Gut bacteria play a key role in the metabolism of dietary isoflavones, thereby influencing the availability and bioactivation of these polyphenols in the intestine. The human intestinal bacterium Slackia isoflavoniconvertens converts the main soybean isoflavones daidzein and genistein to equol and 5-hydroxy-equol, respectively. Cell extracts of S. isoflavoniconvertens catalyzed the conversion of daidzein via dihydrodaidzein to equol and that of genistein to dihydrogenistein. Growth of S. isoflavoniconvertens in the presence of daidzein led to the induction of several proteins as observed by two-dimensional difference gel electrophoresis. Based on determined peptide sequences, we identified a cluster of eight genes encoding the daidzein-induced proteins. Heterologous expression of three of these genes in Escherichia coli and enzyme activity tests with the resulting cell extracts identified the corresponding gene products as a daidzein reductase (DZNR), a dihydrodaidzein reductase (DHDR), and a tetrahydrodaidzein reductase (THDR). The recombinant DZNR also converted genistein to dihydrogenistein at higher rates than were observed for the conversion of daidzein to dihydrodaidzein. Higher rates were also observed with cell extracts of S. isoflavoniconvertens. The recombinant DHDR and THDR catalyzed the reduction of dihydrodaidzein to equol, while the corresponding conversion of dihydrogenistein to 5-hydroxy-equol was not observed. The DZNR, DHDR, and THDR were expressed as Strep-tag fusion proteins and subsequently purified by affinity chromatography. The purified enzymes were further characterized with regard to their activity, stereochemistry, quaternary structure, and content of flavin cofactors.

Recruitment of polymorphonuclear neutrophils (PMNs) remains a paramount prerequisite in innate immune defense and a critical cofounder in inflammatory vascular disease. Neutrophil recruitment comprises a cascade of concerted events allowing for capture, adhesion and extravasation of the leukocyte. Whereas PMN rolling, binding, and diapedesis are well characterized, receptor-mediated processes, mechanisms attenuating the electrostatic repulsion between the negatively charged glycocalyx of leukocyte and endothelium remain poorly understood. We provide evidence for myeloperoxidase (MPO), an abundant PMN-derived heme protein, facilitating PMN recruitment by its positive surface charge. In vitro, MPO evoked highly directed PMN motility, which was solely dependent on electrostatic interactions with the leukocyte's surface. In vivo, PMN recruitment was shown to be MPOdependent in a model of hepatic ischemia and reperfusion, upon intraportal delivery of MPO and in the cremaster muscle exposed to local inflammation or to intraarterial MPO application. Given MPO's affinity to both the endothelial and the leukocyte's surface, MPO evolves as a mediator of PMN recruitment because of its positive surface charge. This electrostatic MPO effect not only displays a so far unrecognized, catalysis-independent function of the enzyme, but also highlights a principal mechanism of PMN attraction driven by physical forces. (Blood. 2011; 117(4):1350-1358) IntroductionRecruitment of polymorphonuclear neutrophils (PMNs) is considered a hallmark in host defense. 1 With vessel-and tissue-infiltrating PMNs also being mechanistically linked to a broad range of vascular inflammatory diseases including coronary artery disease, heart failure and ischemia/reperfusion injury, the pathophysiologic significance of PMN motility reaches far beyond innate immunity. [2][3][4] So far, PMN migration is primarily viewed to be energydependent and cytoskeleton-dependent with G-protein coupled receptors and integrins initiating and orchestrating signaling pathways obligatory for neutrophil adhesion, spreading, diapedesis and chemotactic agitation. [5][6][7] On activation, PMN releases myeloperoxidase (MPO), an abundant heme protein in PMN with potent bactericidal and vascular-inflammatory properties. The enzyme accumulates along the endothelium and in the subendothelial space, 8 where it binds to anionic glycocalyx residues such as heparan sulfate glycosaminoglycans. Given the affinity of MPO to both PMN and endothelial cells, we evaluated whether MPO affects PMN locomotion and recruitment. Methods Isolation of PMNPeripheral blood was drawn from healthy human volunteers and heparinized, and isolation of PMN was performed as previously described. 9 In brief, after sedimentation in dextran solution (45 mg/mL), the supernatant was placed over Histopaque 1077 (Sigma-Aldrich) for density gradient centrifugation. Remaining red blood cells were eliminated by hypotonic lysis and the pellet was resuspended in Hanks balanced salt solution (HBSS; Invitrogen) containing 0...

In our group of patients, ICAM-1 expression was associated with a more aggressive tumour phenotype. Because of its association with malignant progression, ICAM-1 might represent a new target in the treatment of breast cancer patients.

To comprehend emotional prosodic cues in speech is a critical function of human social life. However, it is common in everyday communication that conflicting information in emotional prosody and semantic content co-occur. Here, we sought to specify brain regions involved in conflict monitoring of these interfering communication channels. By means of functional magnetic resonance imaging, we obtained signal increases in the right dorsal anterior cingulate cortex and right superior temporal gyrus (STG) and superior temporal sulcus when participants listened to incongruous compared with congruous sentences. Moreover, valence-specific effects were found in the left inferior frontal gyrus and left STG for happily intoned sentences expressing a negative content. The left caudate nucleus along with the thalamus was active when angrily intoned sentences were coupled with positive semantic content. Our results suggest a brain network that monitors conflict in emotional prosody and emotional semantic content comprising of medial prefrontal areas that have previously been associated with cognitive conflict processing. Furthermore, our study extends the knowledge of these processes by suggesting valence-specific differences of emotional conflict processing.

