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1.
Life Sci ; 241: 117121, 2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31794772

RESUMO

AIMS: This study addressed whether endogenous hepatic stem/progenitor (HSP) cells survival were related to the injured signals during liver cirrhosis. MATERIAL AND METHODS: Liver cirrhosis was induced in C57BL/6 mice by administering diethylnitrosamine (DEN) in drinking water. Hematoxylin-eosin staining and Masson's trichrome staining were used to identify infiltrative cells and connective tissues, respectively. The inflammatory activity grade and fibrotic stage, represented as G and S respectively, and evaluated by the International Simplified Grading and Staging System (ISGSS). Endogenous HSP cells (Ng2+HSP) were identified by immunofluorescence staining with an anti-neuro/glial antigen 2 (Ng2) antibody. All data were analyzed using SPSS 22.0 and GraphPad Prism 6 and Student's t-test was performed to determine statistical significance. p-values < 0.05 were considered statistically significant. KEY FINDINGS: All mice administered oral DEN developed liver fibrosis, liver cirrhosis and hepatocellular carcinoma (HCC). During the fibrosis period, observed a positive correlation of survival of endogenous HSP (Ng2+HSP) cells with inflammatory activity. As the disease developed into the cirrhotic stage, when lost correlation of endogenous HSP cells with inflammatory activity, revealed a dramatically reduced survival rate of endogenous HSP (Ng2+HSP) cells. SIGNIFICANCE: The DEN-induced liver cirrhosis could develop into three time zone of fibrosis, cirrhosis and cancer, and the histological patterns in the model are similar to those described in humans. Dramatic survival and less apoptosis of endogenous HSP (Ng2+HSP) cells was within fibrotic state, where inflammation signals is strong, indicating such time zone (1-6 weeks) during liver cirrhosis is important for mobilizing endogenous HSP-based regeneration.


Assuntos
Carcinoma Hepatocelular/terapia , Inflamação/terapia , Cirrose Hepática/terapia , Neoplasias Hepáticas Experimentais/terapia , Fígado/citologia , Células-Tronco/citologia , Animais , Carcinoma Hepatocelular/etiologia , Dietilnitrosamina/toxicidade , Modelos Animais de Doenças , Inflamação/etiologia , Fígado/imunologia , Cirrose Hepática/induzido quimicamente , Cirrose Hepática/imunologia , Cirrose Hepática/patologia , Neoplasias Hepáticas Experimentais/etiologia , Regeneração Hepática , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Células-Tronco/fisiologia
2.
Chemosphere ; 239: 124747, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31514003

RESUMO

BACKGROUNDS: Polychlorinated biphenyls are persistent environmental pollutants associated with the onset of non-alcoholic fatty liver disease in humans, but there is limited information on the underlying mechanism. In the present study, we investigated the alterations in gene expression profiles in normal human liver cells L-02 following exposure to 2, 3, 3', 4, 4', 5 - hexachlorobiphenyl (PCB 156), a potent compound that may induce non-alcoholic fatty liver disease. METHODS: The L-02 cells were exposed to PCB 156 for 72 h and the contents of intracellular triacylglyceride and total cholesterol were subsequently measured. Microarray analysis of mRNAs and long non-coding RNAs (lncRNAs) in the cells was also performed after 3.4 µM PCB 156 treatment. RESULTS: Exposure to PCB 156 (3.4 µM, 72 h) resulted in significant increases of triacylglyceride and total cholesterol concentrations in L-02 cells. Microarray analysis identified 222 differentially expressed mRNAs and 628 differentially expressed lncRNAs. Gene Ontology and pathway analyses associated the differentially expressed mRNAs with metabolic and inflammatory processes. Moreover, lncRNA-mRNA co-expression network revealed 36 network pairs comprising 10 differentially expressed mRNAs and 34 dysregulated lncRNAs. The results of bioinformatics analysis further indicated that dysregulated lncRNA NONHSAT174696, lncRNA NONHSAT179219, and lncRNA NONHSAT161887, as the regulators of EDAR, CYP1B1, and ALDH3A1 respectively, played an important role in the PCB 156-induced lipid metabolism disorder. CONCLUSION: Our findings provide an overview of differentially expressed mRNAs and lncRNAs in L-02 cells exposed to PCB 156, and contribute to the field of polychlorinated biphenyl-induced non-alcoholic fatty liver disease.


Assuntos
Fígado/efeitos dos fármacos , Bifenilos Policlorados/toxicidade , Transcriptoma/efeitos dos fármacos , Aldeído Desidrogenase/genética , Linhagem Celular , Colesterol/metabolismo , Citocromo P-450 CYP1B1/genética , Receptor Edar/genética , Perfilação da Expressão Gênica , Humanos , Fígado/citologia , Fígado/fisiologia , Hepatopatia Gordurosa não Alcoólica/induzido quimicamente , Hepatopatia Gordurosa não Alcoólica/patologia , Análise de Sequência com Séries de Oligonucleotídeos , RNA Longo não Codificante , RNA Mensageiro/metabolismo , Testes de Toxicidade , Triglicerídeos/metabolismo
3.
Rozhl Chir ; 98(10): 388-393, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31842567

RESUMO

Repopulation of decellularized tissue with cells is a very promising approach in tissue engineering, with liver tissue engineering not being an exception. Decellularized liver scaffolds can serve as an excellent 3D environment for recellularization as it maintain tissue-specific microarchitecture of ECM proteins with important spatial cues for cell adhesion, migration, growth and differentiation. Moreover, by using autologous cells the newly constructed graft should lack immunogenicity in the host organism and thus eliminate the need for immunosuppressive therapy in the post-transplant period. This review provides an overview of liver decellularization and repopulation experiments done so far while highlighting the advances as well as pin-pointing the challenges that remain to be solved.


