Background: To assess the effectiveness and side effects of interferon (IFN)γ therapy for chronic hepatitis C. Materials and methods: A single dose of 1 × 106 IU of natural IFNγ was administered intramuscularly daily for 4 weeks to five patients with chronic active hepatitis C. Alanine aminotransferase levels, 2′-5′ oligoadenylate synthetase activity, β2 microglobulin levels, IFNγ activity and HCV RNA levels were measured in sera. Results: ALT level, 2′-5′ oligoadenylate synthetase (2-5AS) activity, β2 microglobulin (BMG) level, and IFNγ activity increased from 1–2 weeks after the start of IFNγ. However, HCV RNA levels did not decrease during IFNγ administration. There were no serious adverse reactions. Conclusions: IFNγ, which has attracted attention for its immunoenhancement, is worthy to be investigated as a therapy for chronic hepatitis C. However, the use of IFNγ in combination with IFNα, β or other antiviral agents may be more rewarding because of the possibly weak antiviral action of IFNγ.
A new mutation site has been found in a case of cholinesterase (ChE) deficiency diagnosed upon routine blood screening. Genomic DNA was sequenced and four point mutations were found: P1 (exon 2) nucleotide 298 (CCA-TCA), codon 100 (proline-serine), which is a novel mutation site; P4 (exon 2) nucleotide 1410 (CGT-CGG), codon 470 (arginine not changed); PS (exon 3) nucleotide 1543 (CGT-TGT), codon 515 (arginine-threonine); and P6 (exon 4) nucleotide 1615 (GCA-ACA), codon 539 (alanine-threonine). The patient had three (P1, P5, P6) heterozygous and one (P4) homozygous mutations. The three other family members studied had one (P1) or two (P5 and 6) heterozygous mutations in addition to a P4 homozygous mutation but their serum levels of ChE were normal or only slightly decreased. We concluded that three simultaneous mutations at codons 298, 1543 and 1615 are required to reduce serum ChE activity and that the single mutation at codon 298 or two mutations at codon 1543 and 1615 are not enough to reduce ChE activity.
The purpose of this report is to evaluate the usefulness of serum hepatocyte growth factor (HGF) levels as a prognostic factor in hepatectomized patients with early postoperative hyperbilirubinemia (HB). The serum HGF levels of 14 patients with early postoperative HB (serum total bilirubin level above 5 mg/dl lasting at least 3 days), were measured perioperatively (prior to and 6 h, 1, 2, 3, 4, 7 and 14 days following surgery) and the relationship between changes in serum HGF levels and postoperative outcome was analyzed. The 14 patients were divided into two groups: 6 patients (Group A) ameliorated by conservative therapies within 7 days after surgery, and 8 patients (Group B) with prolonged HB more than 2 weeks after surgery. Five patients (83%) in Group A showed relatively low maximum serum HGF levels (< 2.0 ng/ml), whereas 7 patients (88%) in Group B showed high serum HGF levels (> 2.0 ng/ml). Six of the 7 patients in Group B developed hepatic failure and died. These results suggest that, in patients with HE or those in whom it is expected to occur soon after hepatectomy, the measurements of serum HGF levels would be beneficial for determining therapy and predicting postoperative outcomes. (C) 1997 Elsevier Science Ireland Inc.
We detected the hepatitis GB virus C genome by reverse transcription-polymerase chain reaction in three Japanese patients with chronic liver diseases. Partial nucleotide sequences of 5′ noncoding, envelope and NS3 regions had 88.6–90.1, 86.388.0, and 78.6–79.6% nucleotide sequence homology compared with the prototype GBV-C genome. However, they showed higher homology with each other (96.197.1, 88.7–91.2, 84.0–87.0%, respectively), suggesting that they were a genotype of GBV-C. The domains that were highly conserved among all these genomes were present in the 5′ noncoding region. The frequency of detecting the genome by the polymerase chain reaction was higher when we used primers designed on these domains compared with primers designed on the core and NS3 region. The virus genome was detected in eight of 50 (16.0%) consecutive Japanese patients with hepatocellular carcinoma who had received more than ten units of blood transfusion, three of 60 (5.0%) patients with non-B, non-C liver disease and 12 of 35 (34.3%) Malaysian patients with non-B chronic liver disease. Since there are no reliable assays to detect hepatitis GB virus C at present, the detection of the genome by the polymerase chain reaction should be useful for diagnosis. Further nucleotide sequence analysis of the genomes is necessary for epidemiological survey studies and vaccine strategy.
