Effect of Interferon-α2b Administration on Rat Liver Regeneration After Partial Hepatectomy

The purpose of the present study was todelineate the effect of interferon-_2b(IFN-_2b) administration on the liverregenerative capacity after partial hepatectomy in rats.The administration of IFN-_2bsimultaneously with partial hepatectomy did not affect hepatic2b proliferation in a statistically significant manner.When IFN-_2b was administered either 2or 12 hr postoperatively, an inhibition of hepatocyteproliferation was observed 24 hr postoperatively, while atfurther time intervals up to 48 hr, DNA synthesisremained similar to that observed in the simplypartially hepatectomized rats. The enzyme thymidinekinase (TK), has been implicated in the suppression ofproliferation in interferon-treated cell cultures. Inall IFN-_2b-treated groups of rats,alterations of TK activity were observed without beingcorrelated to the liver regenerative status. Additionally,the administration of the polyamine putrescine inpartially hepatectomized rats treated at the time ofsurgery with IFN strongly enhanced TK activity, but did not affect DNA biosynthesis. In theabove-mentioned in vivo model of controlled cellularproliferation, the administration ofIFN-_2b affected the rate of hepatocyteproliferation depending on the time of its administration; this effect was notcorrelated to the enzymatic activity of TK, as inhibitedTK activity is responsible for the suppressed DNAsynthesis in in vitro systems.     

Morphological and Biochemical Effects of a Low Ethanol Dose on Rat Liver Regeneration Role of Route and Timing of Administration

We have demonstrated that in rats subjected topartial hepatectomy (PH), the regenerating liver had anenhanced metabolism of ethanol, which largely dependedon the route and timing of ethanol administration. Therefore, the influence of the administrationroute and timing for ethanol-induced deleterious effectson the regenerating rat liver was evaluated in animalssubjected to 70% PH. Remnant liver showed moderate fatty infiltration, extended distortion ofhepatocellular structure, and high mitotic index.Intragastric ethanol administration (1.5 g/kg bodyweight) considerably reduced the PH-induced changes inliver structures. Ethanol treatment also decreasedliver thymidine kinase activity, serum albumin, andglucose levels. Intraperitoneal administration of thesame ethanol dose to PH rats promoted lesser alterations on liver regeneration. Independently of itsadministration route, ethanol abruptly shortened aPH-induced selective increase in serum enzymeactivities. These data suggest that the inhibitoryeffect of a low dose of ethanol on PH-induced liverregeneration is dependent on the timing and route ofadministration.     

Effect of Granulocyte Colony-Stimulating-Factor Administration on Tissue Regeneration Due to Thioacetamide-Induced Liver Injury in Rats

It has been shown recently that granulocytecolony-stimulating factor (G-CSF) accelerates andenhances the hepatocyte proliferative capacity ofpartially hepatectomized rats. In the present study, weinvestigated the effect of G-CSF administration in a ratmodel of liver injury and regeneration, induced bythioacetamide (TAA) injection. TAA (300 mg/kg bodyweight) was injected intraperitoneally in male Wistarrats, and this was followed by administration ofeither saline (group A) or G-CSF at a dose of 150g/kg body weight (group B), 24 hr later. The animalswere killed at different time points after TAA treatment and the rate of tritiated thymidineincorporation into hepatic DNA, the activity of theenzyme thymidine kinase (EC 2.7.1.21) in the liver, andthe assessment of the mitotic index of hepatocytes, wereemployed to estimate liver regeneration. Theadministration of TAA caused severe hepatic injury,recognized histopathologically and by the raisedactivities of the serum hepatic enzymes aspartate andalanine aminotransferases. The hepatic injury, which peaked 36 hr afterTAA injection, was followed by a regenerative process ofhepatocytes presenting peaks at time points of 48 and 60hr (group A). The administration of G-CSF 24 hr after the injection of TAA (group B) causeda statistically significantly increase in the hepatocyteproliferation indices examined (P < 0.001), comparedto those found in group A at the same time points. It was concluded that G-CSFadministration enhanced the hepatocyte proliferativecapacity in this model of liver injury induced by TAAadministration.     

