RESUMO
Wnt signaling dysfunction and gut dysbiosis may lead to liver fibrosis, yet the underlying mechanisms are not well elucidated. This study demonstrated the role of RSPO4, a Wnt signaling agonist, in liver fibrogenesis and its impact on the gut microbiome. RSPO4 gene in CCl4-induced fibrotic-liver rats was knockout by Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) system, with healthy rats served as the control. Tissue samples and hepatic stellate cells (HSCs) isolated from rats were examined for curative effect of RSPO4-CRISPR treatment. Fecal sample were collected and analyzed with 16 S rRNA sequencing. We found RSPO4-CRISPR relieved liver fibrosis in rats and reversed HSC activation. Further, results showed RSPO4-CRISPR tended to restore the microflora composition. Significance species between groups were identified. Bacteroides and Escherichia-Shigella were the key microbes in the model and negative group, whereas Lactobacillus, Romboutsia, and Lachnospiraceae NK4A136 group were abundant in the control. Notably, Bacteroidales S24-7 group and Ruminococcaceae UCG-005 were the significantly enriched in CRISPR group. We show that the microbiome of rats treated with RSPO4-CRISPR presents a trend towards the restoration of the original condition. Our findings pave a new way to evaluate the curative effect of liver fibrosis treatment.
Assuntos
Sistemas CRISPR-Cas , Doença Hepática Induzida por Substâncias e Drogas/terapia , Microbioma Gastrointestinal , Terapia Genética , Intestinos/microbiologia , Cirrose Hepática Experimental/terapia , Fígado/metabolismo , Trombospondinas/metabolismo , Animais , Tetracloreto de Carbono , Células Cultivadas , Doença Hepática Induzida por Substâncias e Drogas/genética , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Doença Hepática Induzida por Substâncias e Drogas/microbiologia , Disbiose , Células Estreladas do Fígado/metabolismo , Células Estreladas do Fígado/patologia , Fígado/patologia , Cirrose Hepática Experimental/genética , Cirrose Hepática Experimental/metabolismo , Cirrose Hepática Experimental/microbiologia , Masculino , Ratos Sprague-Dawley , Trombospondinas/genética , Via de Sinalização WntRESUMO
BACKGROUND & AIMS: Pathological bacterial translocation (PBT) in cirrhosis is the hallmark of spontaneous bacterial infections, increasing mortality several-fold. Increased intestinal permeability is known to contribute to PBT in cirrhosis, although the role of the mucus layer has not been addressed in detail. A clear route of translocation for luminal intestinal bacteria is yet to be defined, but we hypothesize that the recently described gut-vascular barrier (GVB) is impaired in experimental portal hypertension, leading to increased accessibility of the vascular compartment for translocating bacteria. MATERIALS: Cirrhosis was induced in mouse models using bile-duct ligation (BDL) and CCl4. Pre-hepatic portal-hypertension was induced by partial portal vein ligation (PPVL). Intestinal permeability was compared in these mice after GFP-Escherichia coli or different sized FITC-dextrans were injected into the intestine. RESULTS: Healthy and pre-hepatic portal-hypertensive (PPVL) mice lack translocation of FITC-dextran and GFP-E. coli from the small intestine to the liver, whereas BDL and CCl4-induced cirrhotic mice demonstrate pathological translocation, which is not altered by prior thoracic-duct ligation. The mucus layer is reduced in thickness, with loss of goblet cells and Muc2-staining and expression in cirrhotic but not PPVL mice. These changes are associated with bacterial overgrowth in the inner mucus layer and pathological translocation of GFP-E. coli through the ileal epithelium. GVB is profoundly altered in BDL and CCl4-mice with Ileal extravasation of large-sized 150â¯kDa-FITC-dextran, but only slightly altered in PPVL mice. This pathological endothelial permeability and accessibility in cirrhotic mice is associated with augmented expression of PV1 in intestinal vessels. OCA but not fexaramine stabilizes the GVB, whereas both FXR-agonists ameliorate gut to liver translocation of GFP-E. coli. CONCLUSIONS: Cirrhosis, but not portal hypertension per se, grossly impairs the endothelial and muco-epithelial barriers, promoting PBT to the portal-venous circulation. Both barriers appear to be FXR-modulated, with FXR-agonists reducing PBT via the portal-venous route. LAY SUMMARY: For intestinal bacteria to enter the systemic circulation, they must cross the mucus and epithelial layer, as well as the gut-vascular barrier. Cirrhosis disrupts all 3 of these barriers, giving bacteria access to the portal-venous circulation and thus, the gut-liver axis. Diminished luminal bile acid availability, cirrhosis and the associated reduction in farnesoid x receptor (FXR) signaling seem, at least partly, to mediate these changes, as FXR-agonists reduce bacterial translocation via the portal-venous route to the liver in cirrhosis.
Assuntos
Translocação Bacteriana/efeitos dos fármacos , Permeabilidade Capilar , Dextranos/farmacocinética , Escherichia coli , Microbioma Gastrointestinal/fisiologia , Mucosa Intestinal , Cirrose Hepática Experimental , Receptores Citoplasmáticos e Nucleares , Animais , Ácidos e Sais Biliares/metabolismo , Permeabilidade Capilar/efeitos dos fármacos , Permeabilidade Capilar/fisiologia , Modelos Animais de Doenças , Escherichia coli/isolamento & purificação , Escherichia coli/fisiologia , Hipertensão Portal/metabolismo , Hipertensão Portal/fisiopatologia , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/microbiologia , Cirrose Hepática Experimental/metabolismo , Cirrose Hepática Experimental/microbiologia , Cirrose Hepática Experimental/fisiopatologia , Camundongos , Receptores Citoplasmáticos e Nucleares/agonistas , Receptores Citoplasmáticos e Nucleares/metabolismoRESUMO
BACKGROUND & AIMS: Norfloxacin administration is useful in preventing bacterial infections in cirrhosis but associated to the generation of resistant species. Rifaximin is known to reach high concentrations in the intestinal lumen without generating relevant resistance in the intestinal flora. Our aim was to compare the effect of Norfloxacin and Rifaximin on intestinal flora composition, bacterial translocation and survival in cirrhotic rats. METHODS: Cirrhosis was induced in rats by oral administration of CCl4 . Animals were divided into three groups: only CCl4 (group I, n = 10); CCl4 + Norfloxacin (group II, n = 17) and CCl4 + Rifaximin (group III, n = 14). Gut bacterial composition, bacterial translocation and cytokine levels were measured. RESULTS: Forty-one rats were finally included. The incidence of viable and non-viable bacterial translocation was significantly reduced in animals receiving Norfloxacin; Rifaximin also decreased the incidence of viable and non-viable bacterial translocation, but did not reach statistical significance. Serum TNF-α levels were significantly lower in antibiotic groups. Norfloxacin modified intestinal microbiota, depleting significantly more pathobionts than Rifaximin. CONCLUSION: Norfloxacin is more effective than Rifaximin in preventing bacterial translocation in rats with cirrhosis probably because of its capacity to reduce pathobionts from intestinal microbiota.
