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1.
Braz. j. infect. dis ; 24(1): 13-24, Feb. 2020. tab, graf
Article in English | LILACS | ID: biblio-1089334

ABSTRACT

ABSTRACT Dengue has been a significant public health problem in Colombia since the simultaneous circulation of the four dengue virus serotypes. The replicative fitness of dengue is a biological feature important for virus evolution and contributes to elucidating the behavior of virus populations and viral pathogenesis. However, it has not yet been studied in Colombian isolates. This study aimed to compare the replicative fitness of the four dengue virus serotypes and understand the association between the serotypes, their in vitro infection ability, and their replication in target cells. We used three isolates of each DENV serotype to infect Huh-7 cells at an MOI of 0.5. The percentage of infected cells was evaluated by flow cytometry, cell viability was evaluated by MTT assay, and the pathogenicity index was calculated as a ratio of both parameters. The replicative fitness was measured by the number of viral genome copies produced using quantitative PCR and the production of infectious viral progeny was measured by plaque assay. We showed that Huh-7 cells were susceptible to infection with all the different strain isolates. Nevertheless, the biological characteristics, such as infectious ability and cell viability, were strain-dependent. We also found different degrees of pathogenicity between strains of the four serotypes, representative of the heterogeneity displayed in the circulating population. When we analyzed the replicative fitness using the mean values obtained from RT-qPCR and plaque assay for the different strains, we found serotype-dependent behavior. The highest mean values of replicative fitness were obtained for DENV-1 (log 4.9 PFU/ml) and DENV-4 (log 5.28 PFU/ml), followed by DENV-2 (log 3.9 PFU/ml) and DENV-3 (log 4.31 PFU/ml). The internal heterogeneity of the replicative fitness within each serotype could explain the simultaneous circulation of the four DENV serotypes in Colombia.


Subject(s)
Humans , Virus Replication/genetics , Dengue Virus/genetics , Dengue Virus/pathogenicity , Serogroup , Viral Plaque Assay , Reference Values , Tetrazolium Salts , Time Factors , RNA, Viral/genetics , Cell Line , Cell Survival , Cells, Cultured , Colombia , Reverse Transcriptase Polymerase Chain Reaction , Flow Cytometry , Formazans , Liver/cytology
2.
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
3.
Pesqui. vet. bras ; 38(1): 65-70, Jan. 2018. tab, ilus
Article in English | LILACS, VETINDEX | ID: biblio-895536

ABSTRACT

Histopathological evaluation of liver from 33 pigs slaughtered for human consumption in Amazon region, previously tested by serology and molecular techniques for hepatitis E virus infection (HEV), was analysed in three groups: Group 1, negative for both HEV-RNA and anti-HEV IgG (n=10); Group 2, positive for HEV-RNA (n=13); Group 3, positive for anti-HEV IgG (n=10). Group 2 showed a significant difference among the groups for liver lesions such as lobular activity (P=0.007), periportal interface hepatitis (P=0.004), portal inflammation (P=0.028) hepatitis with lobular, portal and periportal interface activity (P=0.001). HEV detection by immunohistochemistry was performed and 3 of 6 samples of group 2 were positive. Pigs naturally infected by HEV genotype 3 present microscopic necroinflammatory liver lesions similar to HEV in humans. Liver histopathology showed be important in the diagnosis of active asymptomatic HEV infection in pigs slaughtered for human consumption because hepatic liver lesions may present distinct profiles according to molecular and serological diagnosis and in this sense, histopathology and immunohistochemistry may be an important complementary diagnostic tool.(AU)


A avaliação histopatológica hepática de 33 suínos abatidos para consumo humano na região amazônica, previamente testados para infecção pelo vírus da hepatite E (HEV) por sorologia e técnicas moleculares, foi realizada em três grupos: Grupo 1, animais negativos para HEV-RNA e anti-HEV IgG (n=10); Grupo 2, positivos para HEV-RNA (n=13); e Grupo 3, positivos para anti-HEV IgG (n=10). O grupo 2 apresentou diferenças estatísticas significantes entre os grupos em relação à presença de atividade lobular (P=0,007), hepatite periportal de interface (P=0,004), inflamação portal (P= 0.028) e atividade lobular acompanhada por inflamação portal e periportal de interface (P=0,001). A detecção imunohistoquímica do HEV foi realizada e três de seis amostras do Grupo 2 foram positivas. Suínos naturalmente infectados pelo genótipo 3 do HEV apresentam lesões necroinflamatórias no fígado similares a lesão em humanos. A histopatologia hepática demonstrou ser importante no diagnóstico de infecção ativa e assintomática por HEV em suínos abatidos para consumo humano, pois as lesões no fígado apresentaram perfis diferenciados de acordo com o diagnóstico sorológico e molecular da infecção e, neste sentido, a histopatologia e imunohistoquímica podem representar importantes ferramentas complementares de diagnóstico.(AU)


Subject(s)
Animals , Swine/virology , Hepatitis E virus , Genotype , Liver/cytology , Liver/injuries , Immunohistochemistry/veterinary
4.
Braz. j. med. biol. res ; 51(10): e7439, 2018. graf
Article in English | LILACS | ID: biblio-951707