Intra-and Extrahepatic Leukocytes and Cytokine mRNA Expression During Liver Regeneration After Partial Hepatectomy in Rats

We investigated intra- and extrahepaticleukocytes during liver regeneration after a 70% partialhepatectomy in the rats using flow cytometry and RT-PCRfor cytokine mRNA expression. Our study indicated that LFA-1⁺⁺⁺ CD3⁺ cell,NK3.2.3⁺⁺ T cells, and CD5⁺ Bcells, which are activated in autoimmune diseases andmalignancy in humans and mice, were activated in theearly phase of liver regeneration in the liver and the blood after partial hepatectomyin the rats. On measuring cytokine mRNA expression byRT-PCR, only IFN- mRNA was detected in the normalrat liver. IFN- mRNA expression clearly decreased in the liver on day 1 after partial hepatectomyand increased thereafter. IL-2 and IL-4 mRNA were notdetected in the liver regardless of hepatectomy. Everycytokine was detected in normal spleen and increased in the early phase after partial hepatectomy.These were also detected in normal thymus; however, IL-2and IFN- mRNA expressions decreased and IL-4 mRNAexpression increased slightly in the early phase after partial hepatectomy. Thus, the immunesystem dramatically changed both in the liver andelsewhere in the early phase of liver regeneration afterpartial hepatectomy.     

Effect of Vitamin E Deficiency on Inhibition of Liver Regeneration by Long-Term Administration of Alcohol

The effects of vitamin E (VE) deficiency on liver regeneration suppressed by long-term administration of alcohol were studied. Male rats were divided into two groups: the alcohol group and the control group. In addition, each group was subdivided into two groups according to the presence or not of VE. Altogether, four groups were provided: a group maintained on the VE-deficient alcohol diet (group EA), a group maintained on the VE-deficient control diet (group EC), a group maintained on the ordinary alcohol diet (group A), and a group maintained on the ordinary control diet (group C). After pair-feeding for 6 weeks, partial hepatectomy was performed to determine the ornithine decarboxylase (ODC) activity, polyamine levels, lipid peroxide levels, and DNA synthesis. DNA synthesis at 24 hr after partial hepatectomy was suppressed significantly in the alcohol administration group, regardless of the presence or not of VE. DNA synthesis at 48 hr after partial hepatectomy tended to show low values in group EA, compared with group A. As for the hepatic ODC activity, group EA showed the lowest value at 4 hr after partial hepatectomy. Of polyamines, the putrescine level in group EA at 4 hr after partial hepatectomy was significantly low, compared with the other three groups. The levels of spermidine and spermine decreased by long-term administration of alcohol, but the effect of VE deficiency was not found. The lipid peroxide level increased significantly in the VE-deficient diet administration group, but the effect of alcohol administration was not found. These results suggested that the decrease in putrescine after ODC suppression by VE deficiency in addition to the decrease in spermidine and spermine caused by long-term alcohol administration were concerned with suppression of DNA synthesis later.     

Uptake of Extracellular, Dietary Putrescine Is an Important Regulatory Mechanism of Intracellular Polyamine Metabolism DuringCamostate-Induced Pancreatic Growth in Rats

Aim of the present study was to evaluate therole of cellular uptake of dietary [ H]putrescine forthe regulation of pancreatic, hepatic, and smallintestinal polyamine metabolism during normal andcamostate-induced pancreatic growth in rats in vivo. Initiallydose-response and time-course studies of[³H]putrescine uptake were performed. MaleWistar rats were either treated with the synthetictrypsin inhibitor camostate (200 mg/kg body wt orally twice daily),camostate plus the ornithine decarboxylase inhibitoralpha-difluoromethylornithine (DFMO) (2% in drinkingwater plus 3 × 300 mg/kg body wt intraperitoneallyduring daytime) or saline as controls. After 4, 8, 12,24, 36, 48, or 120 hr, five to seven animals per groupwere killed, respectively. Orally fed [ H]putrescine (10nmol/kg body wt. 2 hr prior to death) is rapidly taken up and further metabolized tospermidine in normal growing pancreas, liver, and smallintestine. Feeding of camostate significantly enhanceddietary [³H]putrescine uptake, whilesimultaneous inhibition of de novo synthesis ofintracellular polyamines by DFMO resulted in a highlysignificant further increase in cellular uptake oforally fed [³H]putrescine, which isimmediately metabolized to spermidine. The present in vivo data confirmthe important role of dietary putrescine uptake for themaintenance of intracellular polyamine pool in normaland stimulated pancreatic growth. Furthermore, dietary putrescine uptake is an importantregulatory mechanism to maintain the normal andgrowth-stimulated cellular polyamine pool in thepancreas after potent simultaneous inhibition ofintracellular de novo polyamine synthesis.     

