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Van Gogh-like 2 (Vangl2), a core planar cell polarity component, plays an important role in polarized cellular and tissue morphology induction, growth development, and cancer. However, its role in regulating inflammatory responses remains elusive. Here, we report that Vangl2 is upregulated in patients with sepsis and identify Vangl2 as a negative regulator of The nuclear factor-kappaB (NF-κB) signaling by regulating the protein stability and activation of the core transcription component p65. Mice with myeloid-specific deletion of Vangl2 ( Vangl2 ΔM ) are hypersusceptible to lipopolysaccharide (LPS)-induced septic shock. Vangl2-deficient myeloid cells exhibit enhanced phosphorylation and expression of p65, therefore, promoting the secretion of proinflammatory cytokines after LPS stimulation. Mechanistically, NF-κB signaling-induced-Vangl2 recruits E3 ubiquitin ligase PDLIM2 to catalyze K63-linked ubiquitination on p65, which serves as a recognition signal for cargo receptor NDP52-mediated selective autophagic degradation. Taken together, these findings demonstrate Vangl2 as a suppressor of NF-κB-mediated inflammation and provide insights into the crosstalk between autophagy and inflammatory diseases., Competing Interests: JL, JZ, HJ, ZH, YZ, LH, JY, YX, DW, HL, KZ, PT, QX, ZS, CP, XB, XY No competing interests declared, (© 2023, Lu, Zhang, Jiang et al.)

Introduction: Type I interferon (IFN-I, IFN-α/β), precisely controlled by multiple regulators, including suppressor of cytokine signaling 1 (SOCS1), is critical for host defense against pathogens. However, the impact of IFN-α/β on malaria parasite infections, beneficial or detrimental, remains controversial., Objectives: The contradictory results are suspected to arise from differences in parasite species and host genetic backgrounds. To date, no prior study has employed a comparative approach utilizing two parasite models to investigate the underlying mechanisms of IFN-I response. Moreover, whether and how SOCS1 involves in the distinct IFN-α/β dynamics is still unclear., Methods: Here we perform single-cell RNA sequencing analyses (scRNA-seq) to dissect the dynamics of IFN-α/β responses against P. yoelii 17XL (17XL) and P. berghei ANKA (PbANKA) infections; conduct flow cytometry analysis and functional depletion to identify key cellular players induced by IFN-I; and establish mathematical models to explore the mechanisms underlying the differential IFN-I dynamics regulated by SOCS1., Results: 17XL stimulates an early protective but insufficient toll-like receptor 7 (TLR7)-interferon regulatory factor 7 (IRF7)-dependent IFN-α/β response, resulting in CD11a hi CD49d hi CD4 + T cell activation to enhance anti-malarial immunity. On the contrary, a late IFN-α/β induction through toll-like receptor 9 (TLR9)-IRF7/ stimulator of interferon genes (STING)- interferon regulatory factor 3 (IRF3) dependent pathways expands programmed cell death protein 1 (PD-1) + CD8 + T cells and impairs host immunity during PbANKA infection. Furthermore, functional assay and mathematical modeling show that SOCS1 significantly suppresses IFN-α/β production via negative feedback and incoherent feed-forward loops (I1-FFL). Additionally, differential activation patterns of various transcriptional factors (TFs) synergistically regulate the distinct IFN-I responses., Conclusion: This study reveals the dual functions of IFN-I in anti-malarial immunity: Early IFN-α/β enhances immune responses against Plasmodium infection by promoting CD11a hi CD49d hi CD4 + T cell, while late IFN-α/β suppresses these response by expanding PD-1 + CD8 + T cells. Moreover, both the SOCS1-related network motifs and TFs activation patterns contribute to determine distinct dynamics of IFN-I responses. Hence, our findings suggest therapies targeting SOCS1- or TFs-regulated IFN-I dynamics could be an efficacious approach for preventing malaria and enhancing vaccine efficacy., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Production and hosting by Elsevier B.V.)

