- Email: [email protected]
Normal hepatic iron but increased hepatic copper in nash
144
Poster Sessions
of Hcy and its relationship to the presence of liver damage in a group of 287 chronic alcoholics (197 males and 89 females) in which serum Hcy was measured by HPLC. 116 patients (71 males and 45 females) had a normal liver and 170 (126 males and 44 females) had varying degrees of liver damage. The age of the patients of both groups was simlar. Serum Hey was significantly higher in patients with liver damage than in those with normal liver: males 11.6 -/- 8.6 vs 7.7 + 5.6 umol/l, p < 0.001; females: 10.6 -4- 9.5 vs 6.6 :t: 2.5 umol/l, p < 0.01. Moreover, the percentage of patients with liver damage and abnormal levels of Hcy was significantly higher than the percentage of patients with normal liver (24.1% vs 7.7%, p < 0.001). There were no differences in serum Hey between active alcoholics and those with a period of abstinence. In conclusion, serum levels of Hcy are frequently elevated in chronic alcoholics, being liver damage the main cause of hyperhomocysteinemia in these patients.
~ ' ~ 7 HEPATOPROTECTIVE AND ANTIOXIDANT EFFECT OF BORAGE OIL IN RATS WITH ALCOHOLIC STEATOSIS Vyacheslav Buko, Lubov Sushko, Alex Egorov, Yurji Popov. Institute of
Biochemistry, Grodno, Belarus, Europe Aims: The aim of the present study was to evaluate the effect of the oil extracted from Borago officinalis in rat alcoholic steatosis. Methods: Two groups of rats were treated with ethanol (4 g/kg per day, i.g., 30 days). One of the group received borage oil (120 mg/kg/day, i.g., 30 days). Results: The liver of ethanol-treated animals was characterized by fatty dystrophy. Liver triglyceride and cholesterol ester contents, activities of serum marker enzyme, alanine aminotransferase and gammaglutamyltransferase, were significantly increased. Ethanol increased chemiluminescence enhanced by luminol, hydroxyalkenals, malondialdehyde and cytochrome P-450 (CYP) content in microsomes as well as superoxide dismutase activity in cytosol. The increase in monoenoic fatty acids and the decrease in n-6 acid family in liver phospholipids was observed in ethanol-treated rats. The treatment with borage oil improved liver morphology, decreased triglyceride and cholesterol ester contents, normalized serum marker enzyme activities and fatty acid pattern in liver phospholipids. Borage oil developed the antioxidant effect decreasing all the above free radical-related parameters and normalizing CYP content compared to the ethanol-treated group. Conclusions: CYP is a main source of free oxygen radicals in the liver and we proposed that the antioxidant effect of borage oil is realized via the normalization of CYP content, where borage oil can improve lipid surrounding of CYP. In turn, hepatoprotection by borage oil in alcoholic steatosis is probably connected with its antioxidant properties.
~-]
EXPRESSION OF INDUCIBLE NITRIC OXIDE SYNTHASE IN RAT HEPATOCYTE CULTURES AND DOWN-REGULATION BY CYCLOSPORIN A AND FKS06 OCCUR AT DIFFERENT LEVELS
Nikolina Canova, Eva Kmonickova, Petra Strestikova, Hassan Farghali.
Institute of Pharmacology, 1st Faculty of Medicine, Charles University, Prague, Czech Republic Aim: To compare the effect of cyclosporin A (CsA) and FK506 on the expression ofiNOS in hepatocyte cultures exposed to inflammatory signals. Method: Isolated rat hepatocytes were cultured for 24 hours with or without LPS (5 microg/ml) or cytokine mixture (CM: TNF-alpha 500 U/ml, IL-lbeta 200 U/ml, IFN-gama 100 U/ml) in the absence or presence of CsA or FK506. Total RNA was isolated and the expression of iNOS was evaluated by RT-PCR or by NO2- production in culture supernatant. Urea synthesis and ALT leakage from hepatocytes were measured. Results: LPS or CM increased iNOS mRNA that was not affected by FK506 pre-treatment while CsA pre-treatment reduced its level. CsA (0.1 and 1.0 microg/ml) reduced the LPS- and CM-induced increase of NO2- by
21% and 32% respectively, ALT leakage was increased and urea synthesis was reduced. FK506 (0.1 and 1.0 microg/ml) decreased levels of NO2- by 8% and 21%, respectively. Conclusion: In hepatocytes CsA exhibits its inhibitory effect on a phosphatase at the transcriptional level, while iNOS expression down-regulation by FK506 occurred posttranscriptional. Moreover, NO produced during inflammatory signals might be protective to hepatocytes and the inhibition of iNOS by CsA and b7(506 should be considered in practice.
