Duy, PQ;Weise, SC;Marini, C;Li, XJ;Liang, D;Dahl, PJ;Ma, S;Spajic, A;Dong, W;Juusola, J;Kiziltug, E;Kundishora, AJ;Koundal, S;Pedram, MZ;Torres-Fernández, LA;Händler, K;De Domenico, E;Becker, M;Ulas, T;Juranek, SA;Cuevas, E;Hao, LT;Jux, B;Sousa, AMM;Liu, F;Kim, SK;Li, M;Yang, Y;Takeo, Y;Duque, A;Nelson-Williams, C;Ha, Y;Selvaganesan, K;Robert, SM;Singh, AK;Allington, G;Furey, CG;Timberlake, AT;Reeves, BC;Smith, H;Dunbar, A;DeSpenza, T;Goto, J;Marlier, A;Moreno-De-Luca, A;Yu, X;Butler, WE;Carter, BS;Lake, EMR;Constable, RT;Rakic, P;Lin, H;Deniz, E;Benveniste, H;Malvankar, NS;Estrada-Veras, JI;Walsh, CA;Alper, SL;Schultze, JL;Paeschke, K;Doetzlhofer, A;Wulczyn, FG;Jin, SC;Lifton, RP;Sestan, N;Kolanus, W;Kahle, KT;
PMID: 35379995 | DOI: 10.1038/s41593-022-01043-3
Hydrocephalus, characterized by cerebral ventricular dilatation, is routinely attributed to primary defects in cerebrospinal fluid (CSF) homeostasis. This fosters CSF shunting as the leading reason for brain surgery in children despite considerable disease heterogeneity. In this study, by integrating human brain transcriptomics with whole-exome sequencing of 483 patients with congenital hydrocephalus (CH), we found convergence of CH risk genes in embryonic neuroepithelial stem cells. Of all CH risk genes, TRIM71/lin-41 harbors the most de novo mutations and is most specifically expressed in neuroepithelial cells. Mice harboring neuroepithelial cell-specific Trim71 deletion or CH-specific Trim71 mutation exhibit prenatal hydrocephalus. CH mutations disrupt TRIM71 binding to its RNA targets, causing premature neuroepithelial cell differentiation and reduced neurogenesis. Cortical hypoplasia leads to a hypercompliant cortex and secondary ventricular enlargement without primary defects in CSF circulation. These data highlight the importance of precisely regulated neuroepithelial cell fate for normal brain-CSF biomechanics and support a clinically relevant neuroprogenitor-based paradigm of CH.
Gao, L;Liu, S;Gou, L;Hu, Y;Liu, Y;Deng, L;Ma, D;Wang, H;Yang, Q;Chen, Z;Liu, D;Qiu, S;Wang, X;Wang, D;Wang, X;Ren, B;Liu, Q;Chen, T;Shi, X;Yao, H;Xu, C;Li, CT;Sun, Y;Li, A;Luo, Q;Gong, H;Xu, N;Yan, J;
PMID: 35361973 | DOI: 10.1038/s41593-022-01041-5
Prefrontal cortex (PFC) is the cognitive center that integrates and regulates global brain activity. However, the whole-brain organization of PFC axon projections remains poorly understood. Using single-neuron reconstruction of 6,357 mouse PFC projection neurons, we identified 64 projectome-defined subtypes. Each of four previously known major cortico-cortical subnetworks was targeted by a distinct group of PFC subtypes defined by their first-order axon collaterals. Further analysis unraveled topographic rules of soma distribution within PFC, first-order collateral branch point-dependent target selection and terminal arbor distribution-dependent target subdivision. Furthermore, we obtained a high-precision hierarchical map within PFC and three distinct functionally related PFC modules, each enriched with internal recurrent connectivity. Finally, we showed that each transcriptome subtype corresponds to multiple projectome subtypes found in different PFC subregions. Thus, whole-brain single-neuron projectome analysis reveals organization principles of axon projections within and outside PFC and provides the essential basis for elucidating neuronal connectivity underlying diverse PFC functions.