Enhancement of Liver Regeneration by the Association of Hyptis pectinata with Laser Therapy

Since new molecules that normally would accelerate regeneration can also be potentialized by light, the use of new substances combined with laser therapy seems to be a natural type of experiment. Therefore, the purpose of this study was to assess the effects of Hyptis pectinata leaves on liver regeneration after partial hepatectomy (PH) associated with laser therapy. Twenty-four rats were divided into four groups—PH(control), PHL (laser therapy), PH200 (200 mg/kg of Hyptis pectinata), and PHL200 (200 mg/kg of the plant and laser)—which were submitted to 67% hepatectomy. Laser treatment consisted of focusing the light on the remaining liver after hepatectomy. The data analyzed were serum levels of aminotransferases, liver regeneration, and mitochondrial function. Group PH200 showed a statistically significant decrease in AST levels, and PHL200 disclosed an augmentation in ALT levels. The liver regeneration index was significantly increased in group PHL200. Concerning liver mitochondrial respiratory assay, groups PH200 and PHL200 showed lower state 3 levels than groups PH and PHL. Group PHL showed an increase in state 4 levels and a reduction in membrane potential and RCR. The present study shows that the association of the aqueous extract of Hyptis pectinata leaves at 200 mg/kg with intraoperative laser therapy can stimulate liver regeneration and cause a reduction in liver mitochondrial respiratory function without altering its phosphorylative activity.     

Granulocyte-Colony Stimulating Factor Administration Ameliorates Liver Regeneration in Animal Model of Fulminant Hepatic Failure and Encephalopathy

It has previously been shown that recombinant granulocyte-colony stimulating factor (rG-CSF) accelerates and enhances hepatocyte proliferation in partially hepatectomized rats. In the present study, we examined the effect of rG-CSF administration on liver injury, regeneration, and survival outcome in an experimental rat model of fulminant hepatic failure (FHF) and encephalopathy induced by repeated injections of thioacetamide (TAA). FHF was induced in adult male Wistar rats by three consecutive intraperitoneal injections of TAA, at intervals of 24 hr. The animals were also injected with either saline or rG-CSF. Serum biochemical parameters and blood ammonia levels, liver histology, stage of hepatic encephalopathy, and survival were statistically significantly improved in TAA-intoxicated and rG-CSF-treated rats compared to TAA-intoxicated and saline-treated ones. Furthermore, rG-CSF not only ameliorated the histologically evident liver injury in a statistically significant manner but also enhanced the proliferative capacity of the hepatocytes. Our data confirm the beneficial effect of rG-CSF administration in this animal model of FHF and encephalopathy, supporting evidence for a possible use of rG-CSF as supportive therapy in the management of FHF.     

Pharmacological Inhibition of Protein Kinase C Activity Could Induce Apoptosis in Gastric Cancer Cells by Differential Regulation of Apoptosis-Related Genes

The protein kinase C (PKC) signaling pathwayplays a key role in tumor cell proliferation,differentiation, and apoptosis. Gastric cancer usuallypossesses a higher level of PKC activity than normaltissue. We evaluated inhibition of PKC activity inapoptosis induction of gastric cancer cells and theexpression profile of apoptosis-related genes. Gastriccancer cells (AGS) were incubated with two highlyspecific PKC inhibitors (RO-31-8220 and chelerythrine).Cell viability and cell cycle were determined bymethyl-tetrazolium (MTT) assay and flow cytometry,respectively. Apoptosis was characterized by acridineorange staining, DNA gel electrophoresis, and flowcytometry. The expression of p53,p21^waf/cip1, c-myc, bcl-2, and bax wasdetermined by western blot. The results showed that bothPKC inhibitors hindered cell growth, arrested cells atG₀/G₁ phase and induced apoptosis.The protein level of p53, p21^waf/cip1, c-myc,and bax was elevated while bcl-2 kept unchangedfollowing drug exposure. In conclusion, PKC inhibitorssuppress growth of gastric cancer cells throughapoptosis induction and cell cycle quiescence, which maybe regulated by differential expression ofapoptosis-related genes.     

