Diabetes and dyslipidemia: characterizing lipoprotein metabolism.

Premature atherosclerosis in diabetes accounts for much of the decreased life span. New treatments have reduced this risk considerably. This review explores the relationship among the disturbances in glucose, lipid, and bile salt metabolic pathways that occur in diabetes. In particular, excess nutrient intake and starvation have major metabolic effects, which have allowed us new insights into the disturbance that occurs in diabetes. Metabolic regulators such as the forkhead transcription factors, the farnesyl X transcription factors, and the fibroblast growth factors have become important players in our understanding of the dysregulation of metabolism in diabetes and overnutrition. The disturbed regulation of lipoprotein metabolism in both the intestine and the liver has been more clearly defined over the past few years, and the atherogenicity of the triglyceride-rich lipoproteins, and - in tandem - low levels of high-density lipoproteins, is seen now as very important. New information on the apolipoproteins that control lipoprotein lipase activity has been obtained. This is an exciting time in the battle to defeat diabetic atherosclerosis.

The different effect of pioglitazone as compared to insulin on expression of hepatic and intestinal genes regulating post-prandial lipoproteins in diabetes.

This study investigates lipoprotein composition in diabetes before and after treatment with insulin or pioglitazone and its relationship to gene expression of five genes found in liver and intestine which are involved in cholesterol homeostasis. Thirty zucker diabetic fatty fa/fa and 10 lean rats were examined. mRNA for 3-hydroxy3-methylglutaryl coenzyme A reductase (HMGCoA), microsomal triglyceride transfer protein (MTTP), Niemann Pick C1-like 1 (NPC1L1) and ATP binding cassette transporters (ABC) G5 and G8 was determined using real-time, reverse transcriptase (RT-PCR). Cholesterol, triglyceride, apo B48 and apo B100 were elevated in chylomicrons and very low density lipoproteins (VLDL) of untreated diabetic animals (p<0.02). For similar blood glucose pioglitazone was more effective than insulin in normalising the lipoproteins. In diabetic animals, HMGCoA reductase, MTTP and NPC1L1 mRNA were significantly elevated (p<0.02) and ABCG5 and ABCG8 were significantly reduced (p<0.02) in the liver. Pioglitazone significantly reduced hepatic MTTP and NPC1L1 mRNA (p<0.0001) and significantly increased ABCG5 and G8 mRNA (p<0.0001) as compared to insulin. In conclusion diabetes was associated with major changes in mRNA levels of proteins involved in the regulation of post-prandial lipoproteins. Pioglitazone and insulin have different effects on post-prandial lipoprotein metabolism in part due their effect on genes regulating cholesterol synthesis and lipoprotein assembly.

Messenger RNA levels of genes involved in dysregulation of postprandial lipoproteins in type 2 diabetes: the role of Niemann-Pick C1-like 1, ATP-binding cassette, transporters G5 and G8, and of microsomal triglyceride transfer protein.

AIMS/HYPOTHESIS: The aim of the present study was to examine the relationship between chylomicron composition and expression of genes that regulate chylomicron production in the intestine. We examined expression of the following: (1) Niemann-Pick C1-like 1 (NPC1L1), which regulates cholesterol absorption; (2) ATP-binding cassette transporters G5 and G8 (ABCG5, ABCG8), which regulate cholesterol homeostasis through their ability to excrete enterocyte cholesterol back into the lumen of the intestine; and (3) microsomal triglyceride transfer protein (MTTP), which packages the chylomicron particle by assembling cholesterol, triglyceride, phospholipids and apolipoprotein B48. SUBJECTS, MATERIALS AND METHODS: Type 2 diabetic (26) and non-diabetic (21) patients were examined. Levels of NPC1L1, ABCG5 and ABCG8 and MTTP mRNA were measured in duodenal biopsies by real-time PCR. Lipoproteins were isolated by sequential ultracentrifugation. RESULTS: Diabetic patients had more NPC1L1 mRNA than the control subjects (p<0.02). Expression of ABCG5 and ABCG8 mRNA was lower in the diabetic patients (p<0.05) and MTTP expression was increased (p<0.05). There was a positive correlation between NPLC1L1 and MTTP mRNA (p<0.01) and a negative correlation between NPC1L1 and ABCG5 mRNA (p<0.001). Diabetic patients on statin therapy had increased ABCG5 and ABCG8 mRNA compared to those not on statin (p<0.02 and p<0.05) and less MTTP mRNA than those not on statin (p<0.05). CONCLUSIONS/INTERPRETATION: This study demonstrates that in type 2 diabetes there are important alterations to the expression of intestinal genes that regulate cholesterol absorption and chylomicron synthesis. In diabetic patients statin therapy is associated with reduced MTTP expression and increased ABCG5 and ABCG8 mRNA. The study suggests new mechanisms to explain postprandial diabetic dyslipidaemia and the beneficial effect of statins.

