal function loss and threefold risk of incident macroalbuminuria. CONCLUSIONS/INTERPRETATION: Serum galectin-3 was independently associated with progressive renal disease in type 2 diabetes. Further mechanistic studies are warranted to determine whether galectin-3 is simply a disease biomarker or is also a mediator of the development and progression of diabetic nephropathy.
Subject(s)
Diabetes Mellitus, Type 2/blood , Diabetic Nephropathies/blood , Galectin 3/blood , Albuminuria/blood , Biomarkers/blood , Blood Proteins , Creatinine/blood , Diabetes Mellitus, Type 2/pathology , Diabetic Nephropathies/pathology , Disease Progression , Female , Follow-Up Studies , Galectins , Glomerular Filtration Rate , Humans , Kidney Failure, Chronic/blood , Male , Middle Aged , Prognosis , Risk FactorsABSTRACT
BACKGROUND: The receptor for advanced glycation end products (RAGE) is involved in the pathogenesis of diabetic complications, and soluble forms of the receptor (sRAGE) can counteract the detrimental action of the full-length receptor by acting as decoy. Soluble RAGE is produced by alternative splicing [endogenous secretory RAGE (esRAGE)] and/or by proteolytic cleavage of the membrane-bound receptor. We have investigated the role of A Disintegrin And Metalloproteinase 10 (ADAM10) in the ectodomain shedding of RAGE. METHODS: Constitutive and insulin-induced shedding of RAGE in THP-1 macrophages by ADAM10 was evaluated using an ADAM10-specific metalloproteinase inhibitor. Serum ADAM10 level was measured in type 1 diabetes and control subjects, and the association with serum soluble RAGE was determined. Serum total sRAGE and esRAGE were assayed by ELISA and the difference between total sRAGE and esRAGE gave an estimated measure of soluble RAGE formed by cleavage (cRAGE). RESULTS: RAGE shedding (constitutive and insulin-induced) was significantly reduced after inhibition of ADAM10 in macrophages, and insulin stimulated ADAM10 expression and activity. Diabetic subjects have higher serum total sRAGE and esRAGE (p<0.01) than controls, and serum ADAM10 was also increased (p<0.01). Serum ADAM10 correlated with serum cRAGE in type 1 diabetes (r = 0.40, p<0.01) and in controls (r = 0.31. p<0.01) but no correlations were seen with esRAGE. The association remained significant after adjusting for age, gender, BMI, smoking status and HbA1c. CONCLUSION: Our data suggested that ADAM10 contributed to the shedding of RAGE. Serum ADAM10 level was increased in type 1 diabetes and was a significant determinant of circulating cRAGE.
Subject(s)
ADAM Proteins/blood , Amyloid Precursor Protein Secretases/blood , Diabetes Mellitus, Type 1/blood , Membrane Proteins/blood , Receptor for Advanced Glycation End Products/blood , ADAM10 Protein , Adult , Female , Humans , Male , Middle AgedABSTRACT
BACKGROUND: Lipid accumulation has been shown to accelerate renal injury, and the intracellular accumulation of lipids may be caused by alterations in synthesis as well as lipid uptake and efflux. We have investigated the role of cellular cholesterol transport proteins including adenosine triphosphate binding cassette transporter A1 (ABCA1), G1 (ABCG1) and scavenger receptor class B type I (SR-BI) in diabetic nephropathy. METHODS: Protein expression and the ability to mediate cholesterol efflux of ABCA1, ABCG1 and SR-BI was determined in human renal mesangial cells and proximal tubular epithelial cells cultured under normal or high glucose conditions. Renal expression of these cholesterol transporters was examined in a murine model of streptozotocin-induced type 1 diabetes. RESULTS: ABCA1, ABCG1 and SR-BI were expressed in both human renal mesangial cells and proximal tubular epithelial cells, and mediated cholesterol efflux to apolipoprotein AI and HDL. In vitro, hyperglycemia reduced the expression and the ability to mediate cholesterol efflux of all three cholesterol transporters (p<0.05). In vivo studies showed that intra-renal accumulation of lipids was increased in diabetic mice, particularly in mice with nephropathy. This was associated with a significant reduction in the expression of ABCA1, ABCG1 and SR-BI in the kidneys. These changes were already seen in diabetic mice without nephropathy and preceded the development of nephropathy. Diabetic mice with nephropathy had the lowest level of these cholesterol transporters. CONCLUSION: Inducing diabetes with streptozotocin significantly reduced renal expression of ABCA1, ABCG1 and SR-BI. Defects in cholesterol export pathway in renal cells could therefore promote cholesterol accumulation and might contribute to the development of diabetic nephropathy.
