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P-100 Cerebral glucose metabolism in patients with subclinical hepatic encephalopathy using positron emission tomography
Posters/International Hepatology Communications 3 Suppl. (1995) $37-S169
P-97
Laminin with type IV c o l l a g e n enhances malignant phenotype of h e p a t o m a cells Takuji TORIMURA, Takato UENO, Motoaki KIN, S a d a t a k a INUZUKA, Michio SATA, Kyuichi TANIKAWA 1), H i r o h i s a YANO, Masamichi KOJIRO 2) 2nd dept. of medicine 1) a n d 1st dept of pathology 2) , Kurume University School o f Medicine
Background~Aims : Laminin
(La) and type IV c o l l a g e n (Ty IV) e n h a n c e cell adhesion, migration and proliferation. Ty IV and La are present along t h e sinusoids in hepatocellular carcinoma. These e x t r a c e l l u l a r matrices may promote the m a l i g n a n t phenotype of hepatoma cells. We investigated t h e influences of La and Ty IV on cell migration, a d h e s i o n and p r o l i f e r a t i o n of hepatoma cells. Subjects a n d Methods : Hepatoma cell lines (KYN-I,2,3) were used i n this study. Immunoelectron microscopy was p e r f o r m e d to investigate the production o f L a a n d Ty IV by h e p a t o m a ceils. The influence of La and Ty IV on the m i g r a t i o n , adhesion and proliferation of hepatoma cells was also evaluated. Results : La and Ty IV were produced b y hepatoma cells. The haptotactic migration, adhesion a n d thymidine incorporation of h e p a t o m a cells w e r e stimulated by Ty IV or La. Furthermore, a combination of La and Ty IV enhanced these biological activities of hepatoma ceils much more than Ty IV a l o n e . Conclusions: Our results suggest that the presence of La with Ty IV produced by hepatoma cells enhances t h e malignant phenotype o f h e p a t o m a cells.
P-99
THE MECHANISM OF A M M O N I A - I N D U C E D
INCREASE IN CHLORIDE ION C O N C E N T R A T I O N IN R A T CULTURED H I P P O C A M P A L NEURONS. T.Irie 1)2), M.Hara 2), T.Yasukura 2), K. Inoue 1), C. Inagaki 2). i) Third Department of Internal Medicine and 2)Department of Pharmacology, Kansai Medical University, Osaka 570, Japan. Hepatic encephalopathy is a neuro-psychological syndrome caused by severe liver dysfunction. Hyperammonemia is one of the important pathogenic factors of this syndrome. Previously we reported that chronically applied ammonia induces an increase in intracellular chloride concentration ([Cl]i) in cultured hippocampal neurons, and the increase in [Cl']~ is diminished by an inhibitor of carbonic anhydrase, acetazolamide. In this study, we further examined the mechanisms of ammonia-induced increase in [Cl']i. Hippocampal neuronal cells from 17-day-old rat fetus were cultured and exposed to NH4CI for 1-3 days with or without test reagents. The [Cl]i was fluorometrically estimated using a C1--sensitive fluorescent dye. The exposure to 2mM NH4C1 for 1 day significantly increased [C[]i (13.7-+1.8mM, n=9) as compared to that without exposure (8.1 -+0.5mM, n=19) and that with additional 2raM of NaC1 (7.5_+1.3 mM, n=7) or KC[ (6.3_+1.3mM, n=7). The NH4Cl-induced increase in [Cl]i was abolished by SITS and DIDS (0.1mM each), inhibitors of C l /HCO 3" exchanger, but not by bumetanide (50 # M), an inhibitor of Na+/K÷/2C1 - cotransporter. Actinomycin D (1 # M), a mRNA synthesis inhibitor, and cycloheximide (0.5 # g/ml), a protein synthesis inhibitor, also inhibited the ammonia-induced increase in [Cl']i. These findings suggest that chronic exposure of hippoeampal neurons to ammonia increases [Cl']i mainly through the facilitated exchange of C l and HCO3, in a de novo protein synthesis-dependent mannor.
$61
P-98 LATE ONSET ORNITHINE TRANSCARBAMYLASE DEFICNECY IN MALES M. Yoshino, 1 A. NishiyorL 1 H. Kate, 1 R. Kumashiro, 2and K. Tanikawa.2 1 Department of Pediatrics and Child Health, and 2 Second Department of Medicine, Kurume University School of Medicine, Kurume, Japan Male patients with ornithine transcarbamylase (OTC) deficiency usually develop a hyperammonemic crisis in the neonatal period or in early infancy. In contrast to these classical patients, here we report the occurence of males with the disease who developed a hyperammonemic crisis in adolescence or in late adulthood. The molecular basis for lhe lateness of the onset of this disease in these patients was investigated. Of 5 cases, cases 1 through 4 each developed afirst episode between 16 and 58 years of age, had an OTC activity in the postmortem liver tissue that ranged from 2-9% of control mean, and each had decreased levels of the OTC protein. Analysis of the OTC gene disclosed an R40H mutation in cases 1, 3, and 4, and in agrandson (case 5) of amale sibling of case 1, and a Y55D mutation in case 2. The activity of the mutant OTCs expressed in cosf cells was 20% for the R4OH mulalion, and 30% for the Y55D mutation of the wild type OTC. When the mutant enzyme was treated by freezing and thawing, the R40H OTC decreased to 10% of the mean of the wild type, while the Y55D OTC remained essentially at the 30% level. These results suggested that these mutant enzymes were operating in vivo at a level consistent with low normal urea production, but this level declined due to their structural instability, after an environmental load was applied.
