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Patients with the genetic blistering skin condition epidermolysis bullosa (EB) report severe pain as a consequence of skin and mucous membrane lesions including blisters, wounds, and scars. Adequate symptom alleviation is not often achieved using conventional pharmacologic interventions. Finding novel approaches to pain care in EB is imperative to improve the quality of life of patients living with EB. There are several anecdotal reports on the use of cannabinoid-based medicines (CBMs) by EB patients to reduce the burden of symptoms. However, controlled clinical investigations assessing these reported effects are lacking. As the pain quality "unpleasantness" delineates EB pain, we hypothesize the modulation of affective pain processing in the brain by way of intervention with CBMs comprising the cannabinoids Δ-9-tetrahydrocannabinol and cannabidiol-objectified by functional magnetic resonance imaging (fMRI). The C4EB study is an investigator-initiated, single-centre, randomized, double-blind, placebo-controlled and crossover trial. Adult patients with the diagnosis epidermolysis bullosa, reporting chronic pain will be eligible to participate. Following baseline measurements, participants will be randomized to receive the sublingually administered interventions placebo and Transvamix® in forward or reversed orders, each for two weeks and separated by a washout. The primary outcome is the difference in numeric rating scale pain scores between grouped interventions, using affective descriptors within the Short-form McGill Pain Questionnaire-2. Secondary outcomes include pain self-efficacy, concomitant analgesic medication-use and adverse events. Additionally, fMRI will be employed to assess brain connectivity related to neuroanatomic pain circuits at baseline, placebo and Transvamix® interventions. The study was approved by the ethical committee at the University Medical Center of Groningen in the Netherlands. Results will be submitted for publication in a peer-reviewed journal. Trial registration number: Netherlands Trial Register: NL9347 (Acronym: C4EB).

BackgroundThe ability to accurately distinguish bacterial from viral infection would help clinicians better target antimicrobial therapy during suspected lower respiratory tract infections (LRTI). Although technological developments make it feasible to rapidly generate patient-specific microbiota profiles, evidence is required to show the clinical value of using microbiota data for infection diagnosis. In this study, we investigated whether adding nasal cavity microbiota profiles to readily available clinical information could improve machine learning classifiers to distinguish bacterial from viral infection in patients with LRTI.ResultsVarious multi-parametric Random Forests classifiers were evaluated on the clinical and microbiota data of 293 LRTI patients for their prediction accuracies to differentiate bacterial from viral infection. The most predictive variable was C-reactive protein (CRP). We observed a marginal prediction improvement when 7 most prevalent nasal microbiota genera were added to the CRP model. In contrast, adding three clinical variables, absolute neutrophil count, consolidation on X-ray, and age group to the CRP model significantly improved the prediction. The best model correctly predicted 85% of the 'bacterial' patients and 82% of the 'viral' patients using 13 clinical and 3 nasal cavity microbiota genera (Staphylococcus, Moraxella, and Streptococcus).ConclusionsWe developed high-accuracy multi-parametric machine learning classifiers to differentiate bacterial from viral infections in LRTI patients of various ages. We demonstrated the predictive value of four easy-to-collect clinical variables which facilitate personalized and accurate clinical decision-making. We observed that nasal cavity microbiota correlate with the clinical variables and thus may not add significant value to diagnostic algorithms that aim to differentiate bacterial from viral infections.

Changes in funerary practices are key to the understanding of social transformations of past societies. Over the course of the Nordic Bronze Age, funerary practices changed from inhumation to cremation. The aim of this study is to shed light on this fundamental change through a cross-examination of archaeometric provenance data and archaeological discussions of the context and layouts of early cremation graves. To this end, we conducted 19 new provenance analyses of strontium isotopes from Early Nordic Bronze age contexts in Thisted County and Zealand and Late Bronze Age contexts from Thisted County and Vesthimmerland (Denmark). These data are subsequently compared with data from other extant relevant studies, including those from Late Bronze Age Fraugde on the Danish island of Fyn. Overall, the variations within our provenience data suggest that the integration and establishment of cremation may not have had a one-to-one relationship with in-migration to Nordic Bronze Age Denmark. Moreover, there seems to be no single blanket scenario which dictated the uptake of cremation as a practice within this part of Southern Scandinavia. By addressing habitus in relation to the deposition of cremations as juxtaposed with these provenance data¸ we hypothesize several potential pathways for the uptake of cremation as a new cultural practice within the Danish Nordic Bronze Age and suggest that this may have been a highly individual process, whose tempo may have been dictated by the specificities of the region(s) concerned.

RATIONALE:A subset of patients with idiopathic pulmonary fibrosis (IPF) contains short leukocyte telomeres or telomere related mutations. We previously showed that alveolar type 2 cells have short telomeres in fibrotic lesions. Our objectives were to better understand how telomere shortening associates with fibrosis in IPF lung and identify a subset of patients with telomere-related disease. METHODS:Average telomere length was determined in multiple organs, basal and apical lung, and diagnostic and end-stage fibrotic lung biopsies. Alveolar type 2 cells telomere length was determined in different areas of IPF lungs. RESULTS:In IPF but not in controls, telomere length in lung was shorter than in other organs, providing rationale to focus on telomere length in lung. Telomere length did not correlate with age and no difference in telomere length was found between diagnostic and explant lung or between basal and apical lung, irrespective of the presence of a radiological apicobasal gradient or fibrosis. Fifteen out of 28 IPF patients had average lung telomere length in the range of patients with a telomerase (TERT) mutation, and formed the IPFshort group. Only in this IPFshort and TERT group telomeres of alveolar type 2 cells were extremely short in fibrotic areas. Additionally, whole exome sequencing of IPF patients revealed two genetic variations in RTEL1 and one in PARN in the IPFshort group. CONCLUSIONS:Average lung tissue telomere shortening does not associated with fibrotic patterns in IPF, however, approximately half of IPF patients show excessive lung telomere shortening that is associated with pulmonary fibrosis driven by telomere attrition.

