Ultrastructural Profile Combined with Immunohistochemistry of a Hepatic Progenitor Cell Line in Pediatric Autoimmune Hepatitis: New Insights into the Morphological Pattern of the Disease.

Considering that the heterogenic population of a hepatic progenitor cell line (HPCL) can play a vital role in autoimmune hepatitis (AIH), we decided to conduct pioneering retrospective evaluation of these cells in pediatric AIH by means of transmission electron microscopy (TEM). The aim of the study was to assess the ultrastructure of the HPCL in children with untreated AIH. Ultrastructural analysis of the HPCL population, preceded by immunohistochemical staining for cytokeratin 7 (CK7), was performed using pretreatment liver biopsies from 23 children with clinicopathologically diagnosed AIH. Immunohistochemical assessment for CK7 allowed detection of proliferating immature epithelial cells differentiating towards periportal and intralobular intermediate hepatocytes without marked formation of ductular reactions in AIH children. Using TEM, we distinguished three morphological types of HPCs: I-the most undifferentiated progenitor cells; III-intermediate hepatocyte-like cells; II-intermediate bile duct cells. Most frequent were the cells differentiating towards hepatocytes, most rare-those differentiating towards cholangiocytes. The results indicate that an HPCL may be an important source of hepatocyte regeneration. Ultrastructural analyses of the HPCL population, combined with immunohistochemistry for CK7, might be a useful tool to evaluate liver cell regeneration, including fibrogenesis, and may help better understand the morphological pattern of the disease, in pediatric AIH. Frequent appearance of an HPCL in the vicinity of fibrotic foci, often accompanied by hyperactive Kupffer cells and transitional hepatic stellate cells, may indicate their significant involvement in liver fibrogenesis.

Hepatic and Extrahepatic Sources and Manifestations in Endoplasmic Reticulum Storage Diseases.

Alpha-1-antitrypsin (AAT) and fibrinogen are secretory acute phase reactant proteins. Circulating AAT and fibrinogen are synthesized exclusively in the liver. Mutations in the encoding genes result in conformational abnormalities of the two molecules that aggregate within the rough endoplasmic reticulum (RER) instead of being regularly exported. That results in AAT-deficiency (AATD) and in hereditary hypofibrinogenemia with hepatic storage (HHHS). The association of plasma deficiency and liver storage identifies a new group of pathologies: endoplasmic reticulum storage disease (ERSD).

Biochemical and ultrastructural changes in kidney and liver of African Giant Rats (Cricetomysgambianus, Waterhouse, 1840) exposed to intraperitoneal sodium metavanadate (vanadium) intoxication.

We studied the hepatic and renal impact of sodium metavanadate (SMV) exposure in African giant rats (AGR). Twelve male AGR were used and divided into two groups. The control group received sterile water while the SMV-exposed group received 3 mg/kg SMV intraperitoneally for 14 days. SMV exposed AGR groups showed significantly decreased activities of serum AST, ALT, ALP and creatinine concentration but increased blood urea nitrogen (BUN), albumin and globulin concentrations. Kidney ultrastructure examination revealed atrophy of the glomerular tuft, loss of podocytes, distortions of the endothelium and glomerular basement membrane. The liver sinusoids fenestration phenotypes were abnormal. Hepatocytes exhibited hypertrophy with uneven, crenated and dentate nuclei. SMV exposure induced activation of monocytes, as well as Kupffer and fibrous cells. Alterations in glomerular podocytes and cell-cell and cell matrix contact and inflammatory liver fibrosis are key events in progressive glomerular failure and hepatic damage due to SMV intoxication.

Observation and characterisation of macrophages in zebrafish liver.

Kupffer cells are liver-resident macrophages that play an important role in mediating immune-related functions in mammals and humans. They are well-known for their capacity to phagocytose large amounts of waste complexes, cell debris, microbial particles and even malignant cells. Location, appearance and functional aspects are important features used to identify these characteristic cells of the liver sinusoid. To-date, there is limited information on the occurrence of macrophages in zebrafish liver. Therefore, we aimed to characterise the ultrastructural and functional aspects of liver-associated macrophages in the zebrafish model by taking advantage of the latest advances in zebrafish genetics and multimodal correlative imaging. Herein, we report on the occurrence of macrophages within the zebrafish liver exhibiting conventional ultrastructural features (e.g. presence of pseudopodia, extensive lysosomal apparatus, a phagolysosome and making up ∼3% of the liver volume). Intriguingly, these cells were not located within the sinusoidal vascular bed of hepatic tissue but instead resided between hepatocytes and lacked phagocytic function. While our results demonstrated the presence and structural similarities with liver macrophages from other experimental models, their functional characteristics were distinctly different from Kupffer cells that have been described in rodents and humans. These findings illustrate that the innate immune system of the zebrafish liver has some distinctly different characteristics compared to other animal experimental models. This conclusion underpins our call for future studies in order to have a better understanding of the physiological role of macrophages residing between the parenchymal cells of the zebrafish liver.

