Intracranial haemorrhage in thrombocytopenic haematology patients--a nested case-control study: the InCiTe study protocol.
Intracranial haemorrhage (ICH) is one of the most serious side-effects of severe thrombocytopenia in haematology patients. ICH is rare, but can have devastating consequences (death or major morbidity). It is unknown why some patients with severe thrombocytopenia bleed and others do not. Primary aim was to identify risk factors for ICH in patients with haematological malignancies. Secondary aims were to identify short-term outcomes for these patients at 30 days (major morbidity and mortality) and produce a more accurate estimate of ICH incidence in this population. This information is key to identifying means to improve treatment and quality of care. This is a UK-wide case-control study of ICH nested within a 4-year prospective surveillance study set up specifically for the case-control study. Each case will be matched to one control. Cases will be adult haematology patients (≥16 years) who have had any type or severity of ICH who are receiving, about to receive or have just received myeloablative chemotherapy (defined as chemotherapy expected to cause a significant thrombocytopenia <50×10(9)/L for >5 days) or a haemopoietic stem cell transplant. Only patients being treated with curative intent will be included. Controls will be patients who fulfil the same inclusion criteria as cases (apart from ICH) and were treated at the same hospital immediately before the index case. Cases and controls will be matched to type of treatment (myeloablative chemotherapy or haemopoietic stem cell transplant). Hospitals across the UK will participate in a monthly email reporting strategy (started June 2011), as to whether a case of ICH occurred during the preceding calendar month. Case and control forms will be sent to any hospital reporting an eligible case. Conditional logistic regression will be used to calculate ORs. Denominator data for incidence estimates will use national registry data. ISRCTN05026912 (prospective registration). NIHR Portfolio (UKCRN ID 10712).


Does restructuring hospitals result in greater efficiency?--An empirical test using diachronic data.
Hospitals are being restructured more frequently. Increased cost efficiency is the usual justification given for such changes. All 20 major teaching hospitals in Australia's two most populous states were investigated by classifying each over a 5-6 year period in terms of their cost efficiency (average cost per case weighted by Australian diagnosis-related group [AN-DRG] data and adjusted for inflation) and structure, categorized as traditional-professional (TP), clinical-divisional (CD), or clinical-institute (CI). In all, 12 hospitals changed structure during the study period. There was slight evidence that CD structures were more efficient than TP structures but this was not supported by other evidence. There were no significant differences in efficiency in the first or second years following changes from either TP to CD or TP to CI structures. All four hospitals changing from CD to CI structure became significantly less efficient. This may be due to frequency rather than type of change as they were the only hospitals that implemented two structural changes. Hospitals that changed or did not change structure were similar in efficiency at the beginning and at the end of the study period, in overall efficiency during the period, and in trends toward efficiency over time. The findings challenge those who advocate restructuring hospitals on the grounds of improving cost efficiency.


Mechanically stimulated cytoskeleton rearrangement and cortical contraction in human neutrophils.
A mechanical test with micropipets is used to characterize cytoskeleton rearrangement and contraction induced by mechanical stresses in human neutrophils. The yield shear resultant of the cell cortex is on the order of 0.06 to 0.09 mN.m-1. The measured yield shear resultant suggests that the neutrophil cortex is a weakly cross-linked structure. When a tether is pulled out from the cell surface, a polymer structure starts to fill it and spreads out from the cell body. The rate of advancement of the polymerization front is almost constant and, therefore, is not diffusion limited. The measured rate is much smaller than the one of spontaneous actin polymerization, suggesting that the limiting process is either the dissociation of actin monomers from their dimers with the capping proteins or the rate of formation of new nucleation sites or both. Polymerization is also observed after applying sufficient mechanical stresses on a small portion of the cell surface. The polymerization is followed by mass transfer from the cell into the prestressed region and later on by contraction of the main cell body. The pressure generating the flow is located in the prestressed region and most probably is a result of its "swelling" and contraction. The contraction of the main cell body is very similar (in its time dependence and magnitude) to the contraction during phagocytosis. The measured maximum cortical tension is on the order of 0.5 mN.m-1, which for a 3.5-microns diameter pipet corresponds to a maximum contraction force of 11 nN.


Cytokine fusion constructs as DNA vaccines against tumors.
Various studies have used DNA vaccination as a method of immunizing against tumors (1-12). As with any tumor vaccine, one challenge is to find a truly tumor-specific antigen (13,14). The majority of immunologically targeted tumor antigens are also expressed on a subset of normal host cells. Examples of such antigens include prostate-specific antigen, and CD20, a B cell marker. Some tumor antigens are specific for activated cells of certain types, such as carcinoembryonic antigen (CEA) or the IL-2 receptor. These are often found on embryonic or fetal cells as well as tumor cells. The carbohydrate antigens of melanomas and the immunoglobulin (Ig) idiotype of B cell lymphomas represent tumor-specific antigens (TSA). Unfortunately, TSA have not been identified in more common malignancies. Furthermore, the antigenic determinants of known TSA may differ between patients; for example, the tumor idiotype (Id) of B cell lymphoma is highly patient-specific and must be determined for each case.


Tuberculosis in Turkish patients with systemic lupus erythematosus: increased frequency of extrapulmonary localization.
The objective was to investigate the frequency and characteristics of tuberculosis (TB) in patients with systemic lupus erythematosus (SLE). We reviewed the charts of 556 patients with SLE who were followed up between 1978 and 2001 in our lupus clinic. Patients who developed TB after the diagnosis of SLE were identified (SLE/TB+). Ninety-six consecutive patients with SLE who did not develop TB during the follow-up were evaluated as a control group (SLE/TB-). Clinical, laboratory and management characteristics of these two groups of patients were recorded according to a predefined protocol, and compared. Of the 556 patients evaluated, 20 patients (3.6%) with TB were identified. Nine of the 20 patients (45%) had extrapulmonary TB (vertebral involvement in three patients, meningeal in two, and joint and soft tissue in four). Arthritis and renal involvement were significantly high in the SLE/TB+ group (P = 0.045, P = 0.009, respectively). The mean daily dose of prednisolone before the diagnosis of TB and the cumulative dose of prednisolone were significantly higher in the SLE/TB+ group compared to the SLE/TB- group (27 +/- 22 g versus 16 +/- 16 g, 24 +/- 45 mg versus 11 +/- 8.5 mg, respectively). In conclusion, we found an increased frequency of TB infection and a high prevalence of extrapulmonary TB in a large cohort of SLE patients. The mean daily dose of prednisolone before the diagnosis of TB and the cumulative dose of prednisolone, which possibly related to disease severity, were important determinants for the increased risk of TB in these patients with SLE.


Crystal structure of the stromelysin catalytic domain at 2.0 A resolution: inhibitor-induced conformational changes.
Matrix metalloproteinases are believed to play an important role in pathological conditions such as osteoarthritis, rheumatoid arthritis and tumor invasion. Stromelysin is a zinc-dependent proteinase and a member of the matrix metalloproteinase family. We have solved the crystal structure of an active uninhibited form of truncated stromelysin and a complex with a hydroxamate-based inhibitor. The catalytic domain of the enzyme of residues 83-255 is an active fragment. Two crystallographically independent molecules, A and B, associate as a dimer in the crystals. There are three alpha-helices and one twisted, five-strand beta-sheet in each molecule, as well as one catalytic Zn, one structural Zn and three structural Ca ions. The active site of stromelysin is located in a large, hydrophobic cleft. In particular, the S1' specificity site is a deep and highly hydrophobic cavity. The structure of a hydroxamate-phosphinamide-type inhibitor-bound stromelysin complex, formed by diffusion soaking, has been solved as part of our structure-based design strategy. The most important feature we observed is an inhibitor-induced conformational change in the S1' cavity which is triggered by Tyr223. In the uninhibited enzyme structure, Tyr223 completely covers the S1' cavity, while in the complex, the P1' group of the inhibitor displaces the Tyr223 in order to fit into the S1' cavity. Furthermore, the displacement of Tyr223 induces a major conformational change of the entire loop from residue 222 to residue 231. This finding provides direct evidence that Tyr223 plays the role of gatekeeper of the S1' cavity. Another important intermolecular interaction occurs at the active sit of molecule A, in which the C-terminal tail (residues 251-255) from molecule B inserts. The C-terminal tail interacts extensively with the active site of molecule A, and the last residue (Thr255) coordinated to the catalytic zinc as the fourth ligand, much like a product inhibitor would. The inhibitor-induced conformational change and the intermolecular C-terminal-zinc coordination are significant in understanding the structure-activity relationships of the enzyme.


The Prognostic Impact of the Lymphocyte-to-Monocyte Ratio in Resected Pancreatic Head Adenocarcinoma.
Although the prognostic significance of systematic inflammation-based scores, such as the neutrophil-to-lymphocyte ratio (NLR), the platelet-to-lymphocyte ratio (PLR), and the prognostic nutritional index (PNI), has been explored in pancreatic cancers, few reports have investigated the lymphocyte-to-monocyte ratio (LMR). We aimed to retrospectively investigate the prognostic value of the preoperative LMR in patients with resectable pancreatic head cancer (PHC). From 2005 to 2016, 165 patients underwent pancreatoduodenectomy for PHC. All samples of peripheral blood were collected within 2 weeks prior to surgery. The best cutoff values of the LMR for predicting survival were determined by using a minimum p value approach (cut-off value: 2.8). The clinicopathological features of LMR <2.8 (n = 25) and ≥2.8 (n = 140) were compared. Patients with LMR ≥2.8 showed significantly lower NLR and PLR, and significantly higher PNI. Levels of CEA and CA19-9 were similar, and the pathological findings were comparable between the groups. The overall survival of patients with LMR ≥2.8 (66.2% at 1 year) was superior to that of patients with LMR <2.8 (36.1% at 1 year, p = 0.015). Multivariate analysis identified LMR <2.8 (hazard ratio 1.72, 95% CI 1.02-2.89, p = 0.042), lymphatic and venous invasion and positive surgical margin as independent prognostic factors. LMR may carry important prognostic information for patients with resectable PHC. Preoperative LMR may be considered for use in risk stratification for individual patients with PHC.