Assuntos
Fígado/fisiologia , Engenharia Tecidual/métodos , Tecidos Suporte , Animais , Fenômenos Fisiológicos Celulares , Matriz Extracelular/fisiologia , Humanos , Fígado/citologia , Suínos
4.
Biochemistry (Mosc) ; 84(9): 1093-1106, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31693469

RESUMO

Differential expression of 30,003 genes was studied in the liver of female Wistar rats fed with isocaloric diets with the excess of fat, fructose, or cholesterol, or their combinations for 62 days using the method of whole-transcriptome profiling on a microchip. Relative mRNA expression levels of the Asah2, Crot, Crtc2, Fmo3, GSTA2, LOC1009122026, LOC102551184, NpY, NqO1, Prom1, Retsat, RGD1305464, Tmem104, and Whsc1 genes were also determined by RT-qPCR. All the tested diets affected differently the key metabolic pathways (KEGGs). Significant changes in the expression of steroid metabolism gene were observed in the liver of animals fed with the tested diets (except the high-fat high fructose diet). Both high-fat and high-fructose diets caused a significant decrease in the expression of squalene synthase (FDFT1 gene) responsible for the initial stage of cholesterol synthesis. On the contrary, in animals fed with the high-cholesterol diet (0.5% cholesterol), expression of the FDFT1 gene did not differ from the control group; however, these animals were characterized by changes in the expression of glucose and glycogen synthesis genes, which could lead to the suppression of glycogen synthesis and gluconeogenesis. At the same time, this group demonstrated different liver tissue morphology in comparison with the animals fed with the high-fructose high-fat diet, manifested as the presence of lipid vacuoles of a smaller size in hepatocytes. The high-fructose and high-fructose high-fat diets affected the metabolic pathways associated with intracellular protein catabolism (endocytosis, phagocytosis, proteasomal degradation, protein processing in the endoplasmic reticulum), tight junctions and intercellular contacts, adhesion molecules, and intracellular RNA transport. Rats fed with the high-fructose high-fat or high-cholesterol diets demonstrated consistent changes in the expression of the Crot, Prom1, and RGD1305464 genes, which reflected a coordinated shift in the regulation of lipid and carbohydrate metabolisms.


Assuntos
Colesterol/farmacologia , Gorduras na Dieta/farmacologia , Açúcares da Dieta/farmacologia , Frutose/farmacologia , Fígado/efeitos dos fármacos , Fígado/metabolismo , RNA/genética , Transcriptoma/efeitos dos fármacos , Animais , Colesterol/administração & dosagem , Colesterol/metabolismo , Biologia Computacional , Gorduras na Dieta/administração & dosagem , Gorduras na Dieta/metabolismo , Açúcares da Dieta/administração & dosagem , Açúcares da Dieta/metabolismo , Feminino , Frutose/administração & dosagem , Frutose/metabolismo , Perfilação da Expressão Gênica , Fígado/citologia , RNA/análise , RNA/isolamento & purificação , Ratos , Ratos Wistar , Transcriptoma/genética
5.
Immunity ; 51(6): 1088-1101.e5, 2019 12 17.
Artigo em Inglês | MEDLINE | ID: mdl-31732168

RESUMO

The B cell response to Ehrlichia muris is dominated by plasmablasts (PBs), with few-if any-germinal centers (GCs), yet it generates protective immunoglobulin M (IgM) memory B cells (MBCs) that express the transcription factor T-bet and harbor V-region mutations. Because Ehrlichia prominently infects the liver, we investigated the nature of liver B cell response and that of the spleen. B cells within infected livers proliferated and underwent somatic hypermutation (SHM). Vh-region sequencing revealed trafficking of clones between the spleen and liver and often subsequent local clonal expansion and intraparenchymal localization of T-bet+ MBCs. T-bet+ MBCs expressed MBC subset markers CD80 and PD-L2. Many T-bet+ MBCs lacked CD11b or CD11c expression but had marginal zone (MZ) B cell phenotypes and colonized the splenic MZ, revealing T-bet+ MBC plasticity. Hence, liver and spleen are generative sites of B cell responses, and they include V-region mutation and result in liver MBC localization.


Assuntos
Linfócitos B/imunologia , Ehrlichia/imunologia , Ehrlichiose/imunologia , Imunoglobulina M/imunologia , Fígado/imunologia , Baço/imunologia , Animais , Antígeno B7-1/biossíntese , Região Variável de Imunoglobulina/genética , Memória Imunológica/imunologia , Fígado/citologia , Camundongos , Camundongos Endogâmicos C57BL , Proteína 2 Ligante de Morte Celular Programada 1/biossíntese , Hipermutação Somática de Imunoglobulina/genética , Baço/citologia , Proteínas com Domínio T/metabolismo
6.
Nature ; 574(7779): 538-542, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31645727

RESUMO

The most common causes of chronic liver disease are excess alcohol intake, viral hepatitis and non-alcoholic fatty liver disease, with the clinical spectrum ranging in severity from hepatic inflammation to cirrhosis, liver failure or hepatocellular carcinoma (HCC). The genome of HCC exhibits diverse mutational signatures, resulting in recurrent mutations across more than 30 cancer genes1-7. Stem cells from normal livers have a low mutational burden and limited diversity of signatures8, which suggests that the complexity of HCC arises during the progression to chronic liver disease and subsequent malignant transformation. Here, by sequencing whole genomes of 482 microdissections of 100-500 hepatocytes from 5 normal and 9 cirrhotic livers, we show that cirrhotic liver has a higher mutational burden than normal liver. Although rare in normal hepatocytes, structural variants, including chromothripsis, were prominent in cirrhosis. Driver mutations, such as point mutations and structural variants, affected 1-5% of clones. Clonal expansions of millimetres in diameter occurred in cirrhosis, with clones sequestered by the bands of fibrosis that surround regenerative nodules. Some mutational signatures were universal and equally active in both non-malignant hepatocytes and HCCs; some were substantially more active in HCCs than chronic liver disease; and others-arising from exogenous exposures-were present in a subset of patients. The activity of exogenous signatures between adjacent cirrhotic nodules varied by up to tenfold within each patient, as a result of clone-specific and microenvironmental forces. Synchronous HCCs exhibited the same mutational signatures as background cirrhotic liver, but with higher burden. Somatic mutations chronicle the exposures, toxicity, regeneration and clonal structure of liver tissue as it progresses from health to disease.


