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Vascular staplers are routinely used in laparoscopic liver resection, which has become a standard procedure in advanced medical facilities. Although previous reports have outlined the benefits of staple line reinforcement (SLR), its application in Glissonean pedicle transection during hepatic resection remains poorly studied. This study investigated surgical SLR as a tool to enhance staple line strength and improve perioperative hemostasis. Here, 10 patients who underwent laparoscopic liver resection using the Tri-StapleTM2.0 Reinforced Reload were included. Patient characteristics, surgical details, and outcomes were assessed. The results demonstrated successful outcomes with no complications related to bile leakage or injuries during staple insertion. Overall, our findings suggest that SLR can be safely utilized in Glissonean pedicle transection during laparoscopic liver resections. Further studies are required to comprehensively evaluate its benefits compared with conventional surgical staplers., (© 2024 Asia Endosurgery Task Force and Japan Society of Endoscopic Surgery and John Wiley & Sons Australia, Ltd.)

Non-alcoholic steatohepatitis (NASH) occurrence has been increasing and is becoming a major cause of liver cirrhosis and liver cancer. However, effective treatments for NASH are still lacking. We examined the benefits of serum-free conditioned medium from stem cells derived from human exfoliated deciduous teeth (SHED-CM) on a murine non-alcoholic steatohepatitis (NASH) model induced by a combination of Western diet (WD) and repeated administration of low doses of carbon tetrachloride intraperitoneally, focusing on the gut-liver axis. We showed that repeated intravenous administration of SHED-CM significantly ameliorated histological liver fibrosis and inflammation in a murine NASH model. SHED-CM inhibited parenchymal cell apoptosis and reduced the activation of inflammatory macrophages. Gene expression of pro-inflammatory and pro-fibrotic mediators (such as Tnf-α, Tgf-β, and Ccl-2) in the liver was reduced in mice treated with SHED-CM. Furthermore, SHED-CM protected intestinal tight junctions and maintained intestinal barrier function, while suppressing gene expression of the receptor for endotoxin, Toll-like receptor 4, in the liver. SHED-CM promoted the recovery of Caco-2 monolayer dysfunction induced by IFN-γ and TNF-α in vitro. Our findings suggest that SHED-CM may inhibit NASH fibrosis via the gut-liver axis, in addition to its protective effect on hepatocytes and the induction of macrophages with unique anti-inflammatory phenotypes., (© 2021. The Author(s).)

Warm ischemia and reperfusion injury (IRI) is a prognostic factor in donation after cardiac death donor transplantation. However, a reliable method to predict IRI before transplantation has not been established. The aim of this study was to identify predictive markers of hepatic IRI by simultaneous measurement of endogenous molecules using matrix-assisted laser desorption/ionization-imaging mass spectrometry (MALDI-IMS). Rats were subjected to hepatic warm ischemia (70%) for 30 or 90 minutes and subsequent reperfusion. The livers were collected at the end of ischemia and 1 hour, 6 hours, and 24 hours after reperfusion. The liver tissue sections were applied to IMS (m/z 200-2000). Candidate molecules were identified by tandem mass spectrometry. Imaging mass spectrometry (IMS) revealed a significant increase in the taurine conjugates of dihydroxycholanoic acid (TDHCA) during ischemia and a tendency to return to the basal level after reperfusion. Notably, high-resolution measurements revealed focal accumulation of TDHCA in the intrahepatic bile duct with ischemic time. In conclusion, IMS is a useful method to detect minute changes provoked by ischemia, which are barely detectable in assays involving homogenization. Accordingly, focal accumulation of TDHCA during ischemia may be a candidate marker for predicting later IRI., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The effect of selective transcatheter arterial embolisation (TAE) using trisacryl gelatine microspheres (TGMs) in the normal canine liver was investigated. Selective embolisation was achieved by injecting TGMs into the left hepatic artery through a microcatheter in four healthy dogs. After embolisation, computed tomography (CT), biochemical analysis and histological examination were performed during a 12-week observation period. Embolisation was successful in all four dogs. Postoperative CT revealed consistent embolisation of the artery within the experimental period in three dogs. Hepatic enzyme levels slightly increased after embolisation but tapered to normal ranges. Histological examinations revealed no abnormal changes. Thus, selective TAE with TGMs was well tolerated in normal dogs and may be applicable to canine hepatocellular carcinoma., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Choline is used to synthesize phospholipids and a lack of choline induces a number of liver‑related diseases, including non‑alcoholic steatohepatitis. The current study characterized the choline uptake system, at molecular and functional levels, in the immortalized human hepatic cell line, Fa2N‑4, to identify the specific choline transporter involved in choline uptake. The present study also assesed whether choline deficiency or the inhibited choline uptake affected cell viability and apoptosis. Reverse transcription‑quantitative polymerase chain reaction (PCR) revealed choline transporter‑like protein 1 (CTL1) and CTL2 mRNA and protein expression in Fa2N‑4 cells. [Methyl‑3H]choline studies revealed choline uptake was saturable and mediated by a single transport system that functioned in a Na+‑independent but pH‑dependent manner, which was similar to CTL1. Hemicholinium‑3 (HC‑3), which is a choline uptake inhibitor, and choline deficiency inhibited cell viability, increased caspase‑3 and ‑7 activities, and increased fluorescein isothiocyanate‑Annexin V immunofluorescent staining indicated apoptosis. Immunofluorescent staining also revealed CTL1 and CTL2 localized in plasma and mitochondrial membranes, respectively. [Methyl‑3H]choline uptake was enhanced by a protein kinase C (PKC) activator, phorbol‑12‑myristate 13‑acetate (PMA). Immunofluorescence staining and western blot analysis demonstrated increased CTL1 expression on the cell membrane following PMA treatment. The results of current study indicated that extracellular choline is primarily transported via CTL1, relying on a direct H+ gradient that functions as a driving force in Fa2N‑4 cells. Furthermore, it was hypothesized that CTL1 and the choline uptake system are strongly associated with cell survival, and that the choline uptake system is modulated by PKC signaling via increased CTL1 expression on the cell surface. These findings provide further insights into the pathogenesis of liver disease involving choline metabolism.

