Spatial transcriptome mapping of the desmoplastic growth pattern of colorectal liver metastases by in situ sequencing reveals a biologically relevant zonation of the desmoplastic rim.

PURPOSE: We describe the fibrotic rim formed in the desmoplastic growth pattern (DHGP) of colorectal cancer liver metastasis (CLM) using in situ sequencing (ISS). The origin of the desmoplastic rim is still a matter of debate, and the detailed cellular organization has not yet been fully elucidated. Understanding the biology of the DHGP in CLM can explore targeted treatment to improve survival. EXPERIMENTAL DESIGN: We used ISS, targeting 150 genes, to characterize the desmoplastic rim by unsupervised clustering of gene co-expression patterns. The cohort comprised 10 chemo-naïve liver metastasis resection samples with a DHGP. RESULTS: Unsupervised clustering of spatially mapped genes revealed molecular and cellular diversity within the desmoplastic rim. We confirmed the presence of the ductular reaction and cancer-associated fibroblasts. Importantly, we discovered angiogenesis and outer and inner zonation in the rim, characterized by NGFR and POSTN expression. CONCLUSIONS: ISS enabled the analysis of the cellular organization of the fibrous rim surrounding CLM with a DHGP and suggests a transition from the outer part of the rim, with nonspecific liver injury response, into the inner part, with gene expression indicating collagen synthesis and extracellular matrix remodeling influenced by the interaction with cancer cells, creating a cancer cell supportive environment. Moreover, we found angiogenic processes in the rim. Our results provide a potential explanation of the origin of the rim in DHGP and lead to exploring novel targeted treatments for patients with CLM to improve survival.

Interneuron diversity in the human dorsal striatum.

Deciphering the striatal interneuron diversity is key to understanding the basal ganglia circuit and to untangling the complex neurological and psychiatric diseases affecting this brain structure. We performed snRNA-seq and spatial transcriptomics of postmortem human caudate nucleus and putamen samples to elucidate the diversity and abundance of interneuron populations and their inherent transcriptional structure in the human dorsal striatum. We propose a comprehensive taxonomy of striatal interneurons with eight main classes and fourteen subclasses, providing their full transcriptomic identity and spatial expression profile as well as additional quantitative FISH validation for specific populations. We have also delineated the correspondence of our taxonomy with previous standardized classifications and shown the main transcriptomic and class abundance differences between caudate nucleus and putamen. Notably, based on key functional genes such as ion channels and synaptic receptors, we found matching known mouse interneuron populations for the most abundant populations, the recently described PTHLH and TAC3 interneurons. Finally, we were able to integrate other published datasets with ours, supporting the generalizability of this harmonized taxonomy.

BirthSeq, a new method to isolate and analyze dated cells in different vertebrates.

Embryonic development is a complex and dynamic process that unfolds over time and involves the production and diversification of increasing numbers of cells. The impact of developmental time on the formation of the central nervous system is well documented, with evidence showing that time plays a crucial role in establishing the identity of neuronal subtypes. However, the study of how time translates into genetic instructions driving cell fate is limited by the scarcity of suitable experimental tools. We introduce BirthSeq, a new method for isolating and analyzing cells based on their birth date. This innovative technique allows for in vivo labeling of cells, isolation via fluorescence-activated cell sorting, and analysis using high-throughput techniques. We calibrated the BirthSeq method for developmental organs across three vertebrate species (mouse, chick and gecko), and utilized it for single-cell RNA sequencing and novel spatially resolved transcriptomic approaches in mouse and chick, respectively. Overall, BirthSeq provides a versatile tool for studying virtually any tissue in different vertebrate organisms, aiding developmental biology research by targeting cells and their temporal cues.

Deconwolf enables high-performance deconvolution of widefield fluorescence microscopy images.

