Role of peroxisome proliferator-activated receptor-gamma (PPARgamma) during liver regeneration in rats

Background and Aim:  Peroxisome proliferator-activated receptor-gamma (PPARγ), a member of the nuclear receptor superfamily, is widely expressed in adipocytes and other tissues, including the liver. Several reports have shown that PPARγ activation induced cell-cycle arrest and apoptosis in tumor cells. We investigated the role of the PPARγ/ligand system and the effect of the PPARγ agonist during liver regeneration.
Methods:  Expression of PPARγ and serum levels of 15-deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2) by enzyme immunoassay were evaluated in rats following partial hepatectomy (PH group). Further, the effect of the PPARγ agonist, pioglitazone, on liver regeneration (PH + PGZ group) was evaluated by proliferating cell nuclear antigen labeling index, relative liver weight, and expression of cell-cycle regulators.
Results:  The number of PPARγ-stained hepatocytes decreased at 24 h (PH, 15.8 ± 2.2%; sham, 35.5 ± 2.4%; P  < 0.001) and increased in the late phase of liver regeneration compared to the sham-operated group ( P  < 0.001 at 48–120 h). The peaks of serum 15d-PGJ2 (627.0 ± 91.1 pg/ml) and PPARγ expression (90.6 ± 3.1%) coincided in the late phase of liver regeneration. Also, oral administration of pioglitazone inhibited hepatocyte proliferation, in terms of the proliferating cell nuclear antigen (PCNA) labeling index and p27 expression during the late phase of liver regeneration, and caused a transient reduction in liver mass when compared to the PH group.
Conclusions:  These results indicate that the PPARγ/ligand system may be one of the key negative regulators of hepatocyte proliferation and may be responsible for the inhibition of liver growth in the late phase of liver regeneration.     

Vascular endothelial growth factor does not improve liver regeneration and survival after 90% subtotal liver resection

Aim: Vascular endothelial growth factor (VEGF) has been shown to stimulate liver regeneration after 70% partial hepatectomy (PH). It is unclear, however, whether exogenous administration of VEGF can also be used to improve liver regeneration and survival after 90% subtotal liver resection. The aim of this study was to determine the effect of exogenous and endogenous VEGF after 90% subtotal hepatectomy (SH). Methods: Rats were subjected to 90% SH and treated with VEGF, anti-VEGF or NaCl. Postoperatively (3 h - 5 days) liver body weight ratio (LBR), hepatocyte proliferation and biochemical markers were assessed. ELISA was performed to measure protein levels for VEGF. Gene expression was determined by customized cDNA arrays and quantitative RT-PCR. Results: Administration of VEGF did not enhance LBR or hepatic proliferation, or reduce the serum parameters. VEGF levels were the highest in VEGF-treated animals. The overall survival after 90% SH reached 78% in VEGF-treated animals, but did not differ significantly from that of anti-VEGF or NaCl-treated animals (74% and 75%, respectively). Gene expression analysis showed a modulation of anti-apoptotic and cell cycle control genes that was independent of VEGF. Conclusions: In contrast to PH, liver regeneration and survival after SH cannot be modulated by VEGF. This indicates that the relevant mechanisms that stimulate liver regeneration after hepatectomy at least partially depend upon the extent of liver resection.     

Erythropoietin promotes hepatic regeneration after extended liver resection in rats

Background and Aim:  It has been proven in various animal studies that recombinant human erythropoietin (rHuEPO) protects renal, cardiac and neuronal, as well as hepatic, tissue from ischemia, and promotes regeneration of adult central nervous system neurons. To date, no data are available as to whether rHuEPO has the ability to stimulate liver regeneration after liver resection.
Methods:  Rats undergoing 70% or 90% hepatectomy received an intraportalvenous administration (i.p.) of rHuEPO prior to resection or a subcutaneous injection (s.c.) for 3 days postoperatively, control animals were treated with surgery and saline injection only. Regeneration capacity of remnant livers was studied over 7 days by histology and immunohistochemistry (Ki-67, proliferating cell nuclear antigen [PCNA]). Polymerase chain reaction was carried out to measure transforming growth factor β (TGF-β), hypoxia induced factor (HIF), signal transducing activator 3 and vascular endothelial growth factor.
Results:  Ten-day survival in rats undergoing 90% hepatectomy significantly increased in i.p.-pretreated animals. After 70% hepatectomy the mitotic index was significantly increased in both rHuEPO-treated groups. These data were confirmed by PCNA and Ki-67 expression, which was significantly increased in the treated groups 24 h and 2 days after liver resection. TGF-β and HIF mRNA both were upregulated in control animals 3 h after surgery.
Conclusion:  rHuEPO effectively increased liver regeneration in rats after 70% liver resection and enhanced survival after 90% hepatectomy. Thus, rHuEPO may increase the regenerative capacity after major hepatectomy.     

