Tumourigenic non-small-cell lung cancer mesenchymal circulating tumour cells: a clinical case study.

BACKGROUND: Over the past decade, numerous reports describe the generation and increasing utility of non-small-cell lung cancer (NSCLC) patient-derived xenografts (PDX) from tissue biopsies. While PDX have proven useful for genetic profiling and preclinical drug testing, the requirement of a tissue biopsy limits the available patient population, particularly those with advanced oligometastatic disease. Conversely, 'liquid biopsies' such as circulating tumour cells (CTCs) are minimally invasive and easier to obtain. Here, we present a clinical case study of a NSCLC patient with advanced metastatic disease, a never smoker whose primary tumour was EGFR and ALK wild-type. We demonstrate for the first time, tumorigenicity of their CTCs to generate a patient CTC-derived eXplant (CDX). PATIENTS AND METHODS: CTCs were enriched at diagnosis and again 2 months later during disease progression from 10 ml blood from a 48-year-old NSCLC patient and implanted into immunocompromised mice. Resultant tumours were morphologically, immunohistochemically, and genetically compared with the donor patient's diagnostic specimen. Mice were treated with cisplatin and pemetrexed to assess preclinical efficacy of the chemotherapy regimen given to the donor patient. RESULTS: The NSCLC CDX expressed lung lineage markers TTF1 and CK7 and was unresponsive to cisplatin and pemetrexed. Examination of blood samples matched to that used for CDX generation revealed absence of CTCs using the CellSearch EpCAM-dependent platform, whereas size-based CTC enrichment revealed abundant heterogeneous CTCs of which ∼80% were mesenchymal marker vimentin positive. Molecular analysis of the CDX, mesenchymal and epithelial CTCs revealed a common somatic mutation confirming tumour origin and showed CDX RNA and protein profiles consistent with the predominantly mesenchymal phenotype. CONCLUSIONS: This study shows that the absence of NSCLC CTCs detected by CellSearch (EpCAM(+)) does not preclude CDX generation, highlighting epithelial to mesenchymal transition and the functional importance of mesenchymal CTCs in dissemination of this disease.

[The Lymphoid Variant of HES (L-HES) as Differential Diagnose of Severe Asthma in Childhood]. / Ein T-Zell assoziiertes Hypereosinophilie-Syndrom (L-HES) als Differenzialdiagnose eines Asthma bronchiale.

Based on a case report an overview on the differential diagnostic considerations with respect to blood hypereosinophilia (HE) and hypereosinophilic syndromes (HES) in childhood is given. A 13-year-old boy was admitted for the clarification of an asthma. In the blood count an increased HE with 3 500/µl (30%) was found along with elevated total serum IgE and IL-5 level (2 000 IU/ml and 17 pg/ml). Lung function showed an obstruction (FEV1 38%). Radiologically the picture of bronchiectasis and mucus pluggine appeared. In the BAL a HE (76%) with raised IL-5 level was apparent. Histologically asthma was diagnosed with mucostasis, hypertrophy of the bronchial wall musculature and a lung HE. Differential-diagnostically an ABPA, a Churg-Strauss-Syndrome, a parasitosis, drug associated HE, allergies and malignant disease could be excluded. An aberrant T-cell clone in peripheral blood was detected by flow cytometry and T-cell receptor clonal rearrangements by PCR, leading to the diagnosis of a lymphoid variant of HES (L-HES). Failure to detect the FIP1L1-PDGFRA gene fusion and a normal bone marrow examination could exclude a neoplastic HES (HESN). After steroid initiation, prompt decrease of blood eosinophilia with resolution of symptoms was observed. Steroid discontinuation led to eosinophilia recurrence associated with disease symptoms. As steroid-sparing agent the immunosuppressive azathioprine was additionally given; steroid doses could be decreased and stopped in the course. This case demonstrated the range of HE evaluation in infancy. With asthma one should also consider the possibility of a L-HES.

Weight-loss strategies used by baby boomer men: a mixed methods approach.