Comparison of diets high in monounsaturated versus polyunsaturated fatty acid on postprandial lipoproteins in diabetes.

BACKGROUND: Postprandial hypertriglyceridaemia is common in diabetes. Fatty acids are regulators of gene expression and may play an important role in regulatingthe postprandial lipoprotein cascade. AIM: To examine postprandial lipoprotein differences between diabetic and control subjects on polyunsaturated (linoleic) and monounsaturated (oleic) fat diets. METHODS: A randomised, crossover study. RESULTS: LDL was greater and HDL less (p < 0.05) in diabetic patients compared to controls on the linoleic acid diet. Apo E per particle was significantly lower in the diabetic patients compared to control subjects on both linoleic (p < 0.05) and oleic acid diets (p < 0.01). HDL apo E was also significantly lower in the diabetic patients compared to controls on the linoleic acid diet (p < 0.02). CONCLUSION: A change from linoleic to oleic acid diet resulted in an improvement in LDL and HDL in the diabetic patients. We suggest that the reduced apo E/particle in the diabetic patients may be an explanation for the delayed lipoprotein clearance.

Microsomal triglyceride transfer protein polymorphisms and lipoprotein levels in type 2 diabetes.

BACKGROUND: Microsomal triglyceride transfer protein (MTP) regulates the assembly of chylomicrons in the intestine and very-low-density lipoprotein (VLDL) in the liver. Common polymorphisms have been described that do not affect lipoproteins in non-diabetic subjects. Their effect in diabetes has not been described in a Caucasian population. AIM: To investigate the association of these three common polymorphisms with lipoproteins in type 2 diabetes. METHODS: Eighty-two patients consumed a high-fat test meal. Chylomicron and VLDL apoB48, apoB100, cholesterol, triglycerides and phospholipids were measured fasting, and at 4 and 6 h postprandially. MTP genotyping was performed by PCR-RFLP. RESULTS: Thirty-three subjects were heterozygous for the -493 G/T substitution. These patients had significantly lower LDL cholesterol (3.0 +/- 0.2 vs. 3.5 +/- 0.1 mmol/l, p < 0.02). In the postprandial period, they had higher levels of apoB48 in the VLDL fraction (4 h, 7.0 +/- 1.4 vs. 2.9 +/- 0.4 microg/ml plasma, p < 0.002; 6 h, 6.4 +/- 1.0 vs. 3.5 +/- 0.5 microg/ml plasma, p < 0.05). In the VLDL fraction there was significantly less cholesterol at 4 and 6 h (p < 0.05). The -400 A/T substitution gave very similar lipoprotein results, but there was significant linkage dysequilibrium between the two polymorphisms. No association was found between the -164 T/C polymorphism and either plasma lipids or the postprandial lipid profile. ApoE genotype was also examined, but did not influence the above results. DISCUSSION: The common -493 G/T MTP polymorphism is associated with changes in VLDL and LDL in Type 2 diabetic patients. The importance of the changes in apoB48-containing small particles requires further investigation. The significantly lower LDL cholesterol suggests that this polymorphism may confer protection against atherosclerosis in type 2 diabetes.

Microsomal triglyceride transfer protein: does insulin resistance play a role in the regulation of chylomicron assembly?