From the bush to the big smoke--development of a hybrid urban community based medical education program in the Northern Territory, Australia.
The Northern Territory (NT) of Australia is a unique setting for training medical students. This learning environment is characterised by Aboriginal health and an emphasis on rural and remote primary care practice. For over a decade the NT Clinical School (NTCS) of Flinders University has been teaching undergraduate medical students in the NT. Community based medical education (CBME) has been demonstrated to be an effective method of learning medicine, particularly in rural settings. As a result, it is rapidly gaining popularity in Australia and other countries. The NTCS adopted this model some years ago with the implementation of its Rural Clinical School; however, urban models of CBME are much less well developed than those in rural areas. There is considerable pressure to better incorporate CBME into medical student teaching environment, particularly because of the projected massive increase in student numbers over the next few years. To date, the community setting of urban Darwin, the NT capital city, has not been well utilised for medical student training. In 2008, the NTCS enrolled its first cohort of students in a new hybrid CBME program based in urban Darwin. This report describes the process and challenges involved in development of the program, including justification for a hybrid model and the adaptation of a rural model to an urban setting. Relationships were established and formalised with key partners and stakeholders, including GPs and general practices, Aboriginal medical services, community based healthcare providers and other general practice and community organisations. Other significant issues included curriculum development and review, development of learning materials and the establishment of robust evaluation methods. Development of the CBME model in Darwin posed a number of key challenges. Although the experience of past rural programs was useful, a number of distinct differences were evident in the urban setting. Change leadership and inter-professional collaboration were key strengths in the implementation and ongoing evaluation of the program. The program will provide important information about medical student training in urban community settings, and help inform other clinical schools considering the adoption of similar models.


Uterine differentiation as a foundation for subsequent fertility.
Uterine differentiation in cattle and sheep begins prenatally, but is completed postnatally. Mechanisms regulating this process are not well defined. However, studies of urogenital tract development in murine systems, particularly those involving tissue recombination and targeted gene mutation, indicate that the ideal uterine organizational programme evolves epigenetically through dynamic cell-cell and cell-matrix interactions that define the microenvironmental context within which gene expression occurs and may ensure adult tissue stability. In the cow and ewe, transient postnatal exposure of the developing uterus to steroids can produce immutable changes in adult uterine tissues that may alter the embryotrophic potential of the uterine environment. Thus, success of steroid-sensitive postnatal events supporting uterine growth and development can dictate the functional potential of the adult uterus. Studies to determine effects of specific steroidal agents on patterns of uterine development during defined neonatal periods, as well as the functional consequences of targeted neonatal steroid exposure in the adult uterus, should enable identification of critical developmental mechanisms and determinants of uterine integrity and function. Extreme adult uterine phenotypes (lesion models) created in cattle and sheep by strategic postnatal steroid exposure hold promise as powerful tools for the study of factors affecting uterine function and the rapid identification of novel uterine genes.


Cuticle integrity and biogenic amine synthesis in Caenorhabditis elegans require the cofactor tetrahydrobiopterin (BH4).
Tetrahydrobiopterin (BH4) is the natural cofactor of several enzymes widely distributed among eukaryotes, including aromatic amino acid hydroxylases (AAAHs), nitric oxide synthases (NOSs), and alkylglycerol monooxygenase (AGMO). We show here that the nematode Caenorhabditis elegans, which has three AAAH genes and one AGMO gene, contains BH4 and has genes that function in BH4 synthesis and regeneration. Knockout mutants for putative BH4 synthetic enzyme genes lack the predicted enzymatic activities, synthesize no BH4, and have indistinguishable behavioral and neurotransmitter phenotypes, including serotonin and dopamine deficiency. The BH4 regeneration enzymes are not required for steady-state levels of biogenic amines, but become rate limiting in conditions of reduced BH4 synthesis. BH4-deficient mutants also have a fragile cuticle and are generally hypersensitive to exogenous agents, a phenotype that is not due to AAAH deficiency, but rather to dysfunction in the lipid metabolic enzyme AGMO, which is expressed in the epidermis. Loss of AGMO or BH4 synthesis also specifically alters the sensitivity of C. elegans to bacterial pathogens, revealing a cuticular function for AGMO-dependent lipid metabolism in host-pathogen interactions.


Combination of Antiretroviral Drugs and Radioimmunotherapy Specifically Kills Infected Cells from HIV-Infected Individuals.
Eliminating virally infected cells is an essential component of any HIV eradication strategy. Radioimmunotherapy (RIT), a clinically established method for killing cells using radiolabeled antibodies, was recently applied to target HIV-1 gp41 antigen expressed on the surface of infected cells. Since gp41 expression by infected cells is likely downregulated in patients on antiretroviral therapy (ART), we evaluated the ability of RIT to kill ART-treated infected cells using both in vitro models and lymphocytes isolated from HIV-infected subjects. Human peripheral blood mononuclear cells (PBMCs) were infected with HIV and cultured in the presence of two clinically relevant ART combinations. Scatchard analysis of the 2556 human monoclonal antibody to HIV gp41 binding to the infected and ART-treated cells demonstrated sufficient residual expression of gp41 on the cell surface to warrant subsequent RIT. This is the first time the quantification of gp41 post-ART is being reported. Cells were then treated with Bismuth-213-labeled 2556 antibody. Cell survival was quantified by Trypan blue and residual viremia by p24 ELISA. Cell surface gp41 expression was assessed by Scatchard analysis. The experiments were repeated using PBMCs isolated from blood specimens obtained from 15 HIV-infected individuals: 10 on ART and 5 ART-naïve. We found that 213Bi-2556 killed ART-treated infected PBMCs and reduced viral production to undetectable levels. ART and RIT co-treatment was more effective at reducing viral load in vitro than either therapy alone, indicating that gp41 expression under ART was sufficient to allow 213Bi-2556 to deliver cytocidal doses of radiation to infected cells. This study provides proof of concept that 213Bi-2556 may represent an innovative and effective targeting method for killing HIV-infected cells treated with ART and supports continued development of 213Bi-2556 for co-administration with ART toward an HIV eradication strategy.


Nucleus tractus solitarii lesions alter the metabolic and hyperthermic response to central prostaglandin E1 in the rat.
1. Given that the nucleus tractus solitarii (NTS) may regulate the ability of brown adipose tissue to evoke non-shivering thermogenesis and that brown fat may mediate the rise in whole-body metabolism observed following central pyrogen administration, we assessed whether interruption of baroreceptor afferents coursing though the NTS would interfere with the ability of prostaglandin E1 to evoke a normal fever response profile. 2. Infusion of 150-600 ng of prostaglandin E1 (PGE1) into a lateral cerebral ventricle of the conscious rat resulted in a rise in core temperature, and also an increase in whole-body metabolic rate, brown adipose tissue temperature, arterial blood pressure and heart rate. 3. Following bilateral electrolytic lesions to the NTS, resting core and brown fat temperatures, metabolic rates, blood pressures and heart rates in the NTS-lesioned animals were comparable to control rats. However, the PGE1-evoked increase in metabolic rate, along with the rise in core and brown adipose tissue temperatures and heart rate were attenuated. The pressor response was, however, enhanced, possibly due to the demonstrated interference by the lesions with normal baroreflex control. 4. The findings suggest that the nucleus tractus solitarii region of the rats' brain may be important in mediating the thermogenesis evoked by central PGE1.


Time-resolved imaging of the MALDI linear-TOF ion cloud: direct visualization and exploitation of ion optical phenomena using a position- and time-sensitive detector.
In this study, we describe the implementation of a position- and time-sensitive detection system (Timepix detector) to directly visualize the spatial distributions of the matrix-assisted laser desorption ionization ion cloud in a linear-time-of-flight (MALDI linear-ToF) as it is projected onto the detector surface. These time-resolved images allow direct visualization of m/z-dependent ion focusing effects that occur within the ion source of the instrument. The influence of key parameters, namely extraction voltage (E(V)), pulsed-ion extraction (PIE) delay, and even the matrix-dependent initial ion velocity was investigated and were found to alter the focusing properties of the ion-optical system. Under certain conditions where the spatial focal plane coincides with the detector plane, so-called x-y space focusing could be observed (i.e., the focusing of the ion cloud to a small, well-defined spot on the detector). Such conditions allow for the stigmatic ion imaging of intact proteins for the first time on a commercial linear ToF-MS system. In combination with the ion-optical magnification of the system (~100×), a spatial resolving power of 11–16 μm with a pixel size of 550 nm was recorded within a laser spot diameter of ~125 μm. This study demonstrates both the diagnostic and analytical advantages offered by the Timepix detector in ToF-MS.


Lycopene Extracts from Different Tomato-Based Food Products Induce Apoptosis in Cultured Human Primary Prostate Cancer Cells and Regulate TP53, Bax and Bcl-2 Transcript Expression
Carotenoids are the main tomato components, especially lycopene. Lycopene is more bioavailable in tomato processed products than in raw tomatos, since formation of lycopene cis-isomers during food processing and storage may increase its biological activity. In the current study, we evaluated the influence of lycopene extracts (5 mg / mL) from different tomato-based food products (paste, sauce, extract and ketchup) on cell viability and apoptosis on primary human prostate cancer cells (PCa cels) for 96h. Using MTT assay, we observed a significant decrease on primary PCa cell viability upon treatment with lycopene extracted from either 4 tomato-based food products. Flow cytometeric analysis revealed that lycopene from tomato extract and tomato sauce promoted up to fifty-fold increase on the proportion of apoptotic cells, when compared to the control group. Using real time PCR assay, we found that lycopene promoted an upregulation of TP53 and Bax transcript expression and also downregulation of Bcl-2 expression in PCa cells. In conclusion, our data demostrate that cis-lycopene promoted a significant inhibition on primary PCa cell viability, as well as an increase on their apoptotic rates, evidencing that cis-lycopene contained in tomato sauce and extract cain mainly modulate of primary human prostate cancer cell survival.


General Practitioners involvement in enteral tube feeding at home: a qualitative study.
Complex medical treatment is moving from hospital to primary care and General Practitioners (GPs) are increasingly asked to undertake new roles. There are now an estimated 19,500 patients being fed in the UK in the community on enteral tube feeding using a variety of different feeding tubes (Percutaneous endoscopic gastrostomy (PEG), Jejunostomy, or nasogastric (NG). The majority of patients are over the age of 65 years when they had artificial feeding initiated and mainly because of dysphagia. The aim of this study was to explore GPs knowledge, attitudes and skills relating to enteral feeding in the community. Semi-structured one-to-one interviews with a convenience sample of GPs in Northern Ireland. Twenty-three GPs in three health boards in Northern Ireland participated in the study. Most found dealing with enteral feeding to be a predominantly negative experience. They had little involvement in patient selection for the procedure and poor or no discharge information. GPs felt inadequately trained, there was poor communication between primary and secondary care and little support. There was anger and frustration among GPs about lack of resources (funding and training), and the perception that primary care was used as a dumping ground. Moving complex medical treatment from secondary to primary care has major implications for GPs who should be included in the patient selection process, have adequate discharge information about their patients, be adequately resourced and have appropriate support and training.