Background/aim: Lenvatinib (LEN) has been developed for the treatment of unresectable hepatocellular carcinoma (u-HCC). We aimed to elucidate the relative change in hepatic reserve function early following LEN treatment in affected patients., Materials/methods: From March 2018 to April 2019, 123 u-HCC patients (median age 71 years; male:female ratio 95:28; Child-Pugh score 5:6:7 = 65:50:8; modified albumin-bilirubin [mALBI] grade 1:2a:2b:3 = 44:28:50:1, Barcelona Clinic Liver Cancer stage A:B:C = 1:49:73) were enrolled. Relative changes in hepatic reserve function at 2 and 4 weeks after starting LEN were retrospectively evaluated., Results: The median survival was 11.3 months. The Child-Pugh score declined from the start to 4 weeks after commencing LEN (score 5:6:7:8:9:≥10 = 65:50:8:0:0:0 vs. 50:39:22:8:0:4, p < 0.001). A comparison among ALBI scores at the start of LEN and those at 2 and 4 weeks revealed significant relative changes (-2.36 ± 0.45 to -2.20 ± 0.49 at 2 weeks, -2.15 ± 0.50 at 4 weeks, p < 0.001, Bonferroni method), while there was no significant difference between those at 2 and 4 weeks (p= 0.210, Bonferroni method). Assessments of relative changes of ALBI score in patients divided by mALBI grade 1, 2a, and 2b or more showed a significant decline in score regardless of grade (-2.82 ± 0.17 to -2.53 ± 0.34, p < 0.001; -2.46 ± 0.10 to -2.31 ± 0.33, p = 0.017; and -1.90 ± 0.26 to -1.75 ± 0.42, p= 0.009, respectively)., Conclusion: Decline in hepatic function is common in the early stage (≤4 weeks, especially within 2 weeks) after introducing LEN. It is important to introduce molecular targeting agent drugs for u-HCC in patients with better hepatic function, who show transarterial catheter chemoembolization failure, as much as possible, along with consideration of the negative influence of LEN on the early response of hepatic function., (© 2019 S. Karger AG, Basel.)

Background and Purpose: We reported previously that hydrogen gas (H 2 ) reduced hepatic ischemia and reperfusion injury (IRI) after prolonged cold storage (CS) of livers retrieved from heart-beating donors. The present study was designed to assess whether H 2 reduced hepatic IRI during donation of a cardiac death (DCD) graft with subsequent CS., Methods: Rat livers were harvested after 30-min cardiac arrest and stored for 4 h in University of Wisconsin solution. The graft was reperfused with oxygenated buffer, with or without H 2 (H 2 or NT groups, respectively), at 37° for 90 min on isolated perfused rat liver apparatus., Results: In the NT group, liver enzyme leakage, apoptosis, necrosis, energy depletion, redox status, impaired microcirculation, and bile production were indicative of severe IRI, whereas in the H 2 group these impairments were significantly suppressed. The phosphorylation of cytoplasmic MKK4 and JNK were enhanced in the NT group and suppressed in the H 2 group. NFkB-p65 and c-Fos in the nucleus were unexpectedly unchanged by IRI regardless of H 2 treatment, indicating the absence of inflammation in this model., Conclusion: H 2 was observed to ameliorate IRI in the DCD liver by maintaining microcirculation, mitochondrial functions, and redox status, as well as suppressing the cytoplasmic MKK4-JNK-mediated cellular death pathway.

Fatty acid desaturase 2 (FADS2) is responsible for the first desaturation reaction in the synthesis of highly unsaturated fatty acids (HUFAs), such as arachidonic acid (20:4n-6) and eicosapentaenoic acid (20:5n-3), and is involved in Mead acid (20:3n-9) production during essential fatty acid deficiency (EFAD). In this study, an obvious hepatic lipid accumulation was observed in EFAD mice treated with a FADS2 inhibitor. FADS2 inhibition in the EFAD state reduced secretion of very low-density lipoprotein (VLDL) and markedly diminished Mead acid in phosphatidylcholine (PC) in the liver and plasma. As the results, the amount of C20 HUFAs in hepatic and plasma PC dramatically reduced in the EFAD mice treated with a FADS2 inhibitor, whereas the decrease of C20 HUFA levels of PC in EFAD mice was not observed because of the increased Mead acid in PC. These results supposed that Mead acid in PC is important as a component of VLDL. It is possible that Mead acid plays the role of a substitute of HUFAs in VLDL secretion during EFAD., (Copyright © 2018 Elsevier Inc. All rights reserved.)

The use of synthetic biomarkers is an emerging technique to improve disease diagnosis. Here, we report a novel design strategy that uses analyte-responsive acetaminophen (APAP) to expand the catalogue of analytes available for synthetic biomarker development. As proof-of-concept, we designed hydrogen peroxide (H 2 O 2 )-responsive APAP (HR-APAP) and succeeded in H 2 O 2 detection with cellular and animal experiments. In fact, for blood samples following HR-APAP injection, we demonstrated that the plasma concentration ratio [APAP+APAP conjugates]/[HR-APAP] accurately reflects in vivo differences in H 2 O 2 levels. We anticipate that our practical methodology will be broadly useful for the preparation of various synthetic biomarkers., (© 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Hydrogen gas reduces ischemia and reperfusion injury (IRI) in the liver and other organs. However, the precise mechanism remains elusive. We investigated whether hydrogen gas ameliorated hepatic I/R injury after cold preservation. Rat liver was subjected to 48-h cold storage in University of Wisconsin solution. The graft was reperfused with oxygenated buffer with or without hydrogen at 37° for 90 min on an isolated perfusion apparatus, comprising the H 2 (+) and H 2 (-) groups, respectively. In the control group (CT), grafts were reperfused immediately without preservation. Graft function, injury, and circulatory status were assessed throughout the perfusion. Tissue samples at the end of perfusion were collected to determine histopathology, oxidative stress, and apoptosis. In the H 2 (-) group, IRI was indicated by a higher aspartate aminotransferase (AST), alanine aminotransferase (ALT) leakage, portal resistance, 8-hydroxy-2-deoxyguanosine-positive cell rate, apoptotic index, and endothelial endothelin-1 expression, together with reduced bile production, oxygen consumption, and GSH/GSSG ratio (vs. CT). In the H 2 (+) group, these harmful changes were significantly suppressed [vs. H 2 (-)]. Hydrogen gas reduced hepatic reperfusion injury after prolonged cold preservation via the maintenance of portal flow, by protecting mitochondrial function during the early phase of reperfusion, and via the suppression of oxidative stress and inflammatory cascades thereafter., (Copyright © 2016 International Center for Artificial Organs and Transplantation and Wiley Periodicals, Inc.)

In the present study, we examined the role of indoleamine 2,3-dioxygenase (IDO) in the development of CCl4-induced hepatic fibrosis. The liver fibrosis induced by repetitive administration with CCl4 was aggravated in IDO-KO mice compared to WT mice. In IDO-KO mice treated with CCl4, the number of several inflammatory cells and the expression of pro-inflammatory cytokines increased in the liver. In the results, activated hepatic stellate cells (HSCs) and fibrogenic factors on HSCs increased after repetitive CCl4 administration in IDO-KO mice compared to WT mice. Moreover, the treatment with l-tryptophan aggravated the CCl4-induced hepatic fibrosis in WT mice. Our findings demonstrated that the IDO deficiency enhanced the inflammation in the liver and aggravated liver fibrosis in repetitive CCl4-treated mice., Competing Interests: The authors have declared that no competing interests exist.

