Hepatocytes In Zone Research Articles

EGFR-mediated crosstalk between vascular endothelial cells and hepatocytes promotes Piezo1-dependent liver regeneration

Hepatocyte proliferation is essential for recovering liver function after injury. In liver surgery, the mechanical stimulation induced by hemodynamic changes triggers vascular endothelial cells (VECs) to secrete large amounts of cytokines that enhance hepatocyte proliferation and play a pivotal role in liver regeneration. Piezo1, a critical mechanosensory ion channel, can detect and convert mechanical forces into chemical signals, importing external stimuli into cells and triggering downstream biological effects. However, the precise role of Piezo1 in VECs, especially in terms of mediating liver regeneration, remains unclear. Here, we report on a potential mechanism by which early changes in hepatic portal hemodynamics activate Piezo1 in VECs to promote hepatocyte proliferation during the process of liver regeneration induced by portal vein ligation in rats. In this liver regeneration model, hepatocyte proliferation is mainly distributed in zone 1 and zone 2 of liver lobules at 24–48 h after surgery, while only a small number of Ki67-positive hepatocytes were observed in zone 3. Activation of Piezo1 promotes increased secretion of epiregulin and amphiregulin from VECs via the PKC/ERK1/2 axis, further activating epidermal growth factor receptor (EGFR) and ERK1/2 signals in hepatocytes and promoting proliferation. In addition, cytokines secreted by Piezo1-activated VECs can induce hepatocytes to undergo epithelial–mesenchymal transition. In the liver lobules, the expression of EGFR in hepatocytes of zone 1 and zone 2 is significantly higher than that in zone 3. The EGFR inhibitor gefitinib inhibits liver regeneration by suppressing the proliferation of hepatocytes in the middle zone. Thus, activation of Piezo1 in VECs promotes hepatocyte proliferation, suggesting mechanical stimulation regulates hepatocyte proliferation in zone 1 and zone 2 during portal vein ligation-induced liver regeneration. These data provide a theoretical basis for the regulation of liver regeneration through chemical signals mediated by mechanical stimulation.

Abstract 5499: Spatially-resolved transcriptomics analyses of liver cancer

Abstract Dissecting and understanding the cancer ecosystem, especially that around the tumor margins, which have strong implications for tumor cell infiltration and invasion, are essential for exploring the mechanisms of tumor metastasis and developing effective new treatments. Using a novel tumor border scanning and digitization model enabled by nanoscale resolution-SpaTial Enhanced REsolution Omics-sequencing (Stereo-seq), we identified a 500 μm-wide zone centered around the tumor border in patients with liver cancer, referred to as “the invasive zone”. We detected strong immunosuppression, metabolic reprogramming, and severely damaged hepatocytes in this zone. We also identified a subpopulation of damaged hepatocytes with increased expression of serum amyloid A1 and A2 (referred to collectively as SAAs) located close to the border on the paratumor side. Overexpression of CXCL6 in adjacent malignant cells could induce activation of the JAK-STAT3 pathway in nearby hepatocytes, which subsequently caused SAAs’ overexpression in these hepatocytes. Furthermore, overexpression and secretion of SAAs by hepatocytes in the invasive zone could lead to the recruitment of macrophages and M2 polarization, further promoting local immunosuppression, potentially resulting in tumor progression. Clinical association analysis in additional five independent cohorts of patients with primary and secondary liver cancer (n = 423) showed that patients with overexpression of SAAs in the invasive zone had a worse prognosis. Further in vivo experiments using mouse liver tumor models in situ confirmed that the knockdown of genes encoding SAAs in hepatocytes decreased macrophage accumulation around the tumor border and delayed tumor growth. The identification and characterization of a novel invasive zone in human cancer patients not only add an important layer of understanding regarding the mechanisms of tumor invasion and metastasis, but may also pave the way for developing novel therapeutic strategies for advanced liver cancer and other solid tumors. Citation Format: Liang Wu. Spatially-resolved transcriptomics analyses of liver cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5499.

