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1.
Arq. bras. med. vet. zootec. (Online) ; 73(4): 868-876, Jul.-Aug. 2021. graf, mapas, ilus
Article in English | ID: biblio-1285271

ABSTRACT

The melanomacrophage centers (MMCs) in the liver of fish are indicators of environmental conditions, as they are involved in xenobiotic biotransformation. The objective of this study was to evaluate the number of MMC in the liver of juveniles and adults of Sciades herzbergii from areas with different levels of contamination. The fish were caught at three points (reference - A1, potentially impacted - A2 and contaminated - A3), in São José bay (Maranhão, Brazil), in four samples. The livers were subjected to the standard histological procedure and 5µm sections were stained with hematoxylin-eosin. In livers of A2 adult individuals (260.50±161.50 MMCs / mm²) they presented a greater number of MMCs when compared to A3 adults (60.00 ± 30.10 MMCs / mm²). Juveniles showed considerable values in A1 (100.00 ± 0.00 MMCs/mm²) and A2 (95.33 ± 33.00 MMCs / mm²) compared to juveniles in A3 (49.00±0.00 MMCs/mm²). These high values are unexpected for young people. The average number of MMC correlated with the rainy season in the region. The use of hepatic MMCs as a biomarker of exposure to pollutants, in particular substances from fisheries systems, such as ammonia and nitrite, proved to be adequate to differentiate areas with different levels of impacts.(AU)


Os centros melanomacrófagos (MMCs) no fígado de peixes são indicadores das condições ambientais, pois estão envolvidos na biotransformação xenobiótica. O objetivo deste estudo foi avaliar o número de MMC no fígado de juvenis e adultos de Sciades herzbergii de áreas com diferentes níveis de contaminação. Os peixes foram capturados em três pontos (referência - A1; potencialmente impactado - A2; e contaminado - A3), na baía de São José (Maranhão, Brasil), em quatro amostras. Os fígados foram submetidos ao procedimento histológico padrão e cortes de 5µm foram corados com hematoxilina-eosina. Em fígados de indivíduos adultos A2 (260,50±161,50 MMCs/mm²), eles apresentaram maior número de MMCs quando comparados aos adultos A3 (60,00±30,10 MMCs/mm²). Os juvenis apresentaram valores elevados em A1 (100,00 ± 0,00 MMCs/mm²) e A2 (95,33±33,00 MMCs/mm²) quando comparados aos juvenis em A3 (49,00±0,00 MMCs/mm²). Esses altos valores são inesperados para os jovens. O número médio de MMC correlacionou-se com a época chuvosa na região. A utilização de MMCs hepáticos como biomarcador de exposição a poluentes, em particular substâncias provenientes de sistemas pesqueiros, como amônia e nitrito, mostrou-se adequada para diferenciar áreas com diferentes níveis de impactos.(AU)


Subject(s)
Animals , Catfishes , Environmental Biomarkers , Biological Monitoring/methods , Kupffer Cells , Kupffer Cells/cytology , Environmental Pollution/analysis
2.
Article in English | WPRIM | ID: wpr-880608

ABSTRACT

The incidence of non-alcohol fatty liver disease (NAFLD) is increasing year by year, and the relevant cardiovascular events have become a major problem in chronic diseases management. The activation of innate immunity is closely related to the development of NAFLD. The immune cells include Kupffer cells, neutrophils, dendritic cells, and natural killer T cells, which acts through the activation of innate immunity-related signals mediated by pattern recognition receptors.


Subject(s)
Humans , Immunity, Innate , Kupffer Cells , Liver , Non-alcoholic Fatty Liver Disease
3.
Article in Chinese | WPRIM | ID: wpr-828870

ABSTRACT

OBJECTIVE@#To investigate the role of endoplasmic reticulum (ER)-stress of Kupffer cells (KCs) and KCs-derived tumor necrosis factor- (TNF-) in medicating apoptosis of hepatic stellate cell (HSC).@*METHODS@#Sixty male SD rats were randomized into control group, model group, ER- stress group, depletion group and KCs block group (=15). The 4 groups of rats were given intraperitoneal injections (twice a week for 8 weeks) of normal saline (2 mg/kg); 40% CCl4 solution (in peanut oil, 2 mg/kg); 40% CCl4 solution (2 mg/kg) and tunicamycin (1 mg/kg); and 40% CCl4 solution (2 mg/kg) and tunicamycin (1 mg/kg) followed by clodronate liposomes (50 mg/kg), respectively. After the treatments, samples of the liver tissue and serum were collected from the rats from the 4 groups to isolate KC cells, which were co-cultured with LX2 cells. In the depletion group, the rats were injected with anti-rat TNFR mAb (0.35 mg/kg) via the portal vein before isolating the KCs. Liver function examination, Eirius red staining, ELISA, immuno- histochemical staining, and RT-PCR were performed to assess the liver function, liver fibrosis, KC phenotypes, expression of the in fl ammatory factors, and the number of active HSC was detected. The isolated KCs were treated with tunicamycin before co-culture with LX2 cells, and ELISA, RT-PCR and Western blot were performed to examine KC phenotypes, in fl ammatory factors, LX2 cell apoptosis and TNFR/caspase8 pathway activity.@*RESULTS@#Compared with the rats in the control group, the rats in the model group had significantly increased ALT and AST levels, Sirius red staining-positive area, and Desmin-positive cells (activated HSCs) ( < 0.05) with significantly lowered number of CD16-positive KCs (M1), and TNF- protein and mRNA expression ( < 0.05). Compared with those in the model group, the rats in ER-stress group showed significantly decreased ALT and AST levels, Sirius red staining positivity and Desmin-positive cells ( < 0.05) and increased number of CD16-positive KCs and TNF- expressions ( < 0.05). In the depletion group, compared with the ER-stress group, the rats had significantly increased ALT and AST levels of, Sirius red staining positivity and Desmin-positive cells ( < 0.05) and reduced CD16- positive KCs and TNF-expressions ( < 0.05). In the cell co-culture experiment, the model group showed significantly reduced TUNEL-positive LX2 cells, CD16-positive cells, and expressions of TNFR1, cleaved caspase- 8 and cleaved caspase- 3 in the KCs ( < 0.05) with increased Desmin-positive LX2 cells ( < 0.05). Compared with the model group, the ER- stress group exhibited significantly increased TUNEL-positive LX2 cells, CD16-positive cells and expressions of TNFR, cleaved caspase-8 and cleaved caspase-3 in the KCs ( < 0.05) and decreased Desmin-positive LX2 cells ( < 0.05). In the depletion group, blocking TNFR resulted in significantly decreased expressions of cleaved caspase-8 and cleaved caspase-3 compared with those in ER- stress group ( < 0.05) although there was no significant changed in TNFR expression.@*CONCLUSIONS@#ER stress of KCs promotes the transformation of KCs towards M1 phenotype and increases the expression of TNF-, which triggers the apoptosis of HSCs through the TNFR/caspase-8 pathway.


