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Serum laminin in patients with chronic active hepatitis and mild liver diseases
InternationalHepatologyCommunications,1994; 2: 52-55 0 1994 Elsevier Science B.V. All rights reserved 0928-4346/94/%07.00
52
HEPCOM 00058
Preliminary note
Serum laminin in patients with chronic active hepatitis and mild liver diseases Julio Collazos and Fernando Diaz Servicio de Medicina Interna, Hospital de Galdhcano, Vizcaya, Spain
(Received 29 July 1993; accepted 3 September 1993)
Laminin is a component of the basement membranes synthesized by Ito cells. We have measured the serum levels of laminin in 15 chronic active hepatitis patients and in 29 patients with mild liver diseases, who also underwent a complete clinical and laboratory evaluation. Abnormal levels of laminin were observed in 13.3% of chronic active hepatitis and in 10.3%of mild liver disease patients. Patients with chronic active hepatitis had higher serum levels of laminin than patients with mild liver diseases (P = 0.016). However, the usefulness of laminin to differentiate between these two groups was limited. Highly significant correlations were found between laminin and total protein (r = 0.79, P < O.OOOl),y-globulins (r = 0.78, P < O.OOOl),and IgG (r = 0.73, P = 0.0001) in the chronic active hepatitis group and lower in the mild liver diseases group. A possible role of defective immunoregulation in the increase of laminin is suggested. Key words: Basement membrane; Chronic liver disease; Liver test; Immunoregulation
Laminin is a component of the basement membranes synthesized by Ito cells which are located in the space of Disse [l]. Serum laminin has been related to portal venous pressure [2], fibrosing activity of the liver [3,4] and alcohol abuse [4,5]. However, some not well known factors have an important influence on laminin serum levels [6]. We have not found any study reported in the literature especially devoted to the evaluation of laminin in patients with chronic active hepatitis (CAH) and mild liver diseases (MLD). We have measured the serum levels of laminin in a group of patients with untreated CAH and MLD, who also underwent a thorough clinical and biochemical evaluation, in order to elucidate the possible usefulness of laminin to differentiate between these two groups.
Correspondence to: Dr. J. Collazos, Servicio de Medicina Interna,
Vizcaya, Spain.
Hospital de GaldBcano, 48960
SERUM LAMININ IN CHRONIC HEPATITIS
53
Materials and Methods Forty-four consecutive nonbiopsied patients that were electively admitted for liver biopsy because of chronic alterations of liver tests persisting for more than 6 months and that were diagnosed as having CAH or MLD after biopsy, were included in the study. Fifteen patients had CAH and the remaining 29 patients had MLD. Of the latter patients, 15 had chronic persistent hepatitis, five had steatosis and nine had minimal changes in liver biopsy. Twenty-seven patients were males (eight CAH and 19 MLD) and 17 females (seven CAH and ten MLD). The ages ranged between 16 and 76 years (mean 40.8 years) with a similar distribution between the two groups. Informed consent was obtained from each patient. Also, the study protocol, previously approved by the Ethical Committee of our hospital, conformed to the ethical guidelines of the Declaration of Helsinki. Blood was collected simultaneously for all the laboratory determinations. Laminin was measured by a radioimmunoassay (Hoechst-Behring, Frankfurt). The upper limit of normal was established at 1.8 U/ml which corresponded to the 95 percentile of a control group composed by 8 1 HBsAg-negative healthy individuals with ages ranging between 18 and 64 years (mean 40.9). Statistical analysis A preliminary test revealed that the distribution of laminin was not Gaussian. Accordingly, nonparametric tests were used. The Spearman’s rank correlation coefficient was used for correlations between continuous variables, the Mann-Whitney U test to compare two groups, and the Kruskal-Wallis test to compare more than two groups. A P < 0.05 level for a two-sided test was considered statistically significant.
Results Five patients (11.4%) had moderately increased serum levels of laminin. Table 1 shows the abnormal values of laminin according to different cut-off levels in CAH and MLD. Age or sex did not significantly influence the laminin levels in our patients. Patients with CAH had higher laminin serum levels than those with MLD (P = 0.016).
TABLE 1 Serum levels of laminin at different cut-offs Total
CAH MLD All
Laminin (U/ml) >l.O
>1.2
>1.4
>1.8
(n)
n (%)
n (o/o)
n (%)
n (%)
15 29 44
11 (73.3) 10 (34.5) 21 (47.7)
6 (40.0) 5 (17.2) 11 (25.0)
5 (33.3) 4 (13.8) 9 (20.5)
2 (13.3) 3 (10.3) 5 (11.4)
CAH denotes chronic active hepatitis, and MLD, mild liver diseases.
