Prolonged inhibition of glycogen phosphorylase in livers of Zucker Diabetic Fatty rats models human glycogen storage diseases.

The preclinical efficacy and safety of GPi921, a glycogen phosphorylase inhibitor, was assessed following twenty-eight days of administration to Zucker Diabetic Fatty (ZDF) rats. The ZDF rat is an animal model of type 2 diabetes mellitus (TTDM) which develops severe hyperglycemia. Inhibition of glycogen phosphorylase throughout the duration of the study was demonstrated by reductions in twenty-four-hour glucose profiles and glycated hemoglobin levels. In addition, progression towards hyperglycemia was halted in treated but not control animals, which developed hyperglycemia over the twenty-eight days of the study. Biochemical and histopathological analysis revealed large increases in hepatic glycogen, which closely paralleled the development of hepatomegaly and ultimately resulted in increases in hepatic lipids. Furthermore, prolonged glycogen phosphorylase inhibition resulted in an increased incidence and severity of other adverse pathological findings in the liver, such as inflammation, fibrosis, hemorrhage, and necrosis. The observed biochemical and histopathological phenotype of the liver closely resembled that seen in severe cases of human glycogen storage diseases (GSD) and hepatic glycogenosis in poorly controlled diabetes mellitus. These findings revealed that although glycogen phosphorylase inhibitors are efficacious agents for the control of hyperglycemia, prolonged treatment might have the potential to cause significant clinical hepatic complications that resemble those seen in GSD and hepatic glycogenosis.

In vivo evidence for alcohol-induced neurochemical changes in rat brain without protracted withdrawal, pronounced thiamine deficiency, or severe liver damage.

Magnetic resonance spectroscopy (MRS) studies in human alcoholics report decreases in N-acetylaspartate (NAA) and choline-containing (Cho) compounds. Whether alterations in brain metabolite levels are attributable to alcohol per se or to physiological effects of protracted withdrawal or impaired nutritional or liver status remains unclear. Longitudinal effects of alcohol on brain metabolites measured in basal ganglia with single-voxel MRS were investigated in sibling pairs of wild-type Wistar rats, with one rat per pair exposed to escalating doses of vaporized alcohol, the other to vapor chamber air. MRS was conducted before alcohol exposure and twice during exposure. After 16 weeks of alcohol exposure, rats achieved average blood alcohol levels (BALs) of approximately 293 mg per 100 ml and had higher Cho and a trend for higher glutamine+glutamate (Glx) than controls. After 24 weeks of alcohol exposure, BALs rose to approximately 445 mg per 100 ml, and alcohol-exposed rats had higher Cho, Glx, and glutamate than controls. Thiamine and thiamine monophosphate levels were significantly lower in the alcohol than the control group but did not reach levels low enough to be considered clinically relevant. Histologically, livers of alcohol-exposed rats exhibited greater steatosis and lower glycogenosis than controls, but were not cirrhotic. This study demonstrates a specific pattern of neurobiochemical changes suggesting excessive membrane turnover or inflammation, indicated by high Cho, and alterations to glutamate homeostasis in the rat brain in response to extended vaporized alcohol exposure. Thus, we provide novel in vivo evidence for alcohol exposure as causing changes in brain chemistry in the absence of protracted withdrawal, pronounced thiamine deficiency, or severe liver damage.

Myocardial glycogen storage disease in farmed rainbow trout, Oncorhynchus mykiss (Walbaum).

This paper is the first description of a spontaneous glycogen-storage disease in a lower vertebrate, as previous descriptions deal with humans and other mammals, or fish where the condition has been experimentally induced. Affected farmed rainbow trout experienced increased mortality from 60 days post-startfeeding and displayed clinical signs of heart failure with abnormal behaviour, exophthalmia, distended abdomen and ventral skin petechiation. Necropsy revealed alterations in cardiac shape with distended atria and rounded ventricles. Microscopically, the compact wall of the ventricle was absent, uneven or thinner than normal. The cardiac myocytes contained extensive amounts of glycogen in cytoplasmic vacuoles as demonstrated by periodic acid-Schiff staining that was abolished by saliva-diastase pretreatment on serial sections. Associated lesions included conspicuous subepicardial and myocardial vascularization, epicardial thickening and necrosis of the ventricular compactum/spongiosum interphase. The lesions in cardiac myocytes had a striking resemblance to glycogenosis type II (Pompe disease), a rare autosomal recessive lysosomal storage disease in humans. This condition was more severe and mortality was higher in a replicate/parallel fish group treated perorally with 17alpha-methyltestosterone to produce all-female progeny, indicating that the hormone treatment aggravated the condition resulting in earlier and more severe manifestation of the disease in this group.

Hepatocellular carcinoma occurring in a patient with Crohn's disease treated with both azathioprine and infliximab.