Subject(s)
Animals , Apoptosis , Caspase 8 , Endoplasmic Reticulum Stress , Hepatic Stellate Cells , Kupffer Cells , Male , Rats , Rats, Sprague-Dawley , Tumor Necrosis Factor-alpha
4.
Article in English | WPRIM | ID: wpr-771434

ABSTRACT

OBJECTIVE@#To investigate the mechanism of inflflammatory-mediated toll-like receptor 4 (TLR4)-p38 mitogen-activated protein kinase (p38 MAPK) pathway in Kupffer cells (KCs) of non-alcoholic steatohepatitis (NASH) rats and the intervention effect of soothing Gan (Liver) and invigorating Pi (Spleen) recipes on this pathway.@*METHODS@#After 1 week of acclimatization, 120 Sprague-Dawley male rats were randomly divided into 8 groups using a random number table (n=15 per group): normal group, model group, low-dose Chaihu Shugan Powder (, CHSG) group (3.2 g/kg), high-dose CHSG group (9.6 g/kg), low-dose Shenling Baizhu Powder (, SLBZ) group (10 g/kg), high-dose SLBZ (30 g/kg) group, and low- and highdose integrated recipe (L-IR, H-IR) groups. All rats in the model and treatment groups were fed with a high-fat diet (HFD). The treatments were administrated by gastrogavage once daily and lasted for 26 weeks. The liver tissues were detected with hematoxylin-eosin (HE) and oil red O staining. Levels of liver lipids, serum lipids and transaminases were measured. KCs were isolated from the livers of rats to evaluate the mRNA expressions of TLR4 and p38 MAPK by real-time flfluorescence quantitative polymerase chain reaction, and proteins expressions of TLR4, p-p38 MAPK and p38 MAPK by Western blot. Levels of inflammatory cytokines including tumor necrosis factor α (TNF-α), interleukin (IL)-1 and IL-6 in KCs were measured by enzyme-linked immunosorbent assay.@*RESULTS@#After 26 weeks of HFD feeding, HE and oil red O staining showed that the NASH model rats successfully reproduced typical pathogenesis and histopathological features. Compared with the normal group, the model group exhibited significant increases in body weight, liver weight, liver index, serum levels of total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol, and aspartate aminotransferase as well as TC and TG levels in liver tissues, and significant decrease in serum level of high-density lipoprotein cholesterol (Plt;0.05 or Plt;0.01), while those indices were significantly ameliorated in the H-IR group (Plt;0.05 or Plt;0.01). Higher levels of TNF-α, IL-1 and IL-6 in KCs were observed in the model group compared with the normal group (Plt;0.01). Significant decreases in TNF-α, IL-1 and IL-6 were observed in the H-SLBZ, H-IR and L-IR groups compared with the model group (Plt;0.05 or Plt;0.01). The mRNA expressions of TLR4 and p38 MAPK and protein expressions of TLR4, p38 MAPK and p-p38 MAPK in KCs in the model group were significantly higher than the normal group (Plt;0.01), while those expression levels in the L-IR and H-IR groups were significantly lower than the model group (Plt;0.05 or Plt;0.01).@*CONCLUSION@#Inflflammation in KCs might play an important role in the pathogenesis of NASH in rats. The data demonstrated the importance of TLR4-p38MAPK signaling pathway in KCs for the anti-inflflammatory effect of soothing Gan and invigorating Pi recipes.


