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Background: Current imaging criteria for categorising disease response in metastatic renal cell carcinoma (mRCC) correlate poorly with overall survival (OS) in patients on anti-angiogenic therapies. We prospectively assess diffusion-weighted and multiphase contrast-enhanced (MCE) MR imaging (MRI) as markers of outcome., Methods: Treatment-naive mRCC patients on a phase II trial using sunitinib completed an MRI substudy. Whole-tumour apparent diffusion coefficient (ADC) maps and histograms were generated, and mean ADC and AUC(low) (proportion of the tumour with ADC values lying below the 25th percentile of the ADC histogram) recorded. On MCE-MRI, regions of interest were drawn around the most avidly enhancing components to analyse enhancement parameters. Baseline (n=26) and treatment-related changes in surviving patients (n=20) were correlated with OS. Imaged metastases were also analysed., Results: Forty-seven per cent of the patients showed significant changes in whole-tumour mean ADC following therapy, but there was no correlation with outcome. Patients with a high baseline AUC(low) and greater-than-median AUC(low) increase had reduced OS (HR=3.67 (95% confidence interval (CI)=1.23-10.9), P=0.012 and HR=3.72 (95% CI=0.98-14.21), P=0.038, respectively). There was no correlation between MCE-MRI parameters and OS. Twenty-eight metastases were analysed and showed positive correlation with primary tumour mean ADC for individual patients (r=0.607; P<0.001)., Conclusion: Primary RCC ADC histogram analysis shows dynamic changes with sunitinib. Patients in whom the tumour ADC histogram demonstrated high baseline AUC(low) or a greater-than-median increase in AUC(low) with treatment had reduced OS.

Background: Autosomal dominant polycystic kidney disease (ADPKD) is clinically heterogenic. Biomarkers are needed to predict prognosis and guide management. We aimed to profile microRNA (miRNA) in ADPKD to gain molecular insight and evaluate biomarker potential., Methods: Small-RNA libraries were generated from urine specimens of ADPKD patients (N = 20) and patients with chronic kidney disease of other etiologies (CKD, N = 20). In this report, we describe the miRNA profiles and baseline characteristics. For reference, we also examined the miRNA transcriptome in primary cultures of ADPKD cyst epithelia (N = 10), normal adult tubule (N = 8) and fetal tubule (N = 7) epithelia., Results: In primary cultures of ADPKD kidney cells, miRNA cistrons mir-143(2) (9.2-fold), let-7i(1) (2.3-fold) and mir-3619(1) (12.1-fold) were significantly elevated compared to normal tubule epithelia, whereas mir-1(4) members (19.7-fold), mir-133b(2) (21.1-fold) and mir-205(1) (3.0-fold) were downregulated (P<0.01). Expression of the dysregulated miRNA in fetal tubule epithelia resembled ADPKD better than normal adult cells, except let-7i, which was lower in fetal cells. In patient biofluid specimens, mir-143(2) members were 2.9-fold higher in urine cells from ADPKD compared to other CKD patients, while expression levels of mir-133b(2) (4.9-fold) and mir-1(4) (4.4-fold) were lower in ADPKD. We also noted increased abundance mir-223(1) (5.6-fold), mir-199a(3) (1.4-fold) and mir-199b(1) (1.8-fold) (P<0.01) in ADPKD urine cells. In ADPKD urine microvesicles, miR-1(2) (7.2-fold) and miR-133a(2) (11.8-fold) were less abundant compared to other CKD patients (P<0.01)., Conclusions: We found that in ADPKD urine specimens, miRNA previously implicated as kidney tumor suppressors (miR-1 and miR-133), as well as miRNA of presumed inflammatory and fibroblast cell origin (miR-223/miR-199), are dysregulated when compared to other CKD patients. Concordant with findings in the primary tubule epithelial cell model, this suggests roles for dysregulated miRNA in ADPKD pathogenesis and potential use as biomarkers. We intend to assess prognostic potential of miRNA in a followup analysis.

Objective: To determine the long-term (12-year) effects of a conservative nurse-led intervention for postnatal urinary incontinence., Design: Follow-up of a randomised controlled trial., Setting: Community-based intervention in three centres (in the UK and New Zealand)., Population: A cohort of 747 women with urinary incontinence at 3 months after childbirth, of whom 471 (63%) were followed up after 12 years., Methods: Women were randomly allocated to active conservative treatment after delivery (pelvic floor muscle training and bladder training), or to a control group receiving standard care., Main Outcome Measures: Prevalence of urinary incontinence (primary outcome) and faecal incontinence, symptoms and signs of prolapse, and performance of pelvic floor muscle training at 12 years., Results: The significant improvements relative to controls that had been found in urinary incontinence (60 versus 69%; risk difference, RD, -9.1%; 95% confidence interval, 95% CI, -17.3 to -1.0%) and faecal incontinence (4 versus 11%; RD -6.1%; 95% CI -10.8 to -1.6%) at 1 year did not persist for urinary incontinence (83 versus 80%; RD 2.1%; 95% CI -4.9 to 9.1%) or faecal incontinence (19 versus 15%; RD 4.3%; 95% CI -2.5 to 11.0%) at the 12-year follow up, irrespective of incontinence severity at trial entry. The prevalence of prolapse symptoms or objectively measured pelvic organ prolapse also did not differ between the groups. In the short term the intervention motivated more women to perform pelvic floor muscle training (83 versus 55%), but this fell in both groups by 12 years (52 versus 49%)., Conclusions: The moderate short-term benefits of a brief nurse-led conservative treatment for postnatal urinary incontinence did not persist. About four-fifths of women with urinary incontinence 3 months after childbirth still had this problem 12 years later., (© 2013 Royal College of Obstetricians and Gynaecologists.)

Background: Historically, the Renal Unit at King's College Hospital used intravenous (IV) iron sucrose (IS) to treat iron deficiency anaemia in patients with chronic kidney disease who were not on dialysis (CKD-ND). As part of a service initiative to improve patient experience, new products were considered as alternatives. This study investigated the potential impact on patient experience and service costs by switching from IS to ferric carboxymaltose (FCM)., Methods: A decision analytical model was used to calculate the impact of switching from IS to FCM for a cohort of CKD-ND patients. Service provision data were collected for 365 patients who received 600 mg IS within a 12 month period, creating the IS data set. The service provision data, along with a clinically relevant FCM administration protocol (stipulating total doses of 500 mg FCM), were used to calculate a corresponding theoretical data set for FCM for the same cohort of patients., Results: The FCM protocol saved each patient two hospital visits and 2.66 hours of time (equating to approximately a saving of £36.21 in loss of earnings) and £19 in travel costs. Direct attributable costs for iron administration (which included drug, disposables, nursing staff, and hospital-provided patient transport costs) were £58,646 for IS vs £46,473 for FCM. Direct overhead costs (which included nursing preparation time, administration staff, clinic space, and consultant time costs) were £40,172 for the IS service vs £15,174 for the FCM service., Limitations: Based on clinical experience with the products, this analysis assumes that 500 mg FCM is therapeutically equivalent to 600 mg IS. Consultant time costs are assumed to be the same between the two treatment groups. IV iron administration protocols and data are specific to King's College Hospital. The design is retrospective and changes to the management of the clinic, including service delivery optimization, may also affect real costs., Conclusion: FCM was associated with fewer hospital visits and reduced transport costs for CKD-ND patients receiving IV iron and has the potential to save 19-37% in service costs. Owing to increased administration efficiency, FCM can improve the overall patient experience while reducing the total cost of the King's College Hospital IV iron service for CKD-ND patients, compared with treatment with IS.

