Discovery of novel coumarin-based KRAS-G12C inhibitors from virtual screening and Rational structural optimization.

KRAS-G12C inhibitors has been made significant progress in the treatment of KRAS-G12C mutant cancers, but their clinical application is limited due to the adaptive resistance, motivating development of novel structural inhibitors. Herein, series of coumarin derivatives as KRAS-G12C inhibitors were found through virtual screening and rational structural optimization. Especially, K45 exhibited strong antiproliferative potency on NCI-H23 and NCI-H358 cancer cells harboring KRAS-G12C with the IC50 values of 0.77 µM and 1.50 µM, which was 15 and 11 times as potent as positive drug ARS1620, respectively. Furthermore, K45 reduced the phosphorylation of KRAS downstream effectors ERK and AKT by reducing the active form of KRAS (KRAS GTP) in NCI-H23 cells. In addition, K45 induced cell apoptosis by increasing the expression of anti-apoptotic protein BAD and BAX in NCI-H23 cells. Docking studies displayed that the 3-naphthylmethoxy moiety of K45 extended into the cryptic pocket formed by the residues Gln99 and Val9, which enhanced the interaction with the KRAS-G12C protein. These results indicated that K45 was a potent KRAS-G12C inhibitor worthy of further study.

1927nm fractional thulium fiber laser combined with 30% salicylic acid for the treatment of acne and acne scars: A prospective, randomized, and split-face study.

OBJECTIVES: Patients with acne usually develops acne scars subsequently, early intervention of scars is crucial in acne management. 1927nm fractional thulium fiber laser (TFL) is effective in scars improvement and chemical peels with 30% supramolecular salicylic acid (SSA) can be applied for the treatment of acne. The purpose of this study is to evaluate and compare the efficacy and safety of TFL monotherapy versus the concomitant application of TFL and 30% SSA on acne and acne scars. MATERIALS AND METHODS: Thirty-three patients with acne and acne scars were enrolled, and two sides of the face were randomly divided to receive either TFL and SSA chemical peeling or TFL. Four sessions of TFL treatments were applied with 4-week intervals for both sides, SSA combined treatment side received eight SSA chemical peels with 2-week intervals additionally. GAGS, ECCA score, the number of acne lesions, melanin index (MI) and erythema index (EI), transepidermal water loss (TEWL), and side effects were recorded at Weeks 0, 4, 8, 12, and 18. Satisfaction of patients was recorded on both sides at the end of the study. RESULTS: Thirty patients completed the study. Both control group (TFL monotherapy) and SSA group (TFL combined with SSA chemical peeling) significantly improved GAGS and ECCA score. SSA group showed higher efficacy in terms of GAGS and ECCA score, acne lesion count, TEWL, MI, EI, and satisfaction than control group. All the side effects were temporary and tolerable, no adverse effects were observed. CONCLUSIONS: Both TFL and the TFL combined with 30% SSA chemical peeling are safe and effective for the treatment and prevention of acne and acne scars, though the combined group has higher efficacy.

Water-Soluble Au25 Clusters with Single-Crystal Structure for Mitochondria-Targeting Radioimmunotherapy.

Atomically precise gold clusters play an important role in the development of high-Z-element-based radiosensitizers, due to their intriguing structural diversity and advantages in correlating structures and properties. However, the synthesis of gold clusters with both water-solubility and single-crystal structure remains a challenge. In this study, atomically precise Au25(S-TPP)18 clusters (TPP-SNa = sodium 3-(triphenylphosphonio)propane-1-thiolate bromide) showing both mitochondria-targeting ability and water-solubility were obtained via ligand design for enhanced radioimmunotherapy. Compared with Au25(SG)18 clusters (SG = glutathione), Au25(S-TPP)18 exhibited higher radiosensitization efficiency due to its mitochondria-targeting ability, higher ROS production capacity, and obvious inhibition upon thioredoxin reductase (TrxR). In addition, the enhanced radiotherapy-triggered abscopal effect in combination with checkpoint blockade displayed effective growth inhibition of distant tumors. This work reveals the ligand-regulated organelle targeting ability of metal clusters by which feasible strategies to promote their application in precise theranostics could be realized.