Range
Median
0.6-3.0 0.5-2.3 0.5-3.0
1.2 1.0
1.05
54
J. COLLAZOS
AND F. DIAZ
We did not find any significant correlation between laminin and hematocrit, mean corpuscular volume, prothrombin activity, AST, ALT, alkaline phosphatase, GGT, LDH, bilirubin, glycocholic acid, albumin, IgA or IgM in either the CAH or the MLD patients. On the contrary, it was found with total protein (CAH r = 0.79, P < 0.0001; MLD not significant), y-globulins (CAH r = 0.78, P < 0.0001; MLD r = 0.37, P = 0.04) and IgG (CAH r = 0.73, P = 0.0001; MLD r = 0.43, P = 0.01). No significant association was found between laminin and etiology, viral markers, smoking or alcoholic habit, cholelithiasis, hepato- or splenomegaly, drug addiction, urinary bilirubin or urobilinogen. Patients with cytolysis, as measured by aminotransferases, did not have significantly higher laminin levels than those without cytolysis, and the same was true for patients with or without cholestasis. The usefulness of laminin to predict CAH as compared with MLD was evaluated at the same cut-offs used in Table 1. Sensitivity was 73.3,40, 33.3 and 13.3%, and specificity was 65.5, 82.8, 86.2 and 89.7%, respectively, with efficiency of 68.2% for all cut-offs. The receiver operating characteristic (ROC) curve showed that the most discriminant value of laminin to differentiate CAH from MLD was 1 U/ml.
Discussion In our series, patients with CAH had significantly higher serum levels of laminin than patients with MLD. However, although the specificity was high using 1.8 U/ml as a cut-off, the sensitivity was too low to be clinically useful. Schneider et al. [3] have also observed higher levels of laminin in CAH and in active cirrhosis than in other chronic liver diseases. The mechanism whereby laminin increases in liver diseases appears to be multifactorial including increased synthesis by Ito cells [l] and poor metabolization due to liver disease [6]. Although alcohol intake has been related to abnormal levels of laminin [4,5] we failed to find any association with alcohol intake or with biochemical alcohol markers. Also, neither cytolysis nor cholestasis appeared to increase the levels of laminin in our patients. Noteworthy is the highly significant correlation found by us with total protein, y-globulins and IgG, especially in the CAH patients, with correlation’s coefficients close to 0.8. We have not found such association to be reported. IgG increases more frequently than other immunoglobulins in CAH patients [7,8]. On the other hand, a disordered immunoregulation with loss of suppressor-cell activity has been found in CAH patients [9] that could be the cause of this increase. It would be interesting to speculate about the role that this defective immunoregulation could have in the release of laminin from Ito cells. However, further studies are necessary to clarify this association. We conclude that although CAH patients have higher laminin serum levels than MLD patients, the usefulness of laminin to differentiate between these two entities is limited. An important correlation appears to exist between laminin levels and IgG, especially in CAH patients.
SERUM LAMININ IN CHRONIC HEPATITIS
55
References 1 Maher JJ, Friedman SL, Roll FJ, Bissell DM. Immunolocalization of laminin in normal rat liver and biosynthesis of laminin by hepatic lipocytes in primary culture. Gastroenterology 1988;94:1053-1062. 2 Gressner AM, Tittor W. Serum laminin - its concentration increases with portal hypertension in cirrhotic liver disease. Klin Wochenschr 1986;64:1240-1248. 3 Schneider M, Voss B, Hogeman B, Eberhardt G, Gerlach U. Evaluation of serum laminin P-l, procollagenpeptides, and N-acetyl-beta-glucosaminidase for monitoring the activity of liver fibrosis. Hepatogastroenterology 1989;36:506-510. 4 NiemelH 0, Risteli J, Blake JE, Risteli L, Compton KV, Orrego H. Markers of fibrogenesis and basement membrane formation in alcoholic liver disease. Relation to severity, presence of hepatitis, and alcohol intake. Gastroenterology 1990;98:1612-1619. 5 Annoni G, Colombo M, Cantaluppi MC, Khlat B, Lampertico P, Rojkind M. Serum type III procollagen peptide and laminin (Lam-Pl) detect alcoholic hepatitis in chronic alcohol abusers. Hepatology 1989;9:693-697. 6 Smedsrd B, Paulsson M, Johansson S. Uptake and degradation in vivo and in vitro of laminin and nidogen by rat liver cells. Biochem J 1989;261:3742. 7 Feizi T. Serum immunoglobulins in chronic liver disease. Gut 1968;9: 193-198. 8 Maddrey WC. Chronic hepatitis. In: Zakim D, Boyer TD, editors. Hepatology. A Textbook of Liver Disease. 2nd edn. Philadelphia: WB Saunders, 1990:1025-1061. 9 Nouri-Aria KT, Hegarty JE, Alexander GJM, Eddleston ALWF, Williams R. Effect of corticosteroids on suppressor-cell activity in ‘autoimmune’ and viral chronic active hepatitis. N Engl J Med 1982;307:1301-1304.
