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Metastasis is the major cause of cancer-related deaths. Neuroblastoma (NB), a childhood tumor has been molecularly defined at the primary cancer site, however, the bone marrow (BM) as the metastatic niche of NB is poorly characterized. Here we perform single-cell transcriptomic and epigenomic profiling of BM aspirates from 11 subjects spanning three major NB subtypes and compare these to five age-matched and metastasis-free BM, followed by in-depth single cell analyses of tissue diversity and cell-cell interactions, as well as functional validation. We show that cellular plasticity of NB tumor cells is conserved upon metastasis and tumor cell type composition is NB subtype-dependent. NB cells signal to the BM microenvironment, rewiring via macrophage mgration inhibitory factor and midkine signaling specifically monocytes, which exhibit M1 and M2 features, are marked by activation of pro- and anti-inflammatory programs, and express tumor-promoting factors, reminiscent of tumor-associated macrophages. The interactions and pathways characterized in our study provide the basis for therapeutic approaches that target tumor-to-microenvironment interactions. The bone marrow is a common site of metastasis for neuroblastoma patients. Here, the authors perform single cell RNA-seq and ATAC-seq of bone marrow aspirates from 16 subjects and show conservation of tumor cell plasticity in metastases and identify tumor-to-bone marrow cell signals that trigger tumor promoting monocytes.

Natural killer (NK) cells have multifaceted roles within the complex tumor milieu. They are pivotal components of innate immunity and shape the dynamic landscape of tumor-immune cell interactions, and thus can be leveraged for use in therapeutic interventions. NK-based immunotherapies have had remarkable success in hematological malignancies, but these therapies are met with many challenges in solid tumors, including neuroblastoma (NB), a childhood tumor arising from the sympathetic nervous system. With a focus on NB, this review outlines the mechanisms employed by NK cells to recognize and eliminate malignant cells, delving into the dynamic relationship between ligand-receptor interactions, cytokines, and other molecules that facilitate the cross talk between NK and NB cells. We discuss the immunomodulatory functions of NK cells and the mechanisms that contribute to loss of this immunosurveillance in NB, with a focus on how this dynamic has been utilized in recent immunotherapy advancements for NB.

Early childhood tumours arise from transformed embryonic cells, which often carry large copy number alterations (CNA). However, it remains unclear how CNAs contribute to embryonic tumourigenesis due to a lack of suitable models. Here we employ female human embryonic stem cell (hESC) differentiation and single-cell transcriptome and epigenome analysis to assess the effects of chromosome 17q/1q gains, which are prevalent in the embryonal tumour neuroblastoma (NB). We show that CNAs impair the specification of trunk neural crest (NC) cells and their sympathoadrenal derivatives, the putative cells-of-origin of NB. This effect is exacerbated upon overexpression of MYCN , whose amplification co-occurs with CNAs in NB. Moreover, CNAs potentiate the pro-tumourigenic effects of MYCN and mutant NC cells resemble NB cells in tumours. These changes correlate with a stepwise aberration of developmental transcription factor networks. Together, our results sketch a mechanistic framework for the CNA-driven initiation of embryonal tumours. Copy number alterations in stem cells impair neural crest differentiation and set the stage for neuroblastoma-like traits and tumours. This study hints at early tumourigenesis mechanisms and finds developmental gene signatures linked to prognosis.

Alzheimer’s disease (AD) accounts for more than 60% of the dementia cases and currently there is no curative treatment for it. With the emergence of potentially disease modifying treatments, early diagnosis is key to identify patient groups that would benefit from such treatments, aiming to prevent severe cognitive decline. We previously identified a set of DNA methylation signatures that allow for accurate diagnosis of AD in cortical neurons and brain tissue, even before clinical manifestation of the disease [1]. Here we investigate 11 of these signature regions via targeted next-generation sequencing in cell-free DNA (cfDNA) isolated from cerebrospinal fluid (CSF) of AD patients homozygous for APOE4 (n=4) and sporadic AD (n=5) cases compared to age-matched control samples (n=5). Our analyses demonstrated that 6/11 of the tested DNA methylation signatures that had initially been identified in cortical neurons and brain tissue were also validated in cfDNA. The remainder of the tested regions either showed opposite trends (3/11) or did not result in any differences (2/11) between control and AD cases. Thus, this presents a direct approach allowing to test for these DNA methylation signatures in CSF-derived cfDNA, and bypasses the need to generate induced pluripotent stem cell-derived cortical neurons from patients.

