Alrp, a survival factor that controls the apoptotic process of regenerating liver after partial hepatectomy in rats

Augmenter of Liver Regeneration (Alrp) enhances, through unknown mechanism/s, hepatocyte proliferation only when administered to partially hepatectomized (PH) rats. Liver resection, besides stimulating hepatocyte proliferation, induces reactive oxygen species (ROS), triggering apoptosis. To clarify the role of Alrp in the process of liver regeneration, hepatocyte proliferation, apoptosis, ROS-induced parameters and morphological findings of regenerating liver were studied from PH rats Alrp-treated for 72 h after the surgery. The same parameters, evaluated on regenerating liver from albumin-treated PH rats, were used as control. The results demonstrated that Alrp administration induces the anti-apoptotic gene expression, inhibits hepatocyte apoptosis and reduces ROS-induced cell damage. These and similar data from in vitro studies and the presence of 'Alrp homologous proteins' in viruses as well as in mammals (i) allow to hypothesize that Alrp activity/ies may not be exclusive for regenerating liver and (ii) suggest the use of Alrp in the treatment of oxidative stress-related diseases.     

Delayed liver regeneration after partial hepatectomy in adiponectin knockout mice

We previously demonstrated that adiponectin has anti-fibrogenic and anti-inflammatory effects in the liver of mouse models of various liver diseases. However, its role in liver regeneration remains unclear. The aim of this study was to determine the role of adiponectin in liver regeneration. We assessed liver regeneration after partial hepatectomy in wild-type (WT) and adiponectin knockout (KO) mice. We analyzed DNA replication and various signaling pathways involved in cell proliferation and metabolism. Adiponectin KO mice exhibited delayed DNA replication and increased lipid accumulation in the regenerating liver. The expression levels of peroxisome proliferator-activated receptor (PPAR) α and carnitine palmitoyltransferase-1 (CPT-1), a key enzyme in mitochondrial fatty acid oxidation, were decreased in adiponectin KO mice, suggesting possible contribution of altered fat metabolism to these phenomena. Collectively, the present results highlight a new role for adiponectin in the process of liver regeneration.     

Effects of carvacrol upon the liver of rats undergoing partial hepatectomy

The present study aims to investigate the possible effects of carvacrol obtained from origanum oil upon the regenerative feature of the liver subsequent to partial hepatectomy in rats.Male Wistar Albino rats, weighing 230±30 g, were divided into three experiment groups. Group I (n=8) were used as sham operation group. Group II (n=8) were applied saline solution and hepatectomy. Carvacrol and hepatectomy (73 mg/kg) were applied to Group III (n=8). One dose of test material was injected 1 h before 68% partial hepatectomy. At the end of the experiments, blood and organs were removed. The liver regeneration rate of the rats was calculated measuring the weights of their liver before and after the hepatectomy. Hematoxylin and eosin, interleukin-6 (IL-6) and proliferating cell nuclear antigen (PCNA) treatments were applied to liver sections. Aspartate transaminase (AST), alanine transaminase (ALT), tumor necrosis factor-alpha (TNF-α) and IL-6 levels were determined in serum samples.The liver regeneration, mitotic index and PCNA index increased significantly in rats of Group III over the Group II at the 72nd hour after partial hepatectomy. Histological evaluations were also similar with these results of PCNA and mitotic indexes. In AST, ALT, TNF-α and IL-6 levels, there was no statistically significant difference.According to these results, it is concluded that carvacrol increases the liver regeneration rate.     

Expression of hepatocyte growth factor mRNA in regenerating rat liver after partial hepatectomy.

Hepatocyte Growth Factor (HGF) is a potent complete mitogen for primary cultures of hepatocytes in vitro. There is strong evidence that this novel growth factor may mediate hepatocyte regeneration after liver damage. We have shown previously that the amount of immunoreactive HGF markedly increases in the serum of rats soon after partial hepatectomy or CCl4 administration. In the present paper, we demonstrate that the level of HGF mRNA in rat liver also dramatically increases from 3 to 6 hours post hepatectomy, peaks at 12 hr and gradually returns to undetectable levels by 72 to 96 hours post hepatectomy. In separate experiments, DNA synthesis (in vivo) was determined in rat liver remnants after partial hepatectomy. DNA synthesis peaked 24 hr after hepatectomy, 12 hr after the peak of HGF mRNA expression. These results suggest that HGF may be one of the major early signals that triggers hepatocyte proliferation during liver regeneration.     

