RESUMEN
Dietary plant sterols and stanols as present in our diet and in functional foods are well-known for their inhibitory effects on intestinal cholesterol absorption, which translates into lower low-density lipoprotein cholesterol concentrations. However, emerging evidence suggests that plant sterols and stanols have numerous additional health effects, which are largely unnoticed in the current scientific literature. Therefore, in this review we pose the intriguing question "What would have occurred if plant sterols and stanols had been discovered and embraced by disciplines such as immunology, hepatology, pulmonology or gastroenterology before being positioned as cholesterol-lowering molecules?" What would then have been the main benefits and fields of application of plant sterols and stanols today? We here discuss potential effects ranging from its presence and function intrauterine and in breast milk towards a potential role in the development of non-alcoholic steatohepatitis (NASH), cardiovascular disease (CVD), inflammatory bowel diseases (IBD) and allergic asthma. Interestingly, effects clearly depend on the route of entrance as observed in intestinal-failure associated liver disease (IFALD) during parenteral nutrition regimens. It is only until recently that effects beyond lowering of cholesterol concentrations are being explored systematically. Thus, there is a clear need to understand the full health effects of plant sterols and stanols.
Asunto(s)
Asma/tratamiento farmacológico , Enfermedades Cardiovasculares/tratamiento farmacológico , Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Enfermedad del Hígado Graso no Alcohólico/tratamiento farmacológico , Fitosteroles/farmacología , Sitoesteroles/farmacología , Asma/metabolismo , Enfermedades Cardiovasculares/metabolismo , Colesterol/metabolismo , LDL-Colesterol/antagonistas & inhibidores , LDL-Colesterol/metabolismo , Humanos , Enfermedades Inflamatorias del Intestino/metabolismo , Absorción Intestinal/efectos de los fármacos , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Fitosteroles/administración & dosificación , Sitoesteroles/administración & dosificaciónRESUMEN
BACKGROUND AND AIM: T cells are key players in the chronic intestinal inflammation that characterises Crohn's disease. Here we aim to map the intestinal T-cell receptor [TCR] repertoire in patients with Crohn's disease, using next-generation sequencing technology to examine the clonality of the T-cell compartment in relation to mucosal inflammation and response to therapy. METHODS: Biopsies were taken from endoscopically inflamed and uninflamed ileum and colon of 19 patients with Crohn's disease. From this cohort, additional biopsies were taken after 8 weeks of remission induction therapy from eight responders and eight non-responders. Control biopsies from 11 patients without inflammatory bowel disease [IBD] were included. The TCRß repertoire was analysed by next-generation sequencing of biopsy RNA. RESULTS: Both in Crohn's disease patients and in non-IBD controls, a broad intestinal T-cell repertoire was found, with a considerable part consisting of expanded clones. Clones in Crohn's disease were more expanded [p = 0.008], with the largest clones representing up to as much as 58% of the total repertoire. There was a substantial overlap of the repertoire between inflamed and uninflamed tissue and between ileum and colon. Following therapy, responders showed larger changes in the T-cell repertoire than non-responders, although a considerable part of the repertoire remained unchanged in both groups. CONCLUSIONS: The intestinal T-cell repertoire distribution in Crohn's disease is different from that in the normal gut, containing profoundly expanded T-cell clones that take up a large part of the repertoire. The T-cell repertoire is fairly stable regardless of endoscopic mucosal inflammation or response to therapy.
Asunto(s)
Enfermedad de Crohn/inmunología , Enfermedad de Crohn/patología , Receptores de Antígenos de Linfocitos T/metabolismo , Linfocitos T/metabolismo , Linfocitos T/patología , Adalimumab/uso terapéutico , Adulto , Antiinflamatorios/uso terapéutico , Biopsia , Budesonida/uso terapéutico , Proteína C-Reactiva/metabolismo , Estudios de Casos y Controles , Células Clonales/efectos de los fármacos , Colon/patología , Enfermedad de Crohn/tratamiento farmacológico , Femenino , Fármacos Gastrointestinales/uso terapéutico , Humanos , Íleon/patología , Inflamación/inmunología , Inflamación/patología , Infliximab/uso terapéutico , Masculino , Persona de Mediana Edad , Índice de Severidad de la Enfermedad , Linfocitos T/efectos de los fármacos , Adulto JovenRESUMEN
Thiopurines are commonly used drugs in the therapy of Crohn's disease, but unfortunately only show a 30% response rate. The biological basis for the thiopurine response is unclear, thus hampering patient selection prior to treatment. A genetic risk factor associated specifically with Crohn's disease is a variant in ATG16L1 that reduces autophagy. We have previously shown that autophagy is involved in dendritic cell (DC)-T-cell interactions and cytoskeletal regulation. Here we further investigated the role of autophagy in DC cytoskeletal modulation and cellular trafficking. Autophagy-deficient DC displayed loss of filopodia, altered podosome distribution, and increased membrane ruffling, all consistent with increased cellular adhesion. Consequently, autophagy-deficient DC showed reduced migration. The cytoskeletal aberrations were mediated through hyperactivation of Rac1, a known thiopurine target. Indeed thiopurines restored the migratory defects in autophagy-deficient DC. Clinically, the ATG16L1 risk variant associated with increased response to thiopurine treatment in patients with Crohn's disease but not ulcerative colitis. These results suggest that the association between ATG16L1 and Crohn's disease is mediated at least in part through Rac1 hyperactivation and subsequent defective DC migration. As this phenotype can be corrected using thiopurines, ATG16L1 genotyping may be useful in the identification of patients that will benefit most from thiopurine treatment.