Background: Vitamin A is necessary for kidney development and has also been linked to regulation of solute and water homeostasis and to protection against kidney stone disease, infection, inflammation, and scarring. Most functions of vitamin A are mediated by its main active form, all-trans retinoic acid (tRA), which binds retinoic acid receptors (RARs) to modulate gene expression. We and others have recently reported that renal tRA/RAR activity is confined to the ureteric bud (UB) and collecting duct (CD) cell lineage, suggesting that endogenous tRA/RARs primarily act through regulating gene expression in these cells in embryonic and adult kidney, respectively., Methodology/principal Findings: To explore target genes of endogenous tRA/RARs, we employed the mIMCD-3 mouse inner medullary CD cell line, which is a model of CD principal cells and exhibits constitutive tRA/RAR activity as CD principal cells do in vivo. Combining antagonism of RARs, inhibition of tRA synthesis, exposure to exogenous tRA, and gene expression profiling techniques, we have identified 125 genes as candidate targets and validated 20 genes that were highly regulated (Dhrs3, Sprr1a, and Ppbp were the top three). Endogenous tRA/RARs were more important in maintaining, rather than suppressing, constitutive gene expression. Although many identified genes were expressed in UBs and/or CDs, their exact functions in this cell lineage are still poorly defined. Nevertheless, gene ontology analysis suggests that these genes are involved in kidney development, renal functioning, and regulation of tRA signaling., Conclusions/significance: A rigorous approach to defining target genes for endogenous tRA/RARs has been established. At the pan-genomic level, genes regulated by endogenous tRA/RARs in a CD cell line have been catalogued for the first time. Such a catalogue will guide further studies on molecular mediators of endogenous tRA/RARs during kidney development and in relation to renal defects associated with vitamin A deficiency.

Microbial pathogens are ancient selective agents that have driven many aspects of multicellular evolution, including genetic, behavioural, chemical and immune defence systems. It appears that fungi specialised to attack insects were already present in the environments in which social insects first evolved and we hypothesise that if the early stages of social evolution required antifungal defences, then covariance between levels of sociality and antifungal defences might be evident in extant lineages, the defences becoming stronger with group size and increasing social organisation. Thus, we compared the activity of cuticular antifungal compounds in thrips species (Insecta: Thysanoptera) representing a gradient of increasing group size and sociality: solitary, communal, social and eusocial, against the entomopathogen Cordyceps bassiana. Solitary and communal species showed little or no activity. In contrast, the social and eusocial species killed this fungus, suggesting that the evolution of sociality has been accompanied by sharp increases in the effectiveness of antifungal compounds. The antiquity of fungal entomopathogens, demonstrated by fossil finds, coupled with the unequivocal response of thrips colonies to them shown here, suggests two new insights into the evolution of thrips sociality: First, traits that enabled nascent colonies to defend themselves against microbial pathogens should be added to those considered essential for social evolution. Second, limits to the strength of antimicrobials, through resource constraints or self-antibiosis, may have been overcome by increase in the numbers of individuals secreting them, thus driving increases in colony size. If this is the case for social thrips, then we may ask: did antimicrobial traits and microbes such as fungal entomopathogens play an integral part in the evolution of insect sociality in general?

Members of the epidermal growth factor (EGF) family bind to ErbB (EGFR) family receptors which play an important role in the regulation of various fundamental cell processes including cell proliferation and differentiation. The normal rodent kidney has been shown to express at least three members of the ErbB receptor family and is a major site of EGF ligand synthesis. Polycystic kidney disease (PKD) is a group of diseases caused by mutations in single genes and is characterized by enlarged kidneys due to the formation of multiple cysts in both kidneys. Tubule cells proliferate, causing segmental dilation, in association with the abnormal deposition of several proteins. One of the first abnormalities described in cell biological studies of PKD pathogenesis was the abnormal mislocalization of the EGFR in cyst lining epithelial cells. The kidney collecting duct (CD) is predominantly an absorptive epithelium where electrogenic Na(+) entry is mediated by the epithelial Na(+) channel (ENaC). ENaC-mediated sodium absorption represents an important ion transport pathway in the CD that might be involved in the development of PKD. A role for EGF in the regulation of ENaC-mediated sodium absorption has been proposed. However, several investigations have reported contradictory results indicating opposite effects of EGF and its related factors on ENaC activity and sodium transport. Recent advances in understanding how proteins in the EGF family regulate the proliferation and sodium transport in normal and PKD epithelial cells are discussed here. This article is part of a Special Issue entitled: Polycystic Kidney Disease., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Epithelial cell polarity is essential for the establishment and maintenance of morphological and functional asymmetries that underlie normal renal structure and function and are brought about by the appropriate delivery of growth factor receptors and ion and fluid transporters and channels to apical or basolateral cell membranes. The fundamental process of cellular polarization is established early during development and is controlled by sets of evolutionarily conserved proteins that integrate intrinsic and extrinsic polarity cues. Specialized structural domains between adjacent cells and cells with their matrix, termed adherens junctions (AJ) and focal adhesions (FA), respectively, are formed that contain specific components of multi-molecular complexes acting as sites to recruit proteins and to activate intracellular mechano-transduction pathways. Regulation of these processes results in tight spatio-temporal control of renal tubule growth and lumen diameter. Abnormalities in macromolecular polarization complexes lead to a variety of diseases in different organs, a common example of which is Polycystic Kidney Disease (PKD), where epithelial cysts replace normal renal tubules. Membrane protein polarity defects in Autosomal Dominant (AD) PKD include the mis-polarization of normally basolateral membrane proteins to apical, lumenal membranes, such as epidermal growth factor (EGFR/ErbB) receptors and Na(+)K(+)-ATPase-α1 subunit; mis-polarization of normally apical membrane proteins to basolateral membranes, including the Na(+)K(+)2Cl(-) (NKCC1) symporter; and the failure to traffic and insert proteins into membranes resulting in their intracellular accumulation, such as E-cadherin and the β1 subunit of Na(+)K(+)-ATPase. Abnormalities in structural AJ, FA and polarity complexes in ADPKD epithelia include loss of E-cadherin, and focal adhesion kinase (FAK), MALS-3, Crb and Dlg complexes as well as disruptions in Rab/sec and syntaxin trafficking and membrane docking pathways. Since proper polarization of epithelial cells lining renal tubules is essential for normal kidney development and differentiation to prevent abnormal cystic dilation, interventions to reverse polarity defects to normal would offer therapeutic opportunities for PKD. This article is part of a Special Issue entitled: Polycystic Kidney Disease., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Autosomal dominant polycystic kidney disease (ADPKD) is caused by mutation of PKD1 and PKD2 that encode polycystin-1 and polycystin-2. Polycystin-1 is tyrosine phosphorylated and modulates multiple signaling pathways including AP-1, and the identity of the phosphatases regulating polycystin-1 are previously uncharacterized. Here we identify members of the LAR protein tyrosine phosphatase (RPTP) superfamily as members of the polycystin-1complex mediated through extra- and intracellular interactions. The first extracellular PKD1 domain of polycystin-1 interacts with the first Ig domain of RPTPσ, while the polycystin-1 C-terminus of polycystin-1 interacts with the regulatory D2 phosphatase domain of RPTPγ. Additional homo- and heterotypic interactions between RPTPs recruit RPTPδ. The multimeric polycystin protein complex is found localised in cilia. RPTPσ and RPTPδ are also part of a polycystin-1/E-cadherin complex known to be important for early events in adherens junction stabilisation. The interaction between polycystin-1 and RPTPγ is disrupted in ADPKD cells, while RPTPσ and RPTPδ remain closely associated with E-cadherin, largely in an intracellular location. The polycystin-1 C-terminus is an in vitro substrate of RPTPγ, which dephosphorylates the c-Src phosphorylated Y4237 residue and activates AP1-mediated transcription. The data identify RPTPs as novel interacting partners of the polycystins both in cilia and at adhesion complexes and demonstrate RPTPγ phosphatase activity is central to the molecular mechanisms governing polycystin-dependent signaling. This article is part of a Special Issue entitled: Polycystic Kidney Disease., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Angiogenesis is a fundamental step in several important physiological events and pathological conditions including embryonic development, wound repair, tumor growth and metastasis. PRKX was identified as a novel type-I cAMP-dependent protein kinase gene expressed in multiple developing tissues. PRKX has also been shown to be phylogenetically and functionally distinct from PKA. This study presents the first evidence that PRKX stimulates endothelial cell proliferation, migration, and vascular-like structure formation, which are the three essential processes for angiogenesis. In contrast, classic PKA demonstrated an inhibitory effect on endothelia vascular-like structure formation. Our findings suggest that PRKX is an important protein kinase engaged in the regulation of angiogenesis and could play critical roles in various physiological and pathological conditions involving angiogenesis. PRKX binds to Pin-1, Magi-1 and Bag-3, which regulate cell proliferation, apoptosis, differentiation and tumorigenesis. The interaction of PRKX with Pin-1, Magi-1 and Bag-3 could contribute to the stimulating role of PRKX in angiogenesis., (Copyright © 2011. Published by Elsevier Inc.)

