Effect and mechanism of Bovis Calculus on ulcerative colitis by inhibiting IL-17/IL-17RA/Act1 signaling pathway / 中国中药杂志

This study aimed to elucidate the effect and underlying mechanism of Bovis Calculus in the treatment of ulcerative colitis(UC) through network pharmacological prediction and animal experimental verification. Databases such as BATMAN-TCM were used to mine the potential targets of Bovis Calculus against UC, and the pathway enrichment analysis was conducted. Seventy healthy C57BL/6J mice were randomly divided into a blank group, a model group, a solvent model(2% polysorbate 80) group, a salazosulfapyridine(SASP, 0.40 g·kg~(-1)) group, and high-, medium-, and low-dose Bovis Calculus Sativus(BCS, 0.20, 0.10, and 0.05 g·kg~(-1)) groups according to the body weight. The UC model was established in mice by drinking 3% dextran sulfate sodium(DSS) solution for 7 days. The mice in the groups with drug intervention received corresponding drugs for 3 days before modeling by gavage, and continued to take drugs for 7 days while modeling(continuous administration for 10 days). During the experiment, the body weight of mice was observed, and the disease activity index(DAI) score was recorded. After 7 days of modeling, the colon length was mea-sured, and the pathological changes in colon tissues were observed by hematoxylin-eosin(HE) staining. The levels of tumor necrosis factor-α(TNF-α), interleukin-1β(IL-1β), interleukin-6(IL-6), and interleukin-17(IL-17) in colon tissues of mice were detected by enzyme-linked immunosorbent assay(ELISA). The mRNA expression of IL-17, IL-17RA, Act1, TRAF2, TRAF5, TNF-α, IL-6, IL-1β, CXCL1, CXCL2, and CXCL10 was evaluated by real-time polymerase chain reaction(RT-PCR). The protein expression of IL-17, IL-17RA, Act1, p-p38 MAPK, and p-ERK1/2 was investigated by Western blot. The results of network pharmacological prediction showed that Bovis Calculus might play a therapeutic role through the IL-17 signaling pathway and the TNF signaling pathway. As revealed by the results of animal experiments, on the 10th day of drug administration, compared with the solvent model group, all the BCS groups showed significantly increased body weight, decreased DAI score, increased colon length, improved pathological damage of colon mucosa, and significantly inhibited expression of TNF-α,IL-6,IL-1β, and IL-17 in colon tissues. The high-dose BCS(0.20 g·kg~(-1)) could significantly reduce the mRNA expression levels of IL-17, Act1, TRAF2, TRAF5, TNF-α, IL-6, IL-1β, CXCL1, and CXCL2 in colon tissues of UC model mice, tend to down-regulate mRNA expression levels of IL-17RA and CXCL10, significantly inhibit the protein expression of IL-17RA,Act1,and p-ERK1/2, and tend to decrease the protein expression of IL-17 and p-p38 MAPK. This study, for the first time from the whole-organ-tissue-molecular level, reveals that BCS may reduce the expression of pro-inflammatory cytokines and chemokines by inhibiting the IL-17/IL-17RA/Act1 signaling pathway, thereby improving the inflammatory injury of colon tissues in DSS-induced UC mice and exerting the effect of clearing heat and removing toxins.

Mechanism of famous classical formula Huaihua Powder in treatment of ulcerative colitis based on metabonomics / 中国中药杂志

Ultra-high performance liquid chromatography-quadrupole-time of flight tandem mass spectrometry(UHPLC-Q-TOF-MS) was employed in this study to observe the effect of Huaihua Powder on the serum metabolites of mice with ulcerative colitis and reveal the mechanism of Huaihua Powder in the treatment of ulcerative colitis. The mouse model of ulcerative colitis was established by dextran sodium sulfate salt(DSS). The therapeutic effect of Huaihua Powder on ulcerative colitis was preliminarily evaluated based on the disease activity index(DAI), colon appearance, colon tissue morphology, and the content of inflammatory cytokines such as tumor necrosis factor-α(TNF-α), interleukin-6(IL-6), and interleukin-1β(IL-1β). UHPLC-Q-TOF-MS was employed to profile the endogenous metabolites of serum samples in blank control group, model group, and low-, medium-, and high-dose Huaihua Powder groups. Multivariate analyses such as principal component analysis(PCA), partial least squares discriminant analysis(PLS-DA), and orthogonal partial least squares discriminant analysis(OPLS-DA) were performed for pattern recognition. Potential biomarkers were screened by Mass Profiler Professional(MPP) B.14.00 with the thresholds of fold change≥2 and P<0.05. The metabolic pathways were enriched by MetaboAnalyst 5.0. The results showed that Huaihua Powder significantly improved the general state and colon tissue morphology of mice with ulcerative colitis, reduced DAI, and lowered the levels of TNF-α, IL-6, and IL-1β in serum. A total of 38 potential biomarkers were predicted to be related to the regulatory effect of Huaihua Powder, which were mainly involved in glycerophospholipid metabolism, glycine, serine, and threonine metabolism, mutual transformation of glucuronic acid, and glutathione metabolism. This study employed metabolomics to analyze the mechanism of Huaihua Powder in the treatment of ulcerative colitis, laying a foundation for the further research.