Ultrastructural zonal heterogeneity of hepatocytes and mitochondria within the hepatic acinus during liver regeneration after partial hepatectomy

BACKGROUND INFORMATION: Partial hepatectomy (70%) induces cell proliferation until the original mass of the liver is restored. In the first 24 h after partial hepatectomy, drastic changes in the metabolism of the remaining liver have been shown to occur. To evaluate changes in hepatocyte ultrastructure within the hepatic acinus during the liver regenerative process, we investigated, by light and electron microscopy observations on specimens taken 0 h, 24 h and 96 h after partial hepatectomy, the hepatocyte structure and ultrastructure in the periportal and pericentral area of the hepatic acinus, with a particular emphasis on mitochondria ultrastructure. Moreover, some biochemical events that could affect the mitochondria ultrastructure and function were investigated. RESULTS: We found that, 24 h after partial hepatectomy, mitochondria with altered ultrastructure were preferentially localized in the periportal area. Periportal hepatocytes showed also an increase in the number of peroxisomes, free ribosomes, lysosomes and autophagosomes. Altered mitochondria showed swelling, an ultrastructural index of increased membrane permeability, a reduction in the number of cristae, and a rarefied, often vacuoled, matrix. Consistently, an increase in the mitochondrial oxidized/reduced glutathione ratio was found as well as calcium release from mitochondria in a manner inhibited by cyclosporin A. Interestingly, light and electron microscopy analysis showed that the hepatocytes in the periportal area were the cells with the major structural attributes to proliferate. At 96 h after partial hepatectomy, the preferential zonation of altered mitochondria was lost and the normal mitochondrial membrane permeability properties were restored. CONCLUSIONS: We suggest that 24 h after partial hepatectomy, a preferential zonation of altered mitochondria in the periportal hepatocytes could be involved in the changes of metabolic and functional heterogeneity of the hepatocytes within the hepatic acinus during the regenerative process.     

Chronobiology of the proliferative events related to angiogenesis in mice liver regeneration after partial hepatectomy

In liver regeneration the formation of new capillary blood vessels is a fundamental requirement for cellular proliferation. Vascular endothelial growth factor (VEGF) is involved in the events of angiogenesis, the mRNA of which is expressed in both hepatocytes and non-parenchymal cells. In this experimental design we try to establish if during liver regeneration in mouse, the expression of VEGF is produced before or after the hepatocytes proliferation. C3H/S adult male mice were divided in three groups in order to study: VEGF expression; S-phase index (SI); and mitotic activity (MA) of hepatocytes. The results that were analyzed by ANOVA, show that VEGF expression starts to increase 26 h after PH with a peak at 28 h. Furthermore, the DNA synthesis (DNAs) reaches maximal level 42 h after pH, meanwhile the MA of the hepatocytes shows an increase 8h after the DNAs peak. In conclusion, it could be argued that the chronobiology of the events related to liver regeneration in mice started with a release of VEGF by the hepatocytes, followed by its DNAs and mitosis.     

Glycosylation and ligand-binding activities of rat plasma fibronectin during liver regeneration after partial hepatectomy

Fibronectin (FN) is a multifunctional glycoprotein present in the extracellular matrix (ECM) and plasma. We previously reported that the glycosylation and ligand-binding of vitronectin (VN) change markedly after partial hepatectomy (PH). Here we show the changes of FN during liver regeneration. The yields of purified sham-operated (SH-) and PH-FN were higher than that of non-operated (NO)-FN, while binding activities of FNs to ECM ligands were changed only slightly by hepatectomy. The carbohydrate concentration of PH-FN decreased to 66% of that of NO- and SH-FN. By using LC/MS(n), eight kinds of complex-type N-glycan structures were found to be present in all FNs, and bi- and trisialobiantennary glycans were the major structures. Fucosylation was markedly increased, while O-acetylation of sialic acid was found to be decreased in PH-FN. The alterations in glycosylation and biological activities of FN after PH are different from those of VN, suggesting that these glycoproteins play different biological functions in tissue remodeling.     

Turnover of ribosomal proteins in regenerating rat liver after partial hepatectomy

The rates of synthesis and degradation of ribosomal proteins, prelabelled with [14C]bicarbonate, were determined as an index of the rate of ribosome turnover in regenerating rat liver. The half-life of ribosomes is about 178 and 75 hr in regenerating and normal liver, respectively. The comparison of turnover rates of ribosomal proteins with the corrected values of rRNA, based on re-utilization of nucleotides, suggests that ribosomes are metabolized as a unit in vivo. There is at least 70% overestimation for ribosome half-life when orotate-labelled RNA is used for turnover determinations. The absolute rate of synthesis is estimated as 3925 and 1081 ribosomes/min per cell in 24 hr regenerating and normal rat liver, respectively.     