Antimicrobial Resistance is a global crisis that veterinarians contribute to through their use of antimicrobials in animals. Antimicrobial stewardship has been shown to be an effective means to reduce antimicrobial resistance in hospital environments. Effective monitoring of antimicrobial usage patterns is an essential part of antimicrobial stewardship and is critical in reducing the development of antimicrobial resistance. The aim of this study is to describe how frequently antimicrobials were used in veterinary consultations and identify the most frequently used antimicrobials. Using VetCompass Australia, Natural Language Processing techniques, and the Australian Strategic Technical Advisory Group's (ASTAG) Rating system to classify the importance of antimicrobials, descriptive analysis was performed on the antimicrobials prescribed in consultations from 137 companion animal veterinary clinics in Australia between 2013 and 2017 (inclusive). Of the 4,400,519 consultations downloaded there were 595,089 consultations where antimicrobials were prescribed to dogs or cats. Antimicrobials were dispensed in 145 of every 1000 canine consultations; and 38 per 1000 consultations involved high importance rated antimicrobials. Similarly with cats, 108 per 1000 consultations had antimicrobials dispensed, and in 47 per 1000 consultations an antimicrobial of high importance rating was administered. The most common antimicrobials given to cats and dogs were cefovecin and amoxycillin clavulanate, respectively. The most common topical antimicrobial and high-rated topical antimicrobial given to dogs and cats was polymyxin B. This study provides a descriptive analysis of the antimicrobial usage patterns in Australia using methods that can be automated to inform antimicrobial use surveillance programs and promote antimicrobial stewardship.

BACKGROUND:Because of a lack of preference-based health-related quality of life (HRQoL) instruments suitable for infants aged 0-12 months, we previously developed the Infant QoL Instrument (IQI). The present study aimed to generate an algorithm to estimate utilities for the IQI. METHODS:Via an online survey, respondents from the general population and primary caregivers from China-Hong Kong, the UK, and the USA were presented 10 discrete choice scenarios based on the IQI classification system. An additional sample of respondents from the general population were also asked if they considered the examined health states to be worse than death. Coefficients for the IQI item levels were obtained with a conditional logit model based on the responses of the primary caregivers for IQI states only. These coefficients were then normalized using the rank-ordered logit model based on the responses from the general population who assessed "death" as a choice option. In this way, the values were rescaled from full health (1.0) to death (0.0), and consequently, they became suitable for the computation of quality-adjusted life years. RESULTS:The total sample consisted of 1409 members of the general population and 1229 primary caregivers. Results indicated that, out of the 7 IQI items ("sleeping," "feeding," "breathing," "stooling/poo," "mood," "skin," and "interaction"), "breathing" had the highest impact on the HRQoL of infants. Moreover, except for "stooling," all item levels were statistically significant. The general population sample considered none of the health states as worse than death. The utility value for the worst health state was 0.015 (State 4444444). CONCLUSIONS:The IQI is the first generic instrument to assess overall HRQoL in 0-1-year-old infants by providing values and utilities. Using discrete choice experiments, we demonstrated that it is possible to derive utilities of infant health states. The next step will be to collect IQI values in a clinical population of infants and to compare these values with those of other instruments.

BackgroundInvasive fungal infection (IFI) detection requires application of complex case definitions by trained staff. Administrative coding data (ICD-10-AM) may provide a simplified method for IFI surveillance, but accuracy of case ascertainment in children with cancer is unknown.ObjectiveTo determine the classification performance of ICD-10-AM codes for detecting IFI using a gold-standard dataset (r-TERIFIC) of confirmed IFIs in paediatric cancer patients at a quaternary referral centre (Royal Children's Hospital) in Victoria, Australia from 1st April 2004 to 31st December 2013.MethodsICD-10-AM codes denoting IFI in paediatric patients (ResultsOf 1,671 evaluable patients, 113 (6.76%) had confirmed IFI diagnoses according to gold-standard criteria, while 114 (6.82%) cases were identified using the codes. Of the clinical IFI cases, 68 were in receipt of ≥1 ICD-10-AM code(s) for IFI, corresponding to an overall sensitivity, PPV and F1 score of 60%, respectively. Sensitivity was highest for proven IFI (77% [95% CI: 58-90]; F1 = 47%) and invasive candidiasis (83% [95% CI: 61-95]; F1 = 76%) and lowest for other/unspecified IFI (20% [95% CI: 5.05-72%]; F1 = 5.00%). The most frequent misclassification was coding of invasive aspergillosis as invasive candidiasis.ConclusionICD-10-AM codes demonstrate moderate sensitivity and PPV to detect IFI in children with cancer. However, specific subsets of proven IFI and invasive candidiasis (codes B37.x) are more accurately coded.

BackgroundIn many countries, medical interventions are reimbursed on the basis of recommendations made by advisory boards and committees that apply multiple criteria in their assessment procedures. Given the diversity of these criteria, it is difficult to find common ground to determine what information is required for setting priorities.ObjectiveTo investigate whether society and patients share the same interests and views concerning healthcare priorities.MethodsWe applied a framework of discrete choice models in which respondents were presented with judgmental tasks to elicit their preferences. They were asked to choose between two hypothetical scenarios of patients receiving a new treatment. The scenarios graphically presented treatment outcomes and patient characteristics. Responses were collected through an online survey administered among respondents from the general population (N = 1,253) and patients (N = 1,389) and were analyzed using conditional logit and mixed logit models.ResultsThe respondents' preferences regarding new medical treatments revealed that they attached the most relative importance to additional survival years, age at treatment, initial health condition, and the cause of disease. Minor differences in the relative importance assigned to three criteria: age at treatment, initial health, and cause of disease were found between the general population and patient samples. Health scenarios in which patients had higher initial health-related quality of life (i.e., a lower burden of disease) were favored over those in which patients' initial health-related quality of life was lower.ConclusionsOverall, respondents within the general population expressed preferences that were similar to those of the patients. Therefore, priority-setting studies that are based on the perspectives of the general population may be useful for informing decisions on reimbursement and other types of priority-setting processes in health care. Incorporating the preferences of the general population may simultaneously increase public acceptance of these decisions.

We present the first comprehensive multi-isotopic data on human and animal remains from the Final Neolithic Corded Ware culture (ca. 2900-2300 cal. BC) in south-eastern Poland. The study focused on communities of two settlement areas located in the Małopolska Upland and in the Subcarpathian region. Carbon and nitrogen isotopes of bone collagen were investigated to obtain insights into human dietary preferences, whereas the strontium isotope composition of human tooth enamel was used to trace the mobility and provenance of individuals. Sr isotope data point to a non-local origin of at least one-quarter of the investigated individuals in the Subcarpathian region, consistent with associated allochthonous grave inventories of eastern or western origins. In contrast, all investigated individuals in the Małopolska Upland were of local origin. Furthermore, our study shows an example that the use of fauna for the assessment of the local 87Sr/86Sr range of an archaeological site can lead to incorrect conclusions and suggests that a detailed Sr isotopic survey of the geological background and its hydrologic elements is necessary to provide conclusive constraints for the identification of local and non-local individuals in prehistoric communities. Carbon and nitrogen isotope composition of bone collagen indicate an omnivorous diet that included C3-based terrestrial plant and animal resources, in which plant food dominated. In both regions, there were no significant sex differences in dietary intakes. Higher δ15Ncoll values of younger infants presumably reflect the effect of weaning.