Assuntos
Antibacterianos/farmacologia , Infecções Bacterianas/prevenção & controle , Translocação Bacteriana/efeitos dos fármacos , Doença Hepática Induzida por Substâncias e Drogas/tratamento farmacológico , Cirrose Hepática Experimental/tratamento farmacológico , Norfloxacino/farmacologia , Rifaximina/farmacologia , Animais , Infecções Bacterianas/sangue , Infecções Bacterianas/microbiologia , Tetracloreto de Carbono , Doença Hepática Induzida por Substâncias e Drogas/sangue , Doença Hepática Induzida por Substâncias e Drogas/microbiologia , Citocinas/sangue , Microbioma Gastrointestinal/efeitos dos fármacos , Mediadores da Inflamação/sangue , Cirrose Hepática Experimental/sangue , Cirrose Hepática Experimental/induzido quimicamente , Cirrose Hepática Experimental/microbiologia , Masculino , Viabilidade Microbiana/efeitos dos fármacos , Ratos Sprague-DawleyRESUMO
Bacterial translocation and lipopolysaccharide (LPS) leakage occur at a very early stage of liver fibrosis in animal models. We studied the role of LPS in hepatic stellate cell (HSC) activation and the underlying mechanisms in vitro and in vivo. Herein, we demonstrated that LPS treatment led to a dramatic increase in autophagosome formation and autophagic flux in LX-2 cells and HSCs, which was mediated through the AKT-MTOR and AMPK-ULK1 pathway. LPS significantly decreased the lipid content, including the lipid droplet (LD) number and lipid staining area in HSCs; pretreatment with macroautophagy/autophagy inhibitors or silencing ATG5 attenuated this decrease. Furthermore, lipophagy was induced by LPS through the autophagy-lysosomal pathway in LX-2 cells and HSCs. Additionally, LPS-induced autophagy further reduced retinoic acid (RA) signaling, as demonstrated by a decrease in the intracellular RA level and Rar target genes, resulting in the downregulation of Bambi and promoting the sensitization of the HSC's fibrosis response to TGFB. Compared with CCl4 injection alone, CCl4 plus LPS injection exaggerated liver fibrosis in mice, as demonstrated by increased Col1a1 (collagen, type I, α 1), Acta2, Tgfb and Timp1 mRNA expression, ACTA2/α-SMA and COL1A1 protein expression, and Sirius Red staining area, which could be attenuated by injection of an autophagy inhibitor. LPS also reduced lipid content in HSCs in vivo, with this change being attenuated by chloroquine (CQ) administration. In conclusion, LPS-induced autophagy resulted in LD loss, RA signaling dysfunction, and downregulation of the TGFB pseudoreceptor Bambi, thus sensitizing HSCs to TGFB signaling.
Assuntos
Autofagia , Células Estreladas do Fígado/metabolismo , Lipopolissacarídeos/metabolismo , Cirrose Hepática Experimental/microbiologia , Tretinoína/metabolismo , Actinas/genética , Actinas/metabolismo , Animais , Translocação Bacteriana , Tetracloreto de Carbono/toxicidade , Células Cultivadas , Colágeno Tipo I/genética , Colágeno Tipo I/metabolismo , Cadeia alfa 1 do Colágeno Tipo I , Regulação para Baixo , Bactérias Gram-Negativas/fisiologia , Gotículas Lipídicas/metabolismo , Lipopolissacarídeos/farmacologia , Cirrose Hepática Experimental/induzido quimicamente , Cirrose Hepática Experimental/metabolismo , Cirrose Hepática Experimental/patologia , Masculino , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Receptores do Ácido Retinoico/genética , Receptores do Ácido Retinoico/metabolismo , Transdução de Sinais , Inibidor Tecidual de Metaloproteinase-1/genética , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/farmacologiaRESUMO
BACKGROUND: Impaired intestinal motility may lead to the disruption of gut microbiota equilibrium, which in turn facilitates bacterial translocation (BT) and endotoxemia in cirrhosis. We evaluated the influence of mosapride, a prokinetic agent, on BT and DNA fingerprints of gut microbiota in cirrhotic rats. METHODS: A rat model of cirrhosis was set up via subcutaneous injection of carbon tetrachloride (CCl4). The portal pressure, liver and intestinal damage, plasma endotoxin, BT, and intestinal transit rate (ITR) of cirrhotic rats were determined. Fecal DNA fingerprints were obtained by ERIC-PCR. The expressions of tight junction proteins were evaluated by western blotting. RESULTS: Mosapride treatment to cirrhotic rats significantly reduced the plasma endotoxin level and incidence of BT, accompanied by increased ITR. Cirrhotic rats (including those treated with mosapride) suffered from BT exhibited significantly lower ITR than those who are free of BT. Pearson coefficient indicated a significant and negative correlation between the plasma endotoxin level and ITR. The genomic fingerprints of intestinal microbiota from the three groups fell into three distinctive clusters. In the mosapride-treated group, Shannon's index was remarkably increased compared to the model group. Significantly positive correlation was detected between Shannon's index and ITR. Mosapride did not improve hepatic and intestinal damages and ileal expressions of occludin and ZO-1. CONCLUSIONS: Mosapride significantly increases intestinal motility in cirrhotic rats, thus to recover the disordered intestinal microbiota, finally resulting in decreased plasma endotoxin and BT.
