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The pharmacokinetics of oxmetidine (SK&F 92994) were investigated in nine cirrhotic patients and compared with ten control subjects with gastroduodenal ulcers, but without any symptoms of hepatic pathology. On two separate occasions each patient received 200 mg oxmetidine as a single oral dose and 100 mg as a single intravenous dose. In the cirrhotics, the bioavailability of the oral dose and the plasma elimination half-life after both oral and intravenous administration were significantly higher than in the controls. Moreover, a positive correlation was found between the plasma elimination half-lives and the biochemical parameters of cholestasis. Such findings indicate that in severe liver disease and in cholestasis the accumulation of oxmetidine in the circulation may limit the use of this drug.

Polymerized human serum albumin (pHSA) binding activity due to antibodies was evaluated by a sensitive and specific enzyme-linked immunosorbent assay (ELISA) in well characterized cases of acute (AVH) and chronic active (CAH) viral hepatitis. Moreover, hepatitis B virus (HBV) infection serum samples were tested for the presence of HBsAg-associated pHSA receptors detected by radioimmunoassay. Specificity of ELISA in detecting pHSA binding activity due to antibodies was assayed by testing IgG fractions from positive serum samples and purified HBsAg particles. A very high prevalence (mean = 87%) of antibody response to pHSA was detected in serum samples of patients with type B, A and post-transfusion non-A, non-B (NANB) AVH and CAH. IgM antibodies were found in 75% of type B AVH, in 77% of type A AVH and in none of the NANB cases at the onset of illness. In CAH, IgM antibodies were found in 37.5% of HBsAg-positive cases and in 44% of NANB cases. In HBV-related AVH and CAH no correlation was found between anti-pHSA antibodies and HBsAg-associated pHSA receptors or the HBeAg/anti-HBe status. The antibody response detected by ELISA showed a cross-reactivity with monomeric human albumin and heterologous polymeric and monomeric albumins. However, in HBV infection the antibody binding was significantly higher for pHSA than for monomeric human albumin or albumins from other species, thus suggesting that the antigenic sites of pHSA evoking the antibody response may differ in relation to the etiology of viral hepatitis.

In order to characterize the effector mechanism of "in vitro' lymphocytotoxicity against isolated rabbit liver cells in acute viral hepatitis (AVH), the test was performed in 4 patients with type A, in 8 with type, B, and in 2 patients with non-A non-B AVH: (a) using peripheral blood lymphocytes; (b) after enzymatic digestion of the surface membrane immunoglobulins of these lymphocytes, and (c) as in b with addition of patients' sera. 78.5% of the patients had an elevated cytotoxicity which did not change when the lymphocytes were treated as in b and c. Only the 2 patients which progressed to chronicity had an elevated cytotoxicity which disappeared after enzymatic exposure and reappeared with the addition of patients' sera. The results suggest that "in vitro' cytoxicity in recovering AVH is not antibody dependent, but probably of spontaneous type. On the contrary an antibody-dependent cytotoxicity is present in the 2 cases of AVH who progressed to chronicity.

"In vivo" and "in vitro" action of 2 MPG on lymphocytotoxic activity was evaluated in 22 patients with CAH, 14 were HBsAg positive and 8 were negative. The test was performed with and without 2 MPG in the colture medium, and before and after treatment. HBsAg+ patients received 2 MPG treatment, while HBsAg- ones received immunosoppressive therapy and 4 of these were on 2 MPG treatment as well. Cytotoxic Index was reduced non-significantly by addition of 2 MPG in colture medium in both groups. 2 MPG treatment does not modify cytotoxic activity which is reduced by immunosoppressive therapy. These findings suggest a positive effect of 2 MPG on liver-cell metabolism with an increased resistence of the epatocyte to the "in vitro" lymphocytes aggression. Hence it may be suggested an association treatment with immunosoppressive agents and 2 MPG.