The inland area of southwestern Sweden is well known for its well-preserved archaeological animal and human remains dating back to the Mesolithic and Neolithic (10000-4000 and 4000-1700 BC). They allow application of multiple bioarchaeological methods, giving insights into various and complementary aspects of prehistoric human life, as well as economic and social structures. One important aspect concerns human mobility and its relation to social networks and to circulation of objects. Here, strontium isotope analysis plays a crucial role. The present study aims to construct a strontium isotope baseline of southwestern Sweden with considerably greater coverage and higher resolution than previously published data. As the region has been affected by glacial events, the relation between bedrock geology and isotope signals of the bioavailable strontium in such areas is given special attention. We determined strontium isotope ratios for 61 water and five archaeological animal samples, and combined the data with previous measurements of two water and 21 non-domestic faunal samples. The results reveal a complex pattern. Several areas with distinct baseline ranges can be distinguished, although with overlaps between some of them. Overall, the bioavailable strontium isotope signals mirror the basement geology of the region. The highest ratios occur in the geologically oldest eastern parts of the Precambrian terrain, while lower ratios are found in the western part, and the lowest ratios occur in the youngest Paleozoic areas. At the same time, there are minor deviations compared to the underlying bedrock, due to glacial transport, overlying sediments, and local intrusions of younger rocks. The background data set now available allows for more nuanced and detailed interpretations of human and animal mobility in the region, in particular by identification of subregions with differing strontium isotope ratios within the Precambrian province. Also, we can now identify long distance mobility with greater confidence.

BACKGROUND:The assessment of health-related quality of life (HRQoL) is important for health outcomes research, disease modeling studies and comparisons of different healthcare interventions. Yet, only a few tools are available to assess HRQoL in 0-1-year-old infants. Furthermore, there is a need for an instrument able to assess HRQoL with a single, standardized, overall score in the first year of life. Here we described the development of the Infant health-related Quality of life Instrument (IQI), a generic, preference-based instrument that can be administered through a mobile application for assessing HRQoL in 0-1-year-old infants. METHODS:A multi-step development process began by extracting candidate health concepts from relevant measures identified by two literature searches. Next, three panels, with experts from Asia, Europe, New Zealand and United States of America, and two surveys, with primary caregivers in New Zealand, Singapore, and the United Kingdom, evaluated the relevance of the candidate health concepts, organized them into attributes based on their similarities, explored alternative attributes and generated response scales. Additional interviews assessed the cross-cultural interpretability, parents' understanding of health attributes, and the usability of the mobile application. RESULTS:The final list of 7 health attributes included in the IQI consisted of sleeping, feeding, breathing, stooling/poo, mood, skin, and interaction. The users' experiences with the mobile application were generally positive. CONCLUSIONS:The IQI is the first generic, preference-based, instrument designed to assess overall HRQoL in 0-1-year old infants. It is short and easy-to-administer through a mobile application. Moreover, close attention was paid to the opinions of the infants' primary caregivers during the instrument and mobile application development process.

Gonad morphogenesis relies on the correct spatiotemporal expression of a number of genes that together fulfill the differentiation of the bipotential gonad into testes or ovaries. As such, the transcription factors WT1 and GATA4 are pivotal for proper gonadal development. Here we address the contributions of GATA4 and WT1 to the sex differentiation phase in testes and ovaries. We applied an ex vivo technique for cultivating gonads in hanging droplets of media that were supplemented with vivo-morpholinos to knockdown WT1 and GATA4 either alone or in combination at the same developmental stage. We show that WT1 is equally important for both, the initial establishment and the maintenance of the sex-specific gene expression signature in testes and ovaries. We further identified Foxl2 as a novel putative downstream target gene of WT1. Moreover, knockdown of WT1 reduced mRNA levels of several molecular components of the hedgehog signaling pathway in XY gonads, whereas Gata4 vivo-morpholino treatment increased transcripts of Dhh and Ptch1 in embryonic testes. The data suggest that for its proper function, WT1 relies on the correct expression of the GATA4 protein. Furthermore, GATA4 down-regulates several ovarian promoting genes in testes, such as Ctnnb1, Fst, and Bmp2, suggesting that this repression is required for maintaining the male phenotype. In conclusion, this study provides novel insights into the role of WT1 and GATA4 during the sex differentiation phase and represents an approach that can be applied to assess other proteins with as yet unknown functions during gonadal development.

Telomere maintenance dysfunction has been implicated in the pathogenesis of Idiopathic Pulmonary Fibrosis (IPF). However, the mechanism of how telomere length is related to fibrosis in the lungs is unknown. Surgical lung biopsies of IPF patients typically show a heterogeneous pattern of non-fibrotic and fibrotic areas. Therefore, telomere length (TL) in both lung areas of patients with IPF and familial interstitial pneumonia was compared, specifically in alveolar type 2 (AT2) cells. Fluorescent in situ hybridization was used to determine TL in non-fibrotic and fibrotic areas of 35 subjects. Monochrome multiplex quantitative polymerase chain reaction (MMqPCR) was used for 51 whole lung biopsies and blood TL measurements. For sporadic IPF subjects, AT2 cell TL in non-fibrotic areas was 56% longer than in fibrotic areas. No such difference was observed in the surrounding lung cells. In subjects carrying a telomerase reverse transcriptase (TERT) mutation, AT2 cell TL was significantly shorter than in sporadic subjects. However, no difference in surrounding cell TL was observed between these subject groups. Finally, using biopsy MMqPCR TL measurements, it was determined that IPF subjects with shortest lung TL had a significantly worse survival than patients with long TL. This study shows that shortening of telomeres critically affects AT2 cells in fibrotic areas, implying TL as a cause of fibrogenesis. Furthermore, short lung telomere length is associated with decreased survival.

