A NOD2-NALP1 complex mediates caspase-1-dependent IL-1beta secretion in response to Bacillus anthracis infection and muramyl dipeptide

NOD2, a NOD-like receptor (NLR), is an intracellular sensor of bacterial muramyl dipeptide (MDP) that was suggested to promote secretion of the proinflammatory cytokine IL-1beta. Yet, the molecular mechanism by which NOD2 can stimulate IL-1beta secretion, and its biological significance were heretofore unknown. We found that NOD2 through its N-terminal caspase recruitment domain directly binds and activates caspase-1 to trigger IL-1beta processing and secretion in MDP-stimulated macrophages, whereas the C-terminal leucine-rich repeats of NOD2 prevent caspase-1 activation in nonstimulated cells. MDP challenge induces the association of NOD2 with another NLR protein, NALP1, and gel filtration analysis revealed the formation of a complex consisting of NOD2, NALP1, and caspase-1. Importantly, Bacillus anthracis infection induces IL-1beta secretion in a manner that depended on caspase-1 and NOD2. In vitro, Anthrax lethal toxin strongly potentiated IL-1beta secretion, and that response was NOD2 and caspase-1-dependent. Thus, NOD2 plays a key role in the B. anthracis-induced inflammatory response by being a critical mediator of IL-1beta secretion.     

Effect of relaxin on aromatase activity in human endometrial stromal cells

Previous studies have shown that the aromatase activity in human endometrial stromal cells was stimulated by progestin and enhanced by estrogen and forskolin (Fk), an agent that stimulates the accumulation of intracellular cAMP. Present study was undertaken to investigate whether any peptide hormone would affect endometrial aromatase activity. Stromal cells were isolated from normal proliferative and secretory endometria and cultured in nutrient medium. Porcine relaxin (RLX) was added to culture medium individually or in combination with medroxyprogesterone acetate (MPA) and estradiol (E2). Cells treated with RLX alone did not affect the aromatase activity. RLX, however, exerted a synergistic effect on aromatase activity in the presence of MPA or MPA plus E2. On the other hand, human CG, epidermal growth factor, human PRL, and insulin did not increase the aromatase activity in the presence or absence of MPA and E2 studied in a limited number of specimens. The progestin-dependent effect of RLX on aromatase activity was dose dependent indicating that the biological effect of RLX is mediated through a saturable mechanism. When RLX was added to MPA-pretreated cells, additional increase of aromatase activity was seen after 24 h incubation indicating that the action of RLX on stromal cells is not an acute effect. Antiprogestin, RU486, inhibited the stimulation of aromatase activity in both MPA and MPA plus RLX treated cells. RLX has either no effect or a moderate increase (up to 2-fold over the control) on intracellular cAMP content. On the other hand, Fk increased the intracellular cAMP level and enhanced the aromatase activity in the presence of progestin. Also RLX did not replace the effect of Fk since additional increase of aromatase activity was noted when stromal cells were incubated with MPA plus RLX plus Fk in comparison to MPA plus RLX or MPA plus Fk. These results suggest that the action of RLX on stromal cells may be mediated through an intracellular messenger independent of cAMP. Present studies provide evidence that RLX exerts a synergistic effect on aromatase activity in the presence of progestin in human endometrial stromal cells. It is evident that human endometrium is a target organ of RLX.     

Renin-angiotensin system component expression in the HL-1 atrial cell line and in a pig model of atrial fibrillation

OBJECTIVES: Local atrial tissue angiotensin II (AngII) level is elevated in atrial fibrillation (AF), but the mechanism is unknown. We hypothesized that atrial myocytes express all components of the renin-angiotensin system (RAS) and investigated whether rapid depolarization alone is sufficient to increase paracrine AngII production by up-regulating RAS component expression. METHODS: In the HL-1 atrial cell line, rapid depolarization was induced by rapid field electrical stimulation (RES) at 1.0 V/cm and 600/min (10 Hz) in atrial HL-1 cells. In a pig model of AF, AF was induced by atrial pacing at 600/min in 10 adult pigs and 10 sham-operated pigs for comparison. RESULTS: In atrial myocytes, RES induced a sustained elevation of intracellular calcium, and up-regulation of angiotensin-converting enzyme (ACE), chymase and angiotensinogen, resulting in increased AngII production. RES-induced AngII production was attenuated by enalapril [ACE inhibitor (ACEI)] and chymostatin (chymase inhibitor). Conditioned medium from RES-stimulated atrial myocytes increased [3H]leucine uptake and atrial natriuretic peptide expression in atrial myocytes, and [3H]proline uptake and collagen type 1 alpha 1 expression in atrial fibroblasts. Both were attenuated by co-incubation with the AngII type 1 receptor blocker (ARB) losartan. In the porcine model, significant structural changes and a similar pattern of changes of RAS components were noted in AF pigs. CONCLUSIONS: Atrial cells expressed all components of RAS and rapid depolarization alone was sufficient to up-regulate RAS components, increase paracrine AngII production and induce atrial structural changes, which are attenuated by ACEI, ARB and chymase inhibitor.     