Asunto(s)
Proteínas Relacionadas con la Autofagia/metabolismo , Autofagia , Enfermedad de Crohn/inmunología , Células Dendríticas/fisiología , Proteína de Unión al GTP rac1/metabolismo , Alelos , Animales , Autofagia/genética , Proteínas Relacionadas con la Autofagia/genética , Estructuras de la Membrana Celular/patología , Movimiento Celular , Células Cultivadas , Colitis Ulcerosa/tratamiento farmacológico , Enfermedad de Crohn/tratamiento farmacológico , Enfermedad de Crohn/genética , Citoesqueleto/metabolismo , Células Dendríticas/patología , Femenino , Predisposición Genética a la Enfermedad , Humanos , Mercaptopurina/uso terapéutico , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Polimorfismo Genético , ARN Interferente Pequeño/genética , RiesgoRESUMEN
This literature review assesses the current knowledge about the immunological aspects of the vermiform appendix in health and disease. An essential part of its immunological function is the interaction with the intestinal bacteria, a trait shown to be preserved during its evolution. The existence of the appendiceal biofilm in particular has proved to have a beneficial effect for the entire gut. In assessing the influence of acute appendicitis and the importance of a normally functioning gut flora, however, multiple immunological aspects point towards the appendix as a priming site for ulcerative colitis. Describing the immunological and microbiotical changes in the appendix during acute and chronic inflammation of the appendix, this review suggests that this association becomes increasingly plausible. Sustained by the distinct composition of cells, molecules and microbiota, as well as by the ever more likely negative correlation between the appendix and ulcerative colitis, the idea of the appendix being a vestigial organ should therefore be discarded.
Asunto(s)
Apéndice/inmunología , Animales , Apendicitis/complicaciones , Apendicitis/inmunología , Apendicitis/metabolismo , Apendicitis/patología , Apéndice/citología , Apéndice/metabolismo , Apéndice/patología , Evolución Biológica , Humanos , Enfermedades Inflamatorias del Intestino/complicaciones , Enfermedades Inflamatorias del Intestino/inmunología , Enfermedades Inflamatorias del Intestino/metabolismo , Enfermedades Inflamatorias del Intestino/patología , Mucosa Intestinal/citología , Mucosa Intestinal/inmunología , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patología , Linfocitos/inmunología , Linfocitos/metabolismo , MicrobiotaRESUMEN
The human appendix has long been considered as a vestigial organ, an organ that has lost its function during evolution. In recent years, however, reports have emerged that link the appendix to numerous immunological functions in humans. Evidence has been presented for an important role of the appendix in maintaining intestinal health. This theory suggests that the appendix may be a reservoir or 'safe house' from which the commensal gut flora can rapidly be reestablished if it is eradicated from the colon. However, the appendix may also have a role in the development of inflammatory bowel disease (IBD). Several large epidemiological cohort studies have demonstrated the preventive effect of appendectomy on the development of ulcerative colitis, a finding that has been confirmed in murine colitis models. In addition, current studies are examining the possible therapeutic effect of an appendectomy to modulate disease course in patients with ulcerative colitis. This literature review assesses the current knowledge about the clinical and immunological aspects of the vermiform appendix in IBD and suggests that the idea of the appendix as a vestigial remnant should be discarded.
