October 2024
STAR Protocols
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October 2024
STAR Protocols
September 2024
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9 Reads
Veterinary Microbiology
August 2024
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13 Reads
Infectious Medicine
April 2024
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86 Reads
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4 Citations
Flaviviruses in the Japanese encephalitis virus (JEV) serogroup, such as JEV, West Nile virus, and St. Louis encephalitis virus, can cause severe neurological diseases. The nonstructural protein 1 (NS1) is a multifunctional protein of flavivirus that can be secreted by infected cells and circulate in the host bloodstream. NS1′ is an additional form of NS1 protein with 52 amino acids extension at its carboxy-terminal and is produced exclusively by flaviviruses in the JEV serogroup. In this study, we demonstrated that the secreted form of both NS1 and NS1′ can disrupt the blood-brain barrier (BBB) of mice, with NS1′ exhibiting a stronger effect. Using the in vitro BBB model, we found that treatment of soluble recombinant JEV NS1 or NS1′ protein increases the permeability of human brain microvascular endothelial cells (hBMECs) and leads to the degradation of tight junction proteins through the autophagy-lysosomal pathway. Consistently, NS1′ protein exhibited a more pronounced effect compared to NS1 in these cellular processes. Further research revealed that the increased expression of macrophage migration inhibitory factor (MIF) is responsible for triggering autophagy after NS1 or NS1′ treatment in hBMECs. In addition, TLR4 and NF-κB signaling was found to be involved in the activation of MIF transcription. Moreover, administering the MIF inhibitor has been shown to decrease viral loads and mitigate inflammation in the brains of mice infected with JEV. This research offers a novel perspective on the pathogenesis of JEV. In addition, the stronger effect of NS1′ on disrupting the BBB compared to NS1 enhances our understanding of the mechanism by which flaviviruses in the JEV serogroup exhibit neurotropism. IMPORTANCE Japanese encephalitis (JE) is a significant viral encephalitis worldwide, caused by the JE virus (JEV). In some patients, the virus cannot be cleared in time, leading to the breach of the blood-brain barrier (BBB) and invasion of the central nervous system. This invasion may result in cognitive impairment, behavioral disturbances, and even death in both humans and animals. However, the mechanism by which JEV crosses the BBB remains unclear. Previous studies have shown that the flavivirus NS1 protein plays an important role in causing endothelial dysfunction. The NS1′ protein is an elongated form of NS1 protein that is particularly produced by flaviviruses in the JEV serogroup. This study revealed that both the secreted NS1 and NS1′ of JEV can disrupt the BBB by breaking down tight junction proteins through the autophagy-lysosomal pathway, and NS1′ is found to have a stronger effect compared to NS1 in this process. In addition, JEV NS1 and NS1′ can stimulate the expression of MIF, which triggers autophagy via the ERK signaling pathway, leading to damage to BBB. Our findings reveal a new function of JEV NS1 and NS1′ in the disruption of BBB, thereby providing the potential therapeutic target for JE.
March 2024
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53 Reads
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2 Citations
Journal of Neuroinflammation
Japanese encephalitis virus (JEV) is a neurotropic pathogen that causes lethal encephalitis. The high susceptibility and massive proliferation of JEV in neurons lead to extensive neuronal damage and inflammation within the central nervous system. Despite extensive research on JEV pathogenesis, the effect of JEV on the cellular composition and viral tropism towards distinct neuronal subtypes in the brain is still not well comprehended. To address these issues, we performed single-cell RNA sequencing (scRNA-seq) on cells isolated from the JEV-highly infected regions of mouse brain. We obtained 88,000 single cells and identified 34 clusters representing 10 major cell types. The scRNA-seq results revealed an increasing amount of activated microglia cells and infiltrating immune cells, including monocytes & macrophages, T cells, and natural killer cells, which were associated with the severity of symptoms. Additionally, we observed enhanced communication between individual cells and significant ligand-receptor pairs related to tight junctions, chemokines and antigen-presenting molecules upon JEV infection, suggesting an upregulation of endothelial permeability, inflammation and antiviral response. Moreover, we identified that Baiap2-positive neurons were highly susceptible to JEV. Our findings provide valuable clues for understanding the mechanism of JEV induced neuro-damage and inflammation as well as developing therapies for Japanese encephalitis.