Spirulina platensis aqueous extracts ameliorate colonic mucosal damage and modulate gut microbiota disorder in mice with ulcerative colitis by inhibiting inflammation and oxidative stress / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Ulcerative colitis (UC) is a chronic and recurrent inflammatory bowel disease (IBD) that has become a major gastroenterologic problem during recent decades. Numerous complicating factors are involved in UC development such as oxidative stress, inflammation, and microbiota disorder. These factors exacerbate damage to the intestinal mucosal barrier. Spirulina platensis is a commercial alga with various biological activity that is widely used as a functional ingredient in food and beverage products. However, there have been few studies on the treatment of UC using S. platensis aqueous extracts (SP), and the underlying mechanism of action of SP against UC has not yet been elucidated. Herein, we aimed to investigate the modulatory effect of SP on microbiota disorders in UC mice and clarify the underlying mechanisms by which SP alleviates damage to the intestinal mucosal barrier. Dextran sulfate sodium (DSS) was used to establish a normal human colonic epithelial cell (NCM460) injury model and UC animal model. The mitochondrial membrane potential assay 3-‍‍(4,5-dimethylthiazol-2-yl)-2,‍5-diphenyltetrazolium bromide (MTT) and staining with Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) and Hoechst 33258 were carried out to determine the effects of SP on the NCM460 cell injury model. Moreover, hematoxylin and eosin (H&E) staining, transmission electron microscopy (TEM), enzyme-linked immunosorbent assay (ELISA), quantitative real-time polymerase chain reaction (qPCR), western blot, and 16S ribosomal DNA (rDNA) sequencing were used to explore the effects and underlying mechanisms of action of SP on UC in C57BL/6 mice. In vitro studies showed that SP alleviated DSS-induced NCM460 cell injury. SP also significantly reduced the excessive generation of intracellular reactive oxygen species (ROS) and prevented mitochondrial membrane potential reduction after DSS challenge. In vivo studies indicated that SP administration could alleviate the severity of DSS-induced colonic mucosal damage compared with the control group. Inhibition of inflammation and oxidative stress was associated with increases in the activity of antioxidant enzymes and the expression of tight junction proteins (TJs) post-SP treatment. SP improved gut microbiota disorder mainly by increasing antioxidant enzyme activity and the expression of TJs in the colon. Our findings demonstrate that the protective effect of SP against UC is based on its inhibition of pro-inflammatory cytokine overproduction, inhibition of DSS-induced ROS production, and enhanced expression of antioxidant enzymes and TJs in the colonic mucosal barrier.

Deficiency in glutathione synthesis and reduction contributes to the pathogenesis of colitis-related liver injury / 中南大学学报(医学版)

OBJECTIVES@#Liver disease is the most common extra-intestinal manifestation of ulcerative colitis (UC), but the underlying pathogenesis is still not clarified. It is well accepted that the occurrence of UC-related liver disease has close correlation with immune activation, intestinal bacterial liver translocation, inflammatory cytokine storm, and the disturbance of bile acid circulation. The occurrence of UC-related liver disease makes the therapy difficult, therefor study on the pathogenesis of UC-related liver injury is of great significance for its prevention and treatment. Glutathione (GSH) shows multiple physiological activities, such as free radical scavenging, detoxification metabolism and immune defense. The synthesis and the oxidation-reduction all contribute to GSH antioxidant function. It is reported that the deficiency in hepatic GSH antioxidant function participates in multiple liver diseases, but whether it participates in the pathogenesis of UC-related liver injury is still not clear. This study aims to investigate the feature and underlying mechanism of GSH synthesis and oxidation-reduction function during the development of UC, which will provide useful information for the pathogenesis study on UC-related liver injury.@*METHODS@#UC model was induced by 2,4,6-trinitrobenzenesulfonic acid (TNBS)-ethanol solution (5 mg/0.8 mL per rat, 50% ethanol) via intra-colonic administration in rats, and the samples of serum, liver, and colon tissue of rats were collected at the 3rd, 5th, and 7th days post TNBS. The severity degree of colitis was evaluated by measuring the disease activity index, colonic myeloperoxidase activity, and histopathological score, and the degree of liver injury was evaluated by histopathological score and the serum content of alanine aminotransferase. Spearman correlation analysis was also conducted between the degree of colonic lesions and index of hepatic histopathological score as well as serum aspartate aminotransferase level to clarify the correlation between liver injury and colitis. To evaluate the hepatic antioxidant function of GSH in UC rats, hepatic GSH content, enzyme activity of GSH peroxidase (GSH-Px), and GSH reductase (GR) were determined in rats at the 3rd, 5th, and 7th days post TNBS, and the protein expressions of glutamine cysteine ligase (GCL), GSH synthase, GSH-Px, and GR in the liver of UC rats were also examined by Western blotting.@*RESULTS@#Compared with the control, the disease activity index, colonic myeloperoxidase activity, and histopathological score were all significantly increased at the 3rd, 5th, and 7th days post TNBS (all P<0.01), the serum aspartate aminotransferase level and hepatic histopathologic score were also obviously elevated at the 7th day post TNBS (all P<0.05). There was a significant positive correlation between the degree of liver injury and the severity of colonic lesions (P=0.000 1). Moreover, compared with the control, hepatic GSH content and the activity of GSH-Px and GR were all significantly decreased at the 3rd and 5th days post TNBS (P<0.05 or P<0.01), and the protein expressions of GCL, GSH-Px, and GR were all obviously down-regulated at the 3rd, 5th, and 7th days post TNBS (P<0.05 or P<0.01).@*CONCLUSIONS@#There is a significant positive correlation between the degree of liver injury and the severity of colonic lesions, and the occurrence of reduced hepatic GSH synthesis and decreased GSH reduction function is obviously earlier than that of the liver injury in UC rats. The reduced hepatic expression of enzymes that responsible for GSH synthesis and reduction may contribute to the deficiency of GSH synthesis and oxidation-reduction function, indicating that the deficiency in GSH antioxidant function may participate in the pathogenesis of UC related liver injury.