The aim of this pilot study was to evaluate the potential of a new noninvasive optical measurement of muscle oxygenation (MOx) to identify shock severity in patients with suspected sepsis.We enrolled 51 adult patients in the emergency department (ED) who presented with possible sepsis using traditional Systematic Inflammatory Response Syndrome criteria or who triggered a "Code Sepsis." Noninvasive MOx measurements were made from the first dorsal interosseous muscles of the hand once potential sepsis/septic shock was identified, as soon as possible after admission to the ED. Shock severity was defined by concurrent systolic blood pressure, heart rate, and serum lactate levels. MOx was also measured in a control group of 17 healthy adults.Mean (± SD) MOx in the healthy control group was 91.0 ± 5.5% (n = 17). Patients with mild, moderate, and severe shock had mean MOx values of 79.4 ± 21.2%, 48.6 ± 28.6%, and 42.2 ± 4.7%, respectively. Mean MOx for the mild and moderate shock severity categories were statistically different from healthy controls and from each other based on two-sample t-tests (p < 0.05).We demonstrate that noninvasive measurement of MOx was associated with clinical assessment of shock severity in suspected severe sepsis or septic shock. The ability of MOx to detect even mild septic shock has meaningful implications for emergency care, where decisions about triage and therapy must be made quickly and accurately. Future longitudinal studies may validate these findings and the value of MOx in monitoring patient status as treatment is administered.

Parkinson's disease (PD) is the second most prevalent neurodegenerative disorder worldwide and characterized by the loss of dopaminergic neurons in the patients' midbrains. Both the presence of the protein α-synuclein in intracellular protein aggregates in surviving neurons and the genetic linking of the α-synuclein encoding gene point towards a major role of α-synuclein in PD etiology. The exact pathogenic mechanisms of PD development are not entirely described to date, neither is the specific role of α-synuclein in this context. Previous studies indicate that one aspect of α-synuclein-related cellular toxicity might be direct proteasome impairment. The 20/26S proteasomal machinery is an important instrument of intracellular protein degradation. Thus, direct proteasome impairment by α-synuclein might explain or at least contribute to the formation of intracellular protein aggregates. Therefore this study investigates direct proteasomal impairment by α-synuclein both in vitro using recombinant α-synuclein and isolated proteasomes as well as in living cells. Our experiments demonstrate that the impairment of proteasome activity by α-synuclein is highly dependent upon the cellular background and origin. We show that recombinant α-synuclein oligomers and fibrils scarcely affect 20S proteasome function in vitro, neither does transient α-synuclein expression in U2OS ps 2042 (Ubi(G76V)-GFP) cells. However, stable expression of both wild-type and mutant α-synuclein in dopaminergic SH-SY5Y and PC12 cells results in a prominent impairment of the chymotrypsin-like 20S/26S proteasomal protein cleavage. Thus, our results support the idea that α-synuclein in a specific cellular environment, potentially present in dopaminergic cells, cannot be processed by the proteasome and thus contributes to a selective vulnerability of dopaminergic cells to α-synuclein pathology.

BACKGROUND:Vitamin D deficiency may be a risk factor for mortality but previous meta-analyses lacked standardization of laboratory methods for 25-hydroxyvitamin D (25[OH]D) concentrations and used aggregate data instead of individual participant data (IPD). We therefore performed an IPD meta-analysis on the association between standardized serum 25(OH)D and mortality. METHODS:In a European consortium of eight prospective studies, including seven general population cohorts, we used the Vitamin D Standardization Program (VDSP) protocols to standardize 25(OH)D data. Meta-analyses using a one step procedure on IPD were performed to study associations of 25(OH)D with all-cause mortality as the primary outcome, and with cardiovascular and cancer mortality as secondary outcomes. This meta-analysis is registered at ClinicalTrials.gov, number NCT02438488. FINDINGS:We analysed 26916 study participants (median age 61.6 years, 58% females) with a median 25(OH)D concentration of 53.8 nmol/L. During a median follow-up time of 10.5 years, 6802 persons died. Compared to participants with 25(OH)D concentrations of 75 to 99.99 nmol/L, the adjusted hazard ratios (with 95% confidence interval) for mortality in the 25(OH)D groups with 40 to 49.99, 30 to 39.99, and

AIMS:There is controversy in the guidelines as to whether patients with atrial fibrillation and a low risk of stroke should be treated with anticoagulation, especially those with a CHA2DS2-VASc score of 1 point. METHODS:In a retrospective, nationwide cohort study, we used the Swedish National Patient Registry, the National Prescribed Drugs Registry, the Swedish Registry of Education and the Population and Housing Census Registry. 48 433 patients were identified between 1 January 2006 and 31 December 2008 with incident atrial fibrillation who were divided in age categories, sex and a CHA2DS2-VASc score of 0, 1, 2 and ≥3 and they were included in a time-varying analysis of warfarin treatment versus no treatment. The primary end-point was cerebral infarction and stroke, and patients were followed until 31 December 2009. RESULTS:Patients with 1 point from the CHA2DS2-VASc score showed the following adjusted hazard ratios (HR) with a 95% confidence interval: men 65-74 years 0.46 (0.25-0.83), men

Much attention has been directed to the physiological effects of nitric oxide (NO)-cGMP signaling, but virtually nothing is known about its hematologic effects. We reported for the first time that cGMP signaling induces human γ-globin gene expression. Aiming at developing novel therapeutics for anemia, we examined here the hematologic effects of NO-cGMP signaling in vivo and in vitro. We treated wild-type mice with NO to activate soluble guanylate cyclase (sGC), a key enzyme of cGMP signaling. Compared to untreated mice, NO-treated mice had higher red blood cell counts and total hemoglobin but reduced leukocyte counts, demonstrating that when activated, NO-cGMP signaling exerts hematopoietic effects on multiple types of blood cells in vivo. We next generated mice which overexpressed rat sGC in erythroid and myeloid cells. The forced expression of sGCs activated cGMP signaling in both lineage cells. Compared with non-transgenic littermates, sGC mice exhibited hematologic changes similar to those of NO-treated mice. Consistently, a membrane-permeable cGMP enhanced the differentiation of hematopoietic progenitors toward erythroid-lineage cells but inhibited them toward myeloid-lineage cells by controlling multiple lineage-specific transcription factors. Human γ-globin gene expression was induced at low but appreciable levels in sGC mice carrying the human β-globin locus. Together, these results demonstrate that NO-cGMP signaling is capable of stimulating erythropoiesis in both in vitro and vivo settings by controlling the expression of multiple lineage-specific transcription factors, suggesting that cGMP signaling upregulates erythropoiesis at the level of gene transcription. The NO-cGMP signaling axis may constitute a novel target to stimulate erythropoiesis in vivo.