Intratumoral injection of 50% acetic acid solution has recently been performed as a new local therapy for hepatocellular carcinoma. The use of ethanol in this therapy has problems of high dosage, frequent injections, and local tumor recurrence. Probably, in these regards the usefulness of acetic acid with potent coagulating and necrotizing effects cannot be denied. We have previously reported the acute toxicity of acetic acid injected into the normal rat liver at various concentrations. In the present study, we assessed the acute toxicity and necrotizing effect of 50% acetic acid solution and its effect on other organs after injection into the normal rat liver in different volumes. With increases in the volume of acetic acid, the size of the necrotic zone tended to increase and the mortality rate rose. The LD50 of 50% acetic acid calculated with Behrens' method was 0.279 ml/kg. Histopathological examination of rats died immediately after acetic acid injection revealed acute pulmonary edema and marked congestion of the kidneys and spleen. Therefore, if this agent is used clinically, sufficient attention must be paid to its dose level and its acute toxicity.
We studied the proliferative activity of hepatocellular carcinoma (HCC) by immunohistochemical staining with Ki-67, a monoclonal antibody to nuclear protein observed in the proliferative period of the cell cycle, and evaluated the usefulness of Ki-67 as a prognostic factor. Of the patients who underwent hepatectomy for HCC at our department, 56 cases in which pathological studies for the tumor showed no degeneration or necrosis were selected. Immunohistochemical staining was performed according to the method of Shi et al., using MIB-1 monoclonal antibody (Immunotech, SA) as a primary antibody. Ki-67 L.I. was significantly higher in chronic hepatitis (CH) and liver cirrhosis (LC) than in normal liver, and the L.I. was also significantly higher in non-cancerous tissue of the patients with HCC than in tissues with other liver diseases. In patients with a history of portal hypertension, the L.I. in non-cancerous liver tissue was significantly lower at the previous operation for portal hypertension without HCC than that at the subsequent operation for HCC. The L.I. was significantly higher in patients who showed a high serum AFP level and in those who had intrahepatic metastasis. It also tended to be higher in patients with vessel invasion, infiltration to capsule, aneuploidy tumor cells, and poor differentiation. The cumulative survival rate was significantly lower in patients with the L.I. of 10% or higher than in those with the L.I. of less than 10% in cancerous region. Recurrence was observed earlier after hepatectomy in patients with higher L.I. and the L.I. was significantly higher in those who had recurrence within 12 months. In the patients who could be followed up, the L.I. was significantly higher in those who had multiple recurrence after hepatectomy than in those who had single-lesion recurrence. The incidence of HCC was higher in patients with CH or LC in which the proliferative activity was abnormally enhanced for a prolonged period, suggesting that hepatocyte hyperproliferation triggers hepatic carcinogenesis. In addition, long-term survival was expected even in patients with advanced HCC if the L.I. in preoperative biopsy specimens was low, while in patients with high L.I., postoperative recurrence and distant metastasis may occur more frequently, and postoperative supplementary therapies may be necessary, even when the tumor is completely resectable. In conclusion, Ki-67 staining is considered to be useful for evaluation of the malignant potential of HCC.
Recent studies indicated that bile duct ligation down-regulates the expression of Na+/taurocholate cotransporting polypeptide and Na+-dependent taurocholate uptake by basolateral membrane vesicles in the rat. These findings suggest that hepatic taurocholate uptake in bile duct-ligated rats is mediated by the organic anion transporting polypeptide, a Na+-independent taurocholate uptake system which is common for sulfobromophthalein uptake. Therefore, the effect of sulfobromophthalein on biliary excretion of taurocholate and pravastatin in bile ductligated rats was studied. Although biliary excretion of pravastatin was markedly inhibited by sulfobromophthalein, biliary taurocholate excretion was not affected by sulfobromophthalein in bile duct-ligated rats. The excretory maximum of sulfobromophthalein in bile duct-ligated rats was reduced to one-fifth of control rats. These findings indicate that, in the bile duct-ligated rats, taurocholate uptake is mediated not by the multispecific organic anion transporter, but by other uptake system(s).