Assuntos
Células Clonais/citologia , Células Clonais/patologia , Fibrose/genética , Fibrose/patologia , Fígado/citologia , Fígado/metabolismo , Mutação , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Células Clonais/metabolismo , Análise Mutacional de DNA , Hepatócitos/citologia , Hepatócitos/metabolismo , Hepatócitos/patologia , Humanos , Fígado/patologia , Masculino , Pessoa de Meia-Idade , Filogenia , Células-Tronco/citologia , Células-Tronco/metabolismo , Células-Tronco/patologia
7.
BMC Complement Altern Med ; 19(1): 283, 2019 Oct 25.
Artigo em Inglês | MEDLINE | ID: mdl-31653214

RESUMO

BACKGROUND: A potato protein hydrolysate, APPH is a potential anti-obesity diet ingredient. Since, obesity leads to deterioration of liver function and associated liver diseases, in this study the effect of APPH on high fat diet (HFD) associated liver damages was investigated. METHODS: Six week old male hamsters were randomly separated to six groups (n = 8) as control, HFD (HFD fed obese), L-APPH (HFD + 15 mg/kg/day of APPH), M-APPH (HFD + 30 mg/kg/day), H-APPH (HFD + 75 mg/kg/day of APPH) and PB (HFD + 500 mg/kg/day of probucol). HFD fed hamsters were administered with APPH 50 days through oral gavage. The animals were euthanized and the number of apoptotic nuclei in liver tissue was determined by TUNEL staining and the extent of interstitial fibrosis was determined by Masson's trichrome staining. Modulation in the molecular events associated with apoptosis and fibrosis were elucidated from the western blotting analysis of the total protein extracts. RESULTS: Hamsters fed with high fat diet showed symptoms of liver damage as measured from serum markers like alanine aminotransferase and aspartate aminotransferase levels. However a 50 day long supplementation of APPH effectively ameliorated the effects of HFD. HFD also modulated the expression of survival and apoptosis proteins in the hamster liver. Further the HFD groups showed elevated levels of fibrosis markers in liver. The increase in fibrosis and apoptosis was correlated with the increase in the levels of phosphorylated extracellular signal-regulated kinases (pERK1/2) revealing a potential role of ERK in the HFD mediated liver damage. However APPH treatment reduced the effect of HFD on the apoptosis and fibrosis markers considerably and provided hepato-protection. CONCLUSION: APPH can therefore be considered as an efficient therapeutic agent to ameliorate high fat diet related liver damages.


Assuntos
Caspase 3/metabolismo , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Obesidade/dietoterapia , Proteínas de Plantas/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Solanum tuberosum/metabolismo , Animais , Apoptose , Caspase 3/genética , Cricetinae , Dieta Hiperlipídica/efeitos adversos , Fibrose/dietoterapia , Fibrose/genética , Fibrose/metabolismo , Fibrose/fisiopatologia , Humanos , Fígado/citologia , Fígado/metabolismo , Fígado/patologia , Masculino , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 9 da Matriz/genética , Mesocricetus , Obesidade/genética , Obesidade/metabolismo , Obesidade/fisiopatologia , Proteínas de Plantas/química , Hidrolisados de Proteína/química , Hidrolisados de Proteína/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Solanum tuberosum/química
8.
Ecotoxicol Environ Saf ; 186: 109749, 2019 Dec 30.
Artigo em Inglês | MEDLINE | ID: mdl-31622878

RESUMO

Hexavalent chromium [Cr(VI)] is a common heavy metal pollutant widely used in various industrial fields. It is well known that mitochondria are the most vulnerable targets of heavy metals, but the key molecule/event that directly mediated mitochondrial dysfunction after Cr(VI) exposure is still unclear. The present study was aimed to explore whether Cr(VI) exposure could affect the mitochondrial fission/fusion process, and whether the related abnormal mitochondrial dynamics have been implicated in Cr(VI)-induced mitochondrial dysfunction. We found that the mitochondrial dysfunction caused by Cr(VI) exposure was characterized by decreased mitochondrial respiratory chain complex (MRCC) I/II activities and levels, collapsed mitochondrial membrane potential (MMP), depleted ATP, and increased reactive oxygen species (ROS) level. Cr(VI) induced abnormal mitochondrial fission/fusion events, the antioxidant Nacetyl-L-cysteine (NAC) restored the abnormal mitochondrial function as well as the fission/fusion dynamics. ROS was the up-stream regulator of extracellular regulated protein kinases (ERK) signaling, and the application of a specific ERK1/2 inhibitor PD98059 confirmed that activation of ERK1/2 signaling was associated with the abnormal mitochondrial fission/fusion and mitochondrial dysfunction. We also demonstrated that treatment with dynamic-like protein 1 (DLP1)-siRNA rescued mitochondrial dysfunction in Cr(VI)-exposed L02 hepatocytes. We reached the conclusion that blockage of ROS-ERK-DLP1 signaling and mitochondrial fission alleviates Cr(VI)-induced mitochondrial dysfunction in L02 hepatocytes, which may provide the new avenue for developing effective strategies to protect against Cr(VI)-induced hepatotoxicity.