Effect of Pancreaticobiliary Diversion on Liver Regeneration

To ascertain whether the increase in cholecystokinin (CCK) level associated with pancreaticobiliary diversion has a cytoproliferative effect on the liver similar to that on the pancreas, we studied three groups of rats: group I (n = 19) had a pancreaticobiliary diversion and 4 weeks later a 70% liver resection, group II (n = 13) had a liver resection only, and group III (n = 6) underwent a sham liver operation. Tissue samples were taken from the liver and pancreas 48 h after the liver operation. Liver regeneration was evaluated on the basis of continuous incorporation of tritiated thymidine into hepatocytes, liver weight, and DNA content. For confirmation of the increase in CCK levels the effect on the pancreas was studied by measuring wet weight, total protein, and total DNA content. The results showed trophic effects on the pancreas, as expected, but no effects whatsoever on liver regeneration. CCK does not seem to have any role in the regulation of liver regeneration.     

Identification of Bile Canalicular Cell Surface Antigen HAM.4 as Dipeptidyl Peptidase IV (DPPIV) and Characterization of Its Role in Hepatic Regeneration After Partial Hepatectomy in Rats

Dipeptidyl peptidase IV (DPPIV) has beenimplicated in the control of cell growth anddifferentiation. A rat hepatocyte membrane antigenrecognized by a monoclonal antibody (HAM.4) has now beenshown to be identical to DPPIV by immunoblot analysisand amino acid sequencing. The amounts of DPPIVimmunoreactive protein and enzymatic activity in serumincreased in a manner independent of de novo proteinsynthesis, and without any biochemical orimmunohistochemical changes in hepatic DPPIV, duringliver regeneration after partial hepatectomy in rats.DPPIV purified from serum by HAM.4 antibody-basedaffinity chromatography lacked the NH₂-terminal 36 aminoacids of the membrane-bound enzyme, suggesting thatproteolytic cleavage may mediate the release of DPPIVinto serum. No significant differences in therestoration of liver mass or in hepatic DNA synthesis were apparentbetween DPPIV-deficient and normal rats after partialhepatectomy, suggesting that DPPIV may not be essentialfor hepatic regeneration.     

Ethanol-Associated Alterations in the Kinetics of Putrescine Uptake and Metabolism by the Regenerating Liver

Biosynthesis of the polyamines, putrescine, spermidine, and spermine is required for DNA synthesis and liver regeneration after partial hepatectomy. We have previously reported that chronic ethanol consumption impairs polyamine synthesis and significantly retards liver regeneration after partial hepatectomy. In those studies, supplementation with putrescine restored hepatic DNA synthesis in ethanol-fed rats but exerted no effect in pair-fed controls. These differences in the response to putrescine treatment may have resulted from ethanol-associated differences in hepatic uptake, release, or metabolism of putrescine. To resolve these issues and define more completely how putrescine treatment affects DNA synthesis, we now assess the kinetics of putrescine uptake and metabolism after intraperitoneal or intravenous injection of radiolabeled putrescine (1.2 mmol/kg, specific activity 1 microCi/mmol) into rats fed 36% ethanol diets or isocaloric, nonethanol diets for 6 weeks prior to partial hepatectomy. After putrescine treatment, hepatic putrescine concentrations were greater in ethanol-fed rats than controls. Differences in post-treatment hepatic putrescine levels between ethanol and pair-fed groups could not be explained by differences in the rates of hepatic putrescine uptake or excretion into bile; residual de novo synthesis of putrescine from ornithine or metabolism of hepatic putrescine to its polyamine products, spermidine and spermine. Indeed, supplemental putrescine was not appreciably converted to spermidine or spermine in either ethanol or control rats. Hence, these latter polyamines are unlikely to be responsible for the treatment-associated improvement in DNA synthesis that has been noted in ethanol-fed rats. This suggests that putrescine itself acts to restore hepatic DNA synthesis in ethanol-fed rats.     

