Utility of a simplified ultrasound assessment to assess interstitial pulmonary fibrosis in connective tissue disorders - preliminary results

Introduction  

Interstitial pulmonary fibrosis (IPF) is a frequent manifestation in patients with connective tissue disorders (CTD). Recently the ultrasound (US) criterion validity for its assessment has been proposed; however, the US scoring systems adopted include the study of several lung intercostal spaces (LIS), which could be time-consuming in daily clinical practice. The aim of this study was to investigate the utility of a simplified US B-lines scoring system compared with both the US comprehensive assessment and the high-resolution computed tomography (HRCT) findings of IPF in CTD patients.     

Applying selected reaction monitoring to targeted proteomics

Selected reaction monitoring (SRM) is a highly selective and sensitive mass spectrometric methodology for precise and accurate quantification of low-abundant proteins in complex mixtures and for characterization of modified peptides, and constitutes the method of choice in targeted proteomics. Owing to its outstanding features, SRM arises as an alternative to antibody-based assays for discovery and validation of clinically relevant biomarkers, a topic that is tackled in this article. Several of the obstacles encountered in SRM experiments, mainly those derived from shared physicochemical properties of peptides (e.g., mass, charge and chromatographic retention time), can compromise selectivity and quantitation. We illustrate how to circumvent these limitations on the basis of using time-scheduled chromatographic approaches and choosing appropriate spectrometric conditions, including the careful selection of the precursor and diagnostic ions.     

Development of a classification and ranking method for the identification of possible biomarkers in two-dimensional gel-electrophoresis based on principal component analysis and variable selection procedures

The identification of biomarkers is one of the leading research areas in proteomics. When biomarkers have to be searched for in spot volume datasets produced by 2D gel-electrophoresis, problems may arise related to the large number of spots present in each map and the small number of samples available in each class (control/pathological). In such cases multivariate methods are usually exploited together with variable selection procedures, to provide a set of possible biomarkers: they are however usually aimed to the selection of the smallest set of variables (spots) providing the best performances in prediction. This approach seems not to be suitable for the identification of potential biomarkers since in this case all the possible candidate biomarkers have to be identified to provide a general picture of the "pathological state": in this case exhaustivity has to be preferred to provide a complete understanding of the mechanisms underlying the pathology. We propose here a ranking and classification method, "Ranking-PCA", based on Principal Component Analysis and variable selection in forward search: the method selects one variable at a time as the one providing the best separation of the two classes investigated in the space given by the relevant PCs. The method was applied to an artificial dataset and a real case-study: Ranking-PCA exhaustively identified the potential biomarkers and provided reliable and robust results.     

Foraging activity of bumblebees (Bombus terrestris L.) on Bt-expressing eggplants

A greenhouse experiment was setup to study foraging behavior of the bumblebee Bombus terrestris L. on Cry3Bb-expressing genetically modified (GM) eggplants and their near-isogenic control. Commonly, more bumblebees visited GM eggplants compared to near-isogenic control, but this difference was only marginally significant. The mean length of feeding bouts was similar on the two treatments. Neither the number of flowers produced nor their size could explain bumblebees?? tendency to prefer GM eggplants. Volatile compounds were extracted from five plants per genotype and separated using gas chromatography. Thirteen compounds were identified and five of them appeared significantly more abundant in GM eggplants. Six of the identified compounds [(+)-limonene, Z-jasmone, p-cymene, ??-pinene, methyl-salicilate, and (?)-limonene] were tested in electrophysiological bioassays with antennas detached from young bumblebees, and a response was recorded in all six cases. Experimental results indicate that pollination activity of bumblebees is compatible with this GM eggplant event as a food source and that chemical cues may have an important role in plant identification. The implications for environmental risk assessment of GM plants are discussed.     

Survival, development, and gonadal differentiation in Rana dalmatina chronically exposed to chlorpyrifos

Chlorpyrifos is an organophosphate pesticide among the most widely used in the world, which is suspected to be an endocrine-disrupting compound. To determine the capacity of chlorpyrifos to affect gonadal differentiation in Rana dalmatina, tadpoles were exposed to this pesticide during larval development until completion of metamorphosis at ecologically relevant concentrations (0.025 and 0.05 mg/L). No effects of chlorpyrifos exposure on survival, development, or metamorphosis were observed. After a 1 month metamorphosis, the gonadal phenotype was determined by gross morphology and histological examination. Morphological and histological analysis revealed normal ovaries or testes in froglets belonging to control group, whereas testes from several froglets exposed to chlorpyrifos were interspersed with testicular oocytes in histological sections. Chlorpyrifos exposure during the entire larval period did not affect sex ratio, but reduced the percentage of males with histologically normal testes. The findings suggest that chlorpyrifos exposure has significant effects on gonadal differentiation in some animals by inducing an intersex condition and alterations to testicular morphology, and that R. dalmatina is sensitive to endocrine disruption. Thereby, this study provides evidence that the ecologically relevant concentrations of chlorpyrifos, although not adversely affect the survival, development, or metamorphosis, may interfere with sex differentiation and reproductive development of R. dalmatina via endocrine-disrupting mechanisms.     

Mesenchymal stromal cells primed with paclitaxel provide a new approach for cancer therapy

Background

Mesenchymal stromal cells may represent an ideal candidate to deliver anti-cancer drugs. In a previous study, we demonstrated that exposure of mouse bone marrow derived stromal cells to Doxorubicin led them to acquire anti-proliferative potential towards co-cultured haematopoietic stem cells (HSCs). We thus hypothesized whether freshly isolated human bone marrow Mesenchymal stem cells (hMSCs) and mature murine stromal cells (SR4987 line) primed in vitro with anti-cancer drugs and then localized near cancer cells, could inhibit proliferation.

Methods and Principal Findings

Paclitaxel (PTX) was used to prime culture of hMSCs and SR4987. Incorporation of PTX into hMSCs was studied by using FICT-labelled-PTX and analyzed by FACS and confocal microscopy. Release of PTX in culture medium by PTX primed hMSCs (hMSCsPTX) was investigated by HPLC. Culture of Endothelial cells (ECs) and aorta ring assay were used to test the anti-angiogenic activity of hMSCsPTX and PTX primed SR4987(SR4987PTX), while anti-tumor activity was tested in vitro on the proliferation of different tumor cell lines and in vivo by co-transplanting hMSCsPTX and SR4987PTX with cancer cells in mice. Nevertheless, despite a loss of cells due to chemo-induced apoptosis, both hMSCs and SR4987 were able to rapidly incorporate PTX and could slowly release PTX in the culture medium in a time dependent manner. PTX primed cells acquired a potent anti-tumor and anti-angiogenic activity in vitro that was dose dependent, and demonstrable by using their conditioned medium or by co-culture assay. Finally, hMSCsPTX and SR4987PTX co-injected with human cancer cells (DU145 and U87MG) and mouse melanoma cells (B16) in immunodeficient and in syngenic mice significantly delayed tumor takes and reduced tumor growth.

Conclusions

These data demonstrate, for the first time, that without any genetic manipulation, mesenchymal stromal cells can uptake and subsequently slowly release PTX. This may lead to potential new tools to increase efficacy of cancer therapy.     

Utility of the tourniquet test and the white blood cell count to differentiate dengue among acute febrile illnesses in the emergency room

Dengue often presents with non-specific clinical signs, and given the current paucity of accurate, rapid diagnostic laboratory tests, identifying easily obtainable bedside markers of dengue remains a priority. Previous studies in febrile Asian children have suggested that the combination of a positive tourniquet test (TT) and leucopenia can distinguish dengue from other febrile illnesses, but little data exists on the usefulness of these tests in adults or in the Americas. We evaluated the diagnostic accuracy of the TT and leucopenia (white blood cell count <5000/mm(3)) in identifying dengue as part of an acute febrile illness (AFI) surveillance study conducted in the Emergency Department of Saint Luke's Hospital in Ponce, Puerto Rico. From September to December 2009, 284 patients presenting to the ED with fever for 2-7 days and no identified source were enrolled. Participants were tested for influenza, dengue, leptospirosis and enteroviruses. Thirty-three (12%) patients were confirmed as having dengue; 2 had dengue co-infection with influenza and leptospirosis, respectively. An infectious etiology was determined for 141 others (136 influenza, 3 enterovirus, 2 urinary tract infections), and 110 patients had no infectious etiology identified. Fifty-two percent of laboratory-positive dengue cases had a positive TT versus 18% of patients without dengue (P<0.001), 87% of dengue cases compared to 28% of non-dengue cases had leucopenia (P<0.001). The presence of either a positive TT or leucopenia correctly identified 94% of dengue patients. The specificity and positive predictive values of these tests was significantly higher in the subset of patients without pandemic influenza A H1N1, suggesting improved discriminatory performance of these tests in the absence of concurrent dengue and influenza outbreaks. However, even during simultaneous AFI outbreaks, the absence of leucopenia combined with a negative tourniquet test may be useful to rule out dengue.