Tumor Suppressor CYLD Acts as a Negative Regulator for Non-Typeable Haemophilus influenza-Induced Inflammation in the Middle Ear and Lung of Mice

Non-typeable Haemophilus influenza (NTHi) is an important human pathogen causing respiratory tract infections in both adults and children. NTHi infections are characterized by inflammation, which is mainly mediated by nuclear transcription factor kappaB (NF-κB)-dependent production of inflammatory mediators. The deubiquitinating enzyme cylindromatosis (CYLD), loss of which was originally reported to cause a benign human syndrome called cylindromatosis, has been identified as a key negative regulator for NF-κB in vitro. However, little is known about the role of CYLD in bacteria-induced inflammation in vivo. Here, we provided direct evidence for the negative role of CYLD in NTHi-induced inflammation of the mice in vivo. Our data demonstrated that CYLD is induced by NTHi in the middle ear and lung of mice. NTHi-induced CYLD, in turn, negatively regulates NTHi-induced NF-κB activation through deubiquitinating TRAF6 and 7 and down-regulates inflammation. Our data thus indicate that CYLD acts as a negative regulator for NF-κB-dependent inflammation in vivo, hence protecting the host against detrimental inflammatory response to NTHi infection

MvaT binds to the PexsC promoter to repress the type III secretion system in Pseudomonas aeruginosa

Pseudomonas aeruginosa is an opportunistic human pathogen capable of causing a variety of acute and chronic infections. Its type III secretion system (T3SS) plays a critical role in pathogenesis during acute infection. ExsA is a master regulator that activates the expression of all T3SS genes. Transcription of exsA is driven by two distinct promoters, its own promoter PexsA and its operon promoter PexsC. Here, in combination with a DNA pull-down assay and mass spectrometric analysis, we found that a histone-like nucleoid-structuring (H-NS) family protein MvaT can bind to the PexsC promoter. Using EMSA and reporter assays, we further found that MvaT directly binds to the PexsC promoter to repress the expression of T3SS genes. The repression of MvaT on PexsC is independent of ExsA, with MvaT binding to the -429 to -380 bp region relative to the transcription start site of the exsC gene. The presented work further reveals the complex regulatory network of the T3SS in P. aeruginosa

Warm Sitz Bath: Are There Benefits after Transurethral Resection of the Prostate?

PURPOSE: We aimed to evaluate the efficacy of warm water sitz baths in patients who have undergone transurethral resection of the prostate (TURP) owing to lower urinary tract symptoms secondary to benign prostatic hyperplasia. MATERIALS AND METHODS: We reviewed the records of 1,783 patients who had undergone TURP between 2001 and 2009. In the warm water sitz bath group, patients were instructed to sit in a tub containing lukewarm water at 40-45degrees C for 10 minutes each time. Patients were advised to perform the procedure for at least 5 days immediately after the removal of a Foley urethral catheter. The differences in post-TURP complications between the warm water sitz bath group and the no sitz bath group were compared. RESULTS: After TURP, 359 of the 1,561 patients performed a warm water sitz bath. Complications after TURP, such as hemorrhage, urinary tract infection, urethral stricture, and acute urinary retention were found in 19 (5.3%) and 75 (6.2%) patients in the sitz bath and no sitz bath groups, respectively (p=0.09). There was a significant difference in postoperative complications such as urethral stricture between the warm sitz bath group and the no sitz bath group (p=0.04). The group that did not undergo warm water sitz bath treatment showed a 1.13-fold increased risk of rehospitalization within 1 month after TURP due to postoperative complications compared with the warm water sitz bath group (odds ratio [OR]=1.134; 95% confidence interval [CI], 1.022 to 1.193; p=0.06). CONCLUSIONS: Warm water sitz bath treatment reduced postoperative complications such as urethral stricture. These results suggest that large-scale prospective studies are needed to establish an ideal method and optimal duration of sitz baths.ope

CRISPLD2 Is a Target of Progesterone Receptor and Its Expression Is Decreased in Women with Endometriosis