•
]
EVALUATION OF THE FUNCTIONALITY AND CYCLOSPORIN A BIOTRANSFORMATION IN LABORATORY SCALE HEPATOCYTE BIOREACTORS
Nikolina Canova, Eva Kmonickova, Dana Lincova, Hassan Farghali.
Institute of Pharmacology, 1st Faculty of Medicine, Charles University, Prague, Czech Republic Background: Apparently hepatocyte culture models are metabolically less active, due to inefficient oxygenation and waste product buildup. Since several years, we are using immobilized and perfused hepatocytes in agarose gel as a prototype of hepatocyte bioreactor in various biomedical studies. Goal: The present study was directed to assess and compare the functionality of recently described flat membrane bioreactors [1] with hepatocyte in monolayer culture. Methods: Flat membrane bioreactors were prepared from isolated rat hepatocytes and compared to hepatocytes in primary culture. The functional and metabolic competence of hepatocytes in various models were tested by urea synthesis, ALT leakage and cyclosporin A (CsA) biotransformation (HPLC-assessed) during 48-72 hours. Results: Urea synthesis increased several-fold, ALT leakage was not increased 20 hours after the operation of the bioreactors and CsA concentration decreased to 20% of the initial value in perfusate. The use of hepatocyte bioreactors demonstrated that cells are more stable in a bioreactor models than in a conventional hepatocyte culture. Conclusion: Hepatocyte bioreactors may become popular in hepatology research due to the better performance of perfused hepatocytes as compared to conventional culture. Moreover their potentials as bioartificial liver for bridged therapy are now under extensive investigation. References [1] Langsch A and Bader A: Biotechnotogy and Bioengineering, 2001; 76, 115-125.
[-~
NORMAL HEPATIC IRON BUT INCREASED HEPATIC COPPER IN NASH
Blanka Cieslarova l , Jiri Horak 1, Jan Stritesky 2, Ivana Putova 3, Vladimir Palicka 4. 1Department of Medicine I., 3rd Faculty of Medicine,
Charles University, Prague; 2Institute of Pathology, 3rd Faculty of Medicine, Charles University, Prague; 3Institute of Biology, 3rd Faculty of Medicine, Charles University,Prague; 41stitute of Biochemistry, Faculty of Medicine Charles University, Hradec Kralove, Czech Republic On the ground of medical history, clinical laboratory examination and liver biopsy the diagnosis of nonalcoholic steatohepatitis (NASH) was established in 18 patients (15 males and 3 females). Obesity, NIDDM and hyperlipidemia as risk factors for steatosis/steatohepatitis were present in this group of patients either all or in various combinations. In females the ratio of AST:ALT was 1.02-2.6, in males 0.46-2.5. Enzyme activities roughly correlated with hepatocyte damage on liver biopsy. Serological markers of iron and copper metabolism were not suggestive of hemochromatosis or Wilson's disease. Liver biopsy specimens were measured for iron (18/18) and copper (7/18) contents. The hepatic levels of iron lay between 70.31345 mikrog/g of dry liver tissue (normal value < 1400 mikrog/g) and the hepatic iron index was 0.143-0.79 (normal value < 1.0). In 8/18 patients genetic examination of the HFE gene was performed. Two out of 8 were
Category 8: Nutrition, metabolism, alcoholic liver disease, pharmacology H63D heterozygotes and one was a C282Y heterozygote. The hepatic Copper content was increased in 6/7 examinated biopsy specimens. The levels ranged between 9-133 mikrog/g dry liver tissue;median was 86 mikrog/g (normal value < 50). Conclusion: In this group of NASH patients, hepatic copper concentration was increased contrary to hepatic iron, which was normal.