Using newly-designed primers for semi-nested reverse transcriptase-polymerase chain reaction to detect 5′-untranslated region of HGVGB-C hepatitis virus RNA, we screened 45 patients and found three HGVGB-C positive patients (with fulminant hepatitis and liver cirrhosis, and on hemodialysis). Restriction fragment length polymorphism and single-strand conformation polymorphism analysis revealed a similar pattern in all positive specimens. The sequence amplified by our primers might be a highly conserved sequence. We believe our primers are useful for screening of HGVGB-C virus RNA.
Of 112 patients with chronic hepatitis C, 12 (11%) tested positive for RNA of GB virus C (GBV-C) by reverse-transcription polymerase chain reaction with primers deduced from the 5′ untranslated region. RNAs of GBV-C and hepatitis C virus (HCV) were followed in the 12 patients before and after they received 19 trials of interferon (IFN) therapy. GBV-C RNA disappeared from serum in 11 (58%) trials on eight patients. However, it stayed negative at 6 months after IFN only in two patients. One of them regained GBV-C RNA at 12 months and kept it thereafter, while an additional patient who failed to clear it at the completion of IFN turned negative at 24 months after therapy. Thus, two patients (17%) became persistently negative for GBV-C RNA. HCV RNA disappeared from serum at the completion of ten trials (53%) on eight patients, and stayed negative in three patients (25%). Two of them did not lose GBV-C RNA but kept normal transaminase levels. These results indicate that GBV-C is susceptible to IFN with a sensitivity comparable to but independent of HCV, and that GBV-C by itself would not elevate transaminases in hepatitis C patients who respond to IFN.
Excess matrix in hepatic fibrosis results from both fibrogenic stimulation of stellate cells by TGFβ1 and cell proliferation due to induction of β-platelet derived growth factor receptor (β-PDGFR). In this paper, treatment of culture-activated rat stellate cells with anti-TGFβ inhibited collagen and fibronectin mRNA expression by 82 and 58%, respectively, versus control cells. In vivo, anti-TGFβ inhibited collagen I gene expression by 86% in stellate cells isolated from rats treated with CC14 compared with control antibody. In contrast to stellate cells, anti-TGFβ had no effect on collagen I gene expression in isolated sinusoidal endothelial cells. Anti-TGFβ administered in vivo to rats with liver injury also reduced expression of stellate cell β-PDGFR mRNA to that of control animals. Anti-TGFβ antibody had no effect on the histologic appearance of the tissue. These data support a role for TGFβ in stellate cell matrix expression and provide evidence for transmodulation of PDGF receptor by TGFβ in vivo. However, inhibition of TGFβ alone may not be adequate to attenuate severe hepatic injury and fibrosis.
This study investigated the relationship between telomerase activity and telomerase RNA component (telomerase RNA) in human liver tissues. Telomerase RNA was detected in all hepatocellular carcinoma (HCC) tissues (2020), and telomerase activity was detected in 18 (90.00%) of the 20 HCC tissues. Telomerase RNA was detected in 17 (89.47%) of 19 livers with liver cirrhosis, and telomerase activity was detected in eight (42.11%) of the 19 tissues. In livers with chronic hepatitis, telomerase RNA was detected in 11 (64.71%) of 17, and telomerase activity was detected in six (35.29%) of the 17 tissues. Telomerase RNA was detected in all the tissues with telomerase activity detected, i.e. 19 HCC tissues, 17 liver cirrhosis tissues and 11 chronic hepatitis tissues. In all normal tissues (n = 3), telomerase RNA and activity were not detected. From our results, one can deduce that the production of telomerase is thought to be under the control of telomerase RNA, the possibility is great that chronic hepatitis tissues and liver cirrhosis tissues become HCC when the telomerase RNA is positive.
The early vascular injury in the liver was characterized in an experimental model of multiple organ failure (MOF). Significant increases of hyaluronic acid levels (660%) and plasma alanine aminotransferase activities (1050%) were observed after 20 min hepatic ischemia followed by 4 h reperfusion and injection of 0.5 mg/kg Salmonella enteritidis endotoxin at 30 min reperfusion. Morphological evaluation of sinusoids with transmission electron microscopy indicated neutrophil and Kupffer cell activation as well as damage or loss of sinusoidal endothelial cells. Hepatocellular injury was evident from fused microvilli and blebbed plasma membranes. Treatment with the 21-aminosteroid tirilazad mesylate (U-74006F) (2 × 3 mg/kg) reduced plasma hyaluronic acid levels by 61% and plasma transaminase activities by 69% suggesting a beneficial effect on sinusoidal endothelial cell and parenchymal cell injury. This was confirmed by morphology. Our data provide morphological and functional evidence for severe injury to sinusoidal endothelium and the vascular pole of hepatocytes in this model of MOF. U-74006F significantly protected the liver against this Kupffer cell- and neutrophil-mediated injury. Thus, U-74006F may be a promising therapeutic for liver dysfunction and failure during a local or systemic inflammatory response.