Microscopy-based spatially resolved omic methods are transforming the life sciences. However, these methods rely on high numerical aperture objectives and cannot resolve crowded molecular targets, limiting the amount of extractable biological information. To overcome these limitations, here we develop Deconwolf, an open-source, user-friendly software for high-performance deconvolution of widefield fluorescence microscopy images, which efficiently runs on laptop computers. Deconwolf enables accurate quantification of crowded diffraction limited fluorescence dots in DNA and RNA fluorescence in situ hybridization images and allows robust detection of individual transcripts in tissue sections imaged with ×20 air objectives. Deconvolution of in situ spatial transcriptomics images with Deconwolf increased the number of transcripts identified more than threefold, while the application of Deconwolf to images obtained by fluorescence in situ sequencing of barcoded Oligopaint probes drastically improved chromosome tracing. Deconwolf greatly facilitates the use of deconvolution in many bioimaging applications.

Associations between food intake and psychosomatic symptoms in 16-year-old adolescents.

AIMS: An increase in psychosomatic symptoms among adolescents has recently been reported. Few studies have examined the relation between food intake and psychosomatic symptoms. The aim was to study the association between food intake and overall psychosomatic burden and separate psychosomatic symptoms. METHODS: In this cross-sectional study, we used data from 6248 girls and 7153 boys in south-east Sweden who turned 16 years of age during the academic years 2009/2010 to 2015/2016 and responded to a health questionnaire at the school health services. The association between overall healthy food intake and a low psychosomatic burden was calculated as odds ratios (95% confidence interval) and stratified for other lifestyle habits and gender. RESULTS: Sixty-nine per cent of the boys and 35% of the girls had a low psychosomatic burden. There was a positive association between an overall healthy food intake and a low psychosomatic burden (P<0.0001), regardless of other lifestyle habits and gender. An overall healthy food intake was also positively associated with a lower frequency of the separate psychosomatic symptoms of concentration difficulties, sleep difficulties, a poor appetite or dizziness (P<0.0001). CONCLUSIONS: A healthy food intake seems to be associated with a low psychosomatic burden among adolescents. Further knowledge is needed to explore whether an improved food intake can reduce psychosomatic symptoms and enhance mental health.

Why the Single-N Design Should Be the Default in Affective Neuroscience.

Many studies in affective neuroscience rely on statistical procedures designed to estimate population averages and base their main conclusions on group averages. However, the obvious unit of analysis in affective neuroscience is the individual, not the group, because emotions are individual phenomena that typically vary across individuals. Conclusions based on group averages may therefore be misleading or wrong, if interpreted as statements about emotions of an individual, or meaningless, if interpreted as statements about the group, which has no emotions. We therefore advocate the Single-N design as the default strategy in research on emotions, testing one or several individuals extensively with the primary purpose of obtaining results at the individual level. In neuroscience, the equivalent to the Single-N design is deep imaging, the emerging trend of extensive measurements of activity in single brains. Apart from the fact that individuals react differently to emotional stimuli, they also vary in shape and size of their brains. Group-based analysis of brain imaging data therefore refers to an "average brain" that was activated in a way that may not be representative of the physiology of any of the tested individual brains, nor of how these brains responded to the experimental stimuli. Deep imaging avoids such group-averaging artifacts by simply focusing on the individual brain. This methodological shift toward individual analysis has already opened new research areas in fields like vision science. Inspired by this, we call for a corresponding shift in affective neuroscience, away from group averages, and toward experimental designs targeting the individual.

Cellular architecture of evolving neuroinflammatory lesions and multiple sclerosis pathology.

Multiple sclerosis (MS) is a neurological disease characterized by multifocal lesions and smoldering pathology. Although single-cell analyses provided insights into cytopathology, evolving cellular processes underlying MS remain poorly understood. We investigated the cellular dynamics of MS by modeling temporal and regional rates of disease progression in mouse experimental autoimmune encephalomyelitis (EAE). By performing single-cell spatial expression profiling using in situ sequencing (ISS), we annotated disease neighborhoods and found centrifugal evolution of active lesions. We demonstrated that disease-associated (DA)-glia arise independently of lesions and are dynamically induced and resolved over the disease course. Single-cell spatial mapping of human archival MS spinal cords confirmed the differential distribution of homeostatic and DA-glia, enabled deconvolution of active and inactive lesions into sub-compartments, and identified new lesion areas. By establishing a spatial resource of mouse and human MS neuropathology at a single-cell resolution, our study unveils the intricate cellular dynamics underlying MS.

A drained nutrient-poor peatland forest in boreal Sweden constitutes a net carbon sink after integrating terrestrial and aquatic fluxes.

Northern peatlands provide a globally important carbon (C) store. Since the beginning of the 20th century, however, large areas of natural peatlands have been drained for biomass production across Fennoscandia. Today, drained peatland forests constitute a common feature of the managed boreal landscape, yet their ecosystem C balance and associated climate impact are not well understood, particularly within the nutrient-poor boreal region. In this study, we estimated the net ecosystem carbon balance (NECB) from a nutrient-poor drained peatland forest and an adjacent natural mire in northern Sweden by integrating terrestrial carbon dioxide (CO2 ) and methane (CH4 ) fluxes with aquatic losses of dissolved organic C (DOC) and inorganic C based on eddy covariance and stream discharge measurements, respectively, over two hydrological years. Since the forest included a dense spruce-birch area and a sparse pine area, we were able to further evaluate the effect of contrasting forest structure on the NECB and component fluxes. We found that the drained peatland forest was a net C sink with a 2-year mean NECB of -115 ± 5 g C m-2 year-1 while the adjacent mire was close to C neutral with 14.6 ± 1.7 g C m-2 year-1 . The NECB of the drained peatland forest was dominated by the net CO2 exchange (net ecosystem exchange [NEE]), whereas NEE and DOC export fluxes contributed equally to the mire NECB. We further found that the C sink strength in the sparse pine forest area (-153 ± 8 g C m-2 year-1 ) was about 1.5 times as high as in the dense spruce-birch forest area (-95 ± 8 g C m-2 year-1 ) due to enhanced C uptake by ground vegetation and lower DOC export. Our study suggests that historically drained peatland forests in nutrient-poor boreal regions may provide a significant net ecosystem C sink and associated climate benefits.

Harmonised statistics and maps of forest biomass and increment in Europe.

Forest biomass is an essential resource in relation to the green transition and its assessment is key for the sustainable management of forest resources. Here, we present a forest biomass dataset for Europe based on the best available inventory and satellite data, with a higher level of harmonisation and spatial resolution than other existing data. This database provides statistics and maps of the forest area, biomass stock and their share available for wood supply in the year 2020, and statistics on gross and net volume increment in 2010-2020, for 38 European countries. The statistics of most countries are available at a sub-national scale and are derived from National Forest Inventory data, harmonised using common reference definitions and estimation methodology, and updated to a common year using a modelling approach. For those counties without harmonised statistics, data were derived from the State of Europe's Forest 2020 Report at the national scale. The maps are coherent with the statistics and depict the spatial distribution of the forest variables at 100 m resolution.

Mercury deposition and redox transformation processes in peatland constrained by mercury stable isotopes.

Peatland vegetation takes up mercury (Hg) from the atmosphere, typically contributing to net production and export of neurotoxic methyl-Hg to downstream ecosystems. Chemical reduction processes can slow down methyl-Hg production by releasing Hg from peat back to the atmosphere. The extent of these processes remains, however, unclear. Here we present results from a comprehensive study covering concentrations and isotopic signatures of Hg in an open boreal peatland system to identify post-depositional Hg redox transformation processes. Isotope mass balances suggest photoreduction of HgII is the predominant process by which 30% of annually deposited Hg is emitted back to the atmosphere. Isotopic analyses indicate that above the water table, dark abiotic oxidation decreases peat soil gaseous Hg0 concentrations. Below the water table, supersaturation of gaseous Hg is likely created more by direct photoreduction of rainfall rather than by reduction and release of Hg from the peat soil. Identification and quantification of these light-driven and dark redox processes advance our understanding of the fate of Hg in peatlands, including the potential for mobilization and methylation of HgII.

Spatial multimodal analysis of transcriptomes and metabolomes in tissues.

We present a spatial omics approach that combines histology, mass spectrometry imaging and spatial transcriptomics to facilitate precise measurements of mRNA transcripts and low-molecular-weight metabolites across tissue regions. The workflow is compatible with commercially available Visium glass slides. We demonstrate the potential of our method using mouse and human brain samples in the context of dopamine and Parkinson's disease.

Spatial tumour gene signature discriminates neoplastic from non-neoplastic compartments in colon cancer: unravelling predictive biomarkers for relapse.

BACKGROUND: Opting for or against the administration of adjuvant chemotherapy in therapeutic management of stage II colon cancer remains challenging. Several studies report few survival benefits for patients treated with adjuvant therapy and additionally revealing potential side effects of overtreatment, including unnecessary exposure to chemotherapy-induced toxicities and reduced quality of life. Predictive biomarkers are urgently needed. We, therefore, hypothesise that the spatial tissue composition of relapsed and non-relapsed colon cancer stage II patients reveals relevant biomarkers. METHODS: The spatial tissue composition of stage II colon cancer patients was examined by a novel spatial transcriptomics technology with sub-cellular resolution, namely in situ sequencing. A panel of 176 genes investigating specific cancer-associated processes such as apoptosis, proliferation, angiogenesis, stemness, oxidative stress, hypoxia, invasion and components of the tumour microenvironment was designed to examine differentially expressed genes in tissue of relapsed versus non-relapsed patients. Therefore, FFPE slides of 10 colon cancer stage II patients either classified as relapsed (5 patients) or non-relapsed (5 patients) were in situ sequenced and computationally analysed. RESULTS: We identified a tumour gene signature that enables the subclassification of tissue into neoplastic and non-neoplastic compartments based on spatial expression patterns obtained through in situ sequencing. We developed a computational tool called Genes-To-Count (GTC), which automates the quantification of in situ signals, accurately mapping their position onto the spatial tissue map and automatically identifies neoplastic and non-neoplastic tissue compartments. The GTC tool was used to quantify gene expression of biological processes upregulated within the neoplastic tissue in comparison to non-neoplastic tissue and within relapsed versus non-relapsed stage II colon patients. Three differentially expressed genes (FGFR2, MMP11 and OTOP2) in the neoplastic tissue compartments of relapsed patients in comparison to non-relapsed patients were identified predicting recurrence in stage II colon cancer. CONCLUSIONS: In depth spatial in situ sequencing showed potential to provide a deeper understanding of the underlying mechanisms involved in the recurrence of disease and revealed novel potential predictive biomarkers for disease relapse in colon cancer stage II patients. Our open-access GTC-tool allowed us to accurately capture the tumour compartment and quantify spatial gene expression in colon cancer tissue.

Comparison of absorbed doses and organ doses measured with thermoluminescent dosimeters and Gafchromic film for cone beam computed tomography examination of the posterior mandibular region in a head phantom.

OBJECTIVES: We aimed to map the correlation between thermoluminescent dosimeters (TLDs) and Gafchromic film for measuring absorbed doses and to compare minimum, maximum, and mean absorbed doses over larger regions of interest and at various craniofacial organs and tissues during cone beam computed tomography (CBCT) exposure of the mandibular third molar region. STUDY DESIGN: We positioned TLDs at 75 measurement points in a head phantom. Gafchromic film was cut to the same shape as the 5 levels of the phantom and was placed on top of the TLDs. Both dosimetry methods thus included the surface of each level simultaneously. CBCT scans were made using a 5 × 5 cm field of view and a rotation angle of 200°. Measurements included absorbed dose distributions, doses at all 75 points, and minimum, maximum, and mean doses within organs and tissues. RESULTS: The correlation of point-dose measurements at all TLD sites with doses measured on film was strong (R2 = 0.9687), with greatest correlation at lower doses (<2 mGy). Large deviations between TLD and film measurements of minimum and maximum doses and absorbed doses to the organs occurred at all 5 levels. TLD positioning failed to cover several organ sites; for these, only absorbed dose measurements from the film were available. CONCLUSIONS: TLDs were unable to sample dose distributions and gradients accurately. The characteristics of Gafchromic LD-V1 film make it a favorable alternative in dental CBCT dosimetry.

