Hormonal control of glucose production and pyruvate kinase activity in isolated rat liver cells: influence of hypothyroidism

Hormonal control of glucose production and of -pyruvate kinase activity has been measured in isolated liver cells from fed control and thyroidectomized rats. In hypothyroid rats, sensitivity to isoproterenol as measured by these parameters was increased: the apparent K_0.5 for isoproterenol-induced stimulation of glucose production decreased from 8.0 ± 3 × 10⁻⁶ M in control rats to 2.0 ± 0.2 × 10⁻⁸ M in hypothyroid rats (P < 0.001) and the apparent K_0.5 for inhibition of -pyruvate kinase was 5 ± 2 X 10⁻⁷ M vs. 7 ± 2 × 10⁻⁹ M (P < 0.001) in control and thyroidectomized rats, respectively. Utilisation of specific adrenergic antagonists confirmed increased β-adrenergic responsiveness in hypothyroid rats. This phenomenon was not reversed by 3 days of T₃ treatment (10 μg/100 g body weight). Sensitivity to the -agonist was unchanged by thyroid status. Stimulation of glucose production and inhibition of -pyruvate kinase activity by glucagon and their reversal by insulin were not affected by hypothyroidism. The dose-response curve to vasopressin and its maximal effect measured on stimulation of glucose production were unchanged in thyroidectomized rats. Thus, hypothyroidism produces a specific enhancement of liver β-adrenergic responsiveness without affecting sensitivity to glucagon, insulin and vasopressin.     

Glucose and insulin responses to intravenous glucose challenge in relatives of Nigerian patients with non-insulin-dependent diabetes mellitus

We analysed blood insulin and glucose concentrations before and during frequently sampled intravenous glucose tolerance tests (FSIGT) in 2 groups of Nigerian subjects: (A) Control group (n = 18), without a positive family history of diabetes mellitus, and (B) Experimental group (n = 16), comprising age-, sex- and body mass-matched first-degree relatives of patients with non-insulin-dependent diabetes mellitus (NIDDM). In comparison with Group A subjects, those in Group B had: (i) higher fasting plasma glucose level (mean ± S.E.M., 4.1 ± 0.1 vs. 3.8 ± 0.11 mmol/l, P < 0.05); (ii) similar fasting serum insulin levels (6.7 ± 5.0 vs. 5.8 ± 5.6 mU/l, P = NS); (iii) lower mean incremental area under the first-phase (t = 0–10 min) post-glucose challenge insulin curve (376.9 ± 8.8 vs. 435.6 ± 5.6 mU/min l⁻¹, P < 0.05); (iv) increased incremental area under the second-phase (t = 10–182 min) post-glucose challenge insulin curve (432.9 ± 11.5 vs. 161.3 ± 8.7 mU/min l⁻¹, P < 0.05); (v) reduced K_G rate constant of glucose elimination (0.97 ± 0.12 vs. 1.41 ± 0.12%/min, P < 0.05). These results suggest that the subjects with a positive family history of NIDDM have a reduced beta-cell insulin secretory reserve (from reduced first-phase insulin response), tendency to rebound hyperinsulinemia during the latter phase of the insulin secretory response, a degree of tissue insulin insensitivity (as evident from high fasting plasma glucose despite similar insulin levels) and a diminished glucose disposal rate, in comparison with subjects without a family history of NIDDM. These features predict subsequent development of diabetes and suggest that as in Caucasians, first-degree relatives of Nigerian patients with NIDDM are at greater risk for future development of the disease.     

In vivo and in vitro studies of GAD-antibody positive subjects with Type 2 diabetes: A distinct sub-phenotype

The purpose of this study was to determine if immune mechanisms in GAD positive patients’ contribute to the pathogenesis of a specific sub-type of Type 2 diabetes. GAD positive (n = 8) and GAD negative (n = 8) subjects diagnosed with Type 2 diabetes were matched for age, gender, body mass index, duration of diabetes and glycaemic control. All subjects underwent an insulin-modified frequently sampled intravenous glucose tolerance test to measure insulin sensitivity and insulin secretory function with minimal model analysis. In addition, BRIN-BD11 clonal β-cells were supplemented with patients’ sera to determine basal and alanine-stimulated insulin secretion and terminal complement complex (TCC) formation. Both groups were severely insulin resistant (0.56 ± 0.17 vs. 0.99 ± 0.33 10⁻⁴ min⁻¹/(μU ml⁻¹) for GADneg and GADpos, respectively) but the GAD negative subjects had a higher basal (87 ± 11 vs. 58 ± 14 pmol l⁻¹, p < 0.05) and glucose-stimulated insulin secretion (ΔAUCins 0.96 ± 0.12 vs. 0.60 ± 0.12 pmol/(l⁻¹ min), p < 0.05). In vivo measures of insulin secretion were negatively correlated with TCC formation, independent of antibody status. In conclusion, GAD positive subjects initially diagnosed with Type 2 diabetes are unable to compensate for insulin resistance due to more pronounced β-cell impairment. TCC formation may be partly responsible for the insulin secretory dysfunction associated with this specific sub-type of Type 2 diabetes.     