ABSTRACT

Nuclear factor erythroid-related factor 2 (Nrf2) has been implicated in several detoxifying and antioxidant defense processes. Nrf2-mediated heme oxygenase-1 (HO-1) expression was demonstrated to play a key role against oxidative stress. Gastrodin (GSTD) is a well-known active compound isolated from the roots of Rhizoma gastrodiae, a plant used in ancient Chinese traditional medicine. The aim of this work was to investigate whether GSTD could alleviate H2O2-induced oxidative stress in mouse liver sinusoidal endothelial cells (LSECs). In LSECs exposed to 1 mM H2O2, treatment with GSTD (1, 10, or 50 µM) resulted in higher cell viability than the untreated control. Treated cells maintained a higher Bcl2/Bax ratio and suppressed caspase-9 expression compared with untreated cells, reducing cell apoptosis. GSTD was protective for H2O2-induced oxidative injury by reducing the generation of intracellular reactive oxygen species and malondialdehyde. HO-1 and Nrf2 expressions were synergistically upregulated by GSTD. Inhibition of HO-1 by 10 µM zinc protoporphyrin resulted in less protective effects on cell viability and malondialdehyde reduction by GSTD treatment in H2O2-exposed LSECs. Additionally, phosphorylated p38 in LSECs exposed to H2O2 was elevated by GSTD. Inhibition of p38 phosphorylation by SB203580 did not induce Nrf2 and HO-1 expression after 1 or 10 µM GSTD treatment and the protective effect on cell viability and malondialdehyde reduction in H2O2-exposed LSECs was reduced. The data conclusively demonstrated that GSTD-induced HO-1 and Nrf2 expression is involved in protection of LSECs from H2O2-induced oxidative injury, which may be regulated by p38 phosphorylation.


Subject(s)
Animals , Rabbits , Benzyl Alcohols/pharmacology , Endothelial Cells/drug effects , p38 Mitogen-Activated Protein Kinases/metabolism , Heme Oxygenase-1/metabolism , Glucosides/pharmacology , Hydrogen Peroxide/pharmacology , Up-Regulation/drug effects , Cell Survival/drug effects , Apoptosis/drug effects , Liver/cytology , Liver/drug effects , Malondialdehyde/metabolism , Models, Theoretical
5.
Int. j. morphol ; 35(4): 1285-1290, Dec. 2017. graf
Article in English | LILACS | ID: biblio-893129

ABSTRACT

SUMMARY: Ghrelin is the endogenous ligand for the growth hormone secretagogue receptor, and has been found in the liver of multiple vertebrates. While ghrelin has been identified in the gastrointestinal tract of African ostrich chicks, little is known regarding its distribution in the liver of the African ostrich. In the present study, the distribution and morphological characteristics of ghrelin-immunopositive (ghrelin-ip) cells in the liver of the African ostrich were investigated using immunohistochemistry. Our results indicate that the liver is divided into two sections: the capsule and the parenchyma, which comprises hepatic lobules and the hepatic portal area. The hepatic lobules include the central vein, hepatocellular cord, and the hepatic sinusoid. The hepatocellular cord is composed of hepatocytes, and Macrophagocytus stellatus (Kupffer cells) as well as endothelial cells reside within the hepatic sinusoid. ghrelin-ip cells were detected among both the Macrophagocytus stellatus and endothelial cells of the hepatic sinusoid in the African ostrich liver. In contrast, no ghrelinip cells were located within the hepatocytes or the hepatic portal area. These results clearly demonstrated the presence of ghrelin-ip cells in the liver of the African ostrich. Therefore, ghrelin may have a physiological function in the liver of the African ostrich.


RESUMEN: La ghrelina es el ligando endógeno para el receptor secretagogo de la hormona del crecimiento, y se ha encontrado en el hígado de múltiples vertebrados. A pesar que la ghrelina ha sido identificada en el tracto gastrointestinal de polluelos de avestruz africanas, poco se sabe sobre su distribución en el hígado de esta ave. En el presente estudio se investigó la distribución y características morfológicas de las células ghrelininmunopositivas (ghrelin-ip) en el hígado del avestruz africana mediante inmunohistoquímica. Nuestros resultados indican que el hígado se divide en dos secciones: la cápsula y el parénquima, que comprende los lóbulos hepáticos y el área portal hepática. Los lóbulos hepáticos incluyen la vena central, el cordón hepatocelular y el sinusoide hepático. El cordón hepatocelular está compuesto de hepatocitos y de Macrophagocytus stellatus (células de Kupffer) y las células endoteliales se localizan dentro del sinusoide hepático. Fueron detectacas células ghrelin-ip entre los Macrophagocytus stellatus y las células endoteliales del sinusoide hepático en el hígado de avestruz africana. En contraste, no se localizaron células de ghrelin-ip dentro de los hepatocitos o en el área portal hepática. Estos resultados demuestran claramente la presencia de células de ghrelin-ip en el hígado. Por lo tanto, la ghrelina puede tener una función fisiológica en el hígado de avestruz africana.