Subject(s)
Animals , Drugs, Chinese Herbal , Pharmacology , Kupffer Cells , Physiology , MAP Kinase Signaling System , Male , Medicine, Chinese Traditional , Non-alcoholic Fatty Liver Disease , Drug Therapy , Plant Extracts , Pharmacology , Rats , Rats, Sprague-Dawley , Signal Transduction , Toll-Like Receptor 4 , Physiology , p38 Mitogen-Activated Protein Kinases , Physiology
5.
Acta cir. bras ; 33(12): 1052-1060, Dec. 2018. graf
Article in English | LILACS | ID: biblio-973489

ABSTRACT

Abstract Purpose: To establish a method for the preparation of zoledronate liposome and to observe its effect on inducing the apoptosis of rat liver Kupffer cells. Methods: Zoledronate was encapsulated in liposomes, and then the entrapment rate was detected on a spectrophotometer. The prepared Zoledronate liposome (0.01 mg/mL) was injected into the tail vein of SD rats. Three days later, the number of Kupffer cells (CD68 positive) in rat liver tissue was detected by immunohistochemistry. Flow cytometry was used to detect the apoptosis rate of the isolated liver Kupffer cell cultured in vitro. Results: The entrapment rate of Zoledronate was 43.4±7.8%. Immunohistochemistry revealed that the number of Kupffer cells was 19.3±2.1 in PBS group and 5.5±1.7 in Zoledronate liposome group, with a significant difference (P<0.05). The apoptosis rate of Kupffer cells was 4.1±0.8% in PBS group, while it was 9±2.2% and 23.3±5.9% in Zoledronate liposomes groups with different concentrations of Zoledronate liposome (P<0.05). Conclusions: Zoledronate liposomes can effectively induce the apoptosis of Kupffer cells in vivo and in vitro, and the apoptosis rate is related to the concentration of Zoledronate liposome. To establish a rat liver Kupffer cell apoptosis model can provide a new means for further study on Kupffer cell function.


Subject(s)
Animals , Male , Apoptosis/drug effects , Zoledronic Acid/pharmacology , Kupffer Cells/drug effects , Liver/cytology , Immunohistochemistry , Random Allocation , Cell Count , Reproducibility of Results , Treatment Outcome , Rats, Sprague-Dawley , Drug Compounding/methods , Flow Cytometry , Zoledronic Acid/administration & dosage , Zoledronic Acid/chemical synthesis , Liposomes/chemical synthesis
6.
Article in English | WPRIM | ID: wpr-713310

ABSTRACT

BACKGROUND/AIMS: Nonalcoholic steatohepatitis (NASH) is prevalent in both economically developed and developing countries. Twenty percent of NASH progresses to cirrhosis with/without hepatocellular carcinoma, and there is an urgent need to find biomarkers for early diagnosis and monitoring progression of the disease. Using immunohistochemical and immunoelectron microscopic examination we previously reported that expression of matrix metalloproteinase-1 (MMP-1) increased in monocytes, Kupffer cells and hepatic stellate cells in early stage NASH. The present study investigated whether serum MMP-1 levels reflect disease activity and pharmaceutical effects in NASH patients. METHODS: We measured the serum levels of MMPs, tissue inhibitors of metalloproteinases (TIMPs), and several cytokines/chemokines in patients with histologically proven early and advanced stages of NASH and compared them with those in healthy controls. RESULTS: Serum MMP-1 levels in stage 1 fibrosis, but not in the more advanced fibrosis stages, were significantly higher than in healthy controls (P=0.019). There was no correlation between serum MMP-1 level and fibrosis stage. Serum MMP- 1 levels in NASH patients represented disease activity estimated by serum aminotransferase values during the follow-up period. In contrast, MMP-2, MMP-9 and TIMPs did not change with disease activity. Consistent with the finding that MMP-1 is expressed predominantly in monocytes and Kupffer cells, serum levels of monocyte chemotactic protein-1 and granulocyte-colony stimulating factor were significantly increased in NASH with stage 1 fibrosis. CONCLUSIONS: These results suggest that serum MMP-1 levels represent disease activity and may serve as a potential biomarker for monitoring the progression of NASH.


Subject(s)
Biomarkers , Carcinoma, Hepatocellular , Chemokine CCL2 , Cytokines , Developing Countries , Early Diagnosis , Fibrosis , Follow-Up Studies , Hepatic Stellate Cells , Humans , Kupffer Cells , Liver Cirrhosis , Matrix Metalloproteinase 1 , Matrix Metalloproteinases , Metalloproteases , Monocytes , Non-alcoholic Fatty Liver Disease
7.
Laboratory Animal Research ; : 133-139, 2018.
Article in English | WPRIM | ID: wpr-719081

ABSTRACT

Nonalcoholic steatohepatitis (NASH) is becoming common chronic liver disease because of the increasing global prevalence of obesity and consequently Nonalcoholic fatty liver disease (NAFLD). However, the mechanism for progression of NAFLD to NASH and then cirrhosis is not completely understood, yet. The triggering of these hepatic diseases is thought from hepatocyte injury caused by over-accumulated lipid toxicity. Injured hepatocytes release damage-associated molecular patterns (DAMPs), which can stimulate the Kupffer cells (KCs), liver-resident macrophages, to release pro-inflammatory cytokines and chemokines, and recruit monocyte-derived macrophages (MDMs). The increased activation of KCs and recruitment of MDMs accelerate the progression of NAFLD to NASH and cirrhosis. Therefore, characterization for activation of hepatic macrophages, both KCs and MDMs, is a baseline to figure out the progression of hepatic diseases. The purpose of this review is to discuss the current understanding of mechanisms of NAFLD and NASH, mainly focusing on characterization and function of hepatic macrophages and suggests the regulators of hepatic macrophages as the therapeutic target in hepatic diseases.