December 2023
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32 Reads
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3 Citations
Virologica Sinica
Ferroptosis is a newly discovered prototype of programmed cell death (PCD) driven by iron-dependent phospholipid peroxidation accumulation, and it has been linked to numerous organ injuries and degenerative pathologies. Although studies have shown that a variety of cell death processes contribute to JEV-induced neuroinflammation and neuronal injury, there is currently limited research on the specific involvement of ferroptosis. In this study, we explored the neuronal ferroptosis induced by JEV infection in vitro and in vivo. Our results indicated that JEV infection induces neuronal ferroptosis through inhibiting the function of the antioxidant system mediated by glutathione (GSH)/glutathione peroxidase 4 (GPX4), as well as by promoting lipid peroxidation mediated by yes-associated protein 1 (YAP1)/long-chain acyl-CoA synthetase 4 (ACSL4). Further analyses revealed that JEV E and prM proteins function as agonists, inducing ferroptosis. Moreover, we found that treatment with a ferroptosis inhibitor in JEV-infected mice reduces the viral titers and inflammation in the mouse brains, ultimately improving the survival rate of infected mice. In conclusion, our study unveils a critical role of ferroptosis in the pathogenesis of JEV, providing new ideas for the prevention and treatment of viral encephalitis.
October 2023
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105 Reads
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6 Citations
eLife
Pancreatic cancer is the seventh leading cause of cancer-related death worldwide, and despite advancements in disease management, the 5-year survival rate stands at only 12%. Triptolides have potent anti-tumor activity against different types of cancers, including pancreatic cancer, however poor solubility and toxicity limit their translation into clinical use. We synthesized a novel pro-drug of triptolide, ( E )-19-[(1'-benzoyloxy-1'-phenyl)-methylidene]-Triptolide (CK21), which was formulated into an emulsion for in vitro and in vivo testing in rats and mice, and using human pancreatic cancer cell lines and patient-derived pancreatic tumor organoids. A time-course transcriptomic profiling of tumor organoids treated with CK21 in vitro was conducted to define its mechanism of action, as well as transcriptomic profiling at a single time point post-CK21 administration in vivo. Intravenous administration of emulsified CK21 resulted in the stable release of triptolide, and potent anti-proliferative effects on human pancreatic cancer cell lines and patient-derived pancreatic tumor organoids in vitro, and with minimal toxicity in vivo . Time course transcriptomic profiling of tumor organoids treated with CK21 in vitro revealed <10 differentially expressed genes (DEGs) at 3 h and ~8,000 DEGs at 12 h. Overall inhibition of general RNA transcription was observed, and Ingenuity pathway analysis together with functional cellular assays confirmed inhibition of the NF-κB pathway, increased oxidative phosphorylation and mitochondrial dysfunction, leading ultimately to increased reactive oxygen species (ROS) production, reduced B-cell-lymphoma protein 2 (BCL2) expression, and mitochondrial-mediated tumor cell apoptosis. CK21 is a novel pro-drug of triptolide that exerts potent anti-proliferative effects on human pancreatic tumors by inhibiting the NF-κB pathway, leading ultimately to mitochondrial-mediated tumor cell apoptosis.
October 2023
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72 Reads
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3 Citations
Cell Reports
The current paradigm indicates that naive T cells are primed in secondary lymphoid organs. Here, we present evidence that intranasal administration of peptide antigens appended to nanofibers primes naive CD8⁺ T cells in the lung independently and prior to priming in the draining mediastinal lymph node (MLN). Notably, comparable accumulation and transcriptomic responses of CD8⁺ T cells in lung and MLN are observed in both Batf3KO and wild-type (WT) mice, indicating that, while cDC1 dendritic cells (DCs) are the major subset for cross-presentation, cDC2 DCs alone are capable of cross-priming CD8⁺ T cells both in the lung and draining MLN. Transcription analyses reveal distinct transcriptional responses in lung cDC1 and cDC2 to intranasal nanofiber immunization. However, both DC subsets acquire shared transcriptional responses upon migration into the lymph node, thus uncovering a stepwise activation process of cDC1 and cDC2 toward their ability to cross-prime effector and functional memory CD8⁺ T cell responses.
