Evaluation of Ubiquitinated Proteins by Proteomics Reveals the Role of the Ubiquitin Proteasome System in the Regulation of Grp75 and Grp78 Chaperone Proteins during Intestinal Inflammation

The ubiquitin proteasome system (UPS) is the major pathway of intracellular protein degradation and may be involved in the pathophysiology of inflammatory bowel diseases or irritable bowel syndrome. UPS specifically degrades proteins tagged with an ubiquitin chain. We aimed to identify polyubiquitinated proteins during inflammatory response in intestinal epithelial HCT-8 cells by a proteomic approach. HCT-8 cells were incubated with interleukin 1β, tumor necrosis factor-α, and interferon-γ for 2 h. Total cellular protein extracts were separated by 2D gel electrophoresis and analyzed by an immunodetection using antiubiquitin antibody. Differential ubiquitinated proteins were then identified by LC-ESI MS/MS. Seven proteins were differentially ubiquitinated between control and inflammatory conditions. Three of them were chaperones: Grp75 and Hsc70 were more ubiquitinated (p < 0.05) and Grp78 was less ubiquitinated (p < 0.05) under inflammatory conditions. The results for Grp75 and Grp78 were then confirmed in HCT-8 cells and in 2-4-6-trinitrobenzen sulfonic acid induced colitis in rats mimicking inflammatory bowel disease by immunoprecipitation. No difference was observed in irritable bowel syndrome like model. In conclusion, we showed that a proteomic approach is suitable to identify ubiquitinated proteins and that UPS-regulated expression of Grp75 and Grp78 may be involved in inflammatory response. Further studies should lead to the identification of ubiquitin ligases responsible for Grp75 and Grp78 ubiquitination

Dietary n-3 PUFA May Attenuate Experimental Colitis

International audienceBackground. Inflammatory bowel diseases (IBD) occurred in genetically predisposed people exposed to environmental triggers. Diet has long been suspected to contribute to the development of IBD. Supplementation with n-3 polyunsaturated fatty acids (PUFA) protects against intestinal inflammation in rodent models while clinical trials showed no benefits. We hypothesized that intervention timing is crucial and dietary fatty acid pattern may influence intestinal environment to modify inflammation genesis. The aim of this study was to evaluate the dietary effect of PUFA composition on intestinal inflammation. Methods. Animals received diet varying in their PUFA composition for four weeks before TNBS-induced colitis. Colon inflammatory markers and gut barrier function parameters were assessed. Inflammatory pathway PCR arrays were determined. Results. n-3 diet significantly decreased colon iNOS, COX-2 expression, IL-6 production, and LTB4 production but tended to decrease colon TNFα production () compared to control diet. Tight junction protein (claudin-1, occludin) expressions and MUC2 and TFF3 mRNA levels were not different among groups. n-9 diet also decreased colon IL-6 production (). Conclusions. Dietary n-3 PUFA influence colitis development by attenuating inflammatory markers. Further research is required to better define dietary advice with a scientific rationale

The association of intraprostatic calcifications and dosimetry parameters with biochemical control after permanent prostate implant

Purpose: The objective of this study was to evaluate the impact of intraprostatic calcifications (IC) on long-term tumor control in patients treated with permanent implant prostate brachytherapy (PIPB). Materials and Methods: Data from 609 I-125 patients treated with PIPB were retrospectively reviewed. The presence of IC was determined by reviewing postimplant CT images. Doses delivered were determined using the Monte Carlo (model-based) calculations and the TG43 approach. Biochemical relapses at 7 and 10 years were determined according to Phoenix definition. Long-term biochemical relapse-free survival (bRFS) was determined using Kaplan–Meier estimates with log rank test. Cox proportional hazard models were used for analysis of predictor factors of biochemical recurrence. Results: IC were observed for 11.1% of patients. Clinical stage, PSA, Gleason score, D'Amico risk group, and ADT use were comparable between IC and no IC groups. The 7- and 10-year bRFS for the entire cohort were 94.1% and 90.6%, respectively. The bRFS at 7 years was 90.5% (with IC) vs. 94.5% (without IC) (p = 0.198); the corresponding values at 10 years were 78.8% vs. 91.8% (p = 0.046). On Cox model, only prostatic calcifications were a significant risk factor for biochemical relapse (HR: 2.30, IC 95%: 1.05–5.00, p = 0.037; and HR: 3.94; IC 95%: 1.00–15.38; p = 0.049 for univariate and multivariate analysis, respectively). Conclusion: The presence of IC in patients treated with PIPB decreases V100 and D90 for postimplant Monte Carlo dosimetry (compared with TG43); correspondingly, IC are associated with a lower 10-y bRFS. Model-based dose calculations are critical to evaluate potential cold spots due to calcifications