The purpose of the present study was to clarify the pathogenetic mechanism of ethanol-induced fatty liver in experimental diabetic rats. Male Wistar strain rats, weighing 140-150 g, were intravenously administered with a single dose of streptozotocin (60 mg/kg) with citrate buffer. They were given an Oriental MF powder diet containing 10% lard and tap water or 20% ethanol solution ad libitum for 5 weeks. All experiments were performed in the morning (i. e., 18 hr after the last meal). The results obtained may be summarized as follows.
1) In normal and diabetic rats given ethanol solution (abbreviated as N-FE, and DM-FE, respectively), the serum FFA concentrations and liver TG contents were remarkably increased compared to those in the normal (N) and diabetic (DM) rats, although the serum TG concentration did not differ among all groups.
2) The lipid-peroxide contents in both the liver mitchondrial and microsomal fractions in the N+E rats were significantly increased compared to those in the N rats. In the DM-1-E rats, the lipid-peroxide content was increased in the microsomal fraction.
3) In a kinetic study of FFA and triglyceride using ¹⁴C-l-palmitate, it was found that the disappearance curve of palmitate in the N rats was similar to that in the N-I-E rats, but the release of labeled VLDL-TG was smaller in the N-I-E rats than that in the N rats. However, these patterns were not different between the DM and DM-FE rats.
4) The release of VLDL-TG induced by Triton WR-1339 was diminished in both the N-I-E and DM-I-E rats as compared to their respective controls. In the DM rats, the post-heparin lipolytic activity (PHLA) was less than that in the N rats. In both groups, however, ethanol had no effect on the PHLA.
These results suggest that ethanol-induced fatty liver in diabetic rats may result in an overproduction of TG in the liver and a relative decrease of VLDL-TG release from the liver.

View full abstract

The vitamin E (Vit E) levels of plasma lipoprotein fractions (VLDL-Vit E, LDL-Vit E and HDL-Vit E) and lipid peroxide (LPO) levels of plasma, aorta, liver and pancreas, were determined in 8 streptozotocin (50 mg/kg body weight)-induced diabetic rats (STZ rats) to elucidate the role of Vit E, especially HDL-Vit E, in the lipid peroxidation in plasma and several tissues. Vit E and LPO were determined by the methods of Abe and Yagi, respectively. STZ rats were given water and normal diet ad libitum for 3 months. Each value for the STZ rats was statistically compared with the data of 7 control rats kept under similar conditions.
In the STZ rats, LDL-Vit E (1.41±0.13 ng/ml, mean±SE), HDL-Vit E (3.58±0.20) and Total-Vit E (Vit E of VLDL, LDL and HDL; 6.17±0.45) were significantly higher than those of control rats, LDL-Vit E (1.96±0.21, p<0.05), HDL-Vit E (4.59±0.13, p<0.001) and Total-Vit E (7.30±0.25, p<0.05), respectively. On the other hand, VLDL-Vit E was similar in both groups.
All LPO levels in the plasma, aorta and pancreas were high but that of the liver was low, as compared to control rats.
The above results indicate that HDL-Vit E does not have a close relationship to the lipid peroxidation in several tissues. The reasons for this are not clear. One possible factor could be a decrease of some unsaturated fatty acid to be peroxidized. Another explanation may involve an increase of other antioxidants without Vit E. Further studies on these points are required.

View full abstract

The effect of small-dose intravenous insulin boluses was compared with that of conventional large-dose insulin therapy by the intravenous and subcutaneous routes in 9 patients with severe diabetic ketoacidosis.
In the treatment with small-dose intravenous insulin boluses, 4 patients presenting semicoma received hourly intravenous injections of 8 units of regular insulin (RI) and one patient presenting coma received an initial intravenous injection of 20 units of RI with subsequent hourly injections of 10 units of RI.
The pretreatment blood glucose level was 892±131 mg/dl (M±SE) in the small-dose group, and 692±121 mg/dl in the large-dose group. The blood glucose dropped to levels of less than 250 mg/dl in 8±2 hr and 6±2 hr, respectively, and the amount of insulin necessary to lower blood glucose to these levels was 72±21 u and 97±19 u, respectively. The rate of fall in blood glucose per hour until then was 91±12 mg/dl in the former and 98±27 mg/dl in the latter. The percent decrease from the initial blood glucose level was 59±8 % and 61±8 % in 6 hr, respectively. These data were not significantly different between the two groups. The IRI concentrations measured at just before the insulin injections during the treatment with small-dose intravenous insulin boluses (n = 3) ranged from 18 to 43 μu/ml. No hypoglycemia or hypokalemia was observed in either of the groups.
These results suggest that small-dose intravenous insulin boluses are simple and as effective as conventional large-dose insulin therapy in the treatment of diabetic coma. Furthermore, it has been confirmed that the amelioration of hyperglycemia seen in diabetic coma can be attained by the same amount of insulin in both groups.