Probe electrospray ionization (PESI) is a recently developed ionization technique that enables the direct detection of endogenous compounds like metabolites without sample preparation. In this study, we have demonstrated the first combination use of PESI with triple quadrupole tandem mass spectrometry (MS/MS), which was then applied to intact endogenous metabolite analysis of mice liver, achieving detection of 26 metabolites including amino acids, organic acids, and sugars. To investigate its practicality, metabolic profiles of control and CCl4-induced acute hepatic injury mouse model were measured by the developed method. Results showed clear separation of the two groups in score plots of principal component analysis and identified taurine as the primary contributor to group separation. The results were further validated by the established gas chromatography/MS/MS method, demonstrating the present method's usefulness. In addition, we preliminarily applied the method to real-time analysis of an intact liver of a living mouse. We successfully achieved monitoring of the real-time changes of two tricarboxylic acid cycle intermediates, α-ketoglutaric acid and fumaric acid, in the liver immediately after pyruvic acid injection via a cannulated tube to the portal vein. The present method achieved an intact analysis of metabolites in liver without sample preparation, and it also demonstrates future possibility to establish in vivo real-time metabolome analysis of living animals by PESI/MS/MS.

For successful mitochondrial transgene expression, two independent processes, i.e., developing a mitochondrial gene delivery system and construction of DNA vector to achieve mitochondrial gene expression, are required. To date, very few studies dealing with mitochondrial gene delivery have been reported and, in most cases, transgene expression was not validated, because the construction of a reporter DNA vector for mitochondrial gene expression is the bottleneck. In this study, mitochondrial transgene expression by the in vivo mitochondrial gene delivery of an artificial mitochondrial reporter DNA vector via hydrodynamic injection is demonstrated. In the procedure, a large volume of naked plasmid DNA (pDNA) is rapidly injected. We designed and constructed pHSP-mtLuc (CGG) as a mitochondrial reporter DNA vector that possesses a mitochondrial heavy strand promoter (HSP) and an artificial mitochondrial genome with the reporter NanoLuc (Nluc) luciferase gene that records adjustments to the mitochondrial codon system. We delivered the pDNA into mouse liver mitochondria by hydrodynamic injection, and detected exogenous mRNA in the liver using reverse transcription PCR analysis. The hydrodynamic injection of pHSP-mtLuc (CGG) resulted in the expression of the Nluc luciferase protein in liver and skeletal muscle. Our mitochondrial transgene expression reporter system would contribute to mitochondrial gene therapy and further studies directed at mitochondrial molecular biology.

Background and Aim: The prevalence of nonalcoholic fatty liver disease (NAFLD) is higher in men than in women, but according to some epidemiological studies, this gender difference disappears after menopause. Estrogen therapy protects against NAFLD and nonalcoholic steatohepatitis (NASH) after menopause. We investigated the therapeutic effect of raloxifene, a second-generation selective estrogen-receptor modulator, on NASH induced by a choline-deficient high-fat (CDHF) diet in female ovariectomized (OVX) mice., Methods: Seven-week-old female C57BL/6J mice were divided into three experimental groups as follows: (1) sham operation (SHAM group), (2) ovariectomy (OVX group), and (3) ovariectomy + raloxifene (intraperitoneal injection, 3 mg/kg body weight/day; OVX + RLX group). These three groups of mice were fed a CDHF diet for 8 weeks; choline-sufficient high-fat (CSHF) diet was used as control diet. Serum biochemical indicators of hepatic function and liver histological changes were evaluated., Results: Compared with CSHF diet, ovariectomy enhances liver injury and fibrosis in CDHF diet-fed mice. Serum alanine aminotransferase (ALT) levels were significantly lower in the OVX + RLX group than in the OVX group. The OVX group developed extensive steatosis with inflammation and fibrosis. Lobular inflammatory scores and fibrosis staging in the OVX + RLX group were significantly lower than in the OVX group. Furthermore, the OVX + RLX group exhibited significantly higher expression of hepatic estrogen receptor-α, which was significantly lower in the OVX group than in the SHAM group., Conclusions: Raloxifene may ameliorate progression of liver fibrosis of NASH induced by CDHF diet in ovariectomized female mice, and up-regulation of estrogen receptor-α may play an important role in the beneficial effects of raloxifene on NASH.

Background and Purpose: Hydrogen sulfide (H2S) ameliorates hepatic ischemia and reperfusion injury (IRI), but the precise mechanism remains elusive. We investigated whether sodium hydrogen sulfide (NaHS), a soluble derivative of H2S, would ameliorate hepatic IRI, and if so, via what mechanism., Methods: Mice were subjected to partial warm ischemia for 75 min followed by reperfusion. Either NaHS or saline was administered intravenously 10 min before reperfusion. The liver and serum were collected 3, 6, and 24 h after reperfusion., Results: In the NaHS(-) group, severe IRI was apparent by the ALT leakage, tissue injury score, apoptosis, lipid peroxidation, and inflammation (higher plasma TNF-α, IL-6, IL-1β, IFN-γ, IL-23, IL-17, and CD40L), whereas IRI was significantly ameliorated in the NaHS(+) group. These effects could be explained by the augmented nuclear translocation of Nrf2, and the resulting up-regulation of HO-1 and thioredoxin-1. Phosphorylation of the PDK-1/Akt/mTOR/p70S6k axis, which is known to mediate pro-survival and anti-apoptotic signals, was significantly augmented in the NaHS(+) group, with a higher rate of PCNA-positive cells thereafter., Conclusion: NaHS ameliorated hepatic IRI by direct and indirect anti-oxidant activities by augmenting pro-survival, anti-apoptotic, and anti-inflammatory signals via mechanisms involving Nrf-2, and by accelerating hepatic regeneration via mechanisms involving Akt-p70S6k.