Subject(s)
Animals , Struthioniformes/anatomy & histology , Ghrelin/metabolism , Liver/cytology , Immunohistochemistry
6.
Ann. hepatol ; 16(2): 297-303, Mar.-Apr. 2017. tab, graf
Article in English | LILACS | ID: biblio-887236

ABSTRACT

ABSTRACT Introduction and aim. The inability to distinguish cancer (CSCs) from normal stem cells (NSCs) has hindered attempts to identify safer, more effective therapies for hepatocellular carcinoma (HCC). The aim of this study was to document and compare cell membrane potential differences (PDs) of CSCs and NSCs derived from human HCC and healthy livers respectively and determine whether altered GABAergic innervation could explain the differences. Material and methods. Epithelial cell adhesion molecule (EpCAM) positive stem cells were isolated from human liver tissues by magnetic bead separations. Cellular PDs were recorded by microelectrode impalement of freshly isolated cells. GABAA receptor subunit expression was documented by reverse transcriptase polymerase chain reaction (RT-PCR) and immunofluorescence. Results. CSCs were significantly depolarized (-7.0 ± 1.3 mV) relative to NSCs (-23.0 ± 1.4 mV, p < 0.01). The depolarized state was associated with different GABAA receptor subunit expression profiles wherein phasic transmission, represented by GAGAA α3 subunit expression, was prevalent in CSCs while tonic transmission, represented by GABAA α6 subunit expression, prevailed in NSCs. In addition, GABAA subunits α3, β3, γ3 and δ were strongly expressed in CSCs while GABAA π expression was dominant in NSCs. CSCs and NSCs responded similarly to GABAA receptor agonists (ΔPD: 12.5 ± 1.2 mV and 11.0 ± 3.5 mV respectively). Conclusion. The results of this study indicate that CSCs are significantly depolarized relative to NSCs and these differences are associated with differences in GABAA receptor subunit expression. Together they provide new insights into the pathogenesis and possible treatment of human HCC.


Subject(s)
Humans , Neoplastic Stem Cells/metabolism , Receptors, GABA-A/metabolism , Carcinoma, Hepatocellular/genetics , Carcinoma, Hepatocellular/metabolism , GABA-A Receptor Agonists/pharmacology , Epithelial Cell Adhesion Molecule/metabolism , Liver/cytology , Liver Neoplasms/metabolism , Phenotype , Stem Cells/drug effects , Neoplastic Stem Cells/drug effects , RNA, Messenger/genetics , RNA, Messenger/metabolism , Biomarkers/metabolism , Fluorescent Antibody Technique , Immunomagnetic Separation , Receptors, GABA-A/drug effects , Receptors, GABA-A/genetics , Reverse Transcriptase Polymerase Chain Reaction , Protein Subunits , Liver Neoplasms/genetics , Membrane Potentials/drug effects
7.
Pesqui. vet. bras ; 36(supl.1): 101-105, June 2016. tab, graf, ilus
Article in English | LILACS, VETINDEX | ID: lil-798008

ABSTRACT

Regarding the importance of starvation and refeeding and the occurrence of refeeding syndrome in various conditions, the present study was conducted to investigate the effects of refeeding on some parameters of energy metabolism and electrolytes and changes of hepatic tissue in male Wistar rats. Fifty-seven rats were divided into six groups, having 6 to 11 rats. Food was provided ad-libitum until three months and then the first group was considered without starvation (day 0). Other rats were fasted for two weeks. Group 2 was applied to a group immediately after starvation (day 14). Groups 3 to 6 were refed in days 16 till 22, respectively. At the end of each period, blood and tissue samples were taken and histopathological and serum analysis, including serum electrolytes (calcium, phosphorus, sodium, potassium), the energy parameters (glucose, insulin, cortisol) and the liver enzymes (ALT, AST, ALP) were determined. Insulin decreased by starvation and then showed an increasing trend compared to starvation period, which the highest amount of this parameter was observed eight days post-refeeding. Serum glucose level showed the opposite pattern of insulin. Histopathological examination of the tissue sections revealed clear vacuoles after starvation and refeeding, in which the severity of lesions gradually decreased during refeeding. The cortisol level decreased by starvation and then increased during refeeding. Also, potassium and phosphorus concentrations declined by refeeding and the serum sodium and potassium levels were changed in the relatively opposite manner. The calcium level decreased by starvation and then increased during refeeding. These results could help recognize and remedy the refeeding syndrome.(AU)


Subject(s)
Animals , Rats , Electrolytes/metabolism , Fasting/metabolism , Liver/anatomy & histology , Liver/cytology , Rats, Wistar/metabolism , Metabolism/physiology
8.
Gut and Liver ; : 166-176, 2016.
Article in English | WPRIM | ID: wpr-25631

ABSTRACT

Mesothelial cells (MCs) cover the surface of visceral organs and the parietal walls of cavities, and they synthesize lubricating fluids to create a slippery surface that facilitates movement between organs without friction. Recent studies have indicated that MCs play active roles in liver development, fibrosis, and regeneration. During liver development, the mesoderm produces MCs that form a single epithelial layer of the mesothelium. MCs exhibit an intermediate phenotype between epithelial cells and mesenchymal cells. Lineage tracing studies have indicated that during liver development, MCs act as mesenchymal progenitor cells that produce hepatic stellate cells, fibroblasts around blood vessels, and smooth muscle cells. Upon liver injury, MCs migrate inward from the liver surface and produce hepatic stellate cells or myofibroblast depending on the etiology, suggesting that MCs are the source of myofibroblasts in capsular fibrosis. Similar to the activation of hepatic stellate cells, transforming growth factor β induces the conversion of MCs into myofibroblasts. Further elucidation of the biological and molecular changes involved in MC activation and fibrogenesis will contribute to the development of novel approaches for the prevention and therapy of liver fibrosis.