Highly multiplexed targeted sequencing strategy for infectious disease surveillance.

BACKGROUND: Global efforts to characterize diseases of poverty are hampered by lack of affordable and comprehensive detection platforms, resulting in suboptimal allocation of health care resources and inefficient disease control. Next generation sequencing (NGS) can provide accurate data and high throughput. However, shotgun and metagenome-based NGS approaches are limited by low concentrations of microbial DNA in clinical samples, requirements for tailored sample and library preparations plus extensive bioinformatics analysis. Here, we adapted molecular inversion probes (MIPs) as a cost-effective target enrichment approach to characterize microbial infections from blood samples using short-read sequencing. We designed a probe panel targeting 2 bacterial genera, 21 bacterial and 6 fungi species and 7 antimicrobial resistance markers (AMRs). RESULTS: Our approach proved to be highly specific to detect down to 1 in a 1000 pathogen DNA targets contained in host DNA. Additionally, we were able to accurately survey pathogens and AMRs in 20 out of 24 samples previously profiled with routine blood culture for sepsis. CONCLUSIONS: Overall, our targeted assay identifies microbial pathogens and AMRs with high specificity at high throughput, without the need for extensive sample preparation or bioinformatics analysis, simplifying its application for characterization and surveillance of infectious diseases in medium- to low- resource settings.

Detection and attribution of an anomaly in terrestrial photosynthesis in Europe during the COVID-19 lockdown.

Carbon dioxide (CO2) uptake by plant photosynthesis, referred to as gross primary production (GPP) at the ecosystem level, is sensitive to environmental factors, including pollutant exposure, pollutant uptake, and changes in the scattering of solar shortwave irradiance (SWin) - the energy source for photosynthesis. The 2020 spring lockdown due to COVID-19 resulted in improved air quality and atmospheric transparency, providing a unique opportunity to assess the impact of air pollutants on terrestrial ecosystem functioning. However, detecting these effects can be challenging as GPP is influenced by other meteorological drivers and management practices. Based on data collected from 44 European ecosystem-scale CO2 flux monitoring stations, we observed significant changes in spring GPP at 34 sites during 2020 compared to 2015-2019. Among these, 14 sites showed an increase in GPP associated with higher SWin, 10 sites had lower GPP linked to atmospheric and soil dryness, and seven sites were subjected to management practices. The remaining three sites exhibited varying dynamics, with one experiencing colder and rainier weather resulting in lower GPP, and two showing higher GPP associated with earlier spring melts. Analysis using the regional atmospheric chemical transport model (LOTOS-EUROS) indicated that the ozone (O3) concentration remained relatively unchanged at the research sites, making it unlikely that O3 exposure was the dominant factor driving the primary production anomaly. In contrast, SWin increased by 9.4 % at 36 sites, suggesting enhanced GPP possibly due to reduced aerosol optical depth and cloudiness. Our findings indicate that air pollution and cloudiness may weaken the terrestrial carbon sink by up to 16 %. Accurate and continuous ground-based observations are crucial for detecting and attributing subtle changes in terrestrial ecosystem functioning in response to environmental and anthropogenic drivers.

Catchment characteristics control boreal mire nutrient regime and vegetation patterns over ~5000 years of landscape development.