Assuntos
Antioxidantes/farmacologia , Cromo/toxicidade , Dinaminas/farmacologia , Fígado/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Acetilcisteína/farmacologia , Carcinógenos Ambientais , Dinaminas/metabolismo , Flavonoides/farmacologia , Hepatócitos/efeitos dos fármacos , Hepatócitos/fisiologia , Humanos , Fígado/citologia , Fígado/metabolismo , Fígado/fisiopatologia , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/fisiologia , Dinâmica Mitocondrial/efeitos dos fármacos , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Transdução de Sinais
9.
Nature ; 574(7776): 112-116, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31554966

RESUMO

Organogenesis is a complex and interconnected process that is orchestrated by multiple boundary tissue interactions1-7. However, it remains unclear how individual, neighbouring components coordinate to establish an integral multi-organ structure. Here we report the continuous patterning and dynamic morphogenesis of hepatic, biliary and pancreatic structures, invaginating from a three-dimensional culture of human pluripotent stem cells. The boundary interactions between anterior and posterior gut spheroids differentiated from human pluripotent stem cells enables retinoic acid-dependent emergence of hepato-biliary-pancreatic organ domains specified at the foregut-midgut boundary organoids in the absence of extrinsic factors. Whereas transplant-derived tissues are dominated by midgut derivatives, long-term-cultured microdissected hepato-biliary-pancreatic organoids develop into segregated multi-organ anlages, which then recapitulate early morphogenetic events including the invagination and branching of three different and interconnected organ structures, reminiscent of tissues derived from mouse explanted foregut-midgut culture. Mis-segregation of multi-organ domains caused by a genetic mutation in HES1 abolishes the biliary specification potential in culture, as seen in vivo8,9. In sum, we demonstrate that the experimental multi-organ integrated model can be established by the juxtapositioning of foregut and midgut tissues, and potentially serves as a tractable, manipulatable and easily accessible model for the study of complex human endoderm organogenesis.


Assuntos
Sistema Biliar/embriologia , Intestinos/embriologia , Fígado/embriologia , Modelos Biológicos , Morfogênese , Pâncreas/embriologia , Animais , Sistema Biliar/citologia , Biomarcadores/análise , Biomarcadores/metabolismo , Padronização Corporal , Endoderma/citologia , Endoderma/embriologia , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Intestinos/citologia , Fígado/citologia , Masculino , Camundongos , Organoides/citologia , Organoides/embriologia , Pâncreas/citologia , Esferoides Celulares/citologia , Esferoides Celulares/metabolismo , Esferoides Celulares/transplante , Fatores de Transcrição HES-1/análise , Fatores de Transcrição HES-1/metabolismo
10.
Aquat Toxicol ; 216: 105291, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31525644

RESUMO

Most pesticides used in agriculture end up in the aquatic environment through runoff and leaching of treated crops. One of the most commonly used herbicides is glyphosate. This compound or its metabolites are frequently detected in surface water in Europe. In the present study, in vivo and in vitro studies were carried out using the early life stages of rainbow trout (Oncorhynchus mykiss) and the cell line RTL-W1 (a liver cell line from rainbow trout) to characterize the toxic effects of glyphosate at environmentally-realistic concentrations. Both studies were performed using the commercial formulation Roundup® GT Max, and technical-grade glyphosate for the in vitro study. Eyed-stage embryos were exposed for 3 weeks to sub-lethal concentrations (0.1 and 1 mg/L) of glyphosate using Roundup. Numerous toxicity endpoints were recorded such as survival, hatching success, larval biometry, developmental abnormalities, swimming activity, genotoxicity (formamidopyrimidine DNA-glycosylase Fpg-modified comet assay), lipid peroxidation (TBARS), protein carbonyls and target gene transcription. Concentrations neither affected embryonic or larval survival nor increased developmental abnormalities. However, a significant decrease was observed in the head size of larvae exposed to 1 mg/L of glyphosate. In addition, a significant increase in mobility was observed for larvae exposed to glyphosate at 0.1 mg/L. TBARS levels were significantly decreased on larvae exposed to 1 mg/L (a.i.), and cat and cox1 genes were differently transcribed from controls. DNA damage was detected by the Fpg-modified comet assay in RTL-W1 cell line exposed to the technical-grade glyphosate and Roundup formulation. The results suggest that chronic exposure to glyphosate, at environmental concentrations, could represent a potential risk for early life stages of fish.


Assuntos
Glicina/análogos & derivados , Herbicidas/toxicidade , Estágios do Ciclo de Vida/efeitos dos fármacos , Fígado/citologia , Oncorhynchus mykiss/crescimento & desenvolvimento , Animais , Células Sanguíneas/metabolismo , Gatos , Morte Celular/efeitos dos fármacos , Linhagem Celular , Ensaio Cometa , Dano ao DNA , Embrião não Mamífero/efeitos dos fármacos , Exposição Ambiental , Regulação da Expressão Gênica/efeitos dos fármacos , Glicina/toxicidade , Larva/efeitos dos fármacos , Peroxidação de Lipídeos/efeitos dos fármacos , Fígado/efeitos dos fármacos , Mutagênicos/toxicidade , Oncorhynchus mykiss/sangue , Oncorhynchus mykiss/embriologia , Carbonilação Proteica/efeitos dos fármacos , Natação , Poluentes Químicos da Água/toxicidade
11.
J Dairy Sci ; 102(11): 10340-10359, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31495618