Toxoplasma gondii (T. gondii)-associated polymorphic effector proteins are crucial in parasite development and regulating host anti-T. gondii immune responses. However, the mechanism remains obscure. Here, it is shown that Toxoplasma effector dense granules 4 (GRA4) restricts host IFN-I activation. Infection with Δgra4 mutant T. gondii strain induces stronger IFN-I responses and poses a severe threat to host health. Mechanistically, GRA4 binds to phosphorylated TBK1 to promote TRIM27-catalyzed K48-ubiquitination at Lys251/Lys372 residues, which enhances its recognition by autophagy receptor p62, ultimately leading to TBK1 autophagic degradation. Furthermore, an avirulent Δgra4 strain (ME49Δompdc/gra4) is constructed for tumor immunotherapy due to its ability to enhance IFN-I production. Earlier vaccination with ME49Δompdc/gra4 confers complete host resistance to the tumor compared with the classical ME49Δompdc treatment. Notably, ME49Δompdc/gra4 vaccination induces a specific CD64 + MAR-1 + CD11b + dendritic cell subset, thereby enhancing T cell anti-tumor responses. Overall, these findings identify the negative role of T. gondii GRA4 in modulating host IFN-I signaling and suggest that GRA4 can be a potential target for the development of T. gondii vaccines and tumor immunotherapy., (© 2024 The Author(s). Advanced Science published by Wiley‐VCH GmbH.)

Comprehensive single-cell metabolic profiling is critical for revealing phenotypic heterogeneity and elucidating the molecular mechanisms underlying biological processes. However, single-cell metabolomics remains challenging because of the limited metabolite coverage and inability to discriminate isomers. Herein, we establish a single-cell metabolomics platform for in-depth organic mass cytometry. Extended single-cell analysis time guarantees sufficient MS/MS acquisition for metabolite identification and the isomers discrimination while online sampling ensures the high-throughput of the method. The largest number of identified metabolites (approximately 600) are achieved in single cells and fine subtyping of MCF-7 cells is first demonstrated by an investigation on the differential levels of 3-hydroxybutanoic acid among clusters. Single-cell transcriptome analysis reveals differences in the expression of 3-hydroxybutanoic acid downstream antioxidative stress genes, such as metallothionein 2 (MT2A), while a fluorescence-activated cell sorting assay confirms the positive relationship between 3-hydroxybutanoic acid and target proteins; these results suggest that the heterogeneity of 3-hydroxybutanoic acid provides cancer cells with different ability to resist surrounding oxidative stress. Our method paves the way for deep single-cell metabolome profiling and investigations on the physiological and pathological processes that occur during cancer., (© 2024. The Author(s).)

Hypertrophic cardiomyopathy (HCM) is a common inherited cardiovascular disease, which can cause heart failure and lead to death. In this study, we performed high-resolution single-cell RNA-sequencing of 2115 individual cardiomyocytes obtained from HCM patients and normal controls. Signature up- and down-regulated genes in HCM were identified by integrative analysis across 37 patients and 41 controls from our data and published human single-cell and single-nucleus RNA-seq datasets, which were further classified into gene modules by single-cell co-expression analysis. Using our high-resolution dataset, we also investigated the heterogeneity among individual cardiomyocytes and revealed five distinct clusters within HCM cardiomyocytes. Interestingly, we showed that some extracellular matrix (ECM) genes were up-regulated in the HCM cardiomyocytes, suggesting that they play a role in cardiac remodelling. Taken together, our study comprehensively profiled the transcriptomic programs of HCM cardiomyocytes and provided insights into molecular mechanisms underlying the pathogenesis of HCM., (© 2023 The Authors. Cell Proliferation published by Beijing Institute for Stem Cell and Regenerative Medicine and John Wiley & Sons Ltd.)