PURPOSE:Self-reported data suggest that older adults with dementia are inactive. The purpose of the present study was to objectively assess the physical activity (PA) levels of community-dwelling and institutionalized ambulatory patients with dementia, and to compare with the PA levels of cognitive healthy older adults. METHODS:We used actigraphy to assess the PA levels in institutionalized (n = 83, age: 83.0 ± 7.6, Mini-Mental-State Examination (MMSE): 15.5 ± 6.5) and community-dwelling dementia patients (n = 37, age: 77.3 ± 5.6, MMSE-score: 20.8 ± 4.8), and healthy older adults (n = 26, age: 79.5 ± 5.6, MMSE-score: 28.2 ± 1.6). We characterized PA levels based on the raw data and classified

Proteolytic processing of amyloid-β precursor protein (APP) by beta-site APP cleaving enzyme 1 (BACE1) is the initial step in the production of amyloid beta (Aβ), which accumulates in senile plaques in Alzheimer's disease (AD). Essential for this cleavage is the transport and sorting of both proteins through endosomal/Golgi compartments. Golgi-localized γ-ear-containing ARF-binding (GGA) proteins have striking cargo-sorting functions in these pathways. Recently, GGA1 and GGA3 were shown to interact with BACE1, to be expressed in neurons, and to be decreased in AD brain, whereas little is known about GGA2. Since GGA1 impacts Aβ generation by confining APP to the Golgi and perinuclear compartments, we tested whether all GGAs modulate BACE1 and APP transport and processing. We observed decreased levels of secreted APP alpha (sAPPα), sAPPβ, and Aβ upon GGA overexpression, which could be reverted by knockdown. GGA-BACE1 co-immunoprecipitation was impaired upon GGA-GAE but not VHS domain deletion. Autoinhibition of the GGA1-VHS domain was irrelevant for BACE1 interaction. Our data suggest that all three GGAs affect APP processing via the GGA-GAE domain.

Familial adenomatous polyposis (FAP) is often due to adenomatous polyposis coli (APC) gene germline mutations. Somatic APC defects are found in about 80% of colorectal cancers (CRCs) and adenomas. Rapamycin inhibits mammalian target of rapamycin (mTOR) protein, which is often expressed in human adenomas and CRCs. We sought to assess the effects of rapamycin in a mouse polyposis model in which both Apc alleles were conditionally inactivated in colon epithelium. Two days after inactivating Apc, mice were given rapamycin or vehicle in cycles of two weeks on and two weeks off. Polyps were scored endoscopically. Mice were euthanized at time points or when moribund, and tissue analyses were performed. In other studies, mice with demonstrable Apc-defective colon polyps were given rapamycin, followed by analysis of their colon tissues. The median survival of mice receiving rapamycin treatment cycles was 21.5 versus 6.5 weeks in control mice (p = 0.03), and rapamycin-treated mice had a significantly lower percentage of their colon covered with polyps (4.3+/- 2 vs 56.5+/- 10.8 percent, p = 0.001). Mice with Apc-deficient colon tissues that developed high grade dysplasia treated with rapamycin underwent treatment for significantly longer than mice treated with vehicle (15.8 vs 5.1 weeks, p = 0.003). In Apc-defective colon tissues, rapamycin treatment was linked to decreased levels of β-catenin and Sox9 at 7 weeks. Other effects of rapamycin in Apc-defectivecolon tissues included decreased proliferation and increased numbers of differentiated goblet cells at 7 weeks. Rapamycin did not affect β-catenin-regulated gene expression in cultured intestinal epithelial cells. Rapamycin has potent inhibitory effects in a mouse colon polyposis model, and mTOR inhibition is linked to decreased proliferation and increased expression of differentiation markers in Apc-mutant colon epithelium and delays development of dysplasia. Our findings highlight the possibility that mTOR inhibitors may have relevance for polyposis inhibition approaches in FAP patients.

The objectives of this study were to develop a rat model of gastrointestinal colonization with vancomycin-resistant Enterococcus faecalis (VRE) and extended-spectrum beta-lactamase (ESBL)-producing E. coli and to evaluate intestinal translocation to blood and tissues after total and partial hepatic ischemia. Methods - We developed a model of rat colonization with VRE and ESBL-E coli. Then we studied four groups of colonized rats: Group I (with hepatic pedicle occlusion causing complete liver ischemia and intestinal stasis); Group II (with partial liver ischemia without intestinal stasis); Group III (surgical manipulation without hepatic ischemia or intestinal stasis); Group IV (anesthetized without surgical manipulation). After sacrifice, portal and systemic blood, large intestine, small intestine, spleen, liver, lungs, and cervical and mesenteric lymph nodes were cultured. Endotoxin concentrations in portal and systemic blood were determined. Results - The best inocula were: VRE: 2.4×10(10) cfu and ESBL-E. coli: 1.12×10(10) cfu. The best results occurred 24 hours after inoculation and antibiotic doses of 750 µg/mL of water for vancomycin and 2.1 mg/mL for ceftriaxone. There was a significantly higher proportion of positive cultures for ESBL-E. coli in the lungs in Groups I, II and III when compared with Group IV (67%; 60%; 75% and 13%, respectively; p:0.04). VRE growth was more frequent in mesenteric lymph nodes for Groups I (67%) and III (38%) than for Groups II (13%) and IV (none) (p:0.002). LPS was significantly higher in systemic blood of Group I (9.761 ± 13.804 EU/mL-p:0.01). No differences for endotoxin occurred in portal blood. Conclusion -We developed a model of rats colonized with resistant bacteria useful to study intestinal translocation. Translocation occurred in surgical procedures with and without hepatic ischemia-reperfusion and probably occurred via the bloodstream. Translocation was probably lymphatic in the ischemia-reperfusion groups. Systemic blood endotoxin levels were higher in the group with complete hepatic ischemia.

Hsp90 inhibitors such as geldanamycin potently induce Hsp70 and reduce cytotoxicity due to α-synuclein expression, although their use has been limited due to toxicity, brain permeability, and drug design. We recently described the effects of a novel class of potent, small molecule Hsp90 inhibitors in cells overexpressing α-synuclein. Screening yielded several candidate compounds that significantly reduced α-synuclein oligomer formation and cytotoxicity associated with Hsp70 induction. In this study we examined whether chronic treatment with candidate Hsp90 inhibitors could protect against α-synuclein toxicity in a rat model of parkinsonism. Rats were injected unilaterally in the substantia nigra with AAV8 expressing human α-synuclein and then treated with drug for approximately 8 weeks by oral gavage. Chronic treatment with SNX-0723 or the more potent, SNX-9114 failed to reduce dopaminergic toxicity in the substantia nigra compared to vehicle. However, SNX-9114 significantly increased striatal dopamine content suggesting a positive neuromodulatory effect on striatal terminals. Treatment was generally well tolerated, but higher dose SNX-0723 (6-10 mg/kg) resulted in systemic toxicity, weight loss, and early death. Although still limited by potential toxicity, Hsp90 inhibitors tested herein demonstrate oral efficacy and possible beneficial effects on dopamine production in a vertebrate model of parkinsonism that warrant further study.

