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J M Neuberger

Publications and source records attributed to J M Neuberger.

At least 73 records · Page 4Linked to original sources
PubMed1991

A 51Cr release cytotoxicity assay for use with human intrahepatic biliary epithelial cells.

Human intrahepatic biliary epithelial cells are important immune targets in a variety of hepatobiliary diseases particularly primary biliary cirrhosis and primary sclerosing cholangitis. The ability to isolate and maintain these cells in short term primary tissue culture has permitted us to develop an in vitro cytotoxicity assay for the study of these cells as potential targets to a variety of toxic stimuli. We have therefore established a chromium-51 (51Cr) release cytotoxicity assay for use with primary cultures of human intrahepatic biliary epithelial cells. The method is simple, reproducible and is more sensitive than dye exclusion, light microscopy and release of lactate dehydrogenase, aspartate aminotransferase and alanine aminotransferase.

Alanine Transaminase↗
PubMed1991

The effect of bile salts on human vascular endothelial cells.

The uptake and release of radiochromium from adult human vascular endothelial cells in culture was employed to determine the relative toxicity of different bile salts. Endothelial cells after pre-incubation with 51Cr for 18 h were incubated with bile salts for 24 h and percentage chromium release was taken as a measure of toxicity to cells. Lithocholic acid (LC) (potassium salt) was cytotoxic at concentrations greater than 50 microM. However, LC glucuronide, sulfate and the beta-epimer were progressively less toxic with toxicity seen at concentrations of 60, 110 and 180 microM, respectively. The greatest cytotoxic effect was observed with glycolithocholic acid (GLC) (potassium salt) which was toxic at every concentration tested (20-200 microM). Sulfation abolished the toxic effect of GLC. At the concentrations employed for the assay (between 20 and 240 microM) GLC sulfate (disodium salt), taurolithocholic acid sulfate (disodium salt), cholic acid (sodium salt), glycocholic acid (sodium salt), deoxycholic acid (sodium salt) and ursodeoxycholic acid (sodium salt) were not cytotoxic. The 51Cr release cytotoxicity assay was validated with lactate dehydrogenase leakage from endothelial cells with a good correlation (r = 0.87). These data confirm in a human cellular system that LC and its conjugates were the most toxic of the bile salts tested and explains its pathophysiological importance in hepatobiliary disease. It also suggests that biotransformation by either sulfation or beta-epimerisation of bile salts especially of LC, as occurs in patients with intrahepatic or extrahepatic biliary obstruction or severe cholestasis, is hepatoprotective.

Bile Acids and Salts↗
PubMed1991

New monoclonal antibodies reacting with bile ducts: further insights into the pathogenesis of bile ductular proliferation in biliary diseases.

We have produced a range of monoclonal antibodies which stain human intrahepatic bile ducts of different sizes. Amongst 26 monoclonal antibodies produced, five clones reacted specifically with bile ducts of different sizes, of which three have been maintained in culture and their viability following freezing and thawing confirmed. Staining patterns varied between normal adult liver tissue, normal fetal liver tissue and a variety of hepatobiliary diseases. The antibodies provide further evidence of the immunological heterogeneity of the human intrahepatic biliary tree and support the hypothesis that proliferating bile ductules are derived from periseptal hepatocytes. The preparation of the antibodies, their staining reactions in normal adult, normal fetal and a variety of liver diseases are described.

Adult↗
PubMed1991

Role of endoscopic retrograde cholangiopancreatography after orthotopic liver transplantation.

Twelve of 178 (7%) liver transplant patients underwent endoscopic retrograde cholangiopancreatography (ERCP) after transplantation. The indications for ERCP were persistent or late onset cholestasis, recurrent cholangitis, and suspected biliary leaks or strictures. The time between transplantation and ERCP ranged from 44 to 330 days (median 153 days). Biliary complications diagnosed by ERCP included biliary sludge in the form of casts, calculi, or debris (n = 7); bile leaks (n = 2); a biliary stricture (n = 1), and complete biliary obstruction (n = 1). One patient had a normal cholangiogram after transplantation. Biliary sludge was detected by ultrasound before ERCP in only one of six patients. Eight patients underwent endoscopic papillotomy, followed by clearance of biliary sludge in four and dilatation of a biliary stricture in one. Two patients bled after papillotomy but neither required surgical intervention. At a median follow up of 1.2 years (range 0.5-2.8 years), nine patients are well and three have died. ERCP provides both accurate diagnosis of biliary complications after liver transplantation and treatment that obviates the need for additional surgery in selected patients.