Macroautophagy/autophagy plays an important role in regulating cellular homeostasis and influences the pathogenesis of degenerative diseases. Tendinopathy is characterized by tendon degeneration and inflammation. However, little is known about the role of selective autophagy in tendinopathy. Here, we find that pristimerin (PM), a quinone methide triterpenoid, is more effective in treating tendinopathy than the first-line drug indomethacin. PM inhibits the AIM2 inflammasome and alleviates inflammation during tendinopathy by promoting the autophagic degradation of AIM2 through a PYCARD/ASC-dependent manner. A mechanistic study shows that PM enhances the K63-linked ubiquitin chains of PYCARD/ASC at K158/161, which serves as a recognition signal for SQSTM1/p62-mediated autophagic degradation of the AIM2-PYCARD/ASC complex. We further identify that PM binds the Cys53 site of deubiquitinase USP50 through the Michael-acceptor and blocks the binding of USP50 to PYCARD/ASC, thereby reducing USP50-mediated cleavage of K63-linked ubiquitin chains of PYCARD/ASC. Finally, PM treatment in vivo generates an effect comparable to inflammasome deficiency in alleviating tendinopathy. Taken together, these findings demonstrate that PM alleviates the progression of tendinopathy by modulating AIM2-PYCARD/ASC stability via SQSTM1/p62-mediated selective autophagic degradation, thus providing a promising autophagy-based therapeutic for tendinopathy. Abbreviations: 3-MA: 3-methyladenine; AIM2: absent in melanoma 2; AT: Achilles tenotomy; ATP: adenosine triphosphate; BMDMs: bone marrow-derived macrophages; CHX: cycloheximide; Col3a1 : collagen, type III, alpha 1; CQ: chloroquine; Cys: cysteine; DARTS: drug affinity responsive target stability; DTT: dithiothreitol; DUB: deubiquitinase; gDNA: genomic DNA; GSH: glutathione; His: histidine; IL1B/IL-1β: interleukin 1 beta; IND: indomethacin; IP: immunoprecipitation; LPS: lipopolysaccharide; MMP: mitochondrial membrane potential; NLRP3: NLR family, pyrin domain containing 3; PM: pristimerin; PYCARD/ASC: PYD and CARD domain containing; SN: supernatants; SOX9: SRY (sex determining region Y)-box 9; SQSTM1: sequestosome 1; Tgfb : transforming growth factor, beta; TIMP3: tissue inhibitor of metalloproteinase 3; TNMD: tenomodulin; TRAF6: TNF receptor-associated factor 6; Ub: ubiquitin; USP50: ubiquitin specific peptidase 50; WCL: whole cell lysates.

Hepatic ischemia reperfusion injury (IRI) is a risk factor for early graft nonfunction and graft rejection after liver transplantation (LT). The process of liver IRI involves inflammatory response, oxidative stress, apoptosis and other pathophysiological processes. So far, there is still a lack of effective drugs to ameliorate liver IRI. Trans-anethole (TA) is an aromatic compound. Many medications as well as natural foods contain TA. TA has multiple effects such as anti-inflammation, anti-oxidative stress and anti-apoptosis. However, the mechanism of TA pretreatment in liver IRI is unclear. The mice hepatic IRI model was constructed after gavage pretreatment with TA (10 mg/kg, 20 mg/kg, 40 mg/kg) for 7 consecutive days. Our study confirmed that TA pretreatment significantly improve liver function and reduce serum AST, ALT in hepatic IRI. HE staining showed that TA pretreatment alleviated liver injury. Meanwhile, TA (20 mg/kg) pretreatment attenuated hepatocyte apoptosis in hepatic IRI. In addition, TA (20 mg/kg) pretreatment reduced the inflammatory factors TNF-α, IL-6 and infiltration of CD11b positive cells in liver tissues during hepatic IRI in mice. TA pretreatment also alleviated oxidative stress in mice hepatic IRI. Our study further indicated that TA pretreatment attenuated mice hepatic IRI through inhibiting NLRP3 inflammasome activation via regulation of soluble epoxide hydrolase (sEH). This study provides a novel and effective potential drug with few side effects for easing liver IRI., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