Sera from 42 patients with acute viral hepatitis (AVH), 97 patients with chronic active liver disease (CALD) and 89 controls were tested by radioimmunoprecipitation for the presence of antibodies against human liver-specific protein (LSP). Anti-LSP were found in all but one patient with AVH type A (93%) and in a smaller percentage of AVH type B (55%). In non-A, non-B cases, anti-LSP were found in low percentages: 27% in acute cases, 10% in chronic cases. Furthermore, in CALD, a significant difference was found between HBsAg-positive CAH and 'autoimmune' CAH, a significant difference was found between HBsAg-positive CAH and 'autoimmune' CAH, both in anti-LSP prevalence (21%, 67%; P less than 0.005) and in anti-LSP titre (1:154 +/- 170, 1:316 +/- 186; P less than 0.005). In HBsAg-negative/anti-HBc-positive CAH, three of 15 patients were anti-LSP positive. Anti-LSP were found only in three of 57 patients with various non-hepatic diseases with autoimmune features. None of the 12 healthy HBsAg carriers was positive. Hence there is evidence for a considerable heterogeneity in anti-LSP response in acute and in chronic inflammatory HBsAg-negative liver diseases. These data suggest that anti-LSP antibodies do not play a prominent role in the process of transition to chronicity of acute viral hepatitis particularly in non-A, non-B cases, whereas these antibodies may be important in the mechanism of ongoing liver cell injury in patients with 'autoimmune' CAH, and can represent a useful diagnostic marker of this type of hepatitis.

After some general preliminary remarks concerning aetiopathogenetic hypoteses and therapeutic possibilities for Wilson's disease, the Authors report the data obtained from a long-term study carried out on a family of nine brothers. These subjects were all affected with Wilson's syndrome and kept under a D-penicillamine treatment. The addition of 4-5 oral adminstrations a day of 30 mg SAMe resulted in highly significant favourable modifications of all the laboratory data considered to test liver function. The progressive worsening of the same data observed after 60, 90 and 120 days from SAMe withdrawal, seems to prove the actual activity of this molecule on liver function. During SAMe therapy no clinical and laboratory side-effects (macular and papular eruption, pruritus, neutropenia, thrombocytopenia, etc.) were observed while they were detectable in some patients treated with D-penicillamine alone.

The action of an anti-alpha 1-AGP antibody on AMLR, anti-T3 and PHA T-lymphocyte proliferative response was evaluated. We observed a strong dose-dependent inhibition on T-lymphocyte proliferative responsiveness to autologous non-T cells and to anti-T3 stimulus, whereas PHA activation was unaffected. A lower degree of inhibition of the proliferative response was also observed on pretreating both T and non-T cells with the antibody; the addition of anti-alpha 1-AGP in the culture containing cells pretreated with the antibody showed a further inhibition of thymidine incorporation. The data suggest a direct influence of the antibody on membrane alpha 1-AGP and support a positive role of this glycoprotein (distinct from Ia and T3 antigens) on both anti-T3 and autologous non-T cell T-lymphocyte responsiveness, thus indicating the involvement of alpha 1-AGP in the T3-Ti antigen-specific pathway of T-cell activation.

The potential roles of Ca2+ ions in the response of T lymphocytes to stimulation with monoclonal antisera to the T3 antigen were investigated by means of pharmacological agents that predominantly inhibit the flux of Ca2+ ions into cells (verapamil, nifedipine) or the activity of Ca2+-dependent kinases (trifluoperazine, polymyxin B). As assessed by uptake of [3H]thymidine, proliferation induced with anti-T3 +/- recombinant IL-2 at 72 h was inhibited by greater than 80% in the presence of nifedipine at 50 microM, and almost completely arrested (greater than 95% inhibition) with the other agents at the same concentration. Further quantitative assays of the effects of polymyxin B and trifluoperazine on C-kinase labelling of exogenous substrate showed a major reduction with both agents, but inhibition was substantially greater with polymyxin B that with trifluoperazine (IC50 = 14 and 70 microM respectively). These results were confirmed by qualitative assessment of Ca2+/phospholipid-dependent phosphorylation of endogenous substrates, which demonstrated major phosphoproteins of MW 56,000, 52,000, 43,000, and 20,000, and dose-dependent reduction in labelling in the presence of polymyxin B. Similar results were obtained under more physiological conditions in intact cells labelled with 32P orthophosphate. These findings indicate several possible roles for Ca2+ in T-cell activation, and several possible levels of activity, including modulation of calmodulin-dependent kinases and effects on Ca2+/phospholipid-dependent kinases and Ca2+ channels.

