Zonal image analysis of tumour vascular perfusion, hypoxia, and necrosis

A number of laboratories are utilising both hypoxia and perfusion markers to spatially quantify tumour oxygenation and vascular distributions, and scientists are increasingly turning to automated image analysis methods to quantify such interrelationships. In these studies, the presence of regions of necrosis in the immunohistochemical sections remains a potentially significant source of error. In the present work, frozen MCa-4 mammary tumour sections were used to obtain a series of corresponding image montages. Total vessels were identified using CD31 staining, perfused vessels by DiOC7 staining, hypoxia by EF5/Cy3 uptake, and necrosis by haematoxylin and eosin staining. Our goal was to utilise image analysis techniques to spatially quantitate hypoxic marker binding as a function of distance from the nearest blood vessel. Several refinements to previous imaging methods are described: (1) hypoxia marker images are quantified in terms of their intensity levels, thus providing an analysis of the gradients in hypoxia with increasing distances from blood vessels, (2) zonal imaging masks are derived, which permit spatial sampling of images at precisely defined distances from blood vessels, as well as the omission of necrotic artifacts, (3) thresholding techniques are applied to omit holes in the tissue sections, and (4) distance mapping is utilised to define vascular spacing

Using Bayesian networks to guide the assessment of new evidence in an appeal case.

When new forensic evidence becomes available after a conviction there is no systematic framework to help lawyers to determine whether it raises sufficient questions about the verdict in order to launch an appeal. This paper presents such a framework driven by a recent case, in which a defendant was convicted primarily on the basis of audio evidence, but where subsequent analysis of the evidence revealed additional sounds that were not considered during the trial. The framework is intended to overcome the gap between what is generally known from scientific analyses and what is hypothesized in a legal setting. It is based on Bayesian networks (BNs) which have the potential to be a structured and understandable way to evaluate the evidence in a specific case context. However, BN methods suffered a setback with regards to the use in court due to the confusing way they have been used in some legal cases in the past. To address this concern, we show the extent to which the reasoning and decisions within the particular case can be made explicit and transparent. The BN approach enables us to clearly define the relevant propositions and evidence, and uses sensitivity analysis to assess the impact of the evidence under different assumptions. The results show that such a framework is suitable to identify information that is currently missing, yet clearly crucial for a valid and complete reasoning process. Furthermore, a method is provided whereby BNs can serve as a guide to not only reason with incomplete evidence in forensic cases, but also identify very specific research questions that should be addressed to extend the evidence base and solve similar issues in the future.This research was funded by the Engineering and Physical Sciences Research Council of the UK through the Security Science Doctoral Research Training Centre (UCL SECReT) based at University College London (EP/G037264/1), and the European Research Council (ERC-2013-AdG339182-BAYES_KNOWLEDGE)

Quantification of tumour vasculature and hypoxia by immunohistochemical staining and HbO2 saturation measurements

Despite the possibility that tumour hypoxia may limit radiotherapeutic response, the underlying mechanisms remain poorly understood. A new methodology has been developed in which information from several sophisticated techniques is combined and analysed at a microregional level. First, tumour oxygen availability is spatially defined by measuring intravascular blood oxygen saturations (HbO2) cryospectrophotometrically in frozen tumour blocks. Second, hypoxic development is quantified in adjacent sections using immunohistochemical detection of a fluorescently conjugated monoclonal antibody (ELK3-51) to a nitroheterocyclic hypoxia marker (EF5), thereby providing information relating to both the oxygen consumption rates and the effective oxygen diffusion distances. Third, a combination of fluorescent (Hoechst 33342 or DiOC7(3)) and immunohistological (PECAM-1/CD31) stains is used to define the anatomical vascular densities and the fraction of blood vessels containing flow. Using a computer-interfaced microscope stage, image analysis software and a 3-CCD colour video camera, multiple images are digitized, combined to form a photo-montage and revisited after each of the three staining protocols. By applying image registration techniques, the spatial distribution of HbO2 saturations is matched to corresponding hypoxic marker intensities in adjacent sections. This permits vascular configuration to be related to oxygen availability and allows the hypoxic marker intensities to be quantitated in situ. © 1999 Cancer Research Campaig

Polysaccharide peptide from Coriolus versicolor induces interleukin 6-related extension of endotoxin fever in rats

Purpose: Polysaccharide peptide (PSP) extracted from the Coriolus versicolor mushroom is frequently suggested as an adjunct to the chemo- or radiotherapy in cancer patients. In a previous study we showed that PSP induced a tumour necrosis factor-a (TNF-a)-dependent anapyrexia-like response in rats. Thus, PSP appears to be a factor which modifies a number of pathophysiological responses. Because of this, PSP is suggested as a potential adjuvant for cancer therapy during which cancer patients frequently contract microbial infections accompanied by fever. The aim of the present study was to investigate whether or not PSP can modulate the course of the fever in response to an antigen such as lipopolysaccharide (LPS). Materials and methods: Body temperature (Tb) of male Wistar rats was measured by biotelemetry. PSP was injected intraperitoneally (i.p.) at a dose of 100mgkg 1, 2 h before LPS administration (50 mgkg 1, i.p.). The levels of interleukin (IL)-6 and TNF-a in the plasma of rats were estimated 3 h and 14 h post-injection of PSP using a standard sandwich ELISA kit. Results: We report that i.p. pre-injection of PSP 2 h before LPS administration expanded the duration of endotoxin fever in rats. This phenomenon was accompanied by a significant elevation of the blood IL-6 level of rats both 3 h and 14 h post-injection of PSP. Pre-treatment i.p. of the rats with anti-IL-6 antibody (30 mg/rat) prevented the PSP-induced prolongation of endotoxin fever. Conclusions: Based on these data, we conclude that PSP modifies the LPS-induced fever in IL-6-related fashion

Ensemble Composition and Activity Levels of Insectivorous Bats in Response to Management Intensification in Coffee Agroforestry Systems

Shade coffee plantations have received attention for their role in biodiversity conservation. Bats are among the most diverse mammalian taxa in these systems; however, previous studies of bats in coffee plantations have focused on the largely herbivorous leaf-nosed bats (Phyllostomidae). In contrast, we have virtually no information on how ensembles of aerial insectivorous bats – nearly half the Neotropical bat species – change in response to habitat modification. To evaluate the effects of agroecosystem management on insectivorous bats, we studied their diversity and activity in southern Chiapas, Mexico, a landscape dominated by coffee agroforestry. We used acoustic monitoring and live captures to characterize the insectivorous bat ensemble in forest fragments and coffee plantations differing in the structural and taxonomic complexity of shade trees. We captured bats of 12 non-phyllostomid species; acoustic monitoring revealed the presence of at least 12 more species of aerial insectivores. Richness of forest bats was the same across all land-use types; in contrast, species richness of open-space bats increased in low shade, intensively managed coffee plantations. Conversely, only forest bats demonstrated significant differences in ensemble structure (as measured by similarity indices) across land-use types. Both overall activity and feeding activity of forest bats declined significantly with increasing management intensity, while the overall activity, but not feeding activity, of open-space bats increased. We conclude that diverse shade coffee plantations in our study area serve as valuable foraging and commuting habitat for aerial insectivorous bats, and several species also commute through or forage in low shade coffee monocultures

OX40 and 4-1BB delineate distinct immune profiles in sarcoma.

Systemic relapse after radiotherapy and surgery is the major cause of disease-related mortality in sarcoma patients. Combining radiotherapy and immunotherapy is under investigation as a means to improve response rates. However, the immune contexture of sarcoma is understudied. Here, we use a retrospective cohort of sarcoma patients, treated with neoadjuvant radiotherapy, and TCGA data. We explore therapeutic targets of relevance to sarcoma, using genomics and multispectral immunohistochemistry to provide insights into the tumor immune microenvironment across sarcoma subtypes. Differential gene expression between radioresponsive myxoid liposarcoma (MLPS) and more radioresistant undifferentiated pleomorphic sarcoma (UPS) indicated UPS contained higher transcript levels of a number of immunotherapy targets (CD73/NT5E, CD39/ENTPD1, CD25/IL2RA, and 4-1BB/TNFRSF9). We focused on 4-1BB/TNFRSF9 and other costimulatory molecules. In TCGA data, 4-1BB correlated to an inflamed and exhausted phenotype. OX40/TNFRSF4 and 4-1BB/TNFRSF9 were highly expressed in sarcoma subtypes versus other cancers. Despite OX40 and 4-1BB being described as Treg markers, we identified that they delineate distinct tumor immune profiles. This was true for sarcoma and other cancers. While only a limited number of samples could be analyzed, spatial analysis of OX40 expression identified two diverse phenotypes of OX40+ Tregs, one associated with and one independent of tertiary lymphoid structures (TLSs). Patient stratification is of intense interest for immunotherapies. We provide data supporting the viewpoint that a cohort of sarcoma patients, appropriately selected, are promising candidates for immunotherapies. Spatial profiling of OX40+ Tregs, in relation to TLSs, could be an additional metric to improve future patient stratification

Ensemble Composition and Activity Levels of Insectivorous Bats in Response to Management Intensification in Coffee Agroforestry Systems