The high-order three-dimensional (3D) organization of regulatory genomic elements provides a topological basis for gene regulation, but it remains unclear how multiple regulatory elements across the mammalian genome interact within an individual cell. To address this, herein, we developed scNanoHi-C, which applies Nanopore long-read sequencing to explore genome-wide proximal high-order chromatin contacts within individual cells. We show that scNanoHi-C can reliably and effectively profile 3D chromatin structures and distinguish structure subtypes among individual cells. This method could also be used to detect genomic variations, including copy-number variations and structural variations, as well as to scaffold the de novo assembly of single-cell genomes. Notably, our results suggest that extensive high-order chromatin structures exist in active chromatin regions across the genome, and multiway interactions between enhancers and their target promoters were systematically identified within individual cells. Altogether, scNanoHi-C offers new opportunities to investigate high-order 3D genome structures at the single-cell level., (© 2023. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Stringent control of the type I interferon (IFN-I) signaling is critical for host immune defense against infectious diseases, yet the molecular mechanisms that regulate this pathway remain elusive. Here, we show that Src homology 2 containing inositol phosphatase 1 (SHIP1) suppresses IFN-I signaling by promoting IRF3 degradation during malaria infection. Genetic ablation of Ship1 in mice leads to high levels of IFN-I and confers resistance to Plasmodium yoelii nigeriensis ( P.y. ) N67 infection. Mechanistically, SHIP1 promotes the selective autophagic degradation of IRF3 by enhancing K63-linked ubiquitination of IRF3 at lysine 313, which serves as a recognition signal for NDP52-mediated selective autophagic degradation. In addition, SHIP1 is downregulated by IFN-I-induced miR-155-5p upon P.y . N67 infection and severs as a feedback loop of the signaling crosstalk. This study reveals a regulatory mechanism between IFN-I signaling and autophagy, and verifies SHIP1 can be a potential target for therapeutic intervention against malaria and other infectious diseases. IMPORTANCE Malaria remains a serious disease affecting millions of people worldwide. Malaria parasite infection triggers tightly controlled type I interferon (IFN-I) signaling that plays a critical role in host innate immunity; however, the molecular mechanisms underlying the immune responses are still elusive. Here, we discover a host gene [Src homology 2-containing inositol phosphatase 1 (SHIP1)] that can regulate IFN-I signaling by modulating NDP52-mediated selective autophagic degradation of IRF3 and significantly affect parasitemia and resistance of Plasmodium -infected mice. This study identifies SHIP1 as a potential target for immunotherapies in malaria and highlights the crosstalk between IFN-I signaling and autophagy in preventing related infectious diseases. SHIP1 functions as a negative regulator during malaria infection by targeting IRF3 for autophagic degradation., Competing Interests: The authors declare no conflict of interest.

Although somatic cells can be reprogrammed to pluripotent stem cells (PSCs) with pure chemicals, authentic pluripotency of chemically induced pluripotent stem cells (CiPSCs) has never been achieved through tetraploid complementation assay. Spontaneous reprogramming of spermatogonial stem cells (SSCs) was another non-transgenic way to obtain PSCs, but this process lacks mechanistic explanation. Here, we reconstructed the trajectory of mouse SSC reprogramming and developed a five-chemical combination, boosting the reprogramming efficiency by nearly 80- to 100-folds. More importantly, chemical induced germline-derived PSCs (5C-gPSCs), but not gPSCs and chemical induced pluripotent stem cells, had authentic pluripotency, as determined by tetraploid complementation. Mechanistically, SSCs traversed through an inverted pathway of in vivo germ cell development, exhibiting the expression signatures and DNA methylation dynamics from spermatogonia to primordial germ cells and further to epiblasts. Besides, SSC-specific imprinting control regions switched from biallelic methylated states to monoallelic methylated states by imprinting demethylation and then re-methylation on one of the two alleles in 5C-gPSCs, which was apparently distinct with the imprinting reprogramming in vivo as DNA methylation simultaneously occurred on both alleles. Our work sheds light on the unique regulatory network underpinning SSC reprogramming, providing insights to understand generic mechanisms for cell-fate decision and epigenetic-related disorders in regenerative medicine., (©The Author(s) 2022. Published by Oxford University Press on behalf of Higher Education Press.)

Stringent control of type I interferon (IFN-I) signaling is critical to potent innate immune responses against viral infection, yet the underlying molecular mechanisms are still elusive. Here, we found that Van Gogh-like 2 (VANGL2) acts as an IFN-inducible negative feedback regulator to suppress IFN-I signaling during vesicular stomatitis virus (VSV) infection. Mechanistically, VANGL2 interacted with TBK1 and promoted the selective autophagic degradation of TBK1 via K48-linked polyubiquitination at Lys 372 by the E3 ligase TRIP, which serves as a recognition signal for the cargo receptor OPTN. Furthermore, myeloid-specific deletion of VANGL2 in mice showed enhanced IFN-I production against VSV infection and improved survival. In general, these findings revealed a negative feedback loop of IFN-I signaling through the VANGL2-TRIP-TBK1-OPTN axis and highlighted the cross-talk between IFN-I and autophagy in preventing viral infection. VANGL2 could be a potential clinical therapeutic target for viral infectious diseases, including COVID-19.