Subject(s)
Chemokines , Cytokines , Fatty Liver , Fibrosis , Hepatocytes , Kupffer Cells , Liver Diseases , Macrophages , Non-alcoholic Fatty Liver Disease , Obesity , Prevalence
8.
Immune Network ; : e24-2018.
Article in English | WPRIM | ID: wpr-715077

ABSTRACT

Ischemia-reperfusion injury (IRI) is a major complication in liver transplantation (LT) and it is closely related to the recovery of grafts' function. Researches has verified that both innate and adaptive immune system are involved in the development of IRI and Kupffer cell (KC), the resident macrophages in the liver, play a pivotal role both in triggering and sustaining the sterile inflammation. Damage-associated molecular patterns (DAMPs), released by the initial dead cell because of the ischemia insult, firstly activate the KC through pattern recognition receptors (PRRs) such as toll-like receptors. Activated KCs is the dominant players in the IRI as it can secret various pro-inflammatory cytokines to exacerbate the injury and recruit other types of immune cells from the circulation. On the other hand, KCs can also serve in a contrary way to ameliorate IRI by upregulating the anti-inflammatory factors. Moreover, new standpoint has been put forward that KCs and macrophages from the circulation may function in different way to influence the inflammation. Managements towards KCs are expected to be the effective way to improve the IRI.


Subject(s)
Cytokines , Hand , Immune System , Inflammation , Ischemia , Kupffer Cells , Liver Transplantation , Liver , Macrophages , Receptors, Pattern Recognition , Reperfusion Injury , Reperfusion , Toll-Like Receptors
9.
Article in Chinese | WPRIM | ID: wpr-771451

ABSTRACT

OBJECTIVE@#To investigate the role of cathepsin B in hepatic Kupffer cells (KCs) in activating Toll-like receptor 4(TLR- 4)-independent inflammatory pathways in mice with lipopolysaccharide (LPS)-induced sepsis.@*METHODS@#Eighteen wild-type (WT) mice and 18 TLR4-knockout (TLR4) mice were both divided into 3 groups for intraperitoneal injections of a lethal dose (54 mg/kg) of LPS, LPS and CA-074(a cathepsin B inhibitor), or normal saline, and the survival of the mice were observed. Another 36 WT mice and 36 TLR4mice were also divided into 3 groups and subjected to intraperitoneal injections of normal saline, 20 mg/kg LPS, or LPS with CA-074 pretreatment.After the treatments, KCs were collected from the mice for assessing the protein level and activity of cathepsin B.The histopathological changes of the liver were observed with HE staining, and the serum levels of IL-1α, IL-1β, TNF-α and IL-18 were detected.@*RESULTS@#Compared with the WT mice,TLR4mice receiving the lethal dose of LPS had significantly longer survival time (up to 84 h) after the injection,but were still unable to fully resist LPS challenge.CA-074 pretreatment prolonged the survival time of WT mice and TLR4mice to 60 h and 132 h,respectively.In the mouse models of sepsis,20 mg/kg LPS induced significantly enhanced activity of cathepsin B without affecting its expression level in the KCs (<0.05) and increased the serum levels of the inflammatory cytokines.CA-074 pretreatment of the mice obviously lessened the detrimental effects of LPS in TLR4mice by significantly lowering cathepsin B activity in the KCs,alleviating hepatocyte apoptosis and reducing the serum levels of inflammatory cytokines.@*CONCLUSIONS@#Cathepsin B plays an important role in activating TLR4-independent inflammatory pathways in mice with LPS-induced sepsis.


Subject(s)
Animals , Cathepsin B , Physiology , Dipeptides , Pharmacology , Gene Knockout Techniques , Hepatocytes , Inflammation , Metabolism , Interleukin-18 , Blood , Interleukin-1alpha , Blood , Interleukin-1beta , Blood , Kupffer Cells , Metabolism , Lipopolysaccharides , Liver , Pathology , Mice , Sepsis , Metabolism , Toll-Like Receptor 4 , Genetics , Tumor Necrosis Factor-alpha , Blood
10.
Article in English | WPRIM | ID: wpr-60682

ABSTRACT

Temporal and subcellular distributions of hyaluronic acid (HA) as a degradable nanoparticle (NP) in animals were investigated to determine if HA-NP could be utilized as an appropriate drug delivery system. After mice were intravenously injected with 5 mg/kg of Cy5.5-labeled HA-NP sized 350–400 nm or larger HA-polymers, the fluorescence intensity was measured in all homogenized organs from 0.5 h to 28 days. HA-NP was greatly detected in spleen, liver and kidney until day 28, while it was maintained at low levels in other organs. HA-polymer was observed at low levels in all organs. HA-NP quantities in spleen and liver were reduced until day 3, but increased sharply between days 3 and 7, then decreased again, while their HA-polymers were maintained at low levels until day 28. In kidneys, both HA-NP and HA-polymer showed high levels after 0.5 h of administration, but steadily decreased until day 28. According to ultrastructural analyses, HA-NP was engulfed in Kupffer cells of liver and macrophages of spleen and kidney at day 1 and was accumulated in the cytoplasm of kidney tubular cells at day 7. Overall, these findings suggest that HA-NP could be considered a desirable drug carrier in the liver, kidney, or spleen.


