NAFLD exacerbates the effect of dietary sugar on liver fat and development of an atherogenic lipoprotein phenotype

This is the author accepted manuscript. It is currently under an indefinite embargo pending publication by Springer.${\bf Aims/hypothesis:}$ We aimed to test the hypothesis that the effects of dietary sugar on lipoprotein metabolism are influenced by non-alcoholic fatty liver disease (NAFLD). ${\bf Methods:}$ The effect of two 12 week, iso-energetic diets, high and low in non-milk extrinsic sugars (26% and 6% total energy), matched for macronutrient content, was examined in a randomised, cross-over study in men with NAFLD (n=11) and controls (n= 14). Lipoprotein kinetics and the sources of fatty acids for triacylglycerol (TAG) production were measured using stable isotope tracers. ${\bf Results:}$ Liver fat was higher after the high versus low-sugar diet in both groups (p<0.02), but men with NAFLD showed a relatively greater response than controls (p<0.05). After the high versus low-sugar diet, VLDL1-TAG production rate was higher in the controls (p <0.002) due to a greater contribution from splanchnic fatty acids (p<0.02) and de novo lipogenesis (p <0.002), whereas in NAFLD, VLDL2-TAG production rate was higher (p <0.05), due to a greater contribution from splanchnic fatty acids (p<0.02). There was no difference in the contribution of systemic NEFA to VLDL1 and VLDL2-TAG production rate between diets in either group. Intermediate density lipoprotein (IDL), LDL2 and LDL3-apolipoprotein B production rates and post-heparin hepatic lipase activity were all higher (p<0.05) on the high-sugar diet in NAFLD. ${\bf Conclusions:}$ A high sugar intake promoted a greater accumulation of liver fat in NAFLD than controls and increased VLDL-TAG production in both groups, due mainly to an increased contribution of fatty acids from splanchnic sources, which includes hepatic TAG storage pools. These effects may drive the formation of atherogenic lipoproteins.The work was supported by a UK government grant from the Biological Biotechnology Scientific Research Council (Grant no. BB/G009899/1); University of Surrey PhD scholarship for AM; Medical Research Council (body composition measurements) and infrastructure support from the National Institute of Health Research at the Cambridge Biomedical Research Centre

Lipolysis in Diabetic and Non-Diabetic Participants, Effects of Nutrition and Insulin Treatment.

Obesity, metabolic syndrome and type 2 diabetes are all characterised by insulin resistance and in most cases large intra-abdominal fat stores. Insulin resistance is associated with the dysregulation of adipose tissue (AT) Metab including a reduced effectiveness for insulin to suppress lipolysis. Although patients with type 2 diabetes are treated with metformin initially, the majority of patients eventually need treatment with insulin to maintain glucose control. Insulin treatment with most types of insulin therapy is associated with weight gain. However the insulin analogue detemir is weight neutral or causes weight loss and it has been suggested that this may be related to a reduced effect of this insulin in peripheral tissues such as AT. A high sugar intake (fructose and sucrose) is associated with insulin resistance but the effects on AT Metab are not established. Methods were developed to measure cell sizes, in vitro basal, stimulated and insulin inhibited lipolysis and lipoprotein lipase (LPL) activity in AT biopsies. These methods were then applied to study a) subcutaneous AT (SCAT) and omental AT (OMAT) biopsies from twelve participants, seven non-diabetic subjects and five patients with type 2 diabetes treated with metformin (T20H) undergoing abdominal surgery and b) the effect of treatment of patients with type 2 diabetes with insulin detemir (n=7) or insulin neutral protamine hagedorn (NPH) (n=5), in a 16 week parallel group study. The effect of sugar intake on the in vivo rate of lipolysis and post-heparin lipolysis was investigated in 25 men at risk of metabolic syndrome. In a cross-over study design participants were studied after a 12 week diet high in extrinsic sugars and low in extrinsic sugars. Adipocyte size of SCAT and OMAT was significantly larger in T20H than healthy participants (p<0. 001, p<0. 05 respectively). In healthy subjects OMAT had higher basal, isoproterenol stimulated and insulin inhibited lipolysis than SCAT (p<0. 01, p<0. 001, p<0. 05 respectively). In participants with T20H, stimulated lipolysis with isoproterenol in SCAT was significantly higher than OMAT, but there was no significant difference in basal and insulin inhibited lipolysis. There was no significant difference between healthy and T20H in SCAT lipolytic activity, but OMAT has higher basal, isoproterenol stimulated and insulin inhibited lipolysis than SCAT (p<0. 05, p<0. 001, p<0. 01 respectively) in healthy participants. LPL activity in OMAT was significantly higher than SCAT in both healthy and T20H group (p<0. 001,p<0. 05 respectively). In the insulin detemir clinical trial, fasting non-esterified fatty acids (NEFA) decreased significantly with detemir compared to NPH (p<0. 05). Basal lipolysis in the fat biopsies showed a significant reduction with detemir treatment (p<0. 05) with no significant change following NPH treatment. LPL mass and activity increased significantly after 16 weeks treatment with detemir compared to NPH (p=0. 006, p=0. 005). No significant differences were observed on stimulated lipolysis or adipocyte size following either of the treatments. In the dietary intervention study after the high sugar diet compared with low sugar diet, liver fat, fasting plasma glucose and NEFA concentrations were significantly higher in men with higher liver fat at screening (all p<0. 05). In the low-liver fat group, plasma TG (p=0. 07) NEFA (p=0. 04) and LPL activity (p<0. 05) were higher after the low versus high sugar diet, whereas, in the high-liver fat group, hepatic lipase (HL) activity was higher after the high versus the low sugar diet (p<0. 05). The palmitate (PA) concentration decreased significantly after the high sugar diet compared with low sugar diet (p<0. 001) in the high liver fat group. In this group, the percentage of palmitate to total NEFA in the circulation (PA:NEFA) decreased significantly after the high sugar diet, this value was significantly lower than the value after the high sugar diet in the low liver fat group (p<0. 05). However, the palmitate rate of appearance increased after the high sugar diet in the high liver fat group, but did not reach significance (p=0. 06). Conclusion: This study showed that OMAT had less sensitivity to insulin action than SCAT in T20H. In the clinical trial of insulin detemir, the decrease in basal NEFA levels and basal lipolysis and the increased LPL mass and activity suggests that detemir improves fasting insulin sensitivity. This may be due to a greater hepatoselective action of detemir reducing over-insulinisation of adipose tissue and skeletal muscle. The change in plasma lipid Metab and pattern of response in lipase activities in response to a diet high in extrinsic sugars increased the availability of NEFA from peripheral and endothelial lipolysis; this may be due to reduced insulin sensitivity in participants with high liver fat