During embryo development, DNA methylation is established by DNMT3A/3B and subsequently maintained by DNMT1. While much research has been done in this field, the functional significance of DNA methylation in embryogenesis remains unknown. Here, we establish a system of simultaneous inactivation of multiple endogenous genes in zygotes through screening for base editors that can efficiently introduce a stop codon. Embryos with mutations in Dnmts and/or Tets can be generated in one step with IMGZ. Dnmt-null embryos display gastrulation failure at E7.5. Interestingly, although DNA methylation is absent, gastrulation-related pathways are down-regulated in Dnmt-null embryos. Moreover, DNMT1, DNMT3A, and DNMT3B are critical for gastrulation, and their functions are independent of TET proteins. Hypermethylation can be sustained by either DNMT1 or DNMT3A/3B at some promoters, which are related to the suppression of miRNAs. The introduction of a single mutant allele of six miRNAs and paternal IG-DMR partially restores primitive streak elongation in Dnmt-null embryos. Thus, our results unveil an epigenetic correlation between promoter methylation and suppression of miRNA expression for gastrulation and demonstrate that IMGZ can accelerate deciphering the functions of multiple genes in vivo., (© 2023. The Author(s).)

Background: Intestinal transplantation (IT) has become an important procedure for the treatment of irreversible intestinal failure. However, IT is extremely vulnerable to ischemia-reperfusion injury (IRI). Due to the limitations of static cold storage (SCS), hypothermic machine perfusion (HMP) is rapidly gaining popularity. In this study, the intestinal HMP system is established and HMP is compared with SCS., Methods: An intestinal HMP system was built. Ten miniature pigs were randomly divided into the HMP and SCS groups, and their intestines were perfused using the HMP device and SCS, respectively, followed by orthotopic auto-transplantation. Analysis was done on the grafts between the two groups., Results: Operation success rates of the surgery were 100% in both groups. The 7-day survival rate was 100% in the HMP group, which was significantly higher than that of the SCS group (20%, P < 0.05). The pathological results showed that fewer injuries of grafts were in the HMP group. Endotoxin (ET), IL-1, IL-6, IFN-γ and TNF-α levels in the HMP group were significantly lower than in the SCS group (P<0.05), whereas IL-10 levels were significantly higher (P<0.05).The intestinal expression levels of ZO-1 and Occludin were higher in the HMP group compared to the SCS group, whereas Toll-like receptor 4 (TLR4), nuclear factor kappa B (NFκB), and caspase-3 were lower., Conclusions: In this study, we established a stable intestinal HMP system and demonstrated that HMP could significantly alleviate intestinal IRI and improve the outcome after IT., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Hou, Yang, Lu, Shi, Chen, Chen, Wang and Liu.)

Innate immunity acts as the first line of defense against pathogen invasion. During Toxoplasma gondii infection, multiple innate immune sensors are activated by invading microbes or pathogen-associated molecular patterns (PAMPs). However, how inflammasome is activated and its regulatory mechanisms during T. gondii infection remain elusive. Here, we showed that the infection of PRU, a lethal type II T. gondii strain, activates inflammasome at the early stage of infection. PRU tachyzoites, RNA and soluble tachyzoite antigen (STAg) mainly triggered the NLRP3 inflammasome, while PRU genomic DNA (gDNA) specially activated the AIM2 inflammasome. Furthermore, mice deficient in AIM2, NLRP3, or caspase-1/11 were more susceptible to T. gondii PRU infection, and the ablation of inflammasome signaling impaired antitoxoplasmosis immune responses by enhancing type I interferon (IFN-I) production. Blockage of IFN-I receptor fulfilled inflammasome-deficient mice competent immune responses as WT mice. Moreover, we have identified that the suppressor of cytokine signaling 1 (SOCS1) is a key negative regulator induced by inflammasome-activated IL-1β signaling and inhibits IFN-I production by targeting interferon regulatory factor 3 (IRF3). In general, our study defines a novel protective role of inflammasome activation during toxoplasmosis and identifies a critical regulatory mechanism of the cross talk between inflammasome and IFN-I signaling for understanding infectious diseases. IMPORTANCE As a key component of innate immunity, inflammasome is critical for host antitoxoplasmosis immunity, but the underlying mechanisms are still elusive. In this study, we found that inflammasome signaling was activated by PAMPs of T. gondii, which generated a protective immunity against T. gondii invasion by suppressing type I interferon (IFN-I) production. Mechanically, inflammasome-coupled IL-1β signaling triggered the expression of negative regulator SOCS1, which bound to IRF3 to inhibit IFN-I production. The role of IFN-I in anti-T. gondii immunity is little studied and controversial, and here we also found IFN-I is harmful to host antitoxoplasmosis immunity by using knockout mice and recombinant proteins. In general, our study identifies a protective role of inflammasomes to the host during T. gondii infection and a novel mechanism by which inflammasome suppresses IFN-I signaling in antitoxoplasmosis immunity, which will likely provide new insights into therapeutic targets for toxoplasmosis and highlight the cross talk between innate immune signaling in infectious diseases prevention.