It is yet not fully uncovered where and how translation starts in GB virus C or hepatitis G virus (GBV-CHGV), and, accordingly, information on structural proteins of this virus has been scarce. To know a vertebral architecture of GBV-CHGV genome at least for its 5′ portion, we analyzed 31 Japanese isolates of GBV-CHGV. Our results indicated that most of the Japanese isolates were classifiable into ‘Japanese type’, two isolates into ‘North American type’, one into ‘African type’, and another into the fourth distinct type. Despite such sequence diversity across individual types, however, several common characteristics existed. (1) The AUG that resides just downstream of an oligopyrimidine tract was conserved by all isolates as the first in-frame initiation codon for the polyprotein. (2) The sequence upstream of this AUG had many covariant nucleotide substitutions, suggesting conserved secondary structure. (3) The ORF that starts with this AUG did not code for core. (4) Predicted amino acid sequence of putative E1 and E2 was highly conserved among different types, and did not contain hypervariable regions.
Organic anions can uncouple bile salt secretion from the secretion of phospholipid and cholesterol. The uncoupling mechanism appears to be dependent on the type of anion present. To investigate the effect that different uncoupling mechanisms may have on the flow of bile and its composition, rats were depleted of bile salt pool by overnight biliary diversion and reinfused intravenously with sodium taurocholate (STC) at a constant rate (160 nmol/min per 100 g b.w.). After bile salt secretion had achieved a steady state, sulfobromophthalein (BSP) or papaverine was continuously administered at a rate of 100 nmol/min per 100 g b.w. with STC. During the intravenous infusion of BSP or papaverine, the output of biliary lecithin and cholesterol decreased without affecting bile salt output. The degree of fatty acyl chain saturation in biliary lecithin was increased by BSP infusion and decreased by papaverine infusion. The mechanism by which bile salt secretion is uncoupled from the secretion of phospholipid and cholesterol affects the acyl chain composition of secreted lecithin. The changes in composition of lecithin acyl chains may reflect the intervention by different anions at different points in the lipid transport pathway.
The role of cytokines in the etiology of liver injury and their contribution to the systemic manifestations that occur in patients with liver disease, are not clearly understood. Aim: To study if serum levels and in vitro blood mononuclear cell (BMC) production of IL-Iβ and TNFα are related to the severity of alcoholic liver cirrhosis, and identify potential factors that can modify cytokine production in these patients. Serum levels, spontaneous and in vitro LPS stimulated BMC production of Interleukin-Iβ (IL-Iβ) and Tumor Necrosis Factor α (TNFα), were measured in 38 patients with alcoholic cirrhosis Child B or C, and nine normal volunteers. Serum levels and spontaneous in vitro production of IL-Iβ and TNFα were below detection limits. There were no differences between normal controls and Child B or C patients in LPS-stimulated production of IL-Iβ or TNFα. However eight patients with alcoholic hepatitis or infections superimposed on cirrhosis, had higher levels on LPS-stimulated BMC production of IL-Iβ (12.9 ± 5.8 ng/ml) and TNFα (4.9 ± 2.3 ng/ml) than the rest of cirrhotic patients (5.3 ± 3.5 and 1.8 ± 0.9 ng/ml). There was no association between IL-Iβ and TNFα BMC production and parameters of liver function or recent alcohol ingestion. Increased levels of IL-Iβ and TNFα production by stimulated BMC are associated with acute inflammatory events in cirrhotic alcoholic patients and not with the severity of liver disease.
Effects of interleukin-4 (IL-4) and interferon-γ (IFN-γ) on rat Kupffer cell (KC) functions were investigated. Lipopolysaccharide (LPS)-stimulated KCs pretreated with IL-4 produced much more prostaglandin E2 than those pretreated with IFN-γ. In contrast, LPS-stimulated KCs pretreated with IFN-γ produced much more tumor necrosis factor-α, and nitrate and nitrite than those pretreated with IL-4. Different morphologic changes were induced after the culture with each cytokine; transformation of KCs into multinucleated giant cells with IL-4 was especially noted. These results suggest that IFN-γ and IL-4, produced by different subsets of T-helper lymphocytes, differently induce KC into specific morphology, and change KC response to endotoxins, possibly resulting in a modulation of hepatic inflammation.