Multiple P2X and P2Y receptor subtypes in mouse J774, spleen and peritoneal macrophages.
We investigated P2 receptor expression and function in macrophages from mouse, and in the J774 cell line, and revealed a larger spectrum of P2 receptor subtypes than previously recognised. The nucleotides adenosine triphosphate (ATP), adenosine diphosphate, uridine triphosphate and uridine diphosphate evoked an increase in intracellular calcium and the activation of a potassium current. The sensitivity of these responses to the antagonists suramin, PPADS, MRS 2179 and Cibacron blue suggest the presence of at least three functional P2Y receptor subtypes, most probably P2Y(2), P2Y(4) and P2Y(6). ATP also activated P2X receptors, giving rise to a rapidly activating cation conductance. This response was insensitive to the antagonists suramin and Cibacron blue, was potentiated by Zn(2+) and inhibited by acidification suggesting involvement of P2X(4) receptors. In low divalent cation solution, responses to ATP became larger, and dibenzoyl-ATP became more potent than ATP, indicating the presence of P2X(7) receptors. Immunofluorescence, flow cytometry, Western blots and RT-PCR show that P2X(4) and P2X(7) receptors are the most prominent in both macrophage types, while the expression of the other P2X subunits is variable and sometimes weak or undetectable. These techniques also demonstrated the presence of mRNA for P2Y(1), P2Y(2), P2Y(4) and P2Y(6) receptors along with protein expression for the three subtypes we investigated, namely, P2Y(1), P2Y(2) and P2Y(4).


GABAB-receptor-mediated inhibition of the N-methyl-D-aspartate component of synaptic transmission in the rat hippocampus.
GABA receptor regulation of NMDA-receptor-mediated synaptic responses was studied in area CA1 of the rat hippocampus using extracellular and intracellular recording techniques. Picrotoxin (PTX) was used to suppress GABAA inhibition and 6,7-dinitroquinoxaline-2,3-dione (DNQX) was used to suppress non-NMDA receptor-mediated responses. In this manner, we were able to avoid the complicating factors caused by potentials induced by other excitatory and inhibitory amino acid receptors. Under these conditions, large NMDA-receptor-mediated EPSPs were observed. When paired stimuli were given at interstimulus intervals from 100 to 400 msec, powerful inhibition of the second response was observed. This inhibition was reversed by the GABAB antagonists phaclofen and 2-hydroxy-saclofen; it was also depressed by removal of Mg2+ from the bath. Examination of non-NMDA receptor-mediated synaptic responses (determined in the presence of D-2-amino-5-phosphonovalerate and PTX) showed no such inhibition, thereby supporting the hypothesis that GABAB inhibition of NMDA EPSPs is postsynaptic. This difference in paired-pulse inhibition of NMDA and non-NMDA EPSPs leads us to conclude that there was no evidence of GABAB-mediated presynaptic inhibition of excitatory transmitter release. Intracellular recordings in the presence of DNQX and PTX revealed a phaclofen-sensitive late IPSP that correlated in time with the period of inhibition of NMDA responses. Taken together, these data suggest that paired-pulse-inhibition of NMDA responses is produced by a GABAB-receptor-mediated hyperpolarization of the postsynaptic membrane, causing an enhanced block of the NMDA channels by Mg2+. Regulation of NMDA-mediated synaptic responses by GABAB receptors constitutes a powerful mechanism for control of a major excitatory system in hippocampal pyramidal cells.


Multiple introductions from multiple sources: invasion patterns for an important Eucalyptus leaf pathogen.
Many population studies on invasive plant pathogens are undertaken without knowing the center of origin of the pathogen. Most leaf pathogens of Eucalyptus originate in Australia and consequently with indigenous populations available, and it is possible to study the pathways of invasion. Teratosphaeria suttonii is a commonly occurring leaf pathogen of Eucalyptus species, naturally distributed in tropical and subtropical regions of eastern Australia where it is regarded as a minor pathogen infecting older leaves; however, repeated infections, especially in exotic plantations, can result in severe defoliation and tree deaths. Nine polymorphic microsatellite markers were used to assess the genetic structure of 11 populations of T. suttonii of which four where from within its native range in eastern Australia and the remaining seven from exotic Eucalyptus plantations. Indigenous populations exhibited high allele and haplotype diversity, predominantly clonal reproduction, high population differentiation, and low gene flow. The diversity of the invasive populations varied widely, but in general, the younger the plantation industry in a country or region, the lower the diversity of T. suttonii. Historical gene flow was from Australia, and while self-recruitment was dominant in all populations, there was evidence for contemporary gene flow, with South Africa being the most common source and Uruguay the most common sink population. This points distinctly to human activities underlying long-distance spread of this pathogen, and it highlights lessons to be learned regarding quarantine.


TLR2 modulates inflammation in zymosan-induced arthritis in mice.
The interplay between the innate and acquired immune systems in chronic inflammation is not well documented. We have investigated the mechanisms of inflammation in murine zymosan-induced arthritis (ZIA) in the light of recent data on the roles of Toll-like receptor 2 (TLR2) and Dectin-1 in the activation of monocyte/macrophages by zymosan. The severity of inflammation, joint histology, lymphocyte proliferation and antibody production in response to zymosan were analyzed in mice deficient in TLR2 and complement C3, and the effects of Dectin-1 inhibition by laminarin were studied. In comparison with wild-type animals, TLR2-deficient mice showed a significant decrease in the early (day 1) and late phases (day 24) of joint inflammation. C3-deficient mice showed no differences in technetium uptake or histological scoring. TLR2-deficient mice also showed a significant decrease in lymph node cell proliferation in response to zymosan and a lower IgG antibody response to zymosan at day 25 in comparison with wild-type controls, indicating that TLR2 signalling has a role in the development of acquired immune responses to zymosan. Although laminarin, a soluble beta-glucan, was able to significantly inhibit zymosan uptake by macrophages in vitro, it had no effect on ZIA in vivo. These results show that ZIA is more prolonged than was originally described and involves both the innate and acquired immune pathways. C3 does not seem to have a major role in this model of joint inflammation.


Evaluation of serum sphingolipids and the influence of genetic risk factors in age-related macular degeneration.
Sphingolipids are bioactive molecules associated with oxidative stress, inflammation, and neurodegenerative diseases, but poorly studied in the context of age-related macular degeneration (AMD), a prevalent sight-threatening disease of the ageing retina. Here, we found higher serum levels of hexosylceramide (HexCer) d18:1/16:0 in patients with choroidal neovascularization (CNV) and geographic atrophy (GA), two manifestations of late stage AMD, and higher ceramide (Cer) d18:1/16:0 levels in GA patients. A sensitivity analysis of genetic variants known to be associated with late stage AMD showed that rs1061170 (p.Y402H) in the complement factor H (CFH) gene influences the association of Cer d18:1/16:0 with GA. To understand the possible influence of this genetic variant on ceramide levels, we established a cell-based assay to test the modulation of genes in the ceramide metabolism by factor H-like protein 1 (FHL-1), an alternative splicing variant of CFH that also harbors the 402 residue. We first showed that malondialdehyde-acetaldehyde adducts, an oxidation product commonly found in AMD retinas, induces an increase in ceramide levels in WERI-Rb1 cells in accordance with an increased expression of ceramide synthesis genes. Then, we observed that cells exposed to the non-risk FHL-1:Y402, but not the risk associated variant FHL-1:H402 or full-length CFH, downregulated ceramide synthase 2 and ceramide glucosyltransferase gene expression. Together, our findings show that serum ceramide and hexosylceramide species are altered in AMD patients and that ceramide levels may be influenced by AMD associated risk variants.


A consumer-resource approach to the density-dependent population dynamics of mutualism.
Like predation and competition, mutualism is now recognized as a consumer-resource (C-R) interaction, including, in particular, bi-directional (e.g., coral, plant-mycorrhizae) and uni-directional (e.g., ant-plant defense, plant-pollinator) C-R mutualisms. Here, we develop general theory for the density-dependent population dynamics of mutualism based on the C-R mechanism of interspecific interaction. To test the influence of C-R interactions on the dynamics and stability of bi- and uni-directional C-R mutualisms, we developed simple models that link consumer functional response of one mutualistic species with the resources supplied by another. Phase-plane analyses show that the ecological dynamics of C-R mutualisms are stable in general. Most transient behavior leads to an equilibrium of mutualistic coexistence, at which both species densities are greater than in the absence of interactions. However, due to the basic nature of C-R interactions, certain density-dependent conditions can lead to C-R dynamics characteristic of predator-prey interactions, in which one species overexploits and causes the other to go extinct. Consistent with empirical phenomena, these results suggest that the C-R interaction can provide a broad mechanism for understanding density-dependent population dynamics of mutualism. By unifying predation, competition, and mutualism under the common ecological framework of consumer-resource theory, we may also gain a better understanding of the universal features of interspecific interactions in general.


The mitochondrial genome of Phallusia mammillata and Phallusia fumigata (Tunicata, Ascidiacea): high genome plasticity at intra-genus level.
Within Chordata, the subphyla Vertebrata and Cephalochordata (lancelets) are characterized by a remarkable stability of the mitochondrial (mt) genome, with constancy of gene content and almost invariant gene order, whereas the limited mitochondrial data on the subphylum Tunicata suggest frequent and extensive gene rearrangements, observed also within ascidians of the same genus. To confirm this evolutionary trend and to better understand the evolutionary dynamics of the mitochondrial genome in Tunicata Ascidiacea, we have sequenced and characterized the complete mt genome of two congeneric ascidian species, Phallusia mammillata and Phallusia fumigata (Phlebobranchiata, Ascidiidae). The two mtDNAs are surprisingly rearranged, both with respect to one another and relative to those of other tunicates and chordates, with gene rearrangements affecting both protein-coding and tRNA genes. The new data highlight the extraordinary variability of ascidian mt genome in base composition, tRNA secondary structure, tRNA gene content, and non-coding regions (number, size, sequence and location). Indeed, both Phallusia genomes lack the trnD gene, show loss/acquisition of DHU-arm in two tRNAs, and have a G+C content two-fold higher than other ascidians. Moreover, the mt genome of P. fumigata presents two identical copies of trnI, an extra tRNA gene with uncertain amino acid specificity, and four almost identical sequence regions. In addition, a truncated cytochrome b, lacking a C-terminal tail that commonly protrudes into the mt matrix, has been identified as a new mt feature probably shared by all tunicates. The frequent occurrence of major gene order rearrangements in ascidians both at high taxonomic level and within the same genus makes this taxon an excellent model to study the mechanisms of gene rearrangement, and renders the mt genome an invaluable phylogenetic marker to investigate molecular biodiversity and speciation events in this largely unexplored group of basal chordates.


