TMT proteomics analysis of intestinal tissue from patients of irritable bowel syndrome with diarrhea: Implications for multiple nutrient ingestion abnormality.

Diarrheal irritable bowel syndrome (IBS-D) is a chronic functional bowel disease with no clear diagnostic markers and no satisfactory treatment strategies. In recent years, the importance of intestinal microstructure and function in IBS-D has been emphasized. However, the intestinal tissue proteomics of IBS-D patients has not been analyzed. Here, we systematically analyzed the molecule profiling of the intestinal tissues in IBS-D patients through tandem mass tag (TMT)-based proteomics for the first time, aiming to reveal the pathogenesis and provide evidence for diagnosis and treatment of IBS-D. Five IBS-D patients and five healthy subjects were selected, biopsy tissue samples from the junction of sigmoid and rectum were analyzed by TMT proteomics. Differentially expressed proteins were obtained and bioinformatics analysis was performed. Furthermore, parallel reaction monitoring (PRM) and q-PCR detection were applied to validate the differentially expressed proteins. Eighty differentially expressed proteins were screened, 48 of which were up-regulated and 32 were down-regulated (fold change >1.2, P < 0.05). Bioinformatics analysis showed that these proteins were significantly enriched in the nutrient ingestion pathways which are related to immune molecules. SELENBP1, VSIG2, HMGB1, DHCR7, BCAP31 and other molecules were significantly changed. Our study revealed the underlying mechanisms of IBS-D intestinal dysfunction. SIGNIFICANCE: Irritable bowel syndrome with diarrhea (IBS-D) is a worldwide chronic intestinal disease with no definite diagnostic markers. It is still a challenge to accurately locate the pathogenesis of patients for appropriate treatment strategy. Established proteomics studies of IBS-D are only based on urine, blood, or tissue samples from animals. Our study was the first TMT proteomics analysis on intestinal biopsy tissues of patients with IBS-D, which revealed the changes of molecular spectrum of actual intestinal conditions in patients with IBS-D. Some important molecules and signaling pathways have been found abnormal in our study, which were related with nutrient uptake. They not only provided preliminary clues for low fermentable oligosaccharides, disaccharides, monosaccharides, and polyols (FODMAP) intolerance, an unsolved conundrum of IBS-D, but also revealed obscure problems of protein, lipid, and other nutrients ingestion in IBS-D patients. Some of these differentially expressed molecules have been preliminarily verified, and will may be potential candidate molecules for diagnostic markers of IBS-D.

Sestrin2 protects dendritic cells against endoplasmic reticulum stress-related apoptosis induced by high mobility group box-1 protein.

Sestrin2 (SESN2) is a highly evolutionary conserved protein and involved in different cellular responses to various stresses. However, the potential function of SESN2 in immune system remains unclear. The present study was designed to test whether dendritic cells (DCs) could express SESN2, and investigate the underlying molecular mechanism as well as its potential significance. Herein, we firstly reported that SESN2 was expressed in DCs after high mobility group box-1 protein (HMGB1) stimulation and the apoptosis of DCs was obviously increased when SESN2 gene silenced by siRNA. Cells undergone SESN2-knockdown promoted endoplasmic reticulum (ER) stress (ERS)-related cell death, markedly exacerbated ER disruption as well as the formation of dilated and aggregated structures, and they significantly aggravated the extent of ERS response. Conversely, overexpressing SESN2 DCs markedly decreased apoptotic rates and attenuated HMGB1-induced ER morphology fragment together with inhibition of ERS-related protein translation. Furthermore, sesn2-/--deficient mice manifested increased DC apoptosis and aggravated ERS extent in septic model. These results indicate that SESN2 appears to be a potential regulator to inhibit apoptotic ERS signaling that exerts a protective effect on apoptosis of DCs in the setting of septic challenge.

Role of dendritic cells in the host response to biomaterials and their signaling pathways.

Strategies to enhance, inhibit, or qualitatively modulate immune responses are important for diverse biomedical applications such as vaccine adjuvant, drug delivery, immunotherapy, cell transplant, tissue engineering, and regenerative medicine. However, the clinical efficiency of these biomaterial systems is affected by the limited understanding of their interaction with complex host microenvironments, for example, excessive foreign body reaction and immunotoxicity. Biomaterials and biomedical devices implanted in the body may induce a highly complicated and orchestrated series of host responses. As macrophages are among the first cells to infiltrate and respond to implanted biomaterials, the macrophage-mediated host response to biomaterials has been well studied. Dendritic cells (DCs) are the most potent antigen-presenting cells that activate naive T cells and bridge innate and adaptive immunity. The potential interaction of DCs with biomaterials appears to be critical for exerting the function of biomaterials and has become an important, developing area of investigation. Herein, we summarize the effects of the physicochemical properties of biomaterials on the immune function of DCs together with their receptors and signaling pathways. This review might provide a complete understanding of the interaction of DCs with biomaterials and serve as a reference for the design and selection of biomaterials with particular effects on targeted cells. STATEMENT OF SIGNIFICANCE: Biomaterials implanted in the body are increasingly applied in clinical practice. The performance of these implanted biomaterials is largely dependent on their interaction with the host immune system. As antigen-presenting cells, dendritic cells (DCs) directly interact with biomaterials through pattern recognition receptors (PRRs) recognizing "biomaterial-associated molecular patterns" and generate a battery of immune responses. In this review, the physicochemical properties of biomaterials that regulate the immune function of DCs together with their receptors and signaling pathways of biomaterial-DC interactions are summarized and discussed. We believe that knowledge of the interplay of DC and biomaterials may spur clinical translation by guiding the design and selection of biomaterials with particular effects on targeted cell for tissue engineering, vaccine delivery, and cancer therapy.

