DestVI identifies continuums of cell types in spatial transcriptomics data.

Most spatial transcriptomics technologies are limited by their resolution, with spot sizes larger than that of a single cell. Although joint analysis with single-cell RNA sequencing can alleviate this problem, current methods are limited to assessing discrete cell types, revealing the proportion of cell types inside each spot. To identify continuous variation of the transcriptome within cells of the same type, we developed Deconvolution of Spatial Transcriptomics profiles using Variational Inference (DestVI). Using simulations, we demonstrate that DestVI outperforms existing methods for estimating gene expression for every cell type inside every spot. Applied to a study of infected lymph nodes and of a mouse tumor model, DestVI provides high-resolution, accurate spatial characterization of the cellular organization of these tissues and identifies cell-type-specific changes in gene expression between different tissue regions or between conditions. DestVI is available as part of the open-source software package scvi-tools ( https://scvi-tools.org ).

Liver fibrosis affects the targeting properties of drug delivery systems to macrophage subsets in vivo.

Myeloid immune cells promote inflammation and fibrosis in chronic liver diseases. Drug delivery systems, such as polymers, liposomes and microbubbles, efficiently target myeloid cells in healthy liver, but their targeting properties in hepatic fibrosis remain elusive. We therefore studied the biodistribution of three intravenously injected carrier material, i.e. 10 nm poly(N-(2-hydroxypropyl)methacrylamide) polymers, 100 nm PEGylated liposomes and 2000 nm poly(butyl cyanoacrylate) microbubbles, in two fibrosis models in immunocompetent mice. While whole-body imaging confirmed preferential hepatic uptake even after induction of liver fibrosis, flow cytometry and immunofluorescence analysis revealed markedly decreased carrier uptake by liver macrophage subsets in fibrosis, particularly for microbubbles and polymers. Importantly, carrier uptake co-localized with immune infiltrates in fibrotic livers, corroborating the intrinsic ability of the carriers to target myeloid cells in areas of inflammation. Of the tested carrier systems liposomes had the highest uptake efficiency among hepatic myeloid cells, but the lowest specificity for cellular subsets. Hepatic fibrosis affected carrier uptake in liver and partially in spleen, but not in other tissues (blood, bone marrow, lung, kidney). In conclusion, while drug carrier systems target distinct myeloid cell populations in diseased and healthy livers, hepatic fibrosis profoundly affects their targeting efficiency, supporting the need to adapt nanomedicine-based approaches in chronic liver disease.

The CCR2+ Macrophage Subset Promotes Pathogenic Angiogenesis for Tumor Vascularization in Fibrotic Livers.

BACKGROUND & AIMS: Hepatocellular carcinoma (HCC) typically arises in fibrotic or cirrhotic livers, which are characterized by pathogenic angiogenesis. Myeloid immune cells, specifically tumor-associated macrophages (TAMs), may represent potential novel therapeutic targets in HCC, complementing current ablative or immune therapies. However, the detailed functions of TAM subsets in hepatocarcinogenesis have remained obscure. METHODS: TAM subsets were analyzed in-depth in human HCC samples and a combined fibrosis-HCC mouse model, established by i.p. injection with diethylnitrosamine after birth and repetitive carbon tetrachloride (CCl4) treatment for 16 weeks. Based on comprehensively phenotyping TAM subsets (fluorescence-activated cell sorter, transcriptomics) in mice, the function of CCR2+ TAM was assessed by a pharmacologic chemokine inhibitor. Angiogenesis was evaluated by contrast-enhanced micro-computed tomography and histology. RESULTS: We show that human CCR2+ TAM accumulate at the highly vascularized HCC border and express the inflammatory marker S100A9, whereas CD163+ immune-suppressive TAM accrue in the HCC center. In the fibrosis-cancer mouse model, we identified 3 major hepatic myeloid cell populations with distinct messenger RNA profiles, of which CCR2+ TAM particularly showed activated inflammatory and angiogenic pathways. Inhibiting CCR2+ TAM infiltration using a pharmacologic chemokine CCL2 antagonist in the fibrosis-HCC model significantly reduced pathogenic vascularization and hepatic blood volume, alongside attenuated tumor volume. CONCLUSIONS: The HCC microenvironment in human patients and mice is characterized by functionally distinct macrophage populations, of which the CCR2+ inflammatory TAM subset has pro-angiogenic properties. Understanding the functional differentiation of myeloid cell subsets in chronically inflamed liver may provide novel opportunities for modulating hepatic macrophages to inhibit tumor-promoting pathogenic angiogenesis.

M-CSF and GM-CSF Receptor Signaling Differentially Regulate Monocyte Maturation and Macrophage Polarization in the Tumor Microenvironment.

Tumors contain a heterogeneous myeloid fraction comprised of discrete MHC-II(hi) and MHC-II(lo) tumor-associated macrophage (TAM) subpopulations that originate from Ly6C(hi) monocytes. However, the mechanisms regulating the abundance and phenotype of distinct TAM subsets remain unknown. Here, we investigated the role of macrophage colony-stimulating factor (M-CSF) in TAM differentiation and polarization in different mouse tumor models. We demonstrate that treatment of tumor-bearing mice with a blocking anti-M-CSFR monoclonal antibody resulted in a reduction of mature TAMs due to impaired recruitment, extravasation, proliferation, and maturation of their Ly6C(hi) monocytic precursors. M-CSFR signaling blockade shifted the MHC-II(lo)/MHC-II(hi) TAM balance in favor of the latter as observed by the preferential differentiation of Ly6C(hi) monocytes into MHC-II(hi) TAMs. In addition, the genetic and functional signatures of MHC-II(lo) TAMs were downregulated upon M-CSFR blockade, indicating that M-CSFR signaling shapes the MHC-II(lo) TAM phenotype. Conversely, granulocyte macrophage (GM)-CSFR had no effect on the mononuclear tumor infiltrate or relative abundance of TAM subsets. However, GM-CSFR signaling played an important role in fine-tuning the MHC-II(hi) phenotype. Overall, our data uncover the multifaceted and opposing roles of M-CSFR and GM-CSFR signaling in governing the phenotype of macrophage subsets in tumors, and provide new insight into the mechanism of action underlying M-CSFR blockade.