Spatial-temporal proliferation of hepatocytes during pregnancy revealed by genetic lineage tracing.

The maternal liver undergoes dramatic enlargement to adapt to the increased metabolic demands during pregnancy. However, the cellular sources for liver growth during pregnancy remain largely elusive. Here, we employed a proliferation recording system, ProTracer, to examine the spatial-temporal proliferation of hepatocytes during pregnancy. We discovered that during early to late pregnancy, hepatocyte proliferation initiated from zone 1, to zone 2, and lastly to zone 3, with the majority of new hepatocytes being generated in zone 2. Additionally, using single-cell RNA sequencing, we observed that Ccnd1 was highly enriched in zone 2 hepatocytes. We further applied dual-recombinase-mediated genetic lineage tracing to reveal that Ccnd1+ hepatocytes expanded preferentially during pregnancy. Moreover, we demonstrated that estrogen induces liver enlargement during pregnancy, which was abolished in Ccnd1 knockout mice. Our work revealed a unique spatial-temporal hepatocyte proliferation pattern during pregnancy, with Ccnd1+ hepatocytes in zone 2 serving as the major cellular source for hepatic enlargement.

An invasive zone in human liver cancer identified by Stereo-seq promotes hepatocyte–tumor cell crosstalk, local immunosuppression and tumor progression

Dissecting and understanding the cancer ecosystem, especially that around the tumor margins, which have strong implications for tumor cell infiltration and invasion, are essential for exploring the mechanisms of tumor metastasis and developing effective new treatments. Using a novel tumor border scanning and digitization model enabled by nanoscale resolution-SpaTial Enhanced REsolution Omics-sequencing (Stereo-seq), we identified a 500 µm-wide zone centered around the tumor border in patients with liver cancer, referred to as “the invasive zone”. We detected strong immunosuppression, metabolic reprogramming, and severely damaged hepatocytes in this zone. We also identified a subpopulation of damaged hepatocytes with increased expression of serum amyloid A1 and A2 (referred to collectively as SAAs) located close to the border on the paratumor side. Overexpression of CXCL6 in adjacent malignant cells could induce activation of the JAK-STAT3 pathway in nearby hepatocytes, which subsequently caused SAAs’ overexpression in these hepatocytes. Furthermore, overexpression and secretion of SAAs by hepatocytes in the invasive zone could lead to the recruitment of macrophages and M2 polarization, further promoting local immunosuppression, potentially resulting in tumor progression. Clinical association analysis in additional five independent cohorts of patients with primary and secondary liver cancer (n = 423) showed that patients with overexpression of SAAs in the invasive zone had a worse prognosis. Further in vivo experiments using mouse liver tumor models in situ confirmed that the knockdown of genes encoding SAAs in hepatocytes decreased macrophage accumulation around the tumor border and delayed tumor growth. The identification and characterization of a novel invasive zone in human cancer patients not only add an important layer of understanding regarding the mechanisms of tumor invasion and metastasis, but may also pave the way for developing novel therapeutic strategies for advanced liver cancer and other solid tumors.

IGFBP2 expressing midlobular hepatocytes preferentially contribute to liver homeostasis and regeneration

Although midlobular hepatocytes in zone 2 are a recently identified cellular source for liver homeostasis and regeneration, these cells have not been exclusively fate mapped. We generated an Igfbp2-CreER knockin strain that specifically labels midlobular hepatocytes. During homeostasis over 1 year, zone 2 hepatocytes increased in abundance from occupying 21%-41% of the lobular area. After either pericentral injury with carbon tetrachloride or periportal injury with 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC), IGFBP2+ cells replenished lost hepatocytes in zones 3 and 1, respectively. IGFBP2+ cells also preferentially contributed to regeneration after 70% partial hepatectomy, as well as liver growth during pregnancy. Because IGFBP2 labeling increased substantially with fasting, we used single nuclear transcriptomics to explore zonation as a function of nutrition, revealing that the zonal division of labor shifts dramatically with fasting. These studies demonstrate the contribution of IGFBP2-labeled zone 2 hepatocytes to liver homeostasis and regeneration.

