Inhibition of glucose production and stimulation of bile flow by R (+)-alpha-lipoic acid enantiomer in rat liver

AIMS/BACKGROUND: R (+)-alpha-lipoic acid (RLA) has been suggested for the treatment of liver diseases, but has also been shown to improve glucose utilization in diabetic patients. Because detailed information of RLA action on carbohydrate metabolism in intact liver is lacking, we examined concentration-dependent effects of RLA on hepatic glucose production. METHODS: RLA (10(-6-)10(-3) mol L(-1)) or buffer (control) was infused in isolated livers of fasted rats during recirculating perfusion for 90 min (n = 4-6/group). Hepatic glucose and lactate fluxes and bile secretion were continuously monitored. RESULTS: RLA reduced lactate (10 mmol L(-1))-dependent glucose production in concentration-dependent fashion (R = - 0.780, P < 0.001) by up to 67% compared with control (0.36 +/- 0.02 micromol min(-1) g(-1)). In parallel, RLA dose dependently decreased lactate uptake (R = - 0.592, P < 0.001) also by up to 67% (control: 0.58 +/- 0.08 micromol min(-1) g(-1)). RLA (10(-4) mol L(-1) and 10(-3) mol L(-1)) stimulated bile flow by approximately 20 and approximately 50%, respectively (P < 0.02 vs. control). After 10(-3) mol L(-1) RLA infusion, liver glycogen was approximately 3 fold higher (5.2 +/- 1.1 vs. control: 1.8 +/- 0.2 micromol g(-1), P < 0.002). Also at low lactate concentrations (1 mmol L(-1)), 10(-3) mol L(-1) RLA reduced glucose production by approximately 53% and lactate uptake by approximately 60%, but stimulated bile secretion by approximately 50% (P < 0.05). CONCLUSION: RLA reduces hepatic glucose release by inhibiting lactate-dependent glucose production in a concentration-dependent fashion.     

Regulation of amino acid metabolism and α‐cell proliferation by glucagon

Both glucagon and glucagon‐like peptide‐1 (GLP‐1) are produced from proglucagon through proteolytic cleavage. Blocking glucagon action increases the circulating levels of glucagon and GLP‐1, reduces the blood glucose level, and induces the proliferation of islet α‐cells. Glucagon blockade also suppresses hepatic amino acid catabolism and increases the serum amino acid level. In animal models defective in both glucagon and GLP‐1, the blood glucose level is not reduced, indicating that GLP‐1 is required for glucagon blockade to reduce the blood glucose level. In contrast, hyperplasia of α‐cells and hyperaminoacidemia are observed in such animal models, indicating that GLP‐1 is not required for the regulation of α‐cell proliferation or amino acid metabolism. These findings suggest that the regulation of amino acid metabolism is a more important specific physiological role of glucagon than the regulation of glucose metabolism. Although the effects of glucagon deficiency on glucose metabolism are compensated by the suppression of insulin secretion, the effects on amino acid metabolism are not. Recently, data showing a feedback regulatory mechanism between the liver and islet α‐cells, which is mediated by glucagon and amino acids, are accumulating. However, a number of questions on the mechanism of this regulation remain to be addressed. The profile of glucagon as a regulator of amino acid metabolism must be carefully considered for glucagon blockade to be applied therapeutically in the treatment of patients with diabetes.     

Stimulation of bile acid 6α-hydroxylation by rifampin

Background: Rifampin was shown to relieve pruritus in cholestatic liver diseases. There has been much speculation about the origin of pruritus, but it has not yet been comprehensively explained. The role of bile acids in producing pruritus is obscure and still under debate. Since rifampin both inhibits the uptake of bile acids into the hepatocyte and strongly induces mixed-function oxidases in the liver, the beneficial effects of this drug might be a consequence of altered bile acid metabolism.Methods: We investigated the influence of rifampin on urinary bile acid excretion with special respect to glucuronide and sulphate conjugates in 14 healthy volunteers before and after administration of rifampin, 600 mg×7 days, using each subject as his or her own control.Results: Bile acid glucuronide excretion increased from 0.55 to 1.19 μmol/24 h. This was in particular due to a significant increase of the urinary excretion of the 6α-hydroxylated hyocholic and hyodeoxycholic acids, the relative amounts of which accounted for about two thirds of the urinary bile acid excretion. Excretion of sulphates, however, decreased from 1.40 to 0.86 μmol/24 h due to a significantly reduced excretion of lithocholic acid sulphate. No changes in the excretion rates of other primary and secondary bile acids and no changes in their conjugation patterns were observed.Conclusions: The results provide evidence that rifampin induces 6α-hydroxylation of bile acids. The products are subsequently glucuronidated at the 6α-hydroxy groups, thus stimulating renal excretion of potentially toxic bile acids.     