c-Src is a non-receptor tyrosine kinase whose activity is induced by phosphorylation at Y418 and translocation from the cytoplasm to the cell membrane. Increased activity of c-Src has been associated with cell proliferation, matrix adhesion, motility, and apoptosis in tumors. Immunohistochemistry suggested that activated (pY(418))-Src activity is increased in cyst-lining autosomal dominant polycystic kidney disease (ADPKD) epithelial cells in human and mouse ADPKD. Western blot analysis showed that SKI-606 (Wyeth) is a specific inhibitor of pY(418)-Src without demonstrable effects on epidermal growth factor receptor or ErbB2 activity in renal epithelia. In vitro studies on mouse inner medullary collecting duct (mIMCD) cells and human ADPKD cyst-lining epithelial cells showed that SKI-606 inhibited epithelial cell proliferation over a 24-h time frame. In addition, SKI-606 treatment caused a striking statistically significant decrease in adhesion of mIMCD and human ADPKD to extracellular collagen matrix. Retained viability of unattached cells was consistent with a primary effect on epithelial cell anchorage dependence mediated by the loss of extracellular matrix (ECM)-attachment due to α(2)β(1)-integrin function. SKI-606-mediated attenuation of the human ADPKD hyperproliferative and hyper-ECM-adhesive epithelial cell phenotype in vitro was paralleled by retardation of the renal cystic phenotype of Pkd1 orthologous ADPKD heterozygous mice in vivo. This suggests that SKI-606 has dual effects on cystic epithelial cell proliferation and ECM adhesion and may have therapeutic potential for ADPKD patients.

Microbial diseases are important selective agents in social insects and one major defense mechanism is the secretion of cuticular antimicrobial compounds. We hypothesized that given differences in group size, social complexity, and nest type the secretions of these antimicrobials will be under different selective pressures. To test this we extracted secretions from nine wasp species of varying social complexity and nesting habits and assayed their antimicrobial compounds against cultures of Staphylococcus aureus. These data were then combined with phylogenetic data to provide an evolutionary context. Social species showed significantly higher (18x) antimicrobial activity than solitary species and species with paper nests showed significantly higher (11x) antimicrobial activity than those which excavated burrows. Mud-nest species showed no antimicrobial activity. Solitary, burrow-provisioning wasps diverged at more basal nodes of the phylogenetic trees, while social wasps diverged from the most recent nodes. These data suggest that antimicrobial defences may have evolved in response to ground-dwelling pathogens but the most important variable leading to increased antimicrobial strength was increase in group size and social complexity.

Autosomal recessive polycystic kidney disease (ARPKD) is a significant hereditary renal disease occurring in infancy and childhood, which presents with greatly enlarged echogenic kidneys, ultimately leading to renal insufficiency and end-stage renal disease. ARPKD is caused by mutations in a single gene PKHD1, which encodes fibrocystin/polyductin (FPC), a large single transmembrane protein generally known to be on the primary cilium, basal body and plasma membrane. Here, using our newly generated antibody raised against the entire C-terminal intracellular cytoplasmic domain (ICD) of FPC, as well as our previously well-characterized antibody against a peptide of ICD, we report for the first time that at least one isoform of FPC is localized to the centrosome and mitotic spindle of dividing cells in multiple cell lines, including MDCK, mIMCD3, LLC-PK1, HEK293, RCTEC and HFCT cells. Using short-hairpin-mediated RNA interference, we show that the inhibition of FPC function in MDCK and mIMCD3 cells leads to centrosome amplification, chromosome lagging and multipolar spindle formation. Consistent with our in vitro findings, we also observed centrosome amplification in the kidneys from human ARPKD patients. These findings demonstrate a novel function of FPC in centrosome duplication and mitotic spindle assembly during cell division. We propose that mitotic defects due to FPC dysfunction contribute to cystogenesis in ARPKD.

The activation of heterotrimeric G protein signaling is a key feature in the pathophysiology of polycystic kidney diseases (PKD). In this study, we report abnormal overexpression of activator of G protein signaling 3 (AGS3), a receptor-independent regulator of heterotrimeric G proteins, in rodents and humans with both autosomal recessive and autosomal dominant PKD. Increased AGS3 expression correlated with kidney size, which is an index of severity of cystic kidney disease. AGS3 expression localized exclusively to distal tubular segments in both normal and cystic kidneys. Short hairpin RNA-induced knockdown of endogenous AGS3 protein significantly reduced proliferation of cystic renal epithelial cells by 26 +/- 2% (P < 0.001) compared with vehicle-treated and control short hairpin RNA-expressing epithelial cells. In summary, this study suggests a relationship between aberrantly increased AGS3 expression in renal tubular epithelia affected by PKD and epithelial cell proliferation. AGS3 may play a receptor-independent role to regulate Galpha subunit function and control epithelial cell function in PKD.

Amiloride-sensitive sodium entry, via the epithelial sodium channel (ENaC), is the rate-limiting step for Na(+) absorption. Epidermal growth factor (EGF) is involved in the regulation of Na(+) transport and ENaC activity. However it is still controversial exactly how EGF regulates ENaC and Na(+) absorption. The aim of the present study was to characterize the EGF regulation of Na(+) transport in cultured mouse renal collecting duct principal mpkCCD(c14) cells, a highly differentiated cell line which retains many characteristics of the cortical collecting duct (CCD). EGF dose dependently regulates basal transepithelial Na(+) transport in two phases: an acute phase (<4 h) and a chronic phase (>8 h). Similar effects were observed with TGF-alpha, HB-EGF, and amphiregulin which also belong to the EGF-related peptide growth factor family. Inhibition of MEK1/2 by PD98059 or U0126 increased acute effects and disrupted chronic effects of EGF on Na(+) reabsorption. Inhibition of PI3-kinase with LY294002 abolished acute effect of EGF. As assessed by Western blotting, ErbB2 is the most predominant member of the ErbB family detected in mpkCCD(c14) cells. Immunohistochemistry analysis revealed localization of ErbB2 in the CCD in Sprague-Dawley rat kidneys. Both acute and long-term effects of EGF were abolished when cells were treated with tyrphostin AG-825 and ErbB2 inhibitor II, chemically dissimilar selective inhibitors of the ErbB2 receptor. Thus, we conclude that EGF and its related growth factors are important for maintaining transepithelial Na(+) transport and that EGF biphasically modulates sodium transport in mpkCCD(c14) cells via the ErbB2 receptor., (J. Cell. Physiol. 223: 252-259, 2010. (c) 2009 Wiley-Liss, Inc.)

Integrin-associated focal adhesion complex formation and turnover plays an essential role in directing interactions between epithelial cells and the extracellular matrix during organogenesis, leading to appropriate cell spreading, cell-matrix adhesion, and migration. Autosomal recessive polycystic kidney disease (ARPKD) is associated with loss of function of PKHD1-encoded protein fibrocystin-1 and is characterized by cystic dilation of renal collecting tubules (CT) in utero and loss of renal function in patients if they survive the perinatal period. Normal polycystin-1 (PC-1)/focal adhesion complex function is required for control of CT diameter during renal development, and abnormalities in these complexes have been demonstrated in human PC-1 mutant cystic cells. To determine whether loss of fibrocystin-1 was associated with focal adhesion abnormalities, ARPKD cells or normal age-matched human fetal (HF)CT cells in which fibrocystin-1 had been decreased by 85% by small interfering RNA inhibition were compared with normal HFCT. Accelerated attachment and spreading on collagen matrix and decreased motility of fibrocystin-1-deficient cells were associated with longer paxillin-containing focal adhesions, more complex actin-cytoskeletal rearrangements, and increased levels of total beta(1)-integrin, c-Src, and paxillin. Immunoblot analysis of adhesive cells using site-specific phospho-antibodies demonstrated ARPKD-associated loss of activation of focal adhesion kinase (FAK) by phosphorylation at its autophosphorylation site (Y397); accelerated FAK inhibition by phosphorylation at Y407, S843, and S910; as well as increased activation of c-Src at pY418. Paxillin coimmunoprecipitation analyses suggested that fibrocystin-1 was a component of the normal focal adhesion complex and that actin and fibrocystin-1 were lost from ARPKD complexes.