Beneficial effects of splenectomy on massive hepatectomy model in rats

Aim:  Possible spleno-hepatic relationships during hepatectomy remain unclear. The purpose of this study was to investigate the impact of splenectomy during massive hepatectomy in rats.
Methods:  Rats were divided into the following two groups: 90% hepatectomy (Hx group), hepatectomy with splenectomy (Hx+Sp group). The following parameters were evaluated; survival rate, biochemical parameters, quantitative RT-PCR for hemeoxygenase-1 (HO-1) and tumor necrosing factor α (TNFα), immunohistochemical staining for HO-1, proliferating cell nuclear antigen labeling index and liver weights.
Results:  The survival rate after massive hepatectomy significantly improved in Hx+Sp group as well as serum biochemical parameters, compared with Hx group ( P  < 0.05). HO-1 positive hepatocytes and its mRNA expression significantly increased and TNFα mRNA expression significantly decreased in Hx+Sp group compared with Hx group ( P  < 0.05). Moreover, liver regeneration was significantly accelerated at 48 and 72 h after hepatectomy in Hx+Sp group.
Conclusions:  Splenectomy had beneficial effects on massive hepatectomy by ameliorating liver injuries and promoting preferable liver regeneration.     

High expression of angiotensin-converting enzyme and angiotensin-converting enzyme 2 in preservation injury after liver transplantation in rats

Aim:  To explore the possible role of local renin-angiotensin system (RAS) in preservation injury (PI) after liver transplantation by studying expression of angiotensin-converting enzyme (ACE) and angiotensin-converting enzyme 2 (ACE2) in the transplanted liver of rats and its relationship with PI.
Methods:  The animals receiving liver transplantation were assigned to cold preservation group (CP group) and non-cold preservation group (NCP group). The sham-operation group was used as the control. The severity of PI was assessed by histology. The mRNA and protein expressions of ACE and ACE2 were detected by real-time PCR, Western blot, respectively. Tissue hypoxia was assessed by pimonidazole staining.
Results:  Various degrees of tissue injury were observed after liver transplantation, especially in CP group. ACE2 mRNA and protein expressions in the transplantation groups were elevated significantly compared with those of the control group ( P  < 0.01, P  < 0.05), and higher in CP group than those in NCP group ( P  < 0.05). There was a close positive correlation between PI and mRNA expression of ACE and ACE2. Positive pimonidazole staining distributed around the hepatic central vein, and became darker and more extensive with deterioration of PI.
Conclusion:  ACE2 was closely related to tissue hypoxia due to CP-induced PI of the transplanted liver, and ACE may aggravate the inflammation in PI. Local RAS may play an important role in PI of the transplanted liver.     

Hepatoma-derived growth factor is induced in liver regeneration

Aim:  Hepatoma-derived growth factor (HDGF) is a heparin-binding protein, which has been suggested to be involved in the development of kidneys, the cardiovascular system and the liver. We have shown that HDGF is highly expressed in parenchymal hepatocytes in the developing liver and promotes fetal hepatocyte proliferation. In the present study, we asked whether HDGF expression was related to liver regeneration.
Methods:  We examined the mRNA and protein expressions of HDGF in two liver regeneration models. In addition, cellular distribution of HDGF in the regenerating liver was investigated by immunohistochemistry.
Results:  In the carbon tetrachloride (CCl₄)-treated liver, HDGF expression was induced and the peak was detected at 24 h after the CCl₄ injection. HDGF expression was also enhanced in the hepatectomy model and the peak was detected at 12 h after surgery. The increased expression of HDGF protein was also confirmed by western blotting. Expression of the HDGF gene in the regenerating liver was dominantly detected in parenchymal hepatocytes.
Conclusion:  These findings showed that HDGF expression was induced in parenchymal hepatocytes before the DNA synthesis in the regenerating liver, suggesting the possible involvement of HDGF in liver regeneration as an autocrine factor.     