Assuntos
Translocação Bacteriana/efeitos dos fármacos , Benzamidas/farmacologia , Tetracloreto de Carbono , Doença Hepática Induzida por Substâncias e Drogas/prevenção & controle , Endotoxemia/prevenção & controle , Fármacos Gastrointestinais/farmacologia , Microbioma Gastrointestinal/efeitos dos fármacos , Intestinos/efeitos dos fármacos , Cirrose Hepática Experimental/prevenção & controle , Fígado/efeitos dos fármacos , Morfolinas/farmacologia , Animais , Doença Hepática Induzida por Substâncias e Drogas/sangue , Doença Hepática Induzida por Substâncias e Drogas/microbiologia , Doença Hepática Induzida por Substâncias e Drogas/patologia , DNA Bacteriano/genética , Endotoxemia/sangue , Endotoxemia/induzido quimicamente , Endotoxemia/microbiologia , Endotoxemia/patologia , Fezes/microbiologia , Motilidade Gastrointestinal/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Intestinos/microbiologia , Intestinos/patologia , Fígado/metabolismo , Fígado/microbiologia , Fígado/patologia , Cirrose Hepática Experimental/sangue , Cirrose Hepática Experimental/microbiologia , Cirrose Hepática Experimental/patologia , Masculino , Ratos Sprague-Dawley , Proteínas de Junções Íntimas/metabolismoRESUMO
Patients with liver cirrhosis present an increased incidence of infections. The main cause has been founded in alterations of the enteric flora and of the intestinal barrier probably due to portal hypertension, in addition to a reticulo-endothelial system dysfunction. Furthermore, those living with cirrhosis can report a high predisposition to sepsis and septic shock, due to the excessive response of pro-inflammatory cytokines and a complessive hemodynamic derangement. By the analysis in the experimental model of the cirrhotic rat, it was demonstrated that radio-labelled Escherichia coli given by the oral route resulted in the location of the bacteria in the gut, the ascitic fluid and mesenteric lymph nodes, a phenomenon known as bacterial translocation. Bacteria encountered with the highest frequency are those colonizing the intestinal tract, such as E. coli, Klebsiella pneumoniae and Enterobacteriaceae, intracellular bacteria and parasites are reported with a lower frequency. Multi-drug resistant bacteria are cultured with the highest frequency in those with frequent hospitalisations and report both high septic shock and mortality rates. Spontaneous bacterial peritonitis (SBP) is the commonest infection in cirrhotic, estimated to occur in 10-30% of the cases with ascites. A practical approach may include administration of a protected penicillin, III generation cephalosporin or quinolones in uncomplicated cases. Instead, in complicated cases and in nosocomial SBP, administration of cephalosporin or quinolones can be burned by the high resistance rate and drugs active against ESBL-producing bacteria and multi-drug resistant Gram positive bacteria have to be considered as empiric therapy, until cultures are available. When cultures are not readily available and patients fail to improve a repeated diagnostic paracentesis should be performed. Current investigations suggest that norfloxacin 400 mg/day orally has been reported to successfully prevent SBP in patients with low-protein ascites and patients with prior SBP.
Assuntos
Infecções Bacterianas/etiologia , Cirrose Hepática/complicações , Animais , Antibacterianos/uso terapêutico , Ascite/microbiologia , Infecções Bacterianas/tratamento farmacológico , Infecções Bacterianas/epidemiologia , Translocação Bacteriana , Comorbidade , Citocinas/metabolismo , Suscetibilidade a Doenças , Farmacorresistência Bacteriana Múltipla , Infecções por Enterobacteriaceae/tratamento farmacológico , Infecções por Enterobacteriaceae/epidemiologia , Infecções por Enterobacteriaceae/etiologia , Microbioma Gastrointestinal , Humanos , Hipertensão Portal/etiologia , Hipertensão Portal/microbiologia , Incidência , Enteropatias Parasitárias/epidemiologia , Enteropatias Parasitárias/etiologia , Cirrose Hepática/epidemiologia , Cirrose Hepática/imunologia , Cirrose Hepática/microbiologia , Cirrose Hepática Experimental/complicações , Cirrose Hepática Experimental/imunologia , Cirrose Hepática Experimental/microbiologia , Sistema Fagocitário Mononuclear/fisiologia , Peritonite/etiologia , Peritonite/microbiologia , RatosRESUMO
AIM: To evaluate the effect of artesunate (AS) supplementation on bacterial translocation (BT) and gut microbiota in a rat model of liver cirrhosis. METHODS: Fifty-four male Sprague-Dawley rats were randomly divided into a normal control group (N), a liver cirrhosis group (M) and a liver cirrhosis group intervened with AS (MA). Each group was sampled at 4, 6 and 8 wk. Liver cirrhosis was induced by injection of carbon tetrachloride (CCl4), intragastric administration of 10% ethanol, and feeding a high fat diet. Rats in the MA group were intragastrically administered with AS (25 mg/kg body weight, once daily). Injuries of the liver and intestinal mucosa were assessed by hematoxylin-eosin or Masson's trichrome staining. Liver index was calculated as a ratio of the organ weight (g) to body weight (g). The gut microbiota was examined by automated ribosomal intergenic-spacer analysis of fecal DNA. BT was assessed by standard microbiological techniques in the blood, mesenteric lymph nodes (MLNs), liver, spleen, and kidney. RESULTS: Compared to group N, the body weight was reduced significantly in groups M and MA due to the development of liver cirrhosis over the period of 8 wk. The body weight was higher in group MA than in group M. The liver indices were significantly elevated at 4, 6 and 8 wk in groups M and MA compared to group N. AS supplementation partially decreased the liver indices in group MA. Marked histopathologic changes in the liver and small intestinal mucosa in group M were observed, which were alleviated in group MA. Levels of pro-inflammatory interleukin-6 and tumor necrosis factor-α were significantly elevated at 8 wk in ileal homogenates in group M compared to group N, which were decreased after AS supplementation in group MA. The dysbiosis of gut microbiota indicated by the mean diversity (Shannon index) and mean similarity (Sorenson index) was severe as the liver cirrhosis developed, and AS supplementation had an apparent intervention effect on the dysbiosis of gut microbiota at 4 wk. The occurrence of BT was increased in the liver of group M compared to that of group N. AS supplementation reduced BT in group MA at 8 wk. BT also occurred in the MLNs, spleen, and kidney, which was reduced by AS supplementation. BT was not detected in the blood in any group. CONCLUSION: Dysbiosis of gut microbiota, injury of intestinal mucosal barrier and BT occurred as liver cirrhosis progressed, which might enhance inflammation and aggravate liver injury. AS may have other non-antimalarial effects that modulate gut microbiota, inhibit BT and alleviate inflammation, resulting in a reduction in CCl4, alcohol and high fat-caused damages to the liver and intestine.