52
HEPCOM 00058
Preliminary note
Serum laminin in patients with chronic active hepatitis and mild liver diseases Julio Collazos and Fernando Diaz Servicio de Medicina Interna, Hospital de Galdhcano, Vizcaya, Spain
(Received 29 July 1993; accepted 3 September 1993)
Laminin is a component of the basement membranes synthesized by Ito cells. We have measured the serum levels of laminin in 15 chronic active hepatitis patients and in 29 patients with mild liver diseases, who also underwent a complete clinical and laboratory evaluation. Abnormal levels of laminin were observed in 13.3% of chronic active hepatitis and in 10.3%of mild liver disease patients. Patients with chronic active hepatitis had higher serum levels of laminin than patients with mild liver diseases (P = 0.016). However, the usefulness of laminin to differentiate between these two groups was limited. Highly significant correlations were found between laminin and total protein (r = 0.79, P < O.OOOl),y-globulins (r = 0.78, P < O.OOOl),and IgG (r = 0.73, P = 0.0001) in the chronic active hepatitis group and lower in the mild liver diseases group. A possible role of defective immunoregulation in the increase of laminin is suggested. Key words: Basement membrane; Chronic liver disease; Liver test; Immunoregulation
Laminin is a component of the basement membranes synthesized by Ito cells which are located in the space of Disse [l]. Serum laminin has been related to portal venous pressure [2], fibrosing activity of the liver [3,4] and alcohol abuse [4,5]. However, some not well known factors have an important influence on laminin serum levels [6]. We have not found any study reported in the literature especially devoted to the evaluation of laminin in patients with chronic active hepatitis (CAH) and mild liver diseases (MLD). We have measured the serum levels of laminin in a group of patients with untreated CAH and MLD, who also underwent a thorough clinical and biochemical evaluation, in order to elucidate the possible usefulness of laminin to differentiate between these two groups.
Correspondence to: Dr. J. Collazos, Servicio de Medicina Interna,
Vizcaya, Spain.
Hospital de GaldBcano, 48960
SERUM LAMININ IN CHRONIC HEPATITIS
53
Materials and Methods Forty-four consecutive nonbiopsied patients that were electively admitted for liver biopsy because of chronic alterations of liver tests persisting for more than 6 months and that were diagnosed as having CAH or MLD after biopsy, were included in the study. Fifteen patients had CAH and the remaining 29 patients had MLD. Of the latter patients, 15 had chronic persistent hepatitis, five had steatosis and nine had minimal changes in liver biopsy. Twenty-seven patients were males (eight CAH and 19 MLD) and 17 females (seven CAH and ten MLD). The ages ranged between 16 and 76 years (mean 40.8 years) with a similar distribution between the two groups. Informed consent was obtained from each patient. Also, the study protocol, previously approved by the Ethical Committee of our hospital, conformed to the ethical guidelines of the Declaration of Helsinki. Blood was collected simultaneously for all the laboratory determinations. Laminin was measured by a radioimmunoassay (Hoechst-Behring, Frankfurt). The upper limit of normal was established at 1.8 U/ml which corresponded to the 95 percentile of a control group composed by 8 1 HBsAg-negative healthy individuals with ages ranging between 18 and 64 years (mean 40.9). Statistical analysis A preliminary test revealed that the distribution of laminin was not Gaussian. Accordingly, nonparametric tests were used. The Spearman’s rank correlation coefficient was used for correlations between continuous variables, the Mann-Whitney U test to compare two groups, and the Kruskal-Wallis test to compare more than two groups. A P < 0.05 level for a two-sided test was considered statistically significant.
Results Five patients (11.4%) had moderately increased serum levels of laminin. Table 1 shows the abnormal values of laminin according to different cut-off levels in CAH and MLD. Age or sex did not significantly influence the laminin levels in our patients. Patients with CAH had higher laminin serum levels than those with MLD (P = 0.016).
TABLE 1 Serum levels of laminin at different cut-offs Total
CAH MLD All
Laminin (U/ml) >l.O
>1.2
>1.4
>1.8
(n)
n (%)
n (o/o)
n (%)
n (%)
15 29 44
11 (73.3) 10 (34.5) 21 (47.7)
6 (40.0) 5 (17.2) 11 (25.0)
5 (33.3) 4 (13.8) 9 (20.5)
2 (13.3) 3 (10.3) 5 (11.4)
CAH denotes chronic active hepatitis, and MLD, mild liver diseases.