Immunohistochemical Localization of Alogliptin, a DPP-4 Inhibitor, in Tissues of Normal and Type 2 Diabetes Model Rat.

We investigated the pharmacokinetics of alogliptin (AG) at the cell and tissue level in healthy Wistar rats and a type 2 diabetic Goto-Kakizaki (GK) rat model. Immunohistochemistry of the renal tissue in these rats, post 1 hr of AG administration, showed that the signal was observed in the glomeruli, proximal tubule S3 segments, distal tubules, collecting ducts, and only in the brush border of the epithelial cells of the proximal tubule S1, S2 segments. After 6 hr of AG administration, the staining intensity of the regions other than the S3 segments was considerably reduced in Wistar rats, with no change observed in GK rats. At 24 hr, the staining intensity was considerably reduced, even in GK rats; however, the staining of the S3 segment remained unaltered in both. Hepatocytes in zone III of the hepatic lobule were more intensely stained than those in zone I in Wistar rats at 1 hr. However, almost no staining was observed in the hepatocytes of GK rats at 1 hr. Complete loss of signal was observed in the hepatocytes of the Wistar rats after 6 hr. This study revealed that the pharmacokinetics of AG in GK rats are different from those in Wistar rats.

Massive iron overload and acute-on-chronic liver failure in a patient with Diamond\u2013Blackfan anaemia: a case report

BackgroundGenetic anaemias lead us to reflect on the classic ‘trolley dilemma’, when there are two choices but neither one is satisfactory. Either we do not treat anaemia and the patient suffers from chronic tiredness and fatigue, or we do treat it through blood transfusions, leading to iron overload, which is a quite harmful consequence.Case presentationWe present the case of a 34-year-old woman with Diamond–Blackfan anaemia (DBA). Bone marrow stem cell transplantation had not been accessible during her childhood, so she had been submitted to monthly blood transfusions throughout her life, leading to a hepatitis C virus infection (which was treated, achieving a sustained virological response when she was 18 years old), and secondary haemochromatosis. Despite chelation therapy, diffuse iron deposition was occurring in multiple organs, markedly in the heart and liver. Her serum ferritin was higher than 21,000 ng/mL and transferrin saturation reached 102%. When she faced heart decompensation, this congestive condition led to an acute liver injury overlapping pre-existing hepatic fibrosis. She progressed to haemodynamic and hepatic failure, with clinical features of acute-on-chronic liver failure (ACLF). Despite therapeutic optimisation, she died of respiratory insufficiency. An autopsy was performed and revealed the macroscopic and microscopic findings of a massive iron deposition in the liver, heart, lungs, spleen, bone marrow, thyroid and adrenal glands. We found marked advance of liver fibrosis (chronic damage), as well as necrosis of hepatocytes in zone 3 of the Rappaport acinus (acute damage), supporting the hypothesis of ACLF. The main feature responsible for acute liver decompensation seemed to be heart insufficiency.ConclusionThis is the first case reporting the sequence: DBA, multiple blood transfusions, secondary haemochromatosis, advanced liver fibrosis, heart failure, ACLF and death. A multidisciplinary team is essential to care for DBA patients, since there is a significant emotional burden related to the disease, which might impair an effective chelation therapy and lead to severe consequences due to iron deposition.