Adult↗
PubMed1991

Increased expression of intercellular adhesion molecule 1 on bile ducts in primary biliary cirrhosis and primary sclerosing cholangitis.

It has been suggested that immunological mechanisms involving lymphocyte-mediated damage are important in the characteristic bile-duct damage that occurs in primary biliary cirrhosis and primary sclerosing cholangitis. Because adhesion is necessary for the interaction of lymphocytes with their target structures, we have studied the expression of intercellular adhesion molecule 1, a ligand for the leukocyte adhesion receptor lymphocyte function-associated antigen 1 in the liver of patients with primary biliary cirrhosis and primary sclerosing cholangitis. Strong expression of intercellular adhesion molecule 1 was seen on interlobular bile ducts and proliferating bile ductules in both conditions. In primary biliary cirrhosis, medium-sized ducts, which are spared by the disease, were negative. Minimal bile-duct staining was seen in conditions in which bile-duct damage is not a major feature, such as nonbiliary cirrhosis and acute liver diseases. In patients with cirrhosis from any cause, strong expression of intercellular adhesion molecule 1 was detected on the periseptal hepatocytes adjacent to new connective tissue. The intensity of immunohistochemical staining was recorded using a semiquantitative visual scoring system that was subsequently validated quantitatively by confocal laser scanning microscopy. The expression/induction of intercellular adhesion molecule 1 on bile ducts may be important in the pathogenesis of bile-duct damage in primary biliary cirrhosis and primary sclerosing cholangitis and is further evidence to support an immune pathogenesis in these two conditions. Furthermore, the induction of intercellular adhesion molecule 1 on hepatocytes may be an important factor in the liver-cell damage and fibrosis that occur during the development of cirrhosis.

Bile Ducts↗
PubMed1991

Distribution of dihydrolipoamide acetyltransferase (E2) in the liver and portal lymph nodes of patients with primary biliary cirrhosis: an immunohistochemical study.

The reason for the close association between primary biliary cirrhosis and the appearance of antibodies that recognize the E2 component of pyruvate dehydrogenase complex is not understood. The distribution of the three pyruvate dehydrogenase complex subunits was examined in the liver and lymph nodes of patients with primary biliary cirrhosis, patients with other liver diseases and normal subjects by immunohistochemistry using affinity-purified antibodies. Intensity of staining was assessed semiquantitatively and validated by scanning laser confocal microscopy. In primary biliary cirrhosis tissue, the E2 staining pattern did not parallel the reported distribution of mitochondria. E2 staining in biliary epithelial cells was consistently stronger than in hepatocytes. In primary biliary cirrhotic liver, staining of biliary epithelium was significantly stronger than in normal or other liver disease controls; many bile ducts in primary biliary cirrhotic liver demonstrated very high intensity, diffuse distribution of stain. No differences in staining intensity were seen between perivenular hepatocytes in primary biliary cirrhotic liver and those in controls; periportal hepatocytes in primary biliary cirrhotic liver were, however, more intensely stained than perivenular cells. In primary biliary cirrhotic portal lymph nodes, a subset of macrophages showed high-intensity, diffuse distribution of stain. By contrast, staining with antibodies to E1 and E3 (other components of pyruvate dehydrogenase complex) produced uniform-intensity, mitochondrial distribution both in primary biliary cirrhosis and control tissue. The increased intensity of E2 in primary biliary cirrhotic tissue could be explained in terms of abnormal metabolism of E2 by biliary epithelial cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetyltransferases↗
PubMed1991

Growth of normal human hepatocytes in primary culture: effect of hormones and growth factors on DNA synthesis.