Effect of Sishen Pills and its split prescriptions on Tfr/Tfh9/Tfh17 cells in colitis mice / 中国中药杂志

This study aims to investigate the regulatory effect of Sishen Pills(SSP) and its split prescriptions Ershen Pills(EP) and Wuweizi Powder(WP) on T follicular helper(Tfh) cell subset in the dextran sodium sulfate(DSS)-induced colitis mice and the mechanism. A total of 60 male SPF BALB/c mice were used, 10 of which were randomly selected as the normal group. The rest 50 were induced with 3% DSS solution for colitis modeling. After modeling, they were randomized into 5 groups: model group, SSP group, EP group, WP group, and mesalazine group. Body mass, colon mass, colon mass index, colon length, and unit colon mass index in each group were observed. After hematoxylin-eosin(HE) staining, the pathological injury of colon tissue was scored. The expression levels of molecules related to the STAT/SOCS signaling pathway in colon tissues were analyzed by Western blot. Differentiation levels of Tfh cells such as CD4~+CXCR5~+IL-9~+(Tfh9), CD4~+CXCR5~+IL-17~+(Tfh17), and CD4~+CXCR5~+Foxp3~+(Tfr) in peripheral blood of mice were detected by flow cytometry. The results showed each treatment group demonstrated significant increase in body mass and colon length, decrease in colon mass, colon mass index, unit colon mass index, and histopathological score(P<0.05, P<0.01), reduction of the expression of p-STAT3, STAT3, p-STAT6, and STAT6(P<0.05, P<0.01), rise of the expression of SOCS1 and SOCS3(P<0.05, P<0.01), decrease of Tfh9 and Tfh17 cells, and increase of Tfr cells(P<0.05, P<0.01) compared with the model group. These results indicated that SSP and the split EP and WP may alleviate ulcerative colitis by inhibiting the activation of STAT/SOCS signaling pathway and regulating the balance of Tfr/Tfh9/Tfh17 cells.

Effects of astragaloside Ⅳ on inflammatory response and percentage of peripheral blood Th17 cells in mice with ulcerative colitis / 中国中药杂志

This study aimed to investigate the anti-inflammatory effect of astragaloside Ⅳ in mice with ulcerative colitis(UC) and its effect on the percentage of peripheral blood T helper(Th17) cells. Following the establishment of UC mouse model with 2% sodium dextran sulfate(DSS), mice in the positive control group and low-and high-dose astragaloside Ⅳ groups were treated with corresponding drugs by gavage. Disease activity index(DAI) was calculated, and serum interleukin-17(IL-17), tumor necrosis factor-α(TNF-α), and transforming growth factor-β(TGF-β) levels were assayed by ELISA. The pathological changes in colon tissue were observed by HE staining, and Th17/regulatory T cells(Treg) ratio in the peripheral blood was determined by flow cytometry. Western blot was conducted for detecting the relative protein expression levels of forkhead box protein P3(Foxp3) and retinoic acid-related orphan nuclear receptor γT(ROR-γt). The findings demonstrated that in normal mice, the colonic structure was intact. The goblet cells were not reduced and the glands were neatly arranged, with no mucosal erosion, bleeding, or positive cell infiltration. In the model group, the colonic mucosal structure was seriously damaged, manifested as disordered arrangement or missing of glands, vascular dilatation, congestion, and massive inflammatory cell infiltration. The pathological injury of colon tissue was alleviated to varying degrees in drug treatment groups. Compared with the normal group, the model group exhibited elevated percentage of Th17 cells, increased IL-17 and TNF-α content, up-regulated relative ROR-γt protein expression, lowered TGF-β, reduced percentage of Treg cells, and down-regulated relative Foxp3 protein expression. The comparison with the model group showed that DAI score, pathological score, percentage of Th17 cells, IL-17 and TNF-α content, and relative ROR-γt protein expression in the positive control group, low-dose astragaloside Ⅳ group, and high-dose astragaloside Ⅳ group were decreased, while TGF-β content, percentage of Treg cells, and relative Foxp3 protein expression were increased. The DAI score, pathological score, percentage of Th17 cells, IL-17 and TNF-α content, and relative ROR-γt protein expression in the low-dose astragaloside Ⅳ group were higher than those in the positive control group, whereas the content of TGF-β, percentage of Treg cells, and relative Foxp3 protein expression were lower. DAI score, pathological score, percentage of Th17 cells, IL-17 and TNF-α content, relative ROR-γt protein expression in the high-dose astragaloside Ⅳ group declined in contrast to those in the low-dose astragaloside Ⅳ group, while the TGF-β content, percentage of Treg cells, and relative Foxp3 protein expression rose. There was no significant difference between the positive control group and the high-dose astragaloside Ⅳ group. Astragaloside Ⅳ is able to inhibit inflammatory response and diminish the percentage of Th17 cells in mice with UC.