•
VITRO- FELODIPIN 0PHAGOCYTOSIS 7ANDEFFECTS 1 NON LEUCOCYTES BACTERICIDAL ACTIVITY IN ALCOHOLIC LIVER DISEASE (ALD)
145
1.3% - AC) expression than in controls and ALS patients. The length of alcohol abuse was significant higher on CAH (17.7 4- 0.4 yrs) and AC patients (22.3 4- 1.6 yrs) than those of ALS patients (10.8 4- 0.8 yrs). The significant increase (p < 0.05) of activation markers expression, on CAH and ALC patients is correlated with the long exposure, more than 17 years, to antigenic products resulted from the alcohol metabolism and with the intensity of liver injuries.
2-'] DYSLIPIDEMIAS AS MARKER OF CHRONIC ALCOHOLIC LIVER DISEASES
Daniella Cocarla 1, Cora Uram-Tuculescu 1 Aurelian Duran I, Claudiu Cornea 2, Dan Stef 2. 1Department of Physiology, University of
Elena Adorata Coman 1, Rodica Petrovanu l, Georgeta Palel 1, Cynthia Petrovanu t , Didona Ungureanu 2. 1Ambulatory Care Department,
Medicine and Pharmacy, Timisoara; 2University of Medicine and Pharmacy, ~misoara, Romania
University of Medicine Iasi; 2Biochemistry Department, University of Medicine, Iasi, Romania
We investigated in the peripheral blood of 12 alcoholic liver steatosis (ALS), 21 chronic alcoholic hepatitis (CAH), 8 alcoholic chirrosis (AC) patients and 10 healthy people as controls the effects of Felodipin (calcium channels antagonist) on leukocyte phagocytosis and bactericidal activity. Phagocytic capacity was assessed by Latex Phagocytic Index (LPI%) and nitrobhie tetrasolium (NBT%) tests and the bactericidal activity by indirect measuring of reactive oxygen species (ROS) production, using SOD and GSH tests. Testes were performed in basal conditions, after 30 min. preincubation with 0.1, 0.2, 1 ml solutions of felodipin and after 45 min with 0.1 ml solution. There were significant decreased (p < 0.05) on phagocytic tests (PT) in AC patients than those in controls, ALS and CAH patients, without significant decreases of PT on CAH and ALS patients than those in controls. Preincubation of controls samples with felodipin for 30' and 45' leaded to significant decreases (p < 0.0001) of PT than those in basal conditions (NBT = 10.5 4- 0.8%; LPI = 61.63 4- 2.5%). There weren't significant variations on SOD and GSH levels after preincubation of felodipin solutions than those in basal conditions (SOD = 5.59 4- 0.7 U/l; GSH = 537 4- 26.5 microM/l). The PT of ALD patients registered a significant decrease (p < 0.05) in a dose dependent manner after preincubation with felodipin solutions, than those in basal conditions, but not in a time manner. There weren't significant variations (p > 0.05) on SOD and GSH levels after felodipin preincubation in the blood of ALD patients than those in controls and basal conditions. Felodipin decreases leukocyte phagocytosis already decreased by alcohol effects but don't modify the ROS production.
Aims: We studied the seric lipoproteins on a population with intake of alcohol. Method: We considered 140 cases selected from the patients who benefited of medical care in our services (1999-2000). We measured cholesterol, tryglicerides, LDLchol, HDLchol, apolipoproteins A, B and made correlation with another markers of chronic alcoholic diseases, MVC (mean red cell volume) ALAT, ASAT, gammaGT, IgA (measured by immunelectroforesis). We also made a correct anamnesis of alcohol intake and abdominal echography. We considered chronic intake of alcohol, more than 50 g per day. 50 persons were for control, without intake of alcohol. Results: 78% of cases had high level of tryglicerides (210 4- 48 mg/dl), only 34% high cholesterol (230 4- 46 mg/dl), 48% high LDLchol (180 423 mg/dl) and apoB (3.24 mg/dl), and only 12% low level of HDLchol (46 + 37 mg/dl) and apoA (1.28 mg/dl). We obtained good correlation between level of tryglicerides and intake of alcohol (p < 0.0001), high level of IgA (p < 0.001), high ALAT (mean values 78 4- 24UI/1) (p < 0.001), fatty liver on echography (0.001), MCV > 98 micro-meterscubes (p < 0.01). Conclusions: Our findings underline that dyslipidernias with high level of tryglicerides, moderate level of cholesterol, without low level of HDL chol and apoA are a good marker of chronic alcohol intake with fatty liver, as good as the other markers. We have to consider the constitutional dyslipidemias that could influences the values.
