Investigation on clinical risk factors of bladder lesion by machine learning based interpretable model.

Bladder lesion commonly occurs in patients with benign prostatic hyperplasia (BPH), and the routine screening of bladder lesion is vital for its timely detection and treatment, in which the risk of bladder lesion progression can be effectively alleviated. However, current clinical methods are inconvenient for routine screening. In this study, we proposed a convenient routine screening method to diagnose bladder lesions based on several clinical risk factors, which can be obtained through non-invasive, easy-to-operate, and low-cost examinations. The contribution of each clinical risk factor was further quantitatively analyzed to understand their impact on diagnostic decision-making. Based on a cohort study of 253 BPH patients with or without bladder lesions, the proposed diagnostic model achieved high accuracy using these clinical risk factors. Bladder compliance, maximum flow rate (Qmax), prostate specific antigen (PSA), and postvoid residual (PVR) were identified as the four most important clinical risk factors. To the best of our knowledge, this is the innovative research to predict bladder lesions based on the risk factors and quantitatively reveal their contributions to diagnostic decision-making. The proposed model has the potential to serve as an effective routine screening tool for bladder lesions in BPH patients, enabling early intervention to prevent lesion progression and improve the quality of life.

Exosomal CTHRC1 from cancer-associated fibroblasts facilitates endometrial cancer progression via ITGB3/FAK signaling pathway.

The emerging tumor microenvironment (TME) is a complex and constantly evolving entity. Cancer-associated fibroblasts (CAFs) are a vital component of the TME with diverse functions. They interact closely with cancer cells through reciprocal signaling and play a crucial role in tumor progression. Exosomes, which contain diverse biological information, are identified as an important mediator of cell-cell communication. This study aimed to investigate how CAF-derived exosomes promote metastasis of endometrial cancer (EC). Our findings revealed that CAF-derived exosomes significantly enhanced EC cell proliferation and migration compared to normal fibroblast-derived exosomes. Quantitative proteomics analysis of CAF/NF-derived exosomes demonstrated differential expression of CTHRC1, a protein overexpressed in multiple tumors, promoting cancer progression through enhanced cell migration and invasion. Exosomal overload of CTHRC1 significantly contributes to EC cell migration. Mechanically, we determined that ITGB3 was immunoprecipitated by CTHRC1 and phosphorylated FAK on Tyr397, which was important for exosomal CTHRC1 mediated migratory ability of EC cells. Overexpression of CTHRC1 in secreted exosomes promotes the metastatic ability of EC cells in mouse models and may be eliminated by Defactinib, an inhibitor of FAK Tyr397 phosphorylation. Moreover, overexpression of CTHRC1 was increased in EC patients, elevating with cancer progression, and correlated with negative tumor prognosis. Our results revealed that CAF mediated endometrial cancer progression is related to high levels of CTHRC1 and exosomal CTHRC1 derived from CAF may be a promising therapeutic strategy for metastatic endometrial cancer.

Crystal structure of the ATPase domain of porcine circovirus type 2 Rep protein.

PCV2 belongs to the genus Circovirus in the family Circoviridae, whose genome is replicated by rolling circle replication (RCR). PCV2 Rep is a multifunctional enzyme that performs essential functions at multiple stages of viral replication. Rep is responsible for nicking and ligating single-stranded DNA and unwinding double-stranded DNA (dsDNA). However, the structure and function of the Rep are still poorly understood, which significantly impedes viral replication research. This study successfully resolved the structure of the PCV2 Rep ATPase domain (PRAD) using X-ray crystallography. Homologous structure search revealed that Rep belonged to the superfamily 3 (SF3) helicase, and multiple conserved residues were identified during sequence alignment with SF3 family members. Simultaneously, a hexameric PRAD model was generated for analysing characteristic structures and sites. Mutation of the conserved site and measurement of its activity showed that the hallmark motifs of the SF3 family influenced helicase activity by affecting ATPase activity and ß-hairpin just caused the loss of helicase activity. The structural and functional analyses of the PRAD provide valuable insights for future research on PCV2 replication and antiviral strategies.