Subject(s)
ATP-Binding Cassette Transporters/metabolism , Cholesterol/metabolism , Diabetic Nephropathies/mortality , Scavenger Receptors, Class B/metabolism , Animals , Biological Transport , Blood Glucose/metabolism , Diabetic Nephropathies/blood , Disease Models, Animal , Humans , Immunohistochemistry , Kidney/metabolism , Kidney/pathology , Lipids/blood , Male , Mice, Inbred DBAABSTRACT
LDL (low-density lipoprotein) is subjected to pro-atherogenic modifications in the circulation. A novel uraemia-independent mechanism of carbamylation of lipoproteins mediated by MPO (myeloperoxidase) has recently been reported. We have investigated whether carbamylation of LDL was increased in patients with Type 2 diabetes without renal impairment and the role of MPO. cLDL (carbamylated LDL) and MPO were measured by ELISA in a cross-sectional study of 198 patients and 174 non-diabetic controls. The impact of lowering MPO on plasma cLDL was determined by assaying cLDL and MPO in archived samples from a previous randomized open-label parallel group study comparing rosiglitazone (n=20) and sulfonylurea (n=24). Both plasma cLDL (P<0.05) and MPO levels (P<0.01) were higher in patients with Type 2 diabetes than controls in the cross-sectional study. Plasma cLDL correlated with MPO (r=0.42 and P<0.01) in subjects with diabetes, and plasma MPO was an independent determinant of plasma cLDL even after adjusting for age, gender, BMI (body mass index), apoB (apolipoprotein B), urea and HbA1c (glycated haemoglobin). In the randomized trial, rosiglitazone significantly lowered MPO (P<0.01) and cLDL (P<0.05), whereas no changes were observed in the sulfonylurea group despite a similar reduction in HbA1c. The magnitude of reduction in plasma cLDL correlated with changes in MPO, but not with HbA1c in the rosiglitazone group, suggesting that lowering MPO reduced plasma cLDL. Plasma cLDL is increased in patients with Type 2 diabetes even in the absence of renal impairment and carbamylation of LDL in these subjects is mainly mediated by MPO and not by urea.