Asunto(s)
Apéndice/inmunología , Colitis Ulcerosa/inmunología , Disbiosis/inmunología , Microbioma Gastrointestinal/inmunología , Apendicectomía , Apéndice/microbiología , Linfocitos B/inmunología , Colitis Ulcerosa/epidemiología , Enfermedad de Crohn/epidemiología , Enfermedad de Crohn/inmunología , Progresión de la Enfermedad , Disbiosis/epidemiología , Humanos , Inmunoglobulina A/inmunología , Células T Asesinas Naturales/inmunología , Factores Protectores , Índice de Severidad de la EnfermedadRESUMEN
MiR-511-3p is embedded in intron 5 of the CD206/MRC1 gene Mrc1, expressed by macrophage and dendritic cell populations. CD206 and miR-511-3p expression are co-regulated, and their contribution to intestinal inflammation is unclear. We investigated their roles in intestinal inflammation in both mouse and human systems. Colons of CD206-deficient mice displayed normal numbers of monocytes, macrophage, and dendritic cells. In experimental colitis, CD206-deficient mice had attenuated inflammation compared with wild-type (WT) mice. However, neither a CD206 antagonist nor a blocking antibody reproduced this phenotype, suggesting that CD206 was not involved in this response. Macrophages isolated from CD206-deficient mice had reduced levels of miR-511-3p and Tlr4 compared with WT, which was associated with reduced pro-inflammatory cytokine production upon lipopolysaccharides (LPS) and fecal supernatant stimulation. Macrophages overexpressing miR-511-3p showed 50% increase of Tlr4 mRNA, whereas knockdown of miR-511-3p reduced Tlr4 mRNA levels by 60%, compared with scrambled microRNA (miRNA)-transduced cells. Response to anti-tumor necrosis factor (TNF) treatment has been associated with elevated macrophage CD206 expression in the mucosa. However, in colon biopsies no statistically significant change in miR-511-3p was detected. Taken together, our data show that miR-511-3p controls macrophage-mediated microbial responses and is involved in the regulation of intestinal inflammation.
Asunto(s)
Colitis/inmunología , Colon/inmunología , Macrófagos/inmunología , Glicoproteínas de Membrana/genética , MicroARNs/genética , Receptores de Superficie Celular/genética , Animales , Células Cultivadas , Colitis/inducido químicamente , Sulfato de Dextran , Femenino , Regulación de la Expresión Génica , Humanos , Lipopolisacáridos/inmunología , Masculino , Glicoproteínas de Membrana/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptores de Superficie Celular/metabolismo , Receptores Inmunológicos , Receptor Toll-Like 4/genética , Receptor Toll-Like 4/metabolismoRESUMEN
AIM: Various types of cholinergic receptors are expressed on intestinal epithelia. Their function is not completely understood. We hypothesize that cholinergic receptor activation on epithelium may serve a protective function in cytokine-induced barrier dysfunction. METHODS: The effect of cholinergic receptor activation on cellular barrier function in epithelial cells was assessed by measuring electrical impedance, and by determining para-cellular transport in transwell experiments. Cell lysates treated with cytokine and/or cholinergic agonists were analysed for cyto- and chemokine production, and tight junction (TJ) protein rearrangement was assessed. Primary colonic epithelial cells were isolated from surgically resected colon tissue of patients with inflammatory bowel disease. RESULTS: IL-1ß induced production of chemokines (CXCL-1, CXCL-10, IL-8, CCL-7) and led to a rearrangement of TJ proteins (occludin and ZO-1). This response was inhibited by pre-treatment with muscarinic, rather than nicotinic, acetylcholine receptor agonists. Treatment with IL-1ß enhanced paracellular permeability (4kD dextran) and reduced impedance across the monolayer, which was counteracted by pre-incubation with acetylcholine, or muscarinic receptor agonist bethanechol. The protective effect of acetylcholine was antagonized by atropine, underscoring muscarinic receptor involvement. IL-1ß induced transcription of myosin light chain kinase and phosphorylation of myosin light chain, and this cytokine-induced phosphorylation of MLC was inhibited by muscarinic receptor agonists. Furthermore, in epithelial cells from resection material of patients with Crohn's disease and ulcerative colitis, high expression of CXCL-8 was associated with a reduced choline acetyl transferase expression, suggesting an aberrant epithelial production of ACh in inflammatory context. CONCLUSION: Acetylcholine acts on muscarinic receptors on epithelial cells to maintain epithelial barrier function under inflammatory conditions.