Single-Cell Transcriptome Analysis of Small Cell Neuroendocrine Carcinoma of the Endometrium Reveals ISL1 as a Potential Biomarker for Diagnosis and Treatment.

BACKGROUND: As a dedifferentiated tumor, small cell endometrial neuroendocrine tumors (NETs) are rare and frequently diagnosed at an advanced stage with a poor prognosis. Current treatment recommendations are often extrapolated from histologically similar tumors in other sites or based on retrospective studies. The exploration for diagnostic and therapeutic markers in small cell NETs is of great significance. METHODS: In this study, we conducted single-cell RNA sequencing on a specimen obtained from a patient diagnosed with small cell endometrial neuroendocrine carcinoma (SCNEC) based on pathology. We revealed the cell map and intratumoral heterogeneity of the cancer cells through data analysis. Further, we validated the function of ISL LIM Homeobox 1 (ISL1) in vitro in an established neuroendocrine cell line. Finally, we examined the association between ISL1 and tumor staging in small cell lung cancer (SCLC) patient samples. RESULTS: We observed the significant upregulation of ISL1 expression in tumor cells that showed high expression of the neuroepithelial markers. Additionally, in vitro cell function experiments demonstrated that the high ISL1 expression group exhibited markedly higher cell proliferation and migration abilities compared to the low expression group. Finally, we showed that the expression level of ISL1 was correlated with SCLC stages. CONCLUSIONS: ISL1 protein in NETs shows promise as a potential biomarker for diagnosis and treatment.

A review on the recycling of spent lithium iron phosphate batteries.

Lithium iron phosphate (LFP) batteries have gained widespread recognition for their exceptional thermal stability, remarkable cycling performance, non-toxic attributes, and cost-effectiveness. However, the increased adoption of LFP batteries has led to a surge in spent LFP battery disposal. Improper handling of waste LFP batteries could result in adverse consequences, including environmental degradation and the mismanagement of valuable secondary resources. This paper presents a comprehensive examination of waste LFP battery treatment methods, encompassing a holistic analysis of their recycling impact across five dimensions: resources, energy, environment, economy, and society. The recycling of waste LFP batteries is not only crucial for reducing the environmental pollution caused by hazardous components but also enables the valuable components to be efficiently recycled, promoting resource utilization. This, in turn, benefits the sustainable development of the energy industry, contributes to economic gains, stimulates social development, and enhances employment rates. Therefore, the recycling of discarded LFP batteries is both essential and inevitable. In addition, the roles and responsibilities of various stakeholders, including governments, corporations, and communities, in the realm of waste LFP battery recycling are also scrutinized, underscoring their pivotal engagement and collaboration. Notably, this paper concentrates on surveying the current research status and technological advancements within the waste LFP battery lifecycle, and juxtaposes their respective merits and drawbacks, thus furnishing a comprehensive evaluation and foresight for future progress.

Loss-of-function mutations of SOX17 lead to YAP/TEAD activation-dependent malignant transformation in endometrial cancer.

Aberrant hyperactivation of the Hippo pathway effector YAP/TEAD complex causes tissue overgrowth and tumorigenesis in various cancers, including endometrial cancer (EC). The transcription factor SOX17 (SRY [sex-determining region Y]-box 17) is frequently mutated in EC; however, SOX17 mutations are rare in other cancer types. The molecular mechanisms underlying SOX17 mutation-induced EC tumorigenesis remain poorly understood. Here, we demonstrate that SOX17 serves as a tumor suppressor to restrict the proliferation, migration, invasion, and anchorage-independent growth of EC cells, partly by suppressing the transcriptional outputs of the Hippo-YAP/TEAD pathway. SOX17 binds to TEAD transcription factors through its HMG domain and attenuates the DNA-binding ability of TEAD. SOX17 loss by inactivating mutations leads to the malignant transformation of EC cells, which can be reversed by small-molecule inhibitors of YAP/TEAD or cabozantinib, an FDA-approved drug targeting the YAP/TEAD transcriptional target AXL. Our findings reveal novel molecular mechanisms underlying Hippo-YAP/TEAD pathway-driven EC tumorigenesis, and suggest potential therapeutic strategies targeting the Hippo-YAP/TEAD pathway in SOX17-mutated EC.