Impact of renal ischemia/reperfusion injury on the rat Kupffer cell as a remote cell: A biochemical, histological, immunohistochemical, and electron microscopic study.

Almost all transplanted solid organs are exposed to some degree of ischemia-reperfusion (IR) damage. It is interesting to know that this IR damage affects various remote tissues including the liver and resulted in serious adverse effects. Liver injury triggers different responses of liver tissue especially Kupffer cells (KCs). The goal of this current study is to assess the biochemical and morphological changes of hepatic KCs after the induction of renal ischemia-reperfusion (RIR) and point out their role in remote liver injury after RIR. Sixteen male Sprague-Dawley rats were randomly divided into two equal groups: Group I; sham group. Group II; renal ischemia reperfusion (IR) group in which rats were exposed to renal ischemia for 45 min followed by renal reperfusion for 48 h. Three rats from each group were subjected to charcoal injection to evaluate KCs activity. Specimens of rat liver from each group were obtained and processed for biochemical, light microscopic and ultramicroscopic examination. The current results showed elevated serum levels of AST and ALT. The liver HGF-α protein expression increased in IR group compared to the sham group. In IR group, numerous charcoal labeled KCs were observed mainly localized around the central vein. Scanning electron micrographs showed complex primary and secondary foot process of the KCs. Ultrastructural study showed KCs with multiple cytoplasmic vacuoles, lysosomes and mitochondria, rough endoplasmic reticulum and ribosomes. Immuno-histochemical study showed more tumor necrosis factor-α (TNF-α) expression in KCs than the sham group. These results collectively demonstrated that renal IR produced biochemical and morphological changes in the liver KCs and theses cells might have a role in the remote liver injury after renal IR. This might be one of the mechanisms through which RIR affects the liver.

Dietary Lipids Differentially Shape Nonalcoholic Steatohepatitis Progression and the Transcriptome of Kupffer Cells and Infiltrating Macrophages.

A crucial component of nonalcoholic fatty liver disease (NAFLD) pathogenesis is lipid stress, which may contribute to hepatic inflammation and activation of innate immunity in the liver. However, little is known regarding how dietary lipids, including fat and cholesterol, may facilitate innate immune activation in vivo. We hypothesized that dietary fat and cholesterol drive NAFLD progression to steatohepatitis and hepatic fibrosis by altering the transcription and phenotype of hepatic macrophages. This hypothesis was tested by using RNA-sequencing methods to characterize and analyze sort-purified hepatic macrophage populations that were isolated from mice fed diets with varying amounts of fat and cholesterol. The addition of cholesterol to a high-fat diet triggered hepatic pathology reminiscent of advanced nonalcoholic steatohepatitis (NASH) in humans characterized by signs of cholesterol dysregulation, generation of oxidized low-density lipoprotein, increased recruitment of hepatic macrophages, and significant fibrosis. RNA-sequencing analyses of hepatic macrophages in this model revealed that dietary cholesterol induced a tissue repair and regeneration phenotype in Kupffer cells (KCs) and recruited infiltrating macrophages to a greater degree than fat. Furthermore, comparison of diseased KCs and infiltrating macrophages revealed that these two macrophage subsets are transcriptionally diverse. Finally, direct stimulation of murine and human macrophages with oxidized low-density lipoprotein recapitulated some of the transcriptional changes observed in the RNA-sequencing study. These findings indicate that fat and cholesterol synergize to alter macrophage phenotype, and they also challenge the dogma that KCs are purely proinflammatory in NASH. Conclusion: This comprehensive view of macrophage populations in NASH indicates mechanisms by which cholesterol contributes to NASH progression and identifies potential therapeutic targets for this common disease.

Cellular and stromal elements organization in the liver of grass carp, Ctenopharyngodon idella (Cypriniformes: Cyprinidae).