Comparison of p.o. or i.v. proton pump inhibitors on 72-h intragastric pH in bleeding peptic ulcer

Background and Aims:  After successful endoscopic hemostasis in bleeding peptic ulcer, addition of proton pump inhibitors reduce the rate of recurrent bleeding by maintaining intragastric pH at neutral level. The aim of the present study was to evaluate the effect of various proton pump inhibitors given through different routes on intragastric pH over 72 h after endoscopic hemostasis in bleeding peptic ulcer.
Methods:  Ninety consecutive patients who had successful endoscopic therapy of bleeding peptic ulcer underwent 72-h continuous ambulatory intragastric pH study, were randomly assigned to receive p.o. omeprazole 80 mg bolus followed by 40 mg every 12 h for 72 h or i.v. 80 mg omeprazole followed by infusion 8 mg/h for 72 h. Oral pantoprazole 80 mg bolus followed by 80 mg every 12 h for 72 h or i.v. 80 mg pantoprazole followed by infusion of 8 mg/h for 72 h. Oral rabeprazole 80 mg bolus followed by 40 mg every 12 h for 72 h or i.v. 80 mg rabeprazole followed by infusion 8 mg/h for 72 h. Five patients received no treatment after successful endoscopic therapy and underwent 72-h pH study.
Results:  Mean 72-h intragastric pH for p.o. omeprazole was 6.56 versus 6.93 for omeprazole infusion ( P  = 0.48). Mean 72-h intragastric pH for p.o. pantoprazole was 6.34 versus 6.32 for pantoprazole infusion ( P  = 0.62). Mean 72-h intragastric pH for rabeprazole p.o. was 6.11 versus 6.18 rabeprazole i.v. ( P  = 0.55). Mean 72-h pH for the no proton pump inhibitor group was 2.04.
Conclusion:  There was no significant difference among various proton pump inhibitors given through different routes on raising intragastric pH above 6 for 72 h after successful endoscopic hemostasis in bleeding peptic ulcer.     

Effect of granulocyte-macrophage colony-stimulating factor on hepatic regeneration after 70% hepatectomy in normal and cirrhotic rats

BackgroundPost-hepatectomy liver insufficiency is one of the most serious postoperative problems and its prevention is important after major hepatic resection, especially in the cirrhotic liver. Some growth factors and cytokines appear to play important roles in liver regeneration. In the present study we have investigated the effects of granulocyte-macrophage colony-stimulating factor (GM-CSF) on hepatic regeneration after 70% partial hepatectomy (PH) in cirrhotic and non-cirrhotic rats.MethodsA rat model of liver cirrhosis was prepared using thioacetamide (TAA) (a dose of 20 mg/100 g body w, intra-peritoneally) on three days a week for 12 weeks. Adult male rats were divided into four groups:Group 1 (n=10) no cirrhosis and no GM-CSF; Group 2 (n=10) no cirrhosis and GM-CSF; Group 3 (n=10) cirrhosis and no GM-CSF; and Group 4 (n=10) cirrhosis and GM-CSF. All the rats underwent a 70% hepatectomy, and GM-CSF was administrated immediately after operation in Groups 2 and 4. On postoperative days 2 and 7, fresh samples from the remnant liver were obtained to evaluate its regenerative capacity.The liver regenerative process was estimated by DNA synthesis, using flow cytometry.ResultsProliferation index (PI) of hepatocytes at 48 h was higher in Group 4 rats than Group 3 rats (p<0.05). On postoperative day 7, PI was elevated in Group 3 rats compared with Group 4 rats, but this difference was not statistically significant. In non-cirrhotic rats given GM-CSF, PI was increased compared with Group 1 rats at day 2 (p<0.05), but not at day 7.ConclusionsThe findings suggest that the proliferative capacity of liver cells is impaired and delayed after 70% PH in cirrhotic rat liver. GM-CSF administration might enhance the liver PI in both normal and TAA-induced cirrhotic rats.     