Subject(s)
Diabetes Mellitus, Type 2/blood , Lipoproteins, LDL/blood , Peroxidase/metabolism , Adult , Cross-Sectional Studies , Diabetes Mellitus, Type 2/drug therapy , Female , Glyburide/therapeutic use , Glycated Hemoglobin/metabolism , Humans , Lipoproteins, LDL/biosynthesis , Male , Middle Aged , Rosiglitazone , Thiazolidinediones/therapeutic useABSTRACT
AIM: Lectin-like oxidized LDL receptor-1 (LOX-1) is a class E oxidized LDL specific scavenger receptor that recognizes multiple ligands. Advanced glycation end products (AGEs) have been recently identified as other ligands to LOX-1 and shown to increase LOX-1 expressions in diabetes; therefore, we investigated the underlying mechanism involved. METHODS: Confluent human aortic endothelial cells were treated with a fixed concentration of AGE-BSA or BSA as a control in the presence or absence of either antibody of the receptor for advanced glycation end products, mammalian target of rapamycin (mTOR) inhibitor rapamycin, NF-kB inhibitor, phosphoinositide 3-kinases (PI3K) inhibitor or anti-diabetic drug metformin. After stimulation, cells were lysed and Western blot protein expression on LOX-1, rapamycin-insensitive companion of mTOR (RICTOR), the phosphorylation status of p-mTOR, p-P70S6 kinase and p-Akt were determined. RESULTS: AGEs induced LOX-1 expression in endothelial cells. Pretreatment either with anti-RAGE antibody or LY294002 prior to AGE-BSA decreases LOX-1 and p-mTOR expressions. Incubating endothelial cells with AGE-BSA in the presence of rapamycin down-regulated the protein expression-level of p-mTOR by 41% (p<0.05) and LOX-1 expression by 61.5% (p<0.01). Knockdown of RICTOR by RNA silencing showed a 41.5% (p<0.01) and 71.2% (p<0.01) reduction in LOX-1 and p-Akt expressions, respectively. Preincubation of endothelial cells with AGE-BSA and metformin, an anti-diabetic drug known to have an mTOR inhibition effect, significantly reduced AGE-stimulated LOX-1 expression. CONCLUSION: Our results indicated that LOX-1 up-regulation induced by AGE-BSA was a receptor mediated through RAGE and is via the PI3K/PDK1/mTORC2 pathway. Metformincan reduce AGE-stimulated LOX-1 expression in endothelial cells in vitro.
Subject(s)
Endothelial Cells/metabolism , Glycation End Products, Advanced/metabolism , Scavenger Receptors, Class E/metabolism , Serum Albumin, Bovine/metabolism , Aorta/cytology , Cells, Cultured , Dose-Response Relationship, Drug , Gene Expression Regulation, Enzymologic , Humans , Ligands , Metformin/pharmacology , Models, Biological , Phosphoinositide-3 Kinase Inhibitors , RNA, Small Interfering/metabolism , Signal Transduction , TOR Serine-Threonine Kinases/metabolism , Transfection , Up-RegulationABSTRACT
BACKGROUND: In addition to the important role of advanced glycation end products (AGEs) in the pathogenesis of diabetic vascular complications, recent data suggest that advanced glycation end products can also impair insulin action in vitro. We have investigated whether circulating advanced glycation end products are associated with insulin resistance in human subjects independent of metabolic parameters. METHODS: Two hundred and seven healthy non-obese non-diabetic subjects (97 male, 110 female) were recruited from the community. Serum levels of advanced glycation end products, adiponectin, malondialdehyde and high sensitivity C-reactive protein were assayed. Insulin resistance was determined by the homeostasis model assessment index (HOMA-IR). RESULTS: Male subjects had significantly higher body mass index, waist circumference and lower adiponectin level than female subjects and were more insulin resistant. Serum advanced glycation end products (3.67 ± 1.15 unit/mL versus 3.23 ± 1.15, p < 0.05) and malondialdehyde levels (p < 0.05) were also higher in male than in female subjects. Serum advanced glycation end products correlated with HOMA-IR in both male (r = 0.32, p = 0.004) and female subjects (r = 0.28, p = 0.003). Serum adiponectin inversely correlated with HOMA-IR in female (r = - 0.38, p < 0.001) but not in male subjects. On multiple regression analysis, serum AGEs remained an independent determinant of HOMA-IR even after adjusting for age, gender, body mass index, waist, smoking, adiponectin and markers of oxidative stress and inflammation. CONCLUSIONS: Formation and accumulation of advanced glycation end products progress during normal ageing. We have demonstrated that the circulating level of advanced glycation end products is associated with insulin resistance even in non-obese, non-diabetic subjects independent of adiponectin.