Aim: To determine whether fluid injection during radiofrequency ablation (RFA) can increase the coagulation area., Methods: Bovine liver (1-2 kg) was placed on an aluminum tray with a return electrode affixed to the base, and the liver was punctured by an expandable electrode. During RFA, 5% glucose; 50% glucose; or saline fluid was infused continuously at a rate of 1.0 mL/min through the infusion line connected to the infusion port. The area and volume of the thermocoagulated region of bovine liver were determined after RFA. The Joule heat generated was determined from the temporal change in output during the RFA experiment., Results: No liquid infusion was 17.3 ± 1.6 mL, similar to the volume of a 3-cm diameter sphere (14.1 mL). Mean thermocoagulated volume was significantly larger with continuous infusion of saline (29.3 ± 3.3 mL) than with 5% glucose (21.4 ± 2.2 mL), 50% glucose (16.5 ± 0.9 mL) or no liquid infusion (17.3 ± 1.6 mL). The ablated volume for RFA with saline was approximately 1.7-times greater than for RFA with no liquid infusion, representing a significant difference between these two conditions. Total Joule heat generated during RFA was highest with saline, and lowest with 50% glucose., Conclusion: RFA with continuous saline infusion achieves a large ablation zone, and may help inhibit local recurrence by obtaining sufficient ablation margins. RFA during continuous saline infusion can extend ablation margins, and may be prevent local recurrence.

Patients with liver disease may be at risk of zinc depletion. Zinc supplementation has been shown to contribute to inhibition of liver fibrosis and improvement in hepatic encephalopathy. However, little is known about the anti-inflammatory effect of zinc on hepatitis C virus (HCV)-related chronic liver disease. The standard of care for chronic HCV has improved markedly since the approval of interferon (IFN) therapy more than a decade ago. Over the past 20 years, IFN therapy has improved to more effectively eliminate the virus, progressing from single IFN therapy to combination therapy with ribavirin (RBV) and finally to pegylated IFN (PEG-IFN) therapy. However, even combined therapy with PEG-IFN and RBV for 48 wk is unable to eliminate the virus in some 40% of hepatitis C cases, particularly those with genotype 1b and high viral load. Treatment options for patients who have relapsed or are refractory to treatment with PEG-IFN and RBV therefore need to be critically assessed. This paper overviews the relationship between chronic liver disease and zinc metabolism.

A method for the generation of human induced pluripotent stem (iPS) cells was established. This method employs adenovirus carrying the ecotropic retrovirus receptor mCAT1 and Moloney murine leukemia virus (MMLV)-based retroviral vectors carrying the four transcription factors POU5F1 (OCT3/4), KLF4, SOX2, and MYC (c-Myc) (Masaki H & Ishikawa T Stem Cell Res 1:105-15, 2007). The differentiation of human iPS cells into hepatic cells was performed by a stepwise protocol (Song Z et al. Cell Res 19:1233-42, 2009). These cells have potential as patient-specific in vitro models for studying disease etiology and could be used in drug discovery programs tailored to deal with genetic variations in drug efficacy and toxicity.

Background: Previous work has established that HGF/c-Met signaling plays a pivotal role in regulating the onset of S phase following partial hepatectomy (PH). In this study, we used Met(fl/fl);Alb-Cre(+/-) conditional knockout mice to determine the effects of c-Met dysfunction in hepatocytes on kinetics of liver regeneration., Methodology/principal Finding: The priming events appeared to be intact in Met(fl/fl);Alb-Cre(+/-) livers. Up-regulation of stress response (MAFK, IKBZ, SOCS3) and early growth response (c-Myc, c-Jun, c-Fos, DUSP1 and 6) genes as assessed by RT-qPCR and/or microarray profiling was unchanged. This was consistent with an early induction of MAPK/Erk and STAT3. However, after a successful completion of the first round of DNA replication, c-Met deficient hepatocytes were blocked in early/mid G2 phase as shown by staining with phosphorylated form of histone H3. Furthermore, loss of c-Met in hepatocytes diminished the subsequent G1/S progression and delayed liver recovery after partial hepatectomy. Upstream signaling pathways involved in the blockage of G2/M transition included lack of persistent Erk1/2 activation and inability to up-regulate the levels of Cdk1, Plk1, Aurora A and B, and Mad2 along with a defective histone 3 phosphorylation and lack of chromatin condensation. Continuous supplementation with EGF in vitro increased proliferation of Met(fl/fl);Alb-Cre(+/-) primary hepatocytes and partially restored expression levels of mitotic cell cycle regulators albeit to a lesser degree as compared to control cultures., Conclusion/significance: In conclusion, our results assign a novel non-redundant function for HGF/c-Met signaling in regulation of G2/M gene expression program via maintaining a persistent Erk1/2 activation throughout liver regeneration.

Valpha14 natural killer T (Valpha14 NKT) cells activated by alpha-galactosylceramide (alpha-GalCer) secrete a large amount of Th1 and Th2 cytokines. IFN-gamma plays a crucial role in the inflammation response, and is also known as an activator of nitric oxide (NO) production. We previously reported that lipopolysaccharide (LPS)-induced NO production is augmented by alpha-GalCer in mouse peritoneal cells. Since the liver is susceptible to LPS stimulation via the portal vein, we examined the effect of alpha-GalCer on LPS-induced NO production in murine intra-hepatic lymphocytes (IHLs). Although IHLs augmented LPS-induced NO production by alpha-GalCer administration, such an augmentation was not observed in non-treated mice. Furthermore, alpha-GalCer did not augment LPS-induced NO production in IHLs from IFN-gamma knockout mice. In flow cytometry analysis of IHLs from alpha-GalCer-treated mice, the ratio and number of F4/80- and TLR4-positive cells rose as compared with non-treated mice. The liver injury may be induced by LPS and NO under the condition where Valpha14 NKT cells were activated.

Liver-infiltrating T cells play an essential role in the immunopathogenesis of autoimmune liver disease. Programmed death-1 (PD-1) and its ligands, B7-H1/PD-L1 and B7-DC/PD-L2, are new CD28-B7 family members that are involved in the regulation of immune responses. The ligation of PD-1 inhibits T-cell receptor-mediated T cell proliferation and cytokine production, and PD-1-deficient mice develop various organ-specific autoimmune diseases. To investigate the expressions of PD-1 and its ligands in autoimmune liver disease, in particular autoimmune hepatitis (AIH) and primary biliary cirrhosis (PBC), immunohistochemical analysis was performed. Liver biopsy specimens obtained from 17 patients with AIH and PBC were studied. PD-1 was expressed on more than half of the liver-infiltrating T cells within the portal tract. Some of the intrahepatic T cells expressed B7-H1 in patients with AIH and PBC. B7-H1 and B7-DC were mainly expressed on some Kupffer cells (KC) and liver sinusoidal endothelial cells (LSEC) within the sinusoids and their expression was upregulated in autoimmune liver disease. These results suggest that the interaction of PD-1 on T cells with increased expression of B7-H1 and B7-DC on KC and LSEC might be involved in the downregulation of autoreactive lymphocytes and result in the regulation of pathogenesis in autoimmune liver disease.

