Bujko et al. describe four distinct subsets of macrophages in human small intestine that are completely replaced in transplanted duodenum. These subsets show graduated changes in their phenotypes, function, and transcriptome profiles, suggesting that they represent stages of monocyte-derived macrophage maturation in tissue.

Diffuse brain infiltration by glioma cells causes detrimental disease progression, however its multicellular coordination is poorly understood. We here show that glioma cells infiltrate brain collectively, as multicellular networks. Contacts between moving glioma cells were adaptive epithelial-like or filamentous junctions stabilized by N-cadherin, β-catenin and p120-catenin, which underwent kinetic turnover , transmitted intercellular calcium transients and mediated directional persistence. Downregulation of p120-catenin compromised cell-cell interaction and communication, disrupted collective networks, and both the cadherin and RhoA binding domains of p120-catenin were required for network formation and migration. Deregulating p120-catenin further prevented diffuse glioma cell infiltration of the mouse brain with marginalized Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:

The tissue dendritic cell (DC) compartment is heterogeneous, and the ontogeny and functional specialization of human tissue conventional DC (cDC) subsets and their relationship with monocytes is unresolved. Here we identify monocyte-related CSF1RFlt3 antigen presenting cells (APCs) that constitute about half of the cells classically defined as SIRPα DCs in the steady-state human small intestine. CSF1RFlt3 APCs express calprotectin and very low levels of CD14, are transcriptionally related to monocyte-derived cells, and accumulate during inflammation. CSF1RFlt3 APCs show typical macrophage characteristics functionally distinct from their Flt3 cDC counterparts: under steady-state conditions they excel at antigen uptake, have a lower migratory potential, and are inefficient activators of naïve T cells. These results have important implications for the understanding of the ontogenetic and functional heterogeneity within human tissue DCs and their relation to the monocyte lineage.

Celiac disease is an autoimmune disorder in which ingestion of dietary gluten triggers an immune reaction in the small intestine leading to destruction of the lining epithelium. Current treatment focusses on lifelong adherence to a gluten-free diet. Gluten-specific CD4+ T cells and cytotoxic intraepithelial CD8+ T cells have been proposed to be central in disease pathogenesis. Here we use unbiased single-cell RNA-sequencing and explore the heterogeneity of CD45+ immune cells in the human small intestine. We show altered myeloid cell transcriptomes present in active celiac lesions. CD4+ and CD8+ T cells transcriptomes show extensive changes and we define a natural intraepithelial lymphocyte population that is reduced in celiac disease. We show that the immune landscape in Celiac patients on a gluten-free diet is only partially restored compared to control samples. Altogether, we provide a single cell transcriptomic resource that can inform the immune landscape of the small intestine during Celiac disease.

Background Acute myeloid leukemia (AML) is a heterogeneous and aggressive blood cancer that results from diverse genetic aberrations in the hematopoietic stem or progenitor cells (HSPCs) leading to the expansion of blasts in the hematopoietic system. The heterogeneity and evolution of cancer blasts can render therapeutic interventions ineffective in a yet poorly understood patient-specific manner. In this study, we investigated the clonal heterogeneity of diagnosis (Dx) and relapse (Re) pairs at genetic and transcriptional levels, and unveiled the underlying pathways and genes contributing to recurrence. Methods Whole-exome sequencing was used to detect somatic mutations and large copy number variations (CNVs). Single cell RNA-seq was performed to investigate the clonal heterogeneity between Dx-Re pairs and amongst patients. Results scRNA-seq analysis revealed extensive expression differences between patients and Dx-Re pairs, even for those with the same -presumed- initiating events. Transcriptional differences between and within patients are associated with clonal composition and evolution, with the most striking differences in patients that gained large-scale copy number variations at relapse. These differences appear to have significant molecular implications, exemplified by a DNMT3A/FLT3-ITD patient where the leukemia switched from an AP-1 regulated clone at Dx to a mTOR signaling driven clone at Re. The two distinct AML1-ETO pairs share genes related to hematopoietic stem cell maintenance and cell migration suggesting that the Re leukemic stem cell-like (LSC-like) cells evolved from the Dx cells. Conclusions In summary, the single cell RNA data underpinned the tumor heterogeneity not only amongst patient blasts with similar initiating mutations but also between each Dx-Re pair. Our results suggest alternatively and currently unappreciated and unexplored mechanisms leading to therapeutic resistance and AML recurrence.

22 23 24 25 Celiac disease (CeD) is an autoimmune disorder in which ingestion of dietary gluten 26 triggers an immune reaction in the small intestine 1,2 . The CeD lesion is characterized by 27 crypt hyperplasia, villous atrophy and chronic inflammation with accumulation of 28 leukocytes both in the lamina propria (LP) and in the epithelium 3 , which eventually 29 leads to destruction of the intestinal epithelium 1 and subsequent digestive complications 30 and higher risk of non-hodgkin lymphoma 4 . A lifetime gluten-free diet is currently the 31 only available treatment 5 . Gluten-specific LP CD4 T cells and cytotoxic intraepithelial 32 CD8+ T cells are thought to be central in disease pathology 1,6-8 , however, CeD is a 33 complex immune-mediated disorder and to date the findings are mostly based on 34 analysis of heterogeneous cell populations and on animal models. Here, we 35comprehensively explore the cellular heterogeneity of CD45+ immune cells in human 36 small intestine using index-sorting single-cell RNA-sequencing 9,10 . We find that myeloid 37

3D in vitro culture models of cancer cells in extracellular matrix (ECM) have been developed to investigate drug targeting and resistance or, alternatively, mechanisms of invasion; however, models allowing analysis of shared pathways mediating invasion and therapy resistance are lacking. To evaluate therapy response associated with cancer cell invasion, we here used 3D invasion culture of tumor spheroids in 3D fibrillar collagen and applied Ethanol-Ethyl cinnamate (EtOH-ECi) based optical clearing to detect both spheroid core and invasion zone by subcellular-resolved 3D microscopy. When subjected to a single dose of irradiation (4 Gy), we detected significant cell survival in the invasion zone. By physical separation of the core and invasion zone, we identified differentially regulated genes preferentially engaged in invading cells controlling cell division, repair, and survival. This imaging-based 3D invasion culture may be useful for the analysis of complex therapy-response patterns in cancer cells in drug discovery and invasion-associated resistance development.

The heterogeneity and evolution of AML blasts can render therapeutic interventions ineffective in a yet poorly understood patient-specific manner. To gain insight into the clonal heterogeneity of diagnosis (Dx) and relapse (Re) pairs, we employed whole-exome sequencing and single-cell RNA-seq to longitudinally profile two t(8;21) (AML1-ETO = RUNX1-RUNX1T1), and four FLT3-ITD AML cases. The single cell RNA data underpinned the tumor heterogeneity amongst patient blasts. The Dx-Re transcriptomes of high risk FLT3-ITD pairs formed a continuum from extensively changed in the absence of significantly mutational changes in AML-associated genes to rather similar Dx-Re pair of an intermediate risk FLT3-ITD. In one high risk FLT3-ITD pair, a pathway switched from an AP-1 regulated network in Dx to mTOR signaling in Re. The distinct AML1-ETO pairs comprise clusters that share genes related to hematopoietic stem cell maintenance and cell migration suggesting that the Re leukemic stem cell-like (LSC-like) cells probably evolved from the Dx LSC-like cells. In summary, our study revealed a continuum from drastic transcriptional changes to extensive similarities between respective Dx-Re pairs that are poorly explained by the well-established model of clonal evolution. Our results suggest alternative and currently unappreciated and unexplored mechanisms leading to therapeutic resistance and AML recurrence.