Subject(s)
Glycation End Products, Advanced/blood , Insulin Resistance/physiology , Adiponectin/blood , Adult , C-Reactive Protein/metabolism , Female , Humans , Male , Middle AgedABSTRACT
The sRAGE [soluble RAGE (receptor for advanced glycation end-products)] lack the transmembrane and cytoplasmic domain of the full-length receptor and can function as a decoy for RAGE ligands. Recent evidence suggests that sRAGE may be a potential biomarker of RAGE-mediated pathology. The present study aimed to examine the relationship between RAGE expression in peripheral blood monocytes and circulating sRAGE and esRAGE (endogenous sRAGE, a splice variant of sRAGE) in Type 2 diabetes. Protein expression of RAGE and esRAGE in monocyte cell lysate was determined by Western blot in 53 diabetic patients and 52 controls. Monocyte cell-surface-bound full-length RAGE expression was measured using flow cytometry. Serum sRAGE, esRAGE and AGE (advanced glycation end products) were assayed by ELISA. The mean HbA1c (glycated haemoglobin) of the diabetic patients was 9.74% and serum AGEs was increased. Monocyte full-length RAGE expression was significantly higher in diabetic patients whereas esRAGE expression was reduced, and serum AGEs concentration was an independent determinant of monocyte cell surface full-length RAGE expression. Serum levels of sRAGE [573.3 (375.7-754.3) compared with 608.1 (405.3-940.8) pg/ml, P<0.05] and esRAGE [241.8 (154.6-356.6) compared with 286.5 (202.6-390.0) pg/ml, P<0.05; values are medians (interquartile range)] were decreased. There was an inverse association between monocyte RAGE expression and log(serum sRAGE) (r=-0.34, P=0.01) but not with esRAGE. In conclusion, despite an increase in full-length RAGE expression, esRAGE expression was down-regulated in the diabetic patients, and serum sRAGE and esRAGE was also reduced. Hence increased full-length RAGE levels are not associated with a similar increase in sRAGE isoforms levels.
Subject(s)
Diabetes Mellitus, Type 2/blood , Receptors, Immunologic/blood , Blotting, Western/methods , Cross-Sectional Studies , Down-Regulation , Female , Glycated Hemoglobin/metabolism , Glycation End Products, Advanced/blood , Humans , Male , Middle Aged , Monocytes/metabolism , Protein Isoforms/blood , Receptor for Advanced Glycation End Products , Receptors, Immunologic/geneticsABSTRACT
OBJECTIVES: The uptake of oxidized LDL (oxLDL) by macrophages is a key initial event in atherogenesis, and the removal of oxidized lipids from artery wall via reverse cholesterol transport is considered antiatherogenic. The aims of this study were to investigate the pathways mediating the removal of oxysterols from oxLDL-loaded macrophages, and the subsequent uptake of the oxysterols by hepatocytes. METHODS: LDL was labeled with [3H]cholesterol, and LDL-[3H]cholesterol was oxidized by copper using a standard method. [3H]oxysterol formation in oxLDL was analyzed by thin layer chromatography. oxLDL-[3H]sterol was incubated with macrophages, allowing the uptake of [3H]sterol by macrophages. [3H]sterol efflux from macrophages mediated by ATP binding cassette transporters (ABCA1, ABCG1), or scavenger receptor class B type I (SR-BI) was measured. The subsequent uptake of the [3H]sterol by hepatocytes was also determined. RESULTS: 7-Ketocholesterol was the major oxysterol formed in oxLDL, and it was significantly higher in oxLDL compared with that in native LDL (naLDL). oxLDL-derived sterol efflux to HDL from macrophages was significantly increased compared with naLDL-derived sterol, and it was mainly mediated by ABCG1, but not by ABCA1 or SR-BI. Moreover, although HDL dose-dependently induced sterol efflux from macrophages, only the exported sterol by ABCG1 pathway was efficiently taken up by hepatocytes. CONCLUSIONS: ABCG1 mediates oxysterol efflux from oxLDL-loaded macrophages, and the exported oxysterol by ABCG1 pathway can be selectively taken up by hepatocytes.
