Richard G. Larkins

Determinants of restenosis and lack of effect of dietary supplementation with eicosapentaenoic acid on the incidence of coronary artery restenosis after angioplasty

The effect of an eicosapentaenoic acid-rich encapsulated preparation of fish oil on the incidence of early restenosis after coronary angioplasty was assessed by a randomized double-blind placebo-controlled study. A total of 108 patients received either 10 capsules of fish oil (1.8 g eicosapentaenoic acid, 1.2 g docosahexaenoic acid) or 10 control capsules (50% olive oil, 50% corn oil), commencing the day before angioplasty and continuing for 4 months after angioplasty, in addition to treatment with aspirin and verapamil. In 101 (94%) of the 108 patients, follow-up angiographic or postmortem result was evaluated at a mean (+/- SD) of 100 (+/- 22) days. Angiographic restenosis was observed in 34% of patients (29% of lesions) in the fish oil-treated group and 33% of patients (31% of lesions) in the control group (no significant difference). The overall incidence of angiographic restenosis was significantly higher in patients with 1) recurrent angina pectoris, 2) a positive exercise test at follow-up after angioplasty, 3) residual stenosis greater than 30% immediately after angioplasty, and 4) dilation of the left anterior descending or right coronary artery. Biochemical investigations showed a greater decrease in the serum triglyceride levels in the fish oil-treated group versus the control group (p less than 0.05) but no differences between the two groups in cholesterol levels or platelet counts over the 4 month period. In conclusion, in this study, the administration of fish oil at a dose of 10 capsules/day did not reduce the incidence of early restenosis after coronary angioplasty.

Effect of inhibition of aldose reductase on glucose flux, diacylglycerol formation, protein kinase C, and phospholipase A2 activation☆

Activation of the polyol pathway under hyperglycemic conditions is proposed to contribute to the development of diabetic nephropathy. The mechanisms by which this activation may lead to functional and structural changes within the kidney are yet to be definitively established. We have examined in vitro the steps linking increased polyol pathway activity resulting from hyperglycemia to prostaglandin production. Following the demonstration of increased prostaglandin E (PGE) levels in glomeruli from diabetic rats (14.9 +/- 2.5 v 59.1 +/- 19.4 ng PGE/mg protein), a specific inhibitor of aldose reductase, HOE-843, was used in vitro to analyze the response to hyperglycemia of the steps preceding prostaglandin production. In explants of glomeruli from control animals, increasing the glucose concentration in vitro from 5.6 mmol/L to 25 mmol/L resulted in a significant increase in the flux of glucose through the pentose phosphate pathway ([PPP] 1.29 +/- 0.08 v 2.00 +/- 0.11 nmol/h), de novo diacylglycerol synthesis (2.2 +/- 0.1 v 3.1 +/- 0.2 micromol/mg protein), membrane protein kinase C (PKC) activity (18.7 +/- 0.5 v 24.3 +/- 0.75 pmol/microg protein), and in vitro phospholipase A2 (PLA2) activity (2.18 +/- 0.46 v 3.83 +/- 1.07 nmol arachidonic acid hydrolyzed/min/mg cytosolic protein). For all parameters measured, the increase resulting from the increased glucose concentration could be prevented by in vitro addition of HOE-843 for 24 hours before measurement. These findings provide evidence to suggest a mechanism linking increased polyol pathway activity and an increase in PLA2 activity to increased prostaglandin production, which is observed in diabetes of recent onset and may ultimately lead to changes associated with the development of diabetic nephropathy.