Early childhood tumours arise from transformed embryonic cells, which often carry large copy number alterations (CNA). However, it remains unclear how CNAs contribute to embryonic tumourigenesis due to a lack of suitable models. Here we employ female human embryonic stem cell (hESC) differentiation and single-cell transcriptome and epigenome analysis to assess the effects of chromosome 17q/1q gains, which are prevalent in the embryonal tumour neuroblastoma (NB). We show that CNAs impair the specification of trunk neural crest (NC) cells and their sympathoadrenal derivatives, the putative cells-of-origin of NB. This effect is exacerbated upon overexpression of MYCN, whose amplification co-occurs with CNAs in NB. Moreover, CNAs potentiate the pro-tumourigenic effects of MYCN and mutant NC cells resemble NB cells in tumours. These changes correlate with a stepwise aberration of developmental transcription factor networks. Together, our results sketch a mechanistic framework for the CNA-driven initiation of embryonal tumours.Competing Interest StatementThe authors have declared no competing interest.Footnotes* The entire manuscript has been substantially revised. In particular, we added data based on cellular assays (apoptosis, DNA damage), scRNA-seq (10x Genomics and Parse Bio), WES (SNV and CNA analyses), integration with public genomics data (SEQC and others), and mouse xenografts using the previously used stem cell lines and new derivatives (inducing MYCN in WT and 17q-only background). All figures have been updated and we added a new figure (Fig. 5) and several supplementary figures.

Familial Dysautonomia (FD) is an autosomal recessive disorder caused by a splice site mutation in the gene ELP1, which disproportionally affects neurons. While classically characterized by deficits in sensory and autonomic neurons, neuronal defects in the central nervous system have been described. ELP1 is highly expressed in the normal developing and adult cerebellum, but its role in cerebellum development is unknown. To investigate the cerebellar function of Elp1, we knocked out Elp1 in cerebellar granule cell progenitors (GCPs) and examined the outcome on animal behavior and cellular composition. We found that GCP-specific conditional knockout of Elp1 (Elp1cKO) resulted in ataxia by 8 weeks of age. Cellular characterization showed that the animals had smaller cerebella with fewer granule cells. This defect was already apparent 7 days after birth, when Elp1cKO animals also exhibited fewer mitotic GCPs and shorter Purkinje dendrites. Through molecular characterization, we found that loss of Elp1 was associated with an increase in apoptotic cell death and cell stress pathways in GCPs. Our study demonstrates the importance of ELP1 within the developing cerebellum, and suggests that Elp1 loss in the GC lineage may also play a role in the progressive ataxia phenotypes of FD patients.

Background: Since February 24th, 2022, the beginning of Russia’s aggression against Ukraine, more than 80,000 women were expected to give birth. Therefore, understanding the impact of war on the perinatal health of women is an important requisite to improve perinatal care. This narrative synthesis has two main purposes: on one hand, it aims to summarize the current evidence available based on perinatal health outcomes and care among perinatal women; on the other, it attempts to identify the gaps still present in research in relation to perinatal care. Method: A literature search was completed in diverse databases (e.g., Medline, PsychInfo). Results: Emergent matters related to practice and research in perinatal refugee women have been discussed. Conclusions: In the face of the war in Ukraine, we need to build up further research to provide an evidence-based foundation for preventing and treating the psychological consequences of pregnant women exposed directly to war and those who have been forced into a refugee status during this vulnerable period. Also, it is essential to support not only women transitioning to motherhood, but also supporting midwives and nurses in their work.