Subject(s)
ATP-Binding Cassette Transporters/metabolism , Hepatocytes/metabolism , Lipoproteins, LDL/metabolism , Macrophages/metabolism , ATP Binding Cassette Transporter, Subfamily G, Member 1 , Cell Line, Tumor , Humans , Liver/metabolism , Liver X Receptors , Orphan Nuclear Receptors/metabolism , PPAR gamma/metabolismABSTRACT
OBJECTIVE: Reverse cholesterol transport (RCT) plays a protective role against atherosclerosis and cholesterol efflux from cells is an early step in the RCT pathway. We investigated whether the capacity of serum to induce cholesterol efflux was associated with endothelial dysfunction in type 2 diabetes. METHODS: Endothelium-dependent and -independent vasodilation of the brachial artery was measured by high-resolution vascular ultrasound and serum cholesterol efflux capacity was determined by measuring the transfer of [3H]cholesterol from Fu5AH cells to serum in 137 patients with type 2 diabetes and 75 controls. RESULTS: Serum cholesterol efflux capacity was lower in diabetic patients than in the controls (13.6+/-2.5% vs. 14.6+/-3.4%, respectively, p=0.02), and both endothelium-dependent vasodilation (4.9+/-2.2% vs. 8.8+/-4.1%, respectively, p<0.01) and endothelium-independent vasodilation were impaired (13.4+/-4.3% vs. 16.3+/-5.5%. respectively, p<0.01). Endothelium-dependent vasodilation correlated with serum cholesterol efflux capacity (r=0.26, p=0.003) in diabetic patients and controls (r=0.24, p=0.037). On general linear model analysis, the presence of diabetes, brachial artery diameter and serum cholesterol efflux capacity were significant independent determinants of endothelium-dependent vasodilation. CONCLUSION: Impaired serum cholesterol efflux capacity was associated with endothelial dysfunction independent of other cardiovascular risk factors.
Subject(s)
Brachial Artery/physiopathology , Cholesterol/blood , Diabetes Mellitus, Type 2/blood , Diabetes Mellitus, Type 2/physiopathology , Endothelium, Vascular/physiopathology , Vasodilation , Adult , Animals , Biological Transport , Biomarkers/blood , Brachial Artery/diagnostic imaging , Carcinoma, Hepatocellular/metabolism , Case-Control Studies , Cell Line, Tumor , Diabetes Mellitus, Type 2/diagnostic imaging , Endothelium, Vascular/diagnostic imaging , Female , Humans , Linear Models , Liver Neoplasms/metabolism , Male , Middle Aged , Rats , UltrasonographyABSTRACT
AIMS: LDL is subjected to glycoxidation in diabetes mellitus. We have evaluated the effect of glycoxidized LDL on lectin-like oxidized low density lipoprotein receptor-1 (LOX-1) expression in endothelial cells in vitro, as well as the relationship between glycoxidzied LDL and LOX-1 in type 2 diabetic patients with and without microalbuminuria in vivo. METHODS: Endothelial cells were incubated with modified LDL including glycoxidized LDL, oxidized LDL (oxLDL), glycated LDL, and acetylated LDL, and cellular LOX-1 and the soluble forms of LOX-1 (sLOX-1) in cell medium was measured. Glycoxidized LDL in diabetic patients was determined by measuring the glycoxidation product N(epsilon)-(carboxymethyl)lysine (CML) in apolipoprotein (apo) B. Serum oxLDL and sLOX-1 was determined by ELISA. RESULTS: Only glycoxidized LDL and oxLDL significantly increased LOX-1 expression (p<0.05) and the production of sLOX-1 (p<0.05), and the effect of glycoxidized LDL was greater than that of oxLDL. Both normoalbuminuric (n=110) and microalbuminuric (n=91) patients had higher serum apoB-CML than controls (n=105) (p<0.01), but oxLDL was only elevated in the microalbuminuric patients (p<0.05). Serum sLOX-1 was significantly increased in both groups of patients compared to controls (p<0.01). Serum sLOX-1 correlated with apoB-CML (r=0.36, p<0.001) but not with oxLDL. The relationship between sLOX-1 and apoB-CML was independent of HbA1c, age, gender, BMI and smoking status. CONCLUSION: Glycoxidized LDL was more potent than oxLDL in inducing LOX-1 in vitro. Serum concentration of apoB-CML, a marker of glycoxidized LDL, was increased in type 2 diabetic patients with and without microalbuminuria, and this was associated with an increase in serum sLOX-1.