Robust excitation and Raman conversion of guided vortices in a chiral gas-filled photonic crystal fiber.
The unique ring-shaped intensity patterns and helical phase fronts of optical vortices make them useful in many applications. Here we report for the first time, to the best of our knowledge, efficient Raman frequency conversion between vortex modes in a twisted hydrogen-filled single-ring hollow core photonic crystal fiber (SR-PCF). High-fidelity transmission of optical vortices in an untwisted SR-PCF becomes more and more difficult as the orbital angular momentum (OAM) order increases, due to scattering at structural imperfections in the fiber microstructure. In a helically twisted SR-PCF, however, the degeneracy between left- and right-handed versions of the same mode is lifted, with the result that they are topologically protected from such scattering. With launch efficiencies of ${\sim}{75}\% $∼75%, a high damage threshold and broadband guidance, these fibers are ideal for performing nonlinear experiments that require the polarization state and azimuthal order of the interacting modes to be preserved over long distances. Vortex coherence waves of internal molecular motion carrying angular momentum are excited in the gas, permitting the polarization and OAM of the Raman bands to be tailored, even in spectral regions where conventional solid-core waveguides are opaque or susceptible to optical damage.


Carbohydrate-derived amino-alcohol ligands for asymmetric alkynylation of aldehydes.
[reaction: see text] Conformationally restricted amino alcohols based on carbohydrate scaffolds provide flexible and fine-tuneable libraries that greatly expand the range of ligands available in the Zn(OTf)(2)-mediated addition of alkynes to aldehydes, in some cases with very high stereoselectivities.


In-stream bioreactor for agricultural nitrate treatment.
Nitrate from agricultural activity contributes to nutrient loading in surface water bodies such as the Mississippi River. This study demonstrates a novel in-stream bioreactor that uses carbonaceous solids (woodchips) to promote denitrification of agricultural drainage. The reactor (40 m3) was trenched into the bottom of an existing agricultural drainage ditch in southern Ontario (Avon site), and flow was induced through the reactor by construction of a gravel riffle in the streambed. Over the first 1.5 yr of operation, mean influent NO3-N of 4.8 mg L(-1) was attenuated to 1.04 mg L(-1) at a mean reactor flow rate of 24 L min(-1). A series of flow-step tests, facilitated by an adjustable height outlet pipe, demonstrated that nitrate mass removal generally increased with increasing flow rate. When removal rates were not nitrate-limited, areal mass removal ranged from 11 mg N m(-2) h(-1) at 3 degrees C to 220 mg N m(-2) h(-1) at 14 degrees C (n = 27), exceeding rates reported for some surface-flow constructed wetlands in this climatic region by a factor of about 40. Over the course of the field trial, reactor flow rates decreased as a result of silt accumulation on top of the gravel infiltration gallery. Design modifications are currently being implemented to mitigate the effects of siltation. In-stream reactors have the potential to be scaled larger and could be more manageable than attempting to address nitrate loading from individual tile drains. They could also work well in combination with other nitrate control techniques.


Hydrocephalus shunt technology: 20 years of experience from the Cambridge Shunt Evaluation Laboratory.
The Cambridge Shunt Evaluation Laboratory was established 20 years ago. This paper summarizes the findings of that laboratory for the clinician. Twenty-six models of valves have been tested long-term in the shunt laboratory according to the expanded International Organization for Standardization 7197 standard protocol. The majority of the valves had a nonphysiologically low hydrodynamic resistance (from 1.5 to 3 mm Hg/[ml/min]), which may result in overdrainage related to posture and during nocturnal cerebral vasogenic waves. A long distal catheter increases the resistance of these valves by 100%-200%. Drainage through valves without a siphon-preventing mechanism is very sensitive to body posture, which may result in grossly negative intracranial pressure. Siphon-preventing accessories offer a reasonable resistance to negative outlet pressure; however, accessories with membrane devices may be blocked by raised subcutaneous pressure. In adjustable valves, the settings may be changed by external magnetic fields of intensity above 40 mT (exceptions: ProGAV, Polaris, and Certas). Most of the magnetically adjustable valves produce large distortions on MRI studies. The behavior of a valve revealed during testing is of relevance to the surgeon and may not be adequately described in the manufacturer's product information. The results of shunt testing are helpful in many circumstances, such as the initial choice of shunt and the evaluation of the shunt when its dysfunction is suspected.


InlB: an invasion protein of Listeria monocytogenes with a novel type of surface association.
Listeria monocytogenes is an intracellular bacterial pathogen that expresses several surface proteins critical for the infectious process. Such proteins include InlA (internalin) and InlB, involved in bacterial entry into the host cell, and ActA, required for bacterially induced actin-based motility. Although the molecular mechanisms of attachment of InlA and ActA have been characterized, essentially nothing is known about how InlB is anchored to the bacterial surface. Using a genetic approach, we demonstrate that the last 232 amino acids of InlB are both necessary and sufficient for anchoring this protein to the bacterial surface. An InlB mutant protein deleted for the last 232 amino acids was secreted and not detected at the cell surface. A 'domain-swapping' strategy in which these 232 amino acids were used to replace the normal cell wall-anchoring domain of InlA resulted in a chimeric protein that was anchored to the cell surface and able to confer entry. Interestingly, surface association of InlB also occurred when InlB was added externally to bacteria, suggesting that association may be able to occur after secretion. This association was productive for invasion, as it conferred bacterial entry into host cells. The C-terminal anchoring region in InlB contains 80-amino-acid repeats beginning with the sequence GW that is also present in a newly identified surface-associated bacteriolysin of L. monocytogenes, called Ami. Addition of GW repeats to the C-terminal of InlB improves anchoring of the protein to the cell surface. These and other data suggest that such 'GW' repeats may constitute a novel motif for cell-surface anchoring in Listeria and other Gram-positive bacteria. This motif may have important consequences for the release of surface proteins involved in interactions with eukaryotic cells.


Detection of CDKN2 deletions in tumor cell lines and primary glioma by interphase fluorescence in situ hybridization.
Deletions of chromosomal band 9p21 have been detected in various tumor types including melanoma, glioma, lung cancer, mesothelioma, and bladder cancer. Recently, the CDKN2 gene (p16INK4A, MTS I, CDK41) has been proposed as a candidate tumor suppressor gene because it is frequently deleted in cell lines derived from multiple tumor types. We performed fluorescence in situ hybridization (FISH) with interphase cells using yeast artificial chromosome clones and a cosmid contig of the CDKN2 region. In 10 cell lines (4 glioma, 2 melanoma, 2 non-small cell lung cancer, 2 bladder cancer) with 9p alterations detected by molecular or cytogenetic analysis, interphase FISH with the CDKN2 cosmid contig detected all 9p deletions previously identified by molecular analysis. Using this probe, FISH analysis of primary glioblastoma tumors revealed homozygous deletions of the CDKN2 region in 6 of 9 tumors (67%) whereas a yeast artificial chromosome probe containing the interferon type I (IFN) gene cluster was deleted in only 4 cases (44%). Thus, it is likely that the CDKN2 region is the target of 9p deletions in gliomas. Interphase FISH will play an important role in defining the clinical significance of 9p deletions in primary tumors because it is especially applicable to clinical samples which may be contaminated by normal cells.


A case-control study of determinants for high and low dental caries prevalence in Nevada youth.
The main purpose of this study was to compare the 30% of Nevada Youth who presented with the highest Decayed Missing and Filled Teeth (DMFT) index to a cohort who were caries free and to national NHANES data. Secondly, to explore the factors associated with higher caries prevalence in those with the highest DMFT scores compared to the caries-free group. Over 4000 adolescents between ages 12 and 19 (Case Group: N = 2124; N = 2045) received oral health screenings conducted in public/private middle and high schools in Nevada in 2008/2009 academic year. Caries prevalence was computed (Untreated decay scores [D-Score] and DMFT scores) for the 30% of Nevada Youth who presented with the highest DMFT score (case group) and compared to the control group (caries-free) and to national averages. Bivariate and multivariate logistic regression was used to analyze the relationship between selected variables and caries prevalence. A majority of the sample was non-Hispanic (62%), non-smokers (80%), and had dental insurance (70%). With the exception of gender, significant differences in mean D-scores were found in seven of the eight variables. All variables produced significant differences between the case and control groups in mean DMFT Scores. With the exception of smoking status, there were significant differences in seven of the eight variables in the bivariate logistic regression. All of the independent variables remained in the multivariate logistic regression model contributing significantly to over 40% of the variation in the increased DMFT status. The strongest predictors for the high DMFT status were racial background, age, fluoridated community, and applied sealants respectively. Gender, second hand smoke, insurance status, and tobacco use were significant, but to a lesser extent. Findings from this study will aid in creating educational programs and other primary and secondary interventions to help promote oral health for Nevada youth, especially focusing on the subgroup that presents with the highest mean DMFT scores.


Medicinal smoke reduces airborne bacteria.
This study represents a comprehensive analysis and scientific validation of our ancient knowledge about the effect of ethnopharmacological aspects of natural products' smoke for therapy and health care on airborne bacterial composition and dynamics, using the Biolog microplate panels and Microlog database. We have observed that 1h treatment of medicinal smoke emanated by burning wood and a mixture of odoriferous and medicinal herbs (havan sámagri=material used in oblation to fire all over India), on aerial bacterial population caused over 94% reduction of bacterial counts by 60 min and the ability of the smoke to purify or disinfect the air and to make the environment cleaner was maintained up to 24h in the closed room. Absence of pathogenic bacteria Corynebacterium urealyticum, Curtobacterium flaccumfaciens, Enterobacter aerogenes (Klebsiella mobilis), Kocuria rosea, Pseudomonas syringae pv. persicae, Staphylococcus lentus, and Xanthomonas campestris pv. tardicrescens in the open room even after 30 days is indicative of the bactericidal potential of the medicinal smoke treatment. We have demonstrated that using medicinal smoke it is possible to completely eliminate diverse plant and human pathogenic bacteria of the air within confined space.