Assuntos
Anti-Inflamatórios/farmacologia , Artemisininas/farmacologia , Bactérias/efeitos dos fármacos , Translocação Bacteriana/efeitos dos fármacos , Disbiose , Microbioma Gastrointestinal/efeitos dos fármacos , Intestinos/efeitos dos fármacos , Cirrose Hepática Experimental/tratamento farmacológico , Animais , Artesunato , Bactérias/imunologia , Bactérias/metabolismo , Tetracloreto de Carbono , Citocinas/imunologia , Citocinas/metabolismo , Progressão da Doença , Fezes/microbiologia , Mediadores da Inflamação/imunologia , Mediadores da Inflamação/metabolismo , Mucosa Intestinal/metabolismo , Intestinos/imunologia , Intestinos/microbiologia , Cirrose Hepática Experimental/induzido quimicamente , Cirrose Hepática Experimental/imunologia , Cirrose Hepática Experimental/microbiologia , Masculino , Ratos Sprague-Dawley , Fatores de TempoRESUMO
BACKGROUND & AIMS: In advanced cirrhosis, gut bacterial translocation is the consequence of intestinal barrier disruption and leads to bacterial infection. Bile acid abnormalities in cirrhosis could play a role in the integrity of the intestinal barrier and the control of microbiota, mainly through the farnesoid X receptor. We investigated the long-term effects of the farnesoid X receptor agonist, obeticholic acid, on gut bacterial translocation, intestinal microbiota composition, barrier integrity and inflammation in rats with CCl4-induced cirrhosis with ascites. METHODS: Cirrhotic rats received a 2-week course of obeticholic acid or vehicle starting once ascites developed. We then determined: bacterial translocation by mesenteric lymph node culture, ileum expression of antimicrobial peptides and tight junction proteins by qPCR, fecal albumin loss, enteric bacterial load and microbiota composition by qPCR and pyrosequencing of ileum mucosa-attached contents, and intestinal inflammation by cytometry of the inflammatory infiltrate. RESULTS: Obeticholic acid reduced bacterial translocation from 78.3% to 33.3% (p<0.01) and upregulated the expression of the farnesoid X receptor-associated gene small heterodimer partner. Treatment improved ileum expression of antimicrobial peptides, angiogenin-1 and alpha-5-defensin, tight junction proteins zonulin-1 and occludin, and reduced fecal albumin loss and liver fibrosis. Enteric bacterial load normalized, and the distinctive mucosal microbiota of cirrhosis was reduced. Gut immune cell infiltration was reduced and inflammatory cytokine and Toll-like receptor 4 expression normalized. CONCLUSIONS: In ascitic cirrhotic rats, obeticholic acid reduces gut bacterial translocation via several complementary mechanisms at the intestinal level. This agent could be used as an alternative to antibiotics to prevent bacterial infection in cirrhosis.
Assuntos
Ácido Quenodesoxicólico/análogos & derivados , Inflamação/metabolismo , Intestinos/efeitos dos fármacos , Cirrose Hepática Experimental/tratamento farmacológico , Animais , Ácido Quenodesoxicólico/farmacologia , Citocinas/metabolismo , Inflamação/patologia , Mucosa Intestinal/metabolismo , Intestinos/patologia , Cirrose Hepática Experimental/microbiologia , Cirrose Hepática Experimental/patologia , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
Qualitative and quantitative changes in gut microbiota play a very important role in cirrhosis. Humans harbour around 100 quintillion gut bacteria, thus representing around 10 times more microbial cells than eukaryotic ones. The gastrointestinal tract is the largest surface area in the body and it is subject to constant exposure to these living microorganisms. The existing symbiosis, proven by the lack of proinflammatory response against commensal bacteria, implies the presence of clearly defined communication lines that contribute to the maintenance of homeostasis of the host. Therefore, alterations of gut flora seem to play a role in the pathogenesis and progress of multiple liver and gastrointestinal diseases. This has made its selective modification into an area of high therapeutic interest. Bacterial translocation is defined as the migration of bacteria or bacterial products from the intestines to the mesenteric lymph nodes. It follows that alteration in gut microbiota have shown importance, at least to some extent, in the pathogenesis of several complications arising from terminal liver disease, such as hepatic encephalopathy, portal hypertension and spontaneous bacterial peritonitis. This review sums up, firstly, how liver disease can alter the common composition of gut microbiota, and secondly, how this alteration contributes to the development of complications in cirrhosis.