Soluble alpha 1 acid-glycoprotein is considered an "acute phase protein" with an inhibitory effect on lymphocyte activity; it has recently been shown that a lymphocyte modulatory variant of alpha 1 acid-glycoprotein has a positive role on T cell activation. It is not clear whether the presence of this glycoprotein on lymphocyte membranes is due to an endogenous production or to a passive uptake of soluble alpha 1 acid-glycoprotein by its carbohydrate moiety. Our data show an increase of membrane alpha 1 acid-glycoprotein both in peripheral blood lymphocyte and T-enriched lymphocytes after phytohemagglutinin stimulation. Peripheral blood lymphocyte enzymatic treatment by neuraminidase does not affect alpha 1 acid-glycoprotein expression while pronase digestion induces a strong decrease of alpha 1 acid-glycoprotein positive lymphocytes and a resynthesis after phytohemagglutinin stimulation. Furthermore, the presence of alpha 1 acid-glycoprotein was prevalently, found on helper/inducer lymphocytes. These data support the hypothesis of a synthesis of alpha 1 acid-glycoprotein by T lymphocytes during their activation process.

The relative distribution of T lymphocyte subsets, as defined by the monoclonal antibodies OKT, was determined by cytofluorimetric analysis in peripheral blood and in cells isolated from liver biopsies of 31 patients with chronic active hepatitis (CAH). The percentage of peripheral blood lymphocytes binding OKT8 (directed against cytotoxic/suppressor T cells) was found to be elevated in patients with HBsAg and HBeAg positive chronic active hepatitis. Patients with CAH who had seroconverted to anti-HBe, had an increased number of OKT3-positive cells in their blood, which was directed against a common T cell surface antigen, associated with a decreased number of OKT8 positive cells. Lymphocytes isolated from liver biopsies of patients with CAH presented a general increase of OKT8-positive cells associated with a decreased number of OKT4-positive (helper/inducer) T cells. It is likely that OKT8-positive cells found in liver biopsies represent cytotoxic T cells directed against either viral or liver cell determinants.

Reaction of the T cell membrane with monoclonal antibodies to T3 can initiate cellular activation, and this is associated with increased intracellular Ca2+ and inositol-trisphosphate (IP3) release. We therefore studied the possible involvement of Ca2+/phospholipid-dependent kinase (C-kinase) in these phenomena. Quantitative assays of exogenous substrate phosphorylation in unstimulated cells showed Ca2+/phospholipid-dependent kinase activity in the cytosol, but no comparable activity in the particulate fractions corresponding to membrane and cytoskeleton material. At concentrations of soluble anti-T3 that partially activate T cells in the absence of macrophages, there was a 50 to 60% decrease in C-kinase activity in the cytosol, with a comparable increase in activity in the membrane fraction. A similar transfer of activity was also induced with the known C-kinase activator, 12-O-tetradecanoyl-phorbol-13-acetate, although redistribution was more rapid in onset, more complete, and more sustained. Redistribution of enzyme activity was additionally confirmed by qualitative assays of endogenous substrate phosphorylation. Labeling of intact cells followed by immunoprecipitation analysis with anti-T3 indicated signal-dependent phosphorylation of two components of the T3 complex and an unidentified 94,000 substrate that was resistant to reduction and alkylation. These findings are consistent with an important role for C-kinase in transduction of membrane events by the T3-Ti complex.

Antibodies directed against glutaraldehyde-polymerized human serum albumin (pHSA) were tested by a sensitive and specific enzyme-linked immunosorbent assay (ELISA) in serum samples from 196 patients with various hepatic and non-hepatic diseases and in 38 healthy control subjects. A very high prevalence (80.1%) of antibody response was found in the patient group, ranging from 60% in type non-A, non-B (NANB) chronic active hepatitis (CAH) and lymphomas to 95.5% in hemodialysis patients. A higher prevalence of anti-pHSA antibodies was found in hepatitis B virus (HBV)-related CAH and hemodialysis patients compared with NANB-related CAH, collagen diseases and lymphomas. Anti-pHSA antibodies were most frequent in HBsAg-positive cases, but no correlation between antibody response and the HBeAg/anti-HBe status was found among these patients. Anti-pHSA antibodies did not correlate with HBsAg-associated pHSA receptors determined by radioimmunoassay (RIA). Moreover, a significantly higher pHSA receptor expression was found in HBV-related CAH and hemodialysis patients compared with chronic HBsAg carriers and in HBeAg-positive patients compared with HBeAg-negative cases. The anti-pHSA response seems to be related to the presence of immunoregulation disorders which may be induced by several causes, such as autoimmunity and viral infections. In particular, as far as the HBV infection is concerned, there is no evidence that circulating anti-pHSA antibodies interfere with the natural course of the disease.