Chemical Constituents of Primulina eburnea (Gesneriaceae) and Their Cytotoxic Activities.

Two new ursane-type triterpenes, eburnealactones A and B (1 and 2), one new flavonoid, eburneatin A (6), and one new phenylethanoid glycoside, chiritoside D (7), along with 9 known compounds (3-5, 8-13) were isolated from the whole plant of Primulina eburnea. Their structures were elucidated by comprehensive spectroscopic data analysis (IR, UV, NMR, and HR-ESI-MS). All the compounds were evaluated for their cytotoxic activities. Compound 1 showed significant cytotoxic activities against MKN-45 cell lines and 5637 cell lines with the IC50 values of 9.57 µM and 8.30 µM, respectively. Compound 1 exhibited moderate cytotoxic activities against A549 and PATU8988T cell lines with the IC50 values of 30.70 µM and 38.22 µM, respectively. Compound 6 exhibited moderate cytotoxic activities against MKN-45, HCT116, PATU8988T, 5637 and A-673 cell lines with the IC50 values of 19.69 µM, 16.44 µM, 18.07 µM, 11.51 µM and 18.15 µM, respectively. Compound 5 showed moderate cytotoxic activities against A549 cell lines with the IC50 values of 24.06 µM.

Hypoxia inducible factor 1α promotes interleukin-1 receptor antagonist expression during hepatic ischemia-reperfusion injury.

BACKGROUND: Ischemia-reperfusion injury (IRI) is a major risk associated with liver surgery and transplantation, and its pathological mechanism is complex. Interleukin-1 receptor antagonist (IL-1ra) can protect the liver from IRI. However, the regulatory mechanism of IL-1ra expression is still unclear. AIM: To identify the mechanism that could protect the liver in the early stage of IRI. METHODS: To screen the key genes in hepatic IRI, we performed RNA sequencing and gene enrichment analysis on liver tissue from mice with hepatic IRI. Subsequently, we verified the expression and effect of IL-1ra in hepatic IRI. We also used promoter mutagenesis and chromatin immunoprecipitation assay to search for the transcriptional regulatory sites of hypoxia-inducible factor (HIF)-1α. Finally, to explore the protective mechanism of ischemic preconditioning (IP), we examined the expression of HIF-1α and IL-1ra after IP. RESULTS: We identified IL-1ra as a key regulator in hepatic IRI. The expression of IL-1ra was significantly upregulated after hepatic IRI both in vivo and in vitro. Furthermore, we found that HIF-1α regulated Il-1ra transcription in response to hypoxia. Increased HIF-1α accumulation promoted IL-1ra expression, whereas inhibition of HIF-1α exhibited the opposite effect. We also confirmed a predominant role for hypoxia response element in the regulation of Il1ra transcription by HIF-1α activation. Of note, we demonstrated that IP protects against hepatic IRI by inducing IL-1ra expression, which is mediated through HIF-1α. CONCLUSION: We demonstrated that ischemia or hypoxia leads to increased expression of IL-1ra through HIF-1α. Importantly, IP protects the liver from IRI via the HIF-1α-IL-1ra pathway.

Special Computed Tomography Imaging Features of Thymic Cyst.

Objective: To explore the features and diagnostic value of computed tomography (CT) imaging in cases of thymic cysts. Methods: A total of 24 cases of the thymic cysts (confirmed by postoperative pathology) were retrospectively analyzed. The location, morphology, and density of the thymic cysts were summarized, and the changes in CT value of the region of interest (ROI) in the thymic cysts between noncontrast enhanced and enhanced chest scans were compared and classified. Results: The average long-axis dimension was 17.50 ± 6.00 mm, the CT value range across the 24 cases was 5-81 HU, and the average CT value of the noncontrast enhanced scans was 39.75 ± 20.66 HU. The CT value in the noncontrast enhanced scan was >20 HU in 79% of the sample cases. The CT value in the ROI of the thymic cysts under enhanced scan showed a significant decrease in 15 cases, a significant increase in 5 cases, and an insignificant change in 4 cases. Conclusion: The CT values of the thymic cysts in the enhanced scans were generally lower than in the noncontrast enhanced scans, which might be a valuable finding for thymic cysts diagnosis.