Subject(s)
Animals , Cytoplasm , Drug Carriers , Drug Delivery Systems , Fluorescence , Hyaluronic Acid , Kidney , Kupffer Cells , Liver , Macrophages , Mice , Nanoparticles , Pharmacokinetics , Spleen
11.
Article in Chinese | WPRIM | ID: wpr-273743

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the effects of inhibiting TIM-4 function in Kupffer cells (KCs) on liver graft rejection in mice and explore the underlying mechanism.</p><p><b>METHODS</b>Mouse models of orthotopic liver transplantation were treated with a control mAb group and TIM-4 mAb. The activated KCs were assayed with immunohistochemistry after operation. The expression of TIM-4 in KCs were assayed with Western blotting and RT-PCR and the levels of AST, ALT, TBIL, TNF-α, IFN-γ and CCL2 were assayed detected. The expression of TIM-4 in KCs was observed with laser confocal microscopy. HE staining was used to observe the microstructure of the liver tissues, and the number of CD25Foxp3T cells was determined using with flow cytometry; the proteins levels of p-P65and p-P38 were assayed with Western blotting. The donor mice were treated with clodronate liposomes to destroy the KCs in the liver before transplantation, and the liver grafts were examined for graft rejection.</p><p><b>RESULTS</b>The number of activated KCs in the liver graft increased progressively over time. Compared with the sham-operated group, the liver graft showed significantly increased TIM-4 protein and mRNA levels at 1, 3, and 7 days after transplantation (P<0.05) and increased levels of AST, ALT, TBIL, TNF-α, IFN-γ and CCL2 at 7 days (P<0.05). The graft in TIM-4 mAb group showed mild pathological changes with a mean RAI score of 2.67∓0.75, which was significantly lower than that in control mAb group (P<0.05). The mean survival time of the recipient mice was 53.8∓6.4 days in TIM-4 mAb group, significantly longer than that in the control mAB group (14.5∓2.9 days, P<0.05). Donor treatment with clodronate liposomes resulted in comparable RAI scores in TIM-4 mAb and control mAb groups (8.01∓0.64 vs 7.93∓0.56, P>0.05). The protein levels of p-P65 and p-P38 in TIM-4 mAb group were significantly lower than those in control mAb group (P<0.05), and CD25Foxp3T cells in the liver graft increased significantly in TIM-4 mAb group.</p><p><b>CONCLUSION</b>Inhibition of TIM-4 function in KCs reduces the production of inflammatory factors after liver transplantation possibly by inhibiting the NF-κB and MAPK signaling pathways and promoting the proliferation of Foxp3Treg cells to induce allograft tolerance.</p>


Subject(s)
Animals , Antibodies, Monoclonal , Pharmacology , Graft Rejection , Immunohistochemistry , Kupffer Cells , Metabolism , Liver , General Surgery , Liver Transplantation , Male , Membrane Proteins , Mice , NF-kappa B , Metabolism , T-Lymphocytes, Regulatory , Allergy and Immunology
12.
Article in Chinese | WPRIM | ID: wpr-815011

ABSTRACT

OBJECTIVE@#To evaluate whether dexmedetomidine hydrochloride, an α(2)-adrenergic receptor agonist, can prevent H(2)O(2)-induced oxidative stress and inflammatory response in Kupffer cells.
@*METHODS@#H(2)O(2)-induced oxidative damage model of Kupffer cell was established. Kupffer cells were pre-conditioned by dexmedetomidine hydrochloride or Yohimbine for 24 h. MTT colorimetry was used to demonstrate the survival rate of Kupffer cells. The levels of lactate dehydrogenase (LDH), malonaldehyde (MDA) and TNF-α in the culture medium were assessed by corresponding kits.
@*RESULTS@#Dexmedetomidine hydrochloride protected Kupffer cells from H(2)O(2)-induced oxidative damage, showing an increase in the cell survival rate while a decrease in LDH, MDA and TNF-α release in the culture supernatant. Yohimbine, an α(2)-adrenergic receptor antagonist, completely neutralized the protective effect of Dexmedetomidine hydrochloride on Kupffer cells. Yohimbine itself had no effect on H(2)O(2)-induced oxidative damage and inflammatory response.
@*CONCLUSION@#Dexmedetomidine hydrochloride can prevent H(2)O(2)-induced oxidative stress and inflammatory response in Kupffer cells through activation of α(2)-adrenergic receptors.


Subject(s)
Adrenergic alpha-2 Receptor Antagonists , Pharmacology , Cell Survival , Cells, Cultured , Dexmedetomidine , Pharmacology , Humans , Hydrogen Peroxide , Pharmacology , Kupffer Cells , Cell Biology , L-Lactate Dehydrogenase , Metabolism , Malondialdehyde , Metabolism , Oxidative Stress , Receptors, Adrenergic, alpha-2 , Metabolism , Tumor Necrosis Factor-alpha , Metabolism , Yohimbine , Pharmacology
13.
Rio de Janeiro; s.n; 2016. 68 p. ilus, tab.
Thesis in Portuguese | LILACS | ID: biblio-983468