•Some types of malformation of cortical development may not be easily detected by conventional visual MRI analysis (CVA).•MAP07 program is a sophisticated diagnostic program that offers several postprocessing morphometric maps.•MAP07 facilitates detection and localization of hippocampal sclerosis, focal cortical dysplasias and other types of cortical malformations.•MAP07 program may be used as a complementary method to CVA.•MAP07 may guide additional re-evaluation of MRI by highlighting suspicious cortical regions and thus increase the sensitivity of the MRI itself This study aimed to determine the role of brain MRI post-processing method MAP07 (Morphometric Analysis Program) in detecting epileptogenic brain lesions in patients with pharmacoresistant epilepsy (PE). MAP07 is a sophisticated diagnostic program that offers several morphometric maps and facilitates the detection and localization of hippocampal sclerosis (HS), focal cortical dysplasias (FCD), and other types of cortical malformations, which could be undetected by conventional visual MRI analysis (CVA). 120 patients aged > 16 years with PE have been recruited. 3 T MRI was performed according to epilepsy imaging protocol followed by image postprocessing with a fully automated MATLAB script, MAP07, by applying SPM5 algorithms. Statistical analysis was performed in IBM SPSS Statistics, version 25.0. Analysis in our patients showed a high sensitivity of MAP07 with low specificity and with a high proportion of false-positive patients. After MRI analysis, out of 120 patients, 32 were found to have no structural abnormalities by conventional visual analysis in whom after MAP07 in 5 patients structural lesions were found (in one HS, in one FCD, in two perinatal vascular lesions, and in one hippocampal hyperintensity). There was a quite high overall coincidence of the findings of MAP07 and MRI for the detection of FCD, HS, perinatal ischemia/chronic vascular lesions, heterotopias, and polymicrogyria (kappa coefficient above 0.700). MAP07 analysis is a useful, additional, and automated method that may guide re-evaluation of MRI by highlighting suspicious cortical regions, as a complementary method to CVA, by enhancing the visualization of cortical malformations and lesions.

Protein aggregation is a well-recognized problem in industrial preparation, including biotherapeutics. These low-energy states constantly compete with a native-like conformation, which is more pronounced in the case of macromolecules of low stability in the solution. A better understanding of the structure and function of such aggregates is generally required for the more rational development of therapeutic proteins, including single-chain fusion cytotoxins to target specific receptors on cancer cells. Here, we identified and purified such particles as side products of the renaturation process of the single-chain fusion cytotoxin, composed of two diphtheria toxin (DT) domains and interleukin 13 (IL-13), and applied various experimental techniques to comprehensively understand their molecular architecture and function. Importantly, we distinguished soluble purified dimeric and fractionated oligomeric particles from aggregates. The oligomers are polydisperse and multimodal, with a distribution favoring lower and even stoichiometries, suggesting they are composed of dimeric building units. Importantly, all these oligomeric particles and the monomer are cystine-dependent as their innate disulfide bonds have structural and functional roles. Their reduction triggers aggregation. Presumably the dimer and lower oligomers represent the metastable state, retaining the native disulfide bond. Although significantly reduced in contrast to the monomer, they preserve some fraction of bioactivity, manifested by their IL-13RA2 receptor affinity and selective cytotoxic potency towards the U-251 glioblastoma cell line. These molecular assemblies probably preserve structural integrity and native-like fold, at least to some extent. As our study demonstrated, the dimeric and oligomeric cytotoxin may be an exciting model protein, introducing a new understanding of its monomeric counterpart’s molecular characteristics.

ObjectiveHere we looked for possible mechanisms regulating the progression of type 1 diabetes mellitus (T1DM). In this disease, autoaggressive T cells (T conventional cells, Tconvs) not properly controlled by T regulatory cells (Tregs) destroy pancreatic islets.Research design and methodsWe compared the T-cell compartment of patients with newly diagnosed T1DM (NDT1DM) with long-duration T1DM (LDT1DM) ones. The third group consisted of patients with LDT1DM treated previously with polyclonal Tregs (LDT1DM with Tregs). We have also looked if the differences might be dependent on the antigen specificity of Tregs expanded for clinical use and autologous sentinel Tconvs.ResultsPatients with LDT1DM were characterized by T-cell immunosenescence-like changes and expansion of similar vβ/T-cell receptor (TCR) clones in Tconvs and Tregs. The treatment with Tregs was associated with some inhibition of these effects. Patients with LDT1DM possessed an increased percentage of various proinsulin-specific T cells but not GAD65-specific ones. The percentages of all antigen-specific subsets were higher in the expansion cultures than in the peripheral blood. The proliferation was more intense in proinsulin-specific Tconvs than in specific Tregs but the levels of some proinsulin-specific Tregs were exceptionally high at baseline and remained higher in the expanded clinical product than the levels of respective Tconvs in sentinel cultures.ConclusionsT1DM is associated with immunosenescence-like changes and reduced diversity of T-cell clones. Preferential expansion of the same TCR families in both Tconvs and Tregs suggests a common trigger/autoantigen responsible. Interestingly, the therapy with polyclonal Tregs was associated with some inhibition of these effects. Proinsulin-specific Tregs appeared to be dominant in the immune responses in patients with T1DM and probably associated with better control over respective autoimmune Tconvs.Trial registration numberEudraCT 2014-004319-35.