Although the ability of hormones and growth factors to stimulate DNA synthesis in rat hepatocytes has been investigated extensively, no such study of human hepatocytes has been reported. Here we describe a series of experiments to identify those factors that regulate human hepatocyte DNA synthesis in vitro and which therefore may play a role in the control of human liver regeneration. Human hepatocytes were isolated from normal liver tissue obtained after graft reduction for transplantation into pediatric recipients. Cells were maintained in culture for up to 5 days, and DNA synthesis was determined. Hydroxyurea reduced [3H]thymidine incorporation into DNA by 95%, indicating replicative DNA synthesis. As previously found with rat hepatocytes, epidermal growth factor and transforming growth factor-alpha stimulated DNA synthesis at low nanomolar concentrations; transforming growth factor-alpha was slightly more potent. Although the overall rate of thymidine incorporation was lower than that for rodent cells, human hepatocytes were sensitive to lower concentrations of these growth factors, and the degree of stimulation was similar. Conversely, transforming growth factor-beta inhibited DNA synthesis at low picomolar levels. By contrast (unlike rat hepatocytes), arginine-vasopressin failed to initiate or potentiate DNA synthesis in human cells. These data indicate that normal human hepatocytes can respond to low concentrations of growth promoters or inhibitors, previously shown to have activity on rat hepatocytes. These factors may then play a role in control of human liver growth. However, important species differences are apparent, highlighting the limitations of extrapolating from animal studies to humans.

Animals↗
PubMed1990

Primary hepatic malignancy: the role of liver transplantation.

Between January 1982 and April 1989, 134 patients with a suspected liver neoplasm were referred to the liver unit, Queen Elizabeth Hospital, Birmingham. In 105 (78 per cent), a primary hepatic neoplasm was histologically confirmed, and 47 patients (45 per cent) proved to have primary hepatocellular carcinoma. Twenty-nine orthotopic liver transplants were performed in 28 of these patients (27 per cent). Twenty patients (71 per cent) survived 30 days or longer (median 11.5 months; range 2-87 months), of whom nine are currently alive. We retrospectively analysed our data to determine the influence of preoperative evaluation, histological type and staging on outcome. Computed tomography proved to be superior to intraoperative assessment (86 versus 58 per cent) in diagnosing tumour positive nodes. Patients with tumour negative lymphadenopathy had a better prognosis. Postoperative stage I/II had a median survival of 16 months (range 3-87 months) compared with 7.5 months (range 2-20 months) for stage III. Non-cirrhotic patients with hepatocellular carcinoma had the best prognosis; cholangiocellular carcinoma and cirrhotic patients with hepatocelluar carcinoma had the worst outcome with no survivors beyond 1 year. Because of the advanced stage of disease at the time of presentation, the value of liver transplantation in primary liver cancer is limited. For those presenting with advanced disease confined to the liver (stage I/II) in whom conventional hepatic resection is not possible, significant benefit can be achieved in selected cases.

Adenoma, Bile Duct↗
PubMed1990

Biliary epithelial cells from the liver of patients with primary biliary cirrhosis: isolation, characterization, and short-term culture.

Since biliary epithelial cells of the middle-sized interlobular bile ducts are targets for lymphocyte-mediated damage in patients with primary biliary cirrhosis (PBC), we have developed a method for isolating and maintaining these cells in short-term tissue culture. Intrahepatic biliary epithelial cells were isolated from small segments of liver removed at the time of transplantation. Cells were separated from a collagenase digest by immunomagnetic separation using Dynabeads coupled to a monoclonal antibody (HEA 125) specific for a biliary epithelial cell surface antigen. The yield was approximately 3 x 10(5) cells/g of liver. The isolated cells were characterized morphologically and ultrastructurally using light and electron microscopy, and immunocytochemically using HEA 125 and anti-cytokeratin, anti-vimentin and anti-asialoglycoprotein receptor antibodies. By these criteria cells were judged to be identical to biliary epithelial cells from normal liver. The cells could be maintained in short-term tissue culture for up to 4 weeks without loss of biliary epithelial cell markers. Availability of interlobular biliary epithelial cells will be of value in future investigations of the pathogenetic mechanisms of PBC.

Bile Ducts, Intrahepatic↗
PubMed1990

Symptom development and prognosis in primary biliary cirrhosis: a study in two centers.