We have previously demonstrated that diabetes is associated with an increase in intestinal microsomal triglyceride transfer protein (MTP) mRNA in both the rat and rabbit models. The present study was designed to investigate the relationship between MTP expression and chylomicron assembly in an insulin resistant non-diabetic animal model. Ten insulin resistant Zucker obese fa/fa rats and ten lean fa/minus sign rats were examined at 8-10 weeks of age. The lymph duct was cannulated and lymph collected for 4 h. Lymph chylomicrons were isolated by ultracentrifugation and their composition determined. RNA was extracted from intestinal mucosa and from the liver. MTP mRNA was measured using the RNase protection assay. Blood sugar in the fatty rats was significantly higher (6.3 +/-1.2 vs. 5.4 +/-0.4 P<0.05) and plasma insulin was almost six times that of the lean rats (P<0.001). Plasma cholesterol and phospholipid but not triglyceride were significantly increased in the obese animals (P<0.01). Obese animals secreted significantly more lymph chylomicron apo B48 (0.05 +/-0.02 vs. 0.02 +/-0.01 mg/h P<0.005), triglyceride (9.7 +/-5.3 vs. 3.8+/-1.9 mg/h P<0.005) and phospholipid (1.5 +/-0.7 vs. 0.4 +/-0.3 mg/h P<0.001). The only difference in the chylomicron particle composition between the two groups was a significant increase in phospholipid (P<0.01). Intestinal MTP mRNA expression was significantly higher in the fatty compared to the lean rats (22.1 +/-9.5 vs. 7.8+/-5.6 amol MTP mRNA/microg total RNA P<0.001) as was hepatic MTP mRNA expression (6.9 +/-3.5 vs. 3.4 +/-1.5 amol MTP mRNA/microg total RNA, P<0.01). Thus in this animal model of insulin resistance, increased MTP, which was associated with increased chylomicron particle number, may play a crucial role in the development of atherosclerosis.

Abnormalities in apo B-containing lipoproteins in diabetes and atherosclerosis.

Atherosclerosis is the major cause of death in patients with diabetes. Low-density lipoprotein (LDL) being the most important cholesterol-carrying lipoprotein has been studied extensively in both diabetes and non-diabetes. This paper reviews the literature but also focuses on the precursors of LDL and in particular the postprandial apo B-containing lipoproteins. Abnormalities in the postprandial lipoproteins and alteration in chylomicron assembly and clearance are discussed and the evidence presented suggesting the importance of dysregulation of these lipoproteins in atherosclerotic progression. The relationship between chylomicron production in the intestine and hepatic release of very low-density lipoproteins (VLDL) is explored, as is the interrelationship between clearance rates of these lipoproteins. The size of LDL influences its atherogenicity. VLDL composition and size in relation to its influence on LDL is discussed. The effect of diet on the composition of lipoproteins and the relationship between fatty acid composition and clearance is reviewed. Evidence that diabetic control beneficially alters lipoprotein composition is presented suggesting how improved diabetic control may reduce atherosclerosis. The review concludes with a discussion on the effect of the apo B-containing lipoproteins and their modification through glycation and oxidation on macrophage and endothelial function.

Dietary unsaturated fatty acids in type 2 diabetes: higher levels of postprandial lipoprotein on a linoleic acid-rich sunflower oil diet compared with an oleic acid-rich olive oil diet.

OBJECTIVE: The present study was undertaken to examine the effect of a polyunsaturated fat diet compared with an isocaloric Mediterranean-style monounsaturated fat diet. RESEARCH DESIGN AND METHODS: This was a randomized 2-week crossover study on either a high-polyunsaturated or a high-monounsaturated fat diet in 11 well-controlled diabetic men. Blood was taken fasting and for up to 8 h after a high fat meal. Lipoproteins were isolated by sequential ultracentrifugation. Apolipoprotein (apo) B48 and apo B100 were separated by PAGE. Fatty acids were analyzed by gas-liquid chromatography RESULTS: Fasting blood glucose and insulin levels were significantly higher on the linoleic acid diet compared with the oleic acid diet (P < 0.01 and P < 0.002, respectively). Plasma cholesterol and LDL cholesterol levels were also significantly higher on the linoleic acid diet (P < 0.001). Likewise, fasting chylomicron apo B48 and apo B100 (P < 0.05) and postprandial chylomicron and VLDL apo B48 and B100 (P < 0.05) were also higher on the linoleic acid diet. CONCLUSIONS: This study suggests that, in type 2 diabetes, an oleic acid-rich Mediterranean-type diet versus a linoleic acid-enriched diet may reduce the risk of atherosclerosis by decreasing the number of chylomicron remnant particles.