Inhibition of Cerebral High-Mobility Group Box 1 Protein Attenuates Multiple Organ Damage and Improves T Cell-Mediated Immunity in Septic Rats.

Sepsis remains one of the leading causes of mortality in intensive care units, but there is a shortage of effective treatments. A dysregulated host immune response and multiple organ injury are major factors for the pathogenesis and progression of sepsis, which require specific mechanism and treatment. In the present study, we performed an intracerebroventricular (ICV) injection of BoxA, a specific antagonist of high-mobility group box 1 protein (HMGB1), in septic rats that were produced by cecal ligation and puncture surgery; we further assessed the functional changes of multiple organs and splenic T lymphocytes. We found that the inhibition of cerebral HMGB1 significantly alleviated multiple organ damage under septic exposure, including damage to the heart, liver, lungs, and kidneys; reversed the immune dysfunction of T cells; and increased the survival of septic rats. These data suggest that central HMGB1 might be a potential therapeutic target for septic challenge and that inhibition of brain HMGB1 can protect against multiple organ dysfunction induced by sepsis.

Early antagonism of cerebral high mobility group box-1 protein is benefit for sepsis induced brain injury.

Sepsis induced brain injury acts as an acute complication and accounts for deterioration and high mortality rate of septic condition. HMGB1 is a late inflammatory mediator that plays a critical role in brain dysfunction and diseases. However, the role of HMGB1 in sepsis induced brain dysfunction remains intricate. The current study investigated the effect of HMGB1 on brain injury in septic mice model with intracerebroventricular injection of BoxA (a specific antagonist of HMGB1). The expression of HMGB1, morphological changes of brain tissues, apoptosis of brain cells, and alteration of behavior were determined. The expressions of HMGB1 in cortex, hippocampus, and striatum were significantly enhanced in the sepsis group when compared with the sham group. In septic conditions, brain tissues showed significant abnormalities in tissue structure, and increased apoptosis of brain cells which was caspase-3 dependent. Septic mice showed suppression of locomotor activity and impairment of memory and learning. Neutralizing brain HMGB1 significantly improved brain injury and apoptosis of brain cells, and further ameliorated disturbed locomotor activities and damaged memory and learning. However, no significant improvement of survival rate was seen after inhibiting central HMGB1. These results reveal that HMGB1 is a potential target for ameliorating sepsis induced brain injury with early antagonizing.

The Protective Effect of Alpha 7 Nicotinic Acetylcholine Receptor Activation on Critical Illness and Its Mechanism.

Critical illnesses and injuries are recognized as major threats to human health, and they are usually accompanied by uncontrolled inflammation and dysfunction of immune response. The alpha 7 nicotinic acetylcholine receptor (α7nAchR), which is a primary receptor of cholinergic anti-inflammatory pathway (CAP), exhibits great benefits for critical ill conditions. It is composed of 5 identical α7 subunits that form a central pore with high permeability for calcium. This putative structure is closely associated with its functional states. Activated α7nAChR exhibits extensive anti-inflammatory and immune modulatory reactions, including lowered pro-inflammatory cytokines levels, decreased expressions of chemokines as well as adhesion molecules, and altered differentiation and activation of immune cells, which are important in maintaining immune homeostasis. Well understanding of the effects and mechanisms of α7nAChR will be of great value in exploring effective targets for treating critical diseases.

The potential mechanism of extracellular high mobility group box-1 protein mediated p53 expression in immune dysfunction of T lymphocytes.