Limited proteolysis and oxidative modification of proteins in the hepatocytes of patients with resistant forms of tuberculosis

Abstract The aim of this paper was to evaluate the oxidative modification of proteins and limited proteolysis in the hepatocytes of patients with resistant forms of pulmonary tuberculosis (TB). Material and method: This is a prospective pathomorphological study of 60 patients with confirmed pulmonary TB (clinically and anatomopathologically) deaths from various causes. To evaluate the intensity of free radical processes in hepatocytes, we used a methodology developed by Davydenko IS, 2003. Histological sections were stained with bromphenol blue by Michel-Calvo. Results: When comparing the digital data indicated in our study, it is evident that changes in proteins properties in average tendencies run virtually identically in the hepatocytes of different zones of the comparison group. Thus, the difference is unlikely (p > 0.05) when comparing red/blue ratio (specific staining of acid and basic proteins by Michel-Calvo) in the hepatocytes of zones I, II and III of the comparison group. A similar trend is observed in the analysis of average values of the optical density of a specific colouration on the free amino group of proteins by A. Yasuma and T. Ichikava. The analysis of all parameters showed a linear increase in the intensity of oxidative protein modification and limited proteolysis from zone I to III (from the periportal to the centrolobular parts of the acinus) in all subgroups of the main group according to the acinus zone (by Rappoport). Conclusions: Pulmonary TB leads to increased processes of free radical oxidation of proteins with characteristic effects – increased proteolysis and oxidation of amino groups of proteins – in the hepatocytes of the liver.

Histology and molecular biology studies on the expression and localization of angiopoietin-like protein 8 in human tissues.

Angiopoietin-like protein (ANGPTL) 8 regulates the partitioning of triglycerides by inhibiting lipoprotein lipase in muscle and adipose tissues. ANGPTL8 is expressed in the liver and adipose tissue and secreted into the blood. However, the precise localization of ANGPTL8-expressing cells in these tissues remains unknown. The aim of the present study was to investigate the localization of ANGPTL8-expressing cells in human tissues. Using formalin-fixed paraffin-embedded human tissue specimens, the expression of ANGPTL8 was investigated by reverse transcription-quantitative polymerase chain reaction (RT-qPCR), immunohistochemistry (IHC) and in situ hybridization (ISH). The expression level of ANGPTL8 mRNA was the highest in the liver, followed by lipoma, hibernoma and normal adipose tissue. In the liver, wild type (KF809856) and two splice variants of ANGPTL8 mRNA (KF809857 and KF809858) were found to be expressed. The expression level of the splice variant KF809858, which produces a short form of ANGPTL8, accounted for <1% of ANGPTL8 in the liver. IHC revealed that ANGPTL8 was expressed in hepatocytes in zone 1 of the hepatic acinus in the liver. In the adipose tissue, mature adipocytes weakly expressed ANGPTL8, while immature adipocytes strongly expressed it. ISH confirmed ANGPTL8 mRNA expression in portal hepatocytes and immature adipocytes. ANGPTL8 was expressed in the cells, which actively uptake and metabolize triglycerides. In hibernoma, the ANGPTL8 protein and mRNA were homogeneously expressed in tumor cells. The expression of ANGPTL8 was associated with the differentiation state and activity of lipid metabolism in a subpopulation of cells in the liver and adipose tissue. The association may be helpful for the understanding of local metabolic state in organs and diseases associated with the lipid metabolism.