Autosomal dominant polycystic kidney disease (ADPKD) is a commonly inherited renal disorder caused by defects in the PKD1 or PKD2 genes. ADPKD is associated with significant morbidity, and is a major underlying cause of end-stage renal failure (ESRF). Commonly, treatment options are limited to the management of hypertension, cardiovascular risk factors, dialysis, and transplantation when ESRF develops, although several new pharmacotherapies, including rapamycin, have shown early promise in animal and human studies. Evidence implicates polycystin-1 (PC-1), the gene product of the PKD1 gene, in regulation of the mTOR pathway. Here we demonstrate a mechanism by which the intracellular, carboxy-terminal tail of polycystin-1 (CP1) regulates mTOR signaling by altering the subcellular localization of the tuberous sclerosis complex 2 (TSC2) tumor suppressor, a gatekeeper for mTOR activity. Phosphorylation of TSC2 at S939 by AKT causes partitioning of TSC2 away from the membrane, its GAP target Rheb, and its activating partner TSC1 to the cytosol via 14-3-3 protein binding. We found that TSC2 and a C-terminal polycystin-1 peptide (CP1) directly interact and that a membrane-tethered CP1 protects TSC2 from AKT phosphorylation at S939, retaining TSC2 at the membrane to inhibit the mTOR pathway. CP1 decreased binding of 14-3-3 proteins to TSC2 and increased the interaction between TSC2 and its activating partner TSC1. Interestingly, while membrane tethering of CP1 was required to activate TSC2 and repress mTOR, the ability of CP1 to inhibit mTOR signaling did not require primary cilia and was independent of AMPK activation. These data identify a unique mechanism for modulation of TSC2 repression of mTOR signaling via membrane retention of this tumor suppressor, and identify PC-1 as a regulator of this downstream component of the PI3K signaling cascade.

The technical principles and practices of the civil nuclear industry are described with particular reference to fission and its products, natural and artificial radioactivity elements principally concerned and their relationships, main types of reactor, safety issues, the fuel cycle, waste management, issues related to weapon proliferation, environmental considerations and possible future developments.

The regulation of epithelial branching morphogenesis by bone morphogenetic protein-7 depends, in part, on functionally defined cyclic adenosine monophosphate (cAMP)-dependent protein kinases. We previously identified protein kinase-X (PRKX), a cAMP-dependent kinase, as a regulator of epithelial morphogenesis during kidney development and found that it binds to and phosphorylates Polycystin-1. Overexpression of PRKX stimulates renal epithelial cell migration, tubulogenesis, ureteric bud branching, and glomerular induction in embryonic mouse kidney explants in organ cultures. Here we determined the physiological functions of endogenous PRKX. Knockdown by siRNA of PRKX gene expression in a human fetal collecting tubule (HFCT) cell line exceeded 70% and resulted in decreased cell migration and increased adhesion of the cells to a collagen I matrix. In embryonic mouse kidney explants, the same degree of knockdown decreased ureteric bud branching and glomerular induction. Because PRKX BAG-3 PIN-1 and MAGI-1 are all expressed in ureteric bud derivatives, we tested for interactions among them and found that PRKX binds to all three proteins through its WW domain as determined by TransSignal domain arrays, and it coimmunoprecipitated with Pin-1 in HFCT cell lysates. These studies suggest that Polycystin-1 and Pin-1 may mediate the function of PRKX in kidney development.

As part of a longitudinal surveillance program, 35 members of a larger cohort of 77 Gulf War I veterans who were victims of depleted uranium (DU) "friendly fire" during combat underwent a 3-day clinical assessment at the Baltimore Veterans Administration Medical Center (VAMC). The assessment included a detailed medical history, exposure history, physical examination, and laboratory studies. Spot and 24-h urine collections were obtained for renal function parameters and for urine uranium (U) measures. Blood U measures were also performed. Urine U excretion was significantly associated with DU retained shrapnel burden (8.821 mug U/g creatinine [creat.] vs. 0.005 mug U/g creat., p = .04). Blood as a U sampling matrix revealed satisfactory results for measures of total U with a high correlation with urine U results (r = .84) when urine U concentrations were >/=0.1 mug/g creatinine. However, isotopic results in blood detected DU in only half of the subcohort who had isotopic signatures for DU detectable in urine. After stratifying the cohort based on urine U concentration, the high-U group showed a trend toward higher concentrations of urine beta(2) microglobulin compared to the low-U group (81.7 v. 69.0 mug/g creat.; p = .11 respectively) and retinol binding protein (48.1 vs. 31.0 mug/g creat.; p = .07 respectively). Bone metabolism parameters showed only subtle differences between groups. Sixteen years after first exposure, this cohort continues to excrete elevated concentrations of urine U as a function of DU shrapnel burden. Although subtle trends emerge in renal proximal tubular function and bone formation, the cohort exhibits few clinically significant U-related health effects.

Purpose: To evaluate change from baseline to 12 months follow-up in study and nonstudy (fellow) eye visual fields from the Ischemic Optic Neuropathy Decompression Trial (IONDT)., Design: Randomized controlled trial and observational study., Participants: The IONDT enrolled patients >or=50 years with acute nonarteritic ischemic optic neuropathy (NAION). Randomized patients (n = 258) had visual acuity 20/64 or refused randomization., Interventions: Optic nerve decompression surgery (n = 127) or careful follow-up (n = 131)., Main Outcome Measures: We measured visual fields at baseline and at 6 and 12 months follow-up. Using a computerized system, we classified visual field defects by pattern, location, and severity. We examined changes over time by treatment group, age, baseline comorbidities, and change in visual acuity. In fellow (nonstudy) eyes, we assessed change by whether NAION was present at baseline and also incidence of NAION by whether a visual field defect was present at baseline., Results: We analyzed 245 study eye visual field pairs (179 and 66, randomized and nonrandomized, respectively) for change from baseline to 12 months. We observed significant changes in defect distribution within the central field (P = 0.02) for randomized eyes. Superior and inferior altitudinal defects were less severe at follow-up in both randomized and nonrandomized eyes. We observed an association between change in central field severity and change in visual acuity from baseline (P<0.001 at 6 months; P = 0.01 at 12 months; Kendall's tau-b), but no association between visual field change and treatment group, age, or baseline comorbidities. Superior and inferior visual field defects present at baseline in nonstudy eyes improved at follow-up. Fellow (nonstudy) eyes with normal fields did not have an increased risk of developing NAION compared with eyes with >or=1 defects., Conclusions: Visual fields of NAION patients enrolled in the IONDT were relatively stable from baseline to follow-up. A visual field defect in the nonstudy eye at baseline was not associated with development of NAION during follow-up compared with eyes with normal fields.

Autosomal dominant polycystic kidney disease (ADPKD) is a major cause of end-stage renal disease in adults. Autosomal recessive (AR) PKD affects approximately 1:20,000 live-born children with high perinatal mortality. Both diseases have abnormalities in epithelial proliferation, secretion, and cell-matrix interactions, leading to progressive cystic expansion and associated interstitial fibrosis. Cell number in a kidney reflects the balance between proliferation and apoptosis. Apoptosis results from extrinsic (ligand-induced, expression of caspase-8) and intrinsic (mitochondrial damage, expression of caspase-9) triggers. Previous studies have suggested a role for apoptosis in PKD cyst formation and parenchymal destruction. Mechanisms underlying apoptosis in human ADPKD and ARPKD were examined by quantitative immunohistochemistry and Western immunoblot analyses of age-matched normal and PKD tissues. Caspase-8 expression was significantly greater in small cysts and normal-appearing tubules than in larger cysts in ADPKD kidneys. Caspase-8 also appeared early in the disease process of ADPKD. In ARPKD, expression of caspase-8 was most pronounced in later stages of the disease and was not confined to a specific cyst size. In conclusion, apoptosis in human ADPKD is an early event, occurring predominantly in normal-appearing tubules and small cysts, and is triggered by an extrinsic factor, but it occurs later in ARPKD.

