Alveolar fluid in acute respiratory distress syndrome promotes fibroblast migration: role of platelet-derived growth factor pathway

OBJECTIVES: Fibroblast migration is an initiating step in fibroproliferation; its involvement during acute lung injury and acute respiratory distress syndrome remains poorly understood. The aims of this study were: 1) to determine whether bronchoalveolar lavage fluids from patients with acute lung injury/acute respiratory distress syndrome modulate lung fibroblast migration; 2) to assess lung fibroblast migration\u27s clinical relevance; and 3) to evaluate the role of the platelet-derived growth factor pathway in this effect. DESIGN: Prospective cohort study. SETTING: Three intensive care units of a large tertiary referral center. PATIENTS: Ninety-three ventilated patients requiring bronchoalveolar lavage fluids were enrolled (48 with acute respiratory distress syndrome, 33 with acute lung injury, and 12 ventilated patients without acute lung injury/acute respiratory distress syndrome). INTERVENTIONS: After bronchoalveolar lavage fluids collection during standard care, the patients were followed up for 28 days and clinical outcomes were recorded. Migration assays were performed by using a Transwell model; bronchoalveolar lavage fluids platelet-derived growth factor and soluble platelet-derived growth factor receptor-alpha were characterized by Western blot and measured by ELISA. MEASUREMENTS AND MAIN RESULTS: Most of the bronchoalveolar lavage fluids inhibited basal fibroblast migration. Bronchoalveolar lavage fluids chemotactic index increased with severity of lung injury (28% in patients without acute lung injury/acute respiratory distress syndrome and with acute lung injury vs. 91% in acute respiratory distress syndrome patients; p = .016). In acute lung injury/acute respiratory distress syndrome patients, inhibition of basal fibroblast migration by bronchoalveolar lavage fluids below 52% was independently associated with a lower 28-day mortality (odds ratio [95% confidence interval] 0.313 [0.10-0.98], p = .046). Platelet-derived growth factor-related peptides and soluble platelet-derived growth factor-Ralpha were detected in all bronchoalveolar lavage fluids from acute lung injury/acute respiratory distress syndrome patients. The effect of bronchoalveolar lavage fluids stimulating migration was inhibited by a specific platelet-derived growth factor receptor inhibitor (AG1296). Bronchoalveolar lavage fluids inhibiting migration reversed the effect of rh-platelet-derived growth factor-BB and reduced by 40% the binding of 125I-platelet-derived growth factor-BB to fibroblast cell surface in favor of a role for platelet-derived growth factor-sRalpha. CONCLUSIONS: : Together, our results suggest that during acute lung injury, fibroblast migration is modulated by bronchoalveolar lavage fluids through a platelet-derived growth factor/platelet-derived growth factor-sRalpha balance. Migration is associated with clinical severity and patient 28-day mortality

Hepatocyte and keratinocyte growth factors and their receptors in human lung emphysema

BACKGROUND: Hepatocyte and keratinocyte growth factors are key growth factors in the process of alveolar repair. We hypothesized that excessive alveolar destruction observed in lung emphysema involves impaired expression of hepatocyte and keratinocyte growth factors or their respective receptors, c-met and keratinocyte growth factor receptor. The aim of our study was to compare the expression of hepatocyte and keratinocyte growth factors and their receptors in lung samples from 3 groups of patients: emphysema; smokers without emphysema and non-smokers without emphysema. METHODS: Hepatocyte and keratinocyte growth factor proteins were analysed by immunoassay and western blot; mRNA expression was measured by real time quantitative polymerase chain reaction. RESULTS: Hepatocyte and keratinocyte growth factors, c-met and keratinocyte growth factor receptor mRNA levels were similar in emphysema and non-emphysema patients. Hepatocyte growth factor mRNA correlated negatively with FEV1 and the FEV1/FVC ratio both in emphysema patients and in smokers with or without emphysema. Hepatocyte and keratinocyte growth factor protein concentrations were similar in all patients' groups. CONCLUSION: The expression of hepatocyte and keratinocyte growth factors and their receptors is preserved in patients with lung emphysema as compared to patients without emphysema. Hepatocyte growth factor mRNA correlates with the severity of airflow obstruction in smokers

Interleukin 6, lipopolysaccharide-binding protein and interleukin 10 in the prediction of risk and etiologic patterns in patients with community-acquired pneumonia: results from the German competence network CAPNETZ