Pancreatic islets synthesize phospholipids de novo from glucose via acyl-dihydroxyacetone phosphate

There is considerable evidence that an increased turnover of phosphoinositides and phosphatidic acid accompanies stimulus-induced insulin release. As glucose metabolism via glycolysis produces precursors for phospholipid synthesis, the time course of incorporation of [U14C] labelled glucose was measured to determine the pathways of triose carbon incorporation into phospholipids in the islet. Cultured islets were stimulated with glucose 2.7 or 33 mM. The labelled phospholipids present after stimulation were acyldihydroxyacetone phosphate, lysophosphatidic acid, phosphatidic acid and phosphatidylinositol. Acyl-dihydroxyacetone phosphate rose promptly within 1 minute of raising the glucose concentration and was the primary acylated triose labelled during the first 15 minutes. It was possible to show in vitro conversion of [U14C] glucose-derived acyl-dihydroxyacetone phosphate to lysophosphatidic acid and phosphatidic acid in the presence of NADPH (100 microM), indicating the presence in the islet of acyl-dihydroxyacetone phosphate: NADP oxidoreductase and acyl CoA:1 acylglycerol-3-phosphate acyl transferase, respectively. This study suggests that de novo synthesis of phosphatidic acid provides a link between glucose metabolism and the release of insulin.

Evolution of Insulin Resistance in New Zealand Obese Mice

The etiology of non-insulin-dependent diabetes mellitus (NIDDM) is not known. Hyperglycemia is due to increased hepatic glucose production (HGP), decreased glucose uptake, and impaired insulin secretion. It is unknown if these defects are coinherited or if one precedes and causes the others. The aim of this study was to determine the earliest defects in the evolution of the syndrome in the New Zealand obese (NZO) mouse, a polygenic model of NIDDM. NZO and control NZC mice were studied at 4–5 and 20 wk of age. Glucose turnover and glucose uptake in individual tissues were measured basally and during a hyperinsulinemic clamp. First-phase insulin secretion was measured after an intravenous glucose load. HGP was higher in the NZO mice both basally and during the clamp at bothages. At 4–5 wk of age, there was evidence of insulin insensitivity in brown adipose tissue, soleus, diaphragm, red quadriceps, and red gastrocnemius but not in heart, white quadriceps, and white gastrocnemius. In 20-wk-old mice, insulin responsiveness was decreased in white and brown adipose tissue and soleus muscle but not in heart, diaphragm, red and white quadriceps, and red and white gastrocnemius. First-phase insulin secretion (percentage rise above basal) 3 min after the glucose bolus was impaired in NZO miceat both ages. We conclude that hepatic glucose overproduction, brown adipose tissue and skeletal muscle insulin resistance, and impaired first-phase insulin secretion are all early abnormalities in the NZO mouse.

Impaired Glucose Tolerance, Hyperinsulinemia, and Hypertriglyceridemia in Australian Aborigines From the Desert

A cross section of adult full-blooded Aborigines from three small isolated communities in the desert region of northwest Australia was surveyed for diabetes, impaired glucose tolerance (IGT), insulin levels, and lipoprotein lipids. Sixty-three men and 86 women from a total adult population of 330 were tested. Of the people tested, 67.6% had normal glucose tolerance, 25% had IGT, and 7.4% had diabetes. Both diabetes and IGT were strongly age related. Fasting insulin levels and insulin responses to oral glucose (elevation above basal) were elevated. Although fasting insulin rose with age, insulin response did not rise after adjustment for body mass index (BMI). Plasma triglyceride levels were high, particularly in men >35 yr old (3.13 ± 0.32 mM), but cholesterol levels were not elevated. Multiple regression analysis of fasting glucose, 2-h glucose, plasma triglyceride, fasting insulin, and insulin response for the nondiabetic subjects revealed 7) BMI was an independent risk factor for elevated 2-h glucose levels in women but not in men and was strongly related to fasting insulin concentrations in both genders; 2) fasting insulin concentration was an independent risk factor for increases in fasting glucose, insulin response, and triglyceride levels; 3) insulin response was related to the 2-h glucose level; 4) fasting and 2-h glucose levels and fasting insulin and triglyceride concentrations all rose with age in both genders, with the rate of increase generally greater in men. The most striking difference between these desert Aborigines and previously studied coastal Aborigines from the same geographical region was the significantly higher insulin response. It is possible that the more severe hyperinsulinemia in the desert Aborigines may indicate a greater degree of susceptibility to type II diabetes than in the coastal Aborigines as the duration of urbanization increases.