Lipolysis in Diabetic and Non-Diabetic Participants, Effects of Nutrition and Insulin Treatment.

Obesity, metabolic syndrome and type 2 diabetes are all characterised by insulin resistance and in most cases large intra-abdominal fat stores. Insulin resistance is associated with the dysregulation of adipose tissue (AT) Metab including a reduced effectiveness for insulin to suppress lipolysis. Although patients with type 2 diabetes are treated with metformin initially, the majority of patients eventually need treatment with insulin to maintain glucose control. Insulin treatment with most types of insulin therapy is associated with weight gain. However the insulin analogue detemir is weight neutral or causes weight loss and it has been suggested that this may be related to a reduced effect of this insulin in peripheral tissues such as AT. A high sugar intake (fructose and sucrose) is associated with insulin resistance but the effects on AT Metab are not established. Methods were developed to measure cell sizes, in vitro basal, stimulated and insulin inhibited lipolysis and lipoprotein lipase (LPL) activity in AT biopsies. These methods were then applied to study a) subcutaneous AT (SCAT) and omental AT (OMAT) biopsies from twelve participants, seven non-diabetic subjects and five patients with type 2 diabetes treated with metformin (T20H) undergoing abdominal surgery and b) the effect of treatment of patients with type 2 diabetes with insulin detemir (n=7) or insulin neutral protamine hagedorn (NPH) (n=5), in a 16 week parallel group study. The effect of sugar intake on the in vivo rate of lipolysis and post-heparin lipolysis was investigated in 25 men at risk of metabolic syndrome. In a cross-over study design participants were studied after a 12 week diet high in extrinsic sugars and low in extrinsic sugars. Adipocyte size of SCAT and OMAT was significantly larger in T20H than healthy participants (p<0. 001, p<0. 05 respectively). In healthy subjects OMAT had higher basal, isoproterenol stimulated and insulin inhibited lipolysis than SCAT (p<0. 01, p<0. 001, p<0. 05 respectively). In participants with T20H, stimulated lipolysis with isoproterenol in SCAT was significantly higher than OMAT, but there was no significant difference in basal and insulin inhibited lipolysis. There was no significant difference between healthy and T20H in SCAT lipolytic activity, but OMAT has higher basal, isoproterenol stimulated and insulin inhibited lipolysis than SCAT (p<0. 05, p<0. 001, p<0. 01 respectively) in healthy participants. LPL activity in OMAT was significantly higher than SCAT in both healthy and T20H group (p<0. 001,p<0. 05 respectively). In the insulin detemir clinical trial, fasting non-esterified fatty acids (NEFA) decreased significantly with detemir compared to NPH (p<0. 05). Basal lipolysis in the fat biopsies showed a significant reduction with detemir treatment (p<0. 05) with no significant change following NPH treatment. LPL mass and activity increased significantly after 16 weeks treatment with detemir compared to NPH (p=0. 006, p=0. 005). No significant differences were observed on stimulated lipolysis or adipocyte size following either of the treatments. In the dietary intervention study after the high sugar diet compared with low sugar diet, liver fat, fasting plasma glucose and NEFA concentrations were significantly higher in men with higher liver fat at screening (all p<0. 05). In the low-liver fat group, plasma TG (p=0. 07) NEFA (p=0. 04) and LPL activity (p<0. 05) were higher after the low versus high sugar diet, whereas, in the high-liver fat group, hepatic lipase (HL) activity was higher after the high versus the low sugar diet (p<0. 05). The palmitate (PA) concentration decreased significantly after the high sugar diet compared with low sugar diet (p<0. 001) in the high liver fat group. In this group, the percentage of palmitate to total NEFA in the circulation (PA:NEFA) decreased significantly after the high sugar diet, this value was significantly lower than the value after the high sugar diet in the low liver fat group (p<0. 05). However, the palmitate rate of appearance increased after the high sugar diet in the high liver fat group, but did not reach significance (p=0. 06). Conclusion: This study showed that OMAT had less sensitivity to insulin action than SCAT in T20H. In the clinical trial of insulin detemir, the decrease in basal NEFA levels and basal lipolysis and the increased LPL mass and activity suggests that detemir improves fasting insulin sensitivity. This may be due to a greater hepatoselective action of detemir reducing over-insulinisation of adipose tissue and skeletal muscle. The change in plasma lipid Metab and pattern of response in lipase activities in response to a diet high in extrinsic sugars increased the availability of NEFA from peripheral and endothelial lipolysis; this may be due to reduced insulin sensitivity in participants with high liver fat