Assuntos
Translocação Bacteriana , Microbioma Gastrointestinal , Cirrose Hepática/microbiologia , Animais , Encefalopatia Hepática/etiologia , Encefalopatia Hepática/microbiologia , Humanos , Hipertensão Portal/etiologia , Hipertensão Portal/microbiologia , Cirrose Hepática/etiologia , Cirrose Hepática Experimental/etiologia , Cirrose Hepática Experimental/microbiologia , Linfonodos/microbiologia , Mesentério , Peritonite/etiologia , Peritonite/microbiologia , RatosRESUMO
Many lines of research have established a relationship between the gut microbiome and patients with liver disease. For example, patients with cirrhosis have increased bacteremia, increased blood levels of lipopolysaccharide, and increased intestinal permeability. Patients with cirrhosis have bacterial overgrowth in the small intestine. Selective intestinal decontamination with antibiotics is beneficial for patients with decompensated cirrhosis. In experimental models of chronic liver injury with fibrosis, several toll-like receptors (TLR) are required to make mice sensitive to liver fibrosis. The presumed ligand for the TLRs are bacterial products derived from the gut microbiome, and TLR knockout mice are resistant to liver inflammation and fibrosis. We and others have characterized the association between preclinical models of liver disease in mice with the microbial diversity in their gut microbiome. In each model, including intragastric alcohol, bile duct ligation, chronic carbon tetrachloride (CCl4), administration, and genetic obesity, there is a significant change in the gut microbiome from normal control mice. However, there is not a single clear bacterial strain or pattern that distinguish mice with liver injury from controlled mice. So how can the gut microbiota affect liver disease? We can identify at least 6 changes that would result in liver injury, inflammation, and/or fibrosis. These include: (1) changes in caloric yield of diet; (2) regulation of gut permeability to release bacterial products; (3) modulation of choline metabolism; (4) production of endogenous ethanol; (5) regulation of bile acid metabolism; and (6) regulation in lipid metabolism.
Assuntos
Microbioma Gastrointestinal , Intestinos/microbiologia , Cirrose Hepática Experimental/microbiologia , Hepatopatias/microbiologia , Fígado/microbiologia , Animais , Ácidos e Sais Biliares/metabolismo , Colina/metabolismo , Dieta , Etanol/metabolismo , Humanos , Inflamação/complicações , Inflamação/microbiologia , Mucosa Intestinal/metabolismo , Ligantes , Metabolismo dos Lipídeos , Fígado/metabolismo , Camundongos , Permeabilidade , Receptores Toll-Like/metabolismoRESUMO
BACKGROUND: Pringle maneuver (Pm) is advocated for the reduction of blood loss during liver surgery, while postoperative infections continue to be a frequent complication after hepatic resection and liver transplantation. AIM: To investigate the effect of the Pringle maneuver on systemic bacterial translocation and endotoxemia in cirrhotic rats and cirrhotic rats with selective intestinal decontamination. METHODS: A total of 100 male Sprague-Dawley cirrhotic rats were randomly divided into five equal groups: sham operation, Pm of 10 min, Pm of 20 min, Pm of 30 min, and pretreatment. Tissue samples from mesenteric lymph nodes, liver, lungs, portal, and vena cava vein blood were obtained for culture after 30 min and 24 h of the operation. Endotoxin levels were measured in portal vein and vena cava blood. RESULTS: Portal vein and vena cava blood endotoxin concentrations increased significantly after 30 min, especially 24 h of operation in the Pm of 20 min and Pm of 30 min groups. A significant increase in contaminated mesenteric lymph nodes, liver, portal, and vena cava blood was noted 24 h later. The incidence of gut bacterial translocation increased with the duration extension of Pm. Escherichia coli was the most common bacteria isolated from the tissues. There was a significant decrease of portal vein and vena cava blood endotoxin concentrations and the incidence of bacterial translocation by selective intestinal decontamination. CONCLUSIONS: There is endotoxemia immediately after Pringle maneuver and gut bacteria translocation 24 h later. The incidence of gut bacterial translocation increases with the duration extension of Pm. Selective intestinal decontamination can decrease bacteria translocation and endotoxemia.
Assuntos
Translocação Bacteriana , Perda Sanguínea Cirúrgica/prevenção & controle , Procedimentos Cirúrgicos do Sistema Digestório/efeitos adversos , Endotoxemia/microbiologia , Intestinos/microbiologia , Cirrose Hepática Experimental/microbiologia , Animais , Tetracloreto de Carbono , Constrição , Endotoxemia/sangue , Endotoxinas/sangue , Cirrose Hepática Experimental/induzido quimicamente , Cirrose Hepática Experimental/patologia , Masculino , Duração da Cirurgia , Ratos , Ratos Sprague-Dawley , Fatores de Risco , Fatores de TempoRESUMO
BACKGROUND & AIMS: Bacterial endotoxin is present in patients with advanced cirrhosis and can induce an immunogenic response without an overt infection. Norfloxacin is a gram-negative bactericidal drug able to maintain low endotoxin levels and stimulate IL-10 production. We aimed at investigating the role of IL-10 in decreasing endotoxin absorption in cirrhotic mice treated with norfloxacin. METHODS: Cirrhosis was induced by carbon tetrachloride or bile duct ligation in wild type and IL10-deficient mice with or without norfloxacin prior to an intragastrical administration of E. coli, K. pneumonia or E. faecalis. Spontaneous and induced bacterial translocation, free endotoxin and cytokine levels were evaluated in mesenteric lymph nodes. Intestinal permeability was followed by fluorimetry and barrier integrity markers were measured in disrupted intestinal samples. The inflammatory-modulating mechanism was characterized in purified intestinal mononuclear cells. RESULTS: Norfloxacin reduced spontaneous and induced MLN positive-cultures in wild type and IL-10-deficient animals. However, reduction of free endotoxin levels was associated with norfloxacin in wild type but not in IL-10-deficient mice. Wild type but not IL-10-deficient mice treated with norfloxacin significantly normalized intestinal permeability and improved gut barrier integrity markers. The toll-like receptor 4-mediated pro-inflammatory milieu was modulated by norfloxacin in a concentration-dependent manner in cultured intestinal mononuclear cells of wild type mice but not of IL-10-deficient mice. The restoration of IL-10 levels in IL-10-deficient animals reactivated the norfloxacin effect on inflammatory-modulation, gut barrier permeability, and luminal endotoxin absorption. CONCLUSION: Norfloxacin not only reduces gram-negative intestinal flora but also participates in an IL-10-driven modulation of gut barrier permeability, thus reducing luminal free endotoxin absorption in experimental cirrhosis.