Body composition in patients with crohn's disease and ulcerative colitis / Composição corporal de pacientes com doença de Crohn e colite ulcerativa

ABSTRACT BACKGROUND The nutritional status of individuals with inflammatory bowel diseases is directly related to the severity of the disease and is associated with poor prognosis and the deterioration of immune competence. OBJECTIVE To assess the nutritional status and the body composition of outpatients with inflammatory bowel diseases. METHODS A cross-sectional study was conducted with clinical and nutritional assessment of patients with Crohn’s disease and ulcerative colitis. Patients were classified according to the clinical activity through Crohn’s Disease Activity Index and Mayo Score. Nutritional assessment consisted of anthropometric measurements of current weight, height, mid-arm circumference, triceps skinfold thickness and thickness of adductor policis muscle, with subsequent calculation of BMI, arm muscle circumference and the mid-arm muscle area (MAMA). The phase angle (PhA) and lean and fat mass were obtained with the use of electrical bioimpedance. Descriptive statistics, chi-square test or Fisher exact test, ANOVA and t-test. RESULTS We evaluated 141 patients of which 54 (38.29%) had Crohn’s disease and 87 (61.70%) ulcerative colitis. The mean age was 43.98 (±15.68) years in Crohn’s disease and 44.28 (±16.29) years for ulcerative colitis. Most of the patients were in clinical remission of the disease (Crohn’s disease: 88.89%; ulcerative colitis: 87.36%). Regarding the nutritional classification using BMI, it was found that 48.15% of Crohn’s disease patients were eutrophic and 40.74% were overweight or obese; among patients with ulcerative colitis, 52.87% were classified as overweight or obese. When considering the triceps skinfold, it was observed in both groups a high percentage of overweight and obesity (Crohn’s disease: 75.93%; ulcerative colitis: 72.42%). Crohn’s disease patients showed the most affected nutritional status according to the nutritional variables when compared to patients with ulcerative colitis (BMI: 24.88 kg/m² x BMI: 26.56 kg/m², P=0.054; MAMA: 35.11 mm x MAMA: 40.39 mm, P=0.040; PhA: 6.46° x PhA: 6.83°, P=0.006). CONCLUSION Patients with inflammatory bowel diseases have a high prevalence of overweight and obesity. Crohn’s disease patients had more impaired anthropometric and body composition indicators when compared to patients with ulcerative colitis.

RESUMO CONTEXTO O estado nutricional de indivíduos com doença inflamatória intestinal está diretamente relacionado à gravidade da doença e associado a mau prognóstico e deterioração da competência imune. OBJETIVO Avaliar o status e a composição corporal de pacientes ambulatoriais com doença inflamatória intestinal. METÓDOS Foi conduzido um estudo transversal com avaliação clínica e nutricional de pacientes com doença de Crohn e colite ulcerativa. Pacientes foram classificados de acordo com o índice de atividade clínica Crohn’s Disease Activity Index e escore de Mayo. Avaliação nutricional foi composta peso atual, estatura, circunferência do braço, dobra cutânea tricipital e espessura do músculo adutor do polegar. Posteriormente, foram calculados índice de massa corporal, circunferência muscular do braço e área muscular do braço corrigida. O ângulo de fase e massa magra e massa gorda foram derivadas da bioimpedância elétrica. Foram realizados análise descritiva, teste de qui-quadrado ou exato de Fisher, teste t e ANOVA. RESULTADOS Foram avaliados 141 pacientes, sendo 54 (38,29%) com doença de Crohn e 87 (61,70%) com colite ulcerativa. A idade média foi de 43,98 (±15,68) anos em pacientes com doença de Crohn e 44,28 (±16,29) anos em pacientes com colite ulcerativa. A maioria dos pacientes estava em remissão clínica da doença (doença de Crohn: 88,89%; colite ulcerativa: 87,36%). O estado nutricional de acordo com o IMC foi 48,15% eutrófico e 40,74% sobrepeso/obesidade para doença de Crohn; entre os indivíduos com colite ulcerativa, 52,87% foram classificados como sobrepeso/obesidade. Ao se considerar dobra cutânea do tríceps, observou-se obesidade em ambos os grupos (doença de Crohn 75,93%; colite ulcerativa: 72,42%). Pacientes com doença de Crohn apresentam maiores variações de composição corporal quando comparados com pacientes com colite ulcerativa (IMC: 24,88 kg/m² x IMC: 26,56 kg/m², P=0,054; área do músculo do braço: 35,11mm x área do músculo do braço: 40,39 mm, P=0,040; ângulo de fase: 6,46° x ângulo de fase: 6,83°, P=0,006). CONCLUSÃO Pacientes com doença inflamatória intestinal apresentaram alta prevalência de sobrepeso e obesidade. Indivíduos com doença de Crohn apresentaram parâmetros de composição corporal e de antropometria mais comprometidos, quando comparados com indivíduos com colite ulcerativa.