To Identify Adenomatous Polyposis Coli Gene Mutation as a Predictive Marker of Endometrial Cancer Immunotherapy.

The adenomatous polyposis coli (APC) gene is the chromatin-remodeling-related gene and a typical tumor suppressor. Patients with a high expression of programmed death-ligand 1 (PD-L1) or a high level of tumor mutational burden (TMB) may benefit from immunotherapy in endometrial cancer (EC). This study aimed to demonstrate the role of APC in the diagnosis and immunotherapy treatment of EC. We performed an integrative analysis of a commercial panel including 520 cancer-related genes on 99 tumors from an endometrial cancer cohort in China and DNA-seq data from The Cancer Genome Atlas (TCGA) to identify new gene mutations as endometrial cancer immunotherapy markers. We found that the significant mutant genes that correlated with the PD-L1 expression and TMB were related to the chromatin state and generated a discovery set having 12 mutated genes, including the APC gene, which was identified as a new marker for immunotherapy. Further analysis revealed that tumors with the APC mutation had high TMB, increased expression of PD-L1, and increased lymphocytic infiltration. Next, we verified that APC has an inactive mutation in EC, which may affect the immune response, including PD-L1 expression, microsatellite instability, and lymphocytic infiltrate. Furthermore, patients with the APC mutation had longer overall survival. Our study demonstrates that APC could play an important role in enhancing the response to endometrial cancer treatment, particularly immunotherapy.

Therapeutic exosomes loaded with SERPINA5 attenuated endometrial cancer cell migration via the integrin ß1/FAK signaling pathway.

BACKGROUND: Metastasis is still the major cause of endometrial cancer (EC)-related death. Because of their biological function and regenerative properties, exosomes have been applied to therapeutic regimens. SERPINA5 expression is downregulated in several tumors and linked to tumor cell migration and invasion. However, the expression and biological functions of SERPINA5 in EC remain unclear. METHODS: The levels of SERPINA5 in plasma exosomes were determined with ELISAs. SERPINA5 expression in EC and its relationship with survival outcomes were analyzed using the TCGA database and clinical EC tissue samples. The effect of SERPINA5 overexpression or exosomal SERPINA5 on EC metastasis was examined by cell migration and invasion assays in vitro. Mechanistically, overexpression of SERPINA5 or high exosomal SERPINA5 levels mediated the regulation of the integrin ß1/FAK signaling pathway in EC cell lines. The therapeutic effect of exosomal SERPINA5 was determined with xenograft models. RESULTS: This study revealed that the level of exosomal SERPINA5 was increased in the circulating plasma of EC patients. In addition, the expression of SERPINA5 was decreased in EC patients with distant metastasis, and low expression of SERPINA5 indicated worse survival. In addition, SERPINA5 was elevated in normal tissues adjacent to EC tumors. Moreover, overexpression of SERPINA5 inhibited metastatic potential of EC cell lines in vitro. Furthermore, SERPINA5 loaded on secreted exosomes reduced the metastatic ability of EC cells. Notably, overexpression of SERPINA5 or high exosomal SERPINA5 levels suppressed EC metastatic potential by suppressing integrin ß1/FAK signaling pathway activation. Finally, exosomal SERPINA5 impeded tumor growth and metastasis in xenograft models. CONCLUSIONS: Our findings revealed that a low level of SERPINA5 expression indicated poor survival outcomes in EC and that exogenous SERPINA5 loading of exosomes may be a novel therapeutic strategy for metastatic EC.

Japanese Encephalitis Virus NS1' Protein Interacts with Host CDK1 Protein to Regulate Antiviral Response.