Assuntos
Endotoxinas/sangue , Escherichia coli , Interleucina-10 , Mucosa Intestinal , Intestinos , Klebsiella pneumoniae , Cirrose Hepática Experimental , Norfloxacino/farmacologia , Animais , Antibacterianos/farmacologia , Translocação Bacteriana/efeitos dos fármacos , Translocação Bacteriana/imunologia , Escherichia coli/efeitos dos fármacos , Escherichia coli/fisiologia , Inflamação/imunologia , Inflamação/prevenção & controle , Interleucina-10/sangue , Interleucina-10/imunologia , Mucosa Intestinal/metabolismo , Intestinos/imunologia , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/fisiologia , Cirrose Hepática Experimental/imunologia , Cirrose Hepática Experimental/microbiologia , Camundongos , Camundongos Endogâmicos BALB C , Permeabilidade/efeitos dos fármacos , Receptor 4 Toll-Like/metabolismo , Resultado do TratamentoRESUMO
BACKGROUND & AIMS: Intestinal hyperpermeability, impaired peritoneal macrophages (PMs) phagocytosis, and bacterial translocation (BT), resulting in increased systemic and local infection/inflammation such as spontaneous bacterial peritonitis (SBP) together with increased tumor necrosis factor-α (TNFα) levels, are all implicated in the pathogenesis of cirrhosis-related complications. Manipulation of the cannabinoid receptors (CB1R and CB2R), which are expressed on the gut mucosa and PMs, has been reported to modulate intestinal inflammation and systemic inflammatory cytokine release. Our study aims to explore the effects of chronic CB1R/CB2R agonist/antagonist treatments on relevant abnormalities in cirrhotic ascitic rats. METHODS: Vehicle, archidonyl-2-chloroethylamide (ACEA, CB1R agonist), JWH133 (CB2R agonist), and AM630 (CB2R antagonist) were given to thioacetamide (TAA) and common bile duct ligation (BDL) cirrhotic rats with ascites for two weeks and various measurement were performed. RESULTS: Compared to sham rats, CB2Rs were downregulated in cirrhotic rat intestines and PMs. The two-week JWH133 treatment significantly decreased systemic/intestinal oxidative stress, TNFα and inflammatory mediators, infection, intestinal mucosal damage and hyperpermeability; the JWH133 treatment also decreased bacterial overgrowth/adhesion, BT and SBP, upregulated intestinal tight junctions and downregulated the PM TNFα receptor/NFκBp65 protein expression in cirrhotic rats. Acute and chronic JWH133 treatment corrected the TNFα-induced suppression of phagocytosis of cirrhotic rat PMs, which then could be reversed by concomitant AM630 treatment. CONCLUSIONS: Our study suggests that CB2R agonists have the potential to treat BT and various relevant abnormalities through inhibition of systemic/intestinal oxidative stress, inflammatory cytokines and TNFα release in cirrhosis. Overall, the chronic CB2R agonist treatment affects multiple approach mechanisms, and its direct effect on the hyperdynamic circulation is only minor.
Assuntos
Ascite/tratamento farmacológico , Translocação Bacteriana/efeitos dos fármacos , Canabinoides/administração & dosagem , Cirrose Hepática Experimental/tratamento farmacológico , Receptor CB2 de Canabinoide/agonistas , Animais , Ácidos Araquidônicos/administração & dosagem , Ascite/complicações , Ascite/microbiologia , Infecções Bacterianas/etiologia , Infecções Bacterianas/prevenção & controle , Citocinas/metabolismo , Células Hep G2 , Humanos , Indóis/administração & dosagem , Mediadores da Inflamação/metabolismo , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/microbiologia , Circulação Hepática/efeitos dos fármacos , Cirrose Hepática Experimental/complicações , Cirrose Hepática Experimental/microbiologia , Macrófagos Peritoneais/efeitos dos fármacos , Macrófagos Peritoneais/fisiologia , Estresse Oxidativo/efeitos dos fármacos , Peritonite/etiologia , Peritonite/prevenção & controle , Fagocitose/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Receptor CB1 de Canabinoide/agonistas , Receptor CB2 de Canabinoide/antagonistas & inibidores , Receptor CB2 de Canabinoide/metabolismoRESUMO
BACKGROUND & AIMS: Intervention in the gut ecosystem is considered as a potential strategy to treat liver diseases and their complications. We have evaluated the effects of Bifidobacterium pseudocatenulatum CECT7765 on bacterial translocation and the liver status in experimental cirrhosis. ANIMALS & METHODS: Liver damage was induced in Balb/c mice by weight-controlled oral administration of carbon tetrachloride. Laparotomies were performed at week 12. One week prior to laparotomy, animals received B. pseudocatenulatum CECT7765 (10(9) cfu/daily) or placebo intragastrically. All animals received Escherichia coli (10(7) cfu/single dose) intragastrically 24 hours before laparotomy. A group of naïve non-treated animals was included as control. Liver tissue specimens, mesenteric lymph nodes, intestinal content and blood were collected. Liver histology, profibrogenic genes expression, bacterial DNA translocation, serum endotoxaemia and liver cytokine levels were measured. RESULTS: Bifidobacterium pseudocatenulatum CECT7765 showed no significant effect on structural liver damage, as determined by histological evaluation, alpha-smooth muscle actin distribution, profibrogenic gene expression levels, total hydroxyproline levels and malon dialdehyde production compared with mice receiving placebo. Interestingly, bacterial DNA translocation and serum endotoxin levels were significantly decreased in mice receiving the Bifidobacterium strain compared with placebo. Gut barrier integrity markers were up-regulated in mice receiving B. pseudocatenulatum CECT7765 and quantitatively correlated with intestinal gene copy numbers of the bifidobacterial strain. Gene expression levels of several anti-inflammatory mediators were also increased in mice receiving B. pseudocatenulatum CECT7765 compared with placebo. CONCLUSION: Oral administration of B. pseudocatenulatum CECT7765 is associated with improved gut barrier integrity and shows a beneficial effect against induced bacterial antigen translocation in the CCl4 -model of cirrhosis.