<p>Abstract</p> <p>Background</p> <p>The aim of our study was to investigate the predictive value of the biomarkers interleukin 6 (IL-6), interleukin 10 (IL-10) and lipopolysaccharide-binding protein (LBP) compared with clinical CRB and CRB-65 severity scores in patients with community-acquired pneumonia (CAP).</p> <p>Methods</p> <p>Samples and data were obtained from patients enrolled into the German CAPNETZ study group. Samples (blood, sputum and urine) were collected within 24 h of first presentation and inclusion in the CAPNETZ study, and CRB and CRB-65 scores were determined for all patients at the time of enrollment. The combined end point representative of a severe course of CAP was defined as mechanical ventilation, intensive care unit treatment and/or death within 30 days. Overall, a total of 1,000 patients were enrolled in the study. A severe course of CAP was observed in 105 (10.5%) patients.</p> <p>Results</p> <p>The highest IL-6, IL-10 and LBP concentrations were found in patients with CRB-65 scores of 3-4 or CRB scores of 2-3. IL-6 and LBP levels on enrollment in the study were significantly higher for patients with a severe course of CAP than for those who did not have severe CAP. In receiver operating characteristic analyses, the area under the curve values for of IL-6 (0.689), IL-10 (0.665) and LPB (0.624) in a severe course of CAP were lower than that of CRB-65 (0.764) and similar to that of CRB (0.69). The accuracy of both CRB and CRB-65 was increased significantly by including IL-6 measurements. In addition, higher cytokine concentrations were found in patients with typical bacterial infections compared with patients with atypical or viral infections and those with infection of unknown etiology. LBP showed the highest discriminatory power with respect to the etiology of infection.</p> <p>Conclusions</p> <p>IL-6, IL-10 and LBP concentrations were increased in patients with a CRB-65 score of 3-4 and a severe course of CAP. The concentrations of IL-6 and IL-10 reflected the severity of disease in patients with CAP. The predictive power of IL-6, IL-10 and LBP for a severe course of pneumonia was lower than that of CRB-65. Typical bacterial pathogens induced the highest LBP, IL-6 and IL-10 concentrations.</p

SNARE Protein Mimicry by an Intracellular Bacterium

Many intracellular pathogens rely on host cell membrane compartments for their survival. The strategies they have developed to subvert intracellular trafficking are often unknown, and SNARE proteins, which are essential for membrane fusion, are possible targets. The obligate intracellular bacteria Chlamydia replicate within an intracellular vacuole, termed an inclusion. A large family of bacterial proteins is inserted in the inclusion membrane, and the role of these inclusion proteins is mostly unknown. Here we identify SNARE-like motifs in the inclusion protein IncA, which are conserved among most Chlamydia species. We show that IncA can bind directly to several host SNARE proteins. A subset of SNAREs is specifically recruited to the immediate vicinity of the inclusion membrane, and their accumulation is reduced around inclusions that lack IncA, demonstrating that IncA plays a predominant role in SNARE recruitment. However, interaction with the SNARE machinery is probably not restricted to IncA as at least another inclusion protein shows similarities with SNARE motifs and can interact with SNAREs. We modelled IncA's association with host SNAREs. The analysis of intermolecular contacts showed that the IncA SNARE-like motif can make specific interactions with host SNARE motifs similar to those found in a bona fide SNARE complex. Moreover, point mutations in the central layer of IncA SNARE-like motifs resulted in the loss of binding to host SNAREs. Altogether, our data demonstrate for the first time mimicry of the SNARE motif by a bacterium

STAT3 Regulates Monocyte TNF-Alpha Production in Systemic Inflammation Caused by Cardiac Surgery with Cardiopulmonary Bypass