5-羟色胺2A受体阻断剂对大鼠肝脏再生的影响

目的观察5-羟色胺2A受体阻断剂酮色林对大鼠肝部分切除后肝再生的影响,了解5-羟色胺及其受体在肝脏再生中的作用。方法 80只雄性Wistar大鼠随机分为实验组和对照组。采用肝大部分切除术建立肝再生模型,术后16 h分别给予腹腔内注射酮色林（实验组）和生理盐水（对照组）,采用免疫组化及流式细胞技术动态观察并比较两组大鼠术后24、36、48、72 h肝脏Ki67、增殖细胞核抗原的表达情况。结果大鼠肝大部切除术后24、36 h肝脏表达Ki67、增殖细胞核抗原最为活跃,而后表达逐渐下降。实验组大鼠肝脏表达Ki67、增殖细胞核抗原较对照组显著下降（P〈0.05）。结论 5-羟色胺2A受体阻断剂酮色林显著抑制大鼠肝大部切除术后的肝脏再生,说明5-羟色胺具有一定的促进肝再生的作用,2A受体是其重要的信号传导受体之一。     

Regeneration of liver with marked fatty change following partial hepatectomy in rats

Resection of liver for primary and metastatic tumors and living donor liver transplantation has become a common clinical practice. The success of recovery depends on the regeneration and functions of the remnant liver. However, information on the regenerative potential of liver with steatosis and steatohepatitis, a common clinical problem in this country, is incomplete. Therefore, we evaluated regeneration after two-thirds partial hepatectomy (PH) in male F-344 rats with marked steatosis and mild steatohepatitis induced by feeding choline-deficient diet. Choline-deficient rats and control rats were killed at 24, 36, 48, 72, and 96 h after PH. Liver regeneration was determined by measuring mitotic activity and bromodeoxyuridine (BrdU) labeling in hepatocytes. Livers of rats maintained on the choline-deficient diet showed marked steatosis and mild steatohepatitis. In these animals the levels of serum and liver triacylglycerols (TG) were low and very high, respectively, when compared to controls. In control rats mitotic and BrdU labeling indices were maximal at 24 h followed by a rapid decline, whereas in choline-deficient rats both these indices increased significantly at 36 h and decreased gradually over the next 60 h. By 96 h the size of livers in both groups was comparable. The results of this study indicate that regeneration in the liver of rats with marked steatosis is not impaired.     

肝大部切除大鼠所致的IETM对血浆与肝组织IL-1β的影响

目的 探讨肝大部分切除(PH)诱导的肠源性内毒素血症(IETM)对血浆及残余肝组织IL-1β的影响或可能存在的关系.方法 102只健康雄性成年wistar大鼠随机分为正常对照(NC)组、假手术(SO)组和肝大部切除(PH)组,在不同时间点检测血浆内毒素(ET)、IL-1β、ALT的水平以及残余肝组织IL-1β的蛋白表达...     

Procalcitonin is a valuable prognostic marker in ARDS caused by community-acquired pneumonia

Background and objective:   ARDS is life-threatening acute respiratory failure, and pneumonia is one of the most common causes of direct ARDS. Procalcitonin (PCT) has been evaluated for its utility in determining the aetiology of community-acquired pneumonia (CAP), choice of antibiotics and prediction of outcome. This study evaluated the role of PCT in predicting the outcome of patients with ARDS caused by severe CAP.
Methods:   This was a prospective observational study conducted from September 2002 to December 2003. The plasma PCT was analysed at baseline, 24 and 72 h after enrolment and measured by ELISA.
Results:   Of the 22 patients with ARDS caused by CAP and enrolled in the study, 17 (77.3%) were alive 14 days after admission and five (22.7%) had died. The survivors had lower APACHE II scores (22.2 ± 4.6 vs 30.6 ± 9.6, P  = 0.031), pneumonia severity index (141.9 ± 2.2 vs 195.6 ± 23.8, P  = 0.005) and lower plasma PCT at baseline (9.83 ± 3.54 vs 106.70 ± 67.86, P  = 0.004), at 24 h (10.51 ± 5.39 vs 81.32 ± 57.68, P  = 0.014) and at 72 h (2.03 ± 0.76 vs 19.57 ± 6.67, P  = 0.005).
Conclusion:   PCT analysed within 72 h of the onset of ARDS predicted mortality of patients with ARDS caused by severe CAP.     