Optically detected cross-relaxation spectroscopy of electron spins in diamond.
The application of magnetic resonance spectroscopy at progressively smaller length scales may eventually permit 'chemical imaging' of spins at the surfaces of materials and biological complexes. In particular, the negatively charged nitrogen-vacancy (NV(-)) centre in diamond has been exploited as an optical transducer for nanoscale nuclear magnetic resonance. However, the spectra of detected spins are generally broadened by their interaction with proximate paramagnetic NV(-) centres through coherent and incoherent mechanisms. Here we demonstrate a detection technique that can resolve the spectra of electron spins coupled to NV(-) centres, in this case, substitutional nitrogen and neutral nitrogen-vacancy centres in diamond, through optically detected cross-relaxation. The hyperfine spectra of these spins are a unique chemical identifier, suggesting the possibility, in combination with recent results in diamonds harbouring shallow NV(-) implants, that the spectra of spins external to the diamond can be similarly detected.


Enhanced recovery of Salmonella Typhimurium DT104 from exposure to stress at low temperature.
Salmonella enterica serovar Typhimurium (S. Typhimurium) remains an important cause of food-borne infection in the developed world. In order to establish infection within a host, Salmonella must survive and recover from a range of environmental stresses. S. Typhimurium strain SL1344 is among the most extensively studied pathogenic Salmonella strains, while S. Typhimurium phage type DT104 is an important type that has been associated with pandemic spread and a high number of food-borne disease outbreaks over the last two decades. In this study, we have compared the abilities of these two S. Typhimurium types to recover from stress exposures commonly encountered in food production, including 685 mM NaCl, pH 3.8, low temperature (6 °C) and combinations thereof. Following removal from prolonged (8 days) stress, DT104 cultures that had been exposed to low temperature, with or without additional stress, resumed exponential growth more rapidly than SL1344 cultures exposed to the same conditions. SL1344 showed higher levels of filamentation than DT104 in response to NaCl exposure at low temperature. Further, SL1344 incurred higher levels of membrane damage in response to elevated NaCl and pH 3.8 at both temperatures compared with DT104. However, both strains recovered normal cell division and membrane integrity within 6 h when all stresses were removed. Expression of the Salmonella pathogenicity island 1 gene prgH, the first gene in the prg/org operon, was monitored using a chromosomal reporter in which gfp(+) expression was driven by the prgH promoter. Recovery of prgH expression was comparable for SL1344 and DT104 exposed to stress at 22 °C. However, DT104 cultures exposed to pH 3.8 or combined NaCl and low-pH stress at low temperature resumed prgH expression more rapidly than SL1344. Both strains recovered maximal levels of prgH expression after 6 h recovery from all stresses and, interestingly, maximal levels of prgH expression were significantly higher in SL1344, consistent with prgH expression in late-exponential, non-stressed SL1344 and DT104 cultures. Together, these data show that S. Typhimurium is capable of rapid recovery from environmental and food-related stresses, and give insight into the enhanced ability of DT104 compared with SL1344 to adapt to such stresses, which may contribute to the success of this globally disseminated pathogenic phage type.


Systematically searching for and assessing the literature for self-management of chronic pain: a lay users' perspective.
The Engaging with older adults in the development of strategies for the self management of chronic pain (EOPIC) study aims to design and develop self management strategies to enable older adults to manage their own pain. Involving older adults in research into chronic pain management will better enable the identification and development of strategies that are more appropriate for their use, but how can perspectives really be utilised to the best possible outcomes? Seven older adults were recruited through a local advertising campaign to take part. We also invited participants from the local pain services, individuals who had been involved in earlier phase of the EOPIC study and a previous ESRC funded project. The group undertook library training and research skills training to facilitate searching of the literature and identified sources of material. A grading tool was developed using perceived essential criteria identified by the older adults and material was graded according to the criteria within this scale. Fifty-seven resources from over twenty-eight sources were identified. These materials were identified as being easily accessible, readable and relevant. Many of the web based materials were not always easy to find or readily available so they were excluded by the participants. All but one were UK based. Forty-four items were identified as meeting the key criteria for inclusion in the study. This included five key categories as follows; books, internet, magazines, leaflets, CD's/Tapes. This project was able to identify a number of exemplars of self management material along with some general rules regarding the categories identified. We must point out that the materials identified were not age specific, were often locally developed and would need to be adapted to older adults with chronic pain. For copyright issues we have not included them in this paper. The key message is really related to the format rather than the content. However, the group acknowledge that these may vary according to the requirements of each individual older adult and therefore recommend the development of a leaflet to help others in their search for resources. This leaflet has been developed as part of Phase IV of the EOPIC study.


Osteoclasts express the B2 isoform of vacuolar H(+)-ATPase intracellularly and on their plasma membranes.
Osteoclasts express high levels of vacuolar H(+)-ATPase (V-ATPase) in their ruffled membranes, driving the secretion of H+ required for normal bone resorption. Previous reports have suggested that the B subunit of the osteoclast V-ATPase differs from those expressed in kidney and other tissues. In this study, B subunit isoform-specific antibodies and cDNA probes were used to examine which B subunit isoform is expressed in osteoclasts and osteoclast-like cells. Immunoblotting and RNA hybridization analysis were used to demonstrate that cells from an osteoclast-rich mouse bone marrow culture model express the B2 but not the B1 subunit isoform. Immunocytochemical staining of murine osteoclasts generated in vitro and of native rat osteoclasts in bone sections showed that the B2 but not the B1 isoform was expressed at high levels and was polarized to the ruffled membrane. Human marrow cultures and monocyte-derived macrophages, used as models for osteoclasts, also expressed the B2 but not the B1 subunit isoform. These results indicate that V-ATPases containing the B2 subunit isoform mediate osteoclast bone resorption.


Long-Term Temperature Stress in the Coral Model Aiptasia Supports the "Anna Karenina Principle" for Bacterial Microbiomes.
The understanding of host-microbial partnerships has become a hot topic during the last decade as it has been shown that associated microbiota play critical roles in the host physiological functions and susceptibility to diseases. Moreover, the microbiome may contribute to host resilience to environmental stressors. The sea anemone Aiptasia is a good laboratory model system to study corals and their microbial symbiosis. In this regard, studying its bacterial microbiota provides a better understanding of cnidarian metaorganisms as a whole. Here, we investigated the bacterial communities of different Aiptasia host-symbiont combinations under long-term heat stress in laboratory conditions. Following a 16S rRNA gene sequencing approach we were able to detect significant differences in the bacterial composition and structure of Aiptasia reared at different temperatures. A higher number of taxa (i.e., species richness), and consequently increased α-diversity and β-dispersion, were observed in the microbiomes of heat-stressed individuals across all host strains and experimental batches. Our findings are in line with the recently proposed Anna Karenina principle (AKP) for animal microbiomes, which states that dysbiotic or stressed organisms have a more variable and unstable microbiome than healthy ones. Microbial interactions affect the fitness and survival of their hosts, thus exploring the AKP effect on animal microbiomes is important to understand host resilience. Our data contributes to the current knowledge of the Aiptasia holobiont and to the growing field of study of host-associated microbiomes.


E. coli Nissle 1917 Affects Salmonella adhesion to porcine intestinal epithelial cells.
The probiotic Escherichia coli strain Nissle 1917 (EcN) has been shown to interfere in a human in vitro model with the invasion of several bacterial pathogens into epithelial cells, but the underlying molecular mechanisms are not known. In this study, we investigated the inhibitory effects of EcN on Salmonella Typhimurium invasion of porcine intestinal epithelial cells, focusing on EcN effects on the various stages of Salmonella infection including intracellular and extracellular Salmonella growth rates, virulence gene regulation, and adhesion. We show that EcN affects the initial Salmonella invasion steps by modulating Salmonella virulence gene regulation and Salmonella SiiE-mediated adhesion, but not extra- and intracellular Salmonella growth. However, the inhibitory activity of EcN against Salmonella invasion always correlated with EcN adhesion capacities. EcN mutants defective in the expression of F1C fimbriae and flagellae were less adherent and less inhibitory toward Salmonella invasion. Another E. coli strain expressing F1C fimbriae was also adherent to IPEC-J2 cells, and was similarly inhibitory against Salmonella invasion like EcN. We propose that EcN affects Salmonella adhesion through secretory components. This mechanism appears to be common to many E. coli strains, with strong adherence being a prerequisite for an effective reduction of SiiE-mediated Salmonella adhesion.


Bar homeobox genes are latitudinal prepattern genes in the developing Drosophila notum whose expression is regulated by the concerted functions of decapentaplegic and wingless.
In Drosophila notum, the expression of achaete-scute proneural genes and bristle formation have been shown to be regulated by putative prepattern genes expressed longitudinally. Here, we show that two homeobox genes at the Bar locus (BarH1 and BarH2) may belong to a different class of prepattern genes expressed latitudinally, and suggest that the developing notum consists of checker-square-like subdomains, each governed by a different combination of prepattern genes. BarH1 and BarH2 are coexpressed in the anterior-most notal region and regulate the formation of microchaetae within the region of BarH1/BarH2 expression through activating achaete-scute. Presutural macrochaetae formation also requires Bar homeobox gene activity. Bar homeobox gene expression is restricted dorsally and posteriorly by Decapentaplegic signaling, while the ventral limit of the expression domain of Bar homeobox genes is determined by wingless whose expression is under the control of Decapentaplegic signaling.