Neurotransmitters partially restore glucose sensitivity of insulin and glucagon secretion from perfused streptozotocin-induced diabetic rat pancreas

Summary To elucidate the mechanisms of insensitivity of hormone secretion to glucose in streptozotocin-induced diabetic rat islets, we investigated the effects of acetylcholine (ACh) and norepinephrine on insulin and glucagon secretion in response to changes in glucose concentration, using perfused pancreas preparations. Basal insulin secretion at a blood glucose level of 5.6 mmol/l was significantly higher and basal glucagon secretion significantly lower in streptozotocin-induced diabetic rats than in controls, and neither high (16.7 mmol/l) nor low (1.4 mmol/l) blood glucose concentrations influenced insulin or glucagon secretion. Addition of 10^–6 mol/l ACh to the perfusate increased glucose-stimulated insulin secretion. Also, 10^–6 mol/l ACh, 10^–7 mol/l norepinephrine, as well as a combination of both, induced marked glucagon secretion, this was suppressed by high blood glucose level. Although simultaneous addition of 10^–6 mol/l ACh and 10^–7 mol/l norepinephrine induced only a slight increase in glucagon secretion in response to glucopenia, there was a significant increase in glucagon secretion in conjunction with an ambient decrease in insulin. Histopathological examination revealed a marked decline in acetylcholinesterase and monoamine-oxidase activities in the islets of streptozotocin-induced diabetic rats. We speculate that reduction of the potentiating effects of ACh and norepinephrine lessens glucose sensitivity of islet beta and alpha cells in this rat model of diabetes.Abbreviations STZ Streptozotocin - STZD streptozotocin-induce diabetic - ACh acetylcholine - AChE acetylcholinesterase - NE norepinephrine - MAO monoamine-oxidase     

Reduction of urinary albumin excretion by thromboxane synthetase inhibitor, OKY-046, through modulating renal prostaglandins in patients with diabetic nephropathy

We evaluated the effects of thromboxane synthetase inhibitor, OKY-046, on urinary albumin and prostaglandin (PG) excretion in 14 patients with non-insulin-dependent diabetes mellitus (NIDDM).

Urinary excretion of 6-keto-PGF₁ (a stable metabolite of PGI₂), TXB₂ (a stable metabolite of TXA₂) and PGE₂ in NIDDM patients was comparable with that in control subjects. However, the urinary 6-keto-PGF₁/TXB₂ ratio in NIDDM patients with both micro- and macroalbuminuria was significantly (P < 0.001) lower than that in the controls.

By a single administration of OKY-046 (40 mg, i.v.) to the diabetic patients, urinary TXB₂ excretion significantly (P < 0.05) decreased from 169.7 ± 23.9 to 140.2 ± 17.9 ng/gCr, but urinary 6-keto-PGF₁ and PGE₂ excretion did not change significantly. The urinary 6-keto-PGF₁/TXB₂ ratio thus significantly (P < 0.01) increased from 1.02 ± 0.13 to 1.73 ± 0.41 as associated with significant increments in urine volume (P < 0.05), urinary sodium excretion (P < 0.01) and creatinine clearance (P < 0.05).

Of 14 diabetic patients, 7 with macroalbuminuria (albumin index exceeding 100 mg/gCr) were orally given OKY-046 (600 mg/day) for 8 weeks. After this period, the urinary albumin index significantly (P < 0.05) decreased from 524.9 ± 149.6 to 317.6 ± 90.6 mg/gCr. Urinary PG excretion did not change significantly, although the urinary 6-keto-PGF₁/TXB₂ ratio significantly (P < 0.01) increased from 1.33 ± 0.16 to 2.42 ± 0.65 and decreased to 1.09 ± 0.15 (P < 0.01) following termination of this drug. Similarly, creatinine clearance significantly (P < 0.01) increased, from 56.3 ± 14.1 to 69.4 ± 15.1 ml/min. During the period of 8 weeks, no significant alteration occurred in systemic blood pressure or glycemic control.