~--~ ~ 2 " ~ THE CORRELATION OF SPECIFIC IMMUNE SYSTEM ACTIVATION WITH LENGTH OF ALCOHOL ADDICTION IN PATIENTS WITH ALCOHOLIC LIVER DISEASE D. Cocarla, A. Duran, C. Uram-Tuculescu, C. Cornea, D. Stef.
Department of Physiology, University of Medicine and Pharmacy, Timisoara, Romania The peripheral blood lyrnphocytes from alcoholic liver disease (ALD) patients disclose an overexpression of T lymphocytes activation markers as it has been described in autoimmune diseases. We investigated the correlation between the length of alcohol misuse, the degree of T lymphocytes activation and the level of hepatic injuries. We investigated in peripheral blood of 12 alcoholic liver steatosis (ALS) patients, 21 chronic alcoholic hepatitis (CAH), 8 alcoholic cirrhosis (AC) patients and 10 healthy people as controls the expression of T lymphocytes subsets markers CD3, CD3/CD4+, CD3/CD8+ and the activation markers HLA-DR and CD3/CD25 by flowcytometry. We assessed spectrophotometricaly the GGT, AST and ALT and the duration of alcohol abuse were anamnestic obtained. The results indicated no significant increases (p > 0.05) in transaminases blood levels and AST/ALT and GGT/ALT ratio > 1 when AST < 100 U/L underlined the chronic alcohol abuse. In the peripheral blood of CAH and AC patients there were significant increases (p < 0.05) of CD3/CDS+ cells (26.59 41.13% - CAH; 27.62 d- 1.9% - AC) and HLA-DR (10.06 4- 3.66% CAH; 15.09 4- 3.3% - AC) CD3/CD25 (25.4 4- 1.18% - CAH; 24.88 4-
LIVER FREE FATTY ACIDS IN HUMAN NONALCOHOLIC STEATOHEPATITIS (NASH)
Isabel Tavares de Almeida 1, Helena Cortez Pinto 2, Miguel Carneiro de Moura 2, Ermelinda Camilo 3. 1Centro de Patogenese Molecular, FFUL,
Lisboa; 2Dept. Medicina 2, Hosp. Sta. Maria, Lisboa; 3Centro de Nutricao e Metabolismo, FMUL, Lisboa, Portugal Liver function damage in NASH has been ascribed to hepatic lipid accumulation. The present study was performed to evaluate the free fatty acid (FFA) composition of liver extracts from NASH patients' biopsies with different degrees of histological liver damage: liver biopsies from 11 patients classified as NASH were analysed. Among them 4 had steatohepatitis (Group A), 7 steatosis (Group B, n = 5), and in the latter group 2 had minor alterations (Group C). FFAs were extracted from liver bomogenate and further derivatised. FFA derivatives were analysed by gaschromatography/mass spectrometry, SIM mode. An increased total FFA concentration (mean 4- sem; nmoles/mg protein) was observed in accordance to the lesion severity: Group A: 238 4- 63.5; Group B: 151 4- 18.7; Group C: 96 4- 6.7. Similarly, we did observe a trend towards an increased concentration of palmitic acid (C16:0) and oleic acid (C18:1), the predominant FFA within the groups: Group A: 58.7 4- 15.4; 49.5 + 19.7; Group B: 51.0 4- 2.0; 38.0 4- 3.7; Group C: 27.0 4- 2.1; 25.0 + 6.3. linoleic acid (C18:2) was moderately increased (15.3 > 11.8 > 8.6). Our results suggest that FFA may represent potential bid-markers for liver dysfunction progression
Poster Sessions
of Hcy and its relationship to the presence of liver damage in a group of 287 chronic alcoholics (197 males and 89 females) in which serum Hcy was measured by HPLC. 116 patients (71 males and 45 females) had a normal liver and 170 (126 males and 44 females) had varying degrees of liver damage. The age of the patients of both groups was simlar. Serum Hey was significantly higher in patients with liver damage than in those with normal liver: males 11.6 -/- 8.6 vs 7.7 + 5.6 umol/l, p < 0.001; females: 10.6 -4- 9.5 vs 6.6 :t: 2.5 umol/l, p < 0.01. Moreover, the percentage of patients with liver damage and abnormal levels of Hcy was significantly higher than the percentage of patients with normal liver (24.1% vs 7.7%, p < 0.001). There were no differences in serum Hey between active alcoholics and those with a period of abstinence. In conclusion, serum levels of Hcy are frequently elevated in chronic alcoholics, being liver damage the main cause of hyperhomocysteinemia in these patients.