Dlflfl
x-~t~v CEREBRAL GLUCOSE METABOLISM IN PATIENTS WITH SUBCLINICAL HEPATIC ENCEPHALOPATHY USING POSITRON EMISSION TOMOGRAPHY A.Kato, Y.Kaneta, H.Obara, R.Endo, M.Iwai, O.Moriai, l.Nakadate, T.Yoshida, K.Suzuki, and S.Sato First Dep. of Int. Med., Iwate Med. Univ., Morioka, Japan Although it is assessed that abnormalities of glucose metabolism occur substantially throughout the brain in patients with subclinical hepatic encephalopathy (SHE), liule attention has been paid to these abnormalities in situ. by means of technical difficulties. Thus our AIM is to measure the regional cerebral glucose metabolism(CMRglu) in patients with SHE using positron emission tomography(PET). METHODS: Three patients with non-alcoholic liver cirrhosis and SHE and 13 control subjects underwent PET imaging of the brain using 2-(~SF)-fluoro-2-deoxy-Dglucose). Neurological tests ; the digit symbol and block design subtests from Wechsler Adult Intelligence Scale, auditory brain stem reaction, and electro encephalogram were also undergone for diagnosis of SHE. Regions were defined in frontal, parietal, occipital, basal ganglia, and the white matter. RESULTS: CMRglu value in the gray matter of SHE are higher than that of control, especially significant in frontal and occipital CMRglu (p<0.05, p<0.01 respectively). There was no difference in basal ganglia and white matter. CONCLUSION: CMRglu value in the gray matter of SHE are depressed as compared to control. These data suggest that the abnormalities of cerebral glucose metabolism may be a contributing factor in SHE. Table: Regional Cerebral Glucose Metabolism frontal temporal occipital basal ganglia white matter SHE 5.5_+1.5" 5.1_+t.7 4.9_+0.6** 6.4-+0.6 4.0_+0.9 control 7.1_+0.8 7.0-+0.8 7.0_+0.8 6.4_+0.1 4.9+_0.8 *; P<0.05 **; p<0.01 (Unpaired t-test)
P-97
Laminin with type IV c o l l a g e n enhances malignant phenotype of h e p a t o m a cells Takuji TORIMURA, Takato UENO, Motoaki KIN, S a d a t a k a INUZUKA, Michio SATA, Kyuichi TANIKAWA 1), H i r o h i s a YANO, Masamichi KOJIRO 2) 2nd dept. of medicine 1) a n d 1st dept of pathology 2) , Kurume University School o f Medicine
Background~Aims : Laminin
(La) and type IV c o l l a g e n (Ty IV) e n h a n c e cell adhesion, migration and proliferation. Ty IV and La are present along t h e sinusoids in hepatocellular carcinoma. These e x t r a c e l l u l a r matrices may promote the m a l i g n a n t phenotype of hepatoma cells. We investigated t h e influences of La and Ty IV on cell migration, a d h e s i o n and p r o l i f e r a t i o n of hepatoma cells. Subjects a n d Methods : Hepatoma cell lines (KYN-I,2,3) were used i n this study. Immunoelectron microscopy was p e r f o r m e d to investigate the production o f L a a n d Ty IV by h e p a t o m a ceils. The influence of La and Ty IV on the m i g r a t i o n , adhesion and proliferation of hepatoma cells was also evaluated. Results : La and Ty IV were produced b y hepatoma cells. The haptotactic migration, adhesion a n d thymidine incorporation of h e p a t o m a cells w e r e stimulated by Ty IV or La. Furthermore, a combination of La and Ty IV enhanced these biological activities of hepatoma ceils much more than Ty IV a l o n e . Conclusions: Our results suggest that the presence of La with Ty IV produced by hepatoma cells enhances t h e malignant phenotype o f h e p a t o m a cells.
