Influence of TNF gene polymorphism in patients with acute and fulminant hepatitis

Background Tumor necrosis factor (TNF) is involved in liver damage, especially in fulminant hepatitis (FH). Our previous data showed that the serum level of TNF- was markedly increased in FH. To investigate the mechanism of the overproduction of TNF in FH patients, polymorphism of the TNF gene was studied.Methods We analyzed 120 healthy subjects (controls), 63 patients with acute hepatitis (AH), and 32 patients with FH. Of the 32 FH patients, 21 died or received liver transplantation (FH-D), and 11 survived with intensive therapy (FH-S). The TNF- promoter region at –1031, –863, –857, –308, and –238, and TNF- Nco1 polymorphism sites were studied.Results (1) The four groups showed no differences in polymorphisms of positions –857, –308, and –238. The allelic frequencies of positions –1031C and –863A in the FH-D patients were significantly higher compared to findings in control subjects. (2) The allelic frequency of B2 in the TNF- gene was significantly higher in FH patients, and particularly in the FH-D patients, compared to control subjects. (3) When the patients were divided into four groups by etiology, hepatitis A virus (HAV), HBV, HCV, and non-A non-B non-C, the allelic frequencies of positions –863A and TNF- B2 in FH patients were increased in the non-A non-B non-C group compared to controls.Conclusions FH patients with a poor prognosis had higher frequencies of positions –1031C and –863A in the TNF- promoter region, and higher frequencies of the B2 allele of the TNF- gene. These data suggest that the genomic background may be associated with the prognosis of acute liver failure.     

Family study and frequency of blood group with strong B transferase accompanied by decreased A and H antigens

Summary Subgroups of type A blood, named A₁, A₂ and A₁–A₂ intermediate (A_int), are specifically characterized by their peculiar A alleles and have their own A₁-, A₂- or A_int-forms of -N-acetyl-D-galactosaminyltransferase (A-transferase). It is known, however, that certain type A₂B persons exhibit A₁-transferase. The reason may be an unusual -galactosyltransferase (B-transferase). This strong B-transferase competes with A-transferase for the substrate, H antigen, so as to decrease the A and H antigens on the red cells. We studied this blood group over three generations and found that the strong B-transferase is, in fact, inherited with the B gene and is dominant over normal B-transferase. In AB blood groups in Tokyo, the frequency of people with a strong B-transferase is 5% for A₁B and 22% for A₂B. This enzyme does not always cause weak H or A antigens.     

Gabexate mesilate, a synthetic protease inhibitor, attenuates carbon tetrachloride-induced liver injury in rats

Background Gabexate mesilate, a synthetic protease inhibitor, is used to treat acute pancreatitis and disseminated intravascular coagulation because it inhibits various serine proteases; however, whether gabexate mesilate prevents acute liver failure has not yet been studied. The aim of the present study was to investigate the effect of gabexate mesilate in carbon tetrachloride (CCl₄)-induced liver injury in rats.Methods Acute hepatic failure was induced by administration of CCl₄ intragastrically to male Sprague–Dawley rats. The effects of gabexate mesilate were examined in terms of serum transaminase levels, liver histology, and the prognosis of rats.Results Gabexate mesilate treatment significantly decreased the elevation of serum transaminase levels and improved liver histology 24h after the administration of CCl₄ (0.2ml/100g rat weight). Plasma tumor necrosis factor- (TNF-) and interleukin-1 (IL-1) decreased significantly in the gabexate mesilate-treated rats compared with saline-treated rats. Gabexate mesilate treatment also significantly improved survival rate after a lethal dose of CCl₄ (0.5ml/100g rat weight) from 0% to 20%.Conclusions Gabexate mesilate treatment attenuated CCl₄-induced liver injury via a suppression of proinflammatory cytokine production. In addition, these investigations suggest that gabexate mesilate treatment may provide therapeutic strategies for human acute liver failure.     

Acute hematologic effects of interferon alpha,interferon gamma,tumor necrosis factor alpha and Interleukin 2

Summary This study was designed to investigate acute effects of various doses of the cytokines IFN-alpha, IFN-gamma Interleukin 2 and tumor necrosis factor alpha on white blood cell differential counts. Before initiation of phase II trials, a dose-determination phase was performed, where three different dose levels of each cytokine were applied as a single dose. White blood cell differential counts were assessed immediately before and 2, 12, 24, 48 and 168 h after injection. Patients enrolled suffered from metastatic cancer or chronic active hepatitis. In addition, IFN-alpha was administered to five healthy volunteers. Results indicate that cytokines cause rapid and transient changes in the numbers of leukocyte subsets. Hematologic changes were cell-type- and cytokine-specific: transient lymphopenia was observed after administration of all four cytokines, reaching a nadir 12 to 24 h after subcutaneous injection. Administration of TNF-alpha and IFN-gamma also caused transient monocytopenia. Neutrophilia developed after administration of Interleukin 2, IFN-alpha and TNF-alpha. We conclude that cytokines play a key role in the regulation of peripheral blood cell traffic by their capacity to influence homing patterns of peripheral blood leukocytes.     