Introduction: Since xenografts offer a wide range of incomparable advantages, they can be a better option than allografts but only if the possibility of immunological rejection can be eliminated. In this study, we investigated the ability of α1,3-galactosyltransferase (α1,3-GT) gene knockout (GTKO) pig cancellous bone to promote the repair of a femoral condyle bone defect and its influence on heterologous immune rejection. Materials and methods: Cylindrical bone defects created in a rhesus monkey model were transplanted with GTKO bone, WT bone or left empty. For immunological evaluation, T lymphocyte subsets CD4 + and CD8 + in peripheral blood were assayed by flow cytometry, and the IL-2 and IFN-γ contents of peripheral blood serum were analyzed by ELISA at 2, 5, 7, 10, and 14 days post-surgery. Micro-CT scans and histological assessment were conducted at 4 and 8 weeks after implantation. Results: Compared with WT-pig bone, the heterologous immunogenicity of GTKO-pig bone was reduced. The defect filled with fresh GTKO-pig bone was tightly integrated with the graft. Histological analysis showed that GTKO-pig cancellous bone showed better osseointegration and an appropriate rate of resorption. Osteoblast phenotype progression in the GTKO group was not affected, which revealed that GTKO-pig bone could not only fill and maintain the bone defect, but also promote new bone formation. Conclusion: GTKO-pig cancellous bone decreased the ratio of CD4 + to CD8 + T cells and cytokines (IFN-γ and IL-2) to inhibit xenotransplant rejection. Moreover, GTKO group increased more bone formation by micro-CT analysis and osteoblastic markers (Runx2, OSX and OCN). Together, GTKO-pig cancellous bone showed better bone repair than WT-pig cancellous bone., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Wang, Lu, Song, Zeng, Wang, Yang, Huang, Dai, Yang, Lai, Wang, Cai and Bai.)

Cyclic GMP-AMP synthase (cGAS) and stimulator of interferon genes (STING) play critical roles in the innate immunity against infectious diseases and are required to link pathogen DNA sensing to immune responses. However, the mechanisms by which cGAS-STING-induced cytokines suppress the adaptive immune response against malaria infections remain poorly understood. Here, cGAS-STING signaling is identified to play a detrimental role in regulating anti-malaria immunity. cGAS or STING deficiency in mice markedly prolongs mouse survival during lethal malaria Plasmodium yoelii nigeriensis N67C infections by reducing late interleukin (IL)-6 production. Mechanistically, cGAS/STING recruits myeloid differentiation factor 88 (MyD88) and specifically induces the p38-dependent signaling pathway for late IL-6 production, which, in turn, expands CD11b + Ly6C hi proinflammatory monocytes to inhibit immunity. Moreover, the blockage or ablation of the cGAS-STING-MyD88-p38-IL-6 signaling axis or the depletion of CD11b + Ly6C hi proinflammatory monocytes provides mice a significant survival benefit during N67C and other lethal malaria-strain infections. Taken together, these findings identify a previously unrecognized detrimental role of cGAS-STING-MyD88-p38 axis in infectious diseases through triggering the late IL-6 production and proinflammatory monocyte expansion and provide insight into how targeting the DNA sensing pathway, dysregulated cytokines, and proinflammatory monocytes enhances immunity against infection., (© 2022 The Authors. Advanced Science published by Wiley-VCH GmbH.)

Genome assembly has been benefited from long-read sequencing technologies with higher accuracy and higher continuity. However, most human genome assembly require large amount of DNAs from homogeneous cell lines without keeping cell heterogeneities, since cell heterogeneity could profoundly affect haplotype assembly results. Herein, using single-cell genome long-read sequencing technology (SMOOTH-seq), we have sequenced K562 and HG002 cells on PacBio HiFi and Oxford Nanopore Technologies (ONT) platforms and conducted de novo genome assembly. For the first time, we have completed the human genome assembly with high continuity (with NG50 of ∼2 Mb using 95 individual K562 cells) at single-cell levels, and explored the impact of different assemblers and sequencing strategies on genome assembly. With sequencing data from 30 diploid individual HG002 cells of relatively high genome coverage (average coverage ∼41.7%) on ONT platform, the NG50 can reach over 1.3 Mb. Furthermore, with the assembled genome from K562 single-cell dataset, more complete and accurate set of insertion events and complex structural variations could be identified. This study opened a new chapter on the practice of single-cell genome de novo assembly., (© The Author(s) 2022. Published by Oxford University Press on behalf of Nucleic Acids Research.)