ABSTRACT

É conhecido que processos inflamatórios podem modular a expressão e atividade deenzimas CYPs. Não é claro, entretanto, o modo pelo qual estímulos inflamatórios regulam aexpressão dessas enzimas. Neste trabalho investigamos a hipótese de que as células deKupffer do fígado exerceriam um papel na modulação dos CYPs hepáticos em resposta àinflamação exacerbada ou sepse. O cloreto de gadolínio, é um inibidor seletivo das células deKupffer, conhecido por atenuar o quadro de inflamação exacerbada, quandoadministradopreviamente ao estímulo inflamatório, em diferentes modelos animais. Algunsautores sugeriram que as células de Kupffer atuariam como intermediários na modulação daatividade de CYPs hepáticos desencadeada por estímulos inflamatórios. Há estudos quesugerem que a diminuição da população das células de Kupfferatenua ou elimina a regulaçãodas CYPs hepáticas por estímulos inflamatórios.Além disso, estudos em culturas dehepatócitos in vitro, na ausência de células de Kupffer, tem constatado a regulação negativa daexpressão de CYPs hepáticas após estimulação com LPS. Nessa linha, o objetivo destepresente trabalho é investigar o papel das células de Kupffer na regulação da atividade deenzimas hepáticas de biotransformação de xenobióticos (CYPs) após estimulação inflamatóriacom LPS. Para isso, os níveis séricos de transaminases, e a histopatologia foram empregadospara avaliar o efeito do tratamento com diferentes doses de GdCl3sobre o tecido hepático.Noexperimento principal para investigar o papel das células de Kupffer, os ratos foram alocadosao acaso em quatro grupos. Foram quantificadosmarcadores bioquímicos no soro dos animaispara evidenciar danos ao tecido hepático causados pelos tratamentos e realizado o examehistopatológico...


It is known that inflammatory processes may modulate the expression and activity ofCYP enzymes. The mode by which inflammatory stimuli regulate CYP expression andactivity, however, remains unclear. Kupffer cells are resident macrophages in the liver andthus play an important role in a systemic inflammatory process or in sepsis. Gadoliniumchloride (GdCl3) has been reported to selectively kill an/or inhibit the activity of Kupffercells. Along this line, it has also been described that Gd decreases exacerbated inflammatoryresponses when it is administered prior to inflammatory stimuli in various animal models.Therole of Kupffer cells in the modulation of CYPs activity triggered by inflammatory stimuli,however, is not entirely clear in the literature. There are studies suggesting that a reduction inthe population of Kupffer cells attenuates the down-regulation of hepatic CYPs induced byinflammatory stimuli. However, GdCl3 was also described to decrease liver CYP activityirrespective of whether it depletes or not Kupffer cells. Moreover, in vitro studies showed thatLPS down-regulates the expression of CYP forms in hepatocyte cell lines in culture (in theabsence of Kupffer cells). To investigate whether Kupffer cells play a role in the regulation ofthe activity of liver xenobiotic biotransformation enzymes (CYPs) by inflammatorystimulation with LPS. The activity of transaminases in the blood serum (a marker for liverinjury) was determined and liver histopathology was evaluated in female Wistar rats. In a setof preliminary tests, rats were treated with different doses of GdCl3 or with LPS for selectingdoses and euthanasia time in the main experiment. In the main study experimental groups.Treatment associated liver injury was evaluated by levels of transaminases and alkalinephosphatase in the blood serum and by liver histopathology examination...


Subject(s)
Animals , Mice , Kupffer Cells/enzymology , Liver/enzymology , Xenobiotics/metabolism , Endotoxins , Euthanasia , Gadolinium
14.
Article in Chinese | WPRIM | ID: wpr-232544

ABSTRACT

<p><b>OBJECTIVE</b>To compare the efficiency and cytotoxicity of different transfection reagents used in transfection of RIP140-siRNA into Kupffer cells to optimize the transfection conditions.</p><p><b>METHODS</b>Kupffer cells were transfected with RIP140-siRNA labeled with GFP as the reporter gene using lipofectamine 2000, Roche reagent (X-treme GENE siRNA Transfection Reagent) and puro screening lentivirus (1.0×10(8) TU/mL) as the transfection reagents. The transfection effect was observed under a fluorescent inverted microscope, and laser scanning confocal microscopy was used to analyze RIP140 expression in trasnfected Kupffer cells. Flow cytometry was performed to detect cell apoptosis, and CCK-8 test was used to evaluate the cell proliferation inhibition. RT-RCR and Western blotting were performed to detect the expressions of RIP140 mRNA and protein in the trasnfected cells.</p><p><b>RESULTS</b>Puro screening lentivirus yielded the highest cell transfection efficiency, which exceeded 90%, followed by Roche reagent and then by lipofectamine 2000. Flow cytometry and CCK-8 test showed that the cytotoxicity was the mildest with Roche reagent, moderate with lentivirus, and severe with lipofectamine 2000. The cells trasnfected with lentivirus showed a significantly lower RIP140 expression than cells trasnfected with lipofectamine 2000 and Roche reagent (P<0.05).</p><p><b>CONCLUSION</b>In Kupffer cells, lentivirus-mediated transfection, as compared with the other two trasnfection reagents, can achieve good transfection efficiency with a relativelty low cytotoxicity, and allows for better controllability and stability of the trasnfectiion conditions.</p>


Subject(s)
Adaptor Proteins, Signal Transducing , Genetics , Apoptosis , Cell Proliferation , Flow Cytometry , Humans , Indicators and Reagents , Chemistry , Kupffer Cells , Lentivirus , Lipids , Chemistry , Nuclear Proteins , Genetics , RNA, Messenger , RNA, Small Interfering , Genetics , Transfection
15.
Article in English | WPRIM | ID: wpr-52646

ABSTRACT

The inducible form of nitric oxide synthase (iNOS) is expressed in hepatic cells in pathological conditions. Its induction is involved in the development of liver fibrosis, and thus iNOS could be a therapeutic target for liver fibrosis. This review summarizes the role of iNOS in liver fibrosis, focusing on 1) iNOS biology, 2) iNOS-expressing liver cells, 3) iNOS-related therapeutic strategies, and 4) future directions.