~ ' ~ 7 HEPATOPROTECTIVE AND ANTIOXIDANT EFFECT OF BORAGE OIL IN RATS WITH ALCOHOLIC STEATOSIS Vyacheslav Buko, Lubov Sushko, Alex Egorov, Yurji Popov. Institute of
Biochemistry, Grodno, Belarus, Europe Aims: The aim of the present study was to evaluate the effect of the oil extracted from Borago officinalis in rat alcoholic steatosis. Methods: Two groups of rats were treated with ethanol (4 g/kg per day, i.g., 30 days). One of the group received borage oil (120 mg/kg/day, i.g., 30 days). Results: The liver of ethanol-treated animals was characterized by fatty dystrophy. Liver triglyceride and cholesterol ester contents, activities of serum marker enzyme, alanine aminotransferase and gammaglutamyltransferase, were significantly increased. Ethanol increased chemiluminescence enhanced by luminol, hydroxyalkenals, malondialdehyde and cytochrome P-450 (CYP) content in microsomes as well as superoxide dismutase activity in cytosol. The increase in monoenoic fatty acids and the decrease in n-6 acid family in liver phospholipids was observed in ethanol-treated rats. The treatment with borage oil improved liver morphology, decreased triglyceride and cholesterol ester contents, normalized serum marker enzyme activities and fatty acid pattern in liver phospholipids. Borage oil developed the antioxidant effect decreasing all the above free radical-related parameters and normalizing CYP content compared to the ethanol-treated group. Conclusions: CYP is a main source of free oxygen radicals in the liver and we proposed that the antioxidant effect of borage oil is realized via the normalization of CYP content, where borage oil can improve lipid surrounding of CYP. In turn, hepatoprotection by borage oil in alcoholic steatosis is probably connected with its antioxidant properties.
~-]
EXPRESSION OF INDUCIBLE NITRIC OXIDE SYNTHASE IN RAT HEPATOCYTE CULTURES AND DOWN-REGULATION BY CYCLOSPORIN A AND FKS06 OCCUR AT DIFFERENT LEVELS
Nikolina Canova, Eva Kmonickova, Petra Strestikova, Hassan Farghali.
Institute of Pharmacology, 1st Faculty of Medicine, Charles University, Prague, Czech Republic Aim: To compare the effect of cyclosporin A (CsA) and FK506 on the expression ofiNOS in hepatocyte cultures exposed to inflammatory signals. Method: Isolated rat hepatocytes were cultured for 24 hours with or without LPS (5 microg/ml) or cytokine mixture (CM: TNF-alpha 500 U/ml, IL-lbeta 200 U/ml, IFN-gama 100 U/ml) in the absence or presence of CsA or FK506. Total RNA was isolated and the expression of iNOS was evaluated by RT-PCR or by NO2- production in culture supernatant. Urea synthesis and ALT leakage from hepatocytes were measured. Results: LPS or CM increased iNOS mRNA that was not affected by FK506 pre-treatment while CsA pre-treatment reduced its level. CsA (0.1 and 1.0 microg/ml) reduced the LPS- and CM-induced increase of NO2- by
21% and 32% respectively, ALT leakage was increased and urea synthesis was reduced. FK506 (0.1 and 1.0 microg/ml) decreased levels of NO2- by 8% and 21%, respectively. Conclusion: In hepatocytes CsA exhibits its inhibitory effect on a phosphatase at the transcriptional level, while iNOS expression down-regulation by FK506 occurred posttranscriptional. Moreover, NO produced during inflammatory signals might be protective to hepatocytes and the inhibition of iNOS by CsA and b7(506 should be considered in practice.