Dopaminergic nuclei in the basal ganglia are highly sensitive to damage from oxidative stress, inflammation, and environmental neurotoxins. Disruption of adenosine triphosphate (ATP)-dependent calcium (Ca2+) transients in astrocytes may represent an important target of such stressors that contributes to neuronal injury by disrupting critical Ca2+-dependent trophic functions. We therefore postulated that plasma membrane cation channels might be a common site of inhibition by structurally distinct cationic neurotoxicants that could modulate ATP-induced Ca2+ signals in astrocytes. To test this, we examined the capacity of two dopaminergic neurotoxicants to alter ATP-dependent Ca2+ waves and transients in primary murine striatal astrocytes: MPP+, the active metabolite of 1-methyl 4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), and 6-hydroxydopamine (6-OHDA). Both compounds acutely decreased ATP-induced Ca2+ transients and waves in astrocytes and blocked OAG-induced Ca2+ influx at micromolar concentrations, suggesting the transient receptor potential channel, TRPC3, as an acute target. MPP+ inhibited 1-oleoyl-2-acetyl-sn-glycerol (OAG)-induced Ca2+ transients similarly to the TRPC3 antagonist, pyrazole-3, whereas 6-OHDA only partly suppressed OAG-induced transients. RNAi directed against TRPC3 inhibited the ATP-induced transient as well as entry of extracellular Ca2+, which was augmented by MPP+. Whole-cell patch clamp experiments in primary astrocytes and TRPC3-overexpressing cells demonstrated that acute application of MPP+ completely blocked OAG-induced TRPC3 currents, whereas 6-OHDA only partially inhibited OAG currents. These findings indicate that MPP+ and 6-OHDA inhibit ATP-induced Ca2+ signals in astrocytes in part by interfering with purinergic receptor mediated activation of TRPC3, suggesting a novel pathway in glia that could contribute to neurotoxic injury.

BackgroundThe increasing occurrence of livestock-associated (LA) methicillin-resistant Staphylococcus aureus (MRSA) associated with the clonal complex (CC) 398 within the past years shows the importance of standardized and comparable typing methods for the purposes of molecular surveillance and outbreak detection. Multiple-locus variable number of tandem repeats analysis (MLVA) has recently been described as an alternative and highly discriminative tool for S. aureus. However, until now the applicability of MLVA for the typing of LA-MRSA isolates from different geographic origin has not been investigated in detail. We therefore compared MLVA and S. aureus protein A (spa) typing for characterizing porcine MRSA from distinct Dutch and German farms.Methodology/principal findingsOverall, 134 MRSA isolates originating from 21 different pig-farms in the Netherlands and 36 farms in Germany comprising 21 different spa types were subjected to MLVA-typing. Amplification and subsequent automated fragment sizing of the tandem repeat loci on a capillary sequencer differentiated these 134 isolates into 20 distinct MLVA types. Whereas overall MLVA and spa typing showed the same discriminatory power to type LA-MRSA (p = 0.102), MLVA was more discriminatory than spa typing for isolates associated with the prevalent spa types t011 and t034 (Simpson's Index of Diversity 0.564 vs. 0.429, respectively; pConclusionAlthough the applied MLVA scheme was not more discriminatory than spa typing in general, it added valuable information to spa typing results for specific spa types (t011, t034) which are highly prevalent in the study area, i.e. Dutch-German border area. Thus, both methods may complement each other to increase the discriminatory power to resolute highly conserved clones such as CC398 (spa types t011, t034) for the detection of outbreaks and molecular surveillance of zoonotic MRSA.

Multiple myeloma is a hematological cancer that is considered incurable despite advances in treatment strategy during the last decade. Therapies targeting single pathways are unlikely to succeed due to the heterogeneous nature of the malignancy. Proliferating cell nuclear antigen (PCNA) is a multifunctional protein essential for DNA replication and repair that is often overexpressed in cancer cells. Many proteins involved in the cellular stress response interact with PCNA through the five amino acid sequence AlkB homologue 2 PCNA-interacting motif (APIM). Thus inhibiting PCNA's protein interactions may be a good strategy to target multiple pathways simultaneously. We initially found that overexpression of peptides containing the APIM sequence increases the sensitivity of cancer cells to contemporary therapeutics. Here we have designed a cell-penetrating APIM-containing peptide, ATX-101, that targets PCNA and show that it has anti-myeloma activity. We found that ATX-101 induced apoptosis in multiple myeloma cell lines and primary cancer cells, while bone marrow stromal cells and primary healthy lymphocytes were much less sensitive. ATX-101-induced apoptosis was caspase-dependent and cell cycle phase-independent. ATX-101 also increased multiple myeloma cells' sensitivity against melphalan, a DNA damaging agent commonly used for treatment of multiple myeloma. In a xenograft mouse model, ATX-101 was well tolerated and increased the anti-tumor activity of melphalan. Therefore, targeting PCNA by ATX-101 may be a novel strategy in multiple myeloma treatment.

OBJECTIVES: To investigate to what extent cardiac MRI derived measurements of right ventricular (RV) volumes using the left ventricular (LV) end-systolic and end-diastolic frame misrepresent RV end-systolic and end-diastolic volumes in patients with tetralogy of Fallot (ToF) and a right bundle branch block. METHODS: Sixty-five cardiac MRI scans of patients with ToF and a right bundle branch block, and 50 cardiac MRI scans of control subjects were analyzed. RV volumes and function using the end-systolic and end-diastolic frame of the RV were compared to using the end-systolic and end-diastolic frame of the LV. RESULTS: Timing of the RV end-systolic frame was delayed compared to the LV end-systolic frame in 94% of patients with ToF and in 50% of control subjects. RV end-systolic volume using the RV end-systolic instead of LV end-systolic frame was smaller in ToF (median -3.3 ml/m(2), interquartile range -1.9 to -5.6 ml/m(2); p