Fish liver is considered as a key organ that controls various life functions. The cellular and stromal elements of liver of eighteen specimens of adult grass carp were investigated by light- and electron- microscopy and enzyme histochemistry. The liver was formed of two lobes with a long process extended from the right lobe. Serial paraffin section of the liver identified different kinds of vascular- biliary structures as follows: 1) pancreatic-venous-biliary-arteriolar tracts (P-VBAT); 2) venous-biliary-arteriolar tracts, (VBAT); 3) pancreatic-venous-biliary tracts (P-VBT); 4) venous-biliary tracts (VBT); 5) venous-arteriolar tracts (VAT); 6) isolated veins named as venous tracts (VT); 7) isolated bile ducts, named as biliary tracts (BT); 8) biliary-arteriolar tracts (BAT); 9) pancreatic-biliary tracts (P-BT); 10) pancreatic- venous tracts (P-VT). Macrophages aggregates were associated with VBT and P-BT. The hepatic parenchyma was consisted of many populations of cells. Histochemically, the hepatocytes were strongly reacted with PAS, and Best's carmine. Moreover, strong staining patterns for acid phosphatase, ATPase, and alkaline phosphatase were demonstrated in hepatocytes. The hepatic satellate (Ito) cells were observed in the space of Disse and between hepatocytes. Rodlet cells and eosinophilic granular/ mast cells were encountered in the liver of grass carp. The sinusoids were lined by fenestrated endothelial cells and Kupffer cells. Moreover, dendritic-like cells were demonstrated in the sinusoids and perisinusoidal connective tissue. The biliary duct system was constituted of bile canaliculi, ductules, and bile ducts. Telocytes with their characteristics telopodes were located around bile ducts. The current findings are offering fundamental data on histology of grass carp liver.

Probing the unseen structure and function of liver cells through atomic force microscopy.

With the arrival of atomic force microscopy (AFM) about thirty years ago, this new imaging tool opened up a new area for the exploration of biological samples, ranging from the tissue and cellular level down to the supramolecular scale. Commercial instruments of this new imaging technique began to appear in the five years following its discovery in 1986 by Binnig, Quate & Gerber. From that point onwards the AFM has attracted many liver biologists, and the number of publications describing structure-function relationships on the diverse set of liver cells has grown steadily ever since. It is therefore timely to reflect on the achievements of AFM in disclosing the cellular architecture of hepatocytes, liver sinusoidal endothelial cells, Kupffer cells, stellate cells and liver-associated natural killer cells. In this thematic paper, we present new data and provide an in-depth overview of the current AFM literature on liver cell biology. We furthermore include a future outlook on how this scanning probe imaging tool and its latest developments can contribute to clarify various structural and functional aspects of cells in liver health and disease.

Glassy droplet inclusions within the cytoplasm of Kupffer cells: A novel ultrastructural feature for the diagnosis of pediatric autoimmune hepatitis.

Since Kupffer cells/macrophages (KCs/MPs) may be involved in the pathogenesis of autoimmune hepatitis (AIH), this pioneer study was undertaken to evaluate KCs/MPs in pediatric AIH in transmission-electron microscope. METHODS: Ultrastructural analyses were performed using liver biopsies from 14 children with clinicopathologically diagnosed AIH. RESULTS: In all AIH children, ultrastructural findings revealed changes in the cells lining sinusoidal vessels, especially KCs/MPs and endothelial cells. KCs/MPs showed increased phagocytic activity and damaged mitochondria, frequently with accompanying intense fibrosis. In 10/14 AIH patients, the cytoplasm of sinusoidal KCs/MPs contained characteristic glassy droplet inclusions. They were round, sharply circumscribed, and contained homogeneous material and distinct translucent fields. Their ultrastructure was identical with the Russel bodies of plasma cells, which were also found in liver biopsies in the same patients. CONCLUSION: Ultrastructural identification of characteristic cytoplasmic droplets with glassy appearance in KCs/MPs, never before described in AIH, provides a useful novel morphological feature in the diagnosis of this disease.

[Inhibition of nuclear factor kappa B mediated by miRNA plasmid carried by novel polyethyleneimine nanogel particles in Kupffer cells of rats].