Mutations of cilia-expressed proteins are associated with an attenuated shear-induced increase in intracellular Ca(2+) concentration ([Ca(2+)](i)) in renal epithelial cell lines derived from murine models of autosomal recessive polycystic kidney disease (ARPKD). We hypothesized that human ARPKD cyst-lining renal epithelial cells also exhibited dysregulated mechanosensation. To test this, conditionally immortalized cell lines derived from human fetal ARPKD cyst-lining (pool and clone 5E) cell lines with low levels of fibrocystin/polyductin expression and age-matched normal collecting tubule [human fetal collecting tubule (HFCT) pool and clone 2C] cell lines were grown in culture, loaded with a Ca(2+) indicator dye, and subjected to laminar shear. Clonal cell lines were derived from single cells present in pools of cells from cyst-lining and collecting tubules, microdissected from human kidney. Resting and peak [Ca(2+)](i) were similar between ARPKD 5E and pool, and HFCT 2C and pool; however, the flow-induced peak [Ca(2+)](i) was greater in ARPKD 5E (700 +/- 87 nM, n = 21) than in HFCT 2C (315 +/- 58 nM, n = 12; P < 0.01) cells. ARPKD 5E cells treated with Gd(3+), an inhibitor of nonselective cation channels, inhibited but did not abolish the shear-induced [Ca(2+)](i) transient. Cilia were approximately 20% shorter in ARPKD than HFCT cells, but no difference in ciliary localization or total cellular expression of polycystin-2, a mechanosenory Gd(3+)-sensitive cation channel, was detected between ARPKD and HFCT cells. The intracellular Ca(2+) stores were similar between cells. In summary, human ARPKD cells exhibit an exaggerated Gd(3+)-sensitive mechano-induced Ca(2+) response compared with controls; whether this represents dysregulated polycystin-2 activity in ARPKD cells remains to be explored.

Autosomal dominant polycystic kidney disease (ADPKD) is a common, genetically determined developmental disorder of the kidney that is characterized by cystic expansion of renal tubules and is caused by truncating mutations and haplo-insufficiency of the PKD1 gene. Several defects in cAMP-mediated proliferation and ion secretion have been detected in ADPKD cyst-lining epithelia. Unlike the ubiquitous PKA, the cAMP-dependent CREB-kinase, Protein Kinase X (PRKX) is developmentally regulated, tissue restricted and induces renal epithelial cell migration, and tubulogenesis in vitro as well as branching morphogenesis of ureteric bud in developing kidneys. The possibility of functional interactions between PKD1-encoded polycystin-1 and PRKX was suggested by the renal co-distribution of PRKX and polycystin-1 and the binding and phosphorylation of the C-terminal of polycystin-1 by PRKX at S4166 in vitro. Early consequences of PKD1 mutation include increased tubule epithelial cell-matrix adhesion, decreased migration, reduced ureteric bud branching and aberrant renal tubule dilation. To determine whether PRKX might counteract the adverse effects of PKD1 mutation, human ADPKD epithelial cell lines were transfected with constitutively active PRKX and shown to rescue characteristic adhesion and migration defects. In addition, the co-injection of constitutively active PRKX with inhibitory pMyr-EGFP-PKD1 into the ureteric buds of mouse embryonic kidneys in organ culture resulted in restoration of normal branching morphogenesis without cystic tubular dilations. These results suggest that PRKX can restore normal function to PKD1-deficient kidneys and have implications for the development of preventative therapy for ADPKD.

Bioprospecting for novel antimicrobials increasingly relies on extremely small samples unsuitable for conventional bulk extraction and assay. We developed a microtitre plate assay for minimal amounts of test materials which is rapid, extremely sensitive, allows time-course analysis and reduces false negatives. Developed for the analyses of antimicrobial sensitivity and resistance, the technique is appropriate for assays where source materials are scarce.

Polycystic kidney disease (PKD) is a diverse group of human monogenic lethal conditions inherited as autosomal dominant (AD) or recessive (AR) traits. Recent development of genetically engineered mouse models of ADPKD, ARPKD, and nephronophthisis/medullary cystic disease (NPHP) are providing additional insights into the molecular mechanisms governing of these disease processes as well as the developmental differentiation of the normal kidney. Genotypic and phenotypic mouse models are discussed and provide evidence for the fundamental involvement of cell-matrix, cell-cell, and primary cilia-lumen interactions, as well as epithelial proliferation, apoptosis, and polarization. Structure/function relationships between the PKD1, PKD2, PKHD1, and NPHP genes and proteins support the notion of a regulatory multiprotein cystic complex with a mechanosensory function that integrates signals from the extracellular environment. The plethora of intracellular signaling cascades that can impact renal cystic development suggest an exquisitely sensitive requirement for integrated downstream transduction and provide potential targets for therapeutic intervention. Appropriate genocopy models that faithfully recapitulate the phenotypic characteristics of the disease will be invaluable tools to analyze the effects of modifier genes and small molecule inhibitor therapies.

A cohort of seventy-four 1991 Gulf War soldiers with known exposure to depleted uranium (DU) resulting from their involvement in friendly-fire incidents with DU munitions is being followed by the Baltimore Veterans Affairs Medical Center. Biennial medical surveillance visits designed to identify uranium-related changes in health have been conducted since 1993. On-going systemic exposure to DU in veterans with embedded metal fragments is indicated by elevated urine uranium (U) excretion at concentrations up to 1,000-fold higher than that seen in the normal population. Health outcome results from the subcohort of this group of veterans attending the 2005 surveillance visit were examined based on two measures of U exposure. As in previous years, current U exposure is measured by determining urine U concentration at the time of their surveillance visit. A cumulative measure of U exposure was also calculated based on each veteran's past urine U concentrations since first exposure in 1991. Using either exposure metric, results continued to show no evidence of clinically significant DU-related health effects. Urine concentrations of retinol binding protein (RBP), a biomarker of renal proximal tubule function, were not significantly different between the low vs. high U groups based on either the current or cumulative exposure metric. Continued evidence of a weak genotoxic effect from the on-going DU exposure as measured at the HPRT (hypoxanthine-guanine phosphoribosyl transferase) locus and suggested by the fluorescent in-situ hybridization (FISH) results in peripheral blood recommends the need for continued surveillance of this population.

This review focuses on the mechanisms that underlie the development of human renal cystic diseases. A pathological, clinical, and pathophysiological overview is given. Initial analysis of the cell biology of inappropriate hyperproliferation accompanied by fluid secretion of cyst-lining epithelia has been followed by the elucidation of fundamental defects in epithelial polarity, cell-matrix and cell-cell interactions, and apoptosis, all of which are discussed. Identification of the genes and proteins responsible for several renal cystic diseases has led to a more complete understanding of defects in renal developmental programming, differentiation, and morphogenesis, all of which underlie cystic diseases of the kidney.

Background: The objective of this report is to describe the methods used to develop and validate a computerized system to analyze Humphrey visual fields obtained from patients with non-arteritic anterior ischemic optic neuropathy (NAION) and enrolled in the Ischemic Optic Neuropathy Decompression Trial (IONDT). The IONDT was a multicenter study that included randomized and non-randomized patients with newly diagnosed NAION in the study eye. At baseline, randomized eyes had visual acuity of 20/64 or worse and non-randomized eyes had visual acuity of better than 20/64 or were associated with patients refusing randomization. Visual fields were measured before treatment using the Humphrey Field Analyzer with the 24-2 program, foveal threshold, and size III stimulus., Methods: We used visual fields from 189 non-IONDT eyes with NAION to develop the computerized classification system. Six neuro-ophthalmologists ("expert panel") described definitions for visual field patterns defects using 19 visual fields representing a range of pattern defect types. The expert panel then used 120 visual fields, classified using these definitions, to refine the rules, generating revised definitions for 13 visual field pattern defects and 3 levels of severity. These definitions were incorporated into a rule-based computerized classification system run on Excel(R) software. The computerized classification system was used to categorize visual field defects for an additional 95 NAION visual fields, and the expert panel was asked to independently classify the new fields and subsequently whether they agreed with the computer classification. To account for test variability over time, we derived an adjustment factor from the pooled short term fluctuation. We examined change in defects with and without adjustment in visual fields of study participants who demonstrated a visual acuity decrease within 30 days of NAION onset (progressive NAION)., Results: Despite an agreed upon set of rules, there was not good agreement among the expert panel when their independent visual classifications were compared. A majority did concur with the computer classification for 91 of 95 visual fields. Remaining classification discrepancies could not be resolved without modifying existing definitions. Without using the adjustment factor, visual fields of 63.6% (14/22) patients with progressive NAION and no central defect, and all (7/7) patients with a paracentral defect, worsened within 30 days of NAION onset. After applying the adjustment factor, the visual fields of the same patients with no initial central defect and 5/7 of the patients with a paracentral defect were seen to worsen., Conclusion: The IONDT developed a rule-based computerized system that consistently defines pattern and severity of visual fields of NAION patients for use in a research setting.