In conclusion, OKY-046 has a beneficial effect on albuminuria in diabetic nephropathy by modulating altered renal PGs synthesis, which leads to a change in renal hemodynamics.     

Binding sites for peptide-histidine-isoleucine (PHI) on rat insulinoma-derived RINm5F cells

Specific binding sites for ¹²⁵I-labelled rat peptide-histidine-isoleucine (PHI) were identified on rat insulinoma-derived RINm5F cells. The concentrations of peptides producing half-maximal displacement of label were rat PHI, 0.36 ± 0.14 nM, vasoactive intestinal polypeptide (VIP), 0.38 ± 0.13 nM and secretin, approximately 0.2 μM. Glucagon and glucagon-like peptide-1(7–36)amide were without effect on binding. PHI and VIP produced dose-dependent increases in cAMP production in the cells that were significantly (P < 0.05) above unstimulated rates for ligand concentrations between 10⁻⁸ and 10 ⁻⁶ M. Both PHI and VIP produced a small but significant (P < 0.05) enhancement in the rate of release of immunoreactive insulin from the cells but the effect was not dose dependent.     

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/− 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±0.01mg/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/μg total RNA P<0.001) as was hepatic MTP mRNA expression (6.9±3.5 vs. 3.4±1.5 amol MTP mRNA/μg 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.     

Effects of the angiotensin II antagonist losartan on endothelin-1 and norepinephrine plasma levels during cold pressor test in patients with chronic heart failure

We evaluate the acute hemodynamic and neurohormonal effects of losartan in 15 patients with symptomatic chronic heart failure (CHF), mean age 72 ± 8 years, which were classified in two subgroups: (A) Patients with left ventricular ejection fraction (LVEF) ≤0.35 (n = 7); (B) subjects with LVEF >0.35 (n = 8). Sympathetic reactivity (blood pressure, heart rate and plasma norepinephrine) and plasma endothelin-1 (ET-1) were evaluated by a cold pressor test (CPT). Single doses of losartan (50 mg p.o.) lowered delta DBP in both subgroups (A, 8 ± 9 to 0 ± 5 mmHg, P < 0.05; B, 10 ± 6 to 3 ± 4 mmHg, P < 0.05) and attenuated the rise of HR in patients with mild (4 ± 6 to −1 ± 2 bpm, P < 0.05) but not with severe (4 ± 5 to 2 ± 5 bpm, n.s.) impairment of left ventricular function. Losartan blunted the response (delta) of PNE during CPT (A, 142 ± 131 to 10 ± 74 pg/ml, P < 0.05; B, 129 ± 72 to 1 ± 144 pg/ml, P < 0.01). A significant rise in plasma ET-1 was observed during CPT in patients from subgroup B (0.64 ± 0.40 to 0.81 ± 0.40 fmol/ml, P < 0.05) but not in patients with LVEF ≤0.35 (1.79 ± 0.44 to 1.51 ± 0.66 fmol/ml, n.s.). Losartan attenuated the rise in ET-1 during CPT in patients with LVEF >0.35 (delta ET-1 0.17 ± 0.86 to 0.03 ± 0.11 fmol/ml, P < 0.05), with no significant changes in subgroup A. Acute effects of losartan were characterized by a more favorable hemodynamic and neurohumoral response in patients with chronic heart failure and preserved systolic ventricular function related to subjects with lower ejection fractions.     

Dynamic change in collateral flow associated with myocardial ischemia in humans

Background: This study sought to investigate how collateral flow changes during myocardial ischemia in patients. Methods: Myocardial contrast echocardiography (MCE) and rapid atrial pacing were performed in 20 patients with angiographically evidenced coronary collaterals from the right coronary artery (RCA) to the occluded left anterior descending coronary artery. Sonicated contrast medium was injected into the RCA before and immediately after atrial pacing to determine the peak background-subtracted contrast intensity (PI) in the collateral territory (PIA) and its ratio to PI in the control territory (PI ratio) as parameters of collateral blood flow. Lactate production in the coronary circulation during pacing was determined to assess myocardial ischemia in the collateral territory. Results: PIA showed a significant correlation with regional wall motion either before (r(squared)=−0.64, P<0.01) or after pacing (r(squared)=−0.65, P<0.01). Similarly, PI ratio was significantly correlated with regional wall motion either before (r(squared)=−0.54, P<0.05) or after pacing (r(squared)=−0.64, P<0.01). Rapid atrial pacing decreased both PIA and PI ratio significantly greater in patients with lactate production than in those without (PIA: −67±53 vs. −15±34%, P<0.05; PI ratio: −68±49 vs. −8.2±32%, P<0.05, respectively), while neither PIA nor PI ratio differ between the two groups of patients before pacing (PIA: 13.8±19. vs. 16.2±13.3U, P=0.75; PI ratio: 0.70±0.71 vs. 0.87±0.65, P=0.58, respectively). Conclusions: We concluded that (1) collateral flow determined by MCE was closely associated with regional cardiac function, and (2) not the amount of collateral flow at rest, but pacing-induced change of collateral flow seemed to be a determinant of regional ischemia in patients with coronary collaterals.     