BACKGROUND: Cardiopulmonary bypass (CPB) surgery initiates a controlled systemic inflammatory response characterized by a cytokine storm, monocytosis and transient monocyte activation. However, the responsiveness of monocytes to Toll-like receptor (TLR)-mediated activation decreases throughout the postoperative course. The purpose of this study was to identify the major signaling pathway involved in plasma-mediated inhibition of LPS-induced tumor necrosis factor (TNF)-α production by monocytes. METHODOLOGY/PRINCIPAL FINDINGS: Pediatric patients that underwent CPB-assisted surgical correction of simple congenital heart defects were enrolled (n = 38). Peripheral blood mononuclear cells (PBMC) and plasma samples were isolated at consecutive time points. Patient plasma samples were added back to monocytes obtained pre-operatively for ex vivo LPS stimulations and TNF-α and IL-6 production was measured by flow cytometry. LPS-induced p38 mitogen-activated protein kinase (MAPK) and nuclear factor (NF)-κB activation by patient plasma was assessed by Western blotting. A cell-permeable peptide inhibitor was used to block STAT3 signaling. We found that plasma samples obtained 4 h after surgery, regardless of pre-operative dexamethasone treatment, potently inhibited LPS-induced TNF-α but not IL-6 synthesis by monocytes. This was not associated with attenuation of p38 MAPK activation or IκB-α degradation. However, abrogation of the IL-10/STAT3 pathway restored LPS-induced TNF-α production in the presence of suppressive patient plasma. CONCLUSIONS/SIGNIFICANCE: Our findings suggest that STAT3 signaling plays a crucial role in the downregulation of TNF-α synthesis by human monocytes in the course of systemic inflammation in vivo. Thus, STAT3 might be a potential molecular target for pharmacological intervention in clinical syndromes characterized by systemic inflammation

Relating circulating thyroid hormone concentrations to serum interleukins-6 and -10 in association with non-thyroidal illnesses including chronic renal insufficiency

<p>Abstract</p> <p>Background</p> <p>Because of the possible role of cytokines including interleukins (IL) in systemic non-thyroidal illnesses' (NTI) pathogenesis and consequently the frequently associated alterations in thyroid hormone (TH) concentrations constituting the euthyroid sick syndrome (ESS), we aimed in this research to elucidate the possible relation between IL-6 & IL-10 and any documented ESS in a cohort of patients with NTI.</p> <p>Methods</p> <p>Sixty patients and twenty healthy volunteers were recruited. The patients were subdivided into three subgroups depending on their underlying NTI and included 20 patients with chronic renal insufficiency (CRI), congestive heart failure (CHF), and ICU patients with myocardial infarction (MI). Determination of the circulating serum levels of IL-6 and IL-10, thyroid stimulating hormone (TSH), as well as total T4 and T3 was carried out.</p> <p>Results</p> <p>In the whole group of patients, we detected a significantly lower T3 and T4 levels compared to control subjects (0.938 ± 0.477 vs 1.345 ± 0.44 nmol/L, p = 0.001 and 47.9 ± 28.41 vs 108 ± 19.49 nmol/L, p < 0.0001 respectively) while the TSH level was normal (1.08+0.518 μIU/L). Further, IL-6 was substantially higher above controls' levels (105.18 ± 72.01 vs 3.35 ± 1.18 ng/L, p < 0.00001) and correlated negatively with both T3 and T4 (r = -0.620, p < 0.0001 & -0.267, p < 0.001, respectively). Similarly was IL-10 level (74.13 ± 52.99 vs 2.64 ± 0.92 ng/ml, p < 0.00001) that correlated negatively with T3 (r = -0.512, p < 0.0001) but not T4. Interestingly, both interleukins correlated positively (r = 0.770, p = <0.001). Moreover, IL-6 (R²= 0.338, p = 0.001) and not IL-10 was a predictor of low T3 levels with only a borderline significance for T4 (R²= 0.082, p = 0.071).</p> <p>By subgroup analysis, the proportion of patients with subnormal T3, T4, and TSH levels was highest in the MI patients (70%, 70%, and 72%, respectively) who displayed the greatest IL-6 and IL-10 concentrations (192.5 ± 45.1 ng/L & 122.95 ± 46.1 ng/L, respectively) compared with CHF (82.95 ± 28.9 ng/L & 69.05 ± 44.0 ng/L, respectively) and CRI patients (40.05 ± 28.9 ng/L & 30.4 ± 10.6 ng/L, respectively). Surprisingly, CRI patients showed the least disturbance in IL-6 and IL-10 despite the lower levels of T3, T4, and TSH in a higher proportion of them compared to CHF patients (40%, 45%, & 26% vs 35%, 25%, & 18%, respectively).</p> <p>Conclusion</p> <p>the high prevalence of ESS we detected in NTI including CRI may be linked to IL-6 and IL-10 alterations. Further, perturbation of IL-6 and not IL-10 might be involved in ESS pathogenesis although it is not the only key player as suggested by our findings in CRI.</p

Is adenomyosis the neglected phenotype of an endomyometrial dysfunction syndrome?