Annals of the rheumatic diseases
Rousselle, A;Sonnemann, J;Amann, K;Mildner, A;Lodka, D;Kling, L;Bieringer, M;Schneider, U;Leutz, A;Enghard, P;Kettritz, R;Schreiber, A;
PMID: 35418479 | DOI: 10.1136/annrheumdis-2021-221984
Myeloid cell activation by antineutrophil cytoplasmic antibody (ANCA) is pivotal for necrotising vasculitis, including necrotising crescentic glomerulonephritis (NCGN). In contrast to neutrophils, the contribution of classical monocyte (CM) and non-classical monocyte (NCM) remains poorly defined. We tested the hypothesis that CMs contribute to antineutrophil cytoplasmic antibody-associated vasculitis (AAV) and that colony-stimulating factor-2 (CSF2, granulocyte-macrophage colony-stimulating factor (GM-CSF)) is an important monocyte-directed disease modifier.Myeloperoxidase (MPO)-immunised MPO-/- mice were transplanted with haematopoietic cells from wild-type (WT) mice, C-C chemokine receptor 2 (CCR2)-/- mice to abrogate CM, or transcription factor CCAAT-enhancer-binding protein beta (C/EBPβ)-/- mice to reduce NCM, respectively. Monocytes were stimulated with CSF2, and CSF2 receptor subunit beta (CSF2rb)-deficient mice were used. Urinary monocytes and CSF2 were quantified and kidney Csf2 expression was analysed. CSF2-blocking antibody was used in the nephrotoxic nephritis (NTN) model.Compared with WT mice, CCR2-/- chimeric mice showed reduced circulating CM and were protected from NCGN. C/EBPβ-/- chimeric mice lacked NCM but developed NCGN similar to WT chimeric mice. Kidney and urinary CSF2 were upregulated in AAV mice. CSF2 increased the ability of ANCA-stimulated monocytes to generate interleukin-1β and to promote TH17 effector cell polarisation. CSF2rb-/- chimeric mice harboured reduced numbers of kidney TH17 cells and were protected from NCGN. CSF2 neutralisation reduced renal damage in the NTN model. Finally, patients with active AAV displayed increased urinary CM numbers, CSF2 levels and expression of GM-CSF in infiltrating renal cells.CMs but not NCMs are important for inducing kidney damage in AAV. CSF2 is a crucial pathological factor by modulating monocyte proinflammatory functions and thereby TH17 cell polarisation.
Signal transduction and targeted therapy
Song, D;Jia, X;Liu, X;Hu, L;Lin, K;Xiao, T;Qiao, Y;Zhang, J;Dan, J;Wong, C;Hu, C;Sai, K;Gong, S;Sander, M;Shen, R;Chen, X;Xiao, X;Chen, J;Zhang, Y;Wei, C;Xiao, X;Liang, J;Zhang, Q;Hu, J;Zhu, W;Yan, G;Lin, Y;Cai, J;
PMID: 35393389 | DOI: 10.1038/s41392-022-00921-3
Over the last decade, oncolytic virus (OV) therapy has shown its promising potential in tumor treatment. The fact that not every patient can benefit from it highlights the importance for defining biomarkers that help predict patients' responses. As particular self-amplifying biotherapeutics, the anti-tumor effects of OVs are highly dependent on the host factors for viral infection and replication. By using weighted gene co-expression network analysis (WGCNA), we found matrix remodeling associated 8 (MXRA8) is positively correlated with the oncolysis induced by oncolytic virus M1 (OVM). Consistently, MXRA8 promotes the oncolytic efficacy of OVM in vitro and in vivo. Moreover, the interaction of MXRA8 and OVM studied by single-particle cryo-electron microscopy (cryo-EM) showed that MXRA8 directly binds to this virus. Therefore, MXRA8 acts as the entry receptor of OVM. Pan-cancer analysis showed that MXRA8 is abundant in most solid tumors and is highly expressed in tumor tissues compared with adjacent normal ones. Further study in cancer cell lines and patient-derived tumor tissues revealed that the tumor selectivity of OVM is predominantly determined by a combinational effect of the cell membrane receptor MXRA8 and the intracellular factor, zinc-finger antiviral protein (ZAP). Taken together, our study may provide a novel dual-biomarker for precision medicine in OVM therapy.
Signal transduction and targeted therapy
Zhang, L;Chen, D;Song, D;Liu, X;Zhang, Y;Xu, X;Wang, X;
PMID: 35365599 | DOI: 10.1038/s41392-022-00960-w
The combination of spatial transcriptomics (ST) and single cell RNA sequencing (scRNA-seq) acts as a pivotal component to bridge the pathological phenomes of human tissues with molecular alterations, defining in situ intercellular molecular communications and knowledge on spatiotemporal molecular medicine. The present article overviews the development of ST and aims to evaluate clinical and translational values for understanding molecular pathogenesis and uncovering disease-specific biomarkers. We compare the advantages and disadvantages of sequencing- and imaging-based technologies and highlight opportunities and challenges of ST. We also describe the bioinformatics tools necessary on dissecting spatial patterns of gene expression and cellular interactions and the potential applications of ST in human diseases for clinical practice as one of important issues in clinical and translational medicine, including neurology, embryo development, oncology, and inflammation. Thus, clear clinical objectives, designs, optimizations of sampling procedure and protocol, repeatability of ST, as well as simplifications of analysis and interpretation are the key to translate ST from bench to clinic.