Branching of ureteric bud-derived epithelial tubes is a key morphogenetic process that shapes development of the kidney. Glial cell line-derived neurotrophic factor (GDNF) initiates ureteric bud formation and promotes subsequent branching morphogenesis. Exactly how GDNF coordinates branching morphogenesis is unclear. Here we show that the absence of the receptor tyrosine kinase antagonist Sprouty1 (Spry1) results in irregular branching morphogenesis characterized by both increased number and size of ureteric bud tips. Deletion of Spry1 specifically in the epithelium is associated with increased epithelial Wnt11 expression as well as increased mesenchymal Gdnf expression. We propose that Spry1 regulates a Gdnf/Ret/Wnt11-positive feedback loop that coordinates mesenchymal-epithelial dialogue during branching morphogenesis. Genetic experiments indicate that the positive (GDNF) and inhibitory (Sprouty1) signals have to be finely balanced throughout renal development to prevent hypoplasia or cystic hyperplasia. Epithelial cysts develop in Spry1-deficient kidneys that share several molecular characteristics with those observed in human disease, suggesting that Spry1 null mice may be useful animal models for cystic hyperplasia.

The mechanisms of action of polycystin-1 (PC1) have been difficult to dissect because of its interaction with multiple factors, the heterogeneity of the genetic mutations, and the complexity of the experimental animal models. Here, stable knockdown of PC1 in MDCK epithelial cells was achieved by lentiviral-mediated delivery of a specific small interfering RNA for PKD1. The reduction of PC1 expression prevented tubulogenesis in three-dimensional collagen type I culture in response to hepatocyte growth factor and induced formation of cysts. PC1 knockdown created a condition of haploinsufficiency that led to hyperproliferation, increased adhesion to collagen type I, and increased apoptosis. It was shown that the suppression of PC1 was associated with the increased expression of integrin-alpha2beta1 and reduced apoptosis in cells grown on collagen type I. The engagement of integrin-alpha2beta1 seemed to be essential for the survival because PC1 knockdown cells were significantly less susceptible to anoikis by a mechanism that was reversible by anti-integrin-alpha2beta1 blocking antibodies. Overall, these data link integrin-alpha2beta1 to some of the biologic functions that are ascribed to PC1 and establish the potential of this approach for the direct study of PC1 functions in a genetically defined background. Furthermore, these findings indicate that reduction of PC1 expression levels, rather than the loss of heterozygosity, may be sufficient to induce cystogenesis.

Background: Laparoscopic colposuspension was one of the first minimal access operations for the treatment of women with stress urinary incontinence, with the presumed advantages over traditional Burch colposuspension of avoiding major incisions, shorter hospital stay, and quicker return to normal activities. A variety of approaches and methods are used., Objectives: To determine the effects of laparoscopic colposuspension for urinary incontinence., Search Strategy: We searched the Cochrane Incontinence Group Specialised Trials Register (searched 21 September 2005). Additional trials were sought from other sources such as reference lists, reviews and researchers and authors were contacted for unpublished data and trials., Selection Criteria: Randomised or quasi-randomised controlled trials in women with symptomatic or urodynamic diagnosis of stress or mixed incontinence that included laparoscopic surgery in at least one arm of the study., Data Collection and Analysis: Trials were evaluated for methodological quality and appropriateness for inclusion by the reviewers. Data were extracted by two of the reviewers and cross checked by another. Trial data were analysed by intervention. Where appropriate, a summary statistic was calculated., Main Results: Twenty-one eligible trials were identified. Nine involved the comparison of laparoscopic with open colposuspension. Whilst the women's subjective impression of cure seemed similar for both procedures in the short and medium term follow-up, there was some evidence of poorer results of laparoscopic colposuspension, within 18 months, on objective outcomes. Two poor quality trials reported conflicting long term results (after five years) for this comparison. No significant differences were observed for post-operative urgency, voiding dysfunction or de novo detrusor overactivity. Trends were shown towards a lower perioperative complication rate, longer operating time, less intraoperative blood loss, less postoperative pain, shorter hospital stay, quicker return to normal activities, and shorter duration of catheterisation for laparoscopic compared with open colposuspension. Benefits did not come without a price, as laparoscopic colposuspension in the short term is more costly.Eight studies compared laparoscopic colposuspension with newer 'self-fixing' vaginal slings. Overall there were no significant differences in the reported subjective cure rates of the two procedures, however vaginal sling procedures did have significantly higher objective cure rates at 18 months. No significant differences were observed for post-operative voiding dysfunction, de novo detrusor activity and perioperative complications. Laparoscopic colposuspension has a significantly longer operation time, longer hospital stay and slower return to normal activities when compared to the sling procedures. Significantly higher subjective and objective (dry on 'ultrashort' pad test) one year cure rates were found for women randomised to two paravaginal sutures compared with one suture in a single trial (89% versus 65% and 83% versus 58% respectively). Two small studies compared sutures with mesh and staples for laparoscopic colposuspension and the comparisons, although showing a trend towards favouring the sutures, were not significant. One study compared transperitoneal with extraperitoneal access for laparoscopic colposuspension but it was also small and of poor quality., Authors' Conclusions: The long-term performance of laparoscopic colposuspension remains uncertain. Currently available evidence suggests that it may be as good as open colposuspension at two years post surgery. Like other laparoscopically performed operations, patients having laparoscopic colposuspension recovered quicker, but the operation itself took longer to perform. However, the newer vaginal sling procedures appear to offer even greater benefits of minimal access surgery and better objective outcomes in the short-term. If laparoscopic colposuspension is performed, two paravaginal sutures appear to be more effective than one. The place of laparoscopic colposuspension in clinical practice should become clearer when ongoing trials are reported and when there are more data available describing long-term cure results.

Autosomal Dominant Polycystic Kidney Disease (ADPKD) is a very common lethal monogenetic disease with significant morbidities and a high likelihood of progression to renal failure for which there is no proven disease-specific therapy currently available for clinical use. Human ADPKD cystic epithelia have proliferative abnormalities mediated by EGFR over-expression and mispolarization leading autocrine response to EGF family ligands. We now show that apical localization of EGFR complexes in normal fetal and ADPKD epithelia is associated with heterodimerization of EGFR(HER-1) with HER-2(neu/ErbB2), while basal membrane localization in normal adult renal epithelia is associated with EGFR(HER-1) homodimers. Since ADPKD epithelial cells have reduced migratory function, this was used as a bioassay to evaluate the ability of compounds to rescue the aberrant human ADPKD phenotype. General tyrosine kinase inhibition by herbimycin and specific inhibition of HER-2(neu/ErbB2) by AG825 or pretreatment with ErbB2 siRNA reversed the migration defect of ADPKD epithelia. Selective inhibition of EGFR(HER-1) showed partial rescue. Increased ADPKD cell migration after inhibition of p38MAP kinase but not of PI3-kinase implicated p38MAPK downstream of HER-2(neu/ErbB2) stimulation. Daily administration of AG825 to PKD1 null heterozygous mice significantly inhibited the development of renal cysts. These studies implicate HER2(neu/ErbB2) as an effector of apical EGFR complex mispolarization and that its inhibition should be considered a candidate for clinical therapy of ADPKD.