Subject(s)
Endothelial Cells/metabolism , Hepatic Stellate Cells/metabolism , Humans , Kupffer Cells/metabolism , Liver Cirrhosis/metabolism , Nitric Oxide/metabolism , Nitric Oxide Synthase Type II/antagonists & inhibitors , Polymorphism, Single Nucleotide , RNA, Untranslated/metabolism
16.
Chinese Journal of Hepatology ; (12): 64-68, 2015.
Article in Chinese | WPRIM | ID: wpr-337047

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the effects and mechanisms of the inflammatory reaction related to nonalcoholic steatohepatitis (NASH) and induced by activation of the cholinergic anti-inflammatory pathway.</p><p><b>METHODS</b>A mouse model of NASH was established by feeding a high-fat and high-sugar diet.Activation of the cholinergic anti-inflammatory pathway was achieved by nicotine administration to the NASH modeled mice and normal controls. Liver biopsies were taken and the concentrations of cytokines were measured. Isolated liver primary Kupffer cells and RAw264.7 cells were cultured, pre-treated or not with lipopolysaccharide (LPS) and exposed to nicotine, after which the supernatant concentrations of IL-6 and TNFa were determined by ELISA. The protein expression levels of phosphorylated (p)-NF-kB and I k B were detected in primary cultured Kupffer cells by western blotting.</p><p><b>RESULTS</b>The mouse model of NASH was successfully established, as evidenced by findings from liver biopsy and serum liver function tests. The degree of liver inflammation in the NASH mice decreased after nicotine administration, and the level of serum TNFa also significantly decreased. The levels of serum TNFa were 21.95+/-0.8 pg/mL in nicotine-treated mice and 38.07+/-1.7 pg/mL in the non-nicotine-treated NASH mice (P less than 0.05). The nicotine treatment also significantly reduced the concentration of TNFa in the culture supernatants of Kupffer cells after LPS stimulation; moreover, the supernatant level of TNFa decreased significantly after the nicotine treatment (Pless than 0.05). LPS stimulation of the RAw264.7 cells led to an increased level ofp-NF-kB and a reduced level ofI-kB, suggesting that the NF-kB pathway had been activated; different doses of nicotine pre-treatment led to down-regulation of the p-NF-kB level and up-regulation of the I-kB level, both in dose-dependent manners.</p><p><b>CONCLUSION</b>Activating the cholinergic anti-inflammatory pathway inhibits the NASH-related inflammatory reaction, and the mechanism for this inhibition involves the NF-kB signaling pathway.</p>


Subject(s)
Animals , Cholinergic Agents , Down-Regulation , Inflammation , Interleukin-6 , Kupffer Cells , Lipopolysaccharides , Mice , NF-kappa B , Non-alcoholic Fatty Liver Disease , Phosphorylation , Up-Regulation
17.
Article in Chinese | WPRIM | ID: wpr-249414

ABSTRACT

<p><b>OBJECTIVE</b>To establish a method for simultaneous isolation and primary culture of rat hepatocytes, hepatic stellate cells, Kupffer's cells and hepatic sinus endothelial cells.</p><p><b>METHODS</b>By combining in situ perfusion, in vitro perfusion, density gradient centrifugation and differential adhesion, primary rat hepatocytes, hepatic stellate cells, Kupffer's cells and hepatic sinus endothelial cells were obtained. The purity of these cells were assessed with morphological observation, immunofluorescent staining and ink phagocytosis assay.</p><p><b>RESULTS</b>We successfully obtained the 4 primary cells simultaneously by combining in situ perfusion with in vitro perfusion, density gradient centrifugation, and differential attachment. The cell yield rate, cell viability and purity all met requirements for the subsequent cell experiment.</p><p><b>CONCLUSION</b>The combined cell isolation and culture method is feasible to isolate primary rat hepatocytes, hepatic stellate cells, Kupffer's cells and hepatic sinus endothelial cells simultaneously.</p>


Subject(s)
Animals , Cell Culture Techniques , Cell Separation , Endothelial Cells , Cell Biology , Hepatic Stellate Cells , Cell Biology , Hepatocytes , Cell Biology , Kupffer Cells , Cell Biology , Male , Rats , Rats, Sprague-Dawley
18.
Qom University of Medical Sciences Journal. 2013; 6 (4): 8-13
in Persian, English | IMEMR | ID: emr-126987

ABSTRACT

Despite widespread use of electromagnetic field [EMF] generator devices in everyday life, their effects on biological tissues are still controversial. In the present study, the effects of EMF on the mouse liver were studied. Thirty female balb/c mice were randomly divided into three groups: 1- EMF-exposed, 2-Sham, and 3-Control. EMF group was exposed to 50 Hz and 0.5 mT EMF 4 hours daily for 2 months while the animals in sham group were placed in EMF device without exposure. At the end of 2-month period, all animals were anesthetized and their livers were removed. After preparation of microscopic slides, the panoramic view of liver, the number of Kupffer and necrotic cells were determined using 'Image Tool' software. The data were statistically analyzed using ANOVA and Dunnett's tests. p<0.05 was considered as statically significant. The comparison of data showed that the mean number of Kupffer cells in EMF group significantly increased compared to sham and control group [p<0.001 for both], while the mean number of Kupffer cells in sham group showed no significant difference compared to control group. The mean number of necrotic cells in experimental groups had no significant increase compared to sham and control groups. There were no significant differences in the mean number of necrotic cells between sham and control groups. Qualitative studies also showed no disorganization in the structure of liver lobules and sinusoids in the exposed group compared to sham and control groups. According to the results of this study, longtime exposure to electromagnetic field could increase Kupffer cells but do not affect the necrosis of liver cells in mice