Fms-like tyrosine kinase 3 ligand (Flt3L) is known as the primary differentiation and survival factor for dendritic cells (DCs). Furthermore, Flt3L is involved in the homeostatic feedback loop between DCs and regulatory T cell (Treg). We have previously shown that Flt3L accumulates in the synovial fluid in rheumatoid arthritis (RA) and that local exposure to Flt3L aggravates arthritis in mice, suggesting a possible involvement in RA pathogenesis. In the present study we investigated the role of Flt3L on DC populations, Tregs as well as inflammatory responses in experimental antigen-induced arthritis. Arthritis was induced in mBSA-immunized mice by local knee injection of mBSA and Flt3L was provided by daily intraperitoneal injections. Flow cytometry analysis of spleen and lymph nodes revealed an increased formation of DCs and subsequently Tregs in mice treated with Flt3L. Flt3L-treatment was also associated with a reduced production of mBSA specific antibodies and reduced levels of the pro-inflammatory cytokines IL-6 and TNF-α. Morphological evaluation of mBSA injected joints revealed reduced joint destruction in Flt3L treated mice. The role of DCs in mBSA arthritis was further challenged in an adoptive transfer experiment. Transfer of DCs in combination with T-cells from mBSA immunized mice, predisposed naïve recipients for arthritis and production of mBSA specific antibodies. We provide experimental evidence that Flt3L has potent immunoregulatory properties. Flt3L facilitates formation of Treg cells and by this mechanism reduces severity of antigen-induced arthritis in mice. We suggest that high systemic levels of Flt3L have potential to modulate autoreactivity and autoimmunity.

Survivin is known as an inhibitor of apoptosis and a positive regulator of cell division. We have recently identified survivin as a predictor of joint destruction in patients with rheumatoid arthritis (RA). Flt3 ligand (Flt3L) is expressed in the inflamed joints and has adjuvant properties in arthritis. Studies on 90 RA patients (median age 60.5 years [range, 24-87], disease duration 10.5 years [range, 0-35]) show a strong positive association between the levels of survivin and Flt3L in blood. Here, we present experimental evidence connecting survivin and Flt3L signaling. Treatment of BALB/c mice with Flt3L led to an increase of survivin in the bone marrow and in splenic dendritic cells. Flt3L changed the profile of survivin splice variants, increasing transcription of the short survivin40 in the bone marrow. Treatment with an Flt3 inhibitor reduced total survivin expression in bone marrow and in the dendritic cell population in spleen. Inhibition of survivin transcription in mice, by shRNA lentiviral constructs, reduced the gene expression of Flt3L. We conclude that expression of survivin is a downstream event of Flt3 signaling, which serves as an essential mechanism supporting survival of leukocytes during their differentiation, and maturation of dendritic cells, in RA.

Standard cancer cell lines do not model the intratumoural heterogeneity situation sufficiently. Clonal selection leads to a homogeneous population of cells by genetic drift. Heterogeneity of tumour cells, however, is particularly critical for therapeutically relevant studies, since it is a prerequisite for acquiring drug resistance and reoccurrence of tumours. Here, we report the isolation of a highly tumourigenic primary pancreatic cancer cell line, called JoPaca-1 and its detailed characterization at multiple levels. Implantation of as few as 100 JoPaca-1 cells into immunodeficient mice gave rise to tumours that were histologically very similar to the primary tumour. The high heterogeneity of JoPaca-1 was reflected by diverse cell morphology and a substantial number of chromosomal aberrations. Comparative whole-genome sequencing of JoPaca-1 and BxPC-3 revealed mutations in genes frequently altered in pancreatic cancer. Exceptionally high expression of cancer stem cell markers and a high clonogenic potential in vitro and in vivo was observed. All of these attributes make this cell line an extremely valuable model to study the biology of and pharmaceutical effects on pancreatic cancer.

Idiopathic pulmonary fibrosis (IPF) is a devastating and progressive lung disease. Its aetiology is thought to involve damage to the epithelium and abnormal repair. Alveolar epithelial cells near areas of remodelling show an increased expression of proapoptotic molecules. Therefore, we investigated the role of genes involved in cell cycle control in IPF. Genotypes for five single nucleotide polymorphisms (SNPs) in the tumour protein 53 (TP53) gene and four SNPs in cyclin-dependent kinase inhibitor 1A (CDKN1A), the gene encoding p21, were determined in 77 IPF patients and 353 controls. In peripheral blood mononuclear cells (PBMC) from 16 healthy controls mRNA expression of TP53 and CDKN1A was determined. Rs12951053 and rs12602273, in TP53, were significantly associated with survival in IPF patients. Carriers of a minor allele had a 4-year survival of 22% versus 57% in the non-carrier group (p = 0.006). Rs2395655 and rs733590, in CDKN1A, were associated with an increased risk of developing IPF. In addition, the rs2395655 G allele correlated with progression of the disease as it increased the risk of a rapid decline in lung function. Functional experiments showed that rs733590 correlated significantly with CDKN1A mRNA expression levels in healthy controls. This is the first study to show that genetic variations in the cell cycle genes encoding p53 and p21 are associated with IPF disease development and progression. These findings support the idea that cell cycle control plays a role in the pathology of IPF. Variations in TP53 and CDKN1A can impair the response to cell damage and increase the loss of alveolar epithelial cells.

Proliferating cell nuclear antigen (PCNA) is an essential protein for DNA replication, DNA repair, cell cycle regulation, chromatin remodeling, and epigenetics. Many proteins interact with PCNA through the PCNA interacting peptide (PIP)-box or the newly identified AlkB homolog 2 PCNA interacting motif (APIM). The xeroderma pigmentosum group A (XPA) protein, with a central but somewhat elusive role in nucleotide excision repair (NER), contains the APIM sequence suggesting an interaction with PCNA. With an in vivo based approach, using modern techniques in live human cells, we show that APIM in XPA is a functional PCNA interacting motif and that efficient NER of UV lesions is dependent on an intact APIM sequence in XPA. We show that XPA(-/-) cells complemented with XPA containing a mutated APIM sequence have increased UV sensitivity, reduced repair of cyclobutane pyrimidine dimers and (6-4) photoproducts, and are consequently more arrested in S phase as compared to XPA(-/-) cells complemented with wild type XPA. Notably, XPA colocalizes with PCNA in replication foci and is loaded on newly synthesized DNA in undamaged cells. In addition, the TFIIH subunit XPD, as well as XPF are loaded on DNA together with XPA, and XPC and XPG colocalize with PCNA in replication foci. Altogether, our results suggest a presence of the NER complex in the vicinity of the replisome and a novel role of NER in post-replicative repair.