Evaluation of the immunoexpression of COX-1, COX-2 and p53 in Crohn's disease / Avaliação da imunoexpressão de COX-1, COX-2 e p53 na doença de Crohn

BACKGROUND: Crohn's disease accompanied by nonspecific or idiopathic ulcerative proctocolitis corresponds to a condition called intestinal inflammatory disease. The immunoexpression of cyclooxygenase 2 (COX-2) in Crohn's disease becomes more marked with progression of the disease and the presence of wild-type p53 suppresses the transcription of COX-2. AIMS: To investigate the immunoexpression of cyclooxygenase 1 (COX-1), COX-2 and p53 in Crohn's ileocolitis and to correlated this expression with clinical and histopathological parameters. METHODS: Forty-five cases of Crohn's disease, 16 cases of actinic colitis (diseased-control group) and 11 cases without a history of intestinal disease (normal control group) were studied. Hematoxylin-eosin-stained sections were submitted to histopathological analysis and the immunohistochemical expression of COX-1, COX-2 and p53 was evaluated by the streptavidin-biotin-peroxidase method. RESULTS: Sixty percent of the Crohn's disease patients were women and 40 percent were men, with 75.5 percent whites and 25.5 percent non-whites. The disease involved the terminal ileum in 44.5 percent of cases, ileum in 33.3 percent, colon in 20 percent and duodenum-ileum in 2.2 percent. A significant association was observed between COX-2 immunoreactivity and age <40 years. Histopathological analysis of Crohn's disease samples showed mild or moderate crypt distortion (57.8 percent and 35.6 percent of cases), atrophy (6.6 percent), mild, moderate and marked chronic inflammation (46.7 percent, 26.7 percent and 20 percent), acute inflammatory activity (93.3 percent), ulceration (24.4 percent), mucin depletion (37.8 percent), Paneth's cells (24.4 percent), intraepithelial lymphocytes (93.3 percent), and subepithelial collagen (6.7 percent). In the CD group, COX-1 immunoreactivity in epithelial and inflammatory cells was observed in 26.7 percent and 22.2 percent of cases, respectively. COX-2 immunoreactivity was detected...

RACIONAL: A doença de Crohn, junto com a colite ulcerativa idiopática ou inespecífica constituem a doença inflamatória intestinal. A imunoexpressão de ciclooxigenase 2 (COX-2) na doença de Crohn acentua-se com a progressão da doença, enquanto que a presença do tipo selvagem de p53 suprime a transcrição de COX-2. OBJETIVOS: Investigar a imunoexpressão de ciclooxigenase 1 (COX-1), COX-2 e p53 na doença de Crohn e correlacionar os achados com parâmetros clínico-histopatológicos. MÉTODOS: Foram estudados 45 casos de doença de Crohn (grupo teste), 16 casos de colite actínica (grupo controle-doente) e 11 casos sem história de doença intestinal (grupo controle normal). A avaliação histopatológica foi feita com lâminas coradas pela hematoxilina-eosina e a imunoexpressão de COX-1, COX-2 e p53 foi avaliada por imunoistoquímica, pelo método da estrepto-avidina-biotina-peroxidase. RESULTADOS: Entre os pacientes com doença de Crohn, 60 por cento eram do sexo feminino e 40 por cento do masculino, 75,5 por cento brancos e 25,5 por cento não-brancos. A doença comprometia o íleo terminal em 44,5 por cento dos casos, íleo em 33,3 por cento, cólon em 20 por cento e duodeno-íleo em 2,2 por cento. Associação significante foi detectada entre a imunoexpressão de COX-2 e pacientes com <40 anos. A histopatologia dos casos de doença de Crohn mostrou distorção críptica em grau leve ou moderado (57,8 por cento e 35,6 por cento dos casos), atrofia (6,6 por cento), inflamação focal, difusa superficial e difusa transmural (46,7 por cento, 26,7 por cento e 20 por cento), inflamação aguda neutrofílica (93,3 por cento), alterações epiteliais: ulceração (24,4 por cento), depleção de mucina (37,8 por cento), células de Paneth (24,4 por cento); alterações epiteliais associadas: linfócitos intra-epiteliais (93,3 por cento) e colágeno subepitelial (6,7 por cento). No grupo doença de Crohn, imunoexpressão de COX-1, em células epiteliais e inflamatórias foi observada em 26,7...