In the present study, we examined the activity of p53 protein in Jurkat cells treated with high mobility group box-1 protein (HMGB1), thereafter we investigated the mechanism of extracellular HMGB1 mediated p53 expression in immune dysfunction of T lymphocytes. mRNA expression of p53, mdm2, and p21 was determined by Real-time reverse transcription-polymerase chain reaction(RT-PCR). The apoptotic rate of Jurkat cells was analyzed by flow cytometry. Expressions of bcl-2, bax, caspase-3, phosphorylated (p) extracellular signal-regulated kinase (ERK)1/2, ERK1/2, p-p38 mitogen-activated protein kinase (MAPK), p38 MAPK, and p-c-jun amino-terminal kinase (JNK)1/2 and JNK1/2 were simultaneously determined by Western blotting. After treatment with HMGB1 (100 ng/ml or 1000 ng/ml), the proliferative activity of Jurkat cells was significantly decreased, and a low and medium concentration of HMGB1 induced an up-regulation of p53 mRNA, p-p53 and p53 protein expression. Meanwhile, levels of mdm2 and p21 were elevated by incubated with HMGB1 (100 ng/ml) for 24 or 48 hours. Moreover, the proliferation of Jurkat cells in response to HMGB1 (100 ng/ml) in the vector group was significantly depressed. The bax and caspase-3 levels in p53 shRNA-expressed cells treated with HMGB1 (100 ng/ml) was markedly decreased, whereas expression of bcl-2 was obviously enhanced. Among ERK1/2, p38 MAPK and JNK1/2 signaling, only p38 MAPK pathway could be significantly activated by treatment with HMGB1, and the specific inhibitor of p38 MAPK was used, p53 and p-p53 expression induced by HMGB1 were significantly down-regulated. Taken together, our data strongly indicated that HMGB1 might enhance p53 expression, which was associated with both the proliferative activity as well as apoptosis of T cells.

Expression of IL-37 contributes to the immunosuppressive property of human CD4+CD25+ regulatory T cells.

Interleukin-37 (IL-37) possesses the function of down-regulate systemic and local inflammation. It is unknown whether IL-37 is expressed in human regulatory T cells (Tregs) and its role in modulating the immune response of Tregs. In the present study, cell surface molecules and secretory cytokines were analyzed in order to determine the function of IL-37 in regulating inhibitory effect of human CD4(+)CD25(+)Tregs. Meanwhile, the effects of IL-37 on T cell differentiation and proliferation as co-culture of CD4(+)CD25(+)Treg/CD4(+)CD25(-)T cell were also investigated. It was showed that IL-37 was expressed in cytoplasm of CD4(+)CD25(+)Tregs, and the levels of IL-37 were gradually elevated with the enhanced activity of CD4(+)CD25(+)Tregs. Secretory cytokines such as transforming growth factor (TGF)-ß and interleukin (IL)-10, and expressions of cell surface molecules, including forkhead/winged helix transcription factor p3 (FOXP3) and cytotoxic T-lymphocyte associated antigen (CTLA)-4, were significantly decreased when IL-37 gene was silenced by siRNA. Furthermore, down-regulation of IL-37 expression in human CD4(+)CD25(+)Tregs obviously promoted proliferation of co-cultured T cell and differentiation, together with observably enhancement of IL-2 formation. These results demonstrated that IL-37 might manifest as a critical protein involving in immunosuppression of human CD4(+)CD25(+)Tregs.

[The preventive and therapeutic effect of advanced airway management on pulmonary infection in patients with inhalation injury after tracheotomy].

OBJECTIVE: To observe the preventive and therapeutic effect of advanced airway management on pulmonary infection in patients with inhalation injury after tracheotomy. METHODS: fourteen burn patients with inhalation injury admitted to our hospital from January 2001 to December 2004 were enrolled as control (C) group, and they were treated with conventional systemic therapy and management of airway. Twenty-seven burn patients with inhalation injury admitted to our hospital from January 2005 to October 2009 were enrolled as advanced (A) group, and they were treated with conventional systemic therapy and advanced airway management, including bedside isolation of airway, fixation of both oxygen supply tube and humidifying tube, humidification in specific body position, thinning of sputum, lavement of airway and procedural sputum elimination, steam inhalation combined with medicine, and suction of sputum with interrupted negative pressure. Result of bacterial culture of sputum (the 7th day after tracheotomy) and chest X-ray (at admission and the 7th day after tracheotomy), pulmonary infection, change in blood gas analysis index and oxygen saturation (SO(2)), (within 7 days after tracheotomy), and the number of patients curd in 2 groups were observed and compared. RESULTS: (1) Positive result of bacterial culture of sputum was observed in 11 (78.6%) patients in C group and 12 (44.4%) patients in A group. The difference between them was statistically significant (chi(2) = 4.36, P < 0.05). The main bacterium detected was Pseudomonas aeruginosa. (2) Pneumonia was suspected in 7 patients (25.9%) in A group by chest X-ray, which was obviously fewer than that in C group (8 Cases, 57.1%, chi(2) = 3.87, P < 0.05). The result was in accordance with the diagnosis of pulmonary infection. (3) No CO(2) retention, SO(2) and PaCO(2) abnormality caused by asphyxia was observed in 2 groups, PaCO(2) value in A group was close to that in C group (t = 0.89, P > 0.05). (4) In C group, 9 (64.3%) patients were cured, 5 patients died of pneumonia, wound sepsis, and MODS. In A group, 25 (92.6%) patients were cured, 2 patients died of MODS. Number of cure was obviously larger in A group than in C group (chi(2)= 5.22, P < 0.05). CONCLUSIONS: The advanced airway management has better effects on isolation and humidification of airway, and thinning, drainage, and elimination of sputum. And it can decrease the probability of blind suction and injury to airway, and it prevents pulmonary infection following tracheotomy.