BACKGROUND: Baby Boomer men (those born in 1946-1964) are becoming obese at an earlier age compared to previous generations. The present study aimed to identify weight-loss strategies used by Baby Boomer men, to determine whether those strategies varied by weight status and to explore their dieting experiences. METHODS: The study used a cross-sectional survey of 211 men and 20 in-depth interviews. RESULTS: Men had a mean (SD) body mass index (BMI) of 29.35 (5.07) kg m(-2), with 82% being overweight or obese. Fifty-six percent were currently trying to lose weight. Healthy weight-loss strategies included reducing portions, increasing physical activity, cutting back on fried foods, cutting back on sweets, cutting back on alcohol, using meal replacement drinks/bars and joining a weight-loss programme. Unhealthy strategies included skipping meals and using over-the-counter 'diet pills'. Men who reduced portions, skipped meals, cut back on sweets, joined a weight-loss programme and used diet pills had significantly higher BMIs than men who said they did not (P < 0.05 for all). Interviews revealed that older men struggle to lose weight, 'I've been struggling for the last 2-3 years'. 'The last time I really tried to lose weight I stayed on the diet for just a day or two'. Wives were considered essential to their weight management success. CONCLUSIONS: Men used a do-it-yourself weight-loss approach using both healthy and unhealthy strategies. Obese men were more likely to use unhealthy practices than overweight men.

Neighbourhood species richness and drought-tolerance traits modulate tree growth and δ13 C responses to drought.

Mixed-species forests are promoted as a forest management strategy for climate change adaptation, but whether they are more resistant to drought than monospecific forests remains contested. In particular, the trait-based mechanisms driving the role of tree diversity under drought remain elusive. Using tree cores from a large-scale biodiversity experiment, we investigated tree growth and physiological stress responses (i.e. increase in wood carbon isotopic ratio; δ13 C) to changes in climate-induced water availability (wet to dry years) along gradients in neighbourhood tree species richness and drought-tolerance traits. We hypothesized that neighbourhood species richness increases growth and decreases δ13 C and that these relationships are modulated by the abiotic (i.e. climatic conditions) and the biotic context. We characterised the biotic context using drought-tolerance traits of focal trees and their neighbours. These traits are related to cavitation resistance versus resource acquisition and stomatal control. Tree growth increased with neighbourhood species richness. However, we did not observe a universal relief of water stress in species-rich neighbourhoods. The effects of neighbourhood species richness and climate on growth and δ13 C were modulated by the traits of focal trees and the traits of their neighbours. At either end of each drought-tolerance gradient, species responded in opposing directions during dry and wet years. We show that species' drought-tolerance traits can explain the strength and nature of biodiversity-ecosystem functioning relationships in experimental tree communities experiencing drought. Mixing tree species can increase growth but may not universally relieve drought stress.

Native human adipose stromal cells: localization, morphology and phenotype.

OBJECTIVES: Beside having roles in energy homeostasis and endocrine modulation, adipose tissue (AT) is now considered a promising source of mesenchymal stromal cells (adipose-derived stromal cells or ASCs) for regenerative medicine. Despite numerous studies on cultured ASCs, native human ASCs are rarely investigated. Indeed, the phenotype of ASCs in their native state, their localization within AT and comparison with bone marrow-derived mesenchymal stromal cells (BM-MSCs) has been poorly investigated. DESIGN: To address these issues, the stroma vascular fraction (SVF) of human AT was extracted and native cell subtypes were isolated by immunoselection to study their clonogenic potential in culture. Immunohistology on samples of human AT in combination with reconstruction of confocal sections were performed in order to localize ASCs. RESULTS: Compared with BM-MNCs, all native ASCs were found in the CD34(+) cell fraction of the AT-SVF. Native ASCs expressed classical mesenchymal markers described for BM-MSCs. Interestingly, CD34 expression decreased during ASC cell culture and was negatively correlated with cell proliferation rate. Immunohistological analysis revealed that native ASCs exhibited specific morphological features with protrusions. They were found scattered in AT stroma and did not express in vivo pericytic markers such as NG2, CD140b or alpha-smooth muscle actin, which appeared during the culture process. Finally, ASCs spontaneous commitment to adipocytic lineage was enhanced in AT from obese humans. CONCLUSIONS: The use of complementary methodological approaches to study native human ASCs revealed their immunophenotype, their specific morphology, their location within AT and their stemness. Furthermore, our data strongly suggest that human ASCs participate in adipogenesis during AT development.

High power narrow-band fiber-based ASE source.

In this paper we describe a high power narrow-band amplified spontaneous emission (ASE) light source at 1030 nm center wavelength generated in an Yb-doped fiber-based experimental setup. By cutting a small region out of a broadband ASE spectrum using two fiber Bragg gratings a strongly constrained bandwidth of 12±2 pm (3.5±0.6 GHz) is formed. A two-stage high power fiber amplifier system is used to boost the output power up to 697 W with a measured beam quality of M2≤1.34. In an additional experiment we demonstrate a stimulated Brillouin scattering (SBS) suppression of at least 17 dB (theoretically predicted ~20 dB), which is only limited by the dynamic range of the measurement and not by the onset of SBS when using the described light source. The presented narrow-band ASE source could be of great interest for brightness scaling applications by beam combination, where SBS is known as a limiting factor.

Advances in transfusion medicine in the first decade of the 21st century: Advances in miniaturized technologies.

Several miniaturized high throughput technologies have been developed in the last decade, primarily to study genomic structures and gene expression patterns under various conditions. At the same time, the microarrays, biosensors, integrated microfluidic lab-on-a-chip devices, next generation sequencing or digital PCR are gradually finding their diagnostic applications, although their suitability for specialised diagnostic fields has still to be assessed. In this review we discuss the potential applications of the new technologies to blood testing.

Whole-body MRI in the childhood form of hypophosphatasia.

Hypophosphatasia (HPP) is a rare inborn error of bone metabolism caused by various defects in the gene coding for the tissue-nonspecific alkaline phosphatase (TNSAP). It results in a reduced activity of the TNSAP and elevated concentrations of its substrates, including inorganic pyrophosphate. Clinical features of HPP include defective bone mineralisation with bone deformities, fractures and chronic non-bacterial osteomyelitis. Renal damage due to calcification, craniosynostosis and dental abnormalities with premature loss of dentition are further complications. Until now, detailed descriptions of whole-body magnetic resonance imaging (WB-MRI) in HPP do not exist. Here, we analysed WB-MRIs of 4 children with the childhood form of HPP. Deformities and defects of the long bones could be seen. All patients showed radiological lesions in the metaphyses of the long bones predominantly in the lower extremities being consistent with hyperaemia and oedema. Differential diagnosis includes an inflammatory process being active in these locations.

The influence of climate and fructification on the inter-annual variability of stem growth and net primary productivity in an old-growth, mixed beech forest.

The periodic production of large seed crops by trees (masting) and its interaction with stem growth has long been the objective of tree physiology research. However, very little is known about the effects of masting on stem growth and total net primary productivity (NPP) at the stand scale. This study was conducted in an old-growth, mixed deciduous forest dominated by Fagus sylvatica (L.) and covers the period from 2003 to 2007, which comprised wet, dry and regular years as well as two masts of Fagus and one mast of the co-dominant tree species Fraxinus excelsior (L.) and Acer pseudoplatanus (L.). We combined analyses of weather conditions and stem growth at the tree level (inter- and intra-annual) with fruit, stem and leaf production, and estimates of total NPP at the stand level. Finally, we compared the annual demand of carbon for biomass production with net canopy assimilation (NCA), derived from eddy covariance flux measurements, chamber measurements and modelling. Annual stem growth of Fagus was most favoured by warm periods in spring and that of Fraxinus by high precipitation in June. For stem growth of Acer and for fruit production, no significant relationships with mean weather conditions were found. Intra-annual stem growth of all species was strongly reduced when the relative plant-available water in soil dropped below a threshold of about 60% between May and July. The inter-annual variations of NCA, total NPP and leaf NPP at the stand level were low (mean values 1313, 662 and 168 g C m(-2) year(-1), respectively), while wood and fruit production varied more and contrarily (wood: 169-241 g C m(-2) year(-1); fruits: 21-142 g C m(-2) year(-1)). In all years, an annual surplus of newly assimilated carbon was calculated (on average 100 g C m(-2) year(-1)). The results suggest that stem growth is generally not limited by insufficient carbon resources; only in mast years a short-term carbon shortage may occur in spring. In contrast to common assumption, stem growth alone is not a sufficient proxy for total biomass production or the control of carbon sequestration by weather extremes.

Spectral beam combination of fiber amplified ns-pulses by means of interference filters.

In this paper we introduce a simple scheme to spectrally combine four single beams using three low-cost dielectric interference filters as combining elements. 25 ns pulses from four independent and actively Q-switched fiber seed-sources are amplified in a single stage fiber-amplifier. Temporally and spatially combined 208 W of average power and 6.3 mJ of pulse energy are obtained at two different repetition frequencies. A detailed analysis of beam quality as well as the thermal behavior of the combining elements is carried out and reveals mutual dependency.

2 kW incoherent beam combining of four narrow-linewidth photonic crystal fiber amplifiers.