Design, synthesis and biological evaluation of coumarin derivatives as potential BRD4 inhibitors.

The bromodomain and extra-terminal (BET) bromodomains, particularly BRD4, have been identified as promising therapeutic targets in the treatment of many human disorders such as cancer. Coumarin is a highly privileged moiety for the development of novel anticancer drugs which has been identified in clinical trials for the treatment of various cancers. Herein, we modified BRD4i ABBV-075 with a coumarin ring and synthesized a novel series of coumarin derivatives as BRD4 inhibitors. Among them, the representative compound 27d showed excellent BRD4 inhibitory activities with an IC50 value of 99 nM in the TR-FRET assay. Compared with ABBV-075, compound 27d displayed a favorable cell proliferation inhibitory activity in solid tumors, such as MCF-7, HGC-27 and HepG-2. Further mechanism investigation illustrated that 27d-treatment resulted in G0/G1 phase arrest and promoted apoptosis of MCF-7 cells. Compound 27d also blocked colony formation in a concentration-dependent manner in McF-7 cell lines. As the downstream-protein of BRD4, the expression of c-Myc was decreased in a dose-dependent manner after the treatment of compound 27d. Moreover, compound 27d also exhibited good in vivo and in vitro metabolic stability. All the findings meaningfully make it as a promising lead compound for further drug development.

Composition-Dependent Enzyme Mimicking Activity and Radiosensitizing Effect of Bimetallic Clusters to Modulate Tumor Hypoxia for Enhanced Cancer Therapy.

Alloying is an efficient chemistry to tailor the properties of metal clusters. As a class of promising radiosensitizers, most previously reported metal clusters exhibit unitary function and cannot overcome radioresistance of hypoxic tumors. Here, atomically precise alloy clusters Pt2 M4 (M = Au, Ag, Cu) are synthesized with bright luminescence and adequate biocompatibility, and their composition-dependent enzyme mimicking activity and radiosensitizing effect is explored. Specifically, only the Pt2 Au4 cluster displays catalase-like activity, while the others do not have clusterzyme properties, and its radiosensitizing effect is the highest among all the alloy clusters tested. By taking advantage of the sustainable production of O2 via the decomposition of endogenous H2 O2 , the Pt2 Au4 cluster modulates tumor hypoxia as well as increases the efficacy of radiotherapy. This work thus advances the cluster alloying strategy to produce multifunctional therapeutic agents for improving hypoxic tumor therapy.

Electrochemical enhancement of high-efficiency wet removal of mercury from flue gas.

Electrochemical wet absorption composite system has an excellent potential to remove Hg0 from flue gas. In this study, ruthenium iridium titanium platinum quaternary composite electrode is used as an anode and titanium electrode is used as the cathode, and KI/I2 absorption solution is introduced into the electrocatalysis system as an electrolyte to form KI/I2 electrochemical catalytic oxidation system. The removal rate of Hg0 in flue gas can be increased to 92.3%. The effects of electrolytic voltage, current, Pt content, I2 concentration, and the ratio of KI/I2 on the removal of Hg0 were discussed. The possible free radicals in the electrochemical cathode, anode, and solution were characterized and tested by XRD, SEM, UV-Vis (detection of H2O2, ·OH, O3), and FTIR (detection of IO3-). Combined with experimental data and theoretical derivation, the mechanism of Hg0 removal from flue gas by electrochemical catalytic oxidation alloy formation wet absorption combined process was studied. The results show that the combined process, which is a promising technology can not only improve the removal efficiency of Hg0, but also realize the resource recovery of Hg0 and I2, and provide a feasibility study for the subsequent regeneration of KI/I2 absorption solution.

A multifunctional AIE gold cluster-based theranostic system: tumor-targeted imaging and Fenton reaction-assisted enhanced radiotherapy.