Isolation precautions for antibiotic-resistant bacteria in healthcare settings.
Emergence of drug-resistant bacteria and new or changing infectious pathogens is an important public health problem. Transmission of these pathogens in an acute care setting may occur frequently if proper precautions are not taken. Despite several guidelines and an abundance of literature on the prevention of transmission of epidemiologically important organisms in the healthcare setting, substantial controversy exists. This review focuses on recent data regarding the use of infection control and isolation precautions. New data are available, but the conflict surrounding the use of active surveillance of methicillin-resistant Staphylococcus aureus (MRSA) has not been resolved. The emergence of multidrug-resistant Gram-negative bacteria has prompted a greater interest in infection control strategies for prevention of their spread. Outbreaks of Clostridium difficile have responded to broad infection control initiatives, but further research is required to determine whether the best infection control precautions are being utilized. Effective prevention of the transmission of pathogens within the healthcare system requires a multifaceted approach. Existing guidelines should be used to create institutional policies specific to individual patient populations, problem pathogens and the ability to practically implement various infection control procedures. Despite ongoing study, the use of active surveillance to prevent transmission of MRSA continues to be a complex, controversial and challenging issue.


The pharmacokinetics of flunixin meglumine in the sheep.
Flunixin meglumine was administered intravenously and intramuscularly in sheep and the pharmacokinetics of the drug studied. Plasma concentrations of flunixin were measured by high performance liquid chromatography. The decline in plasma flunixin concentration with time was best fitted by a triexponential equation. The pharmacokinetics following intravenous administration of 1.0 mg/kg indicate that flunixin has a rapid distribution half-life (t1/2 pi = 2.3 min), a slow body clearance rate (Clb = 0.6 ml/kg/min) and an elimination half-life of 229 min. Similarly, at 2.0 mg/kg, flunixin is rapidly distributed from the plasma, t1/2 pi = 2.7 min, has a slow body clearance rate (Clb = 0.7 ml/kg/min) and an elimination half-life of 205 min. Following intramuscular injection flunixin is rapidly and well absorbed from the injection site. It had a mean maximum concentration (Cmax) of > or = 5.9 micrograms/ml when administered at a dose rate of 1.1 mg/kg, and a relative bioavailability of 70%. Plasma concentrations increase proportionally to dose over the range 1.1 mg/kg-2.2 mg/kg when administered by the intramuscular route.


Structural Basis of Diverse Homophilic Recognition by Clustered α- and β-Protocadherins.
Clustered protocadherin proteins (α-, β-, and γ-Pcdhs) provide a high level of cell-surface diversity to individual vertebrate neurons, engaging in highly specific homophilic interactions to mediate important roles in mammalian neural circuit development. How Pcdhs bind homophilically through their extracellular cadherin (EC) domains among dozens of highly similar isoforms has not been determined. Here, we report crystal structures for extracellular regions from four mouse Pcdh isoforms (α4, α7, β6, and β8), revealing a canonical head-to-tail interaction mode for homophilic trans dimers comprising primary intermolecular EC1:EC4 and EC2:EC3 interactions. A subset of trans interface residues exhibit isoform-specific conservation, suggesting roles in recognition specificity. Mutation of these residues, along with trans-interacting partner residues, altered the specificities of Pcdh interactions. Together, these data show how sequence variation among Pcdh isoforms encodes their diverse strict homophilic recognition specificities, which are required for their key roles in neural circuit assembly.


Association between ambient carbon monoxide levels and hospitalizations for congestive heart failure in the elderly in 10 Canadian cities.
We examined the role that ambient air pollution plays in exacerbating cardiac disease by relating daily fluctuations in admissions to 134 hospitals for congestive heart failure in the elderly to daily variations in ambient concentrations of carbon monoxide, nitrogen dioxide, sulfur dioxide, ozone, and the coefficient of haze in Canada's 10 largest cities for the 11-year period 1981-1991 inclusive. We adjusted the hospitalization time series for seasonal, subseasonal, and weekly cycles and for hospital usage patterns. The logarithm of the daily high-hour ambient carbon monoxide concentration recorded on the day of admission displayed the strongest and most consistent association with hospitalization rates among the pollutants, after stratifying the time series by month of year and adjusting simultaneously for temperature, dew point, and the other ambient air pollutants. The relative risk for a change from 1 ppm to 3 ppm, the 25th and 75th percentiles of the exposure distribution, was 1.065 (95% confidence interval = 1.028-1.104). The regression coefficients of the other air pollutants were much more sensitive to simultaneous adjustment for either multiple pollutant or weather model specifications.


A stable form of delayed-type hypersensitivity.
An antigen dose below the level needed to provoke an antibody response produces in mice a persistent, but minor degree of delayed-type hypersensitivity (dth) to sheep red blood cells. The DTH is unstable. It is erased by larger doses of antigen and cannot be built upon by further antigenic stimulation. The much higher levels of DTH resulting from immunization under the modulating influence of cyclophosphamide (CY) or BCG persist under strong secondary antigenic stimulation, though the former is subject to partial suppression unless CY is used to prevent the secondary humoral response. The DTH produced by a BCG-modulated primary response is not subject to this suppressive effect of a secondary antibody response. In this case the anamnestic T-cell response is very brisk and cannont be potentiated by giving CY at the time of the secondary antigenic stimulus. This effect is not due to the modulating influence of a residual BCG infection. It results from a permanent change induced during the primary response. The mediator cells formed under the influence of BCG are apparently resistant to inhibition by blocking serum containing immune complexes. Even the actively dividing T cells which are susceptible to vinblastine, and most readily blocked in the absence of BCG, are highly resistant to blocking by immune complexes. It is not clear whether these cells are intrinsically different or whether their insensitivity to blocking results from features peculiar to the humoral response that accompanies a BCG-modulated primary response. The mediator cells produced by both BCG- and CY-modulated responses become vinblastine resistant, relatively insensitive to humoral blocking factors, and capable of surviving in a functionally active form in syngeneic recipients with an apparent half-life of about 50 days. There were indications, however, that their effective life-span may be greatly extended in some circumstances by persisting antigenic stimulation; and in the case of BCG-modulated immunity the prevailing level of T-cell activity can be greatly augmented by a further antigenic stimulus without the necessity for renewed exposure to BCG.


NFκB signaling in alveolar rhabdomyosarcoma.
Alveolar rhabdomyosarcoma (aRMS) is a pediatric soft tissue cancer commonly associated with a chromosomal translocation that leads to the expression of a Pax3:Foxo1 or Pax7:Foxo1 fusion protein, the developmental underpinnings of which may give clues to its therapeutic approaches. In aRMS, the NFκB-YY1-miR-29 regulatory circuit is dysregulated, resulting in repression of miR-29 and loss of the associated tumor suppressor activity. To further elucidate the role of NFκB in aRMS, we first tested 55 unique sarcoma cell lines and primary cell cultures in a large-scale chemical screen targeting diverse molecular pathways. We found that pharmacological inhibition of NFκB activity resulted in decreased cell proliferation of many of the aRMS tumor cultures. Surprisingly, mice that were orthotopically allografted with aRMS tumor cells exhibited no difference in tumor growth when administered an NFκB inhibitor, compared to control. Furthermore, inhibition of NFκB by genetically ablating its activating kinase inhibitor, IKKβ, by conditional deletion in a mouse model harboring the Pax3:Foxo1 chimeric oncogene failed to abrogate spontaneous tumor growth. Genetically engineered mice with conditionally deleted IKKβ exhibited a paradoxical decrease in tumor latency compared with those with active NFκB. However, using a synthetic-lethal approach, primary cell cultures derived from tumors with inactivated NFκB showed sensitivity to the BCL-2 inhibitor navitoclax. When used in combination with an NFκB inhibitor, navitoclax was synergistic in decreasing the growth of both human and IKKβ wild-type mouse aRMS cells, indicating that inactivation of NFκB alone may not be sufficient for reducing tumor growth, but, when combined with another targeted therapeutic, may be clinically beneficial.


Job satisfaction of occupational physicians in commercial and other delivery settings: a comparative and explorative study.
Commercialization of occupational health services (OHS) and transition to a supplier market which started in 1995, has affected work and working conditions of occupational physicians (OPs) profoundly. OPs have lost influence on the organization of their work and managers took over control. This study explores the impact of commercialization on job satisfaction of occupational physicians. Data were collected through a questionnaire completed by 797 OPs, members of the Netherlands Association of Occupational Medicine (response: 45%). A range of work satisfiers and dissatisfiers measures was included that, according to the Social Action Approach, could explain the variation in job satisfaction. Stepwise multivariate regression analyses were performed to explore to what extent the differences in job satisfaction can be attributed to the characteristics of the setting, biographical variables, satisfiers and dissatisfiers. Occupational physicians in commercial settings had the lowest average score as regards job satisfaction (6.7) contrasting sharply with their colleagues in private practices (8.7) and in non-commercial settings (7.9). The variation in job satisfaction between delivery settings could largely be attributed to satisfiers and dissatisfiers. Biographical characteristics (age, gender) had no effect. The data suggested that not commercialization as such, but the ability of commercial OHS providers to integrate professional values was the crucial factor to bring about job satisfaction. The challenge for commercial OHS providers is to preserve the professional zeal in OPs by integrating professional values in their organization in order to improve the quality of the services and the attractiveness of the profession.


An individually tailored family-centered intervention for pediatric obesity in primary care: study protocol of a randomized type II hybrid effectiveness-implementation trial (Raising Healthy Children study).
Pediatric obesity is a multi-faceted public health concern that can lead to cardiovascular diseases, cancers, and early mortality. Small changes in diet, physical activity, or BMI can significantly reduce the possibility of developing cardiometabolic risk factors. Family-based behavioral interventions are an underutilized, evidence-based approach that have been found to significantly prevent excess weight gain and obesity in children and adolescents. Poor program availability, low participation rates, and non-adherence are noted barriers to positive outcomes. Effective interventions for pediatric obesity in primary care are hampered by low family functioning, motivation, and adherence to recommendations. This (type II) hybrid effectiveness-implementation randomized trial tests the Family Check-Up 4 Health (FCU4Health) program, which was designed to target health behavior change in children by improving family management practices and parenting skills, with the goal of preventing obesity and excess weight gain. The FCU4Health is assessment driven to tailor services and increase parent motivation. A sample of 350 families with children aged 6 to 12 years who are identified as overweight or obese (BMI ≥ 85th percentile for age and gender) will be enrolled at three primary care clinics [two Federally Qualified Healthcare Centers (FQHCs) and a children's hospital]. All clinics serve predominantly Medicaid patients and a large ethnic minority population, including Latinos, African Americans, and American Indians who face disparities in obesity, cardiometabolic risk, and access to care. The FCU4Health will be coordinated with usual care, using two different delivery strategies: an embedded approach for the two FQHCs and a referral model for the hospital-based clinic. To assess program effectiveness (BMI, body composition, child health behaviors, parenting, and utilization of support services) and implementation outcomes (such outcomes as acceptability, adoption, feasibility, appropriateness, fidelity, and cost), we use a multi-method and multi-informant assessment strategy including electronic health record data, behavioral observation, questionnaires, interviews, and cost capture methods. This study has the potential to prevent excess weight gain, obesity, and health disparities in children by establishing the effectiveness of the FCU4Health and collecting information critical for healthcare decision makers to support sustainable implementation of family-based programs in primary care. NCT03013309 ClinicalTrials.gov.