Dysregulated apoptosis of renal tubular epithelial cells (RTEC) is an important component of the pathogenesis of several renal diseases, including HIV-associated nephropathy (HIVAN), the most common cause of chronic kidney failure in HIV-infected patients. In HIVAN, RTEC become infected by HIV-1 in a focal distribution, and HIV-1 infection has been shown to induce apoptosis in vitro. In microarray studies that used a novel renal tubular epithelial cell line from a patient with HIVAN, it was found that the ubiquitin-like protein FAT10 is one of the most upregulated genes in HIV-infected cells. Previously, FAT10 was shown to induce apoptosis in murine fibroblasts. The expression of FAT10 in HIVAN and the ability of FAT10 to induce apoptosis in human RTEC therefore were studied. This study revealed that FAT10 expression is induced after infection of RTEC by HIV-1 and that expression of FAT10 induces apoptosis in RTEC in vitro. Moreover, it was found that inhibition of endogenous FAT10 expression abrogated HIV-induced apoptosis of RTEC. Immunohistochemical studies demonstrated increased FAT10 expression in a murine model of HIVAN, in HIVAN biopsy samples, and in autosomal dominant polycystic kidney disease, another renal disease that is characterized by cystic tubular enlargement and epithelial apoptosis. These results suggest a novel role for FAT10 in epithelial apoptosis, which is an important component of the pathogenesis of many renal diseases.

[reaction: see text] The first total synthesis of the C(3)-symmetric and biologically active natural product, (-)-xyloketal A, has been accomplished in one step from phloroglucinol (1,3,5-trihydroxybenzene) and (4R)-3-hydroxymethyl-2,4-dimethyl-4,5-dihydrofuran. This remarkably direct process involved an exceedingly facile and diastereoselective boron trifluoride diethyl etherate-promoted triple electrophilic aromatic substitution reaction that was coupled to three bicyclic acetal formation reactions.

The synthesis of a series of chiral nonracemic and C2-symmetric 2,2'-bipyridyl ligands (R = Me, i-Pr and Ph) as well as the syntheses of the corresponding unsymmetric 2,2'-bipyridyl ligands (R = Me and Ph) is described. These bipyridyl ligands were prepared, in a notably direct and modular fashion, from the readily available and corresponding 2-chloropyridine acetals (R = Me, i-Pr and Ph). The bipyridyl ligands were evaluated in copper(I)-catalyzed cyclopropanation reactions of styrene with the ethyl and t-butyl esters of diazoacetic acid. The stereoselectivities, as well as the yields of the cyclopropanation reactions, were dependant on the ratio of the bipyridyl ligands and copper triflate that was employed. The best result was obtained in the asymmetric cyclopropanation reaction of styrene and tert-butyl diazoacetate with the C2-symmetric bipyridyl ligand (R = i-Pr). This afforded the corresponding trans-cyclopropane in good diastereoselectivity (4 : 1) and in moderate enantioselectivity (44% ee). The X-ray structure determination of a complex formed between the C2-symmetric 2,2'-bipyridyl ligand (R = Ph) and copper(I) chloride showed that two bipyridyl ligands had coordinated to the copper(I) ion. This information, along with the results of a series of cyclopropanation reactions and NMR data, led to the conclusion that the 2,2'-bipyridyl ligands had the propensity to form catalytically inactive bis-ligated copper(I) species in solution that were in equilibrium with catalytically active copper(i) triflate and the desired mono-ligated copper(I) species. Moreover, it was observed that the complex of the bipyridyl ligand (R = Ph) and copper(I) chloride had a particularly large optical rotation (sodium D-line). The maximum positive optical rotation was subsequently found to be +1.1 x 10(4) at 304 nm and the maximum negative optical rotation was -1.3 x 10(4) at 329 nm.

A series of demethyl analogues of the natural products xyloketal A, B, C, D, and G have been prepared in a notably direct manner from 3-hydroxymethyl-2-methyl-4,5-dihydrofuran and a series of corresponding phenols. These syntheses featured a boron trifluoride diethyl etherate-promoted electrophilic aromatic substitution reaction as a key step. In the case of the synthesis of analogues of xyloketal A, the process was found to be highly efficient (up to 93% yield). The optimized isolated yield of these reaction products is remarkable in view of the fact that this transformation involves, minimally, six individual reactions. Moreover, these synthetic studies provide significant insight into the possible biogenic origin of the xyloketal natural products.

Objective: To investigate the prevalence of persistent and long term postpartum urinary incontinence and associations with mode of first and subsequent delivery., Design: Longitudinal study., Setting: Maternity units in Aberdeen (Scotland), Birmingham (England) and Dunedin (New Zealand)., Population: Women (4214) who returned postal questionnaires three months and six years after the index birth., Methods: Symptom data were obtained from both questionnaires and obstetric data from case-notes for the index birth and the second questionnaire for subsequent births. Logistic regression investigated the independent effects of mode of first delivery and delivery mode history., Main Outcome Measures: Urinary incontinence-persistent (at three months and six years after index birth) and long term (at six years after index birth)., Results: The prevalence of persistent urinary incontinence was 24%. Delivering exclusively by caesarean section was associated with both less persistent (OR=0.46, 95% CI 0.32-0.68) and long term urinary incontinence (OR=0.50, 95% CI 0.40-0.63). Caesarean section birth in addition to vaginal delivery, however, was not associated with significantly less persistent incontinence (OR 0.93, 95% CI 0.67-1.29). There were no significant associations between persistent or long term urinary incontinence and forceps or vacuum extraction delivery. Other significantly associated factors were increasing number of births and older maternal age., Conclusions: The risk of persistent and long term urinary incontinence is significantly lower following caesarean section deliveries but not if there is another vaginal birth. Even when delivering exclusively by caesarean section, the prevalence of persistent symptoms (14%) is still high.

Objective: To identify obstetric and other risk factors for urinary incontinence that occurs during pregnancy or after childbirth., Design: Questionnaire survey of women., Setting: Maternity units in Aberdeen (Scotland), Birmingham (England) and Dunedin (New Zealand)., Population: A total of 3405 primiparous women with singleton births delivered during 1 year., Methods: Questionnaire responses and obstetric case note data were analysed using multivariate analysis to identify associations with urinary incontinence., Main Outcome Measures: Urinary incontinence at 3 months after delivery first starting in pregnancy or after birth., Results: The prevalence of urinary incontinence was 29%. New incontinence first beginning after delivery was associated with older maternal age (oldest versus youngest group, OR 2.02, 95% CI 1.35-3.02) and method of delivery (caesarean section versus spontaneous vaginal delivery, OR 0.28, 95% CI 0.19-0.41). There were no significant associations with forceps delivery (OR 1.18, 95% CI 0.92-1.51) or vacuum delivery (OR 1.16, 95% CI 0.83-1.63). Incontinence first occurring during pregnancy and still present at 3 months was associated with higher maternal body mass index (BMI>25, OR 1.68, 95% CI 1.16-2.43) and heavier babies (birthweight in top quartile, OR 1.56, 95% CI 1.12-2.19). In these women, caesarean section was associated with less incontinence (OR 0.39, 95% CI 0.27-0.58) but incontinence was not associated with age., Conclusions: Women have less urinary incontinence after a first delivery by caesarean section whether or not that first starts during pregnancy. Older maternal age was associated with new postnatal incontinence, and higher BMI and heavier babies with incontinence first starting during pregnancy. The effect of further deliveries may modify these findings.

The evaluation of a chiral, nonracemic and C2-symmetric 2,2'-bipyridyl ligand in copper(I)-catalyzed asymmetric allylic oxidation reactions of a series of cyclic alkenes with tert-butyl peroxybenzoate is reported (up to 91% ee, the highest reported enantioselectivity for a bipyridyl ligand copper(I) complex to date).