Endometriosis, defined as the presence of endometrial cells outside of the uterine cavity, is a major cause of infertility and pelvic pain, afflicting more than 10% of reproductive age women. Endometriosis is a chronic inflammatory disease and lipopolysaccharide promotes the proliferation and invasion of endometriotic stromal cells. Cysteine-rich secretory protein LCCL domain-containing 2 (CRISPLD2) has high affinity for lipopolysaccharide and plays a critical role in defense against endotoxin shock. However, the function of CRISPLD2 has not been studied in endometriosis and uterine biology. Herein, we examined the expression of CRISPLD2 in endometrium from patients with and without endometriosis using immunohistochemistry. The expression of CRISPLD2 was higher in the secretory phase in human menstrual cycle compared to proliferative phase. The expression of CRISPLD2 was significantly decreased in the endometrium of women with endometriosis in the early secretory phase compared to women without endometriosis. The increase of CRISPLD2 expression at the early secretory and dysregulation of its expression in endometriosis suggest progesterone (P4) regulation of CRISPLD2. To investigate whether CRISPLD2 is regulated by P4, we examined the expression of the CRISPLD2 in the uteri of wild-type and progesterone receptor knock out (PRKO) mice. The expression of CRISPLD2 was significantly increased after P4 treatment in the wild-type mice. However, CRISPLD2 expression was significantly decreased in the (PRKO) mice treated with P4. During early pregnancy, the expression of CRISPLD2 was increased in decidua of implantation and post-implantation stages. CRISPLD2 levels were also increased in cultured human endometrial stromal cells during in vitro decidualization. These results suggest that the CRISPLD2 is a target of the progesterone receptor and may play an important role in pathogenesis of endometriosis

Down-Regulation of NF-κB Target Genes by the AP-1 and STAT Complex during the Innate Immune Response in Drosophila

The activation of several transcription factors is required for the elimination of infectious pathogens via the innate immune response. The transcription factors NF-κB, AP-1, and STAT play major roles in the synthesis of immune effector molecules during innate immune responses. However, the fact that these immune responses can have cytotoxic effects requires their tight regulation to achieve restricted and transient activation, and mis-regulation of the damping process has pathological consequences. Here we show that AP-1 and STAT are themselves the major inhibitors responsible for damping NF-κB–mediated transcriptional activation during the innate immune response in Drosophila. As the levels of dAP-1 and Stat92E increase due to continuous immune signaling, they play a repressive role by forming a repressosome complex with the Drosophila HMG protein, Dsp1. The dAP-1–, Stat92E-, and Dsp1-containing complexes replace Relish at the promoters of diverse immune effector genes by binding to evolutionarily conserved cis-elements, and they recruit histone deacetylase to inhibit transcription. Reduction by mutation of dAP-1, Stat92E, or Dsp1 results in hyperactivation of Relish target genes and reduces the viability of bacterially infected flies despite more efficient pathogen clearance. These defects are rescued by reducing the Relish copy number, thus confirming that mis-regulation of Relish, not inadequate activation of dAP-1, Stat92E, or Dsp1 target genes, is responsible for the reduced survival of the mutants. We conclude that an inhibitory effect of AP-1 and STAT on NF-κB is required for properly balanced immune responses and appears to be evolutionarily conserved

Global patient outcomes after elective surgery: prospective cohort study in 27 low-, middle- and high-income countries.

BACKGROUND: As global initiatives increase patient access to surgical treatments, there remains a need to understand the adverse effects of surgery and define appropriate levels of perioperative care. METHODS: We designed a prospective international 7-day cohort study of outcomes following elective adult inpatient surgery in 27 countries. The primary outcome was in-hospital complications. Secondary outcomes were death following a complication (failure to rescue) and death in hospital. Process measures were admission to critical care immediately after surgery or to treat a complication and duration of hospital stay. A single definition of critical care was used for all countries. RESULTS: A total of 474 hospitals in 19 high-, 7 middle- and 1 low-income country were included in the primary analysis. Data included 44 814 patients with a median hospital stay of 4 (range 2-7) days. A total of 7508 patients (16.8%) developed one or more postoperative complication and 207 died (0.5%). The overall mortality among patients who developed complications was 2.8%. Mortality following complications ranged from 2.4% for pulmonary embolism to 43.9% for cardiac arrest. A total of 4360 (9.7%) patients were admitted to a critical care unit as routine immediately after surgery, of whom 2198 (50.4%) developed a complication, with 105 (2.4%) deaths. A total of 1233 patients (16.4%) were admitted to a critical care unit to treat complications, with 119 (9.7%) deaths. Despite lower baseline risk, outcomes were similar in low- and middle-income compared with high-income countries. CONCLUSIONS: Poor patient outcomes are common after inpatient surgery. Global initiatives to increase access to surgical treatments should also address the need for safe perioperative care. STUDY REGISTRATION: ISRCTN5181700