OBJECTIVE: To test a novel gene carrier, named polyethyleneimine nanogel particles (PEI-NP), for delivering the mircroRNA (miRNA) into Kupffer cells (KCs) for silencing the expression of nuclear transcription factor κB (NF-κB) P65. METHODS: The capacity of PEI-NPs to carry miRNA was tested by gel electrophoresis. RAW264.7 cell line was transfected with the cationic polymers which mixed together PEI-NP and miRNA. Then, the cytotoxicity of the RAW264.7 cells was detected by cell counting kit-8 (CCK-8); the cell apoptosis and transfection efficiency were analyzed by flow cytometry combined with annexin V-FITC/PI staining; the gene and protein levels of NF-κB P65 were determined respectively by real-time quantitative PCR (qRT-PCR) and Western blotting. The distribution of the cationic polymers in RAW264.7 cells and liver KCs was observed by transmission electron microscope (TEM). The protein level of NF-κB P65 in KCs was detected by immunohistochemistry. RESULTS: PEI-NPs were able to completely combine with miRNA at the least mass ratio of 20:1. The highest transfection efficiency of 48 hours was (33.63±1.94)% in vitro. The protein level of NF-κB P65 at 48 hours of PEI-NP/miRNA transfection into RAW264.7 cells was significantly inhibited. TEM showed the cationic polymers in RAW264.7 cytoplasm and only in KCs of liver tissue after transfection. Immunohistochemistry indicated that the protein expression of NF-κB P65 in KCs was significantly lower than that in control group. CONCLUSION: Macrophages and KCs were successfully transfected with the cationic polymers in vitro and in vivo, respectively. The expression of NF-κB P65 was knocked down significantly after transfection.

Liver gender dimorphism--insights from quantitative morphology.

It was shown recently that many genes are differentially expressed in the liver of males and females, thus strengthening the concept of liver gender dimorphism. This dimorphism exists in many pathological scenarios, from regeneration to fibrosis, which has led to the development of gender hepatology. Nevertheless, it is still unknown if gender dimorphism occurs in the structure of the normal liver. In recent years, it has been shown that, compared with male, the female rat liver bears less fibrotic tissue, more Kupffer cells (per volume unit) and has higher hepatocellularity, including binucleated hepatocytes (per volume unit). Our hypothesis is that the human liver also hides a gender dimorphic pattern. Baseline differences in fibrotic tissue would contribute to explain severe liver fibrosis in men. As to the disparity of Kupffer cells, this would clarify the stronger response to post-surgery infections in women, and it could be equated when appraising the higher susceptibility to alcohol. Regarding differences in hepatocytes, they not only justify existing differences in some liver parameters (e.g., transaminases and bilirubin), but they could also account for the higher regenerative potential of the female liver. The structural dimorphism in the human liver would sustain the concept of gender hepatology and, eventually, should be considered in the context of liver transplantation.

Loss of lysosomal membrane protein NCU-G1 in mice results in spontaneous liver fibrosis with accumulation of lipofuscin and iron in Kupffer cells.

Human kidney predominant protein, NCU-G1, is a highly conserved protein with an unknown biological function. Initially described as a nuclear protein, it was later shown to be a bona fide lysosomal integral membrane protein. To gain insight into the physiological function of NCU-G1, mice with no detectable expression of this gene were created using a gene-trap strategy, and Ncu-g1(gt/gt) mice were successfully characterized. Lysosomal disorders are mainly caused by lack of or malfunctioning of proteins in the endosomal-lysosomal pathway. The clinical symptoms vary, but often include liver dysfunction. Persistent liver damage activates fibrogenesis and, if unremedied, eventually leads to liver fibrosis/cirrhosis and death. We demonstrate that the disruption of Ncu-g1 results in spontaneous liver fibrosis in mice as the predominant phenotype. Evidence for an increased rate of hepatic cell death, oxidative stress and active fibrogenesis were detected in Ncu-g1(gt/gt) liver. In addition to collagen deposition, microscopic examination of liver sections revealed accumulation of autofluorescent lipofuscin and iron in Ncu-g1(gt/gt) Kupffer cells. Because only a few transgenic mouse models have been identified with chronic liver injury and spontaneous liver fibrosis development, we propose that the Ncu-g1(gt/gt) mouse could be a valuable new tool in the development of novel treatments for the attenuation of fibrosis due to chronic liver damage.

Kupffer cell structure in the juvenile Nile crocodile, Crocodylus niloticus.

