Immunohistochemical analyses of the kinetics and distribution of macrophages, hepatic stellate cells and bile duct epithelia in the developing rat liver

Golbar, Hossain M.; Izawa, Takeshi; Murai, Fumi; Kuwamura, Mitsuru; Yamate, Jyoji

doi:10.1016/j.etp.2010.05.011

Cited by 28 publications

(41 citation statements)

References 39 publications

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“…Over-digestion resulted in a low yield of viable cells, while under-digestion would made it difficult to separate the cells [26]. The liver is comprised of parenchymal hepatocytes and non-parenchymal cells [27], and density as well as cell size are significantly different between the cell types. Due to this fact, we obtained the non-parenchymal cell fraction via differential centrifugation and cultured it.…”

Section: Discussionmentioning

confidence: 99%

A New Method to Isolate and Culture Rat Kupffer Cells

Zeng

Zhang

et al. 2013

PLoS ONE

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BackgroundPrevious methods for Kupffer cells (KCs) isolation require sophisticated skills and tedious procedures. Few studies have attempted to explore the self-renewal capacity of KCs in vitro. Therefore, the aim of this study was to establish a simple method for rat KCs isolation and further investigate the mitotic potential of KCs in vitro.MethodsKCs were obtained by performing one-step perfusion, enzymatic tissue treatment, differential centrifugation and selective adherence. The proliferation ability of cultured KCs was determined by MTT assay and Propidium Iodide FACS analysis. Phagocytic assay and ED-1, ED-2 immunofluorescence were used to identify cell phenotype. After stimulation with LPS, the expression of surface antigens (MHCII, CD40, CD80, and CD86) and the production of cytokines (NF-κB, TNF-α, IL-6 and IL-10) were measured for cell function identification.ResultsKCs were isolated with certain numbers and reasonable purities. The KCs were able to survive until at least passage 5 (P5), and at P3 showed equally strong phagocytic activity as primary KCs (P0). After stimulation with LPS, the change in the expression of surface antigens and the production of cytokines for P3 cells was similar to that for P0 cells.ConclusionsOur study provides a simple and efficient method for KCs isolation, and reveals that self-renewing KCs have the same phagocytic activity and functions as primary KCs.

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Section: Discussionmentioning

confidence: 99%

A New Method to Isolate and Culture Rat Kupffer Cells

Zeng

Zhang

et al. 2013

PLoS ONE

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“…CD163 antigen is expressed on cell surface of resident macrophages (Kupffer cells) 20,23) ; the antigen is a glycoprotein, which belongs to scavenger receptor cysteine-rich group B family and functions as the scavenger receptor for hemoglobin-haptoglobin complexes 23) . Antibody to CD204 was generated against human type 1 scavenger receptor protein and CD204 expression is related to metabolism of oxidized low density lipoprotein 24) ; in normal rat liver, CD204 is expressed in Kupffer cells existing along the sinusoid, similar to CD163expression 25) . MHC class II molecule may be expressed in dendritic cells and activated macrophages as the antigen-presenting cells 21,26) .…”

Section: Immunophenotypical Detection Of Rat Macrophages With Normal mentioning

confidence: 99%

Histopathological Analysis of Rat Hepatotoxicity Based on Macrophage Functions: in Particular, an Analysis for Thioacetamide-induced Hepatic Lesions

2016

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Hepatic macrophages play an important role in homeostasis. The functional abnormalities of hepatic macrophages primarily or secondarily influence chemically induced hepatotoxicity. However, the evaluation system based on their functions has not yet been established. Recently, a new concept (M1-/M2-macrophage polarization) was proposed; M1macropahges are induced by INF-γ, and show high phagocytosis/tissue damage, whereas M2-macropahges are induced by IL-4 and play roles in reparative fibrosis by releasing IL-10 and TGF-β1. In hepatogenesis, CD68-expressing M1macrophages predominantly exist in embryos; in neonates, in contrast, CD163-/CD204-expressing M2-macrophages appear along the sinusoids and mature as Kupffer cells. Activated Kupffer cells by liposome decrease AST and ALT values, whereas AST and ALT values are increased under Kupffer cells depleted with clodronate treatment. Since Kupffer cells may be involved in clearance of liver enzymes, macrophage condition should be taken into consideration when hepatotoxicity is analyzed. In TAA-induced acute hepatic lesions, INF-γ, TNF-α and IL-6 for M1-factors and IL-4 for M2factors are already increased before histopathological change; the appearance of CD68-expressing M1-macrophages and CD163-expressing M2-macrophages follows in injured centrilobular lesions, and TGF-β1 and IL-10 are increased for reparative fibrosis. CD68-expressing M1-macrophages co-express MHC class II and Iba-1, whereas CD163-expressing M2-macrophages also express CD204 and Galectin-3. Under macrophage depletion by clodoronate, TAA-treated rat livers show prolonged coagulation necrosis of hepatocytes, and then develop dystrophic calcification without reparative fibrosis. The depletion of hepatic macrophages influences hepatic lesion development. Collectively, a histopathological analysis method for hepatotoxicity according to M1-/M2-macrophage polarization would lead to the refinement of hazard characterization of chemicals in food and feed.