DsbA of Pseudomonas aeruginosa Is Essential for Multiple Virulence Factors

DsbA is a periplasmic thiol:disulfide oxidoreductase which contributes to the process of protein folding by catalyzing the formation of disulfide bonds. In this study, we demonstrate that the dsbA gene is required for the expression of the type III secretion system under low-calcium inducing conditions, intracellular survival of P. aeruginosa upon infection of HeLa cells, and twitching motility. The diverse phenotypes of the dsbA mutant are likely due to its defect in the folding of proteins that are involved in various biological processes, such as signal sensing, protein secretion, and defense against host clearing. In light of its effect on various virulence factors, DsbA could be an important target for the control of P. aeruginosa infections

<i>In Vitro</i> Inflammation Inhibition Model Based on Semi-Continuous Toll-Like Receptor Biosensing

<div><p>A chemical inhibition model of inflammation is proposed by semi-continuous monitoring the density of toll-like receptor 1 (TLR1) expressed on mammalian cells following bacterial infection to investigate an <i>in vivo</i>-mimicked drug screening system. The inflammation was induced by adding bacterial lysate (e.g., <i>Pseudomonas aeruginosa</i>) to a mammalian cell culture (e.g., A549 cell line). The TLR1 density on the same cells was immunochemically monitored up to three cycles under optimized cyclic bacterial stimulation-and-restoration conditions. The assay was carried out by adopting a cell-compatible immunoanalytical procedure and signal generation method. Signal intensity relative to the background control obtained without stimulation was employed to plot the standard curve for inflammation. To suppress the inflammatory response, sodium salicylate, which inhibits nuclear factor-κB activity, was used to prepare the standard curve for anti-inflammation. Such measurement of differential TLR densities was used as a biosensing approach discriminating the anti-inflammatory substance from the non-effector, which was simulated by using caffeic acid phenethyl ester and acetaminophen as the two components, respectively. As the same cells exposed to repetitive bacterial stimulation were semi-continuously monitored, the efficacy and toxicity of the inhibitors may further be determined regarding persistency against time. Therefore, this semi-continuous biosensing model could be appropriate as a substitute for animal-based experimentation during drug screening prior to pre-clinical tests.</p></div

HSP70-Homolog DnaK of Pseudomonas aeruginosa Increases the Production of IL-27 through Expression of EBI3 via TLR4-Dependent NF-κB and TLR4-Independent Akt Signaling

IL-27, a heterodimeric cytokine composed of the p28 subunit and Epstein&ndash;Barr virus-induced gene 3 (EBI3), acts as a potent immunosuppressant and thus limits pathogenic inflammatory responses. IL-27 is upregulated upon Pseudomonas aeruginosa infection in septic mice, increasing susceptibility to the infection and decreasing clearance of the pathogen. However, it remains unclear which P. aeruginosa-derived molecules promote production of IL-27. In this study, we explored the mechanism by which P. aeruginosa DnaK, a heat shock protein 70-like protein, induces EBI3 expression, thereby promoting production of IL-27. Upregulation of EBI3 expression did not lead to an increase in IL-35, which consists of the p35 subunit and EBI3. The IL-27 production in response to DnaK was biologically active, as reflected by stimulation of IL-10 production. DnaK-mediated expression of EBI3 was driven by two distinct signaling pathways, NF-&kappa;B and Akt. However, NF-&kappa;B is linked to TLR4-associated signaling pathways, whereas Akt is not. Taken together, our results reveal that P. aeruginosa DnaK potently upregulates EBI3 expression, which in turn drives production of the prominent anti-inflammatory cytokine IL-27, as a consequence of TLR4-dependent activation of NF-&kappa;B and TLR4-independent activation of the Akt signaling pathway