RESUMO

We have shown in 2 independent studies that cows who received recombinant bovine interleukin-8 (rbIL-8) administered intrauterinely shortly after parturition have a significant and long-lasting increase in milk yield. In the present study, we hypothesized that the increased milk production associated with rbIL-8 treatment is a consequence of increased postpartum dry matter intake (DMI) and orchestrated homeorhetic changes that prioritize milk production. Cows were enrolled into 1 of 3 treatment groups: those assigned to the control group (CTR; n = 70) received an intrauterine (IU) administration of 500 mL of Dulbecco's phosphate-buffered saline (DPBS) solution and 1 mL of DPBS solution intravenously (IV; jugular vein), those assigned to the rbIL-8 IV group (rbIL8-IV, n = 70) received an IV injection of 167 µg of rbIL-8 and 500 mL of DPBS solution IU, and cows assigned to the rbIL-8 IU group (rbIL8-IU, n = 70) received an IU administration with 1,195 µg of rbIL-8 diluted in 499.5 mL of DPBS solution and 1 mL of DPBS solution IV. Animals were housed in a tiestall from calving to 30 d in milk (DIM) to measure DMI. Blood samples were collected daily from calving to 7 DIM and weekly until 28 DIM. Insulin resistance was evaluated using an intravenous glucose tolerance test and intravenous insulin challenge test (IVICT) in a subgroup of cows (n = 20/treatment) at 10 and 11 DIM, respectively. Additionally, liver biopsy samples were taken at 14 DIM from the same subgroup of cows to measure triglyceride levels and cell proliferation and apoptosis. Cows treated with rbIL8-IU produced more milk (CTR = 36.9 ± 1.5; rbIL8-IU = 38.5 ± 1.5; rbIL8-IV = 36.6 ± 1.5 kg/d), energy-corrected milk (CTR = 42.9 ± 0.9; rbIL8-IU = 46.1 ± 0.8; rbIL8-IV = 43.7 ± 0.9 kg/d), and fat-corrected milk (CTR = 44.3 ± 0.9; rbIL8-IU = 47.8 ± 0.9; rbIL8-IV = 45.2 ± 0.9 kg/d) yields when compared with CTR cows, and no differences were observed between rbIL8-IV and CTR cows. The administration of rbIL8-IU significantly increased DMI compared with CTR (CTR = 18.8 ± 0.3; rbIL8-IU = 19.9 ± 0.3; rbIL8-IV = 19.3 ± 0.3 kg/d). Recombinant bIL-8 treatment did not affect glucose, insulin, or fatty acids (i.e., IVICT only) concentrations or their area under the curve in response to an intravenous glucose tolerance test and IVICT when compared with CTR. Moreover, rbIL-8 treatment administered IU or IV increased liver triglyceride levels. Additionally, cows treated with rbIL8-IU tended to have lower odds of developing hyperketonemia (odds ratio = 0.46, 95% confidence interval: 0.19 to 1.10), lower odds of clinical ketosis and displaced abomasum combined (odds ratio = 0.17, 95% confidence interval: 0.03 to 0.89), and lower odds of diseases combined (odds ratio = 0.43, 95% confidence interval: 0.21 to 0.86) when compared with CTR. We conclude that the administration of rbIL8-IU increases DMI, milk production, fat-corrected milk, and energy-corrected milk while improving overall health during the postpartum period. This study supports the use of rbIL-8 administered IU shortly after calving to improve health and production responses in lactating cows.


Assuntos
Bovinos/fisiologia , Ingestão de Alimentos/efeitos dos fármacos , Resistência à Insulina , Interleucina-8/administração & dosagem , Lactação/efeitos dos fármacos , Ácido 3-Hidroxibutírico/sangue , Animais , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Dieta/veterinária , Ácidos Graxos/metabolismo , Feminino , Glucose/metabolismo , Hormônio do Crescimento/efeitos dos fármacos , Hormônio do Crescimento/metabolismo , Insulina/sangue , Fator de Crescimento Insulin-Like I/efeitos dos fármacos , Fator de Crescimento Insulin-Like I/metabolismo , Interleucina-8/metabolismo , Interleucina-8/farmacologia , Cetose/veterinária , Lactação/fisiologia , Fígado/citologia , Fígado/efeitos dos fármacos , Leite/metabolismo , Parto , Período Pós-Parto/sangue , Gravidez , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/farmacologia
12.
BMC Complement Altern Med ; 19(1): 237, 2019 Sep 03.
Artigo em Inglês | MEDLINE | ID: mdl-31481128

RESUMO

BACKGROUND: Rice husk, a waste material produced during milling, contains numerous phytochemicals that may be sources of cancer chemopreventive agents. Various biological activities of white and colored rice husk have been reported. However, there are few comparative studies of the cancer chemopreventive effects of white and colored rice husk. METHODS: This study investigated the cancer chemopreventive activities of two different colors of rice husk using in vitro and in vivo models. A bacterial mutation assay using Salmonella typhimurium strains TA98 and TA100 was performed; enzyme induction activity in murine hepatoma cells was measured, and a liver micronucleus test was performed in male Wistar rats. RESULTS: The white rice husk (WRHE) and purple rice husk (PRHE) extracts were not mutagenic in Salmonella typhimurium TA98 or TA100 in the presence or absence of metabolic activation. However, the extracts exhibited antimutagenicity against aflatoxin B1 (AFB1) and 2-amino-3,4 dimethylimidazo[4,5-f]quinolone (MeIQ) in a Salmonella mutation assay. The extracts also induced anticarcinogenic enzyme activity in a murine Hepa1c1c7 hepatoma cell line. Interestingly, PRHE but not WRHE exhibited antigenotoxicity in the rat liver micronucleus test. PRHE significantly decreased the number of micronucleated hepatocytes in AFB1-initiated rats. PRHE contained higher amounts of phenolic compounds and vitamin E than WRHE in both tocopherols and tocotrienols as well as polyphenol such as cyanidin-3-glucoside, protocatechuic acid and vanillic acid. Furthermore, PRHE increased CYP1A1 and 1A2 activities while decreasing CYP3A2 activity in the livers of AFB1-treated rats. PRHE also enhanced various detoxifying enzyme activities, including glutathione S-transferase, NAD(P)H quinone oxidoreductase and heme oxygenase. CONCLUSIONS: PRHE showed potent cancer chemopreventive activity in a rat liver micronucleus assay through modulation of phase I and II xenobiotic metabolizing enzymes involved in AFB1 metabolism. Vitamin E and phenolic compounds may be candidate antimutagens in purple rice husk.