•
]
EVALUATION OF THE FUNCTIONALITY AND CYCLOSPORIN A BIOTRANSFORMATION IN LABORATORY SCALE HEPATOCYTE BIOREACTORS
Nikolina Canova, Eva Kmonickova, Dana Lincova, Hassan Farghali.
Institute of Pharmacology, 1st Faculty of Medicine, Charles University, Prague, Czech Republic Background: Apparently hepatocyte culture models are metabolically less active, due to inefficient oxygenation and waste product buildup. Since several years, we are using immobilized and perfused hepatocytes in agarose gel as a prototype of hepatocyte bioreactor in various biomedical studies. Goal: The present study was directed to assess and compare the functionality of recently described flat membrane bioreactors [1] with hepatocyte in monolayer culture. Methods: Flat membrane bioreactors were prepared from isolated rat hepatocytes and compared to hepatocytes in primary culture. The functional and metabolic competence of hepatocytes in various models were tested by urea synthesis, ALT leakage and cyclosporin A (CsA) biotransformation (HPLC-assessed) during 48-72 hours. Results: Urea synthesis increased several-fold, ALT leakage was not increased 20 hours after the operation of the bioreactors and CsA concentration decreased to 20% of the initial value in perfusate. The use of hepatocyte bioreactors demonstrated that cells are more stable in a bioreactor models than in a conventional hepatocyte culture. Conclusion: Hepatocyte bioreactors may become popular in hepatology research due to the better performance of perfused hepatocytes as compared to conventional culture. Moreover their potentials as bioartificial liver for bridged therapy are now under extensive investigation. References [1] Langsch A and Bader A: Biotechnotogy and Bioengineering, 2001; 76, 115-125.
[-~
NORMAL HEPATIC IRON BUT INCREASED HEPATIC COPPER IN NASH
Blanka Cieslarova l , Jiri Horak 1, Jan Stritesky 2, Ivana Putova 3, Vladimir Palicka 4. 1Department of Medicine I., 3rd Faculty of Medicine,
Charles University, Prague; 2Institute of Pathology, 3rd Faculty of Medicine, Charles University, Prague; 3Institute of Biology, 3rd Faculty of Medicine, Charles University,Prague; 41stitute of Biochemistry, Faculty of Medicine Charles University, Hradec Kralove, Czech Republic On the ground of medical history, clinical laboratory examination and liver biopsy the diagnosis of nonalcoholic steatohepatitis (NASH) was established in 18 patients (15 males and 3 females). Obesity, NIDDM and hyperlipidemia as risk factors for steatosis/steatohepatitis were present in this group of patients either all or in various combinations. In females the ratio of AST:ALT was 1.02-2.6, in males 0.46-2.5. Enzyme activities roughly correlated with hepatocyte damage on liver biopsy. Serological markers of iron and copper metabolism were not suggestive of hemochromatosis or Wilson's disease. Liver biopsy specimens were measured for iron (18/18) and copper (7/18) contents. The hepatic levels of iron lay between 70.31345 mikrog/g of dry liver tissue (normal value < 1400 mikrog/g) and the hepatic iron index was 0.143-0.79 (normal value < 1.0). In 8/18 patients genetic examination of the HFE gene was performed. Two out of 8 were
Category 8: Nutrition, metabolism, alcoholic liver disease, pharmacology H63D heterozygotes and one was a C282Y heterozygote. The hepatic Copper content was increased in 6/7 examinated biopsy specimens. The levels ranged between 9-133 mikrog/g dry liver tissue;median was 86 mikrog/g (normal value < 50). Conclusion: In this group of NASH patients, hepatic copper concentration was increased contrary to hepatic iron, which was normal.