BACKGROUND: As cancer is one of the main leading causes of mortality, a series of monotherapies such as chemotherapy, gene therapy and radiotherapy have been developed to overcome this thorny problem. However, a single treatment approach could not achieve satisfactory effect in many experimental explorations. RESULTS: In this study, we report the fabrication of cyclic RGD peptide (cRGD) modified Au4-iron oxide nanoparticle (Au4-IO NP-cRGD) based on aggregation-induced emission (AIE) as a multifunctional theranostic system. Besides Au4 cluster-based fluorescence imaging and enhanced radiotherapy, iron oxide (IO) nanocluster could realize magnetic resonance (MR) imaging and Fenton reaction-based chemotherapy. Abundant toxic reactive oxygen species generated from X-ray irradiation and in situ tumor-specific Fenton reaction under acidic microenvironment leads to the apoptotic and necrotic death of cancer cells. In vivo studies demonstrated good biocompatibility of Au4-IO NP-cRGD and a high tumor suppression rate of 81.1% in the synergistic therapy group. CONCLUSIONS: The successful dual-modal imaging and combined tumor therapy demonstrated AIE as a promising strategy for constructing multifunctional cancer theranostic platform.

Prognostic impact of tumor length in esophageal Cancer: a systematic review and Meta-analysis.

BACKGROUND: In clinical studies, it has been observed that esophageal cancer (EC) patient prognosis can be very different even for those patients with tumors of the same TNM stage. Tumor length has been analysed as a possible independent prognostic factor in many studies, but no unanimous conclusion has been reached. Therefore, this review used a meta-analysis to evaluate the association between tumor length and prognosis in EC patients. METHODS: A systematic search for relevant articles was performed in PubMed, Web of Science, and Embase. Hazard ratios (HRs) and 95% confidence intervals (CIs) were used as effective measures to estimate the correlation between tumor length and prognosis, including overall survival, disease-free survival, progression-free survival, disease-specific survival, and cancer-specific survival. STATA 15.0 software was used to perform the meta-analysis and the data synthesis. RESULTS: Finally, 41 articles with 28,973 patients were included in our study. The comprehensive statistical results showed that long tumors are an independent prognostic parameter associated with poor overall survival (OS) (HR = 1.30; 95% CI: 1.21-1.40, p < .001) and disease-free survival (DFS) (HR = 1.38; 95% CI: 1.18-1.61, p < .001) in EC patients. Subgroup analyses also suggested a significant correlation between long tumors and poor OS. Sensitivity analysis and publication bias evaluation confirmed the reliability and stability of the results. Similar results were obtained in the analyses of progression-free survival (PFS), disease-specific survival (DSS), and cancer-specific survival (CSS). CONCLUSION: The results of this meta-analysis showed that long tumors were related to poor OS, DFS, PFS, DSS and CSS in EC patients. Tumor length might be an important predictor of prognosis in EC patients, and it can be used as an independent staging index. Further well-designed and large-scale prospective clinical studies are needed to confirm these findings.

PARP1 deficiency protects against hyperglycemia-induced neointimal hyperplasia by upregulating TFPI2 activity in diabetic mice.

Diabetes mellitus (DM) promotes neointimal hyperplasia, characterized by dysregulated proliferation and accumulation of vascular smooth muscle cells (VSMCs), leading to occlusive disorders, such as atherosclerosis and stenosis. Poly (ADP-ribose) polymerase 1 (PARP1), reported as a crucial mediator in tumor proliferation and transformation, has a pivotal role in DM. Nonetheless, the function and potential mechanism of PARP1 in diabetic neointimal hyperplasia remain unclear. In this study, we constructed PARP1 conventional knockout (PARP1-/-) mice, and ligation of the left common carotid artery was performed to induce neointimal hyperplasia in Type I diabetes mellitus (T1DM) mouse models. PARP1 expression in the aorta arteries of T1DM mice increased significantly and genetic deletion of PARP1 showed an inhibitory effect on the neointimal hyperplasia. Furthermore, our results revealed that PARP1 enhanced diabetic neointimal hyperplasia via downregulating tissue factor pathway inhibitor (TFPI2), a suppressor of vascular smooth muscle cell proliferation and migration, in which PARP1 acts as a negative transcription factor augmenting TFPI2 promoter DNA methylation. In conclusion, these results suggested that PARP1 accelerates the process of hyperglycemia-induced neointimal hyperplasia via promoting VSMCs proliferation and migration in a TFPI2 dependent manner.