Type I interferon (IFN-I) is a key component of the host innate immune system. To establish efficient replication, viruses have developed several strategies to escape from the host IFN response. Japanese encephalitis virus (JEV) NS1', a larger NS1-related protein, is known to inhibit the mitochondrial antiviral signaling (MAVS)-mediated IFN-ß induction by increasing the binding of transcription factors (CREB and c-Rel) to the microRNA 22 (miRNA-22) promoter. However, the mechanism by which NS1' induces the recruitment of CREB and c-Rel onto the miRNA-22 promoter is unknown. Here, we found that JEV NS1' protein interacts with the host cyclin-dependent kinase 1 (CDK1) protein. Mechanistically, NS1' interrupts the CDC25C phosphatase-mediated dephosphorylation of CDK1, which prolongs the phosphorylation status of CDK1 and leads to the inhibition of MAVS-mediated IFN-ß induction. Furthermore, the CREB phosphorylation and c-Rel activation through the IκBα phosphorylation were observed to be enhanced upon the augmentation of CDK1 phosphorylation by NS1'. The abrogation of CDK1 activity by a small-molecule inhibitor significantly suppressed the JEV replication in vitro and in vivo. Moreover, the administration of CDK1 inhibitor protected the wild-type mice from JEV-induced lethality but showed no effect on the MAVS-/- mice challenged with JEV. In conclusion, our study provides new insight into the mechanism of JEV immune evasion, which may lead to the development of novel therapeutic options to treat JEV infection. IMPORTANCE Japanese encephalitis virus (JEV) is the main cause of acute human encephalitis in Asia. The unavailability of specific treatment for Japanese encephalitis demands a better understanding of the basic cellular mechanisms that contribute to the onset of disease. The present study identifies a novel interaction between the JEV NS1' protein and the cellular CDK1 protein, which facilitates the JEV replication by dampening the cellular antiviral response. This study sheds light on a novel mechanism of JEV replication, and thus our findings could be employed for developing new therapies against JEV infection.

Integrated Analysis of Microbiome and Transcriptome Data Reveals the Interplay Between Commensal Bacteria and Fibrin Degradation in Endometrial Cancer.

The gut-uterus axis plays a pivotal role in the pathogenesis of endometrial cancer (EC). However, the correlations between the endometrial microbiome and endometrial tumor transcriptome in patients with EC and the impact of the endometrial microbiota on hematological indicators have not been thoroughly clarified. In this prospective study, endometrial tissue samples collected from EC patients (n = 30) and healthy volunteers (n = 10) were subjected to 16S rRNA sequencing of the microbiome. The 30 paired tumor and adjacent nontumor endometrial tissues from the EC group were subjected to RNAseq. We found that Pelomonas and Prevotella were enriched in the EC group with a high tumor burden. By integrating the microbiome and hematological indicators, a correlation was observed between Prevotella and elevated serum D-dimer (DD) and fibrin degradation products (FDPs). Further transcriptome analysis identified 8 robust associations between Prevotella and fibrin degradation-related genes expressed within ECs. Finally, the microbial marker of Prevotella along with DD and FDPs showed a high potential to predict the onset of EC (AUC = 0.86). Our results suggest that the increasing abundance of Prevotella in endometrial tissue combined with high serum DD and FDP contents may be important factors associated with tumor burden. The microbe-associated transcripts of host tumors can partly explain how Prevotella promotes DD and FDPs.

Endometrial Tumors with MSI-H and dMMR Share a Similar Tumor Immune Microenvironment.