Objective: To relate medical surveillance outcomes to uranium biomonitoring results in a group of depleted uranium (DU)-exposed, Gulf War I veterans., Methods: Thirty-two veterans of Gulf War I who were victims of 'friendly fire' involving DU weapons, in whom exposure assessment can accurately be measured, had urine uranium concentrations determined using ICP-MS technology. Clinical laboratory parameters were measured and related to urine uranium concentrations. Data were examined by stratifying the cohort into a low U group, <0.10 mug/g creatinine versus a high U group, >/=0.10 mug/g creatinine and assessing differences between groups., Results: Over a decade after first exposure, soldiers possessing embedded DU fragments continue to excrete elevated concentrations of uranium in urine. No clinically significant uranium related health effects were observed in blood count, blood chemistries including renal markers, neuropsychological measures, and semen quality or genotoxicity measures. Markers of early changes in renal glomerular and tubular function were not statistically different between groups; however, genotoxicity measures continue to show subtle, mixed results., Conclusion: Persistent urine uranium elevations continue to be observed more than 12 years since first exposure. Despite this, renal and other clinical abnormalities were not observed, likely due to the 'relatively' low uranium burden in this cohort compared to historical uranium-exposed occupational groups. Continuing surveillance is indicated, however, due to the on-going nature of the exposure. These results are an important finding in light of the on-going controversy regarding health effects observed in soldiers of the Gulf War and other conflicts, whose uranium exposure assessment is unable to be accurately determined.

The issue of traumatic damage to the pelvic floor in childbirth is attracting more and more attention amongst obstetric caregivers and laypersons alike. This is partly due to the fact that elective caesarean section as a potentially preventative intervention is increasingly available and perceived as safe. As there is a multitude of emotive issues involved, including health economics and the relative roles of healthcare providers, the discussion surrounding pelvic floor trauma in childbirth has not always been completely rational. However, after 25 years of urogynaecological research in this field it should be possible to determine whether pelvic floor trauma in childbirth is myth or reality, and, if real, whether it matters for the pathogenesis of incontinence and prolapse. On reviewing the available evidence, it appears that there are sufficient grounds to assume that vaginal delivery (or even the attempt at vaginal delivery) can cause damage to the pudendal nerve, the inferior aspects of the levator ani muscle and fascial pelvic organ supports. Risk factors for such damage have been defined and variously include operative vaginal delivery, a long second stage, and macrosomia. It is much less clear, however, whether such trauma is clinically relevant, and how important it is in the aetiology of pelvic floor morbidity later in life.

The human protein kinase X (PRKX) gene was identified previously as a cAMP-dependent serine/threonine kinase that is aberrantly expressed in autosomal dominant polycystic disease kidneys and normally expressed in fetal kidneys. The PRKX kinase belongs to a serine/threonine kinase family that is phylogenetically and functionally distinct from classical protein kinase A kinases. Expression of PRKX activates cAMP-dependent renal epithelial cell migration and tubular morphogenesis in cell culture, suggesting that it might regulate branching growth of the collecting duct system in the fetal kidney. With the use of a mouse embryonic kidney organ culture system that recapitulates early kidney development in vitro, it is demonstrated that lentiviral vector-driven expression of a constitutively active, cAMP-independent PRKX in the ureteric bud epithelium stimulates branching morphogenesis and results in a 2.5-fold increase in glomerular number. These results suggest that PRKX stimulates epithelial branching morphogenesis by activating cell migration and support a role for this kinase in the regulation of nephrogenesis and of collecting system development in the fetal kidney.

The polycystic kidney disease (PKD1) gene-encoded protein, polycystin-1, is developmentally regulated, with highest expression levels seen in normal developing kidneys, where it is distributed in a punctate pattern at the basal surface of ureteric bud epithelia. Overexpression in ureteric epithelial cell membranes of an inhibitory pMyr-GFP-PKD1 fusion protein via a retroviral (VVC) delivery system and microinjection into the ureteric bud lumen of embryonic day 11 mouse metanephric kidneys resulted in disrupted branching morphogenesis. Using confocal quantitative analysis, significant reductions were measured in the numbers of ureteric bud branch points and tips, as well as in the total ureteric bud length, volume and area, while significant increases were seen as dilations of the terminal branches, where significant increases in outer diameter and volumes were measured. Microinjection of an activating 5TM-GFP-PKD1 fusion protein had an opposite effect and showed significant increases in ureteric bud length and area. These are the first studies to experimentally manipulate polycystin-1 expression by transduction in the embryonic mouse kidney and suggest that polycystin-1 plays a critical role in the regulation of epithelial morphogenesis during renal development.

Unlabelled: Older black men have higher adjusted BMD than older white men. Using data from a longitudinal cohort study of older men followed for a mean of 18.8 +/- 6.5 (SD) months, we found that older black men have a higher rate of decline in femoral neck and total hip BMD and femoral neck BMAD than older white men., Introduction: Older black men have higher adjusted BMD compared with older white men. The difference in BMD may be caused by having attained higher peak bone mass as young adults and/or having a slower rate of decline in bone mass as adults. There are few published longitudinal data on change in bone mass in older white men and no published data for older black men., Materials and Methods: Three hundred forty-nine white men and 119 black men 65 of age (mean age, 75 +/- 5.7 and 72 +/- 5.6 years, respectively) who participated in the longitudinal component of the Baltimore Men's Osteoporosis Study returned for a second visit after a mean of 18.8 +/- 6.5 (SD) months and were not taking medications used to treat low bone mass at either visit. BMD was measured at the femoral neck and total hip by Hologic-certified technicians using a QDR 2000 at the baseline visit (V1) and QDR 4500 at the first follow-up visit (V2). Participants also completed self-administered and interviewer-administered questionnaires and underwent standardized clinic examinations. Bone mineral apparent density (BMAD) at the femoral neck was calculated as an estimate of volumetric BMD. Annual crude and multiple variable adjusted percent changes in BMD and BMAD were calculated., Results: In univariate analyses, black men had lower percent decline in femoral neck and total hip BMD and femoral neck BMAD than white men. In addition, older age at baseline, lower baseline weight, current smoking, and lower baseline BMD were associated with greater percent decline per year in femoral neck BMD; older age at baseline, current smoking, and lower baseline BMD were associated with greater percent decline per year in total hip BMD; and older age at baseline and lower baseline femoral neck BMAD were associated with greater percent decline per year in femoral neck BMAD. Racial differences in bone loss persisted in multiple variable models that controlled for other factors associated with change in BMD and BMAD., Conclusions: Older black men seem to lose bone mass at a slower rate than older white men. These differences in the rate of bone loss may account, in part, for the racial disparities in BMD and BMAD and risk of osteoporotic fractures among older men.

Objective: To define relationships between hoof-acceleration patterns of galloping horses and dynamic properties of the track., Animals: 8 Thoroughbred horses without lameness., Procedure: Acceleration-time curves were recorded by use of accelerometers attached to each hoof as each horse galloped over the track straightaway. Four sessions were conducted for each horse, with the track surface modified by sequentially adding water before each session. These acceleration-time curves were analyzed to determine peak accelerations during the support phase of the stride. Track dynamic properties (hardness, rebound, deceleration rate, rebound rate, and penetration) were recorded with a track-testing device. Moisture content and dry density were measured from soil samples. Stepwise multiple regression was used to identify relationships between hoof-acceleration variables and track dynamic properties., Results: Track rebound rate was most consistently related to hoof variables, especially through an inverse relationship with negative acceleration peaks for all hooves. Also, rebound rate was related to initial acceleration peak during propulsion of the hooves of the forelimb and the nonlead hind limb as well as to the second acceleration peak during propulsion of the lead hooves of the hind limb and nonlead forelimb., Conclusions and Clinical Relevance: The inverse relationship between track rebound rate and negative acceleration peaks for all hooves reflects the most important dynamic property of a track. Any factor that reduces negative acceleration of the hooves will increase stride efficiency by allowing smoother transition from retardation to propulsion and therefore may be important in determining the safety of racing surfaces.

Enantioselective Friedel-Crafts alkylation reactions of a series of substituted indoles with methyl trifluoropyruvate, catalyzed by a chiral nonracemic C(2)-symmetric 2,2'-bipyridyl copper(II) triflate complex, are described. The corresponding 3,3,3-trifluoro-2-hydroxy-2-indole-3-yl-propionic acid methyl esters were formed in good yield and in high enantiomeric excess (up to 90%). This is the first report of the use of a chiral nonracemic 2,2'-bipyridyl ligand in catalytic and enantioselective Friedel-Crafts alkylation reactions. The structural characterization of a copper(II) chloride complex of the chiral 2,2'-bipyridyl ligand by X-ray crystallography is also presented. [reaction: see text]