The morphology of Kupffer cells was examined in the liver of the juvenile Nile crocodile using light microscopy and transmission electron microscopy. Pleomorphic Kupffer cells were located in the sinusoids, in the space of Disse, in the hepatic parenchyma and often connected adjacent sinusoids. The cell surfaces were irregular due to the presence of filopodia and lamelliapodia with phagocytosis of white blood cells, red blood cells and thrombocytes being evident. The cells were in close contact with endothelial cells and pit cells in the sinusoidal lumen and with stellate cells in the space of Disse. The cytoplasm contained large phagosomes comprising a combination of ceroid pigment, melanosomes and siderosomes. The nuclei were often indented and eccentrically placed due to the presence of the phagosomes. Conspicuous clusters of membrane-bound tubular organelles with a filamentous or crystalline interior were observed in the cytoplasm. The clusters were sometimes separated into smaller groups around phagosomes. A clear zone existed between the limiting membrane and the interior of these tubular organelles with the electron-dense interior profiles being, respectively, circular, angular or divided. The tubular organelles have not previously been described in Kupffer cells and possibly represent lysosomes with specialized functions. Mitochondria, microtubules, Golgi profiles, granular and smooth endoplasmic reticulum, and a few cytoplasmic lipid droplets were also present. The presence of the tubular organelles and the occurrence of the Kupffer cells in different locations in the liver of the juvenile Nile crocodile are indicative of particularly active and mobile cells.

Morphological and functional characterization of non-alcoholic fatty liver disease induced by a methionine-choline-deficient diet in C57BL/6 mice.

BACKGROUND: Non-alcoholic fatty liver disease (NAFLD), including non-alcoholic steatohepatitis (NASH), appears to be increasingly common worldwide. Its histopathology and the effects of nutrition on liver function have not been fully determined. AIM: To elucidate the cellular mechanisms of NAFLD induced by a methionine-choline-deficient (MCD) diet in mice. Particular focus was placed on the role of phagocytic cells. METHODS: Male C57BL/6 mice were fed an MCD diet for 30 weeks. A recovery model was also established wherein a normal control diet was provided for 2 weeks after a period of 8, 16, or 30 weeks. RESULTS: Mice fed the MCD diet for ≥ 2 weeks exhibited severe steatohepatitis with elevated serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels. Steatohepatitis was accompanied by the infiltration of CD68-positive macrophages (Kupffer cells). The severity of steatohepatitis increased in the first 16 weeks but was seen to lessen by week 30. Fibrosis began to develop at 10 weeks and continued thereafter. Steatohepatitis and elevated serum hepatic enzyme concentrations returned to normal levels after switching the diet back to the control within the first 16 weeks, but fibrosis and CD68-positive macrophages remained. CONCLUSIONS: The histopathological changes and irreversible fibrosis seen in this model were caused by prolonged feeding of an MCD diet. These results were accompanied by changes in the activity of CD68-positive cells with temporary elevation of CCL-2, MMP-13, and MMP-9 levels, all of which may trigger early steatohepatitis and late fibrosis through phagocytosis-associated MMP induction.

The mechanism of anti-CD20-mediated B cell depletion revealed by intravital imaging.

Anti-CD20 Ab therapy has proven successful for treating B cell malignancies and a number of autoimmune diseases. However, how anti-CD20 Abs operate in vivo to mediate B cell depletion is not fully understood. In particular, the anatomical location, the type of effector cells, and the mechanism underlying anti-CD20 therapy remain uncertain. Here, we found that the liver is a major site for B cell depletion and that recirculation accounts for the decrease in B cell numbers observed in secondary lymphoid organs. Using intravital imaging, we established that, upon anti-CD20 treatment, Kupffer cells (KCs) mediate the abrupt arrest and subsequent engulfment of B cells circulating in the liver sinusoids. KCs were also effective in depleting malignant B cells in a model of spontaneous lymphoma. Our results identify Ab-dependent cellular phagocytosis by KCs as a primary mechanism of anti-CD20 therapy and provide an experimental framework for optimizing the efficacy of therapeutic Abs.

[Dynamics of ultrastructural organization of liver cells in experimental peritonitis].

In the experiment on Wistar rats by methods of light and electron microscopy an ultrastructural disorders of the liver cells in various stages of peritonitis were investigated. It was shown that the reactive phase of experimental peritonitis associates with dystrophic changes of the hepatocytes and the endotheliocytes of sinusoids. These reversible changes are explained by an adaptive response of cells to inflammation. In the toxic and especially in the terminal stages of peritonitis in submicroscopical architectonics of the hepatocytes, Kupffer cells and sinusoidal endotheliocytes, along with dystrophic changes appear destructive violations of organelles of hepatic cells that acquired an irreversible character. The proposed model let description of ultrastructural and functional disorders of the liver cells at different stages of peritonitis.