Regression of Hepatic Fibrosis in Hepatitis C with Long-Term Interferon Treatment

Cirrhosis occurs in 20-50% of patients with hepatitis C and is thought to be irreversible. We describe two patients with cirrhosis secondary to hepatitis C in whom the extensive fibrosis and cirrhosis appeared to regress in response to treatment with interferon- (IFN-). Both patients were in the early stages of cirrhosis, class A in the Child-Pugh classification, total score 5 for each patient. Both responded fully to IFN- and had normalization of all liver function tests and disappearance of hepatitis C viral RNA. Liver biopsies, performed before and after treatment, were coded unpaired by patient, combined with 21 liver biopsies from eight other patients with chronic hepatitis, and read independently by two pathologists using the Knodell scoring system. Knodell scores decreased from 14 to 3.5 and from 13.5 to 4 in these two patients. Cirrhosis and extensive fibrosis, present at baseline, were not present on follow-up liver biopsies, which were of sufficient size to reduce the likelihood of sampling error. We conclude that hepatic fibrosis and clinically early cirrhosis may be reversible in some patients with hepatitis C who respond to treatment with IFN-.     

Presence of interferon and anti-interferon in patients with systemic lupus erythematosus

Summary Sera from 61 patients with systemic lupus erythematosus (SLE) were serially screened over a period of at least 2 years for IFN and anti-IFN antibodies. IFN concentrations were measured both with a cytopathic effect assay and a more sensitive radioimmunoassay. Of the patients 15% (9/61) had IFN in their serum at one or more occasions as measured in the bioassay (6 IU/ml); employing a RIA (1 IU/ml) 28% (17/61) of the patients studied were positive for IFN-. Fifteen patients had a measurable interferonemia over 2–16 months; only two patients had detectable IFN in their serum at only one occasion. In five patients, hourly and daily variations of the IFN titer as measured by RIA were found to amount to less than 80%. The IFN activity found in these sera was characterized as IFN- by means of acid stability, cross-reactivity on heterologous cells, trypsin sensitivity, and neutralization by homologous and heterologous antisera. IFN antibodies were quantified with a neutralization bioassay, an ELISA, and a radioimmunoassay. Of the 61 patients 5% (3) possessed high titers of anti-IFN antibodies which persisted over 2 years. The IFN- antibody positive patients had an inactive form of the disease over years without visceral involvement but decreased serum complement levels (C4, C3, CH50) and repeated episodes of Quincke-like edema.     

Impaired glucose tolerance is accompanied by decreased insulin sensitivity in tissues of mice implanted with cells that overexpress resistin

Aim/hypothesis Resistin, the expression of which is suppressed by thiazolidinedione treatment in adipocytes, is one of the key molecules for the tight link between adiposity and insulin resistance. Here, we show the in vivo effects of resistin on insulin sensitivity in mature mice using a cell implantation method.Methods Resistin cDNA was transfected into 3T3-L1 pre-adipocytes, which were then implanted into subcutaneous areas of nude mice. Metabolic analyses were performed 4 or 6 weeks after implantation.Results The mice implanted with 3T3-L1 cells overexpressing resistin (R-mice) showed significantly (p<0.05) increased plasma resistin levels. After a glucose load plasma insulin levels were significantly greater in R-mice than in mice implanted with mock-transfected cells (M-mice). The AUC of insulin after glucose loading was positively correlated with circulating resistin levels. Significantly decreased glucose responses after insulin injection were observed in R-mice, compared to M-mice. The insulin-induced phosphorylation level of IRS-1 was significantly lower in muscles of R-mice than M-mice. The expression of TNF- mRNA in intra-peritoneal fat tissues was significantly greater in R-mice than in M-mice, but there was no difference between the two groups with regard to subcutaneous fat tissues. The concentration of TNF- in plasma was positively correlated with resistin levels in R-mice.Conclusions/interpretation Resistin, when actually secreted from cells in mature mice, causes disturbed glucose metabolism, possibly based on decreased insulin sensitivity in muscle. The in vivo effects of resistin on insulin sensitivity might be in part mediated by increased TNF- expression in visceral fat tissues.     