Inhibition of lipoprotein lipase induced cholesterol ester accumulation in human hepatoma HepG2 cells

It has been suggested previously that lipoprotein lipase may act as a ligand to enhance binding and uptake of lipoprotein particles. In the present study we have examined the capacity of bovine milk lipoprotein lipase to induce intracellular accumulation of triglyceride and cholesterol ester by VLDL (S_f 60–400) isolated from Type IV hypertriglyceridemic subject (HTg-VLDL) in HepG2 cells, independent of its lipolytic activity. We have also attempted to elucidate the cellular receptor mechanisms responsible for these effects. HTg-VLDL-mediated increases in intracellular triglyceride and cholesterol ester were dependent on the presence of an active lipase. Bovine milk lipoprotein lipase (LPL) increases triglyceride mass by 301% ± 28% (P < 0.0005) and cholesterol ester mass by 176% ± 12% (P < 0.0005). These HTg-VLDL-mediated increases in intracellular triglyceride and cholesterol ester did not occur when heat-inactivated lipase was used. Rhizopus lipase could replace LPL and cause equivalent increases in intracellular triglyceride and cholesterol ester (472% ± 61% (P < 0.005) and 202% ± 25% (P < 0.025) respectively vs. control). HTg-VLDL treated with LPL and reisolated also caused equivalent increases (274% ± 18% (P < 0.01) and 177% ± 12% (P < 0.005) for triglyceride and cholesterol ester). LDL also caused increases in intracellular cholesterol ester (189% ± 20% (P < 0.005)), although three times more LDL cholesterol had to be added to achieve the same effect. These LDL-induced increases were effectively blocked by monoclonal antibodies directed against the B,E receptor binding domains of apo B (−97% ± 13% (P < 0.0005) with anti-apo B 5E11 and − 68% ± 13% (P < 0.05) for anti-apo B B1B3) or by anti-B,E receptor antibodies (− 77% ± 7% (P < 0.01) antibody C7). These same antibodies had little effect on the HTg-VLDL + LPL-induced increases in cholesterol ester (+21%, + 15% and − 22% for 5E11, B1B3 and C7, respectively). Monoclonal anti-apo E antibodies also had no effect on LDL-mediated increases in intracellular cholesterol ester, but had a small and significant effect on VLDL-mediated increases in cholesterol ester. However, heparin, which interferes with cell surface proteoglycan interaction, was very effective at blocking HTg-VLDL-mediated increases in cholesterol ester in the presence of LPL (− 86% ± 8% P < 0.0005). Heparin was also effective in the presence of Rhizopus lipase (−79%) or lipolyzed re-isolated HTg-VLDL (−95%). These results suggest that lipoprotein lipase may enhance the uptake process beyond its role in lipolytic remodelling but does not appear to be an absolute requirement. In contrast, heparin had no effect on LDL-mediated cholesterol ester accumulation. Lactoferrin, which inhibits interaction with the low density lipoprotein receptor-related protein (LRP), was also very effective at inhibiting HTg-VLDL increases in intracellular cholesterol ester (− 95% ± 6%, P < 0.01). However, there was no effect of either heparin or lactoferrin on HTg-VLDL-mediated triglyceride accumulation. Thus cell surface heparin sulphate may facilitate intracellular lipid acquisition by providing a stabilizing bridge with the lipoproteins and enhance uptake through receptor-mediated processes such as LRP.     