PURPOSE: Microsatellite instability (MSI) and mismatch repair deficiency (dMMR) are important biomarkers for predicting responses to immune checkpoint inhibitor (ICI) therapies. Although PCR-based tests for high MSI (MSI-H) and dMMR yield highly concordant results in endometrial cancer (EC), it is unclear whether this is true for MSI-H and MMR detected by next-generation sequencing (NGS) and immunohistochemistry (IHC), respectively. This study investigated whether EC with MSI-H identified by NGS and dMMR identified by IHC have similar tumor immune microenvironments. PATIENTS AND METHODS: EC tissue and corresponding peripheral blood lymphocyte samples were collected from 99 randomly selected patients. MSI status and tumor mutation burden (TMB) were examined by NGS. MMR protein and programmed death ligand (PD-L)1 expression and tumor-infiltrating lymphocyte (TIL) abundance were evaluated by IHC. RESULTS: Of the 99 EC samples, 29 (29%) had dMMR by IHC, while 18 (18%) had MSI-H by NGS. MSI and MMR status identified by the two methods were discordant in the 99 EC patients, and 2/18 NGS-identified MSI-H patients (11%) retained MMR protein expression. MSI-H and dMMR endometrial tumors had similar numbers of cluster of differentiation (CD)3+ TILs (T cells) and CD8+ TILs (cytotoxic T cells) in the tumor center and periphery, which differed from those in microsatellite stable (MSS) and mismatch repair-proficient (pMMR) EC; they also showed similar TMB, PD-L1 expression, and TIL counts with higher TMB and PD-L1 expression than MSS and pMMR ECs. The abundance of CD3+ and CD8+ TILs was increased in PD-L1-positive EC. CONCLUSION: NGS-identified MSI status and IHC-identified MMR status were inconsistent in EC, and 11% of NGS-identified MSI-H tumors retained MMR protein expression. Conversely, MSI and MMR status determined by the two methods provided similar data on TMB, PD-L1 expression, and TIL abundance, which can guide treatment decisions with ICIs.

Plasma exosomes from endometrial cancer patients contain LGALS3BP to promote endometrial cancer progression.

Endometrial cancer (EC) is a common gynaecological cancer worldwide. Exosomes, secreted by living cells and detected in various body fluids, can exchange information between organs and compartments to affect cellular functions, such as proliferation, apoptosis, migration and angiogenesis. We hypothesise that plasma exosomal contents are altered during cancer progression and promote cancer growth and angiogenesis by delivering biomolecules to cancer and vascular endothelial cells. In this study, circulating exosomes derived from EC patients and age-matched healthy people were acquired by commercial kits. Cell counting kit-8, Transwell and Matrigel tube formation assays showed that circulating exosomes from EC patients promote EC cell growth and human umbilical vein endothelial cell (HUVEC) angiogenesis. Next, proteomic analysis and ELISA revealed that plasma exosomal lectin galactoside-binding soluble 3 binding protein (LGALS3BP) increased during EC progression. Moreover, to explore the function of exosomal LGALS3BP, we acquired exosomes containing high levels of LGALS3BP by overexpressing LGALS3BP in human embryonic kidney 293 cells, and we demonstrated that highly contained exosomal LGALS3BP contributed to EC cell proliferation and migration and HUVEC functions via the activation of the PI3K/AKT/VEGFA signalling pathway both in vitro and in vivo. Finally, high LGALS3BP expression was observed in human EC tissue, which indicated a poor prognosis. In addition, immunohistochemical analysis of human EC tissues revealed that LGALS3BP expression was correlated with VEGFA expression and blood vessel density. Hence, we proposed that plasma exosomes containing LGALS3BP contributed to EC growth and angiogenesis during EC progression, which also provided a novel perspective on EC diagnosis and prognosis.

Identification of Interleukin-9 Producing Immune Cells in Endometrial Carcinoma and Establishment of a Prognostic Nomogram.