We previously reported that fibroblast growth factor 2 (FGF2) facilitated the differentiation of transplanted bone marrow cells (BMCs) into hepatocytes. Our earlier study also demonstrated that administration of FGF2 in combination with bone marrow transplantation (BMT) synergistically activated tumor necrosis factor-alpha signaling and significantly improved liver function and prognosis more than BMT alone. However, the way that it affected the extracellular matrix remained unclear. Here, we investigated the effect of FGF2 treatment together with BMT on liver fibrosis in mice treated with carbon tetrachloride (CCl(4)). Transplantation of BMCs and concurrent treatment with FGF2 caused a statistically significant reduction in CCl(4)-induced liver fibrosis that was accompanied by strong expression of matrix metalloproteinase 9 as compared with FGF2-only treatment or BMT alone. Moreover, in this process, the proliferation of bone-marrow-derived cells was accelerated without causing apoptosis. Thus, the administration of FGF2 in combination with BMT synergistically improves CCl(4)-induced liver fibrosis in mice. This treatment has the potential of being an effective therapy for patients with liver cirrhosis.

Aim: To evaluate sample adequacy, safety, and needle passes of a new biopsy needle device compared to the Quick-Core biopsy needle for transjugular liver biopsy in patients affected by liver disease., Methods: Thirty consecutive liver-disease patients who had major coagulation abnormalities and/or relevant ascites underwent transjugular liver biopsy using either a new needle device (18 patients) or the Quick-Core biopsy needle (12 patients). The length of the specimens was measured before fixation. A pathologist reviewed the histological slides for sample adequacy and pathologic diagnoses. The two methods'specimen adequacy and complication rates were assessed., Results: Liver biopsies were technically successful in all 30 (100%) patients, with diagnostic histological core specimens obtained in 30 of 30 (100%) patients, for an overall success rate of 100%. With the new device, 18 specimens were obtained, with an average of 1.1 passes per patient. Using the Quick-Core biopsy needle, 12 specimens were obtained, with an average of 1.8 passes per patient. Specimen length was significantly longer with the new needle device than with the Quick-Core biopsy needle (P < 0.05). The biopsy tissue was not fragmented in any of the specimens with the new aspiration needle device, but tissue was fragmented in 3 of 12 (25.0%) specimens obtained using the Quick-Core biopsy needle. Complications included cardiac arrhythmia in 3 (10.0%) patients, and transient abdominal pain in 4 (13.3%) patients. There were no cases of subcapsular hematoma, hemoperitoneum, or sepsis, and there was no death secondary to the procedure. In particular, no early or delayed major procedure-related complications were observed in any patient., Conclusion: Transjugular liver biopsy is a safe and effective procedure, and there was significant difference in the adequacy of the specimens obtained using the new needle device compared to the Quick-Core biopsy needle. Using the new biopsy needle device, the specimens showed no tissue fragmentation and no increment in major procedure-related complications was observed.

Metabolomics is an emerging tool that can be used to gain insights into cellular and physiological responses. Here we present a metabolome differential display method based on capillary electrophoresis time-of-flight mass spectrometry to profile liver metabolites following acetaminophen-induced hepatotoxicity. We globally detected 1,859 peaks in mouse liver extracts and highlighted multiple changes in metabolite levels, including an activation of the ophthalmate biosynthesis pathway. We confirmed that ophthalmate was synthesized from 2-aminobutyrate through consecutive reactions with gamma-glutamylcysteine and glutathione synthetase. Changes in ophthalmate level in mouse serum and liver extracts were closely correlated and ophthalmate levels increased significantly in conjunction with glutathione consumption. Overall, our results provide a broad picture of hepatic metabolite changes following acetaminophen treatment. In addition, we specifically found that serum ophthalmate is a sensitive indicator of hepatic GSH depletion, and may be a new biomarker for oxidative stress. Our method can thus pinpoint specific metabolite changes and provide insights into the perturbation of metabolic pathways on a large scale and serve as a powerful new tool for discovering low molecular weight biomarkers.

Prostaglandin E1 (PGE1) has several potential therapeutic effects, including cytoprotection, vasodilation, and inhibition of platelet aggregation. This study investigates the protective action of PGE1 against hepatic ischemia/reperfusion injury in vivo using a complementary DNA microarray. PGE1 or saline was continuously administered intravenously to mice in which the left lobe of the liver was made ischemic for 30 minutes and then reperfused. Livers were harvested 0, 10, and 30 minutes postreperfusion. Messenger RNA was extracted, and the samples were labeled with two different fluorescent dyes and hybridized to the RIKEN set of 18,816 full-length enriched mouse complementary DNA microarrays. Serum alanine aminotransferase and aspartate aminotransferase levels at 180 minutes postreperfusion were significantly lower in the PGE1-treated group than in the saline-treated group. The cDNA microarray analysis revealed that the genes encoding heat-shock protein (HSP) 70, glucose-regulated protein 78, HSP86, and glutathione S-transferase were upregulated at the end of the ischemic period (0 minutes postreperfusion) in the PGE1 group. Our results suggested that PGE1 induces HSPs immediately after ischemia reperfusion. HSPs might therefore play an important role in the protective effects of PGE1 against ischemia/reperfusion injury of the liver.

We report a case of cholecystolithiasis with biliobiliary fistulas from gallbladder to hepatic ducts, which were manifested by worsening liver dysfunction. Although it was not diagnosed preoperatively, it was successfully treated by cholecystectomy with closure of fistulas by the gallbladder wall. This case suggests that an internal biliary fistula may be possible, when the gallbladder wall is thickened and shrunken in the case of cholecystolithiasis, accompanied with liver dysfunction despite no dilatation of the common bile duct.

The mechanism that regulates the plasticity of bone marrow cells (BMCs) into hepatocytes is poorly understood. We developed a green fluorescent protein/carbon tetrachloride model to find that BMC transplantation recovered liver damage. Serum albumin level and liver fibrosis were recovered by BMC transplantation. To understand the mechanism, we used DNA-chip technology to profile the change of transient gene expression before and after BMC transplantation. On the basis of gene expression with self-organizing map using specific equation, genes were classified into 153 clusters. The information is useful to understand the dramatic gene activation during the process of the plasticity of BMC.

Congenital absence of the portal vein is an extremely rare anomaly, in which enteric blood bypasses the liver and drains into the inferior vena cava. A 16-year-old girl was referred to our hospital presenting with liver tumor. Although she had suffered from galactosemia soon after birth, the galactosemia had improved spontaneously 1 year later. Between the ages of 8 and 12 years, chronic hepatitis with a mild elevation of aspartate transaminase (AST) and alanine transaminase (ALT) was observed, but liver tumor had not been detected on computed tomography (CT) in regular medical examinations. However, at age 16, liver tumors, 10 cm in diameter, were found. Abdominal angiography indicated complete absence of the portal vein, suggesting that enteric blood was bypassing the liver and draining into the inferior vena cava. In biopsy specimens obtained under ultrasonographic guidance, liver tumors were confirmed histologically as hyperplastic nodules. In addition to this case report, the clinical features of 25 reported cases of congenital absence of the portal vein are reviewed.