Differential effects of glucagon-like peptide-1 (7-36)amide versus cholecystokinin on arginine-induced islet hormone release in vivo and in vitro

We compared the effects of two incretin hormones, glucagon-like peptide-1 (7-36)amide (GLP-1) and cholecystokinin (CCK), on islet hormone secretion. GLP-1 strongly potentiated glucose-stimulated insulin secretion in the perfused rat pancreas and in vivo in mice (p < 0.001). In contrast, GLP-1 did not enhance arginine-induced insulin release under these experimental conditions. In the perfused rat pancreas, GLP-1 also potentiated glucose-stimulated somatostatin secretion but, again, had no effect on arginine-induced somatostatin release. However, GLP-1 promptly inhibited the arginine-induced glucagon release (p < 0.02). In contrast, CCK enhanced insulin release in response to arginine both in the perfused rat pancreas and in vivo in mice (p < 0.001). In conclusion, GLP-1, in contrast to CCK, failed to enhance arginine-induced insulin release both in vitro and in vivo. This suggests that a signal generated by nutrient metabolism is required for the potentiation of insulin secretion by GLP-1. Furthermore, GLP-1 directly inhibited arginine-induced glucagon release as no concurrent increase in insulin or somatostatin release was noted.     

Increased responsiveness to the hyperglycemic, hyperglucagonemic and hyperinsulinemic effects of circulating norepinephrine in ob/ob mice

OBJECTIVE: Several studies have implicated increased sympathetic tone as a contributing factor to the hyperglycemia and hyperglucagonemia of ob/ob mice. However, the responsiveness of plasma glucose, insulin and glucagon to circulating norepinephrine (NE) in ob/ob vs normal lean mice has never been described. Therefore, the present study investigated the effect of a 15 min intravenous NE infusion (1 pmol/min/g) on plasma glucose, insulin and glucagon in anesthetized lean, ob/ob, ob/ob-concurrent yohimbine (alpha(2) antagonist) treated, and ob/ob-chronically sympatholytic dopamine agonist treated (for 14 days prior to infusion) mice. In an effort to gain insight into a possible relation between norepinephrine, hyperglucagonemia and hyperinsulinemia in ob/ob mice, this study also examined the isolated islet responses to NE and glucagon in lean, ob/ob and ob/ob-sympatholytic dopamine agonist treated mice. RESULTS: Basal humoral values of glucose, insulin and glucagon were all elevated in ob/ob vs lean mice (by 63, 1900 and 63%, respectively, P<0.01). However, NE infusion further increased levels of glucose, insulin and glucagon in ob/ob (by 80, 90 and 60%, respectively, P<0.05) but not in lean mice (between group difference for all parameters P<0.05). Acute concurrent yohimbine treatment as well as chronic prior sympatholytic dopamine agonist treatment (bromocriptine plus SKF38393) simultaneously strongly aborgated or abolished all these humoral hypersensitivity responses to intravenous NE in ob/ob mice (P<0.05). Clamping the plasma glucose level in untreated ob/ob mice at a high level (30 mM) established by NE infusion did not significantly alter the plasma insulin level, suggesting that some other influence of NE was responsible for this insulin effect. Direct NE administration at 1 microM to islets from lean and ob/ob mice inhibited 15 mM glucose-stimulated insulin secretion in both groups, but at 0.1 microM it was inhibitory only in islets from ob/ob mice. However, glucagon (10 nM) increased 15 mM glucose-stimulated insulin secretion in ob/ob (by 170%, P<0.05) but not lean mice (between group difference P<0.05). CONCLUSION: These findings suggest that hypersensitivity to circulating NE may potentiate hyperglycemia and hyperglucagonemia in ob/ob mice, and the subsequent hyperglucagonemia coupled with increased islet beta-cell insulin secretory responsiveness to glucagon in ob/ob mice may support hyperinsulinemia, thus explaining the increased plasma insulin level response to intravenous NE in these animals. These findings further support a role for increased peripheral noradrenergic activities in the development and maintenance of the hyperglycemic, hyperglucagonemic and hyperinsulinemic state, characteristic of type 2 diabetes.     

Reactive oxygen species in essential hypertension and non–insulin-dependent diabetes mellitus