Background: Interleukin-9 (IL9) plays a critical role in immunity and the pathogenesis of endometrial cancer (EC), especially endometrioid EC (EEC). This study aimed to identify the IL9+ immune cell subsets and their pleiotropic functions and establish an optimized prognostic nomogram towards the promotion of personalized treatment of EEC. Methods: 1,417 EC patients were involved in the present study. 143 patients from the tertiary gynecology centers in Shanghai between 2013 and 2019 were recruited, and the study protocol was approved by the Institutional Review Board (IRB) of Shanghai First Maternity and Infant Hospital. The genomic data of the other 1,274 patients were extracted from the TCGA and the MSKCC datasets, respectively. Immune and stromal scores were calculated using the ESTIMATE R tool, and the tumor infiltration of immune cells was analyzed using the TIMER platform. Metascape and GEPIA datasets were used for bioinformatic analysis. P < 0.05 was considered statistically significant. All statistical analyses were performed with GraphPad Prism and R studio. Results: 552 genes that were correlated with leukocyte infiltration, lymphocyte activation, and regulation of innate immune response were up-regulated in the high immune score group. More IL9+ cell infiltration was detected in the highly and moderately differentiated EC (p = 0.04). High IL9+ lymphocyte infiltration was related to a better overall survival (p = 0.0027). IL9 positive cell clusters included ILC2s, Vδ2 γδT cells, mast cells, macrophages, and Th9 cells. Parameters such as FIGO stage, IL9 score, Vδ2 + γδT cell infiltration, classification of differentiation, and diabetes mellitus were assigned a weighted number of points in the nomogram for a specific predicted 3-, 5- and 10-year overall survival (OS). IL9-IL9R axis played a vital role in EEC, IL9R positive cell subgroups were also identified, and the related function was analyzed in the present study. Additionally, PR (Progesterone Receptor, or PGR) expression was relevant to a higher density of IL9+ lymphocyte infiltration. However, PGRMC1 (Progesterone Receptor Membrane Component 1) was negatively relevant to IL9R (p = 4.26e-8). Conclusion: We observed a significant infiltration of IL9+ cells and the overrepresentation of IL-9R in tissue specimens of patients in EC cases. The nomogram incorporating the IL9 could accurately predict individualized survival probability in EEC. Additionally, this study not only established a prognostic nomogram but also assist in the firmer understanding of the relevance of the IL9-IL9R axis and IL9-producing cells in EC immunity.

Identification of a protective epitope in Japanese encephalitis virus NS1 protein.

Japanese encephalitis virus (JEV) is one of the most important culex transmitted-flaviviruses, which can cause encephalitis in humans. Although non-structural protein 1 (NS1) of JEV does not stimulate neutralizing antibodies, this protein can provide high immunoprotection in vivo. The protective epitopes and the protective mechanism of NS1 still remain unclear. In this study, we generated five different monoclonal antibodies (mAbs) targeting the NS1 protein of JEV. In vitro experiments revealed that none of these five antibodies neutralized the JEV infection. In mouse protection studies, one of these mAbs, designated 2B8, provided a therapeutic effect against JEV lethal challenge (70% survival rate). Using peptide mapping analysis, we found that mAb 2B8 reacted with the epitope 225PETHTLWGD233 in the NS1 protein, in which any mutations among amino acid residues T228, H229, L231 or W232 could cause binding failure of 2B8 to the NS1 protein. Furthermore, mice immunized with KLH-polypeptide (225PETHTLWGD233) showed reduced mortality following JEV challenge. Collectively, we found a new protective epitope in the JEV NS1 protein. These results may facilitate the development of therapeutic agent and subunit-based vaccines based on the NS1 protein.

Benzoxazine monomer derived carbon dots as a broad-spectrum agent to block viral infectivity.

Multiple viruses can cause infection and death of millions annually. Of these, flaviviruses are found to be highly prevalent in recent years with no distinctive antiviral therapies. Therefore, there is a desperate need for broad-spectrum antiviral drugs that can be active against a large number of existing and emerging viruses. Herein, we prepared a kind of benzoxazine monomer derived carbon dots (BZM-CDs) and demonstrated their infection-blocking ability against life-threatening flaviviruses (Japanese encephalitis, Zika, and dengue viruses) and non-enveloped viruses (porcine parvovirus and adenovirus-associated virus). It was found that BZM-CDs could directly bind to the surface of the virion, and eventually the first step of virus-cell interaction was impeded. The developed nanoparticles are active against both flaviviruses and non-enveloped viruses in vitro. Thus, the application of BZM-CDs may constitute an intriguing broad-spectrum approach to rein in viral infections.

Japanese Encephalitis Virus NS1' Protein Antagonizes Interferon Beta Production.

Japanese encephalitis virus (JEV) is a mosquito-borne virus and the major cause of viral encephalitis in Asia. NS1', a 52-amino acid C-terminal extension of NS1, is generated with a -1 programmed ribosomal frameshift and is only present in members of the Japanese encephalitis serogroup of flaviviruses. Previous studies demonstrated that NS1' plays a vital role in virulence, but the mechanism is unclear. In this study, an NS1' defected (rG66A) virus was generated. We found that rG66A virus was less virulent than its parent virus (pSA14) in wild-type mice. However, similar mortality caused by the two viruses was observed in an IFNAR knockout mouse model. Moreover, we found that rG66A virus induced a greater type I interferon (IFN) response than that by pSA14, and JEV NS1' significantly inhibited the production of IFN-ß and IFN-stimulated genes. Taken together, our results reveal that NS1' plays a vital role in blocking type I IFN production to help JEV evade antiviral immunity and benefit viral replication.