Asunto(s)
Citocinas/metabolismo , Células Epiteliales/citología , Células Epiteliales/metabolismo , Receptores Colinérgicos/metabolismo , Animales , Línea Celular , Supervivencia Celular , Citocinas/genética , Regulación de la Expresión Génica/fisiología , Humanos , Interleucina-1beta/farmacología , Ratones , Ocludina/genética , Ocludina/metabolismo , Receptores Colinérgicos/genética , Receptores Muscarínicos/genética , Receptores Muscarínicos/metabolismo , Uniones Estrechas/metabolismo , Proteína de la Zonula Occludens-1/genética , Proteína de la Zonula Occludens-1/metabolismoRESUMEN
Honey has been used successfully in wound healing for thousands of years. The peptide hormone human epidermal growth factor (hEGF) is also known to have a beneficial effect in various wound healing processes via mechanisms that differ from those for honey. In this study, we show that hEGF can be incorporated into honey via nectar. Plants of Nicotiana langsdorffii x N. sanderae were transformed with the gene for hEGF, equipped with a nectary-targeted promoter and a signal sequence for secretion to nectar. These plants accumulated hEGF in the nectar. The maximum hEGF concentration recorded with ELISA in these plants is 2.5 ng·ml⻹. There is a significant linear relationship (P<0.001) between hEGF concentration and induction of hEGF-receptor phosphorylation. Since the flower morphology of these plants did not allow production of honey from their nectar, we used feeding solutions, spiked with synthetic hEGF, to study transfer of this peptide into honey through bee activity. Transfer of hEGF from a feeding solution to honey by bees occurred with retention of the hEGF concentration and the capacity to induce hEGF-receptor phosphorylation. These observations indicate that plants can function as a production platform for honey containing biologically active peptides, which may enhance wound healing and other biological processes.
Asunto(s)
Abejas , Factor de Crecimiento Epidérmico/biosíntesis , Factor de Crecimiento Epidérmico/genética , Miel , Nicotiana/genética , Nicotiana/metabolismo , Néctar de las Plantas/metabolismo , Animales , Flores/genética , Flores/metabolismo , Humanos , Plantas Modificadas Genéticamente , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/genética , Cicatrización de Heridas/efectos de los fármacosRESUMEN
Honey has potent activity against both antibiotic-sensitive and -resistant bacteria, and is an interesting agent for topical antimicrobial application to wounds. As honey is diluted by wound exudate, rapid bactericidal activity up to high dilution is a prerequisite for its successful application. We investigated the kinetics of the killing of antibiotic-resistant bacteria by RS honey, the source for the production of Revamil® medical-grade honey, and we aimed to enhance the rapid bactericidal activity of RS honey by enrichment with its endogenous compounds or the addition of antimicrobial peptides (AMPs). RS honey killed antibiotic-resistant isolates of Pseudomonas aeruginosa, Staphylococcus epidermidis, Enterococcus faecium, and Burkholderia cepacia within 2 h, but lacked such rapid activity against methicillin-resistant S. aureus (MRSA) and extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli. It was not feasible to enhance the rapid activity of RS honey by enrichment with endogenous compounds, but RS honey enriched with 75 µM of the synthetic peptide Bactericidal Peptide 2 (BP2) showed rapid bactericidal activity against all species tested, including MRSA and ESBL E. coli, at up to 10-20-fold dilution. RS honey enriched with BP2 rapidly killed all bacteria tested and had a broader spectrum of bactericidal activity than either BP2 or honey alone.
Asunto(s)
Péptidos Catiónicos Antimicrobianos/farmacología , Bacterias/efectos de los fármacos , Miel , Viabilidad Microbiana/efectos de los fármacos , Bacterias/aislamiento & purificación , HumanosRESUMEN
BACKGROUND & AIMS: Interleukin (IL) 18 has proinflammatory effects. IL-18 plays a pivotal role in Th1 responses, but its proinflammatory activities extend beyond Th1 cells, including macrophages and production of tumor necrosis factor (TNF) alpha and IL-1beta. IL-18 is up-regulated in colonic specimens of patients with Crohn's disease. The goal of this study was to evaluate the role of IL-18. METHODS: Activity of IL-18 was neutralized using recombinant human IL-18 binding protein isoform a (rhIL-18BPa) in trinitrobenzene sulfonic acid (TNBS)-induced colitis. RESULTS: Mice treated daily with rhIL-18BPa (8 mg/kg) had significant reductions in clinical score, body weight loss, and colon weight increase compared with saline-treated mice. Histologic analysis showed that rhIL-18BPa-treated mice developed only mild colitis without signs of ulceration, with a mean total score of 9.8 +/- 1.3 points compared with 15.9 +/- 1.1 points observed in saline-treated mice with colitis. Analysis of cytokine levels in colon homogenates showed a significant decrease in TNF-alpha, IL-6, and IL-1beta after rhIL-18BPa treatment but no effect on interferon gamma. The therapeutic potential of rhIL-18BPa treatment was confirmed in TNBS mice that were treated only on days 8 and 9 after the start of the experiment. In these mice, significant reductions in total colitis score and colon weight were also observed. CONCLUSIONS: These findings show that inhibition of rhIL-18BPa bioactivity, via rhIL-18BPa, may be beneficial for the treatment of IBD.