In the present study, we have cloned the cDNA of ABCC13, a novel ABC transporter, from the cDNA library of adult human placenta. The ABCC13 gene spans approximately 70kb on human chromosome 21q11.2 and consists of 14 exons. The open reading frame of the ABCC13 cDNA encodes a peptide consisting of 325 amino acid residues. The amino acid sequence corresponding to putative membrane-spanning domains was remarkably similar to ABCC1, ABCC2, ABCC3, and ABCC6. The ABCC13 gene was expressed in the fetal liver at the highest level among the organs studied. While ABCC13 was expressed in the bone marrow, its expression in peripheral blood leukocytes of adult humans was much lower and no detectable levels were observed in differentiated hematopoietic cells. The expression of ABCC13 in K562 cells decreased during cell differentiation induced by TPA. These results suggest that the expression of human ABCC13 is related with hematopoiesis.

The role of NF-kappaB, the rapid-response transcription factor for multiple genes, in cold ischemia-reperfusion (I/R) injury was examined after syngeneic transplantation of liver grafts. Lewis rat recipients were killed 1-48 h after reperfusion of three different liver grafts: 1) uninfected control, 2) infected ex vivo with control adenoviral vector (AdEGFP), and 3) infected ex vivo with AdIkappaB. In uninfected control livers, NF-kappaB was activated biphasically at 1-3 and 12 h after reperfusion with aspartate transaminase (AST) levels of 4,244 +/- 691 IU/l. The first peak of NF-kappaB activation associated with an increase of mRNA for TNF-alpha, IL-1beta, and IL-10. AdEGFP transfection resulted in similar outcomes. Interestingly, AdIkappaB-transfected liver grafts suffered more severe I/R injury (AST >9,000 IU/l). Transfected IkappaB was detected in transplanted livers as early as 6 h, and this correlated with the abrogation of the second, but not the first, peak of NF-kappaB activation at 12-48 h and increased apoptosis. Thus inhibition of the second wave of NF-kappaB activation in IkappaB-transfected livers resulted in an increase of liver injury, suggesting that NF-kappaB may have a dual role during liver I/R injury.

Background: OK-432 is known to increase the host antitumor response. We previously reported that systemic administration of OK-432 (OK-Lipo) specifically induced hepatic lymphocytes in mice. Here we aimed to investigate the antitumor effect of OK-Lipo on hepatocellular carcinomas (HCC) in experimental rats., Methods: Diethylnitrosamine was administered for 12 weeks to all rats (n = 36). Rats were divided into three groups of 12 rats each. One group was injected with OK-Lipo from week 5 (OK-5w group) and another from week 9 (OK-9w group). A control group was injected with saline from week 5 (Non-OK group). At week 13, five rats from each group were used for histological analysis and immunofluorescence assays (surface phenotypic and intracellular cytokine analysis of the mononuclear cells in the liver, spleen and peripheral blood). The remaining rats were observed for the remainder of their survival period., Results: The mean survival times of Non-OK, OK-5w, and OK-9w groups differed significantly (98.0 +/- 5.3 days, 116.0 +/- 5.8 days, and 106.0 +/- 5.4 days, respectively, P < 0.01). Histological examination revealed many apoptotic tumor cells, infiltration of lymphocytes and macrophages in the OK-5w group. The two-color immunofluorescence assay showed that the proportion of natural killer (NK) cells and IFN-gamma-producing cells in the liver were significantly higher in the OK-5w group., Conclusions: These findings showed that systemic administration of OK-Lipo contributed to prolonging the survival of rats with HCC. OK-Lipo induced NK cells and IFN-gamma-producing cells specifically in the liver and these cells seemed to reduce hepatocarcinogenesis and tumor growth.

We developed a new high-performance liquid chromatographic method using an ODS column and a chiral column for the assay of racemic 4-OH-PL sulfate and enantiomeric 4-OH-PL sulfates, respectively. The method was successfully applied to measure phenolsulfotransferase (PST) activities for 4-OH-PL in cytosolic fractions from livers of Japanese monkeys (Macaca fuscata) and for comparison with its activity of cytosolic fractions from rat, rabbit, dog, and human livers and Hep G2 cells. The activity was ranked as Hep G2 cells > monkeys = humans = dogs = rats > rabbits. To evaluate the Japanese monkey as a nonhuman animal model in drug metabolism studies, we further characterized sulfation of 4-OH-PL as a further metabolic pathway in monkey livers to compare that with human livers. Inhibition studies in which cytosolic fractions were preincubated at 43 degrees C or 2,6-dichloro-4-nitrophenol (DCNP) used as a PST inhibitor indicated that two kinds of PSTs, thermolabile, low-Km and DCNP-resistant PST and thermostable, high-Km and DCNP-sensitive PST were involved in 4-OH-PL sulfation by monkey liver cytosol, which is very similar to the reported profile of 4-OH-PL sulfation by human liver cytosol. Sulfation kinetics in a low concentration range of 4-OH-PL enantiomers demonstrated that apparent Km values were similar between human and monkey liver cytosolic fractions, but the Vmax values were different, so that intrinsic clearance values (Vmax/Km, Clint) were higher in monkeys than in humans. Furthermore, enantiomer selectivity of [R(+)-4-OH-PL > S(-)-4-OH-PL] was observed in the Vmax and CLint values of monkey liver cytosol. These results indicate that the profile of sulfation of 4-OH-PL by liver cytosolic fractions is similar in humans and Japanese monkeys.

The three-dimensional (3D) relationship among the hepatic domains and the efferent central and afferent portal veins was investigated by macroscopy, microscopy, and computer-aided 3D reconstruction methods. To clearly distinguish the pericentral domain from the periportal, we used CCl(4)-treated mice and diabetic house musk shrews, which show typical pericentral necrosis and deposition of fat, respectively. The 3D findings obtained were verified against normal control animals using advantages of our unique observations by light and fluorescent microscopy, which made it possible to differentiate the two domains well. The pericentral domains in the mice and shrews appeared three-dimensionally as continuous branched columns, and the periportal domains exist in a sponge-like network that fills the parenchymal space among the columnar pericentral domains. The efferent central veins were concentrically surrounded by the pericentral domain, and segments of the central veins flowed into large sublobular and lobar veins. The walls of these large veins faced the pericentral domain at the confluence with the central veins; the remaining portions of the walls faced the periportal domain. The afferent portal veins were placed at the two-dimensional center of the network of the periportal domain and gave off smaller portal branches radially at the intersections of the network. Three types of liver lobules-classic, portal, and acinar-have been discussed repeatedly at the (2D) level. At the 3D level, it is reasonable to consider that the liver parenchyma consists of the two continuous domains corresponding to the distribution of the vessels that we found., (Copyright 2000 Wiley-Liss, Inc.)