The nucleolar protein NML regulates hepatic ATP levels during liver regeneration after partial hepatectomy

We previously identified a novel protein complex, eNoSC, which senses intracellular energy status and epigenetically regulates the rDNA locus by changing the ratio between the numbers of active and silent gene clusters. eNoSC contains a novel nucleolar protein, Nucleomethylin (NML), which has a methyltransferase-like domain and binds to Lys9-dimethylated histone H3 at the rDNA locus, along with the NAD⁺-dependent deacetylase SIRT1 and the histone methyltransferase SUV39H. The aim of this study was to determine the role of NML in liver after partial hepatectomy (PHx). We assessed liver regeneration and lipid metabolism after PHx in wild-type (WT) and NML transgenic (NML-TG) mice. Survival rates of NML-TG mice were reduced after PHx. We found that hepatic triglyceride content in NML-TG mice remained elevated 48 h after PHx, but not delayed liver regeneration. Moreover, hepatic ATP levels in NML-TG mice were higher than that in WT 48 h after PHx. These observations suggest that NML may regulate consumption of hepatic triglyceride in liver regeneration after PHx due to storage of excess ATP. The delayed consumption of hepatic triglyceride may be the cause of reduced survival rate in NML-TG mice.     

Increase of transferrin receptors in regenerating rat liver cells after partial hepatectomy

The change of transferrin receptors in regenerating rat liver cells after partial hepatectomy was demonstrated. The binding of 125I-labeled transferrin to the cells began to increase after 24 h of partial hepatectomy and reached about double that of the non-operated normal rat liver cells. A Scatchard analysis of binding parameters showed 1.62 X 10(5) (Ka: 1.04 X 10(7) M-1) in normal cells and 3.36 X 10(5) (Ka: 1.23 X 10(7) M-1) in regenerating cells. This reflected on increase of transferrin receptor number and little change occurred in the binding affinity between transferrin and its surface receptor.     

Formation and accumulating of hypotaurine in rat liver regenerating after partial hepatectomy

Hypotaurine is considered to be an intermediate in the major pathway for the biosynthesis of taurine in mammals yet is rarely detected in mammalian tissue. The activity of cysteinesulfinic acid decarboxylase, the enzyme presumably responsible for the biosynthesis of hypotaurine, is frequently present in great amounts in tissue, whereas the mechanism for the conversion of hypotaurine to taurine is poorly understood, there being some doubt at present if an enzyme exists for such a purpose. This paper reports the accumulation of hypotaurine in the liver of rats regenerating after partial hepatectomy. Further, the formation and accumulation of [³⁵S]hypotaurine from [³⁵S]methionine under the same conditions was observed. No hypotaurine was detected in liver of sham-operated control animals, even after the intraperitoneal injection of authentic hypotaurine. These observations suggest that rat liver normally possesses a mechanism for the rapid conversion of hypotaurine to taurine and that this mechanism is impeded in liver regenerating after partial hepatectomy.     

Liver regeneration after partial hepatectomy in rat is more impaired in a steatotic liver induced by dietary fructose compared to dietary fat

Hepatic steatosis (HS) has a negative effect on liver regeneration, but different pathophysiologies of HS may lead to different outcomes. Male Sprague-Dawley rats were fed a high fructose (66% fructose; H-fruc), high fat (54% fat; H-fat), or control chow diet for 4 weeks. Based on hepatic triglyceride content and oil red O staining, HS developed in the H-fruc group, but was less severe compared to the H-fat group. Hepatic mRNA expression levels of fatty acid synthase and fructokinase were increased and those of carnitine palmitoyltransferase-1 and peroxisome proliferator-activated receptor-α were decreased in the H-fruc group compared to the H-fat group. Liver regeneration after 70% partial hepatectomy (PHx) was evaluated by measuring the increase in postoperative liver mass and PCNA-positive hepatocytes, and was impaired in the H-fruc group compared to the H-fat and control groups on days 3 and 7. Serum levels of tumor necrosis factor-α, interleukin-6 and hepatocyte growth factor did not change significantly after PHx. In contrast, serum TGF-β1 levels were slightly but significantly lower in the control group on day 1 and in the H-fat group on day 3 compared to the level in each group on day 0, and then gradually increased. However, the serum TGF-β1 level did not change after PHx in the H-fruc group. These results indicate that impairment of liver regeneration after PHx in HS is related to the cause, rather than the degree, of steatosis. This difference may result from altered metabolic gene expression profiles and potential dysregulation of TGF-β1 expression.     

In vivo transfer of hepatocyte growth factor gene accelerates proliferation of hepatic oval cells in a 2-acetylaminofluorene/partial hepatectomy model in rats

To clarify the effect of hepatocyte growth factor (HGF) on proliferation of hepatic oval cells, we transferred HGF gene into liver of the Solt-Farber rat model. Male Fisher 344 rats were infected with a recombinant adenovirus carrying the cDNA for HGF (pAxCAHGF) from tail vein. HGF mRNA showed its peak at 4 days, and diminished thereafter. The total and proliferating cell nuclear antigen-positive hepatic oval cells were significantly elevated in HGF-transferred rats, in which stem cell factor and c-kit mRNA increased at each time point. Our results suggest that in vivo transfer of the HGF gene into liver accelerates proliferation of hepatic oval cells in the Solt-Farber model in rats.