The development of the bladder trigone, the center of the anti-reflux mechanism

The urinary tract is an outflow system that conducts urine from the kidneys to the bladder via the ureters that propel urine to the bladder via peristalsis. Once in the bladder, the ureteral valve, a mechanism that is not well understood, prevents backflow of urine to the kidney that can cause severe damage and induce end-stage renal disease. The upper and lower urinary tract compartments form independently, connecting at mid-gestation when the ureters move from their primary insertion site in the Wolffian ducts to the trigone, a muscular structure comprising the bladder floor just above the urethra. Precise connections between the ureters and the trigone are crucial for proper function of the ureteral valve mechanism; however, the developmental events underlying these connections and trigone formation are not well understood. According to established models, the trigone develops independently of the bladder, from the ureters, Wolffian ducts or a combination of both; however, these models have not been tested experimentally. Using the Cre-lox recombination system in lineage studies in mice, we find, unexpectedly, that the trigone is formed mostly from bladder smooth muscle with a more minor contribution from the ureter, and that trigone formation depends at least in part on intercalation of ureteral and bladder muscle. These studies suggest that urinary tract development occurs differently than previously thought, providing new insights into the mechanisms underlying normal and abnormal development.     

A limited screen for protein interactions reveals new roles for protein phosphatase 1 in cell cycle control and apoptosis

Protein phosphatase 1 (PP1) catalytic subunits typically combine with other proteins that modulate their activity, direct them to distinct substrates, or serve as substrates for PP1. More than 50 PP1-interacting proteins (PIPs) have been identified so far. Given there are approximately 10 000 phosphoproteins in mammals, many PIPs remain to be discovered. We have used arrays containing 100 carefully selected antibodies to identify novel PIPs that are important in cell proliferation and cell survival in murine fetal lung epithelial cells and human A549 lung cancer cells. The antibody arrays identified 31 potential novel PIPs and 11 of 17 well-known PIPs included as controls, suggesting a sensitivity of at least 65%. A majority of the interactions between PP1 and putative PIPs were isoform- or cell type-specific. We confirmed by co-immunoprecipitation that 9 of these proteins associate with PP1: APAF-1, Bax, E-cadherin, HSP-70, Id2, p19Skp1, p53, PCNA, and PTEN. We examined two of these interactions in greater detail in A549 cells. Exposure to nicotine enhanced association of PP1 with Bax (and Bad), but also induced inhibitory phosphorylation of PP1. In addition to p19Skp1, PP1alpha antibodies also coprecipitated cullin 1, suggesting that PP1alpha is associated with the SCF1 complex. This interaction was only detectable during the G1/S transition and S phase. Forced loss of PP1 function decreased the levels of p27Kip1, a well-known SCF1 substrate, suggesting that PP1 may rescue proteins from ubiquitin/proteasome-mediated destruction. Both of these novel interactions are consistent with PP1 facilitating cell cycle arrest and/or apoptosis.     

Annexin A2–dependent actin bundling promotes secretory granule docking to the plasma membrane and exocytosis

Annexin A2, a calcium-, actin-, and lipid-binding protein involved in exocytosis, mediates the formation of lipid microdomains required for the structural and spatial organization of fusion sites at the plasma membrane. To understand how annexin A2 promotes this membrane remodeling, the involvement of cortical actin filaments in lipid domain organization was investigated. 3D electron tomography showed that cortical actin bundled by annexin A2 connected docked secretory granules to the plasma membrane and contributed to the formation of GM1-enriched lipid microdomains at the exocytotic sites in chromaffin cells. When an annexin A2 mutant with impaired actin filament–bundling activity was expressed, the formation of plasma membrane lipid microdomains and the number of exocytotic events were decreased and the fusion kinetics were slower, whereas the pharmacological activation of the intrinsic actin-bundling activity of endogenous annexin A2 had the opposite effects. Thus, annexin A2–induced actin bundling is apparently essential for generating active exocytotic sites.     

Nephric duct insertion is a crucial step in urinary tract maturation that is regulated by a Gata3-Raldh2-Ret molecular network in mice

Urinary tract development depends on a complex series of events in which the ureter moves from its initial branch point on the nephric duct (ND) to its final insertion site in the cloaca (the primitive bladder and urethra). Defects in this maturation process can result in malpositioned ureters and hydronephrosis, a common cause of renal disease in children. Here, we report that insertion of the ND into the cloaca is an unrecognized but crucial step that is required for proper positioning of the ureter and that depends on Ret signaling. Analysis of Ret mutant mice at birth reveals hydronephrosis and defective ureter maturation, abnormalities that our results suggest are caused, at least in part, by delayed insertion of the ND. We find a similar set of malformations in mutants lacking either Gata3 or Raldh2. We show that these factors act in parallel to regulate ND insertion via Ret. Morphological analysis of ND extension in wild-type embryos reveals elaborate cellular protrusions at ND tips that are not detected in Ret, Gata3 or Raldh2 mutant embryos, suggesting that these protrusions may normally be important for fusion with the cloaca. Together, our studies reveal a novel Ret-dependent event, ND insertion, that, when abnormal, can cause obstruction and hydronephrosis at birth; whether ND defects underlie similar types of urinary tract abnormalities in humans is an interesting possibility.     