Subject(s)
Diabetes Mellitus, Type 2/blood , Lipoproteins, LDL/metabolism , Scavenger Receptors, Class E/metabolism , Adult , Aged , Albuminuria/blood , Apolipoproteins B/metabolism , Endothelial Cells/metabolism , Enzyme-Linked Immunosorbent Assay , Female , Glycation End Products, Advanced , Glycosylation , Humans , Male , Middle Aged , Models, BiologicalABSTRACT
BACKGROUND: Cholesterol efflux from cells is an early step of reverse cholesterol transport (RCT) and the capacity of serum to induce cellular cholesterol efflux has recently been shown to be an independent predictor of coronary artery atherosclerosis. Our aim is to evaluate the capacity of serum to induce ATP-binding cassette transporter A1 (ABCA1) and scavenger receptor class B type I (SR-BI) mediated cholesterol efflux in type 2 diabetic patients with nephropathy. METHODS: Diabetic patients were recruited according to their urinary albumin excretion rate (normoalbuminuria, microalbuminuria and proteinuria) with 20 subjects in each group and compared with 20 age-matched controls. The ability of the serum to induce cholesterol efflux was measured using a cell culture system. RESULTS: Serum capacity to induce ABCA1-mediated cholesterol efflux was decreased in patients with microalbuminuria or proteinuria (p < 0.05) whereas SR-BI-mediated cholesterol efflux was impaired in all three groups of diabetic patients (p < 0.05). Plasma high-density lipoprotein (HDL) cholesterol and apoAI were reduced in all groups of diabetic patients, but pre-beta-HDL was only significantly decreased in those with microalbuminuria or proteinuria. Serum advanced glycation end products (AGEs) were significantly increased in diabetic patients with microalbuminuria or proteinuria. Serum AGEs and pre-beta-HDL were the significant independent determinants of ABCA1-mediated cholesterol efflux, whereas plasma HDL and log (creatinine) were the significant determinants of SR-BI-mediated cholesterol efflux. CONCLUSION: The capacity of serum to induce ABCA1- and SR-BI-mediated cholesterol efflux was impaired in diabetic patients with incipient or overt nephropathy. These abnormalities may contribute to the accelerated development of atherosclerotic vascular disease in these patients.