We found previously that expression of multidrug resistance-associated protein (MRP) 3 is induced in a mutant rat strain (Eisai hyperbilirubinemic rats) whose canalicular multispecific organic anion transporter (cMOAT/MRP2) function is hereditarily defective and in normal Sprague-Dawley (SD) rats after ligation of the common bile duct. In the present study, the inducible nature of MRP3 was examined, using Northern and Western blot analyses, in comparison with that of other secondary active [Na(+)-taurocholic acid cotransporting polypeptide (Ntcp), organic anion transporting polypeptide 1 (oatp1), and organic cation transporter (OCT1)] and primary active [P-glycoprotein (P-gp), cMOAT/MRP2, and MRP6] transporters. alpha-Naphthylisothiocyanate treatment and common bile duct ligation induced expression of P-gp and MRP3, whereas expression of Ntcp, oatp1, and OCT1 was reduced by the same treatment. Although expression of MRP3 was also induced by administration of phenobarbital, that of cMOAT/MRP2, MRP1, and MRP6 was not affected by any of these treatments. Moreover, the mRNA level of MRP3, but not that of P-gp, was increased in SD rats after administration of bilirubin and in Gunn rats whose hepatic bilirubin concentration is elevated because of a defect in the expression of UDP-glucuronosyl transferase. However, the MRP3 protein level was not affected by bilirubin administration. Although the increased MRP3 mRNA level was associated with the increased concentration of bilirubin and/or its glucuronides in mutant rats and in SD rats that had undergone common bile duct ligation or alpha-naphthylisothiocyanate treatment, we must assume that factor(s) other than these physiological substances are also involved in the increased protein level of MRP3.

Liver fibrosis is commonly observed in chronic liver disease. However, the immunological mechanisms underlying hepatic fibrosis due to chronic inflammation are not well defined, mainly because suitable experimental models have not been established. We have found that weekly i.v. administration of concanavalin A (Con A) in BALB/c mice brought about a striking alanine aminotransferase increase, resulting in piecemeal necrosis with bridging fibrosis in the parenchyma. Using this fibrosis model, we demonstrated the kinetics of cytokine mRNA expression in liver. Transforming growth factor (TGF)-beta1, TGF-alpha, basic fibroblast growth factor (bFGF) and hepatocyte growth factor mRNAs were up-regulated after each Con A administration. Furthermore, either anti-IFN-gamma, anti-tumor necrosis factor (TNF)-alpha or anti-TGF-beta mAb given together with Con A markedly inhibited the development of hepatic fibrosis. Treatment with either anti-IFN-gamma or anti-TNF-alpha mAb also completely prevented hepatic injury; in contrast, treatment with anti-TGF-beta mAb did not. The treatment with anti-TGF-beta mAb did not affect the levels of hepatic mRNAs for either IFN-gamma or TNF-alpha after Con A injection. Treatment with either anti-IFN-gamma or anti-TNF-alpha did not affect the expression levels of TGF-beta in the liver. In conclusion, the continuous presence of both severe liver damage and up-regulation of TGF-beta synthesis is necessary to induce hepatic fibrosis in this model.

We have isolated a cDNA clone for a novel glutathione-dependent dehydroascorbate reductase from a rat liver cDNA library in lambdagt11 by immunoscreening. The authenticity of the clone was confirmed as follows: first, the antibody that had been purified through affinity for the protein expressed by the cloned lambdagt11 phage recognized only the enzyme in a crude extract from rat liver; and second, two internal amino acid sequences of purified enzyme were identified in the protein sequence predicted from the cDNA. The predicted protein consists of 213 amino acids with a molecular weight of 24,929, which is smaller by approximately 3,000 than the value obtained by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. This discrepancy of the molecular weight was explained by post-translational modification because the recombinant protein expressed by a mammalian system (Chinese hamster ovary cells) was of the same size as rat liver enzyme but larger than the protein expressed by a bacterial system (Escherichia coli). Chinese hamster ovary cells, originally devoid of glutathione-dependent dehydroascorbate reductase activity, was made to elicit the enzyme activity (1.5 nmol/min/mg of cytosolic protein) by expression of the recombinant protein. Additionally, the cells expressing the enzyme were found to accumulate 1.7 times as much ascorbate as the parental cells after incubation with dehydroascorbate. This result points to the importance of the dehydroascorbic acid reductase in maintaining a high concentration of ascorbate in the cell.

We previously reported that a host cell glycoprotein, gp180, binds duck hepatitis B virus particles, and is encoded by a member of the carboxypeptidase gene family (Kuroki, K., Eng, F., Ishikawa, T., Turck, C., Harada, F., Ganem, D., 1995. gp180, a host cell glycoprotein that binds duck hepatitis B virus particles, is encoded by a member of the carboxypeptidase gene family. J. Biol. Chem. 270, 15022-15028). After that report, carboxypeptidase D (CPD) was subsequently purified from bovine pituitary and characterized as a novel carboxypeptidase E (CPE)-like enzyme, with many characteristics in common with duck gp180 (Song, L., Fricker, L.D., 1995. Purification and characterization of carboxypeptidase D, a novel carboxypeptidase E-like enzyme, from bovine pituitary. J. Biol. Chem. 270, 25007-25013). CPD is now supposed to play an important role in a secretory pathway. To clarify the function of gp180 further, we have isolated and analyzed human and mouse homologues of duck gp180. cDNA clones derived from human HepG2 cells and mouse livers have been isolated on the basis of homology to the duck gp180. The suggested open reading frames of the human and mouse cDNA encode 1380 and 1377 amino acid proteins, respectively and have three carboxypeptidase homologous domains (A, B, and C). Domains A and B have completely conserved the residues known to have the enzymatic activity of carboxypeptidase, but domain C in each cDNA does not. Northern blotting revealed a ubiquitous tissue distribution of human gp180 mRNA with several transcript species. Expression of human gp180 cDNA in transfected 293Tcells exhibited carboxypeptidase activity upon radiometric assay. The human and mouse homologues of duck gp180 have many characteristics in common with bovine CPD. Fluorescence in-situ hybridization reveals that the gene encoding human gp180 is located in region 17q11.2.