We report on beam combining of four narrow-linewidth fiber amplifier chains, running at different wavelengths and each delivering 500 W optical output power. The main amplifier stage consists of a large mode area photonic crystal fiber. The four output beams of the amplifier chains are spectrally (incoherent) combined using a polarization-independent dielectric reflective diffraction grating to form an output beam of 2 kW continuous-wave optical power with good beam quality (M(2)x = 2.0, M(2)y = 1.8).

Climate change. Managing forests after Kyoto.

The Kyoto protocol aims to reduce carbon emissions into the atmosphere. Part of the strategy is the active management of terrestrial carbon sinks, principally through afforestation and reforestation. In their Perspective, Schulze et al. argue that the preservation of old-growth forests may have a larger positive effect on the carbon cycle than promotion of regrowth.

An alternative treatment method to restore limb-length discrepancy in osteoarthritis with high congenital hip dislocation.

AIM: The purpose of this study was to describe the rationale and to analyse the results of uncemented total hip arthroplasty (THA) in patients with osteoarthritis and congenital dislocation of the hip (CDH) prior to iliofemoral distraction. METHOD: Twenty hips were followed-up for 5.8 years (1-12 years). First the THA components were implanted after soft tissue release. In the interval, iliofemoral distraction with an external distraction apparatus was conducted. The second surgery consisted of an open reduction. An iliofemoral distraction of 0.8 mm/day was monitored. The treatment time was 86 days (50-210 days). In 61 days (32-94 days) the hip joints were distracted for 51 mm (41-75 mm). RESULT: The Harris Hip Score increased significantly by 47 points. The SF-36 health score showed a satisfying increase in all patients. CONCLUSION: In experienced hands this method is a relatively safe procedure for limb-length equalization in patients with severe CDH.

Towards error categorisation in BCI: single-trial EEG classification between different errors.

OBJECTIVE: Error-related potentials (ErrP) are generated in the brain when humans perceive errors. These ErrP signals can be used to classify actions as erroneous or non-erroneous, using single-trial electroencephalography (EEG). A small number of studies have demonstrated the feasibility of using ErrP detection as feedback for reinforcement-learning-based brain-computer interfaces (BCI), confirming the possibility of developing more autonomous BCI. These systems could be made more efficient with specific information about the type of error that occurred. A few studies differentiated the ErrP of different errors from each other, based on direction or severity. However, errors cannot always be categorised in these ways. We aimed to investigate the feasibility of differentiating very similar error conditions from each other, in the absence of previously explored metrics. APPROACH: In this study, we used two data sets with 25 and 14 participants to investigate the differences between errors. The two error conditions in each task were similar in terms of severity, direction and visual processing. The only notable differences between them were the varying cognitive processes involved in perceiving the errors, and differing contexts in which the errors occurred. We used a linear classifier with a small feature set to differentiate the errors on a single-trial basis. MAIN RESULTS: For both data sets, we observed neurophysiological distinctions between the ErrPs related to each error type. We found further distinctions between age groups. Furthermore, we achieved statistically significant single-trial classification rates for most participants included in the classification phase, with mean overall accuracy of 65.2% and 65.6% for the two tasks. SIGNIFICANCE: As a proof of concept our results showed that it is feasible, using single-trial EEG, to classify these similar error types against each other. This study paves the way for more detailed and efficient learning in BCI, and thus for a more autonomous human-machine interaction.

Direct determination of intracellular daunorubicin in intact confluent monolayers of AT1 prostate carcinoma cells using a multiwell-multilabel counter.

The cytostatic drug daunorubicin exerts its toxic action by intercalating into the DNA. The efficacy of daunorubicin depends on the intracellular amount in the tumor cell. Here we have evaluated the use of a multiwell-multilabel reader for the direct determination of the fluorescent cytostatic drug daunorubicin in a prostate carcinoma cell line (AT1 R-3327 Dunning prostate carcinoma cells) grown on 24-well plates. We present evidence that this simple fluorescent parameter is a good measure for the toxicologically relevant amount of the drug intercalated into the DNA and, therefore, is a good predictor for the drug's cytotoxicity. The amount of cationic cytostatics in a tumor cell is primarily a function of the efflux pump protein p-gycoprotein (pGP). Therefore, it is of great value that the assay is also suitable for the estimation of the multidrug resistance efflux pump (pGP) activity.

Numerical investigations of stress shielding in total hip prostheses.