Postprandial apolipoprotein B48-and B100-containing lipoproteins in type 2 diabetes: do statins have a specific effect on triglyceride metabolism?

There is little information about the effect of an alteration of low-density lipoprotein (LDL) turnover on chylomicron and very-low-density lipoprotein (VLDL) metabolism, yet chylomicron remnant particles are thought to be particularly atherogenic. This study examined the effect of inhibition of cholesterol synthesis on postprandial lipoproteins. Eight type 2 diabetic patients were examined before treatment with the 3-hydroxy-3-methyl glutaryl coenzyme A (HMGCoA) reductase inhibitor cerivastatin, after 4 weeks on active treatment, and 4 weeks after stopping treatment. On each occasion, blood was collected fasting and at 2-hour intervals for up to 8 hours after a high-fat meal. Chylomicrons and VLDLs were isolated by sequential ultracentrifugation. Compositional analysis was performed including the measurement of apolipoprotein B48 (apo B48) and apo B100 using polyacrylamide gradient gel electrophoresis. During statin treatment, there was a significant reduction in the postprandial chylomicron apo B48 area under the curve (AUC) from 23 +/- 16 to 17 +/- 10 (P < .01) and apo B100 in the chylomicron fraction from 166 +/- 148 to 70 +/- 70 (P < .05). Postprandial cholesterol (362 +/- 193 to 74 +/- 39, P < .005), triglyceride (2,222 +/- 1,440 to 746 +/- 329), and phospholipid (518 +/- 267 to 205 +/- 94) also decreased (P < .005). In the VLDL fraction, the postprandial cholesterol and triglyceride AUC were significantly reduced by statin (316 +/- 228 to 171 +/- 78, P < .05, and 1,733 +/- 833 to 857 +/- 468, P < .02, respectively). Four weeks after cessation of treatment, the chylomicron fraction triglyceride AUC had returned to the pretreatment level, but postprandial chylomicron cholesterol and VLDL cholesterol, triglyceride, and phospholipid were significantly lower than baseline (P < .05). Plasma total cholesterol and LDL cholesterol were significantly reduced with treatment (6.2 +/- 0.5 to 4.3 +/- 1.0 mmol/L, P < .001, and 4.5 +/- 0.4 to 2.8 +/- 1.0 mmol/L, P < .01, respectively) and returned to baseline following cessation of treatment. Fasting plasma triglycerides decreased significantly on treatment (2.4 +/- 1.0 to 1.7 +/- 0.2 mmol/L, P < .05) but remained significantly lower than baseline 4 weeks later (1.8 +/- 0.3 mmol/L, P < .05). This study suggests major postprandial lipoprotein changes on statin therapy which may account, in part, for the beneficial effects of statins in the prevention of myocardial infarction.

Lipoproteins and low-dose estradiol replacement therapy in post-menopausal Type 2 diabetic patients: the effect of addition of norethisterone acetate.

AIMS: Low-dose continuous oestrogen/progestogen may increase patient compliance long-term but the cardioprotective effects in diabetes are unknown. The aim of this study was to compare the effect of low-dose oral oestrogen (1 mg, 17-beta-estradiol) treatment with oestrogen (1 mg 17-beta-estradiol) in combination with low-dose (0.5 mg) continuous norethisterone acetate (NETA) on lipoproteins in Type 2 diabetic patients. METHODS: Thirty-four post-menopausal Type 2 diabetic patients in moderate control (mean haemoglobin A1c 7.7%) who had a serum oestradiol level of < 50 pg/ml were examined over a 6-month period. Serum lipids, and lipoprotein composition of very low density lipoprotein (VLDL), low density lipoprotein (LDL) and high density lipoprotein (HDL) were measured. Serum lipoprotein(a) was determined by an ELISA method, LDL fatty acids by gas-liquid chromatography and LDL oxidizability by thiobarbituric acid reactive substances (TBARS assay). Cholesteryl ester transfer protein (CETP), and cell cholesterol were measured. RESULTS: There was a reduction in serum cholesterol on both treatments but no significant difference between treatment groups. LDL cholesterol decreased by 17% in each group. There was a no significant difference between the groups in serum VLDL or HDL cholesterol or serum triglycerides during the study. The change in lipoprotein(a) during the study was not significantly different between the groups. There was no significant difference in 4 h LDL oxidizability between groups. Although CETP increased with time in both groups there was no significant difference in the change between the groups. CONCLUSION: In this small study, the addition of continuous low-dose NETA did not reduce the potentially beneficial effects of low-dose 17-beta-estradiol on the progression of atherosclerosis in diabetes.