To evaluate whether increased levels of reactive oxygen species (ROS) are involved in the pathogenesis of essential hypertension (EH) and non–insulin-dependent diabetes mellitus (NIDDM), both resting and stimulated levels of intracellular ROS were measured in lymphocytes from patients with EH (n = 10), NIDDM (n = 16) and age-matched healthy individuals (control subjects, n = 19). ROS was monitored with the dye, dihydrorhodamine-123 (DHR; 1 μmol/L) in the presence or absence of superoxide dismutase (superoxide scavenger), sodium azide (singlet oxygen/hydrogen peroxide scavenger), genistein (tyrosine kinase inhibitor), or bisindolylmaleimide (protein kinase C inhibitor). Simultaneous monitoring of cytosolic [Ca²⁺]_i was done with fura-2. Resting ROS levels were significantly higher in NIDDM (4.71 ± 0.25 nmol/10⁶ cells; mean ± SEM, P < .05) compared with EH (4.03 ± 0.22 nmol/10⁶ cells) or controls (4.05 ± 0.15 nmol/10⁶ cells). The formyl-Met-Leu-Phenylalanine-(fMLP)–induced ROS generation was significantly higher in NIDDM (21.92 ± 2.23 nmol/10⁶ cells; P < .05) compared with EH (14.58 ± 1.90 nmol/10⁶ cells) or control (16.06 ± 1.22 nmol/10⁶ cells). The fMLP-induced ROS increase was significantly reduced in the presence of sodium azide in all groups (P < .01) but was largely unaffected in the presence of SOD. Genistein and bisindolylmaleimide significantly inhibited the fMLP-induced ROS in all groups. The fMLP-induced [Ca²⁺]_i increase was significantly higher in NIDDM (71 ± 12 nmol/L, P < .01) compared with EH (42 ± 4 nmol/L) and control subjects (35 ± 3 nmol/L). Phytohemagglutinin was more effective in increasing [Ca²⁺]_i than ROS. It is concluded that ROS may play a role in the metabolic syndrome of NIDDM but not in EH.     

Improved right ventricular systolic time intravals after digitalis in patients with cor pulmonale and chronic obstructive pulmonary disease

Although digitalis has been used to treat patients with cor pulmonale secondary to chronic obstructive pulmonary disease, its effect on right ventricular performance has not been conclusively determined. This study assessed the effects of acute digitalization on measurement of right ventricular systolic time intervals in patients with chronic obstructive pulmonary disease and cor pulmonale. The intervals were recorded before and 40 minutes after administration of ouabain, 1 mg intravenously, in nine men (mean age 58 ± 5 [standard deviation] years) with chronic obstructive pulmonary disease (mean maximal mid expiratory flow rate 0.29 ± 0.08 1 liters/sec) and electrocardiographic evidence of right ventricular hypertrophy.

Ouabain produced significant reductions in right ventricular systolic time intervals, including the right ventricular preelection period (from 117 ± 23 to 102 ± 16 msec; P < 0.01), right ventricular ejection time index (from 397 ± 33 to 375 ± 24 msec; P < 0.01) and mean Q-P₂ index (from 509 ± 23 to 474 ± 8 msec; P < 0.001). In eight patients with simultaneously measured left ventricular systolic time intervals, similar changes were observed, including shortening of the left ventricular preejection period index (from 135 ± 9 to 117 ± 11 msec; P < 0.01), ejection time index (from 395 ± 18 to 379 ± 20 msec; P < 0.01), and the mean Q-A₂ interval (from 530 ± 16 to 495 ± 16 msec; P < 0.001). There were no significant changes in aortic or pulmonary arterial pressures. The results demonstrate that acute administration of digitalis produces significant improvement in right ventricular performance in patients with chronic obstructive pulmonary disease and cor pulmonale. The simultaneous shortening of right and left ventricular systolic time intervals is of comparable magnitude.     

Preconditioning does not prevent postischemic dysfunction in aging heart

Objectives. This study was performed to investigate the effect of single or multiple brief periods of ischemia and the administration of exogenous norepinephrine before a more prolonged ischemic period and after reperfusion in adult and senescent isolated and perfused rat hearts.

Background. The mortality rate for coronary artery disease is greater in the elderly. Ischemic preconditioning has been proposed as an endogenous form of protection against ischemia-reperfusion injury. However, the role of preconditioning in aging heart is unknown.

Methods. Compared the protective effect of preconditioning transient ischemic and norepinephrine stimuli against 20 min of global normothermic ischemia and 40 min of reperfusion in isolated perfused hearts of adult (6 months old) and senescent (24 months old) rats. Norepinephrine release in coronary effluent was determined by high performance liquid chromatography.

Results. Final recovery of percent developed pressure were improved after single preconditioning transient ischemic and norepinephrine stimuli in adult hearts (87.7 ± 9% and 82.3 ± 8.7%) versus unconditioned control hearts (50.6 ± 4.8%, p < 0.01 [mean ± SD]). The effect of preconditioning on developed pressure recovery was not present in senescent hearts after transient ischemic stimulus (39.8 ± 4.9% vs. 41.6 ± 5.8%, P = NS) but was present after norepinephrine stimulus (74.3 ± 10.5, p < 0.01). Norepinephrine release significantly increased after preconditioning transient ischemic stimulus in adult but not in senescent hearts (p < 0.01 vs. adult). Transient ischemic- and norepinephrine-induced preconditioning was blocked by alpha-adrenergic receptor antagonists in both adult and senescent hearts. Multiple transient ischemic stimuli were able to reduce postischemic dysfunction in adult but not in senescent hearts.