Science translational medicine
Mederacke, I;Filliol, A;Affo, S;Nair, A;Hernandez, C;Sun, Q;Hamberger, F;Brundu, F;Chen, Y;Ravichandra, A;Huebener, P;Anke, H;Shi, H;Martínez García de la Torre, RA;Smith, JR;Henderson, NC;Vondran, FWR;Rothlin, CV;Baehre, H;Tabas, I;Sancho-Bru, P;Schwabe, RF;
PMID: 35385339 | DOI: 10.1126/scitranslmed.abe5795
Fibrosis contributes to ~45% of deaths in western countries. In chronic liver disease, fibrosis is a major factor determining outcomes, but efficient antifibrotic therapies are lacking. Although platelet-derived growth factor and transforming growth factor-β constitute key fibrogenic mediators, they do not account for the well-established link between cell death and fibrosis in the liver. Here, we hypothesized that damage-associated molecular patterns (DAMPs) may link epithelial cell death to fibrogenesis in the injured liver. DAMP receptor screening identified purinergic receptor P2Y14 among several candidates as highly enriched in hepatic stellate cells (HSCs), the main fibrogenic cell type of the liver. Conversely, P2Y14 ligands uridine 5'-diphosphate (UDP)-glucose and UDP-galactose were enriched in hepatocytes and were released upon different modes of cell death. Accordingly, ligand-receptor interaction analysis that combined proteomic and single-cell RNA sequencing data revealed P2Y14 ligands and P2Y14 receptor as a link between dying cells and HSCs, respectively. Treatment with P2Y14 ligands or coculture with dying hepatocytes promoted HSC activation in a P2Y14-dependent manner. P2Y14 ligands activated extracellular signal-regulated kinase (ERK) and Yes-associated protein (YAP) signaling in HSCs, resulting in ERK-dependent HSC activation. Global and HSC-selective P2Y14 deficiency attenuated liver fibrosis in multiple mouse models of liver injury. Functional expression of P2Y14 was confirmed in healthy and diseased human liver and human HSCs. In conclusion, P2Y14 ligands and their receptor constitute a profibrogenic DAMP pathway that directly links cell death to fibrogenesis.
Hoch, T;Schulz, D;Eling, N;Gómez, JM;Levesque, MP;Bodenmiller, B;
PMID: 35363540 | DOI: 10.1126/sciimmunol.abk1692
Intratumoral immune cells are crucial for tumor control and antitumor responses during immunotherapy. Immune cell trafficking into tumors is mediated by binding of specific immune cell receptors to chemokines, a class of secreted chemotactic cytokines. To broadly characterize chemokine expression and function in melanoma, we used multiplexed mass cytometry-based imaging of protein markers and RNA transcripts to analyze the chemokine landscape and immune infiltration in metastatic melanoma samples. Tumors that lacked immune infiltration were devoid of most of the profiled chemokines and exhibited low levels of antigen presentation and markers of inflammation. Infiltrated tumors were characterized by expression of multiple chemokines. CXCL9 and CXCL10 were often localized in patches associated with dysfunctional T cells expressing the B lymphocyte chemoattractant CXCL13. In tumors with B cells but no B cell follicles, T cells were the sole source of CXCL13, suggesting that T cells play a role in B cell recruitment and potentially in B cell follicle formation. B cell patches and follicles were also enriched with TCF7+ naïve-like T cells, a cell type that is predictive of response to immune checkpoint blockade. Our data highlight the strength of targeted RNA and protein codetection to analyze tumor immune microenvironments based on chemokine expression and suggest that the formation of tertiary lymphoid structures may be accompanied by naïve and naïve-like T cell recruitment, which may contribute to antitumor activity.
Zhang, MM;Geng, AQ;Chen, K;Wang, J;Wang, P;Qiu, XT;Gu, JX;Fan, HW;Zhu, DY;Yang, SM;Chen, QY;Zhou, ZX;Fan, BY;Bai, Y;Xing, KK;Feng, JM;Wang, JD;Chen, Y;Lu, YC;Liang, Y;Cao, P;Kaang, BK;Zhuo, M;Li, YQ;Chen, T;
PMID: 35443154 | DOI: 10.1016/j.neuron.2022.03.030
Empathic pain has attracted the interest of a substantial number of researchers studying the social transfer of pain in the sociological, psychological, and neuroscience fields. However, the neural mechanism of empathic pain remains elusive. Here, we establish a long-term observational pain model in mice and find that glutamatergic projection from the insular cortex (IC) to the basolateral amygdala (BLA) is critical for the formation of observational pain. The selective activation or inhibition of the IC-BLA projection pathway strengthens or weakens the intensity of observational pain, respectively. The synaptic molecules are screened, and the upregulated synaptotagmin-2 and RIM3 are identified as key signals in controlling the increased synaptic glutamate transmission from the IC to the BLA. Together, these results reveal the molecular and synaptic mechanisms of a previously unidentified neural pathway that regulates observational pain in mice.