Hepatocellular carcinoma is known to be associated with underlying liver diseases, such as cirrhosis, hemochromatosis, and chronic viral hepatitis. All reported cases of hepatocellular carcinoma in association with Crohn's disease involve patients treated previously with azathioprine or both azathioprine and steroids. However, hepatocellular carcinoma associated with the use of azathioprine and infliximab has not been reported. In this report, we describe an unusual case of hepatocellular carcinoma and focal hepatic glycogenosis (FHG) occurring in a non-cirrhotic Crohn's disease patient who has been treated with both azathioprine and infliximab.

[A biochemical study of an animal model with defective muscle glycolysis, induced by iodoacetate administration].

A rat model produced by intraaortic injection of sodium iodoacetate was analyzed biochemically. Hind leg muscles obtained from iodoacetate-injected site showed a selective inhibition of the glyceraldehyde-3-phosphate dehydrogenase (G3PD) activity less than 10% of control mean. No loss of mitochondrial enzyme activity was observed. Electrical stimulation at the sciatic nerve revealed muscle contractures and this muscle showed a mild degeneration in histochemically. This study indicates that this model will be useful to study the pathophysiology of human muscle glycogen storage diseases.

Fusion of storage lysosomes in experimental lipidosis and glycogenosis.

This ultrastructural investigation on renal collecting duct cells and hepatocytes of rats deals with the question of whether or not lipid-storage lysosomes as induced by cationic amphiphilic compounds retain their ability to fuse with autophagosomes/autolysosomes. These were recognized by their glycogen content which was made to persist by means of acarbose, an inhibitor of lysosomal alpha-glucosidase. To induce lipidosis, rats were pretreated for several weeks with chloroquine or chlorphentermine; they then received combined treatment with the lipidosis-inducing drug plus acarbose. In renal collecting duct cells, mixed storage lysosomes displaying the features of both lipidosis and glycogenosis were found to predominate, indicating that fusion between lipid-laden lysosomes and glycogen-containing autophagosomes/autolysosomes was efficient. Hepatocytes also displayed some mixed storage lysosomes; these were, however, regularly accompanied, within a given hepatocyte, by greater numbers of pure lipidosis-related inclusions and pure glycogen vacuoles. This observation indicates that in hepatocytes lipid-storage lysosomes were rather reluctant to fuse, thus displaying a feature of telolysosomes which are no longer capable of participating in cellular digestion.

Glycogen storage disease type X caused by ochratoxin A in broiler chickens.

Day-old broiler chickens fed graded concentrations of ochratoxin A (0, .5, 1.0, 2.0, 4.0, and 8.0 micrograms/g of diet) for 3 weeks were examined histopathologically for glycogen accumulation in muscle tissue using both formalin and ethanol fixatives with periodic acid-Schiff stain in a duplicate diastatic slide technique. However, glycogen could not be visualized in chicken muscle by these techniques which work with human muscle. Quantitative analysis for the glycogen itself permitted demonstration of a significant (P less than .05) fourfold increase in birds fed 8 micrograms/g. Using startled birds demonstrated that this accumulated glycogen was about 99% physiologically available. These findings coupled with prior reports permit the assignment on morphologic criteria of the hyperglycogenation of ochratoxicosis as a type X glycogen storage disease.

Lysosomal glycogen storage mimicking the cytological picture of Pompe's disease as induced in rats by injection of an alpha-glucosidase inhibitor. I. Alterations in liver.

The present paper describes an animal model of lysosomal glycogenosis as induced by a competitive inhibitor of alpha-glucosidase. Rats received intraperitoneal injections of the inhibitor, a pseudotetrasaccharide (Acarbose, Bay g 5421); liver tissue was examined by light and electron microscopy. Substrate-histochemical and enzyme-cytochemical methods were used to demonstrate intralysosomal glycogen storage within hepatocytes and Kupffer cells. The cytological picture closely resembled that occurring in glycogenosis type II (Pompe's disease) of humans. After cessation of drug treatment, the glycogen storage was slowly reversible. The present results point to the physiological role of the lysosomal apparatus for intracellular glycogen turnover. On the cellular level, this experimentally induced glycogenosis may be useful as a model of Pompe's disease.

Activity of alpha-1, 4-glucosidase in furazolidone-induced glycogenosis.

Furazolidone (FZ) at 700 and 800 p.p.m. was added to feed mixtures fed turkey poults two and three weeks posthatching, respectively, to induce acute experimental cardiomyopathy. Poults in the control pen received the same ration but without FZ. From EKG data obtained at 2, 4, and 5 weeks of age, control unaffected and experimental affected poults were selected for sacrifice. Poults were sacrificed by cervical dislocation and appropriate samples of hepatic tissue were removed for assays of activity of alpha-1, 4-glucosidase. Results indicate that enzyme activity in affected FZ-treated poults is similar to that in unaffected control poults. Lack of significant differences in activity of this lysosomal enzyme suggests that FZ-induced glycogenosis may be related to the adult form of idiopathic generalized glucogenosis, the etiology of which remains unidentified.