Association of the human minK gene 38G allele with atrial fibrillation: evidence of possible genetic control on the pathogenesis of atrial fibrillation

Background Human minK protein is the β-subunit of I_Ks potassium channel and plays an important role in cardiac cellular electrophysiology. We investigated the association between human atrial fibrillation and the polymorphism of minK gene (38G or 38S) with a case-control study. Methods We included 108 patients with atrial fibrillation and 108 control subjects. The case patients and control subjects were matched regarding age, sex, presence of valvular heart disease, and presence of left ventricular dysfunction. The genotype of minK was determined with polymerase chain reaction and restriction fragment analysis. Results The results showed an association between the minK 38G allele and atrial fibrillation. The odds ratios for atrial fibrillation in patients with 1 and 2 minK 38G alleles were 2.16 (95% CI 0.81-5.74) and 3.58 (95% CI 1.38-9.27), respectively, when compared with patients without minK 38G allele. In a logistic regression model, the odds ratio for atrial fibrillation was 1.80 (95% CI 1.20-2.71, P < .0046) for patients with 1 more minK 38G allele. Conclusion We report the association between the minK 38G allele and clinical atrial fibrillation. Our findings suggest possible genetic control on the pathogenesis of atrial fibrillation. (Am Heart J 2002;144:485-90.)     

Effect of autologous blood donation on the central venous pressure, blood loss and blood transfusion during living donor left hepatectomy

AIM: Autologous blood donation (ABD) is mainly used to reduce the use of banked blood. In fact, ABD can be regarded as acute blood loss. Would ABD 2-3 d before operation affect the CVP level and subsequently result in less blood loss during liver resection was to be determined. METHODS: Eighty-four patients undergoing living donor left hepatectomy were retrospectively divided as group Ⅰ (GI) and group Ⅱ (Gil) according to have donated 250-300 mL blood 2-3 d before living donor hepatectomy or not. The changes of the intraoperative CVP, surgical blood loss, blood products used and the changes of perioperative hemoglobin (Hb) between groups were analyzed and compared by using Mann-Whitney U test. RESULTS: The results show that the intraoperative CVP changes between GI (n = 35) and GII (n = 49) up to graft procurement were the same, subsequently the blood loss, but ABD resulted in significantly lower perioperative Hb levels in GI. CONCLUSION: Since none of the patients required any blood products perioperatively, all the predonated bloods were discarded after the patients were discharged from the hospital. It indicates that ABD in current series had no any beneficial effects, in term of cost, lowering the CVP, blood loss and reduce the use of banked blood products, but resulted in significant lower Hb in perioperative period.     

Metallothionein 1 h tumour suppressor activity in prostate cancer is mediated by euchromatin methyltransferase 1

Metallothioneins (MTs) are a group of metal binding proteins thought to play a role in the detoxification of heavy metals. Here we showed by microarray and validation analyses that MT1h, a member of MT, is down‐regulated in many human malignancies. Low expression of MT1h was associated with poor clinical outcomes in both prostate and liver cancer. We found that the promoter region of MT1h was hypermethylated in cancer and that demethylation of the MT1h promoter reversed the suppression of MT1h expression. Forced expression of MT1h induced cell growth arrest, suppressed colony formation, retarded migration, and reduced invasion. SCID mice with tumour xenografts with inducible MT1h expression had lower tumour volumes as well as fewer metastases and deaths than uninduced controls. MT1h was found to interact with euchromatin histone methyltransferase 1 (EHMT1) and enhanced its methyltransferase activity on histone 3. Knocking down of EHMT1 or a mutation in MT1h that abrogates its interaction with EHMT1 abrogated MT1h tumour suppressor activity. This demonstrates tumour suppressor activity in a heavy metal binding protein that is dependent on activation of histone methylation. Copyright © 2013 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Levels of Circulating Microparticles in Lung Cancer Patients and Possible Prognostic Value

Background. Endothelial-derived microparticles (EDMPs) and platelet-derived microparticles (PDMPs) have been reported to be increasing in various diseases including malignant diseases. Here, we investigated whether these MPs may be useful biomarkers for predicting lung cancer (LC) disease status, cell type, or metastasis. Methods and Results. One hundred and thirty LC patients were prospectively enrolled into the study between April 2011 and February 2012. Flow cytometric analysis demonstrated that the circulating levels of platelet-derived activated MPs (PDAc-MPs), platelet-derived apoptotic MPs (PDAp-MPs), endothelial-derived activated MPs (EDAc-MPs), and endothelial-derived apoptotic MPs (EDAp-MPs) were significantly higher in LC patients than in 30 age- and gender-matched normal control subjects (all P < 0.05). Additionally, circulating level of PDAc-MPs was significantly lower (P = 0.031), whereas the circulating levels of the other three biomarkers did not differ (all P > 0.1) in early stage versus late stage LC patients. Furthermore, the circulating levels of the four types of MPs did not differ among patients with different disease statuses (i.e., disease controlled, disease progression, and disease without treatment, i.e., fresh case) (all P > 0.2) or between patients with or without LC metastasis (all P > 0.5). Moreover, only the circulating level of EDAp-MPs was significantly associated with the different cell types (i.e., squamous cell carcinoma, adenocarcinoma, and small cell carcinoma) of LC (P = 0.045). Conclusion. Circulating MP levels are significantly increased in LC patients as compared with normal subjects. Among the MPs, only an increased level of EDAp-MPs was significantly associated with different LC cell types.     

Daily Life Events Influence the Results of the Dexamethasone Suppression Test in Healthy Women

Although the Dexamethasone Suppression Test (DST) plays an important role in psychosomatic research, confounding factors limit the sensitivity and specificity of the DST. The aim of this study was to investigate the relationship between the intensity of daily life stressors and DST results in healthy participants after controlling the confounding factors. The subjects of this study consisted of 75 healthy volunteers. The intensity of daily life events was assessed using the Taiwanese version of the Recent Life Change Questionnaire (RLCQ). Neuroticism was assessed using the Maudsley Personality Inventory (MPI). The Dexamethasone Suppression Test (DST) was also performed. The regression model showed that daily life events (RLCQ score) were correlated significantly with cortisol level on day 1 and D% only in women. This finding implies that daily life events should be considered as an independent variable in women in further studies when the DST is applied.     

Tri-layered chitosan scaffold as a potential skin substitute

A tri-layered chitosan-based scaffold was successfully made to replicate the striation of a full-thickness skin more accurately than a single- or bi-layered scaffold, which needed weeks of co-culturing of fibroblasts and keratinocytes to achieve similar striation. Chitosan solution was freeze-dried and made into porous disks. Chitosan or chitosan–pectin in acetic acid solution was electrospun onto the chitosan disk to form a nanofibrous layer and a thin film. Examinations based on scanning electron spectroscopy showed that the scaffold was composed of a porous layer (2 mm) to simulate the dermis, a thin film (25–45 μm) to mimic the basement membrane, and a layer of nanofibers (100–200 μm) to serve as the protective epidermis. The tensile strength and modulus of the composite scaffold were significantly higher than those of the chitosan disk (p < 0.01). The composite was able to quickly absorb water and stayed intact throughout the course of the 14-day cell culture tests. The fibroblast cells seeded on both sides of the scaffolds were able to proliferate and stayed separated by the thin film.     

Pectin-chitosan-PVA nanofibrous scaffold made by electrospinning and its potential use as a skin tissue scaffold

Scaffolds made of chitosan nanofibers are often too mechanically weak for their application and often their manufacturing processes involve the use of harmful and flammable organic solvents. In the attempt to improve the mechanical properties of nanofibrous scaffolds made of chitosan without the use of harmful chemicals, pectin, an anionic polymer was blended with chitosan, a cationic polymer, to form a polyelectrolyte complex and electrospun into nanofibers for the first time. The electrospun chitosan-pectin scaffolds, when compared to electrospun chitosan scaffolds, had a 58% larger diameter, a 21% higher Young’s modulus, a 162% larger strain at break, and a 104% higher ultimate tensile strength. Compared to the chitosan scaffolds, the chitosan-pectin scaffolds’ swelling ratios decreased by 55% after 60?min in a saline solution and more quickly released the preloaded tetracycline HCl. The L929 fibroblast cells proliferated slightly slower on the chitosan-pectin scaffolds than on the chitosan scaffolds. Nonetheless, cells on both materials deposited similar levels of extracellular type I collagen on a per DNA basis. In conclusion, a novel chitosan-pectin nanofibrous scaffold with superior mechanical properties than a chitosan nanofibrous scaffold was successfully made without the use of harmful solvents.