Subject(s)
Epithelial Cells/physiology , Epithelium/metabolism , Hepatic Stellate Cells/physiology , Humans , Liver/cytology , Liver Cirrhosis/etiology , Liver Regeneration/physiology , Mesenchymal Stem Cells/physiology , Myofibroblasts/physiology
9.
Arq. bras. med. vet. zootec ; 67(4): 1188-1192, July-Aug. 2015. ilus
Article in English | LILACS, VETINDEX | ID: biblio-1095959

ABSTRACT

A cirurgia endoscópica por orifícios naturais (NOTES) representa um novo conceito de cirurgia, caracterizada por ausência de incisões abdominais. Os acessos mais comumente usados são o transvaginal e o transgástrico. Entretanto, as rotas transcolônica e transretal representam alternativas promissoras. O presente estudo objetiva avaliar três diferentes técnicas de sutura retal em três suínos submetidos a NOTES transretal para biópsia hepática, avaliando-se concomitantemente as repercussões clínicas e hematológicas. Sob anestesia geral, foi realizada uma incisão transversal no reto para a passagem do endoscópio até a cavidade abdominal em todos os animais para a realização da biópsia hepática. Cada animal recebeu um tipo de sutura retal: sutura em dois planos; reforço com tela de polipropileno ou reforço com membrana de pericárdio bovino. A NOTES transretal em modelo experimental suíno não apresentou implicações clínicas e hematológicas importantes, o que demonstra um acesso alternativo para biópsia hepática. Nenhum animal apresentou sinais de peritonite, aderências ou deiscência de pontos. O uso de reforço com pericárdio bovino para a sutura retal apresenta um atraso na cicatrização quando comparado com a sutura convencional ou com o uso de tela de polipropileno.(AU)


Subject(s)
Animals , Rectum/diagnostic imaging , Swine/surgery , Biopsy/veterinary , Suture Techniques/veterinary , Endoscopy/veterinary , Liver/cytology
10.
Indian J Biochem Biophys ; 2014 Jun; 51(3): 215-222
Article in English | IMSEAR | ID: sea-154231

ABSTRACT

Alcoholism and obesity are strongly associated with several disorders including heart and liver diseases. This study evaluated the effects of rutin treatment in serum, heart and liver tissues of rats subjected to a combination of hypercaloric diet (HD) and chronic ethanol consumption. Rats were divided into three groups: Control: rats fed a standard diet and drinking water ad libitum; G1: rats fed the HD and receiving a solution of 10% (v/v) ethanol; and G2: rats fed the HD and ethanol solution, followed by injections of 50 mg/kg-1 rutin as treatment. After 53 days of HD and ethanol exposure, the rutin was administered every three days for nine days. At the end of the experimental period (95 days), biochemical analyses were carried out on sera, cardiac and hepatic tissues. Body weight gain and food consumption were reduced in both the G1 and G2 groups compared to control animals. Rutin effectively reduced the total lipids (TL), triglycerides (TG), total cholesterol (TC), VLDL, LDL-cholesterol and glucose levels, while it increased the HDL-cholesterol in the serum of G2 rats, compared to G1. Although rutin had no effect on total protein, albumin, uric acid and cretinine levels, it was able to restore serum activities of alkaline phosphatase (ALP), lactate dehydrogenase (LDH), aspartate aminotransferase (AST), alanine aminotransferase (ALT) and creatine kinase (CK) in animals fed HD and receiving ethanol. Glycogen stores were replenished in both hepatic and cardiac tissues after rutin treatment. Moreover, rutin consistently reduced hepatic levels of TG and TC and cardiac AST, ALT and CK activities. Thus, rutin treatment was effective in reducing the risk factors for cardiac and hepatic disease caused by both HD and chronic ethanol consumption.


Subject(s)
Alanine Transaminase/metabolism , /metabolism , Animals , Aspartate Aminotransferases/metabolism , Central Nervous System Depressants/toxicity , /metabolism , Biomarkers/metabolism , /adverse effects , Ethanol/toxicity , Glycemic Index/drug effects , Heart/drug effects , Heart/physiology , L-Lactate Dehydrogenase/metabolism , Lipids/analysis , Liver/cytology , Liver/metabolism , Male , Rats , Rats, Wistar , Rutin/pharmacology
11.
Indian J Biochem Biophys ; 2014 Feb; 51(1): 37-45
Article in English | IMSEAR | ID: sea-154229