Copper-mediated nuclease activity of jadomycin B

The natural product jadomycin B, isolated from Streptomyces venezeulae ISP5230, has been found to cleave DNA in the presence of Cu(II) ions without the requirement for an external reducing agent. The efficiency of DNA cleavage was probed using supercoiled plasmid DNA in buffered solution as a model environment. EC?? and t(?) values for cleavage were 1.7 μM and 0.75 h, respectively, and varied ± 5% with the particular batch of plasmid and jadomycin employed. While UV-vis spectroscopy indicates that the cleavage event does not involve direct binding of jadomycin B to DNA, a stoichiometric Cu(II) preference for optimum cleavage suggests a weak binding interaction between jadomycin B and Cu(II) in the presence of DNA. The Cu(II)-mediated cleavage is greatly enhanced by UV light, which implicates the jadomycin B radical cation and Cu(I) as potential intermediates in DNA cleavage. Evidence in favor of this hypothesis was derived from a mechanistic assay which showed reduced cleavage as a function of added catalase and EDTA, scavengers of H?O? and Cu(II), respectively. Thus, jadomycin B may serve as a source of electrons for Cu(II) reduction, producing Cu(I) which reacts with H?O? to form hydroxyl radicals that cause DNA strand scission. In addition, scavengers of hydroxyl radicals and superoxide also display inhibitory effects, underscoring the ability of jadomycin B to produce a powerful arsenal of deleterious oxygen species when copper is present.     

Review of Florida Red Tide and Human Health Effects

This paper reviews the literature describing research performed over the past decade on the known and possible exposures and human health effects associated with Florida red tides. These harmful algal blooms are caused by the dinoflagellate, Karenia brevis, and similar organisms, all of which produce a suite of natural toxins known as brevetoxins. Florida red tide research has benefited from a consistently funded, long term research program, that has allowed an interdisciplinary team of researchers to focus their attention on this specific environmental issue-one that is critically important to Gulf of Mexico and other coastal communities. This long-term interdisciplinary approach has allowed the team to engage the local community, identify measures to protect public health, take emerging technologies into the field, forge advances in natural products chemistry, and develop a valuable pharmaceutical product. The Review includes a brief discussion of the Florida red tide organisms and their toxins, and then focuses on the effects of these toxins on animals and humans, including how these effects predict what we might expect to see in exposed people.     

Evaluation of 4-poster acaricide applicators to manage tick populations associated with disease risk in a Tennessee retirement community

In 1993, four residents of a retirement community in middle Tennessee were hospitalized with symptoms of ehrlichiosis causing community managers to implement mitigation methods to reduce tick numbers. For the past four years, managers have utilized 4-poster acaricide applicators that aim to reduce disease risk to residents by killing ticks that feed on deer. To determine the efficacy of this technique, we assessed Amblyomma americanum abundance in the vicinity of the devices by dragging 400 m vegetation transects once per month while ticks were active. In 2009, adult tick activity peaked in May, nymphal tick activity peaked in June, and larval activity peaked in September. Close to 4-poster devices, larval, nymphal, and adult tick abundances were reduced by 91%, 68%, and 49%, respectively (larval and nymphal p<0.001, adult p=0.005), relative to nearby untreated areas. No significant reduction in nymphal or adult A. americanum ticks was evident >300 m from 4-poster devices, however a ～90% reduction in larvae was observed to our sampling limit (400 m). At the low density at which these devices are currently being used (average distance between devices = 6.6 km), we conclude that they will have little large-scale effect on the health risk posed by ticks in this community.     

EGFR and EphA2 are host factors for hepatitis C virus entry and possible targets for antiviral therapy

Hepatitis C virus (HCV) is a major cause of liver disease, but therapeutic options are limited and there are no prevention strategies. Viral entry is the first step of infection and requires the cooperative interaction of several host cell factors. Using a functional RNAi kinase screen, we identified epidermal growth factor receptor and ephrin receptor A2 as host cofactors for HCV entry. Blocking receptor kinase activity by approved inhibitors broadly impaired infection by all major HCV genotypes and viral escape variants in cell culture and in a human liver chimeric mouse model in vivo. The identified receptor tyrosine kinases (RTKs) mediate HCV entry by regulating CD81-claudin-1 co-receptor associations and viral glycoprotein-dependent membrane fusion. These results identify RTKs as previously unknown HCV entry cofactors and show that tyrosine kinase inhibitors have substantial antiviral activity. Inhibition of RTK function may constitute a new approach for prevention and treatment of HCV infection.     

Visual perceptual difficulties and under-achievement at school in a large community-based sample of children

Introduction

Difficulties with visual perception (VP) are often described in children with neurological or developmental problems. However, there are few data regarding the range of visual perceptual abilities in populations of normal children, or on the impact of these abilities on children''s day-to-day functioning.

Methods

Data were obtained for 4512 participants in an ongoing birth cohort study (Avon Longitudinal Study of Parents and Children; ALSPAC). The children''s mothers responded to questions designed to elicit indications of visual perceptual difficulties or immaturity, when their children were aged 13 years. We examined associations with standardised school test results in reading and in mathematics at age 13–14 years (SATS-KS3), accounting for potential confounders including IQ.

Results

Three underlying factors explained half the variance in the VP question responses. These correlated best with questions on interpreting cluttered scenes; guidance of movement and face recognition. The adjusted parameter estimates (95% CI) for the cluttered-scenes factor (0.05; 0.02 to 0.08; p<0.001) suggested positive associations with the reading test results whilst that for the guidance-of-movement factor (0.03; 0.00 to 0.06; p = 0.026) suggested positive association with the mathematics results. The raw scores were associated with both test results.

Discussion

VP abilities were widely distributed in this sample of 13-year old children. Lower levels of VP function were associated with under-achievement in reading and in mathematics. Simple interventions can help children with VP difficulties, so research is needed into practicable, cost-effective strategies for identification and assessment, so that support can be targeted appropriately.