Multidrug resistance gene (MDR-1) expression in the colonic mucosa of patients with refractory ulcerative colitis / Expresión del gen de multirresistencia a drogas (MDR-1) en la mucosa colónica de pacientes con colitis ulcerosa refractaria

BACKGROUND: P-Glycoprotein (P-gp), a product of the MDR-1 gene, is a transmembrane efflux pump involved in drug transport, first described in cancer refractoriness. In the normal bowel P-gp is detectable on superficial epithelial cells, but has not been described in crypt epithelium. The role of P-gp and its intestinal expression in steroid-refractory ulcerative colitis (UC) are controversial. AIM: to compare P-gp immunostaining pattern in colonic epithelial cells of steroid-refractory versus steroid-responder UC patients. METHODS: P-gp was assessed by immunohistochemistry in rectal biopsies obtained from 19 patients with active UC, including pre-surgical samples from 11 refractory patients who underwent colectomy, and 8 responders. We devised a 5-point (0-4) score, according to the percentage of epithelial surface with positive immunostaining in the superficial and crypt epithelium (apical, lateral and cytoplasmic areas). RESULTS: Compared with responders, steroid-refractory patients had significantly higher immunostaining scores in the superficial epithelium, both in apical (2.8+/-0.5 versus 1.1+/-0.5, p=0.023) and cytoplasmic cellular areas (2.7+/-0.5 versus 1.2+/-0.5, p=0.032). Positive immunostaining of the superficial epithelium was frequently detected in refractory patients (apical: 9/11 cases, cytoplasmic: 10/11 cases) but was only observed in 4/8 responders. P-gp was also detected in similar areas of the crypt epithelium in 6/11 refractory patients, while it was infrequent in the group of 8 responders (1 apical 1 case, cytoplasmic 2 cases). Samples from the mucosa of normal ileal pouch-anal anastomoses obtained several years after the surgical procedure had a P-gp immunostaining pattern which was similar to that of rectal samples from patients with refractory UC. CONCLUSIONS: These results suggest a critical role of P-gp overexpression in steroid-refractory UC.

Antecedentes. La glicoproteína P (P-gp), un producto del gen MDR-1, es una bomba de eflujo transmembranainvolucrada en el transporte de drogas, descripta por primera vez en el cáncer refractario. En el intestino normal, P-gp se detecta sobre las célulasepiteliales superficiales, pero no se la ha descripto en el epitelio de las criptas. El papel de P-gp y su expresiónintestinal en la colitis ulcerosa (CU) refractaria a esteroides es controvertido. Objetivo. Comparar elpatrón de inmunotinción de P-gp en células epiteliales colónicas de pacientes con CU refractaria vs.respondedora a esteroides. Métodos. Se estudió P-gp por inmunohistoquímica en biopsias rectales obtenidasde 19 pacientes con CU activa, incluyendo muestras prequirúrgicas de 11 pacientes refractarios que fueronsometidos a una colectomía y muestras de 8 respondedores. Ideamos un score de 5 puntos (0-4), según elporcentaje de superficie epitelial con inmunotinción positiva en el epitelio superficial y críptico (áreas apical,lateral y citoplásmica). Resultados. Comparados con los respondedores, los pacientes refractarios a esteroides tenían scores de inmunotinción significativamente mayores en el epitelio superficial, tanto en lasáreas celulares apical (2.8+0.5 vs. 1.1+0.5, p=0.023) como citoplásmica (2.7+0.5 vs. 1.2+0.5, p=0.032). Se detectó frecuentemente inmunotinción positiva en el epitelio superficial en los pacientes refractarios (apical: 9/11 casos, citoplásmica: 10/11 casos), pero la misma se observó sólo en 4/8 respondedores. P-gp también sedetectó en áreas similares del epitelio de las criptas en 6/11 pacientes refractarios, en tanto que fue infrecuenteen el grupo de los 8 respondedores (1 caso en el área apical y 2 en la citoplásmica). Fuerón estudiadasbiopsias de la mucosa de la anastomosis pouch ileal - anal, obtenidas varios años después del procedimeinto quirúrgico, observándose un patrón de...

Clinical Usefulness of Glucocorticoid Receptor beta Expression and NF-kappa B Activity in Patients with Ulcerative Colitis / 대한소화기학회지

BACKGROUND/AIMS: Glucocorticoid resistance poses a challenging clinical problem in inflammatory bowel disease because more than one fourth of patients with severe ulcerative colitis do not respond to anti-inflammatory steroids. Recently, it has been reported that glucocorticoid response is related to the expression of human glucocorticoid receptor beta (hGRbeta) and nuclear factor-kappa B (NF-kappaB) activity. The aims of this study were to clarify whether these factors may predict the responsiveness before treatment. METHODS: Total RNA was extracted from peripheral blood mononuclear cell (PBMC) and colonic mucosa in 17 patients of ulcerative colitis before steroid administration. RNA was reverse transcribed and the resulting complementary DNA was amplified using specific primers for hGR alpha and beta. Concomitantly, NF-kappaB activity in colonic mucosa was assessed by immunohistochemical stain. RESULTS: The expression of hGRbeta mRNA was detected in 10 patients (58.8%) in PBMC and 8 patients (47.1%) in colon, respectively. Operations were performed in 5 patients due to steroid unresponsiveness. Only 5 of 17 patients (29.4%) were consistent in the expression of hGRbeta between PBMC and colon. Seven of 15 patients (46.7%) showed an alteration in the expression of hGRbeta in PBMC after glucocorticoid treatment. NF-kappaB activity was found in both epithelial cell and lamina propria in 12, epithelial cell alone in 1, lamina propria alone in 1 and all negative in 3 patients, respectively. CONCLUSIONS: The expression of hGRbeta was discordant between PBMC and colon in the same patient and showed a change in the expession after the glucocorticoid treatment in nearly half. The expression of hGRbeta and colonic NF-kappaB activity patterns do not provide useful information about glucocorticoid response in patients with ulcerative colitis.