The role of Kupffer cells in the morphogenesis of nonalcoholic steatohepatitis - ultrastructural findings. The first report in pediatric patients.

OBJECTIVE: Until now studies concerning the involvement of hepatic nonparenchymal cells (NPCs), particularly Kupffer cells/macrophages (KCs/MPs), in the pathogenesis of human nonalcoholic steatohepatitis (NASH) have been limited to adult patients; there are no similar reports referring to children. This study aimed to explore, based on ultrastructural analysis, the role of KCs/MPs in the morphogenesis of nonalcoholic steatohepatitis (NASH) in children. MATERIAL AND METHODS: Ultrastructural investigations of KCs were conducted on liver bioptates obtained from 10 children, aged 2-14 years, with clinicopathologically diagnosed NASH. Bioptatic material was fixed in solution of paraformaldehyde and glutaraldehyde in cacodylate buffer, routinely processed for transmission-electron microscopic analysis and examined using an Opton EM microscope. RESULTS: The current ultrastructural study revealed within the hepatic sinusoids the presence of numerous enlarged KCs with increased phagocytic activity, which reduced or blocked vascular lumen. Interestingly, the activated KCs not only contained primary and secondary lysosomes, altered mitochondria, and well-developed Golgi apparatus, but also absorbed fragments of erythrocytes. Such macrophages were frequently seen very close to the transformed hepatic stellate cells (T-HSCs) and progenitor/oval cells. Intensive fibrosis was observed in the vicinity of activated KCs/MPs. Bundles of collagen fibers were seen directly adhering to these cells and to other NPCs, especially T-HSCs. CONCLUSIONS: KCs are involved in the morphogenesis and development of pediatric NASH. Engulfment of erythrocytes by hepatic macrophages may lead to the accumulation of iron derived from hemoglobin in liver and play a role in triggering the generation of oxidative stress in the disease course.

Immunocytochemistry for vancomycin using a monoclonal antibody that reveals accumulation of the drug in rat kidney and liver.

We prepared monoclonal antibodies against N-(γ-maleimidobutyryloxy)succinimide-conjugated vancomycin (VM). The monoclonal antibody was specific for conjugated or free VM. The monoclonal antibody enabled us to develop an immunocytochemical method for detecting the uptake of VM in the rat kidney and liver. Three hours after a single intravenous (i.v.) injection of VM at the therapeutic dose, the immunocytochemistry revealed that VM accumulated in large amounts in both the S1 and S2 segments and in much smaller amounts in the S3 segment of the proximal tubules as well as in the distal tubules and collecting ducts. The drug was detected in the cytoplasm, cytoplasmic irregular granules, nuclei, and microvilli of the proximal tubule cells. The distal tubules and collecting ducts contained scattered swollen cells in which both the nuclei and cytoplasm were heavily immunostained. Twenty-four hours after injection, most of the swollen cells returned back to normal size and had somewhat decreased immunostaining. Also, significant amounts of VM remained accumulated for as long as 8 days postadministration. In the liver, similar drug accumulation was observed in the Kupffer cells and the endothelial cells of the hepatic sinusoids but not in the hepatocytes, suggesting that vancomycin cannot be eliminated via the liver. Immunoelectron microscopic studies demonstrated that in the collecting ducts, uptake of VM occurred exclusively in the lysosomes and cytoplasm of the principal cells and scarcely in the intercalated cells. Furthermore, double fluorescence staining using rats simultaneously administered with VM and gentamicin strongly suggests that both drugs colocalized in lysosomes in the proximal tubule cells of kidneys.

Electron microscopy of rat liver after intravenous injection of nanosized magnetite suspension.

Rat liver was examined by transmission electron microscopy after a single intravenous injection of nanosized magnetite suspension (0.1 g (Fe(3)O(4))/kg body weight). Magnetite particles were found in Kupffer's cells and hepatocytes. Accumulation of the particles by these two cell types was different. Morphometry of magnetite-containing granules in Kupffer's cells and nanoparticle agglomerations in hepatocytes was carried out. The ultrastructure of Kupffer's cell granules was described and the mechanism of penetration of nanosized magnetite particles into the cells was suggested. Nanosized magnetite particles were not completely eliminated over 40 days after a single injection.

Interaction between Kupffer cells and platelets in the early period of hepatic ischemia-reperfusion injury--an in vivo study.

BACKGROUND: Hepatic 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 METHODS: Sprague-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. RESULTS: In 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. CONCLUSIONS: We 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.