Capecitabine (N4-pentyloxycarbonyl-5'-deoxy-5-fluorocytidine) is a novel oral fluoropyrimidine carbamate, which is converted to 5-fluorouracil (5-FU) selectively in tumours through a cascade of three enzymes. The present study investigated tissue localisation of the three enzymes in humans, which was helpful for us to design the compound. Carboxylesterase was almost exclusively located in the liver and hepatoma, but not in other tumours and normal tissue adjacent to the tumours. Cytidine (Cyd) deaminase was located in high concentrations in the liver and various types of solid tumours. Finally, thymidine phosphorylase (dThdPase) was also more concentrated in various types of tumour tissues than in normal tissues. These unique tissue localisation patterns enabled us to design capecitabine. Oral capecitabine would pass intact through the intestinal tract, but would be converted first by carboxylesterase to 5'-deoxy-5-fluorocytidine (5'-dFCyd) in the liver, then by Cyd deaminase to 5'-deoxy-5-fluorouridine (5'-dFUrd) in the liver and tumour tissues and finally by dThdPase to 5-FU in tumours. In cultures of human cancer cell lines, the highest level of cytotoxicity was shown by 5-FU itself, followed by 5'-dFUrd. Capecitabine and 5'-dFCyd had weak cytotoxic activity only at high concentrations. The cytotoxicity of the intermediate metabolites 5'-dFCyd and 5'-dFCyd was suppressed by inhibitors of Cyd deaminase and dThdPase, respectively, indicating that these metabolites become effective only after their conversion to 5-FU. Capecitabine, which is finally converted to 5-FU by dThdPase in tumours, should be much safer and more effective than 5-FU, and this was indeed the case in the HCT116 human colon cancer and the MX-1 breast cancer xenograft models.

Background & Aims: Activin A, a member of the transforming growth factor (TGF)-beta superfamily, recently has been reported to suppress DNA synthesis and to induce apoptosis of hepatocytes. These biological functions are similar to those of TGF-beta1, which is overexpressed in liver cirrhosis. The aim of this study was to examine whether activin A is involved in liver cirrhosis and fibrosis., Methods: Liver cirrhosis or fibrosis was induced by intraperitoneal injections of dimethylnitrosamine or porcine serum into rats. The kinetics of activin A messenger RNA (mRNA) expression in cirrhotic and fibrotic livers and primary cultured rat hepatocytes were assessed by Northern blotting, and the localization of activin A was determined immunohistochemically. Modulation of type 1 collagen mRNA expression by activin A in rat cultured Ito/fat-storing cells and fibroblasts was also examined., Results: Northern blotting showed that activin A mRNA expression was enhanced in fibrotic livers. The numbers of hepatocytes expressing immunoreactive activin A were significantly greater, especially around the fibrotic areas. Activin A mRNA expression in cultured hepatocytes was increased significantly by TGF-beta1 and by activin A itself. Furthermore, type 1 collagen mRNA expression in cultured cells was enhanced by activin A and TGF-beta1 in a synergistic manner., Conclusions: Activin A is overexpressed in rat cirrhotic and fibrotic livers and may contribute to hepatic fibrogenesis.

Circulating hyaluronan is mostly derived from lymph, fibroblast and Ito cells in the liver, and more than 90% of hyaluronan is degraded in hepatic sinusoidal endothelial cells. Thus, elevated serum hyaluronan is regarded as an indication of hepatic fibrosis with activated Ito cells and dysfunctional sinusoidal endothelial cells. We studied the distribution of hyaluronan in human liver sinusoids to determine the influences on elevated hyaluronan levels in sera. Histochemical examination was made using hyaluronan-binding protein (HABP) and serial sections of liver tissue for staining of alpha-smooth muscle actin (ASMA) (an indicator of activated Ito cells) and of ulex europaeus agglutinin I lectin (UEA-1) (closely related to hepatic sinusoidal capillarization). Positive staining, indicating the presence of hyaluronan, was noted in fibrous regions around the portal tracts, areas of focal necrosis in the liver parenchyma, and walls of the sinusoids in chronic hepatitis. In this group, hyaluronan-positive areas corresponded to positive ASMA staining and faint staining of UEA-1. On the contrary, in liver cirrhosis, UEA-1-positive areas were essentially identical to hyaluronan-positive areas and to ASMA-negative areas in sinusoidal walls. Hyaluronan and ASMA could be detected in the same areas of sinusoidal walls in chronic hepatitis, but not in liver cirrhosis. Hyaluronan appears to be mainly related to the staining of activated Ito cells in chronic hepatitis. Therefore, we concluded that in chronic hepatitis, the production of hyaluronan was accelerated in Ito cells; however, degradation of hyaluronan by sinusoidal endothelial cells continued. On the contrary, in liver cirrhosis, hyaluronan production decreased in Ito cells, and a marked transformation of sinusoidal endothelial cells with hepatic sinusoidal capillarization indicated loss of the ability to degrade hyaluronan. These different mechanisms in chronic hepatitis and liver cirrhosis may operate in the sinusoidal walls and may cause the elevation of hyaluronan in sera.

In the present study, we investigated the role of the spleen in experimental hepatic ischemia/reperfusion in the rat. After a 90-min period of ischemia in the left and middle hepatic lobes, the ischemia was released and the liver was reperfused for up to 24 h. Plasma alanine aminotransferase reached a peak 3 h after the onset of reperfusion, and gradually decreased thereafter. A histological examination revealed evidence of hepatocellular necrosis and degeneration, especially 24 h after the onset of reperfusion. In addition, there was a noticeable accumulation of polymorphonuclear cells in the liver following ischemia/reperfusion. A splenectomy performed just prior to ischemia/reperfusion reduced both biochemical and histological hepatocellular injury. The number of polymorphonuclear cells in the liver following ischemia/reperfusion was significantly reduced in rats subjected to splenectomy, suggesting that the increase in polymorphonuclear cells may contribute to liver injury. The number of mononuclear cells also increased in the marginal zones of the spleen following ischemia/reperfusion, and appeared to be derived from the splenic monocyte/macrophage population, based on immunohistochemical studies. The spleen plays an important role in the pathogenesis of hepatic ischemia/reperfusion injury and the splenic monocyte/ macrophage population contributes to liver damage.

It is widely believed that viral clearance is mediated principally by the destruction of infected cells by CTLs. In this report, we use a transgenic mouse model of HBV replication to demonstrate that this assumption may not be true for all viruses. We find that adoptively transferred virus-specific CTLs can abolish HBV gene expression and replication in the liver without killing the hepatocytes. This antiviral function is mediated by IFN gamma and TNF alpha secreted by the CTL or by the antigen-nonspecific macrophages and T cells that they activate following antigen recognition. These cytokines activate two independent virocidal pathways: the first pathway eliminates HBV nucleocapsid particles and their cargo of replicating viral genomes, while the second pathway destabilizes the viral RNA. Intracellular viral inactivation mechanisms such as these could greatly amplify the protective effects of the immune response, while failure of such mechanisms could lead to viral persistence or to the death of the host.