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“…Immunophenotyping studies recently have demonstrated the existence of divergent types of macrophages, which could be identified by specific molecules that they express depending on microenvironmental conditions evoked by cell-to-cell or cell-to-matrix interactions. For example, the phagolysosomal membrane protein, cluster of differentiation (CD)68, is highly expressed by infiltrating macrophages with activated phagocytosis; although CD163 and CD204 are expressed in resident macrophages (Kupffer cells in the liver) in normal rat livers, their expressions in injured tissues may imply the activation of different scavenger receptors recognized by respective CD163 and CD204; antigen-presenting macrophages such as dendritic cells express major histocompatibility complex (MHC) class II molecule not only in normal tissues but also in pathological lesions, and MHC class II molecule expression may be related to subsequent complicated immune response (Damoiseaux et al 1994;Polfliet et al 2006;Conrad and Dittel 2011;Kiyanagi et al 2011;Golbar et al 2012). Recently, macrophages are functionally classified as classically activated macrophages (M1) and alternatively activated M2 macrophages (Sica and Mantovani 2012;Martinez and Gordon 2014;Ong et al 2015).…”

Section: Introductionmentioning

confidence: 99%

Depletion of Hepatic Macrophages Aggravates Liver Lesions Induced in Rats by Thioacetamide (TAA)

et al. 2016

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Hepatic macrophages play crucial roles in hepatotoxicity. We investigated immunophenotypes of macrophages in liver injury induced in rats by thioacetamide (TAA; 300 mg/kg, intraperitoneal) after hepatic macrophage depletion; hepatic macrophages were depleted by liposomal clodronate (CLD; 10 ml/kg, i.v.) one day before TAA injection. Samples were obtained on post-TAA injection days 0, 1, 2, 3, 5, and 7. TAA injection induced coagulation necrosis of hepatocytes on days 1 through 3 and subsequent reparative fibrosis on days 5 and 7 in the centrilobular area, accompanied by increased numbers of M1 macrophages (expressing cluster of differentiation [CD]68 and major histocompatibility complex class II) and M2 macrophages (expressing CD163 and CD204) mainly on days 1 through 3. TAA þ CLD treatment markedly decreased the numbers of M1 and M2 macrophages mainly on days 1 through 3; CD163 þ Kupffer cells were most sensitive to CLD depletion. In TAA þ CLD-treated rats, interestingly, coagulation necrosis of hepatocytes was prolonged with more increased levels of hepatic enzymes (aspartate transaminase, alanine transaminase, and alkaline phosphatase) to TAA-treated rats; reparative fibrosis was incomplete and replaced by dystrophic calcification in the injured area, indicating the aggravated damage. Furthermore, in TAA þ CLD-treated rats, inflammatory factors (monocyte chemoattractant protein [MCP]-1, interferon-g, tumor necrosis factor-a, and interleukin-10) and fibrosis-related factors (transforming growth factor-b1, matrix metalloproteinase-2, tissue inhibitor of metalloproteinase-1) were decreased at messenger RNA levels, indicating abnormal macrophage functions. It was clearly demonstrated that hepatic macrophages have important roles in tissue damage and remodeling in hepatotoxicity.

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Immunohistochemical analyses of the kinetics and distribution of macrophages, hepatic stellate cells and bile duct epithelia in the developing rat liver

Cited by 28 publications

References 39 publications

A New Method to Isolate and Culture Rat Kupffer Cells

A New Method to Isolate and Culture Rat Kupffer Cells

Histopathological Analysis of Rat Hepatotoxicity Based on Macrophage Functions: in Particular, an Analysis for Thioacetamide-induced Hepatic Lesions

Depletion of Hepatic Macrophages Aggravates Liver Lesions Induced in Rats by Thioacetamide (TAA)

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