Subject(s)
ATP-Binding Cassette Transporters/metabolism , Cholesterol/blood , Cholesterol/metabolism , Diabetes Mellitus, Type 2/metabolism , Diabetic Nephropathies/blood , ATP Binding Cassette Transporter 1 , ATP-Binding Cassette Transporters/blood , ATP-Binding Cassette Transporters/genetics , Albuminuria/blood , Albuminuria/metabolism , Analysis of Variance , Biological Transport , Blood Glucose/analysis , Diabetes Mellitus, Type 2/blood , Female , Glycated Hemoglobin/metabolism , Humans , Lipids/blood , Male , Middle Aged , Proteinuria/blood , Proteinuria/metabolism , RNA, Messenger/genetics , Scavenger Receptors, Class B/blood , Scavenger Receptors, Class B/metabolismABSTRACT
Cellular cholesterol efflux is regulated by cholesterol transporters including adenosine triphosphate-binding cassette transporter A1 (ABCA1), ABCG1, and scavenger receptor class B type I (SR-BI). We have investigated whether the expression of these transporters is affected by type 2 diabetes mellitus and the association with glycemic indexes and oxidized low-density lipoprotein (oxLDL). Messenger RNA of ABCA1, ABCG1, and SR-BI in peripheral monocytes was measured in 30 diabetic patients and 30 matched controls. Plasma oxLDL and advanced glycation end products (AGEs) were assayed by enzyme-linked immunosorbent assay. Cellular cholesterol efflux from monocytes to serum was determined in a subgroup chosen randomly. The expression of ABCG1 was decreased in diabetic patients (P < .05), whereas the levels of ABCA1 and SR-BI were comparable between the 2 groups. Fasting glucose, hemoglobin A(1c), AGEs, and oxLDL were all significantly increased in diabetic patients. There was an inverse correlation between serum AGEs and ABCG1 (r = -0.44, P < .05) that remained significant after adjusting for potential confounding factors. No associations between fasting glucose, hemoglobin A(1c), plasma lipids, or oxLDL and the expression of ABCG1, ABCA1, or SR-BI were found. Cholesterol efflux from monocytes to standard serum or autologous serum was significantly impaired in diabetic patients, and the reduction in efflux to autologous serum correlated with the expression of ABCG1 (r = 0.60, P < .05). The expression of ABCG1 in monocytes is reduced in type 2 diabetes mellitus and is partly related to serum AGEs concentration. The reduction in ABCG1 is associated with impairment in cholesterol efflux and may contribute to accelerated foam cell formation in diabetic patients.
Subject(s)
ATP-Binding Cassette Transporters/biosynthesis , Diabetes Mellitus, Type 2/genetics , Leukocytes, Mononuclear/metabolism , Scavenger Receptors, Class B/biosynthesis , ATP Binding Cassette Transporter 1 , ATP Binding Cassette Transporter, Subfamily G, Member 1 , ATP-Binding Cassette Transporters/blood , ATP-Binding Cassette Transporters/genetics , Biological Transport , Blood Glucose/metabolism , Cholesterol/blood , Diabetes Mellitus, Type 2/blood , Enzyme-Linked Immunosorbent Assay , Female , Gene Expression , Glycated Hemoglobin/analysis , Glycation End Products, Advanced/blood , Humans , Lipoproteins, LDL/blood , Male , Middle Aged , RNA, Messenger/blood , RNA, Messenger/genetics , Reverse Transcriptase Polymerase Chain Reaction , Scavenger Receptors, Class B/blood , Scavenger Receptors, Class B/genetics , Triglycerides/metabolismABSTRACT
OBJECTIVE: Endothelial lipase (EL), a new member of the triacylglycerol lipase family, modulates the metabolism of high-density lipoproteins and is upregulated by inflammatory cytokines. Since type 2 diabetes is associated with chronic subclinical inflammation, we have determined whether serum EL concentration is increased in type 2 diabetes and investigated the effect of insulin on EL. METHODS: 237 type 2 diabetic patients on oral anti-diabetic agents, 111 type 2 diabetic patients on insulin therapy and 226 non-diabetic controls were recruited. Serum EL was measured by ELISA. To investigate the effect of insulin on EL production by endothelial cells, human aortic endothelial cells were incubated with insulin and EL mRNA and protein in cell medium was measured. Serum EL was also measured in 16 diabetic subjects before and after starting insulin therapy. RESULTS: Serum EL level was highest in patients on oral anti-diabetic agents whereas those on insulin had similar EL level as controls (oral: 26.7+/-16.1ng/ml; insulin: 23.3+/-11.6, controls: 23.9+/-12.0; ANOVA p=0.04). In both controls and patients on oral anti-diabetic agents, EL correlated with log(CRP) (r=0.20, p=0.003; r=0.23, p<0.001, respectively) but no correlation was seen in patients on insulin. In vitro experiments showed that insulin significantly reduced EL mRNA and protein in human aortic endothelial cells in a dose-dependent manner. Serum EL concentration also significantly decreased in diabetic patients after starting insulin therapy (p<0.03). CONCLUSION: Serum EL concentration was increased in type 2 diabetic patients and was associated with the degree of subclinical inflammation and exogenous insulin therapy lowered serum EL concentration.