Effects of non-esterified fatty acid availability on insulin stimulated glucose utilisation and tissue pyruvate dehydrogenase activity in the rat

Summary Fatty acids in cardiac muscle compete with glucose for oxidation, thereby inhibiting glucose utilisation. It is not clear whether a similar mechanism is important in resting skeletal muscle. We used the hyperinsulinaemic euglycaemic clamp technique in conscious rats fasted for 20 h to examine the effects of increased plasma non-esterified fatty acid levels (1 mmol/l) on glucose metabolism. Insulin was infused at 75 mU/h (plasma insulin, 2.27±0.21 g/l) or 300mU/h (16.41±0.47 g/l). An increase in non-esterified fatty acid levels decreased clamp glucose requirement and 3–³H-glucose turnover by 35% (p<0.001) when the higher insulin dose was used but there was no change at the lower dose. At both insulin infusion rates, clamp blood lactate and pyruvate responses suggested inhibition of muscle glycolysis by elevated plasma non-esterified fatty acid concentrations. Quadriceps muscle glycogen deposition during the clamps was enhanced by increased non-esterified fatty acid availability at the lower insulin dose (p<0.001) but not at the higher insulin concentration. Activation of pyruvate dehyrogenase during the clamps was partially inhibited by increased plasma non-esterified fatty acid in the heart, adipose tissue and quadriceps muscle. This was evident at both insulin levels in heart but only at the higher insulin concentration in muscle (p<0.002). The findings are consistent with an inhibition of glycolysis in skeletal muscle of mixed fibre type as a result of increased fatty acid availability. At low rates of glucose flux glycogen synthesis may compensate for decreased glycolysis so that glucose turnover is not decreased. The role of pyruvate dehydrogenase in the glucose-fatty acid cycle in muscle may depend on the prevailing plasma insulin concentration and the degree of activation of this enzyme.     

Efficacy of α‐glucosidase inhibitors combined with dipeptidyl‐peptidase‐4 inhibitor (alogliptin) for glucose fluctuation in patients with type 2 diabetes mellitus by continuous glucose monitoring

Aims/Introduction

The combination therapy of dipeptidyl‐peptidase (DPP)‐4 inhibitor and α‐glucosidase inhibitors (α‐GIs) is highly effective in suppressing postprandial hyperglycemia. The aim of the present study was to compare the effects of voglibose and miglitol on glucose fluctuation, when used in combination with DPP‐4 inhibitor by using continuous glucose monitoring (CGM).

Materials and Methods

In a randomized cross‐over study, 16 patients with type 2 diabetes who presented with postprandial hyperglycemia despite alogliptin (25 mg) were treated with voglibose (0.9 mg) or miglitol (150 mg). We measured standard deviation (SD); mean amplitude of glycemic excursions (MAGE), and mean, minimum and maximum glucose measured by CGM during three phases (alogliptin monotherapy, dual therapy of alogliptin and voglibose, and dual therapy of alogliptin and miglitol). The primary outcome measure was SD between α‐GIs.

Results

SD was significantly improved by the addition of either voglibose (18.9 ± 10.1) or miglitol (19.6 ± 8.2) to alogliptin monotherapy (36.2 ± 8.7). MAGE improved significantly with the addition of either voglibose (57.5 ± 26.1, P < 0.01) or miglitol (64.6 ± 26.2, P < 0.01) to alogliptin monotherapy (101.5 ± 21.5). There was no significant difference in glucose fluctuation between α‐GIs. There were no differences between two groups in mean (132.6 ± 21.4 and 138.7 ± 25.4) and maximum (184.3 ± 48.7 and 191.9 ± 38.3). The minimum glucose under alogliptin plus voglibose (94.9 ± 20.2) was significantly lower than that under alogliptin and miglitol (105.3 ± 21.0).

Conclusions

Glucose fluctuation was improved by the addition of voglibose or miglitol to alogliptin. Glucose fluctuations and postprandial hyperglycemia were similar between α‐GIs. This trial was registered with the University Hospital Medical Information Network (no. UMIN R000010028).     

Age‐dependent decline in β‐cell function assessed by an oral glucose tolerance test‐based disposition index

We evaluated age‐dependent changes in β‐cell function as assessed with an oral glucose tolerance test (OGTT)‐based analog of the disposition index (oral disposition index). A total of 110 Japanese normoglycemic subjects (aged 22–59 years) was divided into decadal age groups (20, 30, 40 and 50 s) and subjected to an OGTT. The oral disposition index was calculated as the product of the Matsuda index and the ratio of the area under the insulin curve to the area under the glucose curve for 0–120 min during the OGTT (AUC_ins/gluc120). Although indexes of insulin secretion, including AUC_ins/gluc120 and the insulinogenic index, did not differ among age groups, the oral disposition index differed significantly among decadal ages and declined with age. The oral disposition index is thus a sensitive measure of β‐cell function, and a natural decline in such function likely begins in early adulthood and progresses with age. (J Diabetes Invest, doi: 10.1111/j.2040‐1124.2010.00099.x, 2011)     

Effects of glycogen stores and non-esterified fatty acid availability on insulin-stimulated glucose metabolism and tissue pyruvate dehydrogenase activity in the rat