Enhanced mucosal re-epithelialization induced by short chain fatty acids in experimental colitis

The short chain fatty acids (SCFA) are the best nutrients for the colonocytes. Glucose is poorly used as a fuel but may be transformed into SCFA by colonic bacteria. The aim of this study was to investigate the effect of SCFA or glucose on experimental colitis. Colitis was induced in 30 Wistar rats by colonic instillation of 4 percent acetic acid. Five days later they were randomized to receive twice a day colonic lavage containing saline (controls, N = 10), 10 percent hypertonic glucose (N = 10) or SCFA (N = 10) until day 8 when they were killed. At autopsy, the colon was removed and weighed and the mucosa was evaluated macro- and microscopically and stripped out for DNA assay. Data are reported as mean + or -SD or median [range] as appropriate. All animals lost weight but there was no difference between groups. Colon weight was significantly lower in the SCFA group (3.8 + or - 0.5 g) than in the control (5.3 + or - 2.1 g) and glucose (5.2 + or - 1.3 g) groups (PP<0.05). Macroscopically, the severity of inflammation was less in SCFA (grade 2 [1-5]) than in control (grade 9 [4-10]) and glucose-treated (grade 9 [2-10]) animals (P<0.01). Microscopically, ulceration of the mucosa was more severe in the glucose and control groups than in the SCFA group. The DNA content of the mucosa of SCFA-treated animals (8.2 [5.0-20.2] mg/g of tissue) was higher than in glucose-treated (5.1 [4.2-8.5] mg/g of tissue; P<0.01) and control (6.2 [4.5-8.9] mg/g of tissue; P<0.05) animals. We conclude that SCFA may enhance mucosal re-epithelialization in experimental colitis, whereas hypertonic glucose is of no benefit

Utilizaçäo do metabolismo protéico (15N-glicina) na detecçäo precoce de agravamento da atividade da doença em pacientes portadores de retocolite ulcerativa inespecífica / Utilization of the proteic metabolism (15N-glycine) in the precocious detection of disease exacerbation in patients with unespecific ulcerative colitis

Dez pacientes portadores de retocolite ulcerativa inespecífica em atividade leve e moderada foram acompanhados por um período de quatro meses. Foram avaliados os seguintes indicadores de atividade da doença: clínicos (número de evacuaçoes e presença de sangue nas fezes), laboratoriais (hemoglobina, contagem de plaquetas, VHS, mucoproteínas, alpha1-glicoproteína ácida e PCR), escore retossigmoidoscópico e histopatológico. Avaliou-se também o metabolismo protéico dos pacientes mediante (15)N-glicina e amônia como produto final. Apenas um paciente teve exacerbaçao acentuada da atividade da doença durante o período de estudo. Esse paciente apresentava no início do estudo valores de síntese e catabolismo protéicos superiores aos de todos os outros pacientes, ou seja, 4,51 e 3,47 g de proteína/Kg/dia, respectivamente, mostrando estado de hipermetabolismo, compatível com aumento da atividade da doença. Esse aumento de atividade nao foi, entretanto, detectado pelos demais indicadores utilizados, possibilitando aventar a hipótese de que a avaliaçao do metabolismo protéico consegue detectar precocemente o agravamento da atividade da doença em pacientes com retocolite ulcerativa.

Sensibilidade aos efeitos genotóxicos da bleomicina: colite ulcerativa versus câncer colorretal / Sensibility genotoxic effects of bleomycin: ulcerative colitis versus colorectal cancer

Neste estudo foram selecionados 35 pacientes, sendo 13 sadios (grupo I); 10 pacientes com colite ulcerativa (grupo II); e 12 pacientes com câncer colorretal (grupo III). Para determinar a resposta aos efeitos mutagênicos da bleomicina em cultura de linfócitos de sangue periférico, foram realizados dois tipos de cultura para cada paciente: a primeira, sem tratamento com bleomicina (cultura espontânea) e, a segunda, tratada com bleomicina (cultura induzida). Na avaliaçäo microscópica dos linfócitos em metástase, foram determinados o índice mitótico e as alteraçöes cromossômicas estruturais. Os três grupos de pacientes diferiram entre si quanto às determinaçöes do índice mitótico nas culturas espontâneas (p < 0,01) e significativamente maiores que os dos grupos I e III. Quanto à freqüência de quebras cromatídicas espontâneas por célula, näo houve diferença significativa entre os três grupos de pacientes (p = 0,39). Quanto à freqüência de quebras cromatídicas induzidas por célula, observou-se que havia diferença significativa entre os três grupos de pacientes (p < 0,001). O grupo III apresentou valores significativamente superiores aos verificados no grupo II, e este significativamente maiores que os do grupo I. Com base nos resultados, observou-se que a bleomicina induziu ao aumento significativo de aberraçöes cromossômicas nos linfócitos dos grupos de pacientes com colite ulcerativa e com câncer colorretal. A sensibilidade aos efeitos genotóxicos da bleomicina em pacientes com colite ulcerativa pode expressar maior predisposiçäo ao câncer colorretal