We present an approach that integrates protein structure analysis and text mining for protein functional site prediction, called LEAP-FS (Literature Enhanced Automated Prediction of Functional Sites). The structure analysis was carried out using Dynamics Perturbation Analysis (DPA), which predicts functional sites at control points where interactions greatly perturb protein vibrations. The text mining extracts mentions of residues in the literature, and predicts that residues mentioned are functionally important. We assessed the significance of each of these methods by analyzing their performance in finding known functional sites (specifically, small-molecule binding sites and catalytic sites) in about 100,000 publicly available protein structures. The DPA predictions recapitulated many of the functional site annotations and preferentially recovered binding sites annotated as biologically relevant vs. those annotated as potentially spurious. The text-based predictions were also substantially supported by the functional site annotations: compared to other residues, residues mentioned in text were roughly six times more likely to be found in a functional site. The overlap of predictions with annotations improved when the text-based and structure-based methods agreed. Our analysis also yielded new high-quality predictions of many functional site residues that were not catalogued in the curated data sources we inspected. We conclude that both DPA and text mining independently provide valuable high-throughput protein functional site predictions, and that integrating the two methods using LEAP-FS further improves the quality of these predictions.

A number of studies of copy number imbalances (CNIs) in breast tumors support associations between individual CNIs and patient outcomes. However, no pattern or signature of CNIs has emerged for clinical use. We determined copy number (CN) gains and losses using high-density molecular inversion probe (MIP) arrays for 971 stage I/II breast tumors and applied a boosting strategy to fit hazards models for CN and recurrence, treating chromosomal segments in a dose-specific fashion (-1 [loss], 0 [no change] and +1 [gain]). The concordance index (C-Index) was used to compare prognostic accuracy between a training (n = 728) and test (n = 243) set and across models. Twelve novel prognostic CNIs were identified: losses at 1p12, 12q13.13, 13q12.3, 22q11, and Xp21, and gains at 2p11.1, 3q13.12, 10p11.21, 10q23.1, 11p15, 14q13.2-q13.3, and 17q21.33. In addition, seven CNIs previously implicated as prognostic markers were selected: losses at 8p22 and 16p11.2 and gains at 10p13, 11q13.5, 12p13, 20q13, and Xq28. For all breast cancers combined, the final full model including 19 CNIs, clinical covariates, and tumor marker-approximated subtypes (estrogen receptor [ER], progesterone receptor, ERBB2 amplification, and Ki67) significantly outperformed a model containing only clinical covariates and tumor subtypes (C-Index(full model), train[test] = 0.72[0.71] ± 0.02 vs. C-Index(clinical + subtype model), train[test] = 0.62[0.62] ± 0.02; p

BACKGROUND: Due to increasing numbers of people with diabetes taking part in extreme sports (e.g., high-altitude trekking), reliable handheld blood glucose meters (BGMs) are necessary. Accurate blood glucose measurement under extreme conditions is paramount for safe recreation at altitude. Prior studies reported bias in blood glucose measurements using different BGMs at high altitude. We hypothesized that glucose-oxidase based BGMs are more influenced by the lower atmospheric oxygen pressure at altitude than glucose dehydrogenase based BGMs. METHODOLOGY/PRINCIPAL FINDINGS: Glucose measurements at simulated altitude of nine BGMs (six glucose dehydrogenase and three glucose oxidase BGMs) were compared to glucose measurement on a similar BGM at sea level and to a laboratory glucose reference method. Venous blood samples of four different glucose levels were used. Moreover, two glucose oxidase and two glucose dehydrogenase based BGMs were evaluated at different altitudes on Mount Kilimanjaro. Accuracy criteria were set at a bias 6.5 mmol/L) and

BACKGROUND:Despite low incidence, ovarian cancer is the fifth leading cause of cancer deaths and it has the highest mortality rate of all gynecologic malignancies among US women. The mortality rate would be reduced with an early detection marker. The folate receptor alpha (FRalpha) is one logical choice for a biomarker because of its prevalent overexpression in ovarian cancer and its exclusive expression in only a few normal tissues. In prior work, it was observed that patients with ovarian cancer had elevated serum levels of a protein that bound to a FRalpha-specific monoclonal antibody relative to healthy individuals. However, it was not shown that the protein detected was intact functional FRalpha. In the current study, the goal was to determine whether ovarian cancer patients (n = 30) had elevated serum levels of a fully functional intact FRalpha compared to matched healthy controls (n = 30). METHODOLOGY/PRINCIPAL FINDINGS:FRalpha levels in serum were analyzed by two methods, immunoblotting analysis and a radiolabeled folic acid-based microfiltration binding assay. Using the immunoassay, we observed that levels of FRalpha were higher in serum of ovarian cancer patients as compared to controls. Similar results were also observed using the microfiltration binding assay, which showed that the circulating FRalpha is functional. Importantly, we also found that the levels of FRalpha were comparable between early and advanced stage patients. CONCLUSIONS:Our results demonstrate that ovarian cancer patients have elevated levels of functional intact FRalpha. These findings support the potential use of circulating FRalpha as a biomarker of early ovarian cancer.

OBJECTIVE: Comparative evaluation of costs and effects of laparoscopic hysterectomy (LH) and abdominal hysterectomy (AH). DATA SOURCES: Controlled trials from Cochrane Central register of controlled trials, Medline, Embase and prospective trial registers. SELECTION OF STUDIES: Twelve (randomized) controlled studies including the search terms costs, laparoscopy, laparotomy and hysterectomy were identified. METHODS: The type of cost analysis, perspective of cost analyses and separate cost components were assessed. The direct and indirect costs were extracted from the original studies. For the cost estimation, hospital stay and procedure costs were selected as most important cost drivers. As main outcome the major complication rate was taken. FINDINGS: Analysis was performed on 2226 patients, of which 1013 (45.5%) in the LH group and 1213 (54.5%) in the AH group. Five studies scored > or =10 points (out of 19) for methodological quality. The reported total direct costs in the LH group ($63,997) were 6.1% higher than the AH group ($60,114). The reported total indirect costs of the LH group ($1,609) were half of the total indirect in the AH group ($3,139). The estimated mean major complication rate in the LH group (14.3%) was lower than in the AH group (15.9%). The estimated total costs in the LH group were $3,884 versus $3,312 in the AH group. The incremental costs for reducing one patient with major complication(s) in the LH group compared to the AH group was $35,750. CONCLUSIONS: The shorter hospital stay in the LH group compensates for the increased procedure costs, with less morbidity. LH points in the direction of cost effectiveness, however further research is warranted with a broader costs perspective including long term effects as societal benefit, quality of life and survival.