July 2023
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62 Reads
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1 Citation
Journal of Neuroinflammation
Histone methylation is an important epigenetic modification that affects various biological processes, including the inflammatory response. In this study, we found that infection with Japanese encephalitis virus (JEV) leads to an increase in H3K27me3 in BV2 microglial cell line, primary mouse microglia and mouse brain. Inhibition of H3K27me3 modification through EZH2 knockdown and treatment with EZH2 inhibitor significantly reduces the production of pro-inflammatory cytokines during JEV infection, which suggests that H3K27me3 modification plays a crucial role in the neuroinflammatory response caused by JEV infection. The chromatin immunoprecipitation-sequencing (ChIP-sequencing) assay revealed an increase in H3K27me3 modification of E3 ubiquitin ligases Rnf19a following JEV infection, which leads to downregulation of Rnf19a expression. Furthermore, the results showed that Rnf19a negatively regulates the neuroinflammatory response induced by JEV. This is achieved through the degradation of RIG-I by mediating its ubiquitination. In conclusion, our findings reveal a novel mechanism by which JEV triggers extensive neuroinflammation from an epigenetic perspective.
February 2023
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83 Reads
Background Pancreatic cancer is the seventh leading cause of cancer-related death worldwide, and despite advancements in disease management, the 5-year survival rates stands at only 9%. Triptolides have potent anti-tumor activity against different types of cancers, including pancreatic cancer, however poor solubility and toxicity limit their translation into clinical use. Methods We synthesized a novel pro-drug of triptolide, ( E )-19-[(1’-benzoyloxy-1’-phenyl)-methylidene]-Triptolide (CK21), and formulated into an emulsion for in vitro and in vivo testing in rats and mice, and using human pancreatic cancer cell lines and patient-derived pancreatic tumor organoids. A time-course transcriptomic profiling of tumor organoids treated with CK21 in vitro was conducted to define its mechanism of action, as well as transcriptomic profiling at a single time point post-CK21 administration in vivo. Findings Intravenous administration of emulsified CK21 resulted in the stable release of triptolide, and potent anti-proliferative effects on human pancreatic cancer cell lines and patient-derived pancreatic tumor organoids in vitro, and with minimal toxicity in vivo . Time course transcriptomic profiling of tumor organoids treated with CK21 in vitro revealed <10 differentially expressed genes (DEGs) at 3 h and ∼8,000 DEGs at 12 h. Overall inhibition of general RNA transcription was observed, and Ingenuity pathway analysis together with functional cellular assays confirmed inhibition of the NF-κB pathway, increased oxidative phosphorylation and mitochondrial dysfunction, leading ultimately to increased reactive oxygen species (ROS) production, reduced B-cell-lymphoma protein 2 (BCL2) expression, and mitochondrial-mediated tumor cell apoptosis. Interpretation CK21 is a novel pro-drug of triptolide that exerts potent anti-proliferative effects on human pancreatic tumors by inhibiting the NF-κB pathway, leading ultimately to mitochondrial-mediated tumor cell apoptosis. Funding The study of the anti-tumor efficacy of CK21 supported in part by a research grant from Cinkate Pharmaceutical Corp; the funders had no role in the study design, interpretation or decision to publish. Patient-derived pancreatic tumor organoids were a generous gift from the Organoid and Primary Culture Research Core at University of Chicago.
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... Viral brain infections are characterized by strong expression of type I interferon (IFN) genes and inflammatory cytokines [4][5][6] . This is accompanied by influx of immune cells from the periphery with an early recruitment of monocytes/macrophages and natural killer cells, and later T cells 7,8 . Both brain resident and infiltrating cell types have been described to contribute to the innate immune response to viral CNS infections. ...
March 2024
Journal of Neuroinflammation
... In addition, several neurodegenerative diseases have also shown a link to HSV infection (Zhu and Viejo-Borbolla, 2021). Two neuroinvasive viruses, JEV and HSV, both cause severe encephalitis, and the current studies found a strong correlation between symptoms after JEV and HSV infection and ferroptosis (Xu et al., 2023;Zhu et al., 2024). Ferroptosis is a new non-apoptotic form of programmed cell death discovered in recent years, which is usually accompanied by a large amount of iron accumulation and lipid peroxidation (LPO) during cell death. ...