ABSTRACT

The hepatoprotective potential of aqueous Azadirachta indica leaf extract (AAILE) was assessed against DMBA-induced hepatotoxicity. DMBA  (7,12-dimethylbenz[a] anthracene) treatment (40 mg/kg body weight, ip) to male Balb/c mice resulted in the derailment of liver function as revealed by extremely slow clearance of 99mTc-mebrofenin from liver, elevated levels of alkaline phosphatase (ALP) and alanine transaminase (ALT), compared to control group. In addition, elevated micronuclei score and high apoptotic index indicated hepatogenotoxicity in DMBA-treated mice. DMBA treatment also upregulated cytochrome P450 (CYP), cytochrome b5 (Cyt b5) and decreased glutathione-S-transferase activity in hepatic tissue, compared to control group. Enhanced lipid peroxidation (LPO) levels along with decreased reduced glutathione (GSH) level were also observed in DMBA group, compared to control group. AAILE co-treatment (200 mg/kg body weight, po, thrice a week) for 8 weeks followed by DMBA injection showed significant improvement in hepatic status, as revealed by normalization of 99mTc-mebrofenin clearance rate, decreased ALP and ALT levels, reduced genotoxicity in terms of micronuclei score and apoptotic index. Levels of LPO were significantly decreased along with increased hepatic GST and GSH levels in AAILE + DMBA group, compared to DMBA group. However, no significant change was observed in hepatic CYP and Cyt b5 levels, compared to DMBA group. The results indicated that AAILE effectively ameliorated DMBA-induced hepatotoxicity.


Subject(s)
9,10-Dimethyl-1,2-benzanthracene/toxicity , Animals , Azadirachta/chemistry , Cell Division/drug effects , Cytoprotection/drug effects , Liver/cytology , Liver/drug effects , Liver/metabolism , Liver/toxicity , Male , Mice , Mice, Inbred BALB C , Micronucleus Tests , Oxidative Stress , Plant Extracts/pharmacology , Plant Leaves/chemistry , Radiometry
12.
Egyptian Journal of Hospital Medicine [The]. 2014; 57 (October): 554-564
in English | IMEMR | ID: emr-160253

ABSTRACT

Nanoparticles of silver have many important applications and are among the most commonly used nanomaterials. They are increasingly used in a variety of both medical and consumer products which includes: spectrally selective coating for solar energy absorption and intercalation material for electrical batteries, as optical receptors, polarizing filters, catalysts in chemical reaction and bio-labeling. Nanosilver [Ag-NP] has both antibacterial and antiviral activity. Yet, the knowledge about the systemic toxicity of nanosilver is relatively limited. The aim of work: To evaluate the potential toxicity of small size 10nm silver nanoparticles using two different doses [0.1 ml and 0.4 ml] focusing on the ultrastructural changes occurring in mice hepatocytes. This study was performed using three groups of mice. The animals of the first group were given a daily intravenous injection of 0.1 ml of silver nanoparticles for 28 consecutive days. The second group was treated with 0.4 ml of silver nanoparticles for 28 consecutive days. The third group served as a control group in which the animals did not receive any vehicle. The study was focused on the ultrastructure of the liver. Ultrastructure observations of liver cells of mice Treated with any of the two doses [0.1 and 0.4 ml] of 10 nm Ag-NP indicated severe accumulation of dark deposits of Ag-NP in the cytoplasm and the cell organelles. Our study revealed that nanosilver used in doses of 0.1 and 0.4 ml led to deposits in the cells and induced damage of cell components especially the nucleus, mitochondria and chromatin


Subject(s)
Male , Animals, Laboratory , Metal Nanoparticles/toxicity , Cytotoxicity Tests, Immunologic/methods , Liver/cytology , Liver/ultrastructure , Mice
13.
Indian J Biochem Biophys ; 2013 Dec; 50(6): 511-520
Article in English | IMSEAR | ID: sea-150265

ABSTRACT

The study focuses on the importance of Tyr11 amino acid (AA) and subsequent stereochemistry involved in the binding process of neurotensin (NT) with its receptor (NTR)/binding protein(s) as well as the size heterogeneity. Using the binding of 125I-NT with several chicken tissues, it is identified that one of the crucial factors behind all high affinity (Kd ~10 pM) interactions is due to phenolic-OH (Φ-OH) at the para (p) position of Tyr11 within RRPYIL-CO2H (NT8-13) sequence. Replacing the p-OH only in Tyr11 by substituting with p-Cl, p-F and p-NH2 results in significant change of the binding affinity (Kd); p-OH ≈ p-NH2 (~10 pM), p-Cl (~100 pM), p-F (~120 pM). Interestingly, p-NH2 equals to p-OH displaying the highest affinity. Experiments conducted by binding several of the 125I-azido–NT analogs having azido group attached at different positions within the NT molecule have further confirmed the necessity of RRPYIL sequence for high affinity ligand-receptor interaction. The role of Tryp11 in place of Tyr11 in addition to the results above establishes a significant possibility of H–bonding occurring between p-OH of NT and NTR inside the docking space. Photo labeling of the liver tissue by substituted 125I-Y3-azido-NT analogs shows several specifically labeled bands with considerable range of molecular weight (Mr ~90-30 kDa) variations. These results indicate the existence of molecular heterogeneity concerning the sizes of NTR or else any NT binding proteins in the avian tissues. Further, the study has revealed that besides liver, several other chicken tissues also express similar specific high affinity binding (Kd ~20 pM) with varying capacities (Bmax). The order for Bmax is: liver (1.2 pMol/mg) gall bladder (1.03 pMol/mg) > spleen (0.43 pMol/mg) > brain (0.3 pMol/mg) > colon lung (0.15 pMol/mg). In all cases, the binding was reduced by GTPgS (ED50 ~ 0.05 nM), NEM (ED50 ~ 0.50 mM) and NaCl (ED50 ~30 mM), indicating the existence of NTR identical to the mammalian type-1.