Optimal Administration of Tacrolimus in Reduced-Size Liver

The optimal administration of immunosuppressants such as tacrolimus (Tac) for small-for-size (SFS) grafts, where the functional liver mass is small and must regenerate, has not been reported so far. The aim of this study is to clarify the characteristics of Tac metabolism according to liver volume. Seven-week-old male Wistar rats were randomly divided into three groups: (1) Tac administrated and 70% Hx group (Tac 70% Hx group), (2) Tac administrated and 90% Hx group (Tac 90% Hx group), and (3) vehicle administrated and 90% Hx group (control 90% Hx group). In both the Tac groups, Tac (0.3 mg/kg) was given daily for 3 days before operation, and daily after surgery until sacrifice (each time point; n = 5). The plasma concentration of Tac (trough level), as well as liver toxicity, were measured. The plasma concentration of Tac in the Tac 90% Hx group was significantly higher than in the Tac 70% Hx group from 24 to 72 h after operation. Furthermore, expression of CYP3AII mRNA was significantly lower in the Tac 90% Hx group than in the Tac 70% Hx group. Regarding the liver toxicity, there was no significant difference in both the Tac 90% Hx and the control 90% Hx groups. In this experimental study, the plasma concentration of Tac was dependent on the remnant liver volume. Therefore, special attention in regard to Tac administration should also be taken for patients with SFS grafts in living-donor liver transplantation (LDLT).     

人肝再生增强因子基因组DNA的克隆化与序列分析

目的克隆人肝再生增强因子(ALR)的基因组DNA,并进行序列分析。方法采用人肝再生增强因子cDNA及其编码序列,对基因的核苷酸序列数据库(GenBank)以BLAST为工具进行核苷酸序列同源性比较分析,寻找相应的ALR基因组DNA序列。结果从GenBank核苷酸序列数据库中寻找到人ALR基因组DNA全序列,由1813个核苷酸组成。人ALR基因组DNA序列由3段外显示子(exon)组成,分别为18nt,197nt和163nt。基因的编码产物由125个氨基酸残基组成,与小鼠ALR基因组DNA结构类似。基因组DNA定位于染色体的16p13.3位点上。结论克隆了人ALR基因组DNA全序列。     

人肝再生增强因子的分子克隆及生物学活性研究

在众多参与肝细胞再生调控的细胞因子中,如TGF、EGF及大分子HGF等都缺乏器官特异性,而     

The Role of Mesothelial Cells in Liver Development,Injury, and Regeneration

Mesothelial cells (MCs) cover the surface of visceral organs and the parietal walls of cavities, and they synthesize lubricating fluids to create a slippery surface that facilitates movement between organs without friction. Recent studies have indicated that MCs play active roles in liver development, fibrosis, and regeneration. During liver development, the mesoderm produces MCs that form a single epithelial layer of the mesothelium. MCs exhibit an intermediate phenotype between epithelial cells and mesenchymal cells. Lineage tracing studies have indicated that during liver development, MCs act as mesenchymal progenitor cells that produce hepatic stellate cells, fibroblasts around blood vessels, and smooth muscle cells. Upon liver injury, MCs migrate inward from the liver surface and produce hepatic stellate cells or myofibroblast depending on the etiology, suggesting that MCs are the source of myofibroblasts in capsular fibrosis. Similar to the activation of hepatic stellate cells, transforming growth factor β induces the conversion of MCs into myofibroblasts. Further elucidation of the biological and molecular changes involved in MC activation and fibrogenesis will contribute to the development of novel approaches for the prevention and therapy of liver fibrosis.     

Stimulatory Effect of Epidermal Growth Factor on Liver Regeneration after Partial Hepatectomy in Rats

The effect of epidermal growth factor (EGF) on liver regeneration was investigated in rats subjected to partial hepatectomy. In a dose-response study EGF in doses of 6 and 24 nmol/kg X day increased liver regeneration after treatment for 48 h compared with controls, whereas a dose of 48 nmol/kg X day had no effect. In a subsequent study EGF, 6 nmol/kg X day, accelerated liver regeneration significantly after 36, 48, and 72 h of treatment. A possible influence of EGF on other hepatotrophic factors was investigated. No changes in the concentration of gastrin, insulin, or glucagon was found in portal venous blood. This study has shown that EGF in small doses can stimulate liver regeneration, whereas higher doses are ineffective. The study suggests that EGF should be regarded as a hepatotrophic factor.     