[The fructose induces "glycogenosis". III. Histochemical and morphometrical studies of glycogen metabolism in mouse liver after fructose overload (author's transl)]. / Die Fruktose-induzierte "Glykogenose". III. Histochemische und morphometrische Untersuchungen des Glykogenstoffwechsels in der Mäuseleber nach Fruktosebelastung.

INTRODUCTION: After feeding fructose for 7 days rat liver cells show an accumulation of glycogen, a high activity of glucose-6-phosphatase combined with a SER- and RER-reduction. This result was reviewed by mouse liver cells using histochemical and morphometrical methods. MATERIAL AND METHODS: 60% fructose in drinking water was given mice as only nutritional source. Controls had free access to Altromin-R-standard diet and drinking water. Glycogen and glycogen metabolizing enzymes are demonstrated in the course of an 1-14 days fructose diet. After a 7 days diet liver tissue was analysed morphometrically. RESULTS AND DISCUSSION: Feeding of fructose leads to a high glycogen content, combined with a high activity of glycogen-phosphorylase and glucose-6-phosphatase in the liver parenchyma of mouse. Glycogen-synthetase activity falls to a low level. The SER and RER and the peroxisomes are reduced. The single volume of the hepatic nucleus is decreased and the hepatocellular chondrioma is transformed in a smaller number of larger mitochondria. Compared with the rate the analysed organelles and enzymes of mouse liver show only slight quantitative differences. The increase of glucose-6-phosphatase and simultaneous reduction of endoplasmic reticulum-membranes is illustrated by the dynamic structure of endoplasmic reticulum-membranes, which adapt to metabolic changes. The variable turnover of different parts of endoplasmic reticulum-membranes seems to be very important.

[The fructose induced "glycogenosis". II. Histochemical studies of glycogen metabolism in rat liver after fructose overload and similar diets (author's transl)]. / Die Fruktose-induzierte "Glykogenose". II. Histochemische Untersuchungen zum Glykogenstoffwechsel der Rattenleber nach Fruktosebelastung und bei vergleichbaren Diäten

INTRODUCTION: Feeding of fructose for 7 days has been morphometrically shown to induce a SER-reduction and an accumulation of glycogen in rat liver cells. This hypothetical model "glycogenosis" is investigated with histochemical methods. MATERIAL AND METHODS: Rats are given a solution of 60% fructose in water as only nutritional source. Controls are given a solution of 60% glucose in water, an isocaloric Altromin-R-standard diet and an Altromin-R-standard diet ad libitum. Reversion of fructose induced metabolic changes is investigated by a 7 days fructose diet followed by an 1-4 days Altromin-R-standard diet ad libitum. Glycogen and glycogen metabolizing enzymes are demonstrated after a 7 days diet and in the course of an 1-7 days fructose diet. RESULTS AND DISCUSSION: Feeding of fructose leads to a high glycogen content, combined with a high activity of glycogen-phosphorylase and glucose-6-phosphatase in the liver parenchyma. Glycogen-synthetase activity increases during the first 4 days and then it drops to a low level. A pathological alteration of liver cell metabolism seems to be improbable, for all fructose induced changes are reversibel after 2 days of Altromin-R-standard diet. Glucose-6-phosphatase, as a marker-enzyme of the smooth endoplasmatic reticulum, is discussed to become activated by disruption of SER membranes due to fructose.

[The fructose induced "glycogenosis". I. Ultrastructural and morphometric analysis of rat hepatocytes 7 days after fructose overload (author's transl)]. / Die Fruktose-induzierte "Glykogenose". I. Ultrastrukturelle und morphometrische Untersuchungen an der Rattenleberzelle nach 7tägiger Fruktosebelastung

INTRODUCTION: Infusion of fructose has been shown to stimulate the SER and to reduce the RER in rat liver cells. After feeding fructose of 7 days a glycogenosis of unknown pathogenesis occurs, which was analysed morphometrically. MATERIAL AND METHODS: 60% fructose in water, was given as drinking water to animals deprived of other food. Controls had free access to Altromin-R-standard diet and drinking water. Liver tissue was analysed morphometrically according to the methods described by Weibel. RESULTS AND DISCUSSION: Rat hepatocytes accumulate a material with histochemical properties of glycogen. The enlargement of hepatic nuclei is probably due to a pathological edema. The SER is decreased suggesting a drop of glycogen catabolizing enzymes. The drastic reduction of the RER and the non-membrane bound ribosomes are signs of inpeeded protein synthesis. According to this, the peroxisomes, which arise from the RER, are decreased in volume and number. The hepatocellular chondrioma is transformed morphometrically in a smaller number of larger, pleomorphic and cup-shaped mitochondria. The ATP level drops while, on the other hand, the cristeal membranes increase. This might be caused by a negative feed back mechanism. The hepatocellular cytoarchitecture described is similar to the one found in glycogenosis type I and in the cerebrohepato-renal syndrom.