Subject(s)
Amino Acid Sequence , Amino Acid Substitution , Animals , Azides/chemistry , Binding, Competitive , Cell Membrane/metabolism , Chickens , Ethylmaleimide/pharmacology , Female , Guanosine 5'-O-(3-Thiotriphosphate) , Liver/cytology , Male , Molecular Weight , Neurotensin/chemistry , Neurotensin/genetics , Neurotensin/metabolism , Protein Binding/drug effects , Pyrazoles/pharmacology , Quinolines/pharmacology , Receptors, Neurotensin/antagonists & inhibitors , Receptors, Neurotensin/chemistry , Receptors, Neurotensin/metabolism , Sodium Chloride/pharmacology , Stereoisomerism , Tyrosine
14.
Acta cir. bras ; 28(9): 657-663, Sept. 2013. ilus
Article in English | LILACS | ID: lil-684440

ABSTRACT

PURPOSE: To investigate the impact of selective hepatic artery clamping (SHAC) in hepatocellular function. METHODS: Three groups of Wistar male rats were subjected to SHAC ischemia period of 60min: Group A continuous SHAC were subjected to SHAC ischemia period of 60min, Group B intermittent SHAC of 30min with 5min of reperfusion and Group C intermittent SHAC of 15min with 5min of reperfusion. Animals without SHAC were included-Group D. To evaluate hepatocellular function blood markers and hepatic extraction function (HEF) using 99mTc-mebrofenin were performed before and after surgery. Flow cytometry was used to analyze oxidative stress and cell viability. RESULTS: A mortality rate of 7.6% in Group A was observed. HEF maintained normal values between the groups. Flow cytometry demonstrated no significant differences between the groups in viability, type of cell death as well as in the production of reactive oxygen species. CONCLUSIONS: The selective hepatic artery clamping compared to other clamping techniques results on increased cell viability and decreased hepatocyte death. The SHAC is a potential alternative to decrease per-operative bleeding while maintaining hepatocellular function.


Subject(s)
Animals , Male , Rats , Hepatic Artery/surgery , Hepatocytes/physiology , Liver/blood supply , Liver/cytology , Cell Survival , Constriction , Flow Cytometry , Ischemia/physiopathology , Models, Animal , Oxidative Stress , Peroxides/analysis , Rats, Wistar , Reperfusion , Reactive Oxygen Species/metabolism , Time Factors
15.
Braz. j. med. biol. res ; 46(7): 559-566, ago. 2013. graf
Article in English | LILACS | ID: lil-682396

ABSTRACT

Hepatic progenitor cells (HPCs) are a potential cell source for liver cell transplantation but do not function like mature liver cells. We sought an effective and reliable method to induce HPC maturation. An immortalized HP14.5 albumin promoter-driven Gaussian luciferase (ALB-GLuc) cell line was established from HPCs isolated from fetal mouse liver of post coitus day 14.5 mice to investigate the effect of induction factors on ALB promoter. HP14.5 parental cells were cultured in DMEM with different combinations of 2% horse serum (HS), 0.1 µM dexamethasone (DEX), 10 ng/mL hepatic growth factor (HGF), and/or 20 ng/mL fibroblast growth factor 4 (FGF4). Trypan blue and crystal violet staining were used to assess cell proliferation with different induction conditions. Expression of hepatic markers was measured by semi-quantitative RT-PCR, Western blot, and immunofluorescence. Glycogen storage and metabolism were detected by periodic acid-Schiff and indocyanine green (ICG) staining. GLuc activity indicated ALB expression. The combination of 2% HS+0.1 µM Dex+10 ng/mL HGF+20 ng/mL FGF4 induced the highest ALB-GLuc activity. Cell proliferation decreased in 2% HS but increased by adding FGF4. Upon induction, and consistent with hepatocyte development, DLK, AFP, and CK19 expression decreased, while ALB, CK18, and UGT1A expression increased. The maturity markers tyrosine aminotransferase and apolipoprotein B were detected at days 3 and 6 post-induction, respectively. ICG uptake and glycogen synthesis were detectable at day 6 and increased over time. Therefore, we demonstrated that HPCs were induced to differentiate into functional mature hepatocytes in vitro, suggesting that factor-treated HPCs may be further explored as a means of liver cell transplantation.


Subject(s)
Animals , Mice , Cell Differentiation/drug effects , Embryo, Mammalian/drug effects , Hepatocytes/cytology , Liver/cytology , Stem Cells/drug effects , Antigens, Differentiation/analysis , Apolipoproteins B/isolation & purification , Cell Proliferation , Dexamethasone/administration & dosage , Fibroblast Growth Factors/administration & dosage , Gentian Violet , Glycogen/metabolism , Hepatocyte Growth Factor/administration & dosage , Indocyanine Green/pharmacokinetics , Primary Cell Culture/methods , Reverse Transcriptase Polymerase Chain Reaction , Stem Cells/cytology , Trypan Blue , Tyrosine Transaminase/isolation & purification
16.
Biocell ; 37(2): 45-54, ago. 2013. ilus, graf
Article in English | LILACS | ID: lil-708027