Beneficial Effects of Fluvastatin on Liver Microcirculation and Regeneration After Massive Hepatectomy in Rats

Fluvastatin, the first entirely synthetic statin, has a significant cholesterol-lowing effect comparable with other statins. In addition, it has been shown to inhibit oxidative stress and improve vascular endothelial function. The aim of this study was to clarify the pretreatment effects of fluvastatin on liver function after massive hepatectomy in rats. Six-week-old male Wister rats were divided into two groups: a fluvastatin group (group F), pretreated with oral administration of fluvastatin (20 mg/kg per day) for 2 days before 90% hepatectomy; and a control group (group C), pretreated with vehicle for 2 days before hepatectomy. Animals were sacrificed at 0, 12, 24, 48, and 72 h after hepatectomy. The liver regeneration rate, liver function tests, and hepatic stellate cell activation were examined. The liver regeneration rate in group F was significantly higher at 72 h after hepatectomy (P < 0.05). The serum level of total bilirubin in group F was significantly lower at 48 h after hepatectomy (P < 0.05). Sinusoidal area in group F was maintained histologically. Furthermore, the expression of alpha smooth-muscle actin (α-SMA) protein in the liver was inhibited in group F at 48 h after hepatectomy. This study demonstrated the beneficial effects of fluvastatin in a lethal massive hepatectomy model using rats, with improved hepatic regeneration and microcirculations, by inhibiting the activation of hepatic stellate cells.     

Chronic Ethanol Administration Impairs Degradation of Formaldehyde-Treated Albumin by the Perfused Rat Liver

Nonparenchymal cells of the liver appear to be important in the pathogenesis of various liver diseases, including that caused by ethanol. It is known that chronic ethanol administration impairs the process of receptor-mediated endocytosis in hepatocytes. Liver endothelial cells are also actively endocytic cells, playing a prominent role in the clearance from the circulation of a variety of macro-molecules. In this study, we assessed the effect of ethanol administration on this "scavenger" function of liver endothelial cells by measuring the degradation of formaldehyde-treated albumin in isolated, perfused livers of ethanol-fed rats. Rats were pair-fed for 1 or 4 weeks with a liquid diet containing either ethanol as 36% of total calories or an isocaloric amount of carbohydrate. Chronic ethanol administration in this manner for 1 or 4 weeks significantly impaired the degradation of this endothelial cell iigand (by 60 ± 9% and 37 ± 9%, respectively). Liver perfusions were also performed on rats that had been administered ethanol acutely or in which ethanol was added to the perfusate. No acute effect of ethanol on the degradation of this Iigand was seen. These results demonstrate that chronic ethanol ingestion impairs receptor-mediated endocytosis of formaldehyde-treated albumin by liver endothelial cells, indicating that the adverse effects of ethanol on protein trafficking within the liver are not limited to the hepatocytes.     

Effects of Ethanol Administration on Hepatocellular Ultrastructure of Regenerating Liver Induced by Partial Hepatectomy

Acute ethanol administration partially inhibits DNA and protein syntheses during liver regeneration (LR) induced by partial hepatectomy (PH) in rats. Previous findings that the magnitude of ethanol's deleterious effects on LR are related to the route and timing of its administration led us to perform studies at the ultrastructural level, comparing ethanol effects on PH-induced LR, as a consequence of its administration route. PH promoted alterations on the endoplasmic reticulum and mitochondria, accompanied by decreased glycogen and increased lipid content in cytoplasm. Structural nuclear and nucleolar activities were also evident. Intragastric ethanol administration practically abolished the adaptative changes found in PH-promoted regenerating hepatocytes, whereas its administration through the intraperitoneal route induced later ultrastructural modifications, indicating cellular proliferation. These results suggest that ethanol, under certain conditions, could stimulate liver proliferation triggered by PH. The mechanism underlying this surprising effect of ethanol on LR remains to be elucidated. However, it is suggested that an altered ethanol metabolism by rats subjected to PH could be involved.