Palliative chemotherapy for patients with recurrent hepatocellular carcinoma after liver transplantation

Background and Aim:  The majority of patients with post-transplantation recurrence of hepatocellular carcinoma (HCC) have extrahepatic metastases and multifocal lesions. Therefore, they have few treatment options and may not be amenable for local therapy. The safety and efficacy of palliative chemotherapy in this population has not been reported.
Methods:  We retrospectively analyzed 24 patients who received palliative chemotherapy for recurrent HCC after liver transplantation between January 2000 and December 2006 at the Seoul National University Hospital.
Results:  The mean age of patients was 55 years (range 42–70 years). The most commonly used chemotherapeutic regimens were 5-fluorouracil (5-FU)/cisplatin ( n  = 9), which was followed by capecitabine/cisplatin ( n  = 4), 5-FU/mitomycin ( n  = 3), 5-FU/oxaliplatin ( n  = 1), S-1 ( n  = 1), capecitabine ( n  = 1), gemcitabine/oxaliplatin ( n  = 1), gemcitabine/cisplatin ( n  = 1), 5-FU/interferon ( n  = 1) and sorafenib ( n  = 2). The Grade 3/4 hematological toxicity was neutropenia (29.1%), thrombocytopenia (20.9%) and anemia (20.9%). There were no cases of neutropenic fever or bleeding events. The Grade 3/4 non-hematological toxicity included elevation of liver transaminase (8.4%) and jaundice (16.7%). No patient showed an objective response and four patients (16.7%) demonstrated stable disease. The median time to progression was 7.0 weeks (95% CI 5.8–8.2) and the median overall survival was 16.6 weeks (95% CI 10.1–23.1).
Conclusion:  Palliative chemotherapy can be delivered to patients with recurrent HCC after liver transplantation with tolerable toxicity. However, the efficacy to date is not satisfactory. Therefore, more effective systemic chemotherapy is needed for this group of patients.     

Protective effects of a superoxide dismutase/catalase mimetic compound against paraquat pneumotoxicity in rat lung

Background and objective:   EUK-134 is one of the most promising of the superoxide dismutase (SOD)/catalase mimetic compounds. The antioxidant effects of EUK-134 were tested in a rat model of paraquat pneumotoxicity.
Methods:   Male Wistar rats ( n  = 72) were divided into three groups: group 1, controls; group 2, paraquat alone; group 3, paraquat + EUK-134. Paraquat dichloride was administered per os at a dose of 80 mg/kg. EUK-134 was injected intraperitoneally at 10 mg/kg 2 h before the paraquat and again 4 h later at 5 mg/kg.
Results:   On days 1, 3 and 5 after treatment with paraquat alone the LDH activity increased ( P  = 0.0001, P  = 0.00001 and P  = 0.03, respectively), and the total protein content increased ( P  = 0.00002, P  = 0.001 and P  = 0.01, respectively). The levels of acid phosphatase (AcP) in BAL fluid increased on days 1 and 3 ( P  = 0.006 and P  = 0.04). In lung homogenates paraquat alone increased SOD activity on day 1 and decreased it on days 3 and 5. Combined treatment with paraquat and EUK-134 elevated LDH activity on day 3 (significantly less than paraquat alone) and day 5, elevated the total protein content on day 5 only, and did not change AcP activity. The combination of both agents did not alter SOD activity and decreased catalase activity on day 5 significantly less than treatment with paraquat alone ( P  = 0.05).
Conclusions:   EUK-134, a superoxide dismutase/catalase mimetic compound decreased the pneumotoxic effect of paraquat in rats.     