UEG Week 2016 Poster Presentations

Contact E-mail Address: nakuraokura0922@yahoo.co.jp.IntroductionSinusoidal obstruction syndrome (SOS) is a drug-induced liver injury caused by anticancer treatments, such as oxaliplatin-based chemotherapy. Injury of sinusoidal endothelial cells (SECs) and subsequent extravasated platelet aggregation (EPA) initiate development of SOS. Beraprost sodium (BPS) has a protective effect on SECs and antiplatelet effect. The aim of this study was to examine the protective effect of BPS against SECs injury resulting from monocrotaline (MCT)-induced SOS in mice.Aims & MethodsSOS was induced in Crl:CD1 (ICR) mice by intraperitoneal administration of MCT (270 mg/kg). To evaluate the effect of BPS on SOS, the mice were divided into two groups. In the BPS group, BPS (200 μl/kg) was injected by intraperitoneal administration at 1 hour before, 3 hours after, and 9 hours after MCT administration. The control group received intraperitoneal injections of the same volume of saline at the same time points. All mice were sacrificed at 48 hours after MCT administration and the protective effect on SECs was assessed by determining blood cell count, serum biochemical findings and immunohistochemical stains of sinusoidal endothelial cell antibody (SEC-1) and CD62P (P-selectin). Western blot analysis and real-time polymerase chain reaction (RT-PCR) were also used to examine plasminogen activator inhibitor-1 (PAI-1) and endothelial nitric oxide synthase (eNOS) expression.ResultsPlatelet count was maintained in the BPS group compared to that in the MCT group (48.2 vs. 4.8 ´104/μl, P < 0.001). Serum transaminase levels and hyaluronic acid (HA) in the BPS group were significantly reduced compared to the levels in the MCT group (AST 129 vs. 1013 IU/L, P < 0.05; ALT 90 vs. 923 IU/L, P < 0.01; HA 670 vs. 2243 ng/mL, P < 0.01). In the MCT group, liver histology showed endothelial damage of the central venule, congestion and obstruction of the sinusoids, and necrosis of hepatocytes in zone 3. In the BPS group, these findings were suppressed. Immunohistochemistry showed a protective effect of SECs on SEC-1 and a decrease in platelet infiltration in the space of Disse on CD62P in the BPS group compared to that in the MCT group. Expression of PAI-1 was significantly lower and expression of eNOS was significantly higher in the BPS group.ConclusionBPS preserved SECs and suppressed the progression of MCT-induced SOS in mice, suggesting that BPS is useful for the prevention of SOS.Disclosure of InterestAll authors have declared no conflicts of interest.

“Cardiac Hepatopathy”: A Review of Liver Dysfunction in Heart Failure

The unique dual circulation of the liver confers relative protection against ischemic injury; however, low oxygen tension in the microcirculation (sinusoidal blood of the hepatic acinus) may render hepatocytes in zone 3 relatively vulnerable to ischemic injury and necrosis. Severe congestive heart failure is associated with two distinct forms of liver dysfunction under the umbrella term hepatopathy. The two entities include: jaundice related to passive congestion (congestive hepatopathy from backward cardiac failure) and acute hepatocellular necrosis caused by impaired hepatic perfusion (hypoxic hepatitis from forward cardiac failure). This article provides a comprehensive, up-to-date review on the topic and focuses on the epi- demiology, pathology, pathogenesis, clinical manifestations, diagnostic testing and treatment strategies pertaining to liver disease in circulatory failure.

Weil's disease (leptospirosis) manifesting as fulminant hepatic failure: Report of an autopsy case

We report an autopsy case of a 60-year-old man with Weil's disease who died of fulminant hepatic failure. Ante-mortem blood culture yielded the growth of Leptospira interrogans (serovar icterohaemorrhagiae). At autopsy, the liver weighed 1210g and showed a typical appearance of “acute yellow liver atrophy”. Zone 3 (centrilobular region) showed submassive necrosis of hepatocytes accompanied by marked hemorrhage. Hepatocytes in zones 1 and 2 were well preserved, and the leptospira antigen was immunohistochemically demonstrated in several hepatocytes. Dissociation of liver cell plates was not observed. An immunohistochemical study demonstrated that CD31-positive, sinusoidal endothelial cells had almost completely disappeared in zone 3. This finding suggested that severe and selective damage to endothelial cells in zone 3 was the main cause of the submassive hepatocellular necrosis, which led to fulminant hepatic failure in the present case.