P-99
THE MECHANISM OF A M M O N I A - I N D U C E D
INCREASE IN CHLORIDE ION C O N C E N T R A T I O N IN R A T CULTURED H I P P O C A M P A L NEURONS. T.Irie 1)2), M.Hara 2), T.Yasukura 2), K. Inoue 1), C. Inagaki 2). i) Third Department of Internal Medicine and 2)Department of Pharmacology, Kansai Medical University, Osaka 570, Japan. Hepatic encephalopathy is a neuro-psychological syndrome caused by severe liver dysfunction. Hyperammonemia is one of the important pathogenic factors of this syndrome. Previously we reported that chronically applied ammonia induces an increase in intracellular chloride concentration ([Cl]i) in cultured hippocampal neurons, and the increase in [Cl']~ is diminished by an inhibitor of carbonic anhydrase, acetazolamide. In this study, we further examined the mechanisms of ammonia-induced increase in [Cl']i. Hippocampal neuronal cells from 17-day-old rat fetus were cultured and exposed to NH4CI for 1-3 days with or without test reagents. The [Cl]i was fluorometrically estimated using a C1--sensitive fluorescent dye. The exposure to 2mM NH4C1 for 1 day significantly increased [C[]i (13.7-+1.8mM, n=9) as compared to that without exposure (8.1 -+0.5mM, n=19) and that with additional 2raM of NaC1 (7.5_+1.3 mM, n=7) or KC[ (6.3_+1.3mM, n=7). The NH4Cl-induced increase in [Cl]i was abolished by SITS and DIDS (0.1mM each), inhibitors of C l /HCO 3" exchanger, but not by bumetanide (50 # M), an inhibitor of Na+/K÷/2C1 - cotransporter. Actinomycin D (1 # M), a mRNA synthesis inhibitor, and cycloheximide (0.5 # g/ml), a protein synthesis inhibitor, also inhibited the ammonia-induced increase in [Cl']i. These findings suggest that chronic exposure of hippoeampal neurons to ammonia increases [Cl']i mainly through the facilitated exchange of C l and HCO3, in a de novo protein synthesis-dependent mannor.
$61
P-98 LATE ONSET ORNITHINE TRANSCARBAMYLASE DEFICNECY IN MALES M. Yoshino, 1 A. NishiyorL 1 H. Kate, 1 R. Kumashiro, 2and K. Tanikawa.2 1 Department of Pediatrics and Child Health, and 2 Second Department of Medicine, Kurume University School of Medicine, Kurume, Japan Male patients with ornithine transcarbamylase (OTC) deficiency usually develop a hyperammonemic crisis in the neonatal period or in early infancy. In contrast to these classical patients, here we report the occurence of males with the disease who developed a hyperammonemic crisis in adolescence or in late adulthood. The molecular basis for lhe lateness of the onset of this disease in these patients was investigated. Of 5 cases, cases 1 through 4 each developed afirst episode between 16 and 58 years of age, had an OTC activity in the postmortem liver tissue that ranged from 2-9% of control mean, and each had decreased levels of the OTC protein. Analysis of the OTC gene disclosed an R40H mutation in cases 1, 3, and 4, and in agrandson (case 5) of amale sibling of case 1, and a Y55D mutation in case 2. The activity of the mutant OTCs expressed in cosf cells was 20% for the R4OH mulalion, and 30% for the Y55D mutation of the wild type OTC. When the mutant enzyme was treated by freezing and thawing, the R40H OTC decreased to 10% of the mean of the wild type, while the Y55D OTC remained essentially at the 30% level. These results suggested that these mutant enzymes were operating in vivo at a level consistent with low normal urea production, but this level declined due to their structural instability, after an environmental load was applied.
Dlflfl
x-~t~v CEREBRAL GLUCOSE METABOLISM IN PATIENTS WITH SUBCLINICAL HEPATIC ENCEPHALOPATHY USING POSITRON EMISSION TOMOGRAPHY A.Kato, Y.Kaneta, H.Obara, R.Endo, M.Iwai, O.Moriai, l.Nakadate, T.Yoshida, K.Suzuki, and S.Sato First Dep. of Int. Med., Iwate Med. Univ., Morioka, Japan Although it is assessed that abnormalities of glucose metabolism occur substantially throughout the brain in patients with subclinical hepatic encephalopathy (SHE), liule attention has been paid to these abnormalities in situ. by means of technical difficulties. Thus our AIM is to measure the regional cerebral glucose metabolism(CMRglu) in patients with SHE using positron emission tomography(PET). METHODS: Three patients with non-alcoholic liver cirrhosis and SHE and 13 control subjects underwent PET imaging of the brain using 2-(~SF)-fluoro-2-deoxy-Dglucose). Neurological tests ; the digit symbol and block design subtests from Wechsler Adult Intelligence Scale, auditory brain stem reaction, and electro encephalogram were also undergone for diagnosis of SHE. Regions were defined in frontal, parietal, occipital, basal ganglia, and the white matter. RESULTS: CMRglu value in the gray matter of SHE are higher than that of control, especially significant in frontal and occipital CMRglu (p<0.05, p<0.01 respectively). There was no difference in basal ganglia and white matter. CONCLUSION: CMRglu value in the gray matter of SHE are depressed as compared to control. These data suggest that the abnormalities of cerebral glucose metabolism may be a contributing factor in SHE. Table: Regional Cerebral Glucose Metabolism frontal temporal occipital basal ganglia white matter SHE 5.5_+1.5" 5.1_+t.7 4.9_+0.6** 6.4-+0.6 4.0_+0.9 control 7.1_+0.8 7.0-+0.8 7.0_+0.8 6.4_+0.1 4.9+_0.8 *; P<0.05 **; p<0.01 (Unpaired t-test)