Asunto(s)
Colitis/inducido químicamente , Colitis/fisiopatología , Glicoproteínas/uso terapéutico , Interleucina-18/antagonistas & inhibidores , Ácido Trinitrobencenosulfónico , Factor de Necrosis Tumoral alfa/antagonistas & inhibidores , Animales , Peso Corporal/efectos de los fármacos , Colitis/tratamiento farmacológico , Colitis/patología , Femenino , Humanos , Péptidos y Proteínas de Señalización Intercelular , Mucosa Intestinal/efectos de los fármacos , Mucosa Intestinal/patología , Ratones , Ratones Endogámicos BALB C , Isoformas de Proteínas/farmacología , Isoformas de Proteínas/uso terapéutico , Proteínas Recombinantes/uso terapéuticoRESUMEN
CD4+ T lymphocytes in the lamina propria (LP) of the gut play a central role in the immune response in inflammatory bowel disease (IBD). CXCR3 is a chemokine receptor expressed on activated T lymphocytes, and a key component for the recruitment of T helper (Th1) effector cells to the site of inflammation. To determine if CXCR3 is involved in localization of T cells to the gut in IBD patients, we investigated the expression of CXCR3 on CD4+ T lymphocytes in the LP and in the submucosa of resection specimens from 51 IBD patients and 15 control patients. Positive cells were microscopically scored using a semiquantitative analysis on a five-point scale. We found that CD4+ T cells, CXCR3+ cells, and CD4+CXCR3+ T cells in the LP were slightly increased in both IBD groups compared with control non-IBD specimens. In addition, CD4+ and CXCR3+ cells in the submucosa were significant increased in the CD group compared with the control group. CD4+ and CXCR3+ expression was not statistically different between CD and UC. Flow cytometry was used to analyze the percentage of CXCR3+ cells within the CD4+ T-cell population isolated from biopsy specimens and peripheral blood from IBD patients and control patients. There was no difference in the percentage of CD4+CXCR3+ cells between the different groups in the gut as well as in the circulation. These results suggest that CD4+CXCR3+ T cells migrate to the normal and inflamed intestinal mucosa, indicating a role in maintaining normal gut homeostasis. The selective expression of CXCR3+ cells in the submucosa of CD patients might also indicate that these cells play a role in inflammation.
Asunto(s)
Enfermedades Inflamatorias del Intestino/metabolismo , Receptores de Quimiocina/metabolismo , Subgrupos de Linfocitos T/metabolismo , Adulto , Anciano , Membrana Basal/metabolismo , Linfocitos T CD4-Positivos/citología , Linfocitos T CD4-Positivos/metabolismo , Estudios de Casos y Controles , Femenino , Citometría de Flujo , Humanos , Inmunohistoquímica , Mucosa Intestinal/metabolismo , Masculino , Persona de Mediana Edad , Receptores CXCR3 , Subgrupos de Linfocitos T/citologíaRESUMEN
IFN-gamma is a potent pro-inflammatory cytokine thought to be involved in the pathogenesis of Crohn's disease. To further define the role of IFN-gamma in intestinal inflammation, we studied the effects of intra-colonic 2,4,6-trinitrobenzene sulfonic acid (TNBS) instillation in mice with a functionally inactivated IFN-gamma receptor 1 (IFN-gammaR1(- / -)). Our results indicate that IFN-gamma is not necessary for the induction of hapten-induced colitis: after TNBS administration both wild-type and IFN-gammaR1(- / -) mice lost body weight, and the histological features of TNBS-induced colitis were comparable. Colons of IFN-gammaR1(- / -) mice contained a greater number of cells, represented by macrophages and CD4(+) T cells; caudal lymph node cells produced more IFN-gamma and TNF-alpha upon stimulation in vitro. Moreover, IL-18 and IL-12 p40 RNA levels were comparably up-regulated after TNBS treatment in IFN-gammaR1(- / -) wild-type mice. These findings demonstrate that IFN-gamma is dispensable for the development of TNBS-induced colitis. Importantly, the production of Th1 cytokines (e. g. IFN-gamma and TNF-alpha) by caudal lymph node T lymphocytes was enhanced rather than decreased in IFNgammaR1(- / -) mice with no evidence for default Th2 development.
Asunto(s)
Colitis/inmunología , Interferón gamma/inmunología , Células TH1/inmunología , Animales , Colitis/inducido químicamente , Haptenos , Inflamación/inmunología , Interferón gamma/deficiencia , Interferón gamma/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ácido TrinitrobencenosulfónicoRESUMEN
The receptor for urokinase-type plasminogen activator (uPAR) (CD87) plays an important role in leukocyte adhesion and migration. To assess the effect of endotoxin on cellular uPAR, uPAR expression was determined on leukocytes by fluorescence-activated cell sorter analysis in seven healthy subjects following intravenous injection of endotoxin (lot G; 4 ng/kg). Endotoxin induced a transient increase in uPAR expression on monocytes, reaching a 92% +/- 46% increase over baseline expression after 6 h (P < 0.05). Endotoxin did not influence uPAR expression on granulocytes, while uPAR remained undetectable on lymphocytes. Endotoxin also increased soluble uPAR levels in plasma (P < 0.05). Stimulation of human whole blood with endotoxin or gram-positive stimuli in vitro also resulted in an upregulation of monocyte uPAR expression. Although tumor necrosis factor alpha (TNF) upregulated monocyte uPAR expression, anti-TNF did not influence the endotoxin-induced increase in monocyte uPAR expression. These data suggest that infectious stimuli may influence monocyte function in vivo by enhancing the expression of uPAR.