Cloning and characterization of the gene encoding human NPL4, a protein interacting with the ubiquitin fusion-degradation protein (UFD1L).
The ubiquitin fusion-degradation gene (UFD1L) encodes the human homologue of the yeast ubiquitin fusion-degradation 1 protein, an essential component of the ubiquitin-dependent proteolytic turnover and mRNA processing. Although the UFD1L gene has been mapped in the region commonly deleted in patients with DiGeorge syndrome (DGS)/velocardiofacial syndrome (VCFS), correlation between its haploinsufficiency and the phenotype has not yet been established. The only functional data available about mammalian Ufd1p is the ability to form a complex with the rat Npl4 protein, a component of the nuclear pore complex. In this paper we report the cloning and molecular characterization of the human NPL4 gene. This gene encodes for a protein 96% homologous to the rat Npl4, and 44 and 34% homologous to the C. elegans and S. cerevisiae Npl4 gene products, respectively. Fluorescence in situ hybridization experiments on human metaphases localized the NPL4 gene on the most telomeric region of chromosome 17q. Northern blots analysis on foetal and adult human tissues revealed a major approximately 4.5 kb transcript most abundant in heart, brain, kidney and skeletal muscle. In order to test a potential relationship between nuclear transport defects and some aspect of the DGS/VCFS phenotype, we also exclude the presence of mutations in the NPL4 coding sequence in a subset of patients with DGS/VCFS and no detectable 22q11 deletion or mutations at the UFD1L locus.


End-of-life care in nursing homes: the high cost of staff turnover.
Nursing home staff turnover results in high cost--both economic and personal--and has a negative impact on the quality of care provided to residents at the end of life. Reducing staff turnover in nursing homes would benefit both the cost to the U.S. health care system, and, most importantly, the care residents receive in the vulnerable period leading to death. There is rising pressure on nursing homes to improve their palliative and end-of-life care practices and reduce transfers to hospital for situations and conditions that can be safely managed on site. Nursing care staff deserve an investment in the specific training necessary for them to give the highest quality care to dying residents. This training should be multifaceted and include the physiological, psychological, spiritual, interpersonal, and cultural (including ethnic) aspects of dying. Empowerment with these necessary knowledge, skills, and attitudes will not only result in better care for residents but likely also will reduce the burnout and frustration staff experience in caring for residents near death.


Methods to determine slow diffusion coefficients of biomolecules: applications to Engrailed 2, a partially disordered protein.
We present new NMR methods to measure slow translational diffusion coefficients of biomolecules. Like the heteronuclear stimulated echo experiment (XSTE), these new methods rely on the storage of information about spatial localization during the diffusion delay as longitudinal polarization of nuclei with long T(1) such as nitrogen-15. The new BEST-XSTE sequence combines features of Band-selective Excitation Short-Transient (BEST) and XSTE methods. By avoiding the saturation of all protons except those of amide groups, one can increase the sensitivity by 45% in small proteins. The new experiment which combines band-Selective Optimized Flip-Angle Short-Transient with XSTE (SOFAST-XSTE) offers an alternative when very short recovery delays are desired. A modification of the HSQC-edited version of the XSTE experiment offers enhanced sensitivity and access to higher resolution in the indirect dimension. These new methods have been applied to detect changes in diffusion coefficients due to dimerization or proteolysis of Engrailed 2, a partially disordered protein.


Comparison of birefringent electric split-ring resonator and meanderline structures as quarter-wave plates at terahertz frequencies.
We have fabricated a quarter-wave plate from a single layer of birefringent electric split-ring resonators (ELC). For comparison, an appropriately scaled double layer meanderline structure was fabricated. At the design frequency of 639 GHz, the ELC structure achieves 99.9% circular polarization while the meanderline achieves 99.6%. The me-anderline displays a larger bandwidth of operation, attaining over 99% circular polarization from 615 - 743 GHz, while the ELC achieves 99% from 626 - 660 GHz. However, both are broad enough for use with CW sources making ELCs a more attractive choice due to the ease of fabrication. Both samples are free standing with a total thickness of 70 microm for the meanderline structure and a mere 20 microm for the ELC highlighting the large degree of birefringence exhibited with metamaterial structures.


Validation of Astropecten jarli Madsen, 1950 and implications for A. cingulatus Sladen, 1883 (Paxillosida: Astropectinidae).
Astropecten is a major paxillosid genus with a widely occurring presence in tropical and temperate seas. Astropecten jarli from off the west coast of Africa, in the Eastern Atlantic Region, was placed in synonymy with Astropecten cingulatus, a widespread species from the Western Atlantic Region, producing the only amphi-Atlantic Astropecten species. Examination of the holotypes of both species and other specimens of A. cingulatus and A. jarli revealed that A. jarli has a greater major radius:minor radius than A. cingulatus at all sizes, a unique adambulacral spine configuration, and different spination on the inferomarginal plates. ANCOVA indicates A. cingulatus and A. jarli have similar major radius:number of superomarginal plates at small sizes but are distinguishable by the number of superomarginal plates in large individuals. The ratio of the width of the paxillar area:width of the superomarginal plates is greater in A. jarli. Some specimens identified as A. cingulatus from the Eastern Atlantic Region do not fit the description of either A. cingulatus or A. jarli. We conclude that A. jarli is morphologically distinct from A. cingulatus and should be recognized as a valid species. Astropecten cingulatus does not have an amphi-Atlantic distribution and is limited to the Western Atlantic Region.


Immunohistochemical and ultrastructural study of malignant plasmacytoid myoepithelioma of the maxillary sinus.
Myoepithelioma is a rare salivary gland tumor which is composed exclusively of myoepithelial cells. Histologically, it can be divided into three cell types: spindle, plasmacytoid and mixed type. Malignant myoepithelioma is characterized by invasive growth. In March 1995, a 60-year-old man presented with a left cheek tumor which he had first noted 2 years previously. Computed tomography revealed a large expansile tumor in the maxillary sinus with invasion into the surrounding soft tissue. Partial resection of the tumor was performed because of extensive involvement of the surrounding tissue. The patient died due to spread to the brain 5 months after surgery. Histologically, the tumor was composed exclusively of plasmacytoid cells, with bone destruction. Immunohistochemically, these cells were negative for immunoglobulin light chains (kappa and lambda) and heavy chains (Ig G, A, M) but positive for S-100, cytokeratin, actin and vimentin.Ultrastructurally, the tumor cells contained numerous randomly oriented actin-like microfilaments in the cytoplasm, and had desmosomes on the cell membrane. Malignant plasmacytoid myoepithelioma of the maxillary sinus was diagnosed. In addition to our case, only five cases of pure plasmacytoid myoepithelioma have been reported. Plasmacytoid myoepithelioma tends to occur in the minor salivary glands and has more aggressive behavior than spindle cell myoepithelioma. Morphologically, it is very difficult to differentiate plasmacytoid myoepithelioma from plasmacytoma and immunohistochemical staining is necessary to make a correct diagnosis.


The Preconception Stress and Resiliency Pathways Model: a multi-level framework on maternal, paternal, and child health disparities derived by community-based participatory research.
Emerging evidence supports the theoretical and clinical importance of the preconception period in influencing pregnancy outcomes and child health. Collectively, this evidence affirms the need for a novel, integrative theoretical framework to design future investigations, integrate new findings, and identify promising, evidence-informed interventions to improve intergenerational health and reduce disparities. This article presents a transdisciplinary framework developed by the NIH Community Child Health Network (CCHN) through community-based participatory research processes. CCHN developed a Preconception Stress and Resiliency Pathways (PSRP) model by building local and multi-site community-academic participatory partnerships that established guidelines for research planning and decision-making; reviewed relevant findings diverse disciplinary and community perspectives; and identified the major themes of stress and resilience within the context of families and communities. The PSRP model focuses on inter-relating the multiple, complex, and dynamic biosocial influences theoretically linked to family health disparities. The PSRP model borrowed from and then added original constructs relating to developmental origins of lifelong health, epigenetics, and neighborhood and community influences on pregnancy outcome and family functioning (cf. MCHJ 2014). Novel elements include centrality of the preconception/inter-conception period, role of fathers and the parental relationship, maternal allostatic load (a composite biomarker index of cumulative wear-and-tear of stress), resilience resources of parents, and local neighborhood and community level influences (e.g., employment, housing, education, health care, and stability of basic necessities). CCHN's integrative framework embraces new ways of thinking about how to improve outcomes for future generations, by starting before conception, by including all family members, and by engaging the community vigorously at multiple levels to promote resiliency, reduce chronic and acute stressors, and expand individualized health care that integrates promotive and prevention strategies. If widely adopted, the PSRP model may help realize the goal of sustaining engagement of communities, health and social services providers, and scientists to overcome the siloes, inefficiencies, and lack of innovation in efforts to reduce family health disparities. Model limitations include tremendous breadth and difficulty measuring all elements with precision and sensitivity.