Assuntos
Aflatoxina B1/toxicidade , Inativação Metabólica/efeitos dos fármacos , Fígado/efeitos dos fármacos , Micronúcleos com Defeito Cromossômico/efeitos dos fármacos , Oryza/química , Animais , Antimutagênicos/farmacologia , Linhagem Celular , Fígado/citologia , Fígado/enzimologia , Fígado/metabolismo , Masculino , Testes para Micronúcleos , Extratos Vegetais/farmacologia , Ratos , Ratos Wistar , Salmonella typhimurium/efeitos dos fármacos
13.
Nat Commun ; 10(1): 3929, 2019 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-31477734

RESUMO

AT-1/SLC33A1 is a key member of the endoplasmic reticulum (ER) acetylation machinery, transporting acetyl-CoA from the cytosol into the ER lumen where acetyl-CoA serves as the acetyl-group donor for Nε-lysine acetylation. Dysfunctional ER acetylation, as caused by heterozygous or homozygous mutations as well as gene duplication events of AT-1/SLC33A1, has been linked to both developmental and degenerative diseases. Here, we investigate two models of AT-1 dysregulation and altered acetyl-CoA flux: AT-1S113R/+ mice, a model of AT-1 haploinsufficiency, and AT-1 sTg mice, a model of AT-1 overexpression. The animals display distinct metabolic adaptation across intracellular compartments, including reprogramming of lipid metabolism and mitochondria bioenergetics. Mechanistically, the perturbations to AT-1-dependent acetyl-CoA flux result in global and specific changes in both the proteome and the acetyl-proteome (protein acetylation). Collectively, our results suggest that AT-1 acts as an important metabolic regulator that maintains acetyl-CoA homeostasis by promoting functional crosstalk between different intracellular organelles.


Assuntos
Acetilcoenzima A/metabolismo , Citosol/metabolismo , Metabolismo dos Lipídeos , Proteínas de Membrana Transportadoras/metabolismo , Proteoma/metabolismo , Proteômica/métodos , Acetilação , Animais , Retículo Endoplasmático/metabolismo , Haploinsuficiência , Fígado/citologia , Fígado/metabolismo , Lisina/metabolismo , Proteínas de Membrana Transportadoras/genética , Camundongos Knockout , Camundongos Transgênicos
14.
Nat Commun ; 10(1): 3051, 2019 07 11.
Artigo em Inglês | MEDLINE | ID: mdl-31296864

RESUMO

Treatment of liver metastasis experiences slow progress owing to the severe side effects. In this study, we demonstrate a strategy capable of eliminating metastatic cancer cells in a selective manner. Nucleus-targeting W18O49 nanoparticles (WONPs) are conjugated to mitochondria-selective mesoporous silica nanoparticles (MSNs) containing photosensitizer (Ce6) through a Cathepsin B-cleavable peptide. In hepatocytes, upon the laser irradiation, the generated singlet oxygen species are consumed by WONPs, in turn leading to the loss of their photothermally heating capacity, thereby sparing hepatocyte from thermal damage induced by the laser illumination. By contrast, in cancer cells, the cleaved peptide linker allows WONPs and MSNs to respectively target nucleus and mitochondria, where the therapeutic powers could be unleashed, both photodynamically and photothermally. This ensures the energy production of cancer cells can be abolished. We further assess the underlying molecular mechanism at both gene and protein levels to better understand the therapeutic outcome.


Assuntos
Portadores de Fármacos/metabolismo , Neoplasias Hepáticas/terapia , Fotoquimioterapia/métodos , Fármacos Fotossensibilizantes/administração & dosagem , Animais , Catepsina B/metabolismo , Núcleo Celular/metabolismo , Portadores de Fármacos/química , Feminino , Células HCT116 , Hepatócitos/citologia , Hepatócitos/metabolismo , Humanos , Injeções Intravenosas , Lasers , Fígado/citologia , Fígado/metabolismo , Fígado/patologia , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/secundário , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Mitocôndrias/metabolismo , Nanopartículas/química , Nanopartículas/metabolismo , Fosforilação Oxidativa , Óxidos/química , Fotoquimioterapia/instrumentação , Espécies Reativas de Oxigênio , Dióxido de Silício/química , Resultado do Tratamento , Tungstênio/química , Ensaios Antitumorais Modelo de Xenoenxerto
15.
Nat Commun ; 10(1): 3350, 2019 07 26.
Artigo em Inglês | MEDLINE | ID: mdl-31350390

RESUMO

The liver parenchyma is composed of hepatocytes and bile duct epithelial cells (BECs). Controversy exists regarding the cellular origin of human liver parenchymal tissue generation during embryonic development, homeostasis or repair. Here we report the existence of a hepatobiliary hybrid progenitor (HHyP) population in human foetal liver using single-cell RNA sequencing. HHyPs are anatomically restricted to the ductal plate of foetal liver and maintain a transcriptional profile distinct from foetal hepatocytes, mature hepatocytes and mature BECs. In addition, molecular heterogeneity within the EpCAM+ population of freshly isolated foetal and adult human liver identifies diverse gene expression signatures of hepatic and biliary lineage potential. Finally, we FACS isolate foetal HHyPs and confirm their hybrid progenitor phenotype in vivo. Our study suggests that hepatobiliary progenitor cells previously identified in mice also exist in humans, and can be distinguished from other parenchymal populations, including mature BECs, by distinct gene expression profiles.