Asunto(s)
Endotoxemia/enzimología , Precursores Enzimáticos/sangre , Monocitos/enzimología , Receptores de Superficie Celular/sangre , Regulación hacia Arriba , Adulto , Recuento de Células Sanguíneas , Relación Dosis-Respuesta a Droga , Endotoxemia/inducido químicamente , Citometría de Flujo , Humanos , Inyecciones Intravenosas , Leucocitos/enzimología , Lipopolisacáridos/administración & dosificación , Lipopolisacáridos/farmacología , Masculino , Receptores del Activador de Plasminógeno Tipo Uroquinasa , Factores de Tiempo , Factor de Necrosis Tumoral alfa/fisiologíaRESUMEN
Interleukin (IL)-12 is considered a central regulator of host resistance against a variety of pathogens. Therefore, IL-12 has been advocated as a potential therapeutic agent in infections. To determine the in vivo effects of IL-12 on mononuclear cells involved in the host immune response, four chimpanzees received an intravenous injection of recombinant IL-12 (1 microgram/kg). IL-12 induced a sustained decrease in lymphocyte counts, with decreases in CD3+/CD4+ and CD3+/CD8+ cells, while monocyte counts showed a transient increase. IL-12 injection resulted in a shift toward a Th1-mediated immune response as indicated by increased interferon-gamma production during whole-blood stimulation, while not influencing IL-4 production. IL-12-induced activation of NK cells and phagocytes, as indicated by increased NK cell cytotoxicity and increased plasma levels of granzymes A and B and of chitotriosidase activity. These data support the hypothesis that IL-12 may serve as a useful therapeutic agent in infections where a cell-mediated response is protective.
Asunto(s)
Interleucina-12/farmacología , Leucocitos Mononucleares/efectos de los fármacos , Pan troglodytes/inmunología , Animales , Biomarcadores , Citotoxicidad Inmunológica , Granzimas , Hexosaminidasas/sangre , Células Asesinas Naturales , Recuento de Leucocitos , Recuento de Linfocitos , Monocitos/citología , Fagocitos , Serina Endopeptidasas/sangreRESUMEN
Tumor necrosis factor-alpha (TNF-alpha) is released from the cell surface by cleavage of pro-TNF-alpha by metalloproteinases (MPs). In cell cultures, inhibition of MPs has been found not only to reduce the release of TNF-alpha, but also to enhance the surface expression of TNF-alpha and TNF-alpha receptors, which might lead to a proinflammatory effect. To determine the effect of MP inhibition during inflammation in humans, 7 healthy subjects were studied after intravenous injection of lipopolysaccharide (LPS; 4 ng/kg) preceded (-20 minutes) by an oral dose of the MP inhibitor GI5402 (100 mg) or matching placebo. GI5402 strongly reduced LPS-induced TNF-alpha release (P <.001), but did not influence the increase in monocyte-bound TNF-alpha. In addition, GI5402 attenuated the rise in plasma-soluble TNF-alpha receptors types I and II after LPS injection (both P <.001), but did not change the LPS-induced decreases in granulocyte and monocyte TNF-alpha receptor expression. These data suggest that MP inhibitors may be useful as a treatment modality in diseases in which excessive production of TNF-alpha is considered to play an important role. Furthermore, unlike in vitro, no evidence has been found in vivo with MP inhibition for a potential proinflammatory effect due to increases in membrane-bound TNF-alpha and TNF-alpha receptor number.
Asunto(s)
Endotoxemia/tratamiento farmacológico , Inhibidores de la Metaloproteinasa de la Matriz , Receptores del Factor de Necrosis Tumoral/biosíntesis , Factor de Necrosis Tumoral alfa/biosíntesis , Adulto , Estudios Cruzados , Método Doble Ciego , Endotoxemia/inducido químicamente , Endotoxemia/inmunología , Granulocitos/efectos de los fármacos , Granulocitos/inmunología , Humanos , Cinética , Lipopolisacáridos/toxicidad , Masculino , Monocitos/efectos de los fármacos , Monocitos/inmunologíaRESUMEN
BACKGROUND/AIMS: There is an urgent need for an effective bioartificial liver system to bridge patients with fulminant hepatic failure to liver transplantation or to regeneration of their own liver. Recently, we proposed a bioreactor with a novel design for use as a bioartificial liver (BAL). The reactor comprises a spirally wound nonwoven polyester fabric in which hepatocytes are cultured (40 x 10(6) cells/ml) as small aggregates and homogeneously distributed oxygenation tubing for decentralized oxygen supply and CO2 removal. The aims of this study were to evaluate the treatment efficacy of our original porcine hepatocyte-based BAL in rats with fulminant hepatic failure due to liver ischemia (LIS) and to monitor the viability of the porcine hepatocytes in the bioreactor during treatment. The latter aim is novel and was accomplished by applying a new species-specific enzyme immunoassay (EIA) for the determination of porcine alpha-glutathione S-transferase (alpha-GST), a marker for hepatocellular damage. METHODS: Three experimental groups were studied: the first control group (LIS Control, n = 13) received a glucose infusion only; a second control group (LIS No-Cell-BAL, n = 8) received BAL treatment without cells; and the treated group (LIS Cell-BAL, n = 8) was connected to our BAL which had been seeded with 4.4 x 10(8) viable primary porcine hepatocytes. RESULTS/CONCLUSIONS: In contrast to previous comparable studies, BAL treatment significantly improved survival time in recipients with LIS. In addition, the onset of hepatic encephalopathy was significantly delayed and the mean arterial blood pressure significantly improved. Significantly lower levels of ammonia and lactate in the LIS Cell-BAL group indicated that the porcine hepatocytes in the bioreactor were metabolically activity. Low pig alpha-GST levels suggested that our bioreactor was capable of maintaining hepatocyte viability during treatment. These results provide a rationale for a comparable study in LIS-pigs as a next step towards potential clinical application.
Asunto(s)
Glutatión Transferasa/análisis , Isquemia/cirugía , Circulación Hepática , Hígado Artificial/normas , Animales , Diseño de Equipo , Estudios de Evaluación como Asunto , Técnicas para Inmunoenzimas/métodos , Isomerismo , Circulación Hepática/fisiología , Masculino , Ratas , Ratas Wistar , Especificidad de la Especie , PorcinosRESUMEN
To determine in vivo effects of interleukin (IL)-12 on host inflammatory mediator systems, 4 healthy chimpanzees received recombinant human IL-12 (1 microg/kg) by intravenous injection. IL-12 induced increases in plasma concentrations of IL-15, IL-18, and interferon-gamma (IFN-gamma), plus a marked antiinflammatory cytokine response (IL-10, soluble tumor necrosis factor [TNF] receptors, IL-1 receptor antagonist) and secretion of alpha-chemokines (IL-8, IFN-gamma-inducible protein 10) and beta-chemokines (monocyte chemoattractant protein-1, macrophage inflammatory protein-1beta). In addition, IL-12 elicited neutrophilic leukocytosis, neutrophil degranulation (elastase-alpha1-antitrypsin complexes), coagulation activation (F1 + 2 prothrombin fragment, thrombin-antithrombin III complexes), and fibrinolytic activation (tissue-type plasminogen activator, plasmin-alpha2-antiplasmin complexes). IL-12-induced activation of multiple host mediator systems was found only after 8-24 h, remained detectable until the end of the 48-h observation period, and occurred in the absence of detectable TNF and IL-1beta. These data may contribute to understanding the role of IL-12 in the pathogenesis of sepsis syndrome and the toxicity found after repeated injections of IL-12.