December 2023
Virologica Sinica
... These findings reveal that whereas cDC1 may be primary mediators of tumor antigen presentation in WT mice (Gardner et al., 2020), cDC2 are capable of presenting tumor antigens as well and become more effective than cDC1 in this function in the absence of MHC-II. This is consistent with previous work showing that cDC2 are capable of efficiently cross-presenting antigens to CD8 T lymphocytes in other experimental settings (Ballesteros-Tato et al., 2010;Ji et al., 2013;Sheng et al., 2017;Si et al., 2023;Theisen et al., 2018). ...
October 2023
Cell Reports
... A total of 75% of antitumor compounds currently used to treat human cancers are natural products [24]. With advancements in natural compound extraction and separation technologies, a growing number of compounds with anti-pancreatic cancer activities have been identified in plants [25][26][27][28][29]. For instance, CK21 is a novel pro-drug of triptolide that exerts potent anti-proliferative effects on human pancreatic tumors by inhibiting the NF-κB pathway, ultimately leading to mitochondrial-mediated tumor cell apoptosis [25]. ...
October 2023
eLife
... Some results support antiviral efficacy of favipiravir against ZIKV in nonhuman primates [51]. Previous research has also proven that testosterone can reduce mortality and attenuate testicular damage in ZIKVinfected A129 mice [52]. However, our models only indicate that the testosterone could relieve ZIKV-induced testicular lesion. ...
October 2022
iScience
... (B and C) Data are representative of two placentas for each condition from two dams. of ZIKV infection during pregnancy has been described in animal models Yockey et al., 2016Yockey et al., , 2018, it has not been clear which innate immune responses are most relevant in the fetal compartment in a setting of maternal immunocompetence. While viral sensing via RIG-I/MDA5-MAVS has been shown to restrict ZIKV infection in human placental cell lines Zhao et al., 2022), this pathway has not been studied in vivo; however, deletion of the downstream transcription factors Irf3 and Irf7 facilitated placental and fetal infection in a mouse model of intravaginal ZIKV infection during pregnancy (Yockey et al., 2016). Our breeding schemes with immunocompetent (heterozygous) dams and immunocompetent (heterozygous) or immunodeficient (homozygous) sires enabled the generation of innate immune deficiencies specifically in the fetus. ...
August 2022
eLife
... Namely, the local tissue milieu can elicit different immune profiles, increasing the complexity of successfully eliciting protective antibacterial responses across the myriad of tissues that S. aureus can cause invasive disease. This has been demonstrated in recent studies where unique immune responses have been observed when S. aureus invades the epidermis vs. dermis in mouse models of cutaneous infection [99][100][101][102][103][104] . ...
August 2022
... Due to their rapid processing speed, as well as their heightened sensitivity and specificity, molecular techniques have become the predominant diagnostic method for most viral CNS infections [10,17]. As an accurate and rapid biosensor, the CRISPR/Cas system has expanded to a new use of diagnostics for infectious diseases, including the detection of various viral nucleic acids [18][19][20][21]. For ZIKV detection, the Cas13-based SHERLOCK (specific high-sensitivity enzymatic reporter unlocking) platform can probe and distinguish ZIKV and four dengue virus (DENV) serotypes in infected human-patient bodily fluid samples at concentrations as low as one copy per microliter [22]. ...
Reference:
PfAgo-Based Zika Virus Detection
July 2022
Journal of Virological Methods
... A distinctive m6A site is present in the prM (495-695nt) region of the JEV genome, a feature that is not observed in other Flaviviridae viral genomes. The prM protein is largely responsible for the assembly of viruses and their virulence, suggesting that m6A modification of prM may be involved in this process [29,39,41]. ...
June 2022
Microbiology Spectrum
... 目前, JEV主要在中国、日 本、韩国、菲律宾、印度和俄罗斯东部地区等亚洲地 区流行 [96] . Chen等人 [97] 研发了一种编码经修饰的JEV P3株 prM/E蛋白的mRNA疫苗, 在小鼠中可产生中和抗体, 其抗体的PRNT 50 (50% Plaque Reduction Neutralization Test)滴度达到1/100; 同时引起CD8 + T淋巴细胞介导的 免疫应答. 该疫苗也可保护免疫小鼠免受致死剂量病 毒感染, 并减少病毒引起的神经炎症反应 [97] . ...
May 2022