Assuntos
Translocação Bacteriana , Bifidobacterium/fisiologia , Escherichia coli/imunologia , Intestinos/microbiologia , Cirrose Hepática Experimental/terapia , Fígado/microbiologia , Probióticos , Animais , Antígenos de Bactérias/genética , Antígenos de Bactérias/imunologia , Tetracloreto de Carbono , DNA Bacteriano/genética , Endotoxinas/sangue , Escherichia coli/genética , Feminino , Fígado/patologia , Cirrose Hepática Experimental/induzido quimicamente , Cirrose Hepática Experimental/microbiologia , Cirrose Hepática Experimental/patologia , Camundongos Endogâmicos BALB CRESUMO
AIM: Evidence indicates that intestinal microbiota may participate in both the induction and the progression of liver damage. The aim of our research was the detection and evaluation of the effects of chronic treatment with a symbiotic formulation on CCl4 -induced rat liver fibrosis. RESULTS: CCl4 significantly increased gastric permeability in respect to basal values, and the treatment with symbiotic significantly decreased it. CCl4 per se induced a decrease in intestinal permeability. This effect was also seen in fibrotic rats treated with symbiotic and was still evident when normal rats were treated with symbiotic alone (P < 0.001 in all cases). Circulating levels of pro-inflammatory cytokine TNF-α were significantly increased in rats with liver fibrosis as compared with normal rats, while symbiotic treatment normalized the plasma levels of TNF-α and significantly enhanced anti-inflammatory cytokine IL 10. TNF-α, TGF-ß, TLR4, TLR2, iNOS and α-SMA mRNA expression in the liver were up-regulated in rats with CCl4 -induced liver fibrosis and down-regulated by symbiotic treatment. Moreover, IL-10 and eNOS mRNA levels were increased in the CCL4 (+) symbiotic group. Symbiotic treatment of fibrotic rats normalized serum ALT, AST and improved histology and liver collagen deposition. DGGE analysis of faecal samples revealed that CCl4 administration and symbiotic treatment either alone or in combination produced modifications in faecal profiles vs controls. CONCLUSIONS: Our results provide evidence that in CCl4 -induced liver fibrosis, significant changes in gastro-intestinal permeability and in faecal flora occur. Treatment with a specific symbiotic formulation significantly affects these changes, leading to improvement in both liver inflammation and fibrosis.
Assuntos
Trato Gastrointestinal/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Lactobacillus/metabolismo , Cirrose Hepática Experimental/tratamento farmacológico , Permeabilidade/efeitos dos fármacos , Probióticos/farmacologia , Análise de Variância , Animais , Tetracloreto de Carbono/toxicidade , Cromatografia Líquida de Alta Pressão , Citocinas/metabolismo , Eletroforese em Gel de Gradiente Desnaturante , Fezes/química , Galactanos/farmacologia , Trato Gastrointestinal/microbiologia , Regulação da Expressão Gênica/imunologia , Glutamina/farmacologia , Cirrose Hepática Experimental/induzido quimicamente , Cirrose Hepática Experimental/microbiologia , Masculino , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase em Tempo Real , Fator de Necrose Tumoral alfa/sangueAssuntos
Antioxidantes/farmacologia , Translocação Bacteriana/efeitos dos fármacos , Lactobacillus , Cirrose Hepática Experimental/microbiologia , Probióticos/farmacologia , Animais , Tetracloreto de Carbono/toxicidade , Cirrose Hepática Experimental/induzido quimicamente , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
BACKGROUND: The present study investigated the alteration of intestinal microbiota, innate immunity-related genes, and bacterial translocation in rats with cirrhosis and liver transplantation. METHODS: Specific pathogen-free Sprague-Dawley rats were randomized into 4 groups: (1) normal controls (N); (2) liver cirrhosis (LC); (3) normal control groups with liver transplantation (LTN); and (4) liver cirrhosis with liver transplantation (LTC). We examined plasma endotoxin, bacterial tacslocation, denaturing gradient gel electrophoresis (DGGE) profile of intestinal mucosa-associated bacteria, abundance of key bacterial populations, and expression of innate immunity-related gene. RESULTS: The LTC and LC group, showed higher endotoxin levels (1.08±0.73 EU/mL and 0.74±0.70 EU/mL, respectively) than the N group (0.27±0.13 EU/mL; P<.05). the incidence of bacterial translocation (BT) to liver and mesenteric lymph nodes (MLN), and the number of total bacteria were increased significantly in the LTC and LC groups compared with the N group (P<.05). The counts of Lactobacilli and Bacteroides were lower, whereas Enterobacteria were higher in the LC than the N group (P<.05). Mucins (MUC2, MUC3) and Toll-like receptors (TLR2, TLR4) messenger RNA (mRNA) expression were significantly higher in the LC and LTC groups than the N group (P<.05). The marked difference between the groups in the overall structure of the bacterial community was also generated by DGGE profiles. CONCLUSION: Liver cirrhosis disturbs intestinal microbiota and innate immunity-related genes, which contributes to endotoxemia and bacterial translocation. These had not completely recovered in cirrhotic rats until 1 month after orthotopic liver transplantation.