Beyersdorf, JP;Bawage, S;Iglesias, N;Peck, HE;Hobbs, RA;Wroe, JA;Zurla, C;Gersbach, CA;Santangelo, PJ;
PMID: 35357116 | DOI: 10.1021/acsnano.1c10631
Programmable control of gene expression via nuclease-null Cas9 fusion proteins has enabled the engineering of cellular behaviors. Here, both transcriptional and epigenetic gene activation via synthetic mRNA and lipid nanoparticle delivery was demonstrated in vivo. These highly efficient delivery strategies resulted in high levels of activation in multiple tissues. Finally, we demonstrate durable gene activation in vivo via transient delivery of a single dose of a gene activator that combines VP64, p65, and HSF1 with a SWI/SNF chromatin remodeling complex component SS18, representing an important step toward gene-activation-based therapeutics. This induced sustained gene activation could be inhibited via mRNA-encoded AcrIIA4, further improving the safety profile of this approach.
Advanced drug delivery reviews
Vervaeke, P;Borgos, SE;Sanders, NN;Combes, F;
PMID: 35351470 | DOI: 10.1016/j.addr.2022.114236
The success of the messenger RNA-based COVID-19 vaccines of Moderna and Pfizer/BioNTech marks the beginning of a new chapter in modern medicine. However, the rapid rise of mRNA therapeutics has resulted in a regulatory framework that is somewhat lagging. The current guidelines either do not apply, do not mention RNA therapeutics, or do not have widely accepted definitions. This review describes the guidelines for preclinical biodistribution studies of mRNA/siRNA therapeutics and highlights the relevant differences for mRNA vaccines. We also discuss the role of in vivo RNA imaging techniques and other assays to fulfill and/or complement the regulatory requirements. Specifically, quantitative whole-body autoradiography, microautoradiography, mass spectrometry-based assays, hybridization techniques (FISH, bDNA), PCR-based methods, in vivo fluorescence imaging, and in vivo bioluminescence imaging, are discussed. We conclude that this new and rapidly evolving class of medicines demands a multi-layered approach to fully understand its biodistribution and in vivo characteristics.
Kabir, MF;Karami, AL;Cruz-Acuña, R;Klochkova, A;Saxena, R;Mu, A;Murray, MG;Cruz, J;Fuller, AD;Clevenger, MH;Chitrala, KN;Tan, Y;Keith, K;Madzo, J;Huang, H;Jelinek, J;Karakasheva, T;Hamilton, KE;Muir, AB;Tétreault, MP;Whelan, KA;
PMID: 35443762 | DOI: 10.1038/s41467-022-29747-x
Although morphologic progression coupled with expression of specific molecular markers has been characterized along the esophageal squamous differentiation gradient, the molecular heterogeneity within cell types along this trajectory has yet to be classified at the single cell level. To address this knowledge gap, we perform single cell RNA-sequencing of 44,679 murine esophageal epithelial, to identify 11 distinct cell populations as well as pathways alterations along the basal-superficial axis and in each individual population. We evaluate the impact of aging upon esophageal epithelial cell populations and demonstrate age-associated mitochondrial dysfunction. We compare single cell transcriptomic profiles in 3D murine organoids and human esophageal biopsies with that of murine esophageal epithelium. Finally, we employ pseudotemporal trajectory analysis to develop a working model of cell fate determination in murine esophageal epithelium. These studies provide comprehensive molecular perspective on the cellular heterogeneity of murine esophageal epithelium in the context of homeostasis and aging.
van Gurp, L;Fodoulian, L;Oropeza, D;Furuyama, K;Bru-Tari, E;Vu, AN;Kaddis, JS;Rodríguez, I;Thorel, F;Herrera, PL;
PMID: 35440614 | DOI: 10.1038/s41467-022-29588-8
Generation of surrogate cells with stable functional identities is crucial for developing cell-based therapies. Efforts to produce insulin-secreting replacement cells to treat diabetes require reliable tools to assess islet cellular identity. Here, we conduct a thorough single-cell transcriptomics meta-analysis to identify robustly expressed markers used to build genesets describing the identity of human α-, β-, γ- and δ-cells. These genesets define islet cellular identities better than previously published genesets. We show their efficacy to outline cell identity changes and unravel some of their underlying genetic mechanisms, whether during embryonic pancreas development or in experimental setups aiming at developing glucose-responsive insulin-secreting cells, such as pluripotent stem-cell differentiation or in adult islet cell reprogramming protocols. These islet cell type-specific genesets represent valuable tools that accurately benchmark gain and loss in islet cell identity traits.