With the rapid development of single-cell sequencing techniques, several large-scale cell atlas projects have been launched across the world. However, it is still challenging to integrate single-cell RNA-seq (scRNA-seq) datasets with diverse tissue sources, developmental stages and/or few overlaps, due to the ambiguity in determining the batch information, which is particularly important for current batch-effect correction methods. Here, we present SCORE, a simple network-based integration methodology, which incorporates curated molecular network features to infer cellular states and generate a unified workflow for integrating scRNA-seq datasets. Validating on real single-cell datasets, we showed that regardless of batch information, SCORE outperforms existing methods in accuracy, robustness, scalability and data integration., (© The Author(s) 2021. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Synovial macrophage polarization and inflammation are essential for osteoarthritis (OA) development, yet the molecular mechanisms and regulation responsible for the pathogenesis are still poorly understood. Here, we report that pseudolaric acid B (PAB) attenuated articular cartilage degeneration and synovitis during OA. PAB, a diterpene acid, specifically inhibited NF-κB signalling and reduced the production of pro-inflammatory cytokines, which further decreased M1 polarization and vessel formation. We further provide in vivo and in vitro evidences that PAB suppressed NF-κB signalling by stabilizing PPARγ. Using PPARγ antagonist could abolish anti-inflammatory effect of PAB and rescue the activation of NF-κB signalling during OA. Our findings identify a previously unrecognized role of PAB in the regulation of OA and provide mechanisms by which PAB regulates NF-κB signalling through PPARγ, which further suggest targeting synovial inflammation or inhibiting vessel formation at early stage could be an effective preventive strategy for OA., (© 2021 The Authors. Journal of Cellular and Molecular Medicine published by Foundation for Cellular and Molecular Medicine and John Wiley & Sons Ltd.)

Background: To investigate the role and regulatory mechanisms of fargesin, one of the main components of Magnolia fargesii, in macrophage reprogramming and crosstalk across cartilage and synovium during osteoarthritis (OA) development., Methods: Ten-week-old male C57BL/6 mice were randomized and assigned to vehicle, collagenase-induced OA (CIOA), or CIOA with intra-articular fargesin treatment groups. Articular cartilage degeneration was evaluated using the Osteoarthritis Research Society International (OARSI) score. Immunostaining and western blot analyses were conducted to detect relative protein. Raw264.7 cells were treated with LPS or IL-4 to investigate the role of polarized macrophages. ADTC5 cells were treated with IL-1β and conditioned medium was collected to investigate the crosstalk between chondrocytes and macrophages., Results: Fargesin attenuated articular cartilage degeneration and synovitis, resulting in substantially lower Osteoarthritis Research Society International (OARSI) and synovitis scores. In particular, significantly increased M2 polarization and decreased M1 polarization in synovial macrophages were found in fargesin-treated CIOA mice compared to controls. This was accompanied by downregulation of IL-6 and IL-1β and upregulation of IL-10 in serum. Conditioned medium (CM) from M1 macrophages treated with fargesin reduced the expression of matrix metalloproteinase-13, RUNX2, and type X collagen and increased Col2a1 and SOX9 in OA chondrocytes, but fargesin alone did not affect chondrocyte catabolic processes. Moreover, fargesin exerted protective effects by suppressing p38/ERK MAPK and p65/NF-κB signaling., Conclusions: This study showed that fargesin switched the polarized phenotypes of macrophages from M1 to M2 subtypes and prevented cartilage degeneration partially by downregulating p38/ERK MAPK and p65/NF-κB signaling. Targeting macrophage reprogramming or blocking the crosstalk between macrophages and chondrocytes in early OA may be an effective preventive strategy.