View full abstract

To establish a mechanism for the effect of diet therapy on glucose tolerance in maturity-onset diabetes, insulin binding to peripheral mononuclear cells was compared with the results of 100 g-O-GTT and insulin sensitivity tests. A half ng per ml of ¹²⁵I-insulin specific binding to 10⁷ cells was tested in 16 normal subjects, and 18 with maturity-onset diatetes. In the latter, the tests were repeated after a month of diet theray.
In the 18 with untreated diabetes, the insulin sensitivity index (ISI), insulin binding, and its binding sites were decreased significantly compared to those of the normals (p<0.05). Successful diet therapy as proven by body weight loss and by reduced ΣBS, was accompanied by increased ISI, insulin binding, and its binding sites. A significant correlation was observed between ISI and insulin binding after the therapy. However, no statistical correlation was found between insulin binding and the level of fasting serum insulin or ΣIRI during O-GTT.
In conclusion, it can be said that:
1) Decrease of insulin receptor may play a significant role in the progression of glucose intolerance in maturity-onset diabetes.
2) Increases in cellular insulin binding and in insulin sensitivity are important in the improvement of glucose intolerance seen after diet therapy.

View full abstract

A radioimmunoassay method for glibenclamide was developed and evaluated for clinical application. A derivative of glibenclamide was conjugated with bovine serum albumin by diazocoupling via the most distant portion of the molecule from the cyclohexyl ring to be metabolized. Antiserum was raised in rabbits by repeated injections of this material together with Freund's completeadjuvant. ³H-Glibenclamide was used as a tracer. The antiserum bound 35% of 160 pg ³H-glibenclamide at a dilution of 1: 8000 after incubation for 24 hours at 4°C. The binding of ³H-glibenclamide to the antiserum was significantly decreased by the addition of 50 pg unlabeled glibenclamide per tube. The dextran-coated charcoal method was employed to separate free and bound 3H-glibenclamide. Using 10μl plasma, plasma glidenclamide levels of 5-500 ng/ml could be assayed by this radioimmunoassay technique. Since non-specific binding of glibenclamide by plasma albumin occurs, normal human plasma should be included in the serial dilutions of the standard glibenclamide solution when the drug level in human plasma is to be assayed. With the exception of glipizide, most sulfonylurea drugs cross-reacted very little with this antiserum. The two metabolites of glibenclamide (M₁ and M₂) did not cross-react with the radioimmunoassay system to any significant extent. Recovery tests and dilution tests of the radioimmunoassay mostly gave expected values. Plasma glibenclamide values by this method were highly correlated with those by gas chromatography and liquid chromatography. The plasma glibenclamide levels of normal subjects who took 2.5 mg of this drug orally, increased to 72±32 ng/ml after 1.5 hours by this assay method. The presentmethod appears to represent a new tool for studying the pharmacokinetics of glibenclamide in clinical practice.

