RESUMEN
BACKGROUND & AIMS: Bacterial infections commonly occur in decompensated cirrhosis resulting from bacterial translocation from the intestine. We studied the role of intestinal macrophages and the epithelial barrier in cirrhosis. METHODS: Forty-four patients with NASH/ASH cirrhosis (decompensated n=29, compensated n=15) and nineteen controls undergoing endoscopy were recruited. Serum was obtained and LPS and LBP levels determined. Intestinal macrophages were characterized by flow cytometry, immunohistochemistry, and nitric oxide (NO) production measured in supernatant of cultured duodenal samples. Quantitative RT-PCR was performed on duodenal biopsies assessing 84 inflammatory genes. Protein levels of cytokines/chemokines were assessed in serum and supernatant. The duodenal wall was assessed by electron microscopy, tight junction protein expression determined by RT-PCR, immunohistochemistry, and Western blot and, functional analysis performed by transepithelial resistance measurement and permeability studies. RESULTS: Increased plasma LPS, LBP levels and higher numbers of duodenal CD33(+)/CD14(+)/Trem-1(+) macrophages, synthesizing iNOS and secreting NO were present in decompensated cirrhosis. Upregulation of IL-8, CCL2, CCL13 at the transcriptional level, and increased IL-8, and IL-6 were detected in supernatant and serum in cirrhosis. IL-6 and IL-8 co-localised with iNOS(+) and CD68(+), but not with CD11c(+) cells. Electron microscopy demonstrated an intact epithelial barrier. Increased Claudin-2 was detected by Western blot and immunohistochemistry, while decreased transepithelial resistance and increased duodenal permeability were detected in decompensated cirrhosis. CONCLUSIONS: Our study shows the presence of activated CD14(+)Trem-1(+)iNOS(+) intestinal macrophages, releasing IL-6, NO, and increased intestinal permeability in patients with cirrhosis, suggesting that these cells may produce factors capable of enhancing permeability to bacterial products.
Asunto(s)
Interleucina-6/metabolismo , Intestinos/inmunología , Cirrosis Hepática/inmunología , Activación de Macrófagos , Macrófagos/inmunología , Óxido Nítrico/metabolismo , Anciano , Femenino , Humanos , Mucosa Intestinal/metabolismo , Receptores de Lipopolisacáridos/análisis , Macrófagos/metabolismo , Masculino , Persona de Mediana Edad , Óxido Nítrico Sintasa de Tipo II/metabolismo , PermeabilidadRESUMEN
OBJECTIVES: Tigecycline is the prototype of the recently introduced, intravenously administered glycylcycline class of antibiotics, developed in response to the increasing problem of antibiotic resistance in Gram-positive bacteria, especially Staphylococcus aureus, as well as Gram-negative bacteria and anaerobes. However, relatively little is known about the immunomodulatory potential of tigecycline, specifically its interactions with human neutrophils. In the current study we investigated the effects of tigecycline at therapeutically relevant concentrations and greater (0.625-10 mg/L) on alterations in cytosolic Ca(2+) concentrations, generation of antimicrobial reactive oxygen species (ROS) and release of granule proteases [elastase, matrix metalloproteinase-8 (MMP-8) and matrix metalloproteinase-9 (MMP-9)] by human blood neutrophils activated with the chemoattractant N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP; 1 µM). METHODS: Cytosolic Ca(2+) concentrations were measured using fura-2/AM-based spectrofluorimetry and radiometric procedures, generation of ROS by oxygen consumption and myeloperoxidase-mediated auto-iodination, and protease release by ELISA procedures. RESULTS: Exposure of the cells to fMLP resulted in activation of the generation of ROS, as well as release of the granule proteases, all of which were significantly increased by pre-incubation of the cells with tigecycline in a dose-dependent manner. Tigecycline-mediated enhancement of these neutrophil functions was associated with elevations in the concentrations of cytosolic Ca(2+), which appeared to result from the Ca(2+) ionophore activity of tigecycline. CONCLUSIONS: Tigecycline, by functioning as a Ca(2+) ionophore, and independent of antimicrobial activity, potentiates the pro-inflammatory functions of human neutrophils in vitro.