Activity of endogenous phospholipase C and phospholipase A2 in glucose stimulated pancreatic islets.

In cultured pancreatic islets from neonatal rats labelled with [3H] arachidonic acid, glucose stimulation prompted a fall in the labelled arachidonate concentration of phosphatidylinositol and a concomitant rise in 1,2 diacylglycerol and phosphatidic acid. The time course of glucose stimulation indicated that this early event was followed by an increased liberation of arachidonic acid and incorporation into arachidonate metabolites. Incubation of homogenates of glucose stimulated islets with both phosphatidylinositol and phosphatidylcholine specifically labelled with arachidonate in the 2-position acyl chain generated arachidonic acid. This indicated both phospholipase C with 1,2 diacylglycerol lipase and phospholipase A2 activities in the action of glucose. Calcium dependent arachidonic acid release was also seen from arachidonic acid labelled phosphatidic acid. The findings suggest multiple sources of islet arachidonic acid following glucose stimulation including phospholipase A2 hydrolysis of phosphatidic acid.

Effect of Prostaglandin E2 and Hyaluronan on Mesangial Cell Proliferation: A Potential Contribution to Glomerular Hypercellularity in Diabetes

Proliferation of mesangial cells is a feature of several forms of human and experimental glomerulopathy, including that seen in diabetes. The nonsulfated glycosaminoglycan hyaluronan participates in the regulation of pericellular matrix assembly and is a mitogen in some cell types. We have shown previously that hyaluronan production is increased in the glomerulus in a glucose- and prostaglandin-dependent manner. We have investigated the effect of diabetes and of addition of hyaluronan and prostaglandin E2 (PGE2) on the uptake of [3H]thymidine by glomerular core preparations enriched in mesangial cells. When compared with nondiabetic controls, it was shown that [3H]thymidine uptake was significantly increased in glomerular core preparations from streptozotocin-induced diabetic rats (to 169 ± 5%, P < 0.001). In glomerular cores from both experimental groups, hyaluronan (50–250 ng/ml) or PGE2 (10−12 to 10−8 mol/l) increased the uptake of [3H]thymidine. Further, mesangial cells from nondiabetic control glomerular cores, when maintained in culture in early passage, responded with increased [3H]thymidine uptake to raised glucose (5.6–25 mmol/l) and to added hyaluronan and PGE2. We propose that prostaglandin and hyaluronan production in response to a raised glucose environment in diabetes can contribute to mesangial hypercellularity.

Production of 6-Oxo-Prostaglandin F1α by Rat Aorta: Influence of Diabetes, Insulin Treatment, and Caloric Deprivation

The production of 6-oxo-prostaglandin F1α (6-oxo-PGF1α), a stable metabolite of prostacyclin (PGI2), was examined in three separate series of experiments in aortae from rats with diabetes induced by streptozotocin (SZ) and aortae from nondiabetic control rats. In the first series, it was shown that production of 6-oxo-PGF1α by aortae from rats with diabetes of 4–6-wk duration was significantly decreased compared with nondiabetic rats (0.77 ± 0.07 ng/mg wet weight aorta vs. 1.23 ± 0.11 ng/mg wet weight aorta, P < 0.001). Diabetic rats were treated with two doses of insulin, a lower dose of 8 U/kg/day that only partially corrected their plasma glucose and body weight compared with nondiabetic rats, and a higher dose of 25 U/kg/day that completely corrected their plasma glucose and body weight. Both insulin regimens completely reversed the defect in 6-oxo-PGF1α production. In the second series, the time course of development of the defect in 6-oxo-PGF1α production was examined. It was found that two weeks after induction of diabetes, the aortic production of 6-oxo-PGF1α did not differ between control and diabetic rats. However, 4 and 7 wk after induction of diabetes, 6-oxo-PGF1α production was significantly reduced, even though the degree of hyperglycemia and hypoinsulinemia was the same as at 2 wk. In the third series, the effect on aortic 6-oxo-PGF1α production of weight loss caused by restricted dietary intake was compared with the effect of a comparable weight loss caused by experimental diabetes. Again, the diabetic rats had significantly reduced 6-oxo-PGF1α production compared with control rats, but 6-oxo-PGF1α production in nondiabetic rats which had lost weight as a result of limited feeding was the same as nondiabetic control rats fed ad libitum. It is concluded (1) that insulin treatment, even if insufficient to achieve perfect control of plasma glucose, can restore 6-oxo-PGF1α production to normal, (2) diabetes of greater than 2-wk duration is required to produce the abnormality, and (3) the reduced 6-oxo-PGF1α production is not a nonspecific consequence of weight loss.