Subject(s)
Animals, Laboratory , Liver , Mice, Inbred BALB C , Kupffer Cells , Necrosis
19.
Chinese Journal of Burns ; (6): 158-161, 2013.
Article in Chinese | WPRIM | ID: wpr-284121

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the effect of high mobility group box protein 1 (HMGB1) on the production of pro-inflammatory cytokines by Kupffer cell (KC) of rats with severe burn and the role of receptor for advanced glycation end products (RAGE) in the process.</p><p><b>METHODS</b>Model of 30% TBSA full-thickness burn was reproduced in 32 SD rats through immersing the back in 98°C water for 12 s. KC (32 samples) was isolated from rat liver 24 h after injury and inoculated in 24-well plate in the concentration of 1×10(6) cell per well. (1) Cells were divided into control group (cultured with 1 mL PBS) and HMGB1 group (stimulated with 100 ng/mL HMGB1 in the volume of 1 mL) according to the random number table, with 8 samples in each group. At post culture hour (PCH) 48, the expression of RAGE (denoted as grey value ratio) was detected with Western blotting. (2) Another portion of cells were divided into control group (cultured with 1 mL PBS), HMGB1 group (treated with 100 ng/mL HMGB1 in the volume of 1 mL), HMGB1 + anti-RAGE antibody group (treated with 100 ng/mL HMGB1 in the volume of 1 mL after being pre-incubated with 20 µg/mL anti-RAGE monoclonal antibody in the volume of 1 mL for 2 hours), HMGB1 + recombinant rat RAGE/Fc chimera (rrRAGE/Fc) group (treated with the mixture of 100 ng/mL HMGB1 in the volume of 0.5 mL and 5 µg/mL rrRAGE/Fc in the volume of 0.5 mL which were pre-incubated for 2 hours) according to the random number table, with 8 samples in each group. At PCH 48, the protein levels of TNF-α and IL-1β in supernatant were determined with enzyme-linked immunosorbent assay, while the mRNA expression of TNF-α and IL-1β (denoted as grey value ratio) were determined with Northern blotting. Data were processed with one-way analysis of variance, t test, and LSD test.</p><p><b>RESULTS</b>(1) The expression of RAGE in HMGB1 group (1.036 ± 0.101) was significantly higher than that of control group at PCH 48 (0.191 ± 0.024, t = -23.158, P = 0.000). (2) In HMGB1 group, HMGB1 + anti-RAGE antibody group, and HMGB1 + rrRAGE/Fc group, the contents of TNF-α in supernatant were respectively (10.59 ± 1.39), (9.91 ± 1.68), (11.51 ± 2.27) ng/mL; the contents of IL-1β in supernatant were respectively (2.49 ± 0.33), (2.08 ± 0.32), (2.42 ± 0.42) ng/mL; the mRNA levels of TNF-α in cells were respectively 0.311 ± 0.009, 0.301 ± 0.047, 0.326 ± 0.016; the mRNA levels of IL-1β in cells were respectively 0.237 ± 0.021, 0.244 ± 0.041, 0.245 ± 0.013. There were no statistically significant differences in the above indexes among these three groups (with P values all above 0.05). Their levels were all significantly higher than those of control group [with contents of TNF-α and IL-1β in supernatant respectively (2.69 ± 0.14), (0.43 ± 0.05) ng/mL, and mRNA levels of TNF-α and IL-1β in cells respectively 0.140 ± 0.022, 0.077 ± 0.005, P values all below 0.01].</p><p><b>CONCLUSIONS</b>HMGB1 can induce the production of pro-inflammatory cytokines TNF-α and IL-1β from the KC in rats with severe burn. However, RAGE does not play a predominant role in this process.</p>


Subject(s)
Animals , Burns , Metabolism , Cytokines , Metabolism , Disease Models, Animal , HMGB1 Protein , Pharmacology , Interleukin-1beta , Metabolism , Kupffer Cells , Metabolism , Male , Rats , Rats, Sprague-Dawley , Receptor for Advanced Glycation End Products , Receptors, Immunologic , Metabolism , Tumor Necrosis Factor-alpha , Metabolism
20.
Article in Chinese | WPRIM | ID: wpr-318031

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the effect of RNA interfering TLR4 signal pathway on phagocytosis of Kupffer cells.</p><p><b>METHODS</b>RAW2647 mice mononuclear macrophage leukemia cells were observed. The tested group was interfered by Tlr4-mus-1567 RNA which had the best result confirmed by QPCR, cells interfered by Negative Control RNA as NC group, and normal cell as control. We perform the phagocytosis test on each group.</p><p><b>RESULTS</b>The tested group has lower phagocytes percentage than control (17.67% +/- 3.51% vs 32.00% +/- 3.00%, P < 0.01), and lower phagocytic index (46.33% +/- 7.51% vs 82.00% +/- 6.08%, P < 0.01).</p><p><b>CONCLUSIONS</b>Decreased phagocytic activity was observed on Kupffer cells by RNA interference.</p>


Subject(s)
Animals , Kupffer Cells , Allergy and Immunology , Mice , Phagocytosis , RNA Interference , Signal Transduction , Toll-Like Receptor 4 , Genetics , Allergy and Immunology
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