Six oral administrations per day of 30 mg S-adenosylmethionine (SAMe) for 30 days, in addition to 6000 gamma/day of Vitamine B12 induced marked improvements of biochemical parameters in 20 patients with hepatic cirrhosis or various chronic hepatites. Particularly, the protidemia, bilirubinemia and radial immunodiffusion have shown the highest favorable drug responses. These improvements were still lasting and even further increasing 30 days after the end of therapy. In another group of patients with similar diagnosis and under clinical conditions comparable to the previous group of twenty, the administration of Vitamine B12 alone, in the same doses as above, has not induced any alteration in the biochemical parameters.

Autologous mixed lymphocyte reaction (AMLR) and phenotypical composition of circulating lymphocytes obtained from nine subjects with untreated hepatocellular carcinoma (HCC); three HCC patients locally treated by means of intraarterial chemoembolization; six subjects with gut-derived carcinoma (GDC); nine chronic active liver disease patients (CALD), and 14 normal controls have been evaluated, using monoclonal antibodies (MAB) against CD5, CD8, CD4 glycoproteins and anti-LAK-1 molecule, a novel 120-KD surface antigen that is present on the membrane of large granular lymphocytes, by means of classical indirect immunofluorescence technique. Autologous mixed lymphocyte reaction was significantly reduced in all patients, along with CD4-positive cells that are the responder cells in this reaction. A relative increase in the percentage of LAK cells was also observed in neoplastic patients with a normalization after local treatment of the HCC. In the light of promising results obtained by Rosenberg et al. by adoptive transfer of LAK cells activated with Interleukin-2 (IL2) in various cancers, our results support a similar therapeutic trial in HCC patients.

We investigated inhibitory effect of various monoclonal antibodies on T-cell-mediated cytotoxicity against autologous hepatocytes in 24 patients with hepatitis B surface antigen/hepatitis B e antigen (HBsAg/HBeAg)-positive chronic active hepatitis. A significant reduction of cytotoxicity index occurred after preincubation of T lymphocytes with anti-Leu 7 (killer-natural killer cells), D1/12 (Ia-positive cells), 5/9 (restricted helper/inducer cells), and MLR4 ("activated" and radiosensitive helper cells) monoclonal antibodies (MAb). Anti-Leu 2a (cytotoxic/suppressor cells) and anti-Leu 3a (helper/inducer cells) MAb did not affect cytotoxic activity. This finding supports the hypothesis that the T cytotoxic reaction in this in vitro system is probably due to two mechanisms: first, spontaneous cell membrane cytotoxicity sustained by anti-Leu-7-positive lymphocytes; and second, specific cytotoxicity mediated by activated Ia-positive cells. We also found that the presence of helper/inducer cells (5/9 positive) appears to be a prerequisite for the cytotoxic reaction.

We studied the expression of major histocompatibility complex (MHC) class II antigens in liver biopsies taken from ten patients with clinical and biochemical evidence of liver damage after bone marrow transplantation. In all six patients who had histologically confirmed graft-versus-host disease, MHC class II antigens were detected on intrahepatic bile ducts. In four patients with no histological features of graft-versus-host disease, MHC class II antigens were not detected. In controls, a positive reaction for bile duct MHC class II antigens was only detected in the patients with primary biliary cirrhosis. Characterisation of the lymphocytes surrounding the bile ducts showed a prevalence of Leu 3+ cells in graft-versus-host disease and primary biliary cirrhosis. We propose that the aberrant expression of class II antigens on bile duct epithelium cells may play a role in the pathogenesis of graft-versus-host disease. A similar pattern in primary biliary cirrhosis may suggest a common pathogenetic mechanism.

In order to determine the number of circulating B lymphocytes which can synthetize their surface membrane immunoglobulins, we have evaluated by immunofluorescence their presence before and after enzymatic exposure to pronase. After pronase treatment the majority of lymphocytes carrying SmIg has been disappeared (from 7.8% to 1.9%; p < .0.0005). After 18 hours incubaton only a small proportion (4.6%) of B lymphocytes risynthetized their SmIg. This observation support the hypothesis that a subset of lymphocytes present in the peripheral blood carries on its membrane only cytophilic SmIg and therefore may interfere in the determination of the number of B lymphocytes.