Assuntos
Fígado/citologia , Transcrição Genética , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/metabolismo , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismo , Molécula de Adesão da Célula Epitelial/genética , Molécula de Adesão da Célula Epitelial/metabolismo , Feto/metabolismo , Hepatócitos/citologia , Hepatócitos/metabolismo , Humanos , Fígado/metabolismo , Análise de Célula Única , Células-Tronco/citologia , Células-Tronco/metabolismo
16.
Gut ; 68(12): 2228-2237, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31300517

RESUMO

Organoid cultures have emerged as an alternative in vitro system to recapitulate tissues in a dish. While mouse models and cell lines have furthered our understanding of liver biology and associated diseases, they suffer in replicating key aspects of human liver tissue, in particular its complex architecture and metabolic functions. Liver organoids have now been established for multiple species from induced pluripotent stem cells, embryonic stem cells, hepatoblasts and adult tissue-derived cells. These represent a promising addition to our toolbox to gain a deeper understanding of this complex organ. In this perspective we will review the advances in the liver organoid field, its limitations and potential for biomedical applications.


Assuntos
Pesquisa Biomédica/métodos , Fígado/citologia , Modelos Biológicos , Organoides/fisiologia , Medicina Regenerativa/métodos , Animais , Técnicas de Cultura de Células , Linhagem Celular , Humanos , Organoides/citologia
17.
Cancer Sci ; 110(9): 2748-2759, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31301081

RESUMO

In hepatocarcinogenesis induced by diethylnitrosamine (DEN) in B6C3F1 mice, the BrafV637E mutation, corresponding to the human BRAFV600E mutation, plays a pivotal role. The livers of transgenic mice with a hepatocyte-specific human BRAFV600E mutation weighed 4.5 times more than that of normal mice and consisted entirely of hepatocytes, resembling DEN-induced preneoplastic hepatocytes. However, these transgenic mice spontaneously died 7 wk after birth, therefore this study aimed to clarify the causes of death. In the transgenic mice, the liver showed thrombopoietin (TPO) overexpression, which is associated with eventual megakaryocytosis and thrombocytosis, and activated platelets were deposited in hepatic sinusoids. TPO was also overexpressed in the DEN-induced hepatic tumors, and sinusoidal platelet deposition was observed in the hepatic tumors of humans and mice. Podoplanin was expressed in some of the Kupffer cells in the liver of the transgenic mice, indicating that platelet activation occurred via the interaction of podoplanin with C-type lectin receptor 2 (CLEC-2) on the platelet membrane. Additionally, erythrocyte dyscrasia and glomerulonephropathy/interstitial pneumonia associated with platelet deposition were observed. In the transgenic mice, aspirin (Asp) administration prevented platelet activation, reduced the liver/body weight ratio, decreased the platelet deposition in the liver, kidney, and lung, and prevented erythrocyte dyscrasia and ameliorated the renal/pulmonary changes. Thrombopoietin overproduction by BRAFV600E-mutated hepatocytes may contribute to hepatocyte proliferation via thrombocytosis, platelet activation, and the interaction of platelets with hepatic sinusoidal cells, while hematologic, renal, and pulmonary disorders due to aberrant platelet activation may lead to spontaneous death in the transgenic mice.


Assuntos
Carcinogênese/genética , Neoplasias Hepáticas Experimentais/patologia , Fígado/patologia , Proteínas Proto-Oncogênicas B-raf/genética , Trombopoetina/metabolismo , Animais , Biópsia , Plaquetas/patologia , Medula Óssea/patologia , Capilares/patologia , Carcinógenos/administração & dosagem , Carcinógenos/toxicidade , Proliferação de Células/genética , Dietilnitrosamina/administração & dosagem , Dietilnitrosamina/toxicidade , Feminino , Regulação Neoplásica da Expressão Gênica , Hepatectomia , Hepatócitos/patologia , Humanos , Fígado/irrigação sanguínea , Fígado/citologia , Neoplasias Hepáticas Experimentais/induzido quimicamente , Neoplasias Hepáticas Experimentais/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos , Camundongos Transgênicos , Ativação Plaquetária/genética , Cultura Primária de Células , Proteínas Proto-Oncogênicas B-raf/metabolismo , Células Tumorais Cultivadas
18.
Nature ; 572(7768): 199-204, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31292543

RESUMO

The human liver is an essential multifunctional organ. The incidence of liver diseases is rising and there are limited treatment options. However, the cellular composition of the liver remains poorly understood. Here we performed single-cell RNA sequencing of about 10,000 cells from normal liver tissue from nine human donors to construct a human liver cell atlas. Our analysis identified previously unknown subtypes of endothelial cells, Kupffer cells, and hepatocytes, with transcriptome-wide zonation of some of these populations. We show that the EPCAM+ population is heterogeneous, comprising hepatocyte-biased and cholangiocyte populations as well as a TROP2int progenitor population with strong potential to form bipotent liver organoids. As a proof-of-principle, we used our atlas to unravel the phenotypic changes that occur in hepatocellular carcinoma cells and in human hepatocytes and liver endothelial cells engrafted into a mouse liver. Our human liver cell atlas provides a powerful resource to enable the discovery of previously unknown cell types in normal and diseased livers.