Daphnia need to be gut-cleared too: the effect of exposure to and ingestion of metal-contaminated sediment on the gut-clearance patterns of D. magna.
The presence of sediment particles in the gut indicated that Daphnia magna used in whole-sediment bioassays ingest sediment. If gut contents are not removed prior to whole-body tissue-burden analysis, then the bioavailability of any sediment-associated contaminants (e.g. metals) can be overestimated. Gut clearing patterns were determined for D. magna after exposure to both clean and metal-contaminated (Cu and Zn) field-collected sediments. D. magna exposed to reference sediment had fuller guts than those exposed to metal-contaminated sediment (95% versus 60% full). Neither reference- nor metal-exposed D. magna could clear their gut completely of sediment particles when held in clean water for 24 h. When Daphnia were transferred to clean water after exposure to metal-contaminated sediment, there was no significant decrease in gut-fullness (P>0.05) even after 48 h of purging. By comparison, animals transferred to water containing 5 x 10(5) cells of algae (Pseudokircheriella subcapita) after exposure to contaminated sediment showed a significant drop in gut fullness from 56% immediately after exposure to 17% after 4 h of gut-clearance. Although gut fullness did not change significantly beyond 2 h of purging, data were much less variable after 8 h of gut-clearance than after 2 h or 4 h. The depuration of Cu was well described with a two-compartment first-order kinetic model (r2=0.78, P<0.0001) indicating that D. magna exposed to metal-contaminated sediment have one pool of Cu that is quickly depurated (0.2 h(-1)), and one that has been incorporated into the tissues (<<0.00001 h(-1)). Assuming tissue background of 48 microg/g, an exposed animal which has not been depurated or which has been purged with water alone would yield whole-body tissue Cu concentrations that are 5.6- and 4-fold higher, respectively, than that purged with algae + water (8 h). We recommend that D. magna used to estimate metal bioavailability from sediment be gut-cleared in the presence of algae for 8 h prior to determination of whole-body metal concentrations.


Nonhematological mechanisms of improved sea-level performance after hypoxic exposure.
Altitude training has been used regularly for the past five decades by elite endurance athletes, with the goal of improving performance at sea level. The dominant paradigm is that the improved performance at sea level is due primarily to an accelerated erythropoietic response due to the reduced oxygen available at altitude, leading to an increase in red cell mass, maximal oxygen uptake, and competitive performance. Blood doping and exogenous use of erythropoietin demonstrate the unequivocal performance benefits of more red blood cells to an athlete, but it is perhaps revealing that long-term residence at high altitude does not increase hemoglobin concentration in Tibetans and Ethiopians compared with the polycythemia commonly observed in Andeans. This review also explores evidence of factors other than accelerated erythropoiesis that can contribute to improved athletic performance at sea level after living and/or training in natural or artificial hypoxia. We describe a range of studies that have demonstrated performance improvements after various forms of altitude exposures despite no increase in red cell mass. In addition, the multifactor cascade of responses induced by hypoxia includes angiogenesis, glucose transport, glycolysis, and pH regulation, each of which may partially explain improved endurance performance independent of a larger number of red blood cells. Specific beneficial nonhematological factors include improved muscle efficiency probably at a mitochondrial level, greater muscle buffering, and the ability to tolerate lactic acid production. Future research should examine both hematological and nonhematological mechanisms of adaptation to hypoxia that might enhance the performance of elite athletes at sea level.


Biological gene delivery vehicles: beyond viral vectors.
Gene therapy covers a broad spectrum of applications, from gene replacement and knockdown for genetic or acquired diseases such as cancer, to vaccination, each with different requirements for gene delivery. Viral vectors and synthetic liposomes have emerged as the vehicles of choice for many applications today, but both have limitations and risks, including complexity of production, limited packaging capacity, and unfavorable immunological features, which restrict gene therapy applications and hold back the potential for preventive gene therapy. While continuing to improve these vectors, it is important to investigate other options, particularly nonviral biological agents which include bacteria, bacteriophage, virus-like particles (VLPs), erythrocyte ghosts, and exosomes. Exploiting the natural properties of these biological entities for specific gene delivery applications will expand the repertoire of gene therapy vectors available for clinical use. Here, we review the prospects for nonviral biological delivery vehicles as gene therapy agents with focus on their unique evolved biological properties and respective limitations and potential applications. The potential of these nonviral biological entities to act as clinical gene therapy delivery vehicles has already been shown in clinical trials using bacteria-mediated gene transfer and with sufficient development, these entities will complement the established delivery techniques for gene therapy applications.


Increased mandibular condylar growth in mice with estrogen receptor beta deficiency.
Temporomandibular joint (TMJ) disorders predominantly afflict women of childbearing age, suggesting a role for female hormones in the disease process. In long bones, estrogen acting via estrogen receptor beta (ERβ) inhibits axial skeletal growth in female mice. However, the role of ERβ in the mandibular condyle is largely unknown. We hypothesize that female ERβ-deficient mice will have increased mandibular condylar growth compared to wild-type (WT) female mice. This study examined female 7-day-old, 49-day-old, and 120-day-old WT and ERβ knockout (KO) mice. There was a significant increase in mandibular condylar cartilage thickness as a result of an increased number of cells, in the 49-day-old and 120-day-old female ERβ KO compared with WT controls. Analysis in 49-day-old female ERβ KO mice revealed a significant increase in collagen type X, parathyroid hormone-related protein (Pthrp), and osteoprotegerin gene expression and a significant decrease in receptor activator for nuclear factor κ B ligand (Rankl) and Indian hedgehog (Ihh) gene expression, compared with WT controls. Subchondral bone analysis revealed a significant increase in total condylar volume and a decrease in the number of osteoclasts in the 49-day-old ERβ KO compared with WT female mice. There was no difference in cell proliferation in condylar cartilage between the genotypes. However, there were differences in the expression of proteins that regulate the cell cycle; we found a decrease in the expression of Tieg1 and p57 in the mandibular condylar cartilage from ERβ KO mice compared with WT mice. Taken together, our results suggest that ERβ deficiency increases condylar growth in female mice by inhibiting the turnover of fibrocartilage.


Hemochromatosis: a Neolithic adaptation to cereal grain diets.
The Neolithic period in Europe marked the transition from a hunter-gatherer diet rich in red meat to an iron-reduced cereal grain diet. This dietary shift likely resulted in an increased incidence of iron deficiency anemia, especially in women of reproductive age. I propose that hereditary hemochromatosis and in particular the common HFE C282Y mutation may represent an adaptation to decreased dietary iron in cereal grain-based Neolithic diets. Both homozygous and heterozygous carriers of the HFE C282Y mutation have increased iron stores and therefore possessed an adaptive advantage under Neolithic conditions. An allele age estimate places the origin of the HFE C282Y mutation in the early Neolithic period in Northern Europe and is thus consistent with this hypothesis. The lower incidence of this mutation in other agrarian regions (the Mediterranean and Near East) may be due to higher dietary intakes of the iron uptake cofactor vitamin C in those regions. The HFE C282Y mutation likely only became maladaptive in the past several centuries as dietary sources of iron and vitamin C improved in Northern Europe.


CEACAM6 acts as a receptor for adherent-invasive E. coli, supporting ileal mucosa colonization in Crohn disease.
The ileal mucosa of Crohn disease (CD) patients is abnormally colonized by adherent-invasive E. coli (AIEC) that are able to adhere to and invade intestinal epithelial cells. Here, we show that CD-associated AIEC strains adhere to the brush border of primary ileal enterocytes isolated from CD patients but not controls without inflammatory bowel disease. AIEC adhesion is dependent on type 1 pili expression on the bacterial surface and on carcinoembryonic antigen-related cell adhesion molecule 6 (CEACAM6) expression on the apical surface of ileal epithelial cells. We report also that CEACAM6 acts as a receptor for AIEC adhesion and is abnormally expressed by ileal epithelial cells in CD patients. In addition, our in vitro studies show that there is increased CEACAM6 expression in cultured intestinal epithelial cells after IFN-gamma or TNF-alpha stimulation and after infection with AIEC bacteria, indicating that AIEC can promote its own colonization in CD patients.


Use of residual fraction instead of residual volume in the evaluation of lower urinary tract symptoms.
Assessment of postvoid residual volume (PVR) has become a valuable routine investigation in the evaluation of bladder outlet obstruction. PVR has been shown to have interindividual and intraindividual variation and dependence on prevoid urinary volume, thus raising a question about its significance. The aim of this study was to investigate an alternate parameter more reliable than PVR, described as residual fraction (RF) and calculated as (PVR x 100)/prevoid volume. Ninety-three adult patients with lower urinary tract symptoms (LUTS) presenting to the urology outpatient clinic were evaluated for bladder outlet obstruction. Patients with urinary retention and neurological disorders were excluded. Evaluation was by clinical assessment, uroflowmetry, and ultrasound bladder for prevoid and postvoid urine volume estimation. The latter was compared with RF. Results were statistically analyzed using bivariate analysis and Spearman's test. In 93 evaluable patients, there were 87 (94%) males and 6 (6%) females. Residual volume ranged from 4 to 450 mL (mean 91.4+/-92.7 mL). Peak flow rate (Qmax) varied from 3 to 49 mL/s (mean 9+/-15 mLis). Two-tailed correlation between PVR and Qmax was significant at the .05 level, whereas a more significant R value at .01 was observed between RF and Qmax. There was a strong positive correlation between RF and PVR with Qmax. RF statistically correlated better with Qmax than PVR. It is recommended that RF be used instead of PVR in the routine noninvasive evaluation of LUTS.


Observer detection limits for a dedicated SPECT breast imaging system.
An observer-based contrast-detail study is performed in an effort to evaluate the limits of object detectability using a dedicated CZT-based breast SPECT imaging system under various imaging conditions. A custom geometric contrast-resolution phantom was developed that can be used for both positive ('hot') and negative contrasts ('cold'). The 3 cm long fillable tubes are arranged in six sectors having equal inner diameters ranging from 1 mm to 6 mm with plastic wall thicknesses of <0.25 mm, on a pitch of twice their inner diameters. Scans of the activity filled tubes using simple circular trajectories are obtained in a 215 mL uniform water filled cylinder, varying the rod:background concentration ratios from 10:1 to 1:10 simulating a large range of biological uptake ratios. The rod phantom is then placed inside a non-uniformly shaped 500 mL breast phantom and scans are again acquired using both simple and complex 3D trajectories for similarly varying contrasts. Summed slice and contiguous multi-slice images are evaluated by five independent readers, identifying the smallest distinguishable rod for each concentration and experimental setup. Linear and quadratic regression is used to compare the resulting contrast-detail curves. Results indicate that in a moderately low-noise 500 mL background, using the SPECT camera having 2.5 mm intrinsic pixels, the mean detectable rod was approximately 3.4 mm at a 10:1 ratio, degrading to approximately 5.2 mm with the 2.5:1 concentration ratio. The smallest object detail was observed using a 45 degrees tilted trajectory acquisition. The complex 3D projected sine wave acquisition, however, had the most consistent combined intra- and inter-observer results, making it potentially the best imaging approach for consistent results.