ABSTRACT

Cadmium (Cd) induces several effects in different tissues, but our knowledge of the toxic effects on organelles is insufficient. To observe the progression of Cd effects on organelle structure and function, HuH-7 cells (human hepatic carcinoma cell line) were exposed to CdCl2 in increasing concentrations (1 microM - 20 microM) and exposure times (2 h - 24 h). During Cd treatment, the cells exhibited a progressive decrease in viability that was both time- and dose-dependent. Cd treated cells displayed progressive morphological changes that included cytoplasm retraction and nuclear condensation preceding a total loss of cell adhesion. Treatment with 10 microM for 12 h led to irreversible damages. Before these drastic and irreparable damages, treated cells (5 microM for 12 h) presented a progressive loss of mitochond rial function and cytoplasm acidification as well as dysfunction and disorganization of microfilaments and endoplasmic reticulum. These damages led to the induction of apoptotic events and an increase in autophagic bodies in the cytoplasm. These results revealed that Cd affects multiple intra-cellular targets that induce alterations in the mitochondria, cytoskeleton, endoplasmic reticulum and acidic compartments, ultimately culminating in cell death via apoptotic and autophagic pathways.


Subject(s)
Humans , Apoptosis , Autophagy , Cadmium/toxicity , Liver , Organelles , Carcinoma, Hepatocellular , Cell Line, Tumor , Liver/cytology , Liver Neoplasms
17.
Acta cir. bras ; 28(3): 210-215, Mar. 2013. ilus, tab
Article in English | LILACS | ID: lil-667932

ABSTRACT

PURPOSE: To evaluate if the ileum resection changes the functioning liver cell mass, the hepatic metabolism and the biodistribution of radiopharmaceutical in rats. METHODS: Twelve Wistar rats weighing 285g±34g were randomly divided into the ileum resection group (n = 6) and sham group rats (n = 6). After 30 days, they were anesthetized and 0.1mL of 99m-Tc-phytate (0.66MBq) was injected via femoral vein. After 30 minutes, blood samples were collected for red blood cells radioactive labeling and serum ALT, AST and gammaGT. Liver samples were used for 99m-Tc-phytate percentage of radioactivity/gram of tissue and histopathology. Student 's t test was used with significance 0.05. RESULTS: There was a higher uptake of 99m-Tc-phytate in the liver of sham rats, compared to the ileum resection group (p<0.05). GammaGT, ALT and AST were increased in ileum resection rats compared to sham (p<0.05). The he patocytes count was significantly lower in ileum resection group than in sham (p<0.05). Liver: body mass ratio was lower in experimental animals than in sham group (p<0.05). CONCLUSION: These data support that the ileum has important role in liver function and liver mass regulation, and they have potential clinical implications regarding the pathogenesis of liver injury following lower bowel resection.


Subject(s)
Animals , Rats , Ileum/physiology , Liver/anatomy & histology , Liver/physiology , Alanine Transaminase/blood , Aspartate Aminotransferases/blood , Hepatocytes , Ileum/surgery , Liver/cytology , Liver , Organ Size/physiology , Organotechnetium Compounds , Phytic Acid , Random Allocation , Rats, Wistar , Radiopharmaceuticals , Time Factors , gamma-Glutamyltransferase/blood
18.
AJMB-Avicenna Journal of Medical Biotechnology. 2013; 5 (2): 104-117
in English | IMEMR | ID: emr-142798

ABSTRACT

Despite major progress in our general knowledge related to the application of adult stem cells, finding alternative sources for bone marrow Mesenchymal Stem Cells [MSCs] has remained to be challenged. In this study successful isolation, multilineage differentiation, and proliferation potentials of sheep MSCs derived from bone marrow, adipose tissue, and liver were widely investigated. The primary cell cultures were prepared form tissue samples obtained from sheep 30-35 day fetus. Passage-3 cells were plated either at varying cell densities or different serum concentrations for a week. The Population Doubling Time [PDT], growth curves, and Colony Forming Unit [CFU] of MSCs was determined. The stemness and trilineage differentiation potential of MSCs were analyzed by using molecullar and cytochemical staining approaches. The data was analyzed through one way ANOVA using SigmaStat [ver. 2]. The highest PDT and lowest CFU were observed in adipose tissue group compared with other groups [p<0.001]. Comparing different serum concentrations [5, 10, 15, and 20%], irrespective of cell sources, the highest proliferation rate was achieved in the presence of 20% serum [p<0.001]. Additionally, there was an inverse relation between cell seeding density at culture initiation and proliferation rate, except for L-MSC at 300 cell seeding density. All three sources of fetal sheep MSCs had the identical trilineage differentiation potential. The proliferative capacity of liver and bone marrow derived MSCs were similar at different cell seeding densities except for the higher fold increase in B-MSCs at 2700 cells/cm2 density. Moreover, the adipose tissue derived MSCs had the lowest proliferative indices


Subject(s)
Animals , Bone Marrow Cells/cytology , Cell Differentiation/physiology , Cells, Cultured , Colony-Forming Units Assay , Liver/cytology , Cells , Adipose Tissue/cytology , Microscopy, Electron, Scanning Transmission , Cell Culture Techniques
19.
Acta cir. bras ; 27(7): 460-464, jul. 2012. graf, tab
Article in English | LILACS | ID: lil-640093

ABSTRACT

PURPOSE: To determine the impact of hypertension in liver regeneration, in rats by examining gain in liver mass and the replication of hepatocytes and stellate cells. METHODS: Forty Wistar rats were allocated into two groups of twenty, the control and experiment group. The experiment group animals were submitted to induction of renovascular hypertension. A week later, all the animals underwent a partial hepatectomy. Measurements were taken after 24 hours and seven days, when ten animals in each group were euthanized. Thus, four subgroups were obtained. The livers were excised and sent for histopathological analysis. RESULTS: The control group had a greater gain in liver mass than the experiment group seven days after partial hepatectomy (p=0.0051). The difference in the activate stellate cell count was not statistically significant following analysis after both 24 hours and seven days (p=1.0). A higher number of dividing hepatocytes was observed in the control group seven days after partial hepatectomy (p=0.0014). CONCLUSION: In rats, hypertension had no direct influence on stellate cell replication, but led to a delay in liver mass gain and were shown to be a reduction factor on hepatocyte replication seven7 days after partial hepatectomy.