Background:C-Fos was initially described as oncogene, but was associated with favourable prognosis in ovarian cancer (OvCa) patients. The molecular and functional aspects underlying this effect are still unknown.Methods:Using stable transfectants of SKOV3 and OVCAR8 cells, proliferation, migration, invasion and apoptotic potential of c-FOS-overexpressing clones and controls were compared. Adherence to components of the extracellular matrix was analysed in static assays, and adhesion to E-selectin, endothelial and mesothelial cells in dynamic flow assays. The effect of c-FOS in vivo was studied after intraperitoneal injection of SKOV3 clones into SCID mice, and changes in gene expression were determined by microarray analysis.Results:Tumour growth after injection into SCID mice was strongly delayed by c-FOS overexpression, with reduction of lung metastases and circulating tumour cells. In vitro, c-FOS had only weak influence on proliferation and migration, but was strongly pro-apoptotic. Adhesion to components of the extracellular matrix (collagen I, IV) and to E-selectin, endothelial and mesothelial cells was significantly reduced in c-FOS-overexpressing OvCa cells. This corresponds to deregulation of adhesion proteins and glycosylation enzymes in microarray analysis.Conclusion:In addition to its known pro-apoptotic effect, c-FOS might influence OvCa progression by changing the adhesion of OvCa cells to peritoneal surfaces.

Mesenchymal stromal cells (MSCs) have been isolated from many tissues, including gestational tissue. To date, a study comparing the properties and suitability of these cells in cell-based therapies is lacking. In this study, we compared the phenotype, proliferation rate, migration, immunogenicity, and immunomodulatory capabilities of human MSCs derived from umbilical cord lining (CL-MSCs), umbilical cord blood (CB-MSCs), placenta (P-MSCs), and Wharton's jelly (WJ-MSCs). Differences were noted in differentiation, proliferation, and migration, with CL-MSCs showing the highest proliferation and migration rates resulting in prolonged survival in immunodeficient mice. Moreover, CL-MSCs showed a prolongation in survival in xenogeneic BALB/c mice, which was attributed to their ability to dampen TH1 and TH2 responses. Weaker human cellular immune responses were detected against CL-MSCs and P-MSCs, which were correlated with their lower HLA I expression. Furthermore, HLA II was upregulated less substantially by CL-MSCs and CB-MSCs after IFN-γ stimulation. MSC types did not differ in indolamine 2,3-dioxygenase (IDO) expression after IFN-γ stimulation. Despite their lower IDO, HLA-G, and TGF-β1 expression, only CL-MSCs were able to reduce the release of IFN-γ by lymphocytes in a mixed lymphocyte reaction. In summary, CL-MSCs showed the best characteristics for cell-based strategies, as they are hypo-immunogenic and show high proliferation and migration rates. In addition, these studies show for the first time that although immunomodulatory molecules HLA-G, HLA-E, and TGF-β play an important role in MSC immune evasion, basal and induced HLA expression seems to be decisive in determining the immunogenicity of MSCs.

Activated Leukocyte Cell Adhesion Molecule (ALCAM, also called CD 166, MEMD) as cell surface immunoglobulin is reported as prognostic marker in breast cancer, but its predictive value has not yet been evaluated. We have analyzed ALCAM protein expression by Western Blot analysis (n = 160) and mRNA expression by cDNA microarray analysis (n = 162) in primary mammary carcinomas. Both expression results were obtained in 133 cases, showing a strong positive correlation between protein expression and mRNA expression (P < 0.001). Neither ALCAM protein nor mRNA expression are correlated to histological type, grading, stage or age of patient. However, ALCAM protein expression correlates positively with estrogen receptor status (ER) (P = 0.025). A stratified subgroup analysis showed positive correlation of high ALCAM mRNA expression with longer overall survival (OAS; P = 0.0012) in patients treated with adjuvant chemotherapy (n = 100). In contrast, patients with high ALCAM mRNA expression who did not receive chemotherapy tended to have a worse prognosis. Similar but weaker correlations were found regarding ALCAM protein expression data. The predictive impact of ALCAM mRNA expression in chemotherapy treated patients was corroborated by multivariate Cox regression analysis also including histopathological markers (P = 0.001 for OAS). Our overall results reveal that high ALCAM expression levels in primary tumors might be a suitable marker for prediction of the response to adjuvant chemotherapy in breast cancer.

Nonmotor symptoms in Parkinson's disease (PD) involving cognition and emotionality have progressively received attention. The objective of the present study was to investigate recognition of emotional prosody in patients with PD (n = 14) in comparison to healthy control subjects (HC, n = 14). Event-related brain potentials (ERP) were recorded in a modified oddball paradigm under passive listening and active target detection instructions. Results showed a poorer performance of PD patients in classifying emotional prosody. ERP generated by emotional deviants (happy/sad) during passive listening revealed diminished amplitudes of the mismatch-related negativity for sad deviants, indicating an impairment of early preattentive processing of emotional prosody in PD.