Intrahepatic microcirculatory disorder, parenchymal hypoxia and NOX4 upregulation result in zonal differences in hepatocyte apoptosis following lipopolysaccharide- and D-galactosamine-induced acute liver failure in rats

Although the mechanisms responsible for acute liver failure (ALF) have not yet been fully elucidated, studies have indicated that intrahepatic macrophage activation plays an important role in the pathogenesis of ALF through intrahepatic microcirculatory disorder and consequent parenchymal cell death. Intrahepatic microcirculatory disorder has been demonstrated in animal models using intravital microscopy; however, the limitations of this method include simultaneously evaluating blood flow and the surrounding pathological changes. Therefore, in this study, we devised a novel method involving tetramethylrhodamine isothiocyanate (TRITC)-dextran administration for the pathological assessment of hepatic microcirculation. In addition, we aimed to elucidate the mechanisms through which intrahepatic microcirculatory disorder progresses with relation to activated macrophages. ALF was induced in Wistar rats by exposure to lipopolysaccharide and D-galactosamine. Intrahepatic microcirculation and microcirculatory disorder in zone 3 (pericentral zone) of the livers of rats with ALF was observed. Immunohistochemical examinations in conjunction with TRITC-dextran images revealed that the macrophages were mainly distributed in zone 2 (intermediate zone), while cleaved caspase-3-positive hepatocytes, pimonidazole and hypoxia-inducible factor 1-α were abundant in zone 3. We also found that 4-hydroxy-2-nonenal and nicotinamide adenine dinucleotide phosphate oxidase (NOX)4-positive cells were predominantly located in the zone 3 parenchyma. The majority of apoptotic hepatocytes in zone 3 were co-localized with NOX4. Our results revealed that the apoptotic cells in zone 3 were a result of hypoxic conditions induced by intrahepatic microcirculatory disorder, and were not induced by activated macrophages. The increased levels of oxidative stress in zone 3 may contribute to the progression of hepatocyte apoptosis.

Interaction between Kupffer cells and platelets in the early period of hepatic ischemia–reperfusion injury—An in vivo study

BackgroundHepatic ischemia–reperfusion (I/R) leads to activation of Kupffer cells (KCs). The activated KCs cause platelet and leukocyte adhesion to the sinusoidal endothelium. Previously, we reported that platelet–endothelium interactions occur earlier than leukocyte responses. The aim of this study was to evaluate the interaction between platelets and KCs in the hepatic microcirculation after I/R. Materials and methodsSprague-Dawley rats were divided into three groups: the no-ischemia group (control group; n = 6); the 20-min ischemia group (I/R group; n = 6); and the 20-min ischemia + anti-rat platelet serum group (APS group; n = 6). KCs were labeled using the liposome entrapment method. The number of adherent platelets was observed for up to 120 min after reperfusion by intravital microscopy. To investigate the effects of platelets on I/R injury, rats were injected intravenously with rabbit APS for platelet depletion. ResultsIn the I/R group, the number of adherent platelets increased significantly after I/R. More than 50% of the adherent platelets adhered to KCs. Electron microscopy indicated that the platelets attached to the KCs after hepatic ischemia. The histologic findings indicated liver damage and apoptosis of hepatocytes in zone 1. In the I/R group, but not in the control and APS groups, serum ALT increased immediately after reperfusion. ConclusionsWe succeeded in visualizing the dynamics of both KCs and platelets in the hepatic sinusoids. Liver ischemia induced the adhesion of platelets to KCs in the early period, which could play a key role in reperfusion injury of the liver.

Hepatic STAT1-Nuclear Translocation and Interleukin 28B Polymorphisms Predict Treatment Outcomes in Hepatitis C Virus Genotype 1-Infected Patients

BackgroundWe investigated associations between signal transducer and activator of transcription (STAT) 1 in pretreated liver tissues, interleukin (IL) 28B polymorphism and treatment response in hepatitis C virus (HCV)-infected patients treated with peginterferon and ribavirin.Methods and FindingsWe performed immunostaining analysis of STAT1 in liver tissues and determined IL28B polymorphism at rs8099917. We then compared the results with treatment outcomes in HCV genotype 1 patients with high viral load who were receiving peginterferon plus ribavirin. In univariate analysis, younger age, white blood cell counts, virological responder, early virological responder (EVR), mild activity (A1) of liver inflammation grading, and lower STAT1 nuclear-stain of hepatocytes in zone 1, zone 2 and total zones of liver were associated with sustained virological responder (SVR). Multivariate analysis showed that EVR, age and hepatic STAT1 nuclear-stain in zone 2 of liver were independent predictors of SVR. It was also revealed that IL28B and STAT1-nuclear translocation in hepatocytes are independent predictors of response to treatment with peginterferon and ribavirin in chronic hepatitis C patients.ConclusionsConcomitant assessment of lower STAT1 nuclear-stain of hepatocytes and IL28B polymorphism is useful for prediction of SVR in HCV genotype 1 patients.