Role of zinc in treatment of experimental acute pancreatitis in mice

A study on the effect of zinc feeding on the survival rate as well as the levels of trypsinogen, ₂-macroglobulin, zinc, calcium, and magnesium in the plasma, pancreata, and livers of BALB/c mice fed a choline-deficient diet supplemented with 0.5%dl-ethionine (CDE diet) was undertaken. Feeding them a zinc-excess diet significantly increased the survival rate of mice with pancreatitis induced by CDE diet feeding. Trypsinogen concentrations in plasma and pancreas increased in mice fed a CDE diet and further increased in mice fed a zinc-deficient diet. The plasma ₂-macroglobulin levels in mice fed a zinc-deficient diet decreased compared to those fed a zinc-adequate or a zinc-excess diet. In mice with pancreatitis, zinc and calcium concentrations of pancreata increased and magnesium concentrations decreased compared to those of normal controls. The calcium concentrations in both livers and pancreata increased, but magnesium concentrations in these tissues decreased. These results suggest that altered mineral metabolism in the pancreas may have contributed to the pathophysiology of the mice with acute pancreatitis and that zinc supplementation in the diet may be therapeutic for pancreatitis.This study was supported by the Medical Research Service of the Veterans Administration of the United States.     

Generalised giant-cell tumour of bone: Successful treatment of pulmonary metastases with interferon α, a case report

We report on a 37-year-old female patient with generalised giant-cell tumour of the bone, whose tumour was refractory to conventional chemotherapy. Subsequent treatment with interferon 2a in increasing dosage from 4×10⁶ IU (three times a week) to 9×10⁶ IU (three times a week) led to a significant decrease of pulmonary metastases and a stabilisation of the disease after 12 months of treatment. In progressive chemotherapy refractory giant-cell tumour interferon 2a may be of therapeutic benefit.     

Temperature and Serum Proinflammatory Cytokine Changes in Patients with NSCLC after BAL

We examined the effects of bronchoalveolar lavage (BAL) and BAL fluid characteristics on the systemic proinflammatory cytokine expression and their relation to clinical and laboratory findings. Thirty patients suspected to have lung cancer were subjected to fiber-optic bronchoscopy (FOB) and BAL. Clinical and laboratory findings were determined at baseline, 4 h, and 24 h, including lung auscultation, temperature, chest X-ray, WBC, neutrophils, and serum IL-1, IL-6, and TNF-. BAL fluid characteristics were determined including cytokine levels. Fifteen volunteers served as controls to determine serum variation of the same cytokines. Significant temperature elevation was defined as 1°C increase compared to baseline. BAL was associated with temperature and serum TNF- and IL-6 but not IL-1 increase at 4 h. Four patients (13.3%) developed temperature over 38°C. In controls there were no significant changes between baseline and 24 h measurements for the same cytokines. Eleven patients (36.6%) developed a significant temperature elevation 4 h after BAL. These patients had a statistically significant (p < 0.05) increase in serum IL-6 at 4 h and in TNF- at both 4 and 24 h after BAL compared with the nonsignificant temperature increase group. BAL characteristics were not different between the two groups. On the other hand, BAL fluid IL-6 and TNF- levels were significantly higher (p < 0.05) in the nonfever group. Significant temperature increase was observed in 36.6% of the patients undergoing BAL and associated with significant serum TNF- and IL-6 increase at 4 h. Lung cytokines levels, alveolar macrophages, and BAL fluid characteristics are not related to temperature and serum proinflammatory cytokine increase. The hypothesis of alveolar macrophages derive from cytokine production and shift to the systemic circulation cannot be supported by our data. Abbreviations: NSCLC = non-small-cell lung carcinoma, BAL = bronchoalveolar lavage, FOB = fiber-optic bronchoscopy, IL-6 = interleukin 6, IL-1 = interleukin 1-beta, TNF- = tumor necrosis factor alpha, WBC = white blood cells, G-CSF = granulocyte colony stimulating factor, IM = intramuscular, ECG = electrocardiogram.     

Preventive Effect of FK 506 (Tacrolimus Hydrate) on Experimentally Induced Acute Liver Injury in Rats

The aim of the study was to investigate the effect of the immunosuppressant FK 506 (tacrolimus hydrate) on acute liver injury induced by Propionibacterium acnes and lipopolysaccharide (LPS). Acute liver injury was induced in male Wistar rats by injecting the animals with P. acnes (10 mg/rat), and administering LPS (10 g/rat) seven days later. One group was given FK 506 (1 mg/kg) 24 and 2 hr before administration of LPS, and the other group was given the same dose of saline. The 24-hr survival rate, serum alanine aminotransferase (ALT) concentration, and tumor necrosis factor (TNF) - mRNA and protein concentrations in the liver and spleen were then compared. Hepatic macrophages were also isolated from rats seven days after P. acnes injection, LPS, and FK 506 or saline were added to the culture supernatant, and TNF- production was studied. The 24-hr survival rate was 100% in the FK 506-treated group, in contrast with 16.6% in the saline group. Four hours after LPS injection, the serum ALT concentration was 755 ± 401 in the saline group versus 119 ± 42 units/ml (P < 0.01) in the FK 506-treated group. The serum TNF- concentration was lower in the FK 506-treated group (1419 ± 957 pg/ml) than in the saline group (9205 ± 2215) (P < 0.01). The mRNA and protein concentrations in the liver and spleen in the two groups did not differ significantly 1 hr after LPS injection but were significantly lower in the FK 506-treated group after 4 hr. FK 506 did not directly inhibit TNF- production by isolated cultured hepatic macrophages. FK 506 is unable to inhibit initial TNF- production by hepatic macrophages (or probably that by splenic macrophages either) stimulated by injection of LPS in P. acnes + LPS-induced acute liver injury. However, the immunosuppressant does limit hepatic damage by inhibiting subsequent aggravation of inflammation by the cytokine network.     

Short-term inhibition of peroxisome proliferator-activated receptor-γ coactivator-1α expression reverses diet-induced diabetes mellitus and hepatic steatosis in mice

Aims/hypothesis The coactivator of nuclear receptors, peroxisome proliferator-activated receptor- coactivator-1 (PGC-1) has been implicated in a series of events that contribute to the control of glucose metabolism. We have recently reported the use of a PGC-1 antisense oligonucleotide (PGC-1AS) that inhibits up to 60% of PGC-1 expression in pancreatic islets, leading to increased insulin secretion. This oligonucleotide was used in this study to try to ameliorate diet-induced type 2 diabetes in a genetically predisposed mouse strain (Swiss mice).Materials and methods Glucose and insulin tolerance tests, euglycaemic–hyperinsulinaemic clamp, immunoprecipitation assays, immunoblotting assays and immunohistochemistry were used in this investigation.Results Swiss mice became obese and overtly diabetic after 8 weeks of feeding with chow containing 24% saturated fat. One daily dose (1.0 nmol) of PGC-1AS significantly reduced glucose and increased insulin blood levels without affecting food intake and body weight. These effects were accompanied by a reduced area under the glucose curve during an intraperitoneal glucose tolerance test, an increased constant of glucose decay (K_itt) during an insulin tolerance test, and an increased glucose consumption rate during a euglycaemic–hyperinsulinaemic clamp. Moreover, mice treated with PGC-1AS presented an outstanding reduction of macroscopic and microscopic features of hepatic steatosis. These effects were accompanied by reduced expression or function of a series of proteins involved in lipogenesis.Conclusions/interpretation PGC-1 is an attractive target for pharmacological therapeutics in type 2 diabetes mellitus and diet-induced hepatic steatosis.     

Mucosal protective effects of vitamin E and misoprostol during acute radiation-induced enteritis in rats

Cytotoxic effects of ionizing radiation on gastrointestinal epithelium may be mediated by oxygen free radicals. Therapeutic intervention directed toward oxidant scavenging and increasing tissue oxygen tension may provide a novel approach to management. We investigated the effects of a nonenzymatic oxygen radical scavenger (vitamin E) and an exogenous PGE₁ analog known to increase mucosal blood flow (misoprostol) on acute radiation enteritis. Rats were pretreated with: (1) vitamin E, (2) misoprostol, or (3) a combination of both agents prior to 10 Gy abdominal radiation. Three days following irradiation, net fluid absorption usingin vivo isolated loops, mucosal histology, and mucosal morphometry using a computerized videoplan were determined in jejunum, ileum, and colon. Nonirradiated control intestine demonstrated net fluid absorption in all segments, which was not altered by vitamin E and/or misoprostol treatment. Irradiation significantly reduced net fluid absorption in jejunum, ileum, and colon. Vitamin E administered prior to irradiation maintained jejunal, ileal, and colonic fluid absorption near control levels. In contrast misoprostol or a combination of vitamin E and misoprostol did not provide protection against the injury caused by abdominal irradiation. Alterations in intestinal fluid absorption occurred without significant changes in histologic or morphometric appearance. In conclusion, ionizing radiation reducesin vivo intestinal fluid absorption without significant changes in histologic or morphometric appearance. Treatment with vitamin E, but not misoprostol, protects gastrointestinal mucosa against radiation-induced absorptive injury.This work was supported in part by grants from the Alberta Heritage Foundation for Medical Research and the Canadian Foundation of Ileitis and Colitis.     

Hepatic expression of interferon-α in chronic hepatitis B virus infection

Hepatic expression of interferon- (IFN-) was examined by immunohistochemistry in 90 Chinese patients (M/F 67:23, age: 14–69) with a spectrum of hepatitis B virus (HBV)-related chronic liver diseases. Immunoreactive IFN- was detected in sinusoidal cells in 79 patients (88%) and in mononuclear cells in 59 patients (65.6%). Patients with active liver diseases (chronic active hepatitis, active cirrhosis,N=55) had a higher level of IFN- expression compared to patients with inactive histology (N=35; sinusoidal cells,P<0.01; mononuclear cells,P<0.01). Cytoplasmic HBsAg, nuclear HBcAg, and cytoplasmic HBcAg were detected in 79 (88%), 42 (47%), and 23 (27%) patients respectively. Expression of IFN- in mononuclear cells correlated with the expression of cytoplasmic HBcAg (P<0.05) but not with nuclear HBcAg or cytoplasmic HBsAg. When the patients were divided into four different phases according to the natural history of chronic HBV infection, patients in the active liver disease phase had higher IFN- expression compared to the immunotolerant and late phase patients (P<0.01). Using double immunohistochemical staining, both IFN- and cytoplasmic HBcAg were frequently detected near inflammatory infiltrates but no correlation existed between the hepatic expression of HBsAg and IFN-. These data indicate that IFN- is expressed in the liver in HBV-related active liver diseases and that the reported suboptimal production of IFN- by PBMC in HBV-related chronic active liver diseases may be due to a redistribution of the IFN--producing mononuclear cells into the liver, the site of inflammation.     

Granulocyte elastase in assessment of severity of acute pancreatitis

Complexes of granulocyte elastase and ₁-antitrypsin are markers for granulocyte activation. In 75 patients with acute pancreatitis these complexes were immunologically determined daily in plasma during the first week of hospitalization. Patients were classified into three groups: mild pancreatitis (I, 1 complication, N=34), severe pancreatitis (II, 2 complications, N= 29), lethal outcome (III, N=12). Initially, granulocyte elastase (mean±sem) was lower in group I (348±39 g/liter) as compared to groups II (897±183 g/l) and III (799±244 g/liter), P<0.001 for I vs II + III. Initial elastase concentrations >400 g/liter were consistent with a severe or fatal course of the disease but did not distinguish between severe and lethal pancreatitis. In patients with mild or severe disease, mean elastase concentrations decreased continuously during the following days (197±15 g/liter in mild cases, 325±30 g/liter in severe cases at day 7). In patients with lethal disease, however, mean elastase concentrations even increased at day 2 and remained higher than 700 g/liter during the observation period. At days 1 and 2 the predictive value for severe or lethal disease of raised (>400 g/liter) elastase concentrations [positive predictive value (PPV) 82%, negative predictive value (NPV) 81%] was better than that of elevated (>100 mg/liter) C-reactive protein (PPV 73%, NPV 73%), elevated (>4.0 g/liter) ₁-antitrypsin (PPV 59%, NPV 50%), or decreased (<1.5 g/liter) ₂-macroglobulin (PPV 82%, NPV 67%). When the time course of the concentrations of the acute-phase proteins was studied, it was found that rises of granulocyte elastase were followed by elevated C-reactive protein levels after one day, by elevated ₁-antitrypsin levels after two days and by decreased ₂-macroglobulin levels after three to four days. We conclude that granulocyte elastase is a good early marker for the severity of acute pancreatitis. Compared with elevated levels of C-reactive protein and ₁-antitrypsin release of granulocyte elastase reflects an event that precedes acute-phase protein induction.     

Effects of human α-interferon on granulocyte-macrophage progenitor cells (CFU-GM) in vitro

Summary Two preparations of human interferon (IFN)- were assessed for their influence on granulocyte-macrophage progenitor cells (CFU-GM) in vitro. Both highly purified human IFN- Ly and recombinant IFN- 2a suppressed CFU-GM colony formation in a dose-dependent manner using low-density bone-marrow target cells. Suppression of CFU-GM colony formation was accompanied by an increase in clusters. However, depletion of monocytes, T lymphocytes and B lymphocytes from low-density bone-marrow cells resulted in insensitivity of progenitor cells to IFN-. These results demonstrate that the effects of human IFN- on myeloid progenitor cells (CFU-GM) are mediated by accessory cells within the bone marrow.