Assuntos
Células Epiteliais/citologia , Hepatócitos/citologia , Fígado/citologia , Células-Tronco/citologia , Adulto , Animais , Antígenos de Neoplasias/metabolismo , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/imunologia , Carcinoma Hepatocelular/patologia , Moléculas de Adesão Celular/metabolismo , Quimera/imunologia , Quimera/metabolismo , Células Endoteliais/citologia , Células Endoteliais/imunologia , Células Epiteliais/imunologia , Feminino , Regulação da Expressão Gênica , Hepatócitos/imunologia , Hepatócitos/metabolismo , Humanos , Fígado/imunologia , Masculino , Camundongos , Organoides/metabolismo , RNA Citoplasmático Pequeno/genética , Reprodutibilidade dos Testes , Células-Tronco/imunologia
19.
World J Gastroenterol ; 25(24): 3044-3055, 2019 Jun 28.
Artigo em Inglês | MEDLINE | ID: mdl-31293340

RESUMO

BACKGROUND: The formation of liver fibrosis is mainly caused by the activation of hepatic stellate cells (HSCs) and the imbalance of extracellular matrix (ECM) production and degradation. The treatment of liver fibrosis mainly includes removing the cause, inhibiting the activation of HSCs, and inhibiting inflammation. NOD-like receptor (NLR) family, caspase activation and recruitment domain (CARD) domain containing 5/NOD27/CLR16.1 (NLRC5) is a highly conserved member of the NLR family and is involved in inflammation and immune responses by regulating various signaling pathways such as nuclear factor-κB (NF-κB) signaling. It has been found that NLRC5 plays an important role in liver fibrosis, but its specific effect and possible mechanism remain to be fully elucidated. AIM: To investigate the role of NLRC5 in the activation and reversion of HSCs induced with transforming growth factor-ß (TGF-ß) and MDI, and to explore its relationship with liver fibrosis. METHODS: A total of 24 male C57BL/6 mice were randomly divided into three groups, including normal, fibrosis, and recovery groups. Twenty-four hours after a liver fibrosis and spontaneous reversion model was established, the mice were sacrificed and pathological examination of liver tissue was performed to observe the degree of liver fibrosis in each group. LX-2 cells were cultured in vitro and treated with TGF-ß1 and MDI. Real-time quantitative PCR (qPCR) and Western blot were used to analyze the expression levels of NLRC5, α-smooth muscle actin (α-SMA), and collagen type I alpha1 (Col1a1) in each group. The activity of NF-κB in each group of cells transfected with NLRC5-siRNA was detected. RESULTS: Compared with the normal mice, the expression level of NLRC5 increased significantly (P < 0.01) in the fibrosis group, but decreased significantly in the recovery group (P < 0.01). In in vitro experiments, the content of NLRC5 was enhanced after TGF-ß1 stimulation and decreased to a lower level when treated with MDI (P < 0.01). The expression of α-SMA and Col1a1 proteins and mRNAs in TGF-ß1-mediated cells was suppressed by transfection with NLRC5-siRNA (P < 0.01). Western blot analysis showed that the expression of NF-κB p65 protein and phosphorylated IκBα (p-IκBα) was increased in the liver of mice in the fibrosis group but decreased in the recovery group (P < 0.01), and the protein level of nuclear p65 and p-IκBα was significantly increased after treatment with NLRC5-siRNA (P < 0.01). CONCLUSION: NLRC5 may play a key role in the development and reversal of hepatic fibrosis through the NF-κB signaling pathway, and it is expected to be one of the clinical therapeutic targets.


Assuntos
Células Estreladas do Fígado/patologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Cirrose Hepática Experimental/patologia , NF-kappa B/metabolismo , Animais , Tetracloreto de Carbono/toxicidade , Linhagem Celular , Matriz Extracelular/patologia , Técnicas de Silenciamento de Genes , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Fígado/citologia , Fígado/patologia , Cirrose Hepática Experimental/induzido quimicamente , Masculino , Camundongos , RNA Interferente Pequeno/metabolismo , Proteínas Recombinantes/metabolismo , Transdução de Sinais , Fator de Crescimento Transformador beta1/metabolismo
20.
Environ Mol Mutagen ; 60(8): 704-739, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31294869

RESUMO

Regulatory guidance documents stress the value of assessing the most appropriate endpoints in multiple tissues when evaluating the in vivo genotoxic potential of chemicals. However, conducting several independent studies to evaluate multiple endpoints and/or tissue compartments is resource intensive. Furthermore, when dependent on visual detection, conventional approaches for scoring genotoxicity endpoints can be slow, tedious, and less objective than the ideal. To address these issues with current practices we attempted to (1) devise resource sparing treatment and harvest schedules that are compatible with liver and blood micronucleus endpoints, as well as the Pig-a gene mutation assay, and (2) utilize flow cytometry-based methods to score each of these genotoxicity biomarkers. Proof-of-principle experiments were performed with 4-week-old male and female Crl:CD(SD) rats exposed to aristolochic acids I/II, benzo[a]pyrene, cisplatin, cyclophosphamide, diethylnitrosamine, 1,2-dimethylhydrazine, dimethylnitrosamine, 2,6-dinitrotoluene, hydroxyurea, melphalan, temozolomide, quinoline, or vinblastine. These 13 chemicals were each tested in two treatment regimens: one 3-day exposure cycle, and three 3-day exposure cycles. Each exposure, blood collection, and liver harvest was accomplished during a standard Monday-Friday workweek. Key findings are that even these well-studied, relatively potent genotoxicants were not active in both tissues and all assays (indeed only cisplatin was clearly positive in all three assays); and whereas the sensitivity of the Pig-a assay clearly benefitted from three versus one treatment cycle, micronucleus assays yielded qualitatively similar results across both study designs. Collectively, these results suggest it is possible to significantly reduce animal and other resource requirements while improving assessments of in vivo genotoxicity potential by simultaneously evaluating three endpoints and two important tissue compartments using fit-for-purpose study designs in conjunction with flow cytometric scoring approaches. Environ. Mol. Mutagen., 60:704-739, 2019. © 2019 Wiley Periodicals, Inc.


Assuntos
Dano ao DNA/efeitos dos fármacos , Fígado/citologia , Proteínas de Membrana/genética , Micronúcleos com Defeito Cromossômico/induzido quimicamente , Testes para Micronúcleos/métodos , Animais , Dano ao DNA/genética , Feminino , Masculino , Mutagênicos/toxicidade , Ratos , Projetos de Pesquisa
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