High-Throughput Screening of Escherichia coli O157:H7 Pathogenic Genes via Pathway Enrichment and Operon Analysis.

BACKGROUND: About thirty thousand people globally die every day from infectious diarrhea, mostly caused by pathogenic Escherichia coli (E. coli) O157:H7. METHODS: In order to search for clinical diagnostic biomarkers and novel drug targets for infectious diarrhea, we used a bibliometric method to collect pathogenic genes of E. coli O157:H7 and performed a functional analysis of the important pathogenic genes by pathway enrichment and operon analysis. RESULTS: We found 364 pathogenic genes which may be involved in infection with E. coli O157:H7 including 50 new specific pathogenic genes. It is possible that these newly found pathogenic genes will be of great importance in the treatment of E. coli O157:H7 infected diseases and the discovery of novel diagnostic biomarkers. CONCLUSIONS: Our findings also lay a theoretical foundation for the control, diagnosis, and prognosis of pathogenic E. coli related diseases.

p21-Activated Kinase 4 Signaling Promotes Japanese Encephalitis Virus-Mediated Inflammation in Astrocytes.

Japanese encephalitis virus (JEV) targets central nervous system, resulting in neuroinflammation with typical features of neuronal death along with hyper activation of glial cells. Exploring the mechanisms responsible for the JEV-caused inflammatory response remains a pivotal area of research. In the present study, we have explored the function of p21-activated kinase 4 (PAK4) in JEV-mediated inflammatory response in human astrocytes. The results showed that JEV infection enhances the phosphorylation of PAK4 in U251 cells and mouse brain. Knockdown of PAK4 resulted in decreased expression of inflammatory cytokines that include tumor necrosis factor alpha, interleukin-6, interleukin-1ß, and chemokine (C-C motif) ligand 5 and interferon ß upon JEV infection, suggesting that PAK4 signaling promotes JEV-mediated inflammation. In addition, we found that knockdown of PAK4 led to the inhibition of MAPK signaling including ERK, p38 MAPK and JNK, and also resulted in the reduced nuclear translocation of NF-κB and phosphorylation of AP-1. These results demonstrate that PAK4 signaling actively promotes JEV-mediated inflammation in human astrocytes via MAPK-NF-κB/AP-1 pathway, which will provide a new insight into the molecular mechanism of the JEV-induced inflammatory response.

Transcriptional regulation of miR-15b by c-Rel and CREB in Japanese encephalitis virus infection.

MicroRNAs (miRNAs) have been well known to play diverse roles in viral infection at the level of posttranscriptional repression. However, much less is understood about the mechanism by which miRNAs are regulated during viral infection. It is likely that both host and virus contain factors to modulate miRNA expression. Here we report the up-regulation of microRNA-15b (miR-15b) in vitro upon infection with Japanese encephalitis virus (JEV). Analysis of miR-15b precursor, pri-miR-15b and pre-miR-15b, suggest that the regulation occurs transcriptionally. Further, we identified the transcriptional regulatory region of miR-15b that contains consensus binding motif for NF-κB subunit c-Rel and cAMP-response element binding protein (CREB), which are known as transcription factor to regulate gene expression. By promoter fusion and mutational analyses, we demonstrated that c-Rel and CREB bind directly to the promoter elements of miR-15b, which are responsible for miR-15b transcription in response to JEV infection. Finally, we showed that pharmacological inhibition of ERK and NF-κB signaling pathway blocked induction of miR-15b in JEV infection, suggesting important roles of ERK and NF-κB pathway in the regulation of miR-15b gene. Therefore, our observations indicate that induced expression of miR-15b is modulated by c-Rel and CREB in response to JEV infection.