Autoantibody Signatures as a Biomarker Panel for the Detection of Nasopharyngeal Carcinoma.

OBJECTIVE: The early symptoms of nasopharyngeal carcinoma (NPC) are not obvious, and it is difficult to make early diagnosis. A case-control study was conducted to identify potential biomarkers and established a diagnosis model for nasopharyngeal carcinoma. METHODS: Plasma samples of 131 cases of NPC and 132 cases of healthy individuals were incubated with the Ray Biotech Human Lung Cancer IgG Autoantibody Detection Array G1, and signal values were used to develop a risk prediction model for NPC diagnosis. RESULTS: Of the 30 autoantibodies, high expression of MAGE-A4, NY-ESO-1, HuD, Survivin, IMDH2, Ubiquilin-1, IMP1, PGP9.5, IMP3, C-Myc and low expression of Cyclin B1 were potential biomarkers for NPC diagnosis (p <0.05), among which Survivin, MAGE-A4 and IMP3 shows higher AUC of 0.674, 0.652 and 0.650 respectively, the specificity of them was 89.39% (95% CI: 82.85-94.08%), 90.15% (95% CI: 83.75-94.65%) and 88.64% (81.95-93.50%).The risk probability analysis for NPC diagnosis based on the panel of Cyclin B1, NY-ESO-1, Survivin, and IMP3 displayed the best diagnosis performance with an AUC of 0.779, p (Yi = 1) = 1/(1+EXP[8.316+1.672*CyclinB1-1.152*NY-ESO-1-2.052*Survivin-0.950*IMP3]), the specificity of that was 86.36% (95% CI: 79.31-91.71%). CONCLUSIONS: Our findings demonstrated that the panel of Cyclin B1, NY-ESO-1, Survivin, and IMP3 has a good performance in the detection of NPC, and all 11 autoantibodies may also have a certain significance for the prognosis of NPC.

Protein deglycase DJ-1 deficiency induces phenotypic switching in vascular smooth muscle cells and exacerbates atherosclerotic plaque instability.

Protein deglycase DJ-1 (DJ-1) is a multifunctional protein involved in various biological processes. However, it is unclear whether DJ-1 influences atherosclerosis development and plaque stability. Accordingly, we evaluated the influence of DJ-1 deletion on the progression of atherosclerosis and elucidate the underlying mechanisms. We examine the expression of DJ-1 in atherosclerotic plaques of human and mouse models which showed that DJ-1 expression was significantly decreased in human plaques compared with that in healthy vessels. Consistent with this, the DJ-1 levels were persistently reduced in atherosclerotic lesions of ApoE-/- mice with the increasing time fed by western diet. Furthermore, exposure of vascular smooth muscle cells (VSMCs) to oxidized low-density lipoprotein down-regulated DJ-1 in vitro. The canonical markers of plaque stability and VSMC phenotypes were evaluated in vivo and in vitro. DJ-1 deficiency in Apoe-/- mice promoted the progression of atherosclerosis and exaggerated plaque instability. Moreover, isolated VSMCs from Apoe-/- DJ-1-/- mice showed lower expression of contractile markers (α-smooth muscle actin and calponin) and higher expression of synthetic indicators (osteopontin, vimentin and tropoelastin) and Kruppel-like factor 4 (KLF4) by comparison with Apoe-/- DJ-1+/+ mice. Furthermore, genetic inhibition of KLF4 counteracted the adverse effects of DJ-1 deletion. Therefore, our results showed that DJ-1 deletion caused phenotype switching of VSMCs and exacerbated atherosclerotic plaque instability in a KLF4-dependent manner.

HN1L promotes invasion and metastasis of the esophagogastric junction adenocarcinoma.