Conclusions. Preconditioning transient ischemic stimulus significantly reduces postischemic dysfunction in adult but not in senescent hearts, whereas exogenous norepinephrine is able to mimic preconditioning in both adult and senescent hearts. Ischemic preconditioning induces an increase in norepinephrine release in adult but not in senescent hearts. Preconditioning induced by transient ischemic stimulus and norepinephrine was abolished by alpha-adrenergic receptor blockade in both adult and senescent hearts. Thus, our data demonstrate that preconditioning is absent in aging heart and is probably related to the reduction of norepinephrine release and alpha-adrenergic receptor stimulation in response to ischemic preconditioning.     

Study of the rate of early glucose disappearance following insulin injection: insulin sensitivity index

The euglycaemic hyperinsulinaemic glucose clamp is usually considered as the reference technique to evaluate insulin sensitivity. As it is an-expensive and time-consuming tool, we therefore tried to validate a simple insulin tolerance test (ITT) (IV bolus of 0.1 IU/kg of regular insulin, with glucose sampling at −5, 0, 3, 5, 7, 10 and 15 min) and to demonstrate its usefulness. Insulin sensitivity was measured by DG/G0 ratio (G0 = initial glycaemia, DG is the variation between G0 and the glycaemia obtained at 15 min by the calculation of the regression plot). We confirmed the existence of a correlation between the glucose uptake (mg/kg per min) evaluated by glucose clamp and the DG/G0 index (r = 0.9, P < 0.01). There was no stimulation of hormonal counter regulation during the test. The ITT was significantly correlated both with fasting insulin (r = −0.43, P < 0.01), and post-glucose load insulin concentration (r = −0.67, P < 0.01); each measurement expressing insulin sensitivity. Four groups of patients with different insulin sensitivity; controls, NIDDM, gynoid and android obese subjects, were clearly separated by ITT. We showed that fasting glycaemia and DG/G0 were correlated (y = 2.63/x − 0.093; r = 0.82, P < 0.01). These results suggest that ITT could be an easy, quick and low cost method to evaluate insulin resistance in clinical practice and epidemiological studies.     

CETP is a determinant of serum LDL-cholesterol but not HDL-cholesterol in healthy Japanese

Cholesteryl ester transfer protein (CETP) is one of the factors that regulate plasma levels of HDL-cholesterol. To identify the factors that may regulate CETP activity, and to determine to what extent CETP is correlated with physiologic concentrations of lipoprotein, we performed an epidemiologic study in 586 healthy volunteers (317 males and 269 females, mean age 52.2 ± 10.9 years). CETP activity in these subjects was 192.96 ± 48.73 (mean ± S.D.) nmol/ml/h and distributed to a wide range (60–450 nmol/ml/h). Using multiple regression analysis, we found significant positive correlations between CETP activity and LDL-cholesterol (P < 0.03), apolipoprotein (apo) E (P < 0.005) and LCAT activity (P < 0.001). CETP activities showed significant negative correlation with apo A-I (P < 0.03). However, CETP activity showed no significant correlation either with HDL cholesterol or with apo B. One-way layout analysis of variance showed that alcohol drinking and cigarette smoking significantly reduced CETP activity, but there was no significant association between CETP activity and body mass index. Although CETP activities were significantly higher in females than in males (P < 0.001), multiple regression analysis showed no correlation between CETP activity and age in either the males or the females. Our results suggest that CETP activity regulates the concentration of apo A-I and LDL-cholesterol, and that such activity may be influenced by gender, alcohol consumption and cigarette smoking.     

Overexpression of glucagon-like peptide-1 receptor in an insulin-secreting cell line enhances glucose responsiveness