Assuntos
Translocação Bacteriana , Íleo/microbiologia , Imunidade Inata/genética , Intestinos/microbiologia , Cirrose Hepática Experimental/cirurgia , Transplante de Fígado , Animais , Tetracloreto de Carbono , Eletroforese em Gel de Gradiente Desnaturante , Endotoxinas/sangue , Regulação da Expressão Gênica , Íleo/imunologia , Cirrose Hepática Experimental/induzido quimicamente , Cirrose Hepática Experimental/genética , Cirrose Hepática Experimental/imunologia , Cirrose Hepática Experimental/microbiologia , Masculino , Mucina-2/genética , Mucina-3/genética , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase em Tempo Real , Fatores de Tempo , Receptor 2 Toll-Like/genética , Receptor 4 Toll-Like/genéticaRESUMO
BACKGROUND/PURPOSE: The intestinal microflora plays a major role in human health. Intestinal flora imbalances are seen in clinical settings, such as cirrhosis, in which bacterial translocation (BT) results in migration of bacteria or bacterial products from the intestinal lumen to extraintestinal sites. In this study, we explored the effect of alterations in gut flora on BT and liver function in cirrhotic rats. METHODS: In this study, we used a carbon tetrachloride-induced cirrhotic rat model to compare the abundance of major aerobic and anaerobic bacterial species in healthy and cirrhotic rats. We used antibiotic (norfloxacin) and different probiotic treatments to change the status of gut flora in the cirrhotic rats and evaluated BT, liver function, and endotoxemia in the different models. RESULTS: We found higher levels of Enterobacteriaceae in cirrhotic rats when compared with healthy rats. Bifidobacteria treatment resulted in lower levels of Enterobacteriaceae along with increased levels of Lactobacillus when compared with the normal saline group. Both Bifidobacteria and Enterococcus treatments resulted in lower endotoxin levels than in the normal saline group. CONCLUSION: Gut flora imbalances in cirrhotic rats result in significant changes in BT and liver function in cirrhotic rats.
Assuntos
Translocação Bacteriana , Endotoxemia/microbiologia , Intestinos/microbiologia , Cirrose Hepática Experimental/microbiologia , Fígado/microbiologia , Animais , Antibacterianos/farmacologia , Translocação Bacteriana/efeitos dos fármacos , Bifidobacterium/crescimento & desenvolvimento , Tetracloreto de Carbono , Endotoxemia/sangue , Enterobacteriaceae/crescimento & desenvolvimento , Enterococcus/crescimento & desenvolvimento , Intestinos/efeitos dos fármacos , Lactobacillus/crescimento & desenvolvimento , Fígado/metabolismo , Cirrose Hepática Experimental/sangue , Cirrose Hepática Experimental/induzido quimicamente , Masculino , Norfloxacino/farmacologia , Probióticos , Ratos , Ratos Sprague-DawleyRESUMO
INTRODUCTION: Spontaneous bacterial peritonitis (SBP) is mainly caused by bacterial translocation of enteric Gram-negative bacteria, predominantly Escherichia coli. The sympathetic nervous system (SNS) is activated in advanced cirrhosis, particularly in the splanchic circulation, and exerts potent immunosuppressive actions. However, the role of splanchnic SNS activity in bacterial translocation and bacterial spreading in cirrhosis remains unclear. METHODS: E coli or Stapylococcus aureus (10(6) CFU) were given intraperitoneally. After 6 h, mesenteric lymph nodes (MLN), liver, spleen, lung and peripheral blood were harvested from ascitic cirrhotic rats (LC) and healthy controls with and without splanchnic sympathectomy (SE). The bacterial tissue burden was determined by standard microbiological culture techniques. In vitro phagocytic activity of peritoneal polymorphonuclear leucocytes was assessed by FACS analysis. RESULTS: Under basal conditions SE reduced bacterial translocation to MLN in LC rats from 45% to 17%. LC rats had a marked increase in bacteraemia after E coli and S aureus challenge and an increased incidence and degree of E coli translocation to MLN, liver, spleen and lung compared with control rats. SE prevented bacteraemia in LC rats after E coli but not after S aureus challenge. Prior SE abolished the difference in incidence as well as the bacterial tissue burden in each organ after E coli application in LC rats, being no longer significantly different from control rats with or without SE. The protective effects of SE against E coli were associated with a greater influx of mononuclear cells into the peritoneal cavity and increased phagocytic activity of peritoneal polymorphonuclear leucocytes. CONCLUSIONS: In cirrhosis with bacterial peritonitis, hyperactivity of the splanchnic sympathetic nervous system contributes to the translocation of E coli but not S aureus to MLN and extraintestinal sites. This indicates a key role for sympathetic drive in the impairment in host defence against Gram-negative bacteria in cirrhosis.
Assuntos
Translocação Bacteriana/fisiologia , Escherichia coli/fisiologia , Cirrose Hepática Experimental/microbiologia , Peritonite/microbiologia , Nervos Esplâncnicos/fisiopatologia , Staphylococcus aureus/fisiologia , Animais , Células Cultivadas , Infecções por Escherichia coli/microbiologia , Infecções por Escherichia coli/fisiopatologia , Intestino Delgado/inervação , Cirrose Hepática Experimental/imunologia , Cirrose Hepática Experimental/fisiopatologia , Masculino , Infiltração de Neutrófilos/fisiologia , Neutrófilos/imunologia , Cavidade Peritoneal/citologia , Peritonite/imunologia , Peritonite/fisiopatologia , Fagocitose/imunologia , Ratos , Infecções Estafilocócicas/microbiologia , Infecções Estafilocócicas/fisiopatologia , SimpatectomiaRESUMO
In summary the carbon tetrachloride/phenobarbital of cirrhosis in rats mimics human cirrhosis very closely, with development of ascites and SBP. This model shows us that bacterial overgrowth occurs as cirrhosis progresses and that bacterial translocation from the gut to extra-intestinal sites is part of the early pathogenesis of SBP. SID with norfloxacin dramatically reduced translocation and SBP at the expense of grampositive overgrowth and infection with gram-positives and colonization with strange gram negatives. SID with TMP-SMZ actually delayed development of ascites and prolonged survival.