Hepatic glutamate metabolism: a tale of 2 hepatocytes

Glutamate plays a central role in hepatic amino acid metabolism, both because of its role in the transdeamination of most amino acids and because the catabolism of arginine, ornithine, proline, histidine, and glutamine gives rise to glutamate. It is now appreciated that different hepatic functions are restricted to hepatocyte subpopulations within different acinar zones. This is also a feature of glutamate metabolism. Glutamine catabolism and synthesis are physically separated by zonation, with glutamine synthetase restricted to a narrow band of hepatocytes in zone 3 of the hepatic acinus, whereas glutaminase occurs in zone 1. Arginine and ornithine metabolism is also restricted to particular hepatocyte subpopulations. Ornithine aminotransferase, the regulated enzyme of arginine and ornithine catabolism, is restricted to the same zone 3 cells as glutamine synthetase, whereas the urea cycle is found in the remaining hepatocytes. This separation facilitates the independent regulation of these 2 different metabolic processes. We know the acinar localization of only a small fraction of the ≈15,000 genes expressed in the liver. Knowledge of the acinar localization of metabolic processes is essential for an appreciation of their relation to other hepatic functions and their regulation.

Elevated oxidative stress, iron accumulation around microvessels and increased 4-hydroxynonenal immunostaining in zone 1 of the liver acinus in hypercholesterolemic rabbits

Rabbits were fed a diet containing 1% cholesterol for 8 weeks and the levels of iron and oxidized lipids in liver analysed using atomic absorption spectroscopy and gas chromatography-mass spectrometry. A non-significant trend to an increase in iron level, but significant increases in the lipid peroxidation products, F2-isoprostanes and the cholesterol oxidation products 7 beta hydroxycholesterol, 7 ketocholesterol and cholesterol 5,6-alpha epoxide were detected in the liver of the cholesterol-fed rabbits. Histological analysis showed greater accumulation of lipids by Sudan red labelling in hepatocytes of zone I than zones II and III of the liver acinus. The increase in lipids coincided with an increase in iron staining in macrophages around liver microvessels and increased immunostaining to melanotransferrin and the lipid peroxidation product, 4-hydroxynonenal (4-HNE), in zone 1. The results are suggestive of microvascular damage associated with iron accumulation and oxidative stress in the liver during hypercholesterolemia.

Biliary excretion of bile acids and organic anions in rats with dichloroethylene-induced bile canalicular injury

Hepatocytes in zone 1 of the hepatic lobule play a role in the uptake and biliary excretion of bile acids and organic anions under physiological conditions, and hepatocytes in zone 3 may play a role only when there is a high-dose load. To further elucidate the role of hepatic zonation in the hepatic handling of bile acids and organic anions, the biliary excretion of these compounds was studied in rats with dichloroethylene (DCE)-induced selective zone 3 bile canalicular injury. Biliary excretion of various bile acids and organic anions was studied in rats 1 h after oral administration of DCE (5 mg/100 g). The effect of DCE on the immunostaining of multidrug resistance protein 2 (Mrp2; an important canalicular organic anion transporter) in the liver was also examined. The biliary excretory maximum of taurocholate and tauroursodeoxycholate was decreased in DCE-treated rats, whereas the biliary excretion of taurolithocholate-sulfate and phenolphthalein-glucuronide was unchanged in DCE-treated rats, and DCE treatment decreased the biliary excretion of sulfobromophthalein and pravastatin. DCE decreased Mrp2 staining in the canalicular membrane of zone 3 hepatocytes on immunohistochemistry. These findings indicate that canalicular transport in zone 3 hepatocytes is important in the biliary excretion of bile acids and organic anions, when they are administered at high doses.