Glucagon-like peptide-1 (GLP-1), secreted from intestine in response to food intake, enhances insulin secretion from pancreatic β-cells. In this study, we evaluated the effects of stably transfecting the GLP-1 receptor into an insulinoma cell line, RIN 1046-38, on basal and glucose-mediated insulin secretion and on second messenger pathways involved in insulin secretion. The GLP-1 receptor transfected cells had similar insulin mRNA levels but higher insulin content compared with parental cells. In GLP-1 receptor transfected cells, glucose (0.5 mM)-mediated insulin release was increased compared with parental cells (4.52±0.79 pmol insulin/l per mg protein·h vs. 2.21±0.36 pmol insulin/l per mg protein·h; mean±S.E., n=6, P=0.015, in transfected vs. parental cells, respectively). By hemolytic plaque assay measuring single cell insulin secretion, we observed that in the GLP-1 receptor transfected cells versus parental cells the increased insulin secretion was due to the presence of more glucose-responsive cells as well as more insulin released in response to glucose per cell. Resting intracellular cAMP was higher in the GLP-1 transfected cells (35.96±3.88 vs. 18.6±2.01 nmol/l per mg protein·h; mean±S.E., n=4, P=0.039, in transfected vs. parental cells, respectively). In response to GLP-1, both GLP-1 receptor transfected cells and parental cells showed increased cAMP levels independent of glucose. Resting intracellular calcium was the same in both parental and GLP-1 receptor transfected cells. However, more cells were responsive to glucose in the GLP-1 receptor transfected cells and the calcium transients attained in the presence of glucose developed at a faster rate and reached a higher amplitude than in parental cells. We conclude that having an excess of GLP-1 receptors renders β-cells more sensitive to glucose.     

Endomyocardial biopsy in right ventricular cardiomyopathy

Right ventricular cardiomyopathy is characterized by a progressive myocyte loss and fibro-fatty substitution of the right ventricle. The aim of our study was to assess the diagnostic accuracy of right ventricular endomyocardial biopsy. Using an imaging analyser system, histomorphometric parameters of myocytes, interstitium, fibrous tissue and fatty tissue were evaluated on endomyocardial biopsy from 30 patients with arrhythmogenic right ventricular cardiomyopathy, 29 patients with dilated cardiomyopathy and 30 control patients. The percent area of myocytes decreased from 78.10 ± 7.34 in control to 63.39 ± 9.22 in dilated cardiomyopathy (P < 0.05) and to 47.28 ± 15.01 in arrhythmogenic right ventricular cardiomyopathy (P < 0.01). Fibrous tissue increased from 8.10 ± 3.89 in control to 21.80 ± 9.29 in dilated cardiomyopathy (P < 0.05) and to 24.60 ± 11.37 in arrhythmogenic right ventricular cardiomyopathy (P < 0.05). Fatty tissue varied from 0.33 ± 1.44 in control and 0.07 ± 0.31 in dilated cardiomyopathy to 13.30 ± 17.30 in arrhythmogenic right ventricular cardiomyopathy (P < 0.05). Fatty tissue was a feature of arrhythmogenic right ventricular cardiomyopathy (67% of patients vs. 6% of control and dilated cardiomyopathy patients). Diagnostic values typifying arrhythmogenic right ventricular cardiomyopathy, obtained by excluding any overlapping between confidence intervals in the three groups, were: myocytes <44.95%; fibrous tissue >40.38%, and fatty tissue >3.21%, with 67% sensitivity and 91.53% specificity for at least one parameter. In conclusion, a significant difference between arrhythmogenic right ventricular cardiomyopathy, dilated cardiomyopathy and control exists in terms of amount of myocytes, fibrous tissue and fatty tissue. Presence of fatty tissue and fibrous tissue exceeding 3.21% and 40.38%, respectively should be considered highly suspect for arrhythmogenic right ventricular cardiomyopathy in right ventricular endomyocardial biopsy.     

Insulin and glucagon secretion in essential fatty acid deficient rats

Summary Insulin and glucagon secretion in response to common secretagogues were ascertained in the perfused pancreas isolated from essential fatty acid deficient rats. The pattern of insulin secretory response to glucose (16.7 mmol/l) by isolated rat pancreas perfused for 30 min was biphasic in EFA-deficient and control rat pancreas. The amplitude of glucose-stimulated acute secretion (phase I) was significantly greater (p<0.01) in magnitude and amplitude in EFA-deficient rats than in the control rats. There was no significant difference in the second phase of glucosestimulated insulin secretion in the two groups. Glucagon secretion in EFA-deficient and control rats was inhibited by glucose (16.7 mmol/l). Glucagon secretion induced by L-arginine (10 mmol/l) was not significantly different in EFA-deficient and in control rat pancreata (p>0.05). However, arginine (10 mmol/l)-stimulated insulin release was significantly higher in EFA-deficient than in control rats. Growth hormone (100 nmol/l)-induced glucagon and insulin secretion was variable in the two groups but significantly higher than basal secretion. The level of L-leucine (10 mmol/l)-stimulated glucagon and insulin secretion in EFA-deficient rats was minimal but significant. Our results show that isolated pancreata of rats devoid of precursors for endogenous prostaglandin synthesis secreted insulin and glucagon in response to common secretagogues. On the basis of our data, it is concluded that endogenous prostaglandins are probably not obligatory for normal secretory functions of islets of Langerhans.