Diabetes and the Mediterranean diet: a beneficial effect of oleic acid on insulin sensitivity, adipocyte glucose transport and endothelium-dependent vasoreactivity.

Abnormalities in endothelial function may be associated with increased cardiovascular risk in diabetic patients. We examined the effect of an oleic-acid-rich diet on insulin resistance and endothelium-dependent vasoreactivity in type 2 diabetes. Eleven type 2 diabetic patients were changed from their usual linoleic-acid-rich diet and treated for 2 months with an oleic-acid-rich diet. Insulin-mediated glucose transport was measured in isolated adipocytes. Fatty acid composition of the adipocyte membranes was determined by gas-liquid chromatography and flow-mediated endothelium-dependent and -independent vasodilatation were measured in the superficial femoral artery at the end of each dietary period. There was a significant increase in oleic acid and a decrease in linoleic acid on the oleic-acid-rich diet (p<0.0001). Diabetic control was not different between the diets, but there was a small but significant decrease in fasting glucose/insulin on the oleic-acid-rich diet. Insulin-stimulated (1 ng/ml) glucose transport was significantly greater on the oleic- acid-rich diet (0.56+/-0.17 vs. 0.29+/-0.14 nmol/10(5) cells/3 min, p<0.0001). Endothelium-dependent flow-mediated vasodilatation (FMD) was significantly greater on the oleic-acid-rich diet (3.90+/-0.97% vs. 6.12+/-1.36% p<0.0001). There was a significant correlation between adipocyte membrane oleic/linoleic acid and insulin-mediated glucose transport (p<0.001) but no relationship between insulin-stimulated glucose transport and change in endothelium-dependent FMD. There was a significant positive correlation between adipocyte membrane oleic/linoleic acid and endothelium-dependent FMD (r=0.61, p<0.001). Change from polyunsaturated to monounsaturated diet in type 2 diabetes reduced insulin resistance and restored endothelium-dependent vasodilatation, suggesting an explanation for the anti-atherogenic benefits of a Mediterranean-type diet.

Improved metabolic control reduces the number of postprandial apolipoprotein B-48-containing particles in type 2 diabetes.

Postprandial lipoproteins are raised in diabetes and there is increasing evidence for the atherogenicity of the chylomicron remnant. Increased postprandial cholesteryl ester transfer has also been demonstrated in diabetes and may contribute to the atherogenic lipoprotein profile. The present study examined the effect of improving metabolic control on postprandial lipoproteins in 13 Type 2 diabetic patients. Blood was taken fasting and at 2-h intervals following a high fat, 1100 kcal meal. Patients were brought into good control by intensified dietary advice and oral hyperglycaemic agents or insulin if blood glucose failed to respond. Fasting and postprandial cholesteryl ester transfer protein (CETP) and lecithin:cholesteryl acyltransferase (LCAT) were determined in six patients. Lipoproteins were isolated by sequential ultracentrifugation. Chylomicron and very low density lipoprotein (VLDL) apolipoprotein B-48 and apolipoprotein B-100 were isolated by polyacrylamide gradient gel electrophoresis and quantified by densitometric scanning. CETP and LCAT were determined by an endogenous method which determined cholesterol esterification and transfer between the patients' lipoproteins. There was a significant reduction in postprandial chylomicron apo B-48 (P<0.005), apo B-100 (P<0.0005) and chylomicron cholesterol (P<0.001) following improved diabetic control. The chylomicron lipid/apo B ratio increased with improved control (P<0.01). Postprandial CETP and LCAT were significantly reduced in good control (P<0.01 and P<0.05, respectively) and there were significant changes in HDL composition. The study shows that improvement in metabolic control in Type 2 diabetic patients leads to a reduction in postprandial chylomicron particles and less transfer of cholesterol to apo B-containing lipoproteins.