Summary The effects of increased tissue glycogen stores on insulin sensitivity, and on the response of insulin-stimulated glucose utilisation to an acute elevation in plasma fatty acid levels (1.5mmol/l), were investigated in conscious rats using the hyperinsulinaemic euglycaemic clamp. Studies were performed in two groups of rats; (a) fasted 24 h; (b) fasted 4.5 h, but infused with glucose for 4 h (0.5 g/h) of this period before the clamp (fed, glucose infused rats). Clamp glucose requirement and 3-³H-glucose turnover were 20–25% lower in the fed, glucose-infused rats. In these rats, elevation of plasma fatty acid levels resulted in impaired suppression of hepatic glucose output (residual hepatic glucose output: 41±4 vs 8±6 mol·min^–1·kg^–1. p < 0.001) but did not further decrease 3-³H-glucose turnover. Elevated nonesterified fatty acid levels had no significant effect on glucose kinetics in 24 h fasted rats. In the fed glucose-infused rats, at low plasma fatty acid levels, there was no deposition of glycogen in muscle during the clamp and liver glycogen levels fell. With elevation of non-esterified fatty acid levels muscle glycogen deposition was stimulated in both groups, and there was no fall in liver glycogen during the clamps in the fed glucose-infused rats. Increased non-esterified fatty acid availability during the clamps decreased pyruvate dehydrogenase activity in liver, heart, adipose tissue and quadriceps muscle, in both groups of rats. The findings are consistent with an inhibition of glycolysis in liver, skeletal muscle and heart by increased fatty acid availability. Increased glycogen synthesis, however, compensates for decreased glycolytic flux so that glucose turnover is not decreased. When liver glycogen stores are high, an acute increase in non-esterified fatty acid availability impairs suppression of hepatic glucose output. A chronic increase in non-esteriefid fatty acid availability may lead to insulin resistance by increasing glycogen stores.     

Effects of intranasal insulin on endogenous glucose production in insulin‐resistant men

The effects of intranasal insulin on the regulation of endogenous glucose production (EGP) in individuals with insulin resistance were assessed in a single‐blind, crossover study. Overweight or obese insulin‐resistant men (n = 7; body mass index 35.4 ± 4.4 kg/m², homeostatic model assessment of insulin resistance 5.6 ± 1.6) received intranasal spray of either 40 IU insulin lispro or placebo in 2 randomized visits. Acute systemic spillover of intranasal insulin into the circulation was matched with a 30‐minute intravenous infusion of insulin lispro in the nasal placebo arm. EGP was assessed under conditions of a pancreatic clamp with a primed, constant infusion of glucose tracer. Under these experimental conditions, compared with placebo, intranasal administration of insulin did not significantly affect plasma glucose concentrations, EGP or glucose disposal in overweight/obese, insulin‐resistant men, in contrast to our previous study, in which an equivalent dose of intranasal insulin significantly suppressed EGP in lean, insulin‐sensitive men. Insulin resistance is probably associated with impairment in centrally mediated insulin suppression of EGP.     

Inhibition of interleukin‐1β‐stimulated production of matrix metalloproteinases by hyaluronan via CD44 in human articular cartilage

Objective

To investigate the mechanism of the inhibitory action of hyaluronan (HA) on interleukin‐1β (IL‐1β)‐stimulated production of matrix metalloproteinases (MMPs) in human articular cartilage.

Methods

IL‐1β was added to normal and osteoarthritic (OA) human articular cartilage in explant culture to stimulate MMP production. Articular cartilage was incubated or preincubated with a clinically used form of 800‐kd HA to assess its effect on IL‐1β‐induced MMPs. Levels of secreted MMPs 1, 3, and 13 in conditioned media were detected by immunoblotting; intracellular MMP synthesis in chondrocytes was evaluated by immunofluorescence microscopy. Penetration of HA into cartilage tissue and its binding to CD44 were analyzed by fluorescence microscopy using fluoresceinated HA. Blocking experiments with anti‐CD44 antibody were performed to investigate the mechanism of action of HA.

Results

Treatment and pretreatment with 800‐kd HA at 1 mg/ml resulted in significant suppression of IL‐1β‐stimulated production of MMPs 1, 3, and 13 in normal and OA cartilage explant culture. Fluorescence histocytochemistry revealed that HA penetrated cartilage tissue and localized in the pericellular matrix around chondrocytes. HA‐binding blocking experiments using anti‐CD44 antibody demonstrated that the association of HA with chondrocytes was mediated by CD44. Preincubation with anti‐CD44 antibody, which suppressed IL‐1β‐stimulated MMPs, reversed the inhibitory effect of HA on MMP production that was induced by IL‐1β in normal and OA cartilage.

Conclusion

This study demonstrates that HA effectively inhibits IL‐1β‐stimulated production of MMP‐1, MMP‐3, and MMP‐13, which supports the clinical use of HA in the treatment of OA. The action of HA on IL‐1β may involve direct interaction between HA and CD44 on chondrocytes.