Shade coffee plantations have received attention for their role in biodiversity conservation. Bats are among the most diverse mammalian taxa in these systems; however, previous studies of bats in coffee plantations have focused on the largely herbivorous leaf-nosed bats (Phyllostomidae). In contrast, we have virtually no information on how ensembles of aerial insectivorous bats – nearly half the Neotropical bat species – change in response to habitat modification. To evaluate the effects of agroecosystem management on insectivorous bats, we studied their diversity and activity in southern Chiapas, Mexico, a landscape dominated by coffee agroforestry. We used acoustic monitoring and live captures to characterize the insectivorous bat ensemble in forest fragments and coffee plantations differing in the structural and taxonomic complexity of shade trees. We captured bats of 12 non-phyllostomid species; acoustic monitoring revealed the presence of at least 12 more species of aerial insectivores. Richness of forest bats was the same across all land-use types; in contrast, species richness of open-space bats increased in low shade, intensively managed coffee plantations. Conversely, only forest bats demonstrated significant differences in ensemble structure (as measured by similarity indices) across land-use types. Both overall activity and feeding activity of forest bats declined significantly with increasing management intensity, while the overall activity, but not feeding activity, of open-space bats increased. We conclude that diverse shade coffee plantations in our study area serve as valuable foraging and commuting habitat for aerial insectivorous bats, and several species also commute through or forage in low shade coffee monocultures

Safety, immunogenicity, and transplacental antibody transport of conjugated and polysaccharide pneumococcal vaccines administered to pregnant women with HIV: a multicentre randomised controlled trial

Background: Pneumococcus remains an important cause of morbidity in pregnant women with HIV and their infants. We compared the safety and immunogenicity of PCV-10 and PPV-23 with placebo administered in pregnancy. Methods: This double-blind, multicentre, randomised controlled trial was done at eight outpatient clinics in Brazil. Eligible participants were adult women with HIV who were pregnant at a gestational age between 14 weeks and less than 34 weeks and who were taking antiretroviral therapy at study entry. Participants were randomly assigned (1:1:1) to receive either PCV-10, PPV-23, or placebo. Participants and study teams were unaware of treatment allocation. Antibodies against seven vaccine serotypes in PCV-10 and PPV-23 were measured by ELISA. The primary outcomes were maternal and infant safety assessed by the frequency of adverse events of grade 3 or higher; maternal seroresponse (defined as ≥2-fold increase in antibodies from baseline to 28 days after immunisation) against five or more serotypes; and infant seroprotection (defined as anti-pneumococcus antibody concentration of ≥0·35 μg/mL) against five or more serotypes at 8 weeks of life. The study was powered to detect differences of 20% or higher in the primary immunological outcomes between treatment groups. This trial is registered with ClinicalTrials.gov, NCT02717494. Findings: Between April 1, 2016, and Nov 30, 2017, we enrolled 347 pregnant women with HIV, of whom 116 were randomly assigned to the PCV-10 group, 115 to the PPV-23 group, and 116 to the placebo group. One participant in the PCV-10 group did not receive the vaccine and was excluded from subsequent analyses. The frequency of adverse events of grade 3 or higher during the first 4 weeks was similar in the vaccine and placebo groups (3% [90% CI 1–7] for the PCV-10 group, 2% [0–5] for the PPV-23 group, and 3% [1–8] for the placebo group). However, injection site and systemic grade 2 adverse reactions were reported more frequently during the first 4 weeks in the vaccine groups than in the placebo group (14% [9–20] for the PCV-10 group, 7% [4–12] for the PPV-23 group, and 3% [1–7] for the placebo group). The frequency of grade 3 or higher adverse effects was similar across maternal treatment groups (20% [14–27] for the PCV-10 group, 21% [14–28] for the PPV-23 group, and 20% [14–27] for the placebo group). Seroresponses against five or more serotypes were present in 74 (65%) of 114 women in the PCV-10 group, 72 (65%) of 110 women in the PPV-23 group, and none of the 113 women in the placebo group at 4 weeks post vaccination (p<0·0001 for PPV-23 group vs placebo and PCV-10 group vs placebo). Seroresponse differences of 20% or higher in vaccine compared with placebo recipients persisted up to 24 weeks post partum. At birth, 76 (67%) of 113 infants in the PCV-10 group, 62 (57%) of 109 infants in the PPV-23 group, and 19 (17%) of 115 infants in the placebo group had seroprotection against five or more serotypes (p<0·0001 for PPV-23 vs placebo and PCV-10 vs placebo). At 8 weeks, the outcome was met by 20 (19%) of 108 infants in the PCV-10 group, 24 (23%) of 104 infants in the PPV-23 group, and one (1%) of 109 infants in the placebo group (p<0·0001). Although a difference of 20% or higher compared with placebo was observed only in the infants who received PPV-23 at 8 weeks of life, the difference between the two vaccine groups was not appreciable. Interpretation: PCV-10 and PPV-23 were equally safe and immunogenic in pregnant women with HIV and conferred similar levels of seroprotection to their infants. In areas in which childhood PCV administration decreased the circulation of PCV serotypes, PPV-23 administration to pregnant women with HIV might be more advantageous than PCV by virtue of including a broader range of serotypes