Anti-Influenza A Potential of Tagetes erecta Linn. Extract Based on Bioinformatics Analysis and In Vitro Assays.

Tagetes erecta Linn. (TE) is traditionally used to treat cardiovascular, renal, and gastrointestinal diseases. In this study, we investigated the active compounds and targets of TE extract that may exert antiviral effects against influenza A. Active compounds and targets of TE extract were identified using the Traditional Chinese Medicine Systems Pharmacology database (TCSMP). The influenza A-related gene set was screened using GeneCards and the Kyoto Encyclopedia of Genes and Genomes (KEGG). A protein-protein interaction (PPI) network was built to establish the hub targets. Pathway and target studies were conducted using Gene Expression Omnibus (GEO). The interactions between active compounds and potential targets were assessed by molecular docking. An in vitro study was performed using antiviral and plaque reduction assays. From the compound and target search, we identified 6 active compounds and 95 potential targets. We retrieved 887 influenza-associated target genes and determined 14 intersecting core targets between TE and influenza. After constructing a compound-target network, we discovered lutein and beta-carotene to be the key compounds. Next, PPI network analysis identified the top three hub genes associated with influenza (IL-6, HIF1A, and IL-1ß). Similarly, GEO analysis revealed IL-6, TGFB1, and CXCL8 to be the top three target genes. In our docking study, we identified that lutein and IL-6 had the strongest bindings. Our in vitro experimental results revealed that the TE extract exhibited therapeutic rather than prophylactic effects on influenza disease. We identified lutein as a main active compound in TE extract, and IL-6 as an important target associated with influenza, by using data mining and bioinformatics. Our in vitro findings indicated that TE extract exerted protective properties against the influenza A virus. We speculated that lutein, as a key active component in TE extract, is largely responsible for its antiviral effects. Therefore, we suggest TE extract as an alternative in the treatment of influenza.

Sustained Microglial Activation Promotes Synaptic Loss and Neuronal Dysfunction after Recovery from ZIKV Infection.

Zika virus (ZIKV), transmitted by Aedes mosquitoes, has been a global health concern since 2007. It primarily causes fetal microcephaly and neuronal defects through maternal transmission and induces neurological complications in adults. Recent studies report elevated proinflammatory cytokines and persistent neurological alterations post recovery, but the in vivo mechanisms remain unclear. In our study, viral RNA loads in the brains of mice infected with ZIKV peaked at 7 days post infection and returned to baseline by day 21, indicating recovery. RNA sequencing of the cerebral cortex at 7 and 21 days revealed upregulated genes related to neuroinflammation and microglial activation. Histological analyses indicated neuronal cell death and altered neurite morphology owing to severe neuroinflammation. Additionally, sustained microglial activation was associated with increased phospho-Tau levels, constituting a marker of neurodegeneration. These findings highlight how persistent microglial activation leads to neuronal dysfunction post ZIKV recