RESUMEN
Thiopurines (mercaptopurine, azathioprine and thioguanine) are well-established maintenance treatments for a wide range of diseases such as leukemia, inflammatory bowel disease (IBD), systemic lupus erythematosus (SLE) and other inflammatory and autoimmune diseases in general. Worldwide, millions of patients are treated with thiopurines. The use of thiopurines has been limited because of off-target effects such as myelotoxicity and hepatotoxicity. Therefore, seeking methods to enhance target-based thiopurine-based treatment is relevant, combined with pharmacogenetic testing. Controlled-release formulations for thiopurines have been clinically tested and have shown promising outcomes in inflammatory bowel disease. Latest developments in nano-formulations for thiopurines have shown encouraging pre-clinical results, but further research and development are needed. This review provides an overview of novel drug delivery strategies for thiopurines, reviewing modified release formulations and with a focus on nano-based formulations.
Asunto(s)
Purinas/química , Animales , Química Farmacéutica/métodos , Sistemas de Liberación de Medicamentos/métodos , Humanos , Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Purinas/farmacologíaRESUMEN
Thiopurine drugs continue to be a cornerstone of inflammatory bowel disease (IBD) treatment. Thiopurines are economical compared with many newer medical treatments for IBD, other chronic inflammatory diseases and leukaemia, although they are not without their shortcomings. These include a slow-onset therapeutic action and many adverse drug reactions. This feature article surveys published data, unpublished in vitro and in vivo experiments, as well as clinical experience, underpinning a rationale for bringing a novel thiopurine drug formulation to market. This formulation has a rapid action making it suitable for the induction and maintenance treatment of IBD and avoids most thiopurine-associated adverse reactions.
Asunto(s)
Inmunosupresores/administración & dosificación , Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Tioguanina/administración & dosificación , Administración Rectal , Animales , Enfermedad Hepática Inducida por Sustancias y Drogas , Humanos , Inmunosupresores/efectos adversos , Mucosa Intestinal/metabolismo , Tioguanina/efectos adversosRESUMEN
Thiopurines are anti-inflammatory prodrugs. We hypothesised that bacteria may contribute to conversion to active drug. Escherichia coli strain DH5α was evaluated to determine whether it could metabolise the thiopurine drugs, thioguanine or mercaptopurine, to thioguanine nucleotides. A rapid and reliable high performance liquid chromatography (ultraviolet detection) method was developed to quantify indirectly thioguanine nucleotides, by measuring thioguanine nucleoside.
Asunto(s)
Antiinflamatorios/metabolismo , Cromatografía Líquida de Alta Presión , Tioguanina/metabolismo , Calibración , Escherichia coli/metabolismo , Mercaptopurina/metabolismo , Nucleósidos/análisis , Nucleótidos/análisisRESUMEN
BACKGROUND AND AIM: The etiology of Crohn's disease (CD) implicates both genetic and environmental factors. Smoking behavior is one environmental risk factor to play a role in the development of CD. The study aimed to assess the contribution of the interleukin 23 receptor (IL23R) in determining disease susceptibility in two independent cohorts of CD, and to investigate the interactions between IL23R variants, smoking behavior, and CD-associated genes, NOD2 and ATG16L1. METHODS: Ten IL23R single-nucleotide polymorphisms (SNPs) were genotyped in 675 CD cases, and 1255 controls from Brisbane, Australia (dataset 1). Six of these SNPs were genotyped in 318 CD cases and 533 controls from Canterbury, New Zealand (dataset 2). Case-control analysis of genotype and allele frequencies, and haplotype analysis for all SNPs was conducted. RESULTS: We demonstrate a strong increased CD risk for smokers in both datasets (odds ratio 3.77, 95% confidence interval 2.88-4.94), and an additive interaction between IL23Râ SNPs and cigarette smoking. Ileal involvement was a consistent marker of strong SNP-CD association (P ≤ 0.001), while the lowest minor allele frequencies for location were found in those with colonic CD (L2). Three haplotype blocks were identified across the 10 IL23Râ SNPs conferring different risk of CD. Haplotypes conferred no further risk of CD when compared with single SNP analyses. CONCLUSION: IL23R gene variants determine CD susceptibility in the Australian and New Zealand population, particularly ileal CD. A strong additive interaction exists between IL23Râ SNPs and smoking behavior resulting in a dramatic increase in disease risk depending upon specific genetic background.
Asunto(s)
Enfermedad de Crohn/etiología , Enfermedad de Crohn/genética , Receptores de Interleucina/genética , Fumar/efectos adversos , Adolescente , Adulto , Australia , Estudios de Casos y Controles , Estudios de Cohortes , Femenino , Predisposición Genética a la Enfermedad/genética , Genotipo , Humanos , Masculino , Nueva Zelanda , Polimorfismo de Nucleótido Simple/genética , Factores de Riesgo , Adulto JovenRESUMEN
BACKGROUND: Inosine triphosphate pyrophosphohydrolase (ITPase) is a 'house-cleaning' enzyme that degrades non-canonical ('rogue') nucleotides. Complete deficiency is fatal in knockout mice, but a mutant polymorphism resulting in low enzyme activity with an accumulation of ITP and other non-canonical nucleotides, appears benign in humans. We hypothesised that reduced ITPase activity may cause acquired mitochondrial DNA (mtDNA) defects. Furthermore, we investigated whether accumulating mtDNA defects may then be a risk factor for cell transformation, in adult haematological malignancy (AHM). METHODS: DNA was extracted from peripheral blood and bone marrow samples. Microarray-based sequencing of mtDNA was performed on 13 AHM patients confirmed as carrying the ITPA 94C>A mutation causing low ITPase activity, and 4 AHM patients with wildtype ITPA. The frequencies of ITPA 94C>A and IVS2+21A>C polymorphisms were studied from 85 available AHM patients. RESULTS: ITPA 94C>A was associated with a significant increase in total heteroplasmic/homoplasmic mtDNA mutations (p<0.009) compared with wildtype ITPA, following exclusion of haplogroup variants. This suggested that low ITPase activity may induce mitochondrial abnormalities. Compared to the normal population, frequencies for the 94C>A and IVS2+21A>C mutant alleles among the AHM patients were higher for myelodyplastic syndrome (MDS) - but below significance; were approximately equivalent for chronic lymphoblastic leukemia; and were lower for acute myeloid leukemia. CONCLUSIONS: This study invokes a new paradigm for the evolution of MDS, where nucleotide imbalances produced by defects in 'house-cleaning' genes may induce mitochondrial dysfunction, compromising cell integrity. It supports recent studies which point towards an important role for ITPase in cellular surveillance of rogue nucleotides.
Asunto(s)
ADN Mitocondrial/genética , Neoplasias Hematológicas/enzimología , Neoplasias Hematológicas/genética , Pirofosfatasas/genética , Adulto , Anciano , Anciano de 80 o más Años , Animales , Humanos , Ratones , Ratones Noqueados , Análisis por Micromatrices , Persona de Mediana Edad , Polimorfismo GenéticoRESUMEN
OBJECTIVE: The anti-leukemic drugs, azathioprine and 6-mercaptopurine (6MP), are important in the treatment of inflammatory bowel disease but an alternative faster-acting, less-allergenic thiopurine, 6-thioguanine (6TG), can cause hepatic veno-occlusive disease/sinusoidal obstructive syndrome (SOS). Understanding of SOS has been hindered by inability to ethically perform serial liver biopsies on patients and the lack of an animal model. DESIGN: Normal and C57Bl/6 mice with specific genes altered to elucidate mechanisms responsible for 6TG-SOS, were gavaged daily for upto 28d with 6TG, 6MP or methylated metabolites. Animal survival was monitored and at sacrifice a histological score of SOS, haematology and liver biochemistry were measured. RESULTS: Only 6TG caused SOS, which was dose related. 6TG and to a lesser extent 6MP but not methylated metabolites were associated with dose-dependent haematopoietic toxicity. SOS was not detected with non-lethal doses of 6TG. SOS did not occur in hypoxanthine-phosphoribosyl transferase-deficient C57Bl/6 mice, demonstrating that 6TG-SOS requires thioguanine nucleotides. Hepatic inflammation was characteristic of SOS, and C57Bl/6 mice deficient in P- and E-selectins on the surface of vascular endothelial cells showed markedly reduced SOS, demonstrating a major role for leukocytes recruited from blood. Split dosing of 6TG markedly attenuated SOS but still effected immunosuppression and prevented spontaneous colitis in Winnie mice, which have a single nucleotide polymorphism mutation in Muc2. CONCLUSION: This novel model provides clinically relevant insights into how 6TG induces SOS, and how this dangerous adverse drug reaction may be avoided by either inhibition of endothelial activation or simple changes to dosing regimens of 6TG, while still being effective treatment for colitis.
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Modelos Animales de Enfermedad , Enfermedad Veno-Oclusiva Hepática/inducido químicamente , Tioguanina/toxicidad , Animales , Relación Dosis-Respuesta a Droga , Enfermedad Veno-Oclusiva Hepática/prevención & control , Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Ratones , Ratones Endogámicos C57BL , Estadísticas no Paramétricas , Análisis de Supervivencia , Tioguanina/administración & dosificaciónRESUMEN
The gastrointestinal tract is protected by a mucus barrier with both secreted and cell-surface mucins contributing to the exclusion of luminal microbes and toxins. Alterations in the structure and/or quantity of mucins alter the barrier function of mucus and could play roles in initiating and maintaining mucosal inflammation in inflammatory bowel diseases (IBD), and in driving cancer development in the intestine. The aim of this review is to focus on the roles of the mucins in IBD. The polymorphisms of mucin genes that have been associated with susceptibility to IBD, and alterations in mucin expression as well as factors that regulate production of the mucins in IBD, are summarized. Data from animal models of intestinal inflammation, which support the importance of mucins in IBD and cancer development, are also discussed.
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Neoplasias Colorrectales/metabolismo , Enfermedades Inflamatorias del Intestino/metabolismo , Mucosa Intestinal/metabolismo , Mucinas/metabolismo , Animales , Transformación Celular Neoplásica/metabolismo , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/inmunología , Neoplasias Colorrectales/microbiología , Predisposición Genética a la Enfermedad , Humanos , Enfermedades Inflamatorias del Intestino/genética , Enfermedades Inflamatorias del Intestino/inmunología , Enfermedades Inflamatorias del Intestino/microbiología , Mucosa Intestinal/inmunología , Mucosa Intestinal/microbiología , Modelos Animales , Mucinas/genética , Fenotipo , Polimorfismo GenéticoRESUMEN
BACKGROUND AND AIMS: The MUC13 transmembrane mucin is highly and constitutively expressed in the small and large intestine. Although MUC13 polymorphisms have been associated with human inflammatory bowel diseases and susceptibility to Escherichia coli infection in pigs, the biological functions of MUC13 are unknown. This study aimed to explore whether MUC13 modulates intestinal inflammation. METHODS: Muc13(-/-) mice were generated, phenotyped and challenged with the colitis-inducing agent, dextran sodium sulphate (DSS). Colitis was assessed by clinical symptoms and intestinal histopathology. Intestinal epithelial cell apoptosis and proliferation, macrophage infiltration and cytokine production were also quantified. Apoptosis of human LS513 intestinal epithelial cells in response to apoptotic agents, including DSS, was also measured, following knockdown of MUC13 with siRNA. RESULTS: Muc13(-/-) mice were viable, fertile and developed normally, with no spontaneous intestinal pathology except mild focal neutrophilic inflammation in the small and large intestines of old mice. In response to DSS challenge, Muc13(-/-) mice developed more severe acute colitis, as reflected by increased weight loss, rectal bleeding, diarrhoea and histological colitis scores compared with wild-type mice. Increased numbers of F4/80(+) macrophages in inflamed mucosa of Muc13(-/-) mice were accompanied by increased expression of intestinal IL-1ß and TNFα mRNA. Muc13(-/-) mice had significantly increased intestinal epithelial cell apoptosis within 3 days of DSS exposure. LS513 cells were more susceptible to DSS, actinomycin-D, ultraviolet irradiation and TRAIL-induced apoptosis when MUC13 was knocked down by siRNA. CONCLUSIONS: These novel findings indicate a protective role for Muc13 in the colonic epithelium by inhibiting toxin-induced apoptosis and have important implications for intestinal infections, inflammatory diseases and the development of intestinal cancer.
Asunto(s)
Antígenos de Superficie/fisiología , Apoptosis/fisiología , Colitis/fisiopatología , Factor de Crecimiento Epidérmico/fisiología , Mucosa Intestinal/fisiopatología , Mucinas/fisiología , Animales , Antígenos de Superficie/metabolismo , Línea Celular , Proliferación Celular , Colitis/metabolismo , Colitis/patología , Citocinas/fisiología , Sulfato de Dextran/farmacología , Factor de Crecimiento Epidérmico/metabolismo , Humanos , Enfermedades Inflamatorias del Intestino/metabolismo , Enfermedades Inflamatorias del Intestino/fisiopatología , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patología , Macrófagos Peritoneales/fisiología , Ratones , Ratones Noqueados , Mucinas/metabolismo , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
BACKGROUND: Thiopurines, such as azathioprine and mercaptopurine, are of pivotal importance in the treatment of inflammatory bowel disease. Although these drugs have been used for several decades, still many questions remain unanswered. AIM: To provide an overview of clinically and scientifically challenging topics concerning thiopurine therapy in inflammatory bowel disease treatment. METHODS: The first meeting of the Thiopurine Task Force Interest Group was held during the 2009 United European Gastroenterology Week in London (GASTRO2009). The topics of this meeting were of particular clinical and scientific interest. Additional literature was identified by performing a Pubmed search using the search terms 'inflammatory bowel disease', 'azathioprine', '6-mercaptopurine' and 'thioguanine'. RESULTS: The following topics were discussed: therapeutic drug monitoring; the synergy of thiopurines with aminosalicylates and allopurinol; serious adverse events such as opportunistic infections, hepatotoxicity, carcinogenicity and pancreatitis; prolongation of thiopurines during clinical remission; indications for thiopurines in the postoperative setting; and the potential use of thioguanine. Specific interesting and clinically relevant topics for potential future research are provided. CONCLUSIONS: Thiopurines remain central to inflammatory bowel disease treatment, although future studies are required to substantiate a more personalised medicine approach to their use.
Asunto(s)
Inmunosupresores/efectos adversos , Inmunosupresores/uso terapéutico , Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Purinas/efectos adversos , Purinas/uso terapéutico , Ácidos Aminosalicílicos/uso terapéutico , Quimioterapia Combinada , Humanos , Inmunosupresores/metabolismo , Purinas/metabolismoRESUMEN
BACKGROUND: Inflammatory dermatoses in conjunction with inflammatory bowel disease (IBD) comprise a diverse range of disorders. Some but not all of these respond to conventional treatments for the underlying IBD, such as immunomodulating or antibiotic treatments. We describe our experience with high dose vitamin B12, where conventional therapies have failed. CASE REPORTS: The first case had pouchitis complicated by perianal abscesses and a recto-vaginal fistula. The second case had biopsy proven hidradenitis suppurativa affecting the perianal, inguinal and pubic skin. High dose vitamin B12 appeared to be the major factor in preventing the recurrence of suppuration in both patients. Neither patient had vitamin B12 deficiency. Open label experience: high dose vitamin B12 treatment of a further 10 consecutive IBD patients with dermatoses was thought to provide benefit to six of them, but did not appear useful in four patients with perianal Crohn's disease with fistulae as the only manifestation of cutaneous disease. CONCLUSIONS: There appears to be a subset of IBD patients with perianal and more distant inflammatory dermatoses, who benefit from high dose vitamin B12 treatment. Clinical trials in IBD patients with biopsy-characterised suppurative dermatoses will be required in order to properly define the role of this safe and economical therapy.
Asunto(s)
Enfermedades Inflamatorias del Intestino/complicaciones , Enfermedades de la Piel/complicaciones , Deficiencia de Vitamina B 12/complicaciones , Vitamina B 12/uso terapéutico , Complejo Vitamínico B/uso terapéutico , Adolescente , Adulto , Femenino , Hidradenitis Supurativa/complicaciones , Hidradenitis Supurativa/tratamiento farmacológico , Humanos , Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Masculino , Persona de Mediana Edad , Enfermedades de la Piel/tratamiento farmacológico , Supuración/complicaciones , Vitamina B 12/administración & dosificación , Deficiencia de Vitamina B 12/tratamiento farmacológico , Complejo Vitamínico B/administración & dosificación , Adulto JovenRESUMEN
BACKGROUND AND AIM: Anti-tumor-necrosis-factor-alpha (anti-TNF-α) medications are effective in inflammatory bowel disease (IBD), but have an increased risk of tuberculosis (TB) and serious infections. The aim of this study was to examine the Australian/New Zealand experience of serious infections and TB in IBD patients receiving anti-TNF-α therapy from 1999-2009. METHODS: Serious infections, defined as 'requiring hospital admission' and TB cases in patients receiving, or within 3 months following, anti-TNF-α therapy were analyzed across Australia and New Zealand. Patient demographics, IBD medications, duration of anti-TNF-α therapy, and infection details were collected. RESULTS: A total of 5562 IBD patients were managed across the centers. Of these, 489 (16.8%) Crohn's disease and 137 (5.2%) ulcerative colitis patients received anti-TNF-α therapy. There were three cases of latent TB that received prophylaxis prior to anti-TNF-α therapy. No cases of active TB were reported. Fourteen (2.2%) serious infections occurred. Seven occurred in patients receiving anti-TNF-α therapy for less than 6 months, including two cases of primary Varicella zoster (VZV), two cases of Pneumocystis jiroveci pneumonia, two cases of Staphylococcus aureus bacteremia, and one severe flu-like illness. Six patients were taking additional immunosuppressive medications. The other seven infections occurred after 6 months (mean 32.6 ± 24.3 months) and included one case of primary VZV, one flu-like illness, and five bacterial infections. All infections resolved with treatment. CONCLUSION: TB is a very rare complication of anti-TNF-α therapy in Australia and New Zealand. Serious infections are uncommon but early opportunistic infections with Pneumocystis jiroveci pneumonia suggest a need for vigilance in patients on multiple immunosuppressive medications. VZV vaccination prior to immunosuppressive therapy should be considered in VZV-naïve patients.
Asunto(s)
Antiinflamatorios/efectos adversos , Anticuerpos Monoclonales/efectos adversos , Colitis Ulcerosa/tratamiento farmacológico , Enfermedad de Crohn/tratamiento farmacológico , Infecciones Oportunistas/epidemiología , Factor de Necrosis Tumoral alfa/antagonistas & inhibidores , Adalimumab , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Análisis de Varianza , Antiinflamatorios/inmunología , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales Humanizados , Australia/epidemiología , Infecciones Bacterianas/epidemiología , Infecciones Bacterianas/microbiología , Distribución de Chi-Cuadrado , Niño , Preescolar , Femenino , Herpes Zóster/epidemiología , Humanos , Huésped Inmunocomprometido , Lactante , Infliximab , Masculino , Persona de Mediana Edad , Nueva Zelanda/epidemiología , Tuberculosis/epidemiología , Adulto JovenRESUMEN
OBJECTIVES: Mucosa-associated bacteria are increased in inflammatory bowel disease (IBD), which suggests the possibility of an increased source of digestible endogenous mucus substrate. We hypothesized that mucolytic bacteria are increased in IBD, providing increased substrate to sustain nonmucolytic mucosa-associated bacteria. METHODS: Mucolytic bacteria were characterized by the ability to degrade human secretory mucin (MUC2) in pure and mixed anaerobic cultures. Real-time PCR was used to enumerate mucosa-associated mucolytic bacteria in 46 IBD and 20 control patients. Bacterial mucolytic activity was tested in vitro using purified human MUC2. RESULTS: We confirm increased total mucosa-associated bacteria 16S rRNA gene in macroscopically and histologically normal intestinal epithelium of both Crohn's disease (CD) (mean 1.9-fold) and ulcerative colitis (UC) (mean 1.3-fold). We found a disproportionate increase in some mucolytic bacteria. Mean Ruminococcus gnavus were increased >4-fold and Ruminococcus torques â¼100-fold in macroscopically and histologically normal intestinal epithelium of both CD and UC. The most abundantly detected mucolytic bacterium in controls, Akkermansia muciniphila, was reduced many fold in CD and in UC. Coculture of A. muciniphila with MUC2 as the sole carbon source led to reduction in its abundance while it augmented growth of other bacteria. CONCLUSIONS: Mucolytic bacteria are present in healthy humans, where they are an integral part of the mucosa-associated bacterial consortium. The disproportionate increase in R. gnavus and R. torques could explain increased total mucosa-associated bacteria in IBD.
Asunto(s)
Colon/microbiología , Enfermedades Inflamatorias del Intestino/microbiología , Mucosa Intestinal/microbiología , Mucinas/genética , Ruminococcus/genética , Adulto , Anciano , Anciano de 80 o más Años , Colon/metabolismo , Colon/patología , Femenino , Humanos , Enfermedades Inflamatorias del Intestino/genética , Enfermedades Inflamatorias del Intestino/patología , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patología , Masculino , Persona de Mediana Edad , Mucinas/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Ruminococcus/metabolismoRESUMEN
OBJECTIVES: Crohn's disease (CD; MIM 266600) is one of the most common forms of inflammatory bowel disease (IBD), and represents a significant burden to health care in developed countries. Our aim was to determine whether a gene in the IBD linkage region on chromosome 19q13, with a role in Paneth cell secretion and T-cell activation, conferred genetic susceptibility to the development of CD. METHODS: In total, 792 CD cases and 1,244 controls of Australian origin (Caucasian) were genotyped for seven single-nucleotide polymorphisms (SNPs) in the gene encoding the intermediate conductance calcium-activated potassium channel protein (KCNN4) at 19q13.2. CD cases were phenotyped using the Montreal classification. The replication set comprised an additional 326 CD cases and 951 population-based Caucasian controls. Analysis of the KCNN4 mRNA transcript was carried out using quantitative reverse transcriptase-PCR. RESULTS: KCNN4 SNP rs2306801 was associated with CD (primary P=0.0008, odds ratio (OR) (95% confidence interval (CI)): 0.76 (0.65-0.89); replication P=0.01, OR (95% CI): 0.77 (0.61-0.97). Stratification by disease location identified the association between SNP rs2306801 and ileal CD (P=0.01). Non-inflamed ileal mucosa from CD patients carrying any of the common disease-predisposing NOD2 variants (R702W, G908R, 1007fs) had significantly reduced levels of KCNN4 mRNA expression (P=0.001). KCNN4 protein expression was detected in Paneth cells, and in T cells in inflamed lamina propria. CONCLUSIONS: Our data implicate the role of KCNN4 in ileal CD. The dual roles of KCNN4 in Paneth cell secretion and T-cell activation and also its nature as a potassium channel make it an important and practical therapeutic target.
Asunto(s)
Enfermedad de Crohn/genética , Íleon/patología , Canales de Potasio de Conductancia Intermedia Activados por el Calcio/genética , Adulto , Alelos , Australia , Distribución de Chi-Cuadrado , Colon/patología , Enfermedad de Crohn/patología , Femenino , Frecuencia de los Genes , Estudios de Asociación Genética , Ligamiento Genético , Predisposición Genética a la Enfermedad , Genotipo , Haplotipos , Humanos , Inmunohistoquímica , Inflamación/genética , Inflamación/patología , Masculino , Persona de Mediana Edad , Nueva Zelanda , Polimorfismo de Nucleótido Simple , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Estadísticas no ParamétricasRESUMEN
The bacterium Helicobacter pylori can cause peptic ulcer disease, gastric adenocarcinoma and MALT lymphoma. The cell-surface mucin MUC1 is a large glycoprotein which is highly expressed on the mucosal surface and limits the density of H. pylori in a murine infection model. We now demonstrate that by using the BabA and SabA adhesins, H. pylori bind MUC1 isolated from human gastric cells and MUC1 shed into gastric juice. Both H. pylori carrying these adhesins, and beads coated with MUC1 antibodies, induced shedding of MUC1 from MKN7 human gastric epithelial cells, and shed MUC1 was found bound to H. pylori. Shedding of MUC1 from non-infected cells was not mediated by the known MUC1 sheddases ADAM17 and MMP-14. However, knockdown of MMP-14 partially affected MUC1 release early in infection, whereas ADAM17 had no effect. Thus, it is likely that shedding is mediated both by proteases and by disassociation of the non-covalent interaction between the alpha- and beta-subunits. H. pylori bound more readily to MUC1 depleted cells even when the bacteria lacked the BabA and SabA adhesins, showing that MUC1 inhibits attachment even when bacteria cannot bind to the mucin. Bacteria lacking both the BabA and SabA adhesins caused less apoptosis in MKN7 cells than wild-type bacteria, having a greater effect than deletion of the CagA pathogenicity gene. Deficiency of MUC1/Muc1 resulted in increased epithelial cell apoptosis, both in MKN7 cells in vitro, and in H. pylori infected mice. Thus, MUC1 protects the epithelium from non-MUC1 binding bacteria by inhibiting adhesion to the cell surface by steric hindrance, and from MUC1-binding bacteria by acting as a releasable decoy.
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Infecciones por Helicobacter/prevención & control , Helicobacter pylori/patogenicidad , Mucina-1/fisiología , Animales , Adhesión Bacteriana/fisiología , Células Epiteliales/fisiología , Mucosa Gástrica/microbiología , Glicosilación , Humanos , Ratones , Unión Proteica , Subunidades de Proteína , Estómago/microbiología , Estómago/fisiologíaAsunto(s)
Antiinflamatorios/efectos adversos , Anticuerpos Monoclonales/efectos adversos , Enfermedad de Crohn/tratamiento farmacológico , Linfoma de Células T/etiología , Femenino , Humanos , Huésped Inmunocomprometido , Infliximab , Linfoma de Células T/inmunología , Masculino , Mercaptopurina/efectos adversos , Adulto JovenRESUMEN
The etiology of human inflammatory bowel diseases (IBDs) is believed to involve inappropriate host responses to the complex commensal microbial flora in the gut, although an altered commensal flora is not completely excluded. A multifunctional cellular and secreted barrier separates the microbial flora from host tissues. Altered function of this barrier remains a major largely unexplored pathway to IBD. Although there is evidence of barrier dysfunction in IBD, it remains unclear whether this is a primary contributor to disease or a consequence of mucosal inflammation. Recent evidence from animal models demonstrating that genetic defects restricted to the epithelium can initiate intestinal inflammation in the presence of normal underlying immunity has refocused attention on epithelial dysfunction in IBD. We review the components of the secreted and cellular barrier, their regulation, including interactions with underlying innate and adaptive immunity, evidence from animal models of the barrier's role in preventing intestinal inflammation, and evidence of barrier dysfunction in both Crohn's disease and ulcerative colitis.
Asunto(s)
Traslocación Bacteriana/fisiología , Enfermedades Inflamatorias del Intestino/metabolismo , Mucosa Intestinal/microbiología , Animales , Humanos , Enfermedades Inflamatorias del Intestino/patología , PermeabilidadRESUMEN
BACKGROUND: Bacterial gastroenteritis causes morbidity and mortality in humans worldwide. Murine Citrobacter rodentium infection is a model for gastroenteritis caused by the human pathogens enteropathogenic Escherichia coli and enterohaemorrhagic E. coli. Mucin glycoproteins are the main component of the first barrier that bacteria encounter in the intestinal tract. METHODOLOGY/PRINCIPAL FINDINGS: Using Immunohistochemistry, we investigated intestinal expression of mucins (Alcian blue/PAS, Muc1, Muc2, Muc4, Muc5AC, Muc13 and Muc3/17) in healthy and C. rodentium infected mice. The majority of the C. rodentium infected mice developed systemic infection and colitis in the mid and distal colon by day 12. C. rodentium bound to the major secreted mucin, Muc2, in vitro, and high numbers of bacteria were found in secreted MUC2 in infected animals in vivo, indicating that mucins may limit bacterial access to the epithelial surface. In the small intestine, caecum and proximal colon, the mucin expression was similar in infected and non-infected animals. In the distal colonic epithelium, all secreted and cell surface mucins decreased with the exception of the Muc1 cell surface mucin which increased after infection (p<0.05). Similarly, during human infection Salmonella St Paul, Campylobacter jejuni and Clostridium difficile induced MUC1 in the colon. CONCLUSION: Major changes in both the cell-surface and secreted mucins occur in response to intestinal infection.
Asunto(s)
Infecciones por Enterobacteriaceae/metabolismo , Enfermedades Intestinales/metabolismo , Mucinas/metabolismo , Adulto , Anciano , Animales , Ciego/microbiología , Ciego/patología , Citrobacter rodentium/aislamiento & purificación , Citrobacter rodentium/fisiología , Infecciones por Enterobacteriaceae/complicaciones , Infecciones por Enterobacteriaceae/patología , Femenino , Humanos , Enfermedades Intestinales/etiología , Enfermedades Intestinales/patología , Intestino Grueso/microbiología , Intestino Grueso/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Persona de Mediana Edad , Distribución Tisular , Adulto JovenAsunto(s)
Azatioprina/administración & dosificación , Inmunosupresores/administración & dosificación , Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Administración Oral , Azatioprina/efectos adversos , Azatioprina/sangre , Biomarcadores Farmacológicos/sangre , Monitoreo de Drogas/métodos , Recuento de Eritrocitos , Humanos , Inmunosupresores/efectos adversos , Inmunosupresores/sangre , Enfermedades Inflamatorias del Intestino/sangre , Japón , Recuento de Leucocitos , Mercaptopurina/administración & dosificación , Metiltransferasas/genética , Metiltransferasas/metabolismo , Mutación , Inducción de Remisión , Tioguanina/sangre , Resultado del TratamientoRESUMEN
BACKGROUND: MUC2 mucin produced by intestinal goblet cells is the major component of the intestinal mucus barrier. The inflammatory bowel disease ulcerative colitis is characterized by depleted goblet cells and a reduced mucus layer, but the aetiology remains obscure. In this study we used random mutagenesis to produce two murine models of inflammatory bowel disease, characterised the basis and nature of the inflammation in these mice, and compared the pathology with human ulcerative colitis. METHODS AND FINDINGS: By murine N-ethyl-N-nitrosourea mutagenesis we identified two distinct noncomplementing missense mutations in Muc2 causing an ulcerative colitis-like phenotype. 100% of mice of both strains developed mild spontaneous distal intestinal inflammation by 6 wk (histological colitis scores versus wild-type mice, p < 0.01) and chronic diarrhoea. Monitoring over 300 mice of each strain demonstrated that 25% and 40% of each strain, respectively, developed severe clinical signs of colitis by age 1 y. Mutant mice showed aberrant Muc2 biosynthesis, less stored mucin in goblet cells, a diminished mucus barrier, and increased susceptibility to colitis induced by a luminal toxin. Enhanced local production of IL-1beta, TNF-alpha, and IFN-gamma was seen in the distal colon, and intestinal permeability increased 2-fold. The number of leukocytes within mesenteric lymph nodes increased 5-fold and leukocytes cultured in vitro produced more Th1 and Th2 cytokines (IFN-gamma, TNF-alpha, and IL-13). This pathology was accompanied by accumulation of the Muc2 precursor and ultrastructural and biochemical evidence of endoplasmic reticulum (ER) stress in goblet cells, activation of the unfolded protein response, and altered intestinal expression of genes involved in ER stress, inflammation, apoptosis, and wound repair. Expression of mutated Muc2 oligomerisation domains in vitro demonstrated that aberrant Muc2 oligomerisation underlies the ER stress. In human ulcerative colitis we demonstrate similar accumulation of nonglycosylated MUC2 precursor in goblet cells together with ultrastructural and biochemical evidence of ER stress even in noninflamed intestinal tissue. Although our study demonstrates that mucin misfolding and ER stress initiate colitis in mice, it does not ascertain the genetic or environmental drivers of ER stress in human colitis. CONCLUSIONS: Characterisation of the mouse models we created and comparison with human disease suggest that ER stress-related mucin depletion could be a fundamental component of the pathogenesis of human colitis and that clinical studies combining genetics, ER stress-related pathology and relevant environmental epidemiology are warranted.