Nitrate sensing and metabolism modulate motility, biofilm formation, and virulence in Pseudomonas aeruginosa

Infection by the bacterial opportunist Pseudomonas aeruginosa frequently assumes the form of a biofilm, requiring motility for biofilm formation and dispersal and an ability to grow in nutrient- and oxygen-limited environments. Anaerobic growth by P. aeruginosa is accomplished through the denitrification enzyme pathway that catalyzes the sequential reduction of nitrate to nitrogen gas. Mutants mutated in the two-component nitrate sensor-response regulator and in membrane nitrate reductase displayed altered motility and biofilm formation compared to wild-type P. aeruginosa PAO1. Analysis of additional nitrate dissimilation mutants demonstrated a second level of regulation in P. aeruginosa motility that is independent of nitrate sensor-response regulator function and is associated with nitric oxide production. Because motility and biofilm formation are important for P. aeruginosa pathogenicity, we examined the virulence of selected regulatory and structural gene mutants in the surrogate model host Caenorhabditis elegans. Interestingly, the membrane nitrate reductase mutant was avirulent in C. elegans, while nitrate sensor-response regulator mutants were fully virulent. The data demonstrate that nitrate sensing, response regulation, and metabolism are linked directly to factors important in P. aeruginosa pathogenesis.     

Grooved pegboard test as a biomarker of nigrostriatal denervation in Parkinson's disease

Recent pharmacotherapy trials in Parkinson's disease (PD) using dopaminergic neuroimaging as outcome parameter failed to show significant relationships between imaging and clinical results. One possible explanation is that there is a non-linear relationship between striatal denervation and motor performance reflecting a statistical "floor" effect in the imaging data with advanced disease. Both the motor manifestations and the striatal dopamine denervation of idiopathic PD, however, are typically asymmetric and more meaningful associations may be found by comparing data from the least denervated striatum with motor performance in the corresponding body side. PD patients (n=28) underwent [11C]beta-CFT dopamine transporter (DAT) positron emission tomography (PET) and grooved pegboard testing. Voxel-based analysis of DAT PET and bimanual pegboard scores demonstrated significant correlation clusters within the bilateral striata (P<0.001). However, findings were most prominent in the least denervated striatum. There was a significant inverse correlation between pegboard scores of the least affected arm and DAT binding of the least denervated striatum (Rs=-0.69, P<0.0001) but no significant correlation between pegboard scores of the clinically most affected arm and DAT binding of the most denervated striatum (Rs=-0.15, ns). These data indicate that the robustness of the grooved pegboard test as a biomarker for nigrostriatal denervation in PD mainly reflects the relationship between test performance of the clinically least affected limb and the least denervated striatum. These findings indicate that there is both a statistical "floor" and "ceiling" effect for the most affected striatal and body sides that must be considered when employing imaging as an outcome measure in clinical trials in PD.     

Unusual maternal vasculature in the placental periphery leading to the diagnosis of abdominal pregnancy at 25 weeks' gestation

Abdominal pregnancy is a rare condition in which the fetus and placenta are located within the peritoneal cavity. Sonographic findings include visualization of the fetus separate from the uterus, failure to visualize the uterine wall between the fetus and urinary bladder, close approximation of fetal parts to the maternal abdominal wall, eccentric position or abnormal fetal attitude, and visualization of extrauterine placental tissue. We present an unusual case in which mid-trimester transabdominal color Doppler sonographic findings depicted unusual maternal vasculature in the placental periphery leading to the diagnosis of abdominal pregnancy. Postpartum maternal angiography confirmed these vessels as abnormal maternal arterial perfusion of the extrauterine placenta emanating from the uterine arteries and inferior epigastric arteries. Systematic review of the literature confirms that this is the first report of such sonographic manifestations of an abdominal pregnancy.     

Predictors and consequences of pneumonia in critically ill patients with stroke

OBJECTIVE: Ascertain the incidence, predictors and consequences of early (within 48 hours of admission) and nosocomial pneumonia among critically ill patients with stroke. METHODS: Medical records of critically ill patients with acute stroke were reviewed. Predictors and consequences of pneumonia were determined with analysis of variance. RESULTS: 55 patients, aged 33 to 91 (median 74) years, were admitted. The stroke was located at the brainstem in 14 (26.4%) patients. Nine patients (16.4%) had early pneumonia and additional 17 patients (30.9%) developed nosocomial pneumonia. Patients with brain-stem stroke were more likely to develop early pneumonia (p =.04). Nosocomial pneumonia incidence was higher in patients who failed swallowing evaluation (RR = 6.3, 95% CI: 0.9-43.0) and in those who were intubated (58.6% v 0%, p =.00008). Also, nosocomial pneumonia was associated with longer duration of mechanical ventilation and prolonged hospital stay. Nineteen patients (34.5%) died at the hospital. They were more likely to be older (median 77 versus 69 years, p =.03) with higher admission acuity of illness. CONCLUSION: Pneumonia complicated stroke in 47% of critically ill patients and adversely impacted the duration of mechanical ventilation and overall length of hospital stay. Aggressive preventive measures are needed to reduce pneumonia occurrence in stroke patients.     