Vegetation holds the key to many properties that make natural mires unique, such as surface microtopography, high biodiversity values, effective carbon sequestration and regulation of water and nutrient fluxes across the landscape. Despite this, landscape controls behind mire vegetation patterns have previously been poorly described at large spatial scales, which limits the understanding of basic drivers underpinning mire ecosystem services. We studied catchment controls on mire nutrient regimes and vegetation patterns using a geographically constrained natural mire chronosequence along the isostatically rising coastline in Northern Sweden. By comparing mires of different ages, we can partition vegetation patterns caused by long-term mire succession (<5000 years) and present-day vegetation responses to catchment eco-hydrological settings. We used the remote sensing based normalized difference vegetation index (NDVI) to describe mire vegetation and combined peat physicochemical measures with catchment properties to identify the most important factors that determine mire NDVI. We found strong evidence that mire NDVI depends on nutrient inputs from the catchment area or underlying mineral soil, especially concerning phosphorus and potassium concentrations. Steep mire and catchment slopes, dry conditions and large catchment areas relative to mire areas were associated with higher NDVI. We also found long-term successional patterns, with lower NDVI in older mires. Importantly, the NDVI should be used to describe mire vegetation patterns in open mires if the focus is on surface vegetation, since the canopy cover in tree-covered mires completely dominated the NDVI signal. With our study approach, we can quantitatively describe the connection between landscape properties and mire nutrient regime. Our results confirm that mire vegetation responds to the upslope catchment area, but importantly, also suggest that mire and catchment aging can override the role of catchment influence. This effect was clear across mires of all ages, but was strongest in younger mires.

On the uncertainty in estimates of the carbon balance recovery time after forest clear-cutting.

An essential metric for describing carbon dynamics in managed forest landscapes is the recovery time of the carbon balance after clear-cutting. Here, we demonstrate how the age-dependent mathematical trajectory is affected by both the selected model and data availability, leading to considerable uncertainty in the modelling of the net ecosystem production (NEP) over stand age. We further show that the initial carbon loss estimates associated with the timing of the source-sink transition (SST) are significant, but may have a limited effect on the total carbon sequestration at the end of the standard (RP, 120 years) or optimal (OCS) rotation periods.

Single Nucleotide Polymorphism Directed Antiemetic Treatment in Women With Breast Cancer Treated With Neo- or Adjuvant Chemotherapy: A Randomised Multicentre Phase II Study. (EudraCT: 2015-000658-39).

BACKGROUND/AIM: The role of single nucleotide polymorphisms (SNPs) in the frequency and intensity of chemotherapy-induced nausea and vomiting (CINV) in women with breast cancer (BC) is unclear. The primary purpose of this study was to compare/evaluate the effect of SNP-guided antiemetic treatment versus standard CINV treatment. PATIENTS AND METHODS: A randomised, factorial, phase II multicentre study design was used. Women planned for neoadjuvant or adjuvant chemotherapy with epirubicin, cyclophosphamide and fluorouracil (FEC /EC, with or without fluorouracil) for BC were randomised to SNP-guided antiemetic treatment (based on the results of SNP analyses) versus standard CINV treatment. Blood samples were taken before the treatment was initiated. Patient-reported data on CINV (during 10 days from onset of cancer treatment) and health-related quality of life (HRQoL), were collected before and after the first cancer treatment. RESULTS: A total of 188 women were included. Overall, nausea was reported by 86% (n=129) of the patients during the ten-day period from the start of cancer treatment. The SNP genotype studied varied. In FAS-CD95, the genotypes AG and GG were overrepresented; in RB1-LPAR6, GG was overrepresented, and in CCL2, both AA and GG were overrepresented. We found no statistically significant difference in CINV between SNP-guided antiemetic treatment versus standard CINV treatment. CONCLUSION: SNP-guided antiemetic treatment could be as effective as standard treatment. SNP-guided antiemetic treatment of CINV is possibly useful in detecting patients with a higher or lower risk for CINV and thus may help in avoiding over-treatment with toxic components. CINV negatively affects the HRQL.