•
VITRO- FELODIPIN 0PHAGOCYTOSIS 7ANDEFFECTS 1 NON LEUCOCYTES BACTERICIDAL ACTIVITY IN ALCOHOLIC LIVER DISEASE (ALD)
145
1.3% - AC) expression than in controls and ALS patients. The length of alcohol abuse was significant higher on CAH (17.7 4- 0.4 yrs) and AC patients (22.3 4- 1.6 yrs) than those of ALS patients (10.8 4- 0.8 yrs). The significant increase (p < 0.05) of activation markers expression, on CAH and ALC patients is correlated with the long exposure, more than 17 years, to antigenic products resulted from the alcohol metabolism and with the intensity of liver injuries.
2-'] DYSLIPIDEMIAS AS MARKER OF CHRONIC ALCOHOLIC LIVER DISEASES
Daniella Cocarla 1, Cora Uram-Tuculescu 1 Aurelian Duran I, Claudiu Cornea 2, Dan Stef 2. 1Department of Physiology, University of
Elena Adorata Coman 1, Rodica Petrovanu l, Georgeta Palel 1, Cynthia Petrovanu t , Didona Ungureanu 2. 1Ambulatory Care Department,
Medicine and Pharmacy, Timisoara; 2University of Medicine and Pharmacy, ~misoara, Romania
University of Medicine Iasi; 2Biochemistry Department, University of Medicine, Iasi, Romania
We investigated in the peripheral blood of 12 alcoholic liver steatosis (ALS), 21 chronic alcoholic hepatitis (CAH), 8 alcoholic chirrosis (AC) patients and 10 healthy people as controls the effects of Felodipin (calcium channels antagonist) on leukocyte phagocytosis and bactericidal activity. Phagocytic capacity was assessed by Latex Phagocytic Index (LPI%) and nitrobhie tetrasolium (NBT%) tests and the bactericidal activity by indirect measuring of reactive oxygen species (ROS) production, using SOD and GSH tests. Testes were performed in basal conditions, after 30 min. preincubation with 0.1, 0.2, 1 ml solutions of felodipin and after 45 min with 0.1 ml solution. There were significant decreased (p < 0.05) on phagocytic tests (PT) in AC patients than those in controls, ALS and CAH patients, without significant decreases of PT on CAH and ALS patients than those in controls. Preincubation of controls samples with felodipin for 30' and 45' leaded to significant decreases (p < 0.0001) of PT than those in basal conditions (NBT = 10.5 4- 0.8%; LPI = 61.63 4- 2.5%). There weren't significant variations on SOD and GSH levels after preincubation of felodipin solutions than those in basal conditions (SOD = 5.59 4- 0.7 U/l; GSH = 537 4- 26.5 microM/l). The PT of ALD patients registered a significant decrease (p < 0.05) in a dose dependent manner after preincubation with felodipin solutions, than those in basal conditions, but not in a time manner. There weren't significant variations (p > 0.05) on SOD and GSH levels after felodipin preincubation in the blood of ALD patients than those in controls and basal conditions. Felodipin decreases leukocyte phagocytosis already decreased by alcohol effects but don't modify the ROS production.
Aims: We studied the seric lipoproteins on a population with intake of alcohol. Method: We considered 140 cases selected from the patients who benefited of medical care in our services (1999-2000). We measured cholesterol, tryglicerides, LDLchol, HDLchol, apolipoproteins A, B and made correlation with another markers of chronic alcoholic diseases, MVC (mean red cell volume) ALAT, ASAT, gammaGT, IgA (measured by immunelectroforesis). We also made a correct anamnesis of alcohol intake and abdominal echography. We considered chronic intake of alcohol, more than 50 g per day. 50 persons were for control, without intake of alcohol. Results: 78% of cases had high level of tryglicerides (210 4- 48 mg/dl), only 34% high cholesterol (230 4- 46 mg/dl), 48% high LDLchol (180 423 mg/dl) and apoB (3.24 mg/dl), and only 12% low level of HDLchol (46 + 37 mg/dl) and apoA (1.28 mg/dl). We obtained good correlation between level of tryglicerides and intake of alcohol (p < 0.0001), high level of IgA (p < 0.001), high ALAT (mean values 78 4- 24UI/1) (p < 0.001), fatty liver on echography (0.001), MCV > 98 micro-meterscubes (p < 0.01). Conclusions: Our findings underline that dyslipidernias with high level of tryglicerides, moderate level of cholesterol, without low level of HDL chol and apoA are a good marker of chronic alcohol intake with fatty liver, as good as the other markers. We have to consider the constitutional dyslipidemias that could influences the values.