Objectives: To investigate the roles and regulatory mechanisms of synovial macrophages and their polarisation in the development of osteoarthritis (OA)., Methods: Synovial tissues from normal patients and patients with OA were collected. M1 or M2-polarised macrophages in synovial tissues of patients with OA and OA mice were analysed by immunofluorescence and immunohistochemical staining. Mice with tuberous sclerosis complex 1 (TSC1) or Rheb deletion specifically in the myeloid lineage were generated and subjected to intra-articular injection of collagenase (collagenase-induced osteoarthritis, CIOA) and destabilisation of the medial meniscus (DMM) surgery to induce OA. Cartilage damage and osteophyte size were measured by Osteoarthritis Research Society International score and micro-CT, respectively. mRNA sequencing was performed in M1 and control macrophages. Mice and ATDC5 cells were treated with R-spondin-2 (Rspo2) or anti-Rspo2 to investigate the role of Rspo2 in OA., Results: M1 but not M2-polarised macrophages accumulated in human and mouse OA synovial tissue. TSC1 deletion in the myeloid lineage constitutively activated mechanistic target of rapamycin complex 1 (mTORC1), increased M1 polarisation in synovial macrophages and exacerbated experimental OA in both CIOA and DMM models, while Rheb deletion inhibited mTORC1, enhanced M2 polarisation and alleviated CIOA in mice. The results show that promoting the macrophage M1 polarisation leads to exacerbation of experimental OA partially through secretion of Rspo2 and activation of β-catenin signalling in chondrocytes., Conclusions: Synovial macrophage M1 polarisation exacerbates experimental CIOA partially through Rspo2. M1 macrophages and Rspo2 are potential therapeutic targets for OA treatment., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2018. No commercial re-use. See rights and permissions. Published by BMJ.)

Vascular-invasion-mediated interactions between activated articular chondrocytes and subchondral bone are essential for osteoarthritis (OA) development. Here, we determined the role of nutrient sensing mechanistic target of rapamycin complex 1 (mTORC1) signaling in the crosstalk across the bone cartilage interface and its regulatory mechanisms. Then mice with chondrocyte-specific mTORC1 activation (Tsc1 CKO and Tsc1 CKO ER ) or inhibition (Raptor CKO ER ) and their littermate controls were subjected to OA induced by destabilization of the medial meniscus (DMM) or not. DMM or Tsc1 CKO mice were treated with bevacizumab, a vascular endothelial growth factor (VEGF)-A antibody that blocks angiogenesis. Articular cartilage degeneration was evaluated using the Osteoarthritis Research Society International score. Immunostaining and Western blotting were conducted to detect H-type vessels and protein levels in mice. Primary chondrocytes from mutant mice and ADTC5 cells were treated with interleukin-1β to investigate the role of chondrocyte mTORC1 in VEGF-A secretion and in vitro vascular formation. Clearly, H-type vessels were increased in subchondral bone in DMM-induced OA and aged mice. Cartilage mTORC1 activation stimulated VEGF-A production in articular chondrocyte and H-type vessel formation in subchondral bone. Chondrocyte mTORC1 promoted OA partially through formation of VEGF-A-stimulated subchondral H-type vessels. In particular, vascular-derived nutrients activated chondrocyte mTORC1, and stimulated chondrocyte activation and production of VEGF, resulting in further angiogenesis in subchondral bone. Thus a positive-feedback regulation of H-type vessel formation in subchondral bone by articular chondrocyte nutrient-sensing mTORC1 signaling is essential for the pathogenesis and progression of OA. © 2018 American Society for Bone and Mineral Research., (© 2018 American Society for Bone and Mineral Research.)

Communication between osteoblasts and endothelial cells (ECs) is essential for bone turnover, but the molecular mechanisms of such communication are not well defined. Here we identify Cxcl9 as an angiostatic factor secreted by osteoblasts in the bone marrow microenvironment. We show that Cxcl9 produced by osteoblasts interacts with vascular endothelial growth factor and prevents its binding to ECs and osteoblasts, thus abrogating angiogenesis and osteogenesis both in mouse bone and in vitro. The mechanistic target of rapamycin complex 1 activates Cxcl9 expression by transcriptional upregulation of STAT1 and increases binding of STAT1 to the Cxcl9 promoter in osteoblasts. These findings reveal the essential role of osteoblast-produced Cxcl9 in angiogenesis and osteogenesis in bone, and Cxcl9 can be targeted to elevate bone angiogenesis and prevent bone loss-related diseases.