BACKGROUND: Adenocarcinoma of the esophagogastric junction (AEG) refers to cancer that crosses the line of the gastroesophageal junction and includes distal esophageal cancer and proximal gastric cancer. It is characterized by early metastasis and a poor prognosis and has few treatment options. Here, we report a novel potential therapeutic target, hematological and neurological expressed 1-like (HN1L), in AEG. METHODS: A total of 38 patients who underwent surgical resection of AEG at the Department of Thoracic Surgery of Shandong Provincial Hospital from September 2018 to June 2019 were enrolled into the study. We detected the expression of HN1L in AEG and adjacent nontumor tissues by IHC staining. The clinicopathological characteristics of HN1L were statistically analyzed. Then, the expression of HN1L in different cell lines was detected by RT-q PCR. Finally, AGS and HGC-27 cell lines were performed to inhibit HN1L by shRNA in order to explore its role in the development of AEG. RESULTS: Immunohistochemical staining showed that the expression of HN1L in cancer tissues was higher than that in nontumor tissue (p < 0.001). High expression of HN1L was significantly correlated with TNM stage (p = 0.013) and lymph node metastasis (p = 0.03). The expression of HN1L was upregulated in tumor cell lines compared with normal cell line. Additionally, Cell function studies demonstrated that lentivirus-mediated shRNA silencing of HN1L expression could effectively reduce the proliferation, invasion, and metastasis of tumor cell lines and promote their apoptosis (p < 0.05). CONCLUSIONS: HN1L expression might contribute to the invasion and metastasis of AEG and is a promising therapeutic target.

Synthesis of N-2(5H)-furanonyl sulfonyl hydrazone derivatives and their biological evaluation in vitro and in vivo activity against MCF-7 breast cancer cells.

A series of (E)-N-2(5H)-furanonyl sulfonyl hydrazone derivatives have been rationally designed and efficiently synthesized by one-pot reaction with good yields for the first time. This green approach with wide substrate range and good selectivity can be achieved at room temperature in a short time in the presence of metal-free catalyst. The cytotoxic activities against three human cancer cell lines of all newly obtained compounds have been evaluated by MTT assay. Among them, compound 5 k exhibits high cytotoxic activity against MCF-7 human breast cancer cells with an IC50 value of 14.35 µM. The cytotoxic mechanism may involve G2/M phase arrest pathway, which is probably caused by activating DNA damage. Comet test and immunofluorescence results show that compound 5 k can induce DNA damage in time- and dose-dependent manner. Importantly, 5 k also can effectively inhibit the proliferation of MCF-7 cells and angiogenesis in the zebrafish xenograft model. It is potential to further develop N-2(5H)-furanonyl sulfonyl hydrazone derivatives as potent drugs for breast cancer treatment with higher cytotoxic activity by modifying the structure of the compound.

The Multifaceted Roles of TAM Receptors during Viral Infection.

Tyro3, Axl, and Mertk (TAM) receptors play multiple roles in a myriad of physiological and pathological processes, varying from promoting the phagocytic clearance of apoptotic cells, sustaining the immune and inflammatory homeostasis, maintaining the blood-brain barrier (BBB) integrity and central nervous system (CNS) homeostasis, to mediating cancer malignancy and chemoresistance. Growth arrest-specific protein 6 (Gas6) and protein S (Pros1) are the two ligands that activate TAM receptors. Recently, TAM receptors have been reported to mediate cell entry and infection of multitudinous enveloped viruses in a manner called apoptotic mimicry. Moreover, TAM receptors are revitalized during viral entry and infection, which sequesters innate immune and inflammatory responses, facilitating viral replication and immune evasion. However, accumulating evidence have now proposed that TAM receptors are not required for the infection of these viruses in vivo. In addition, TAM receptors protect mice against the CNS infection of neuroinvasive viruses and relieve the brain lesions during encephalitis. These protective effects are achieved through maintaining BBB integrity, attenuating proinflammatory cytokine production, and promoting neural cell survival. TAM receptors also regulate the programmed cell death modes of virus-infected cells, which have profound impacts on the pathogenesis and outcome of infection. Here, we systematically review the functionalities and underlying mechanisms of TAM receptors and propose the potential application of TAM agonists to prevent severe viral encephalitis.