Immunohistochemical staining of liver grafts with a monoclonal antibody against HCV-Envelope 2 for recurrent hepatitis C after living donor liver transplantation

Aim:  We evaluated the expression of hepatitis C virus (HCV) antigen on liver grafts by immunohistochemical staining (IHS) using IG222 monoclonal antibody (mAb) against HCV-envelope 2 (E2).
Methods:  The study material was 84 liver biopsy specimens obtained from 28 patients who underwent living donor liver transplantation (LDLT) for HCV infection. The biopsy samples were examined histopathologically, and by IHS using IG222 mAb against HCV-E2. Serum HCV-RNA level was measured in all patients. The IHS grades were compared among the three groups classified according to the time elapsed from LDLT (at 1–30, 31–179 and ≥180 days post-LDLT) and among four post-transplant conditions, including acute cellular rejection (ACR).
Results:  Immunoreactivity to IG222 was detected in 78.6% of the specimens obtained during the first month after LDLT, and there were no significant differences on the IHS grades between the three groups classified according to the time elapsed from LDLT. The IHS grades were significantly stronger in definite recurrent HCV ( n  = 12) and probable recurrent HCV ( n  = 7) than in definite ACR ( n  = 7) and other complications ( n  = 8). There were no significant differences in serum HCV-RNA levels among the four post-transplant conditions. There was no significant correlation between the IHS grades using IG222 mAb and serum HCV-RNA levels when data of 84 liver biopsy specimens were analyzed.
Conclusions:  Constant HCV-E2 expression was observed in liver biopsy specimens obtained 1 month or longer. The strong HCV-E2 expression on liver grafts were associated with recurrent hepatitis C after LDLT, but the serum HCV-RNA levels were not.     

FK 506 ameliorates the hepatic injury associated with ischemia and reperfusion in rats.

The effect of FK 506 on regeneration of the liver was studied in rats after a two-thirds partial hepatectomy after 60 min of ischemia of the unresected liver. The animals were divided into three distinct groups of 10 rats each. Group 1 (controls) received 0.5 ml saline solution intravenously 30 min after the induction of ischemia. Groups 2 and 3 were injected with FK 506 (0.3 mg/kg) intravenously 30 min after and 24 min before the induction of hepatic ischemia, respectively. The hepatic content of ATP and serum levels of ALT and lactate dehydrogenase were determined on each animal. In addition, the histological appearance and mitotic activity of the remnant liver was determined at regular 24-hr intervals after hepatic ischemia. All 10 control animals died within 72 hr. Treatment with FK 506 resulted in improved survival in groups 2 and 3 (30% and 80%, respectively). The improved survival seen in the FK 506-treated animals was reflected by a restoration of hepatic ATP content, a reduction in the serum levels of ALT and lactate dehydrogenase, an amelioration of hepatic necrosis and neutrophilic infiltration and an increase in the mitotic activity of the liver. These results suggest that FK 506 ameliorates the hepatic injury associated with ischemia/reperfusion and has a potent stimulatory effect on liver cell regeneration that may make it valuable as a hepatoprotective agent when administered to organ donors before graft harvesting.     

Correlation of routinely used coagulation parameters and presence of portal vein thrombosis in patients with liver cirrhosis

Aim:  Portal vein thrombosis (PVT) is a serious complication in patients with liver cirrhosis. In patients with advanced stages of liver cirrhosis plasmatic coagulation and platelet count are often reduced. However, patients with normal coagulation status might carry a high risk for developing PVT. A correlation between coagulation status used in clinical routine and the incidence of PVT in patients with liver cirrhosis has been evaluated in the present retrospective analysis.
Methods:  88 patients with liver cirrhosis were identified by screening a database. Of these patients, 23 suffered from PVT. Patients were classified according to the Child–Pugh classification. Patients were subdivided into early stages (Child A) and advanced stages (Child B/C) of liver cirrhosis.
Results:  In patients with Child–Pugh A cirrhosis, there was no difference in activated partial thromboplastin time (apTT), international normalized ratio (INR), and platelet count between the PVT ( n  = 7) and the control group ( n  = 35). In contrast, the median apTT and INR were significantly lower in patients with Child B/C cirrhosis and PVT ( n  = 16) in comparison with patients without PVT (37 s vs 43 s [ P  = 0.017] and 1.25 vs 1.40 [ P  = 0.022]), respectively. Platelet count did not differ significantly in patients with advanced liver cirrhosis and PVT from those without PVT.
Conclusion:  Patients with advanced liver cirrhosis and PVT displayed lower apTT and INR compared with those without PVT. Therefore, patients with advanced liver cirrhosis and almost normal coagulation parameters might be at particular risk of developing PVT. The results suggest that regular monitoring using Doppler-ultrasound should be carried out in these patients, especially when liver transplantation is intended.