Serum soluble HLA-class I and CD8 molecules were sequentially (pre, during, and post-therapy) measured by sandwich ELISA, in 20 interferon (IFN)-treated chronic hepatitis C (CHC) patients, who were judged as responders (n = 8) or non-responders (n = 12) with respect to the response to the therapy. Pretreatment levels of sHLA-I and sCD8 did not differ between responders and non-responders. Both values increased just after the start of therapy and peaked at 2–4 weeks. The IFN was discontinued at 24 weeks, when sHLA-I decreased to baseline levels in responders, whereas they remained at levels significantly higher than baseline levels even at 48 weeks in non-responders (P < 0.01). sCD8 decreased significantly under than baseline levels at 48 weeks in responders (P < 0.05), in contrast in non-responders, sCD8 did not decrease beyond baseline levels. In summary, serum levels of sHLA-I and sCD8 were elevated by the IFN treatment, and serial follow-up of these immunological parameters could provide additive information regarding the response to IFN therapy.
The effects of glycyrrihizin on liver-function, amounts of HCV RNA and viral complexity in patients with chronic hepatitis C were examined. The subjects were 27 type C chronic hepatitis patients who did not respond to previous interferon therapy. They were given 60 ml of Stronger Neo-Minophagen C, a preparation of glycyrrhizin combined with glycine and cysteine, three times a week for 16 weeks. The treatment was effective in 20 patients (74.1%), whose serum levels of aminotransferase fell to one half of that before treatment, but was not effective in seven patients (25.9%). No significant differences were noted at the beginning of the therapy in the backgrounds (age, sex, serum levels of aminotransferase, etc.( of the effective and non-effective groups. Also, no significant differences in HCV RNA quantity and viral complexity of HCV RNA were observed between the two groups. The results suggest that glycyrrhizin improves serum ALT levels without changing the amounts of HCV RNA and viral complexity, suggesting that the effect might depend on host factors such as the immune response.
Aim, clinical usefulness of a non radioactive HBV DNA measuring method was evaluated. Subjects and methods, the subjects were 8 anti-HBe positive chronic hepatitis B (CH-B) patients and five CH-B patients who were orally administered reverse transcriptase inhibitor (RTI). Serum HBV DNA and DNA-P were serially determined at 1.5–2 month intervals for 1 year in CH-B with anti-HBe positive patients and before, 2, 4, 8, and 12 weeks after RTI administration. Serum HBV DNA was determined by two different non radioactive method (Viraprobe HB Lumi, Quantiplex HBV DNA). Results, the HBV DNA levels measured with two methods were highly correlated (P < 0.0001). Generally, the results for serum HBV DNA and the DNA-P were also correlated (P < 0.01). While, among a total of 51 determinations in CH-B patients with anti-HBe positive, discrepancies between serum HBV DNA and DNA-P from different sensitivities were noted in 15 determinations, and in all cases the pattern was HBV DNA positive and DNA-P negative. From 2 to 12 weeks after RTI administration, the same pattern was observed in three cases. These results were thought to be mainly due to the high sensitivity of HBV DNA measurement. (Conclusion) The non radioactive methods of measuring of HBV DNA are useful for detecting low level HBV replication.
Serum HBV DNA levels were quantified simultaneously by slot-blot hybridization using digoxigenin-labeled probe (S) and by Abbott HBV DNA assay (A). Samples, 69, from 64 patients seropositive for hepatitis B e antigen (HBeAg) were included and the percentages of hepatocytes containing hepatitis B core antigen (HBcAg) were also compared. The extent of HBcAg staining was only moderately correlated with HBV DNA levels detected by either Abbott (R2 = 0.41; P < 0.01) or slot-blot hybridization (R2 = 0.33; P < 0.01) assays. On the other hand, HBV DNA concentrations assessed by either method were well correlated although the levels differed markedly (S = 3.0 × A1.31; R2 = 0.85; P < 0.01). These results indicated that digoxigenin-labeled probe can be used to quantify serum HBV DNA concentration, although a different range of levels was obtained when compared with Abbott HBV DNA assay.