Microwave cell death: Immunohistochemical and enzyme histochemical evaluation.

In Japan, microwave coagulation therapy (MCT) has been used for the management of primary and metastatic liver cancer. Needle biopsy examination from the lesion has frequently shown the presence of nucleated cancer cells in histopathological examinations, prompting the conclusion that cancer cells are not completely eliminated by microwave therapy, whereas computed tomography and ultrasonography examinations show tumor regression. To determine whether microwave-treated tissue contains functionally viable cells, an examination of the Na+-K+-ATPase protein and its activity using immunohistochemical and enzyme histochemical methods were carried out in microwave-treated rat liver. Four concentric, morphologically identifiable zones around the microwave probe needle appeared 2 days after treatment. Zone A, which was between the innermost spongy zone and the outer necrotic zone, contained only slight morphological alterations in the hepatocytes, which had slightly hyperchromatic nuclei and mildly eosinophilic cytoplasm. The hepatocytes in zone A were found to be positive for the Na+-K+-ATPase antigenicity but negative for enzyme activity, indicating that zone A was undergoing cell death, although morphologically this was not discernible. This type of cell death caused by microwave treatment is morphologically different from previously known types of cell death, either oncosis or apoptosis.

Liver and kidney function and histology in rats exposed to cadmium and ethanol.

The present study was performed to assess the function and histology of the liver and kidney in rats exposed to 50 mg Cd/l (as cadmium chloride) and/or 10% (w/v) ethanol (EtOH) for 12 weeks. The activities of alanine aminotransferase (ALAT) and asparate aminotransferase (AspAT) in serum were measured as indicators of the liver function. As parameters of the kidney function, creatinine, total protein and urea concentrations in serum and urine, as well as urinary alkaline phosphatase (ALP) activity were determined, and creatinine clearance was calculated. Both organs were subjected to histopathological analysis. Daily Cd intake ranged from 3.17 to 4.28 mg/kg body weight and from 2.41 to 3.17 mg/kg body weight in the Cd and Cd + EtOH groups, respectively. The daily intake of 10% EtOH ranged from 47.5 to 86.9 g/kg body weight in the EtOH and from 47.3 to 63.4 g/kg body weight in the Cd + EtOH-exposed rats. Cd and EtOH, independently of separate or combined application, changed liver and kidney function and histology. Rats treated with Cd alone and those co-exposed to both substances showed qualitatively similar, but different magnitudes of changes, in liver and kidney histology. Blurred trabecular structure, vacuolar degeneration and increased density of nuclear chromatin with very compact nuclear structure were found in hepatocytes of zones 2 and 3. Moreover, mononuclear cell infiltrations and necrosis of single cells were evident in zone 1. In the kidney tubules, degeneration and hypertrophy of epithelial cells and dilation in the glomeruli were also observed. Some functional (increased serum AspAT and urinary ALP, decreased urinary urea) and structural changes in the liver and kidney were more evident in the case of combined exposure, while others were more evident after single exposure. However, a decrease in creatinine clearance, noted only in the animals treated with Cd and EtOH, shows that functional changes indicating renal insufficiency are more serious in the co-exposed group. Due to lower Cd and EtOH intake (resulting from a stronger aversion to drinking water containing both substances) in the co-exposed rats, as compared to the Cd- and EtOH-treated groups, it is difficult to draw a definite conclusion from this study. The findings, however, seem to indicate that EtOH increases Cd nephrotoxicity in rats, and thus may suggest a higher risk of kidney damage in alcoholics exposed to Cd. Unfortunately, this study does not provide clear evidence if, and to what extent, EtOH influences Cd hepatotoxicity.