~--~ ~ 2 " ~ THE CORRELATION OF SPECIFIC IMMUNE SYSTEM ACTIVATION WITH LENGTH OF ALCOHOL ADDICTION IN PATIENTS WITH ALCOHOLIC LIVER DISEASE D. Cocarla, A. Duran, C. Uram-Tuculescu, C. Cornea, D. Stef.
Department of Physiology, University of Medicine and Pharmacy, Timisoara, Romania The peripheral blood lyrnphocytes from alcoholic liver disease (ALD) patients disclose an overexpression of T lymphocytes activation markers as it has been described in autoimmune diseases. We investigated the correlation between the length of alcohol misuse, the degree of T lymphocytes activation and the level of hepatic injuries. We investigated in peripheral blood of 12 alcoholic liver steatosis (ALS) patients, 21 chronic alcoholic hepatitis (CAH), 8 alcoholic cirrhosis (AC) patients and 10 healthy people as controls the expression of T lymphocytes subsets markers CD3, CD3/CD4+, CD3/CD8+ and the activation markers HLA-DR and CD3/CD25 by flowcytometry. We assessed spectrophotometricaly the GGT, AST and ALT and the duration of alcohol abuse were anamnestic obtained. The results indicated no significant increases (p > 0.05) in transaminases blood levels and AST/ALT and GGT/ALT ratio > 1 when AST < 100 U/L underlined the chronic alcohol abuse. In the peripheral blood of CAH and AC patients there were significant increases (p < 0.05) of CD3/CDS+ cells (26.59 41.13% - CAH; 27.62 d- 1.9% - AC) and HLA-DR (10.06 4- 3.66% CAH; 15.09 4- 3.3% - AC) CD3/CD25 (25.4 4- 1.18% - CAH; 24.88 4-
LIVER FREE FATTY ACIDS IN HUMAN NONALCOHOLIC STEATOHEPATITIS (NASH)
Isabel Tavares de Almeida 1, Helena Cortez Pinto 2, Miguel Carneiro de Moura 2, Ermelinda Camilo 3. 1Centro de Patogenese Molecular, FFUL,
Lisboa; 2Dept. Medicina 2, Hosp. Sta. Maria, Lisboa; 3Centro de Nutricao e Metabolismo, FMUL, Lisboa, Portugal Liver function damage in NASH has been ascribed to hepatic lipid accumulation. The present study was performed to evaluate the free fatty acid (FFA) composition of liver extracts from NASH patients' biopsies with different degrees of histological liver damage: liver biopsies from 11 patients classified as NASH were analysed. Among them 4 had steatohepatitis (Group A), 7 steatosis (Group B, n = 5), and in the latter group 2 had minor alterations (Group C). FFAs were extracted from liver bomogenate and further derivatised. FFA derivatives were analysed by gaschromatography/mass spectrometry, SIM mode. An increased total FFA concentration (mean 4- sem; nmoles/mg protein) was observed in accordance to the lesion severity: Group A: 238 4- 63.5; Group B: 151 4- 18.7; Group C: 96 4- 6.7. Similarly, we did observe a trend towards an increased concentration of palmitic acid (C16:0) and oleic acid (C18:1), the predominant FFA within the groups: Group A: 58.7 4- 15.4; 49.5 + 19.7; Group B: 51.0 4- 2.0; 38.0 4- 3.7; Group C: 27.0 4- 2.1; 25.0 + 6.3. linoleic acid (C18:2) was moderately increased (15.3 > 11.8 > 8.6). Our results suggest that FFA may represent potential bid-markers for liver dysfunction progression