View full abstract

Isolated hepatocytes were prepared from meal-fed female Wistar rats to determine the effects of various substrates on ketogenesis, fatty acid synthesis and glycolysis.
Inhibition of fatty acid synthesis with dibutyryl cyclic AMP or oleate resulted in increased ketogenesis. On the other hand, octanoate, which is transported by a carnitine independent mechanism into the mitochondrion, showed marked ketogenesis without any effect on fatty acid synthesis. Acetate, which is converted to acetyl-CoA either in cytosol or in mitosol, increased both ketogenesis and fatty acid synthesis. These data indicated that there is not always a reciprocal relationship between ketogenesis and fatty acid synthesis.
Exogenous lactate and pyruvate relieved the enhanced ketogenesis with octanoate in association with increase of fatty acid synthesis and CO₂ formation from octanoate. It was concluded that the flow of acetyl groups in fatty acid synthesis contributed more than CO2 formation in the reduction of ketogenesis by exogenous lactate and pyruvate.
The present data suggest that there is a regulatory link between fatty acid synthesis and ketogenesis. Generally, insufficient activity of fatty acid synthesis permits a large increase in ketogenesis, partly because entry of long-chain fatty acyl-CoA into the mitochondrion is facilitated and partly because utilization of acetyl groups is decreased.
Chylomicrons and their remnants prepared in vivo showed a specific inhibitory effect on fatty acid synthesis without any modulation of ketogenesis. This metabolic effect is very difficult to explain and the mechanism is now under investigation in our laboratory. Chylomicron remnants prepared in vitro were found to release free fatty acids into the medium, probably because of bound lipoprotein lipase activity. Metabolic effects of in vitro remnants can evidently be attributed to released free fatty acids.

View full abstract

Insulin resistance and generalized insulin allergy seldom occur in the same individual during insulin therapy. We report here the occurrence of diabetic coma due to insulin resistance following generalized insulin allergy. The patient was a 57-yr-old woman who was started on NPH insulin therapy for a second time. On the 9th day of insulin therapy, she developed generalized urticaria which lasted for 2 days during which time insulin therapy was continued. On day 13, she developed ketoacidosis. She received 450 units of Regular insulin and a fluid and electrolyte replacement for the first 10 hours, but the arterial pH and base excess did not change and she became comatose. Subsequently, she was begun on an increased dose of Actrapid insulin because of suspected immunologic resistance to Regular insulin. This treatment was followed by a prompt normalization of blood sugar and ketoacidosis, and the patient recovered completely. The intradermal direct skin test, Prausnitz-Küstner test and radioallergosorbent test were all positive against various insulins. On the other hand, the ¹²⁵I-insulin binding rate, total and free IRI levels were 67.1%, over 900 mcU/ml and 32 mcU/ml, respectively, just before Regular insulin was switched to Actrapid insulin but thereafter, the free IRI levels increased to over 320 mcU/ml consistent with rapid normalization of the clinical data. These results indicate that anti-insulin IgE antibody and insulin binding antibody were respectively responsible for the occurrence of insulin allergy and insulin resistance in this patient.

View full abstract

The purpose of this communication is to outline the results of light microscopic and immunohistological studies of the islets of male CHBB rats actively immunized repeatedly at 2-week intervals for a total of 12 times with homologous islet tissue.
Of 8 animals in the group sensitized with islet antigen and complete Freund's adjuvant, 4 gave a diabetic K-value. In the group sensitized with islet antigen alone, one of 7 animals had a diabetic K-value.
The pancreas of rats actively immunized with homologous islets yielded a wide variety of morphological findings in the islets and islet cells: insulitis, disorderly arrangement and pyknosis of islet cells, total replacement of some islets by a granulomatous lesion composed of infiltrating histiocytic cells, phagocytosis of particles of the nuclei of islet cells, a decrease in B cells (A/B shift) and fibrosis of the islets. On the other hand there was also proliferation and hypertrophy of islets with enlargement of the nuclei and nucleoli of islet cells, and an increase in chromatin and nuclear fission.

View full abstract

To evaluate the volume of the pancreas in juvenile onset diabetes (JOD), T values were measured by ultrasonography. The T value was obtained by measurement of the ventro-dorsal diameter at the body portion of the pancreas, and a good relationship was noted between the T value and volume of the pancreas in autopsy cases.
Following this fundamental study, T values were measured in 52 JOD patients and 101 healthy controls. A significant correlation (p<0.01) was observed between the T value and the height in the control group. Although a good correlation was also obtained between the T value and the height in JOD, the coefficient for JOD was clearly less than that in the controls.
Furthermore, a significant relationship existed between the duration of diabetes, daily injected dose of insulin and the T deviation value (= average T value of the control in the same height group-T value in the individual case of JOD).
These results suggest that the growth of the pancreas in JOD did not run parallel with increase of height as compared to the control group, and the T value measured by ultrasonography appeared to represent a useful index for estimating the degree of damage of the pancreas.

View full abstract