The prognosis of patients with asymptomatic primary biliary cirrhosis has been uncertain. Cases of primary biliary cirrhosis in 95 patients from two centers are presented: 70 patients from a regional referral center (Freeman Hospital, FH) and 25 from an international tertiary referral center (King's College Hospital, KCH) with similar mean age at diagnosis and duration of follow-up (median, 12 months). During follow-up, 19 of 70 FH patients and 15 of 25 KCH patients developed symptoms (P less than 0.001); the mean time to symptom appearance was 43 months at FH and 35 months at KCH (P = 0.0012). Twenty-five of 95 patients died, 15 of 34 subjects who had developed symptoms (all liver-related deaths) and 10 of 61 who remained asymptomatic (all non-liver-related deaths) (P less than 0.001). Hepatic mortality was worse than among the normal population (P less than 0.05). A comparison of those patients developing symptoms with 111 at FH and 165 at KCH who presented with symptoms suggests no difference in survival once hepatic symptoms appear. In both centers, hepatic mortality was strongly related to symptom appearance in asymptomatic primary biliary cirrhosis: patients developing symptoms resemble those with symptoms at presentation. Other differences between the centers may reflect different referral patterns.

Adult↗
PubMed1990

Neutrophil activation--an important cause of tissue damage during liver allograft rejection?

Although neutrophils have been implicated as mediators of tissue damage in a variety of conditions, their role in graft rejection has not previously been studied. Peripheral neutrophil activation was assessed sequentially in 16 patients following liver transplantation by measuring the ability of neutrophils to respond to the chemotactic peptide FMLP, to release superoxide radicals, and to cause extracellular proteolysis. All three functions increased 1-2 days before episodes of acute rejection and only returned to normal after treatment with high-dose corticosteroids. Three patients who did not develop rejection showed no increase in neutrophil function. Lymphocytes isolated from patients with acute rejection produced factors that were both chemotactic for and capable of activating normal neutrophils. This study shows that neutrophil activation occurs during acute liver allograft rejection, possibly in response to lymphokine secretion, and suggests an important, and hitherto unrecognized, mechanism of graft damage during rejection.

Adult↗
PubMed1990

Inhibition of leukocyte chemotaxis by immunosuppressive agents. Specific inhibition of lymphocyte chemotaxis by cyclosporine.

Since chemotaxis is an important mechanism by which leukocytes are recruited to allografts during rejection, the effect of immunosuppressant agents on in vitro chemotactic responses of neutrophils, monocytes, and lymphocytes was studied. Cyclosporine caused profound inhibition of lymphocyte chemotaxis to three different chemotactic factors at pharmacologic levels but had no effect on neutrophil or monocyte chemotaxis. Methylprednisolone and azathioprine both inhibited chemotactic responses of neutrophils and monocytes as well as lymphocytes. These data suggest that immunosuppressant agents are potent inhibitors of leukocyte chemotaxis and that drugs differ as to which leukocyte cell type they inhibit. The selective action of cyclosporine might explain why this drug is ineffective in reversing established rejection episodes in which monocytes and neutrophils are also involved. Inhibition of leukocyte chemotaxis may be an important mode of action of immunosuppressant drugs, and drug regimes might be adjusted to cause selective inhibition of particular cell types.

Azathioprine↗
PubMed1990

Referral of patients with primary biliary cirrhosis for liver transplantation.

All patients with primary biliary cirrhosis referred to this unit for consideration for transplantation between April 1981 and January 1989 were analysed retrospectively to assess whether disease stage at referral affects the outcome after grafting and whether greater awareness of the benefits of the procedure means that patients are now being referred at an earlier stage. Seventy of the 107 patients have been grafted, with an overall one year actuarial survival of 62%. A better prognosis at the time of referral, as assessed by both serum bilirubin concentration and a mathematically derived prognostic index, was associated with a greater probability of survival after grafting. Patients in the tertile with the best prognosis (median serum bilirubin concentration at referral 84 mumol/l and estimated survival in the absence of transplantation of more than nine months) had a 78% one year actuarial survival after transplantation, whereas those in the tertile with the worst prognosis (median serum bilirubin concentration 467 mumol/l and estimated survival of less than four months) had a one year actuarial survival of only 50%. No trend towards earlier referral of patients, however, was shown using either of these two markers. This retrospective analysis suggests that many patients are being referred too late for an optimal outcome. We recommend that patients with primary biliary cirrhosis who are potential candidates for liver grafting should be referred to a transplant centre before the serum bilirubin concentration approaches 150 mumol/l.

Adult↗