Axl Deficiency Promotes the Neuroinvasion of Japanese Encephalitis Virus by Enhancing IL-1α Production from Pyroptotic Macrophages.

Japanese encephalitis virus (JEV) is a flavivirus that causes Japanese encephalitis (JE), which has an unclear pathogenesis. Despite vaccination, thousands of deaths attributed to JE are reported annually. In this study, we report that mice deficient for Axl, a receptor tyrosine kinase that plays multiple roles in flaviviral infection, displayed greater mortality upon JEV infection. The effect of Axl deficiency on JEV infection was mediated by markedly elevated serum interleukin-1α (IL-1α) levels, which devastated the blood-brain-barrier and promoted viral neuroinvasion within 24 h postinfection. Using an in situ infection model, we showed that dead macrophages were the primary source of observed increased serum IL-1α levels. Axl deficiency enhanced cell death and caused pyroptosis in 80% of JEV-infected macrophages by disrupting phosphatidylinositol 3-kinase (PI3K)-Akt signaling. Intriguingly, the primary effector released by pyroptotic macrophages in our model was IL-1α rather than IL-1ß. Finally, we assessed the effect of an IL-1α antagonist and demonstrated that it effectively prevented the incidence of JE. Our results indicate that Axl plays a protective role in JEV infection, identify IL-1α released by pyroptotic macrophages as a crucial factor promoting JEV neuroinvasion, and suggest that an IL-1α antagonist may be a candidate for JE therapy.IMPORTANCE Japanese encephalitis virus (JEV) is a mosquito-borne flavivirus that causes Japanese encephalitis (JE), the most commonly diagnosed viral encephalitis worldwide. The fatality rate of JE is 20%, and nearly half of the surviving patients develop neuropsychiatric sequelae. Axl is a receptor tyrosine kinase that plays multiple roles in flaviviral infections. Currently, the involvement of Axl in JEV infection remains enigmatic. In this study, we demonstrate that Axl impedes the pathogenesis of severe JE in mice by maintaining blood-brain-barrier (BBB) integrity and restricting viral neuroinvasion. Furthermore, serum IL-1α is a key mediator of this process and is primarily released by JEV-infected pyroptotic macrophages to elicit BBB breakdown, while an IL-1α antagonist can effectively reduce the incidence of severe JE. Our work uncovers the protective role of Axl in antagonizing severe JE and shows that the use of an IL-1α antagonist may be a promising tactic to prevent severe JE.

Transcriptomic Analysis Suggests the M1 Polarization and Launch of Diverse Programmed Cell Death Pathways in Japanese Encephalitis Virus-Infected Macrophages.

The Japanese encephalitis virus (JEV) is a Culex mosquito-borne flavivirus and is the pathogenic agent of Japanese encephalitis, which is the most important type of viral encephalitis in the world. Macrophages are a type of pivotal innate immunocyte that serve as sentinels and respond quickly to pathogen invasions. However, some viruses like JEV can hijack macrophages as a refuge for viral replication and immune escape. Despite their crucial involvement in early JEV infection, the transcriptomic landscapes of JEV-infected macrophages are void. Here, by using an in situ JEV infection model, we investigate the transcriptomic alteration of JEV-infected peritoneal macrophages. We found that, upon JEV infection, the macrophages underwent M1 polarization and showed the drastic activation of innate immune and inflammatory pathways. Interestingly, almost all the programmed cell death (PCD) pathways were activated, especially the apoptosis, pyroptosis, and necroptosis pathways, which were verified by the immunofluorescent staining of specific markers. Further transcriptomic analysis and TUNEL staining revealed that JEV infection caused apparent DNA damage. The transcriptomic analysis also revealed that JEV infection promoted ROS and RNS generation and caused oxidative stress, which activated multiple cell death pathways. Our work uncovers the pivotal pathogenic roles of oxidative stress and multiple PCD pathways in JEV infection, providing a novel perspective on JEV-host interactions.