Range
Median
0.6-3.0 0.5-2.3 0.5-3.0
1.2 1.0
1.05
54
J. COLLAZOS
AND F. DIAZ
We did not find any significant correlation between laminin and hematocrit, mean corpuscular volume, prothrombin activity, AST, ALT, alkaline phosphatase, GGT, LDH, bilirubin, glycocholic acid, albumin, IgA or IgM in either the CAH or the MLD patients. On the contrary, it was found with total protein (CAH r = 0.79, P < 0.0001; MLD not significant), y-globulins (CAH r = 0.78, P < 0.0001; MLD r = 0.37, P = 0.04) and IgG (CAH r = 0.73, P = 0.0001; MLD r = 0.43, P = 0.01). No significant association was found between laminin and etiology, viral markers, smoking or alcoholic habit, cholelithiasis, hepato- or splenomegaly, drug addiction, urinary bilirubin or urobilinogen. Patients with cytolysis, as measured by aminotransferases, did not have significantly higher laminin levels than those without cytolysis, and the same was true for patients with or without cholestasis. The usefulness of laminin to predict CAH as compared with MLD was evaluated at the same cut-offs used in Table 1. Sensitivity was 73.3,40, 33.3 and 13.3%, and specificity was 65.5, 82.8, 86.2 and 89.7%, respectively, with efficiency of 68.2% for all cut-offs. The receiver operating characteristic (ROC) curve showed that the most discriminant value of laminin to differentiate CAH from MLD was 1 U/ml.
Discussion In our series, patients with CAH had significantly higher serum levels of laminin than patients with MLD. However, although the specificity was high using 1.8 U/ml as a cut-off, the sensitivity was too low to be clinically useful. Schneider et al. [3] have also observed higher levels of laminin in CAH and in active cirrhosis than in other chronic liver diseases. The mechanism whereby laminin increases in liver diseases appears to be multifactorial including increased synthesis by Ito cells [l] and poor metabolization due to liver disease [6]. Although alcohol intake has been related to abnormal levels of laminin [4,5] we failed to find any association with alcohol intake or with biochemical alcohol markers. Also, neither cytolysis nor cholestasis appeared to increase the levels of laminin in our patients. Noteworthy is the highly significant correlation found by us with total protein, y-globulins and IgG, especially in the CAH patients, with correlation’s coefficients close to 0.8. We have not found such association to be reported. IgG increases more frequently than other immunoglobulins in CAH patients [7,8]. On the other hand, a disordered immunoregulation with loss of suppressor-cell activity has been found in CAH patients [9] that could be the cause of this increase. It would be interesting to speculate about the role that this defective immunoregulation could have in the release of laminin from Ito cells. However, further studies are necessary to clarify this association. We conclude that although CAH patients have higher laminin serum levels than MLD patients, the usefulness of laminin to differentiate between these two entities is limited. An important correlation appears to exist between laminin levels and IgG, especially in CAH patients.
SERUM LAMININ IN CHRONIC HEPATITIS
55
References 1 Maher JJ, Friedman SL, Roll FJ, Bissell DM. Immunolocalization of laminin in normal rat liver and biosynthesis of laminin by hepatic lipocytes in primary culture. Gastroenterology 1988;94:1053-1062. 2 Gressner AM, Tittor W. Serum laminin - its concentration increases with portal hypertension in cirrhotic liver disease. Klin Wochenschr 1986;64:1240-1248. 3 Schneider M, Voss B, Hogeman B, Eberhardt G, Gerlach U. Evaluation of serum laminin P-l, procollagenpeptides, and N-acetyl-beta-glucosaminidase for monitoring the activity of liver fibrosis. Hepatogastroenterology 1989;36:506-510. 4 NiemelH 0, Risteli J, Blake JE, Risteli L, Compton KV, Orrego H. Markers of fibrogenesis and basement membrane formation in alcoholic liver disease. Relation to severity, presence of hepatitis, and alcohol intake. Gastroenterology 1990;98:1612-1619. 5 Annoni G, Colombo M, Cantaluppi MC, Khlat B, Lampertico P, Rojkind M. Serum type III procollagen peptide and laminin (Lam-Pl) detect alcoholic hepatitis in chronic alcohol abusers. Hepatology 1989;9:693-697. 6 Smedsrd B, Paulsson M, Johansson S. Uptake and degradation in vivo and in vitro of laminin and nidogen by rat liver cells. Biochem J 1989;261:3742. 7 Feizi T. Serum immunoglobulins in chronic liver disease. Gut 1968;9: 193-198. 8 Maddrey WC. Chronic hepatitis. In: Zakim D, Boyer TD, editors. Hepatology. A Textbook of Liver Disease. 2nd edn. Philadelphia: WB Saunders, 1990:1025-1061. 9 Nouri-Aria KT, Hegarty JE, Alexander GJM, Eddleston ALWF, Williams R. Effect of corticosteroids on suppressor-cell activity in ‘autoimmune’ and viral chronic active hepatitis. N Engl J Med 1982;307:1301-1304.