Efeito comparativo entre ácidos graxos w3 e sulfassalazina sobre indicadores de atividade da doença e a cinética da 15N-Glicina no tratamento de pacientes com retocolite inespecífica / Comparison of omega 3 fatty acids and sulfasalazine in ulcerative colitis

A açäo antiinflamatória da sulfassalazina (SS) e dos ácidos graxos poliinsaturados w3 foi estudada, comparativamente, em 10 pacientes (cinco homens e cinco mulheres) com média de idade de 48ñ12 anos, portadores de retocolite ulcerativa inespecífica nas formas leve e moderada, divididos em 2 grupos. Cada grupo recebeu, durante 2 meses, SS (2g/dia) ou ácidos graxos w3 de óleo de peixe (5,4g/dia), segundo delineamento cruzado em que o tratamento com w3 foi seguido de 2 meses de "washout". A avaliaçäo da atividade da doença (clínico, laboratorial, histológica e retossigmoidoscópica), do estado nutricional e do metabolismo protéico global (15N-glicina) foram realizados no início (M1), após 2 meses (M2) e no final do tratamento (M3). No pré tratamento (M1) nenhum paciente se encontrava desnutrido, permanecendo desta forma até M3. Apenas um paciente descompensou durante o estudo, quando fazia uso de ácidos graxos w3. A menor atividade da doença foi observada nos períodos em que a SS foi empregada. Isto pode ser detectado mediante alguns indicadores clínicos (sangramento retal, número de evacuaçöes e consistência das fezes), laboratoriais (contagem de plaquetas, VHS, PCR e alfa1-glicoproteína ácida) e escore histopatológico (a 15 cm da reborda anal). As principais alteraçöes do metabolismo protéico foram no sentido do menor ritmo metabólico conseqüente ao uso da SS. Desta forma, a atividade da doença parece menos responsiva aos ácidos graxos w3, que à SS, o que pode ser detectado pelos indicadores clássicos mais sensíveis, e também pelo estudo cinético de aminoácidos.

Prostaglandin catabolism and ulcerative colitis: effect of sulphasalazine, 5-aminosalicylic acid and other drugs

1. Evidence is presented for the occurrence of type 1 prostaglandin 15-hidroxydehydrogenase in human rectal mucosa. No evidence of the presence of type 2 dnzyme was found. 2. A 15-keto-prostaglandin reductase, responsible for the breakdown of 13, 14-dihydro 15-keto prostaglandins to 13, 14-dihydro prostaglandins, was also present in human rectal mucosa. 3. Ulcerative colitis patients catabolized prostaglandins to the same extent as the control group. PGE was catabolized significantly better than PGF2 alfa. 4. 5-Aminosalicylic acid and sulphapyridine did not affect prostaglandin catabolism. Sulphasalazine, methilsulphasalazine, indomethacin, flurbiprofen, disodium cromoglycate, sodium salicylate and carbenoxolone sodium inhibited prostaglandin catabolism to the same extent in both groups.Salicylazosulphadimidine was a more potent inhibitor of PGE1 catabolism than of PGF2alfa. 5. The increased prostaglandin synthesis reported for ulcerative colitis patients was not paralleled by increased catabolism, a fact possibly contributing to the accumulation of such compounds in the diseased state

Lectin binding in colorectal mucosa.

Lectin binding of goblet cell mucin in human colonic mucosa was studied in patients with irritable bowel syndrome, colorectal malignancy and ulcerative colitis using plant lectins, Dolichos biflorus agglutinin (DBA) and peanut agglutinin (PNA). Normal colonic mucosa demonstrated a strong binding with DBA (100%) but did not bind to PNA at all. Colonic carcinomas showed strong PNA binding (7 of 15 biopsies) while DBA binding was absent in 14 of 15 biopsies. The transitional mucosa showed reduced or absent DBA binding in 6 and positive PNA binding in 2 of 15 biopsies. During the active phase of ulcerative colitis, there was a loss of DBA binding in 10 of 15 biopsies, which was restored during remission in all. One biopsy with severe dysplasia showed PNA binding. It is concluded that normal colorectal mucosa binds DBA strongly but not PNA. Malignant tissue and transitional mucosa reveal PNA binding often, with decreased DBA binding. In ulcerative colitis DBA binding decreases during phases of activity.

Uma formulaçäo oral para liberaçäo de ácido 5-aminossalicílico no cólon / An oral preparation for release of 5-aminosalicylic acid in the colon

Estudou-se uma preparaçäo oral de ácido 5-aminossalicílico de liberaçäo pH dependente. Participaram do estudo 11 indivíduos com idades variando entre 20 e 63 anos, dos quais três eram voluntários sadios, seis apresentavam retocolite ulcerativa e dois doença de Crohn. Através de um acompanhamento radiológico efetuado na 6ª, 12ª e 24ª hora, constatou-se que a desintegraçäo dos comprimidos se realiza somente na regiäo ileocecal e cólon. Finalmente, com base nos resultados, destaca-se a potencialidade da formulaçäo no controle das doenças inflamatórias intestinais