The relationship between cholesterol absorption and intestinal cholesterol synthesis in the diabetic rat model.

The chylomicron remnant particle is thought to be particularly atherogenic and we have previously shown alterations in post-prandial lipoproteins which could contribute to their atherogenicity. Cholesterol metabolism is disturbed in diabetes, yet the effect of diabetes on intestinal cholesterol synthesis and absorption has rarely been investigated. The aim of this study was to examine cholesterol absorption and intestinal synthesis of cholesterol in the streptozotocin diabetic rat. Twelve diabetic rats were paired with 12 control rats. [14C]-Cholesterol emulsion was administered and the lymph duct was canulated. Lymph was collected for 4h. At sacrifice blood was taken for plasma lipoprotein measurements. Chylomicrons were prepared from the lymph by ultracentrifugation and [14C]-cholesterol content was determined by liquid scintillation counting. Lymph apolipoprotein B48 was isolated by gradient gel electrophoresis, and quantified by densitometric scanning. Serum triglyceride and cholesterol were greatly elevated in diabetic compared to control animals (260+/-90 and 9.8+/-8.0 mg/ml vs. 1.0+/-0.4 and 0.6+/-0.3 mg/ml, p < 0.0001 respectively). Lymph chylomicron apo B48 was similar in the two groups. Cholesterol absorption was not significantly different in diabetic compared to control rats but cholesterol synthesis was significantly higher in the diabetic animals (550+/-352 vs. 322+/-113 microg/h p < 0.03). There was a positive correlation between apo B48 and cholesterol absorption (r = 0.70, p < 0.01) in the diabetic rats and control rats (r = 0.71, p < 0.01) but no correlation between apo B48 and cholesterol synthesis in either group. This study demonstrates that cholesterol synthesis was increased in diabetes whereas cholesterol absorption was unaffected suggesting that intestinal cholesterol synthesis made an important contribution to the hypercholesterolaemia seen in the diabetic animals.

The role of microsomal triglyceride transfer protein and dietary cholesterol in chylomicron production in diabetes.

AIMS/HYPOTHESIS: The aim of this study was to examine factors involved in chylomicron production in the streptozotocin diabetic rat, our hypothesis being that the synthesis of the chylomicron is abnormal in diabetes. METHODS: Diabetic rats (n = 20) were paired with control rats (n = 20). Cholesterol emulsion was given by gavage and the lymph duct was cannulated. Lymph was collected for 4 h. Chylomicrons were prepared from the lymph by ultracentrifugation. Lymph apolipoprotein B48 was isolated by gradient gel electrophoresis and quantified by densitometric scanning. Intestinal microsomal triglycerol transfer protein mRNA was measured by solution hybridisation nuclease protection, using a rat specific [32P]-labelled cRNA probe. RESULTS: Serum triglyceride and cholesterol were greatly increased in diabetic compared with control animals (258 +/- 77 and 8.9 +/- 6.4 mg/ml vs 1.04 +/- 0.37 and 0.54 +/- 0.03 mg/ml, p < 0.0001). Lymph chylomicron triglyceride and cholesterol were also higher in diabetic rats (29.4 +/- 27.3 and 0.28 +/- 0.3 mg/h vs 16.8 +/- 10.6 and 0.18 +/- 0.09 mg/h, p < 0.05). Lymph chylomicron apo B48 was similar in the two groups. Intestinal microsomal triglycerol transfer protein mRNA was higher in the diabetic rats (12.6 +/- 3.2 vs 3.8 +/- 3.0 amol/microgram RNA, p < 0.0001) and there was a positive correlation between lymph triglyceride and microsomal triglycerol transfer protein mRNA in the whole group (r = 0.65, p < 0.01). CONCLUSION/INTERPRETATION: The study shows that microsomal triglycerol transfer protein mRNA is raised in diabetes without an increase in apolipoprotein B48 in the lymph suggesting that microsomal triglycerol transfer protein regulates chylomicron triglyceride content but not particle number.