Asunto(s)
Quimiocinas/sangre , Citocinas/sangre , Interleucina-12/farmacología , Animales , Factores de Coagulación Sanguínea/metabolismo , Quimiocina CCL2/sangre , Quimiocina CCL4 , Quimiocinas/biosíntesis , Citocinas/biosíntesis , Relación Dosis-Respuesta a Droga , Fibrinólisis , Humanos , Inflamación/inmunología , Inyecciones Intravenosas , Interferón gamma/sangre , Interleucina-12/administración & dosificación , Interleucina-12/sangre , Interleucina-15/sangre , Interleucina-18/sangre , Proteínas Inflamatorias de Macrófagos/sangre , Neutrófilos/fisiología , Pan troglodytes , Proteínas Recombinantes/administración & dosificación , Proteínas Recombinantes/farmacología , Factores de Tiempo , Activador de Tejido Plasminógeno/sangreRESUMEN
Experimental data indicate that mucosal CD4+ T cells play an important role in the pathogenesis of inflammatory bowel disease (IBD). Based on the pattern of cytokine production, CD4+ T cells may be distinguished into two different phenotypes. Th1 responses are characterized by secretion of interleukin (IL)-2, tumor necrosis factor (TNF)-alpha, lymphotoxin, and interferon (IFN)-gamma and are associated with delayed-type hypersensitivity reactions, whereas Th2 responses, which are characterized by secretion of IL-4, IL-5, and IL-10, have been associated with humoral immune responses and allergy. To assess the number of IFN-alpha and IL-4 positive cells in IBD and normal intestinal specimens, frozen sections from intestinal specimens from 10 Crohn's disease (CD), 8 ulcerative colitis (UC), and 8 healthy controls were examined by immunohistochemistry. Monoclonal antibodies for CD3, CD8, IFN-gamma, and IL-4 were used. T-lymphocyte infiltration and cytokine expression by epithelial, lamina propria, and submucosal cells were scored on a four-point scale by two independent observers who were blinded for the clinical data. One-way analysis of variance (ANOVA) testing was used for statistical analysis. In intestinal specimens from IBD patients, the number of CD3+ cells was found increased in the lamina propria and, within the submucosa, this increase was significant (p < 0.001). In CD the number of lamina propria IFN-gamma positive cells was significantly increased as compared with controls (p < 0.002). In UC the number of both IFN-gamma and IL-4 producing cells in the lamina propria was not significantly increased as compared with controls. The present results confirm the existence of a Th1-biased pattern production in CD but not in UC.
Asunto(s)
Colitis Ulcerosa/patología , Enfermedad de Crohn/patología , Interferón gamma/análisis , Interleucina-4/análisis , Mucosa Intestinal/química , Adolescente , Adulto , Análisis de Varianza , Antígenos CD/análisis , Colectomía , Colitis Ulcerosa/inmunología , Colitis Ulcerosa/cirugía , Enfermedad de Crohn/inmunología , Enfermedad de Crohn/cirugía , Técnicas de Cultivo , Femenino , Humanos , Inmunohistoquímica , Mucosa Intestinal/patología , Recuento de Linfocitos , Masculino , Persona de Mediana Edad , Valores de Referencia , Linfocitos T/química , Linfocitos T/citologíaRESUMEN
Extracorporeal bioartificial liver (BAL) systems based on hepatocytes need to be flushed before clinical application, as hepatocyte culture media are not approved for medical use. Commercially available 0.9% NaCl solution and hemofiltration solution (both supplemented with 10% human albumin) were investigated in vitro to test their potential to wash BAL systems with minimal stress for the cultured hepatocytes. After a 2 hour incubation, the lidocaine metabolising capacity and release of liver enzymes were assessed. As hepatocytes have been cultured in bioreactors in either two or three dimensional cell configurations, we tested the media in respectively hepatocyte monolayers cultures and in our newly developed bioreactor in which hepatocytes reorganise as small hepatocyte aggregates. The three dimensional hepatocyte cultures tolerated both media well, and no significant differences were seen compared with hepatocytes cultured in Williams' E (reference hepatocyte culture medium). The two dimensional hepatocyte cultures tolerated the supplemented hemofiltration solution and the reference medium equally well, but the condition of the porcine hepatocytes monolayer cultures was significantly impaired when incubated with the supplemented physiological saline solution. In conclusion, as a supplemented physiological saline solution may have detrimental effects on the condition of the hepatocytes, the more complex hemofiltration solution (bicarbonate buffered, glucose, essential minerals) was considered the better alternative for flushing bioartificial liver systems.
Asunto(s)
Medios de Cultivo/farmacología , Hígado/citología , Animales , Órganos Artificiales , Aspartato Aminotransferasas/metabolismo , Células Cultivadas , Cromatografía Líquida de Alta Presión , L-Lactato Deshidrogenasa/metabolismo , Lidocaína/análogos & derivados , Lidocaína/metabolismo , Hígado/enzimología , Masculino , Técnicas de Cultivo de Órganos/métodos , PorcinosRESUMEN
The purpose of this study was to investigate whether the efficacy of our novel extracorporeal bioartificial liver (BAL) to support rats with complete liver ischemia (LIS) could be improved by extending the culture time of freshly isolated porcine hepatocytes from 14 hours to 38 hours. The results showed that survival as well as porcine hepatocyte integrity improved, the onset of coma delayed, and the ammonia levels decreased in LIS rats of the 38 hour group compared to the 14 hour group, but no statistically significant differences were observed. In the 38 hour group, but not the 14 hour group, the onset of hepatic encephalopathy was significantly delayed and ammonia metabolism significantly improved compared to the LIS rats in control groups that only received a glucose infusion or were connected to a BAL without cells. In conclusion, prolonged hepatocyte recovery favoured all investigated parameters, although not all observed effects were statistically significant. More research is required to find out how long primary hepatocytes should be cultured in a bioreactor for optimal BAL support.