Subject(s)
Diabetes Mellitus, Type 2/diagnosis , Diabetes Mellitus, Type 2/drug therapy , Hypoglycemic Agents/therapeutic use , Insulin/therapeutic use , Lipase/blood , Adult , Aorta/enzymology , Endothelium, Vascular/enzymology , Female , Humans , Hypoglycemic Agents/administration & dosage , Insulin/administration & dosage , Lipase/genetics , Lipase/metabolism , Male , Middle Aged , RNA, Messenger/analysis , RNA, Messenger/metabolismABSTRACT
The lectin-like oxidized low density lipoprotein receptor-1 (LOX-1) can be proteolytically cleaved and released as soluble forms (sLOX-1). We have determined serums LOX-1 in type 2 diabetes and evaluated the effect of glucose and advanced glycation end products (AGEs) on sLOX-1 in vitro and in vivo. Endothelial cells were incubated with glucose or AGEs, and sLOX-1 in cell medium was measured. Serum sLOX-1 was measured in 219 diabetic patients and 187 controls by ELISA. The effect of lowering glucose and AGEs on sLOX-1 was determined in 38 poorly controlled diabetic patients after improvement in glycemic control. Incubation of endothelial cells with AGE-BSA led to a dose-dependent increase in sLOX-1, whereas the effect of glucose on sLOX-1 was less marked. Serum sLOX-1 was 9% higher in diabetic patients compared with controls (P<0.01). In the poorly controlled patients, serum sLOX-1 decreased by 12.5% after improvement in glycemic control (P<0.05). The magnitude of reduction in sLOX-1 correlated with the improvement in hemoglobin A1c and AGEs but not with the reduction in oxidized LDL. sLOX-1 level is increased in type 2 diabetes. Both glucose and AGEs are important determinants of LOX-1 expression, and lowering glucose and AGEs leads to a reduction in sLOX-1.
Subject(s)
Diabetes Mellitus, Type 2/blood , Scavenger Receptors, Class E/blood , Female , Humans , Male , Middle Aged , SolubilityABSTRACT
BACKGROUND: Advanced glycation end products (AGEs) play an important role in the pathogenesis of diabetic complications. Recent data suggest that AGEs may also interfere with the function of HDL and the reverse cholesterol transport pathway. We have investigated whether serum AGE level is associated with impairment in the antioxidative capacity of HDL and in the ability of serum to induce cholesterol efflux in type 2 diabetic patients with and without nephropathy. METHODS: A total of 167 controls and 264 diabetic patients was recruited. The ability of serum to induce cellular cholesterol efflux and the capacity of HDL to inhibit LDL oxidation ex vivo was determined. Serum AGEs were assayed by competitive ELISA using a polyclonal rabbit antisera raised against AGE-RNase. RESULTS: Diabetic subjects were subdivided into three groups (normoalbuminuria, microalbuminuria and proteinuria). Serum AGEs were significantly increased in diabetic patients with microalbuminuria or proteinuria (P < 0.001). Cholesterol efflux was significantly decreased in all three groups of diabetic patients compared to controls (P < 0.001) whereas the antioxidative capacity of HDL was significantly impaired in patients with microalbuminuria or proteinuria (P < 0.01). No relationship between serum AGEs and cholesterol efflux was found. However, serum AGE concentration was significantly associated with the antioxidative capacity of HDL and this was partly due to the adverse effect of AGEs on paraoxonase-1 activity. CONCLUSION: In type 2 diabetic patients with incipient or overt nephropathy, increased serum concentration of AGEs was associated with impairment in the antioxidative capacity of HDL. Cholesterol efflux to serum was also reduced but was not related to serum AGEs.