OBJETIVO: Determinar o impacto da hipertensão arterial sistêmica na regeneração hepática, em ratos, através da análise do ganho de massa hepática e da replicação dos hepatócitos e das células estreladas. MÉTODOS: Alocaram-se 40 ratos Wistar em dois grupos de 20 animais, os grupos controle e experimento. Os do grupo experimento submeteram-se a indução da hipertensão renovascular. Uma semana após, realizou-se hepatectomia parcial em todos os animais. Colheram-se os dados com 24 horas e sete dias, quando dez animais de cada grupo submeteram-se a eutanásia. Assim, obtiveram-se quatro subgrupos. Os fígados foram retirados e enviados para análise histopatológica. RESULTADOS: O grupo controle apresentou maior ganho de massa hepática do que o grupo experimento sete dias após a hepatectomia parcial (p=0,0051). A diferença na contagem das células estreladas ativadas não foi estatisticamente significante nas análises de 24 horas e de sete dias (p=1,0). Um maior número de hepatócitos em divisão foi observado no grupo controle, sete dias após a hepatectomia parcial (p=0,0014). CONCLUSÃO: Em ratos, a hipertensão não teve influência direta sobre a replicação de células estreladas, mas levou ao atraso no ganho de massa hepática e mostrou ser um fator de redução na replicação de hepatócitos sete dias após a hepatectomia parcial.


Subject(s)
Animals , Male , Rats , Hepatic Stellate Cells/physiology , Hepatocytes/physiology , Hypertension/physiopathology , Liver Regeneration/physiology , Liver/physiology , Cell Count , Hepatectomy , Liver/cytology , Organ Size , Random Allocation , Rats, Wistar , Time Factors
20.
Int. j. morphol ; 30(2): 467-472, jun. 2012. ilus
Article in Spanish | LILACS | ID: lil-651815

ABSTRACT

Hígados de ratas Sprague Dawley fueron irradiados con dosis diarias de 6 J/cm2 emitida por el láser AsGa equivalente a 904 nm durante 15 días De estos animales previamente anestesiados fueron sacrificados transcurridos 5, 10, 30, 45 y 60 días post irradiación para posteriormente obtener quirúrgicamente muestras de hígado y ser procesadas para microscopía electrónica de transmisión, aplicando técnicas morfométricas utilizando aumentos de 8.500 X con especial énfasis en cuantificar fracciones volumétricas de componentes celulares con el objetivo de precisar la duración de las estimulaciones infrarrojas. El análisis de los resultados entre hepatocitos controles e irradiados con dosis de 6 J/cm2 y tiempo de estimulación infrarroja revela que existen marcadas diferencias entre las fracciones volumétricas de componentes celulares determinantes de funcionalidad celular e involucrados en síntesis proteica, cuantificación que demuestra claramente que el efecto del láser infrarrojo persiste hasta los 30 días post estimulación, evidenciándose modificaciones de organelos que revelan alta funcionalidad, mientras que sobre este tiempo es observada una notable inhibición de dicha funcionalidad, concluyéndose entonces que los efectos de radiación infrarroja persisten en tiempos precisos provocando en los hepatocitos una drástica transformación en sus componentes y por ende en su funcionalidad. en estas células de elevado metabolismo.


Livers of Sprague Dawley rats were irradiated with daily doses of 6 J/cm2 emitted by a laser AsGa, equivalent to 904 nm during 15 days. Experiment animals were anaesthetised and killed after 5, 10, 30, 45 and 60 days post irradiation, in order to obtain samples of liver by surgery. These were processed for transmission electron microscopy, and morphometric techniques were applied using 8,500 X magnification with special emphasis on measuring the volumetric fractions of cell components in order to determine the duration of infrared stimulation. Analysis of the results between control hepatocytes and those irradiated with doses of 6 J/cm2 and by period after infra-red stimulation revealed the existence of marked differences between the volumetric fractions of cell components which determine cell function or are involved in protein synthesis. The measurements show clearly that the effect of the infrared laser persists up to 30 days post stimulation, with evidence of modifications of organelles revealing high functioning, while after 30 days a notable inhibition of this functioning is observed. It is therefore concluded that the effects of infrared radiation persist for precise times, provoking a drastic transformation in hepatocyte components, and thus the functioning of these high-metabolism cells.


Subject(s)
Animals , Rats , Hepatocytes/radiation effects , Infrared Rays , Hepatocytes/ultrastructure , Liver/cytology , Liver/radiation effects , Rats, Sprague-Dawley , Time Factors
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