Aseptic loosening of the prosthesis is still a problem in artificial joint implants. The loosening can be caused by, among other factors, resorption of the bone surrounding the prosthesis owing to stress shielding. In order to find out the influence of the prosthesis type on post-operative stress shielding, a static finite element analysis of a femur provided with the conventional uncemented stem BICONTACT and of one with the femoral neck prosthesis SPIRON was carried out. Strain energy densities and maximal principal strain distributions were calculated and compared with the physiological situation. Here, stress shielding was demonstrated in both periprosthetic femora. To determine the areas of the stress shielding, the bone in each FE model was subdivided into three regions of interest (ROI): proximal, diaphyseal, and distal. The numerical computations show stress shielding in the proximal ROI of both periprosthetic femora. Diaphyseally, the femoral neck prosthesis SPIRON, in contrast to the conventional uncemented long-stem prosthesis BICONTACT, causes no decrease in the strain distribution and thus no stress shielding. Distally, no change in the load distribution of either periprosthetic femur could be found, compared with the physiological situation.

[Radiologic evaluation of cup placement variation in conventional total hip arthroplasty]. / Radiologische Evaluation der Variationsbreite der Pfannenpositionierung bei konventioneller Hüftendoprothesenversorgung.

PURPOSE: Determination of the amount of variation in conventional acetabular cup positioning (radiological inclination and anteversion) in view of different factors that could influence the measured angles. MATERIALS AND METHODS: The intended acetabular cup position of 45 degrees inclination and 20 degrees anteversion was checked radiologically in 950 patients who received a cementless total hip replacement. The qualifications of the surgeon, operated side and implant model were recorded and analyzed with respect to a possible correlation with the results. Since a "safe angle" cannot be defined without consideration of other variables, a tolerable deviation of the target position was investigated. RESULTS: The mean inclination angle was 48.7 degrees (SD 7 degrees, minimum 28 degrees, maximum 75 degrees ). Anteversion was measured with a mean of 18.6 degrees (SD 9 degrees, minimum -9 degrees, maximum 50 degrees ). Assuming an acceptable deviation of +/- 5 degrees from the target position (45 degrees inclination and 20 degrees anteversion), only 22.7 % of the acetabular cups were in this range. In the case of an acceptable deviation of +/- 10 degrees, 34.5 % of the cups were still outside of the acceptable range. The qualifications of the surgeon, the implanted model, as well as the operated side did not have a significant influence on the results. CONCLUSION: The common implantation technique yielded significant variation with respect to anteversion and inclination. The application of computer-aided navigation in the placement of acetabular cups would help to improve accuracy and reproducibility considerably in total hip arthroplasty.

Fitting new technologies into the safety paradigm: use of microarrays in transfusion.

Until the late 1990s, mandatory blood screening for transmissible infectious agents depended entirely on antigen/antibody-based detection assays. The recent emergence of Nucleic acid Amplification Technologies (NAT) has revolutionised viral diagnosis, not only by increasing the level of sensitivity but also by facilitating the detection of several viruses in parallel by multiplexing specific primers. In more complex biological situations, when a broad spectrum of pathogens must be screened, the limitations of these first generation technologies became apparent. High throughput systems, such as DNA Arrays, permit a conceptually new approach. These miniaturised micro systems allow the detection of hundreds of different targets simultaneously, inducing a dramatic decrease in reagent consumption, a reduction in the number of confirmation tests and a simplification of data interpretation. However, the systems currently available require additional instrumentation and reagents for sample preparation and target amplification prior to detection on the DNA array. A major challenge in the area of DNA detection is the development of methods that do not rely on target amplification systems. Likewise, the advances of protein microarrays have lagged because of poor stability of proteins, complex coupling chemistry and weak detection signals. Emerging technologies like Biosensors and nano-particle based DNA or Protein Bio-Barcode Amplification Assays are promising diagnostic tools for a wide range of clinical applications, including blood donation screening.

Chronic symptomatic proximal tibiofibular instability in a 3-year-old girl.

A 3-year-old girl was presented with recurrent pain at the right knee and a "snapping" when walking. A free range of motion of both knee joints, stable cruciate and collateral ligaments and a ventrally subluxating and snapping fibula when flexing the knee were found. Due to a longer course of recurrent subluxation and pain, an operation was performed and the tibiofibular ligaments and joint capsule were tightened by sutures. Other operative techniques proposed in the literature are discussed; most of them could lead to deformities or growth disturbances. The reports of tibiofibular instability in the literature are presented. The girl is the youngest patient with idiopathic proximal tibiofibular instability reported in the literature at least in the last 75 years.