Nandrolone decanoate and intranasal calcitonin as therapy in established osteoporosis

This study used a randomized, 2 × 2 factorial design to evaluate over 2 years the effect of intranasal salmon calcitonin and intramuscular nandrolone decanoate on bone mass in elderly women with established osteoporosis. The study was double masked in relation to calcitonin and open in relation to nandrolone decanoate. One hundred and twenty-three women aged 60–88 years who had sustained a previous osteoporotic fracture, or had osteopenia, were recruited through an outpatient clinic. Women were assigned to one of four groups: (1) daily placebo nasal spray, (2) 400 IU intranasal calcitonin daily, (3) 20 intramuscular injections of 50 mg nandrolone decanoate (given as two courses of 10 injections) plus placebo nasal spray, or (4) 20 injections of 50 mg nandrolone decanoate plus 400 IU intranasal calcitonin daily. All subjects received 1000 mg calcium supplementation daily. Outcomes measured included changes in bone mineral density (BMD) at the lumbar spine, as measured by dual-energy quantitative computed tomography (DEQCT), in BMD of the proximal femur, and BMD and bone mineral content (BMC) of the lumbar spine and forearm, as measured by dual-energy X-ray absorptiometry (DXA). Significant positive changes from baseline in DXA BMC at the lumbar spine were observed over 2 years in the calcitonin group (5.0±1.9%, mean ± SE) and in the nandrolone deconate group (4.7±1.9%) but not in the placebo group (1.1±2.2%) or the combined therapy group (0.7±1.8%). Modelling based on the 2×2 factorial design revealed that nandrolone decanoate was associated with a 3.8±1.8% (p<0.05) gain in DXA BMD at the proximal femur. Modelling also revealed that calcitonin treatment was associated with a loss of 11.5±4.7% in DEQCT BMD at the lumbar spine and a loss of 3.7±1.8% in DXA BMD at the proximal femur (p<0.05). There was in vivo antagonism between the two medications of 7.9±3.9% for DXA BMC at the lumbar spine. Both agents caused positive changes from baseline in lumbar spine BMC. Nandrolone decanoate had beneficial effects on BMD at the proximal femur. This dose of intranasal calcitonin was associated with deleterious effects on trabecular BMD at the lumbar spine and total BMD at the proximal femur. There may be significant clinical antagonism between these two medications.

Thyroid hormone levels and protein binding in patients on long-term diphenylhydantoin treatment.

The effect of long‐term diphenylhydantoin (DPH) treatment on thyroid hormone concentrations and protein binding was determined in a randomized controlled trial. As has been demonstrated previously, total thyroxine (T4) concentrations were significanly depressed in patients on DPH. There was no significant effect on indirect indices of protein binding of thyroid hormones, and the free thyroxine index (FTI) was also significantly depressed. Triiodothyronine (T3) and thyrotrophin (TSH) concentrations were either unaffected, or only very slightly affected by DPH. Significant effects on the FTI were still apparent 4 weeks after discontinuing treatment. It is concluded that the depression of total T4 levels observed in vivo is not due solely to diminished protein binding, but may instead be largely explained by reports suggesting enhanced degradation of T4 following DPH therapy.