Macrophage subtype and cytokine expression characterization during the acute inflammatory phase of mouse bone fracture repair

Fracture repair is a complex process requiring heterotypic interactions between osteogenic cells and immune cells. Recent evidence indicates that macrophages are critically involved in fracture repair. Polarized macrophage populations differentially promote and regulate inflammation in other tissues, but little is known about the various macrophage subtypes and their signaling activities following a bone fracture. The authors hypothesized that classically activated (M1 subtype) and alternatively activated (M2 subtype) macrophages are active during the early repair process to initiate and regulate the inflammatory response. To test our hypothesis, bone marrow was collected from intact femurs (naïve group), contralateral and fractured femurs of mice on days 0, 1, 2, 4, and 7 postfracture. Macrophages were isolated from the bone marrow and macrophage subtypes were identified using flow cytometry with antibodies to F4/80, MHC II, CD86, CD11c, and CD40. Bone marrow cytokine levels were measured using xMAP. Flow cytometry revealed dynamic changes in M1 subtype (F4/80+/MHC II+/CD86+), M2 subtype (F4/80+/MHC II−/CD86−), and dendritic cell (DCs; MHCII+/CD11c+/CD40+) populations following fracture as compared to naïve controls. M1 subtype levels were correlated with IL-1α, IL-1ß, IL-2, IL-17, Eotaxin, and MCP-1, while DCs were correlated with IL-6, G-CSF, LIF, KC, and VEGF-A. The results indicate that M1 and M2 subtypes and DCs are recruited to the fracture site early during the repair process and consequently may work in tandem to regulate the inflammatory response required to recruit osteogenic cells needed for later stages of repair.     

Patupilone (epothilone B) inhibits growth and survival of multiple myeloma cells in vitro and in vivo

In this study, we investigated the in vitro and in vivo efficacy of patupilone (epothilone B, EPO906), a novel nontaxane microtubule stabilizing agent, in treatment of multiple myeloma (MM). Patupilone directly inhibited growth and survival of MM cells, including those resistant to conventional chemotherapies, such as the taxane paclitaxel. Patupilone induced G2M arrest of MM cells, with subsequent apoptosis. Interleukin-6 (IL-6) and insulin-like growth factor-1 (IGF-1), 2 known growth and survival factors for MM, did not protect MM.1S cells against patupilone-induced cell death. Proliferation of MM cells induced by adherence to bone marrow stromal cells (BMSCs) was also inhibited by patupilone and was paralleled by down-regulation of vascular endothelial growth factor (VEGF) secretion. Importantly, stimulation of cells from patients with MM, either with IL-6 or by adherence to BMSCs, enhanced the anti-proliferative and proapoptotic effects of patupilone. Moreover, patupilone was effective against MM cell lines that overexpress the MDR1/P-glycoprotein multidrug efflux pump. In addition, patupilone was effective in slowing tumor growth and prolonging median survival of mice that received orthotopical transplants with MM tumor cells. Taken together, these preclinical findings suggest that patupilone may be a safe and effective drug in the treatment of MM, providing the framework for clinical studies to improve patient outcome in MM.     

5′-Guanylylimidodiphosphate, A Potent Activator of Adenylate Cyclase Systems in Eukaryotic Cells

5'-Guanylylimidodiphosphate (Gpp(NH)-p) stimulates adenylate cyclase [ATP-pyrophosphate-lyase (cyclizing), EC 4.6.1.1] activity in plasma membranes isolated from frog and salmon erythrocytes, from rat adrenal, hepatic, and fat cells, and from bovine thyroid cells. The nucleotide acts cooperatively with the various hormones (glucagon, secretin, ACTH, thyrotropin, and catecholamines) that stimulate these adenylate cyclase systems with resultant activities that equal or exceed those obtained with hormone plus GTP or with fluoride ion. In the absence of hormones, Gpp(NH)p is a considerably more effective activator than GTP, and, under certain conditions of incubation, stimulates rat fat cell adenylate cyclase to levels of activity (about 20 nmoles of 3',5'-adenosine monophosphate mg protein per min) far higher than reported hitherto for any adenylate cyclase system examined. The nucleotide activates frog erythrocyte adenylate cyclase when the catecholamine receptor is blocked by the competitive antagonist, propranolol, and activates the enzyme from an adrenal tumor cell line which lacks functional ACTH receptors. In contrast, Gpp(NH)p does not stimulate adenylate cyclase in extracts from Escherichia coli B. Gpp(NH)p appears to be a useful probe for investigating the mechanism of hormone and nucleotide action on adenylate cyclase systems in eukaryotic cells.