Since the dissociation between adenomyoma and endometriosis in the 1920s and the laparoscopic progress in the diagnosis and surgery of endometriosis, the literature has been greatly focused on the disease endometriosis. The study of adenomyosis, on the other hand, has been neglected as the diagnosis remained based on hysterectomy specimens. However, since the introduction of magnetic resonance and sonographic imaging techniques in the 1980s, the myometrial junctional zone has been identified as a third uterine zone and interest in adenomyosis was renewed. This has also been the start for the interest in the role of the myometrial junctional zone dysfunction and adenomyosis in reproductive and obstetrical disorders

Gene Expression Profiling in the Type 1 Diabetes Rat Diaphragm

BACKGROUND:Respiratory muscle contractile performance is impaired by diabetes, mechanisms of which included altered carbohydrate and lipid metabolism, oxidative stress and changes in membrane electrophysiology. The present study examined to what extent these cellular perturbations involve changes in gene expression. METHODOLOGY/PRINCIPAL FINDINGS:Diaphragm muscle from streptozotocin-diabetic rats was analyzed with Affymetrix gene expression arrays. Diaphragm from diabetic rats had 105 genes with at least +/-2-fold significantly changed expression (55 increased, 50 decreased), and these were assigned to gene ontology groups based on over-representation analysis using DAVID software. There was increased expression of genes involved in palmitoyl-CoA hydrolase activity (a component of lipid metabolism) (P = 0.037, n = 2 genes, fold change 4.2 to 27.5) and reduced expression of genes related to carbohydrate metabolism (P = 0.000061, n = 8 genes, fold change -2.0 to -8.5). Other gene ontology groups among upregulated genes were protein ubiquitination (P = 0.0053, n = 4, fold change 2.2 to 3.4), oxidoreductase activity (P = 0.024, n = 8, fold change 2.1 to 6.0), and morphogenesis (P = 0.012, n = 10, fold change 2.1 to 4.3). Other downregulated gene groups were extracellular region (including extracellular matrix and collagen) (P = 0.00032, n = 13, fold change -2.2 to -3.7) and organogenesis (P = 0.032, n = 7, fold change -2.1 to -3.7). Real-time PCR confirmed the directionality of changes in gene expression for 30 of 31 genes tested. CONCLUSIONS/SIGNIFICANCE:These data indicate that in diaphragm muscle type 1 diabetes increases expression of genes involved in lipid energetics, oxidative stress and protein ubiquitination, decreases expression of genes involved in carbohydrate metabolism, and has little effect on expression of ion channel genes. Reciprocal changes in expression of genes involved in carbohydrate and lipid metabolism may change the availability of energetic substrates and thereby directly modulate fatigue resistance, an important issue for a muscle like the diaphragm which needs to contract without rest for the entire lifetime of the organism

Multi-genome identification and characterization of chlamydiae-specific type III secretion substrates: the Inc proteins

<p>Abstract</p> <p>Background</p> <p><it>Chlamydiae </it>are obligate intracellular bacteria that multiply in a vacuolar compartment, the inclusion. Several chlamydial proteins containing a bilobal hydrophobic domain are translocated by a type III secretion (TTS) mechanism into the inclusion membrane. They form the family of Inc proteins, which is specific to this phylum. Based on their localization, Inc proteins likely play important roles in the interactions between the microbe and the host. In this paper we sought to identify and analyze, using bioinformatics tools, all putative Inc proteins in published chlamydial genomes, including an environmental species.</p> <p>Results</p> <p>Inc proteins contain at least one bilobal hydrophobic domain made of two transmembrane helices separated by a loop of less than 30 amino acids. Using bioinformatics tools we identified 537 putative Inc proteins across seven chlamydial proteomes. The amino-terminal segment of the putative Inc proteins was recognized as a functional TTS signal in 90% of the <it>C. trachomatis </it>and <it>C. pneumoniae </it>sequences tested, validating the data obtained <it>in silico</it>. We identified a <it>macro </it>domain in several putative Inc proteins, and observed that Inc proteins are enriched in segments predicted to form coiled coils. A surprisingly large proportion of the putative Inc proteins are not constitutively translocated to the inclusion membrane in culture conditions.</p> <p>Conclusions</p> <p>The Inc proteins represent 7 to 10% of each proteome and show a great degree of sequence diversity between species. The abundance of segments with a high probability for coiled coil conformation in Inc proteins support the hypothesis that they interact with host proteins. While the large majority of Inc proteins possess a functional TTS signal, less than half may be constitutively translocated to the inclusion surface in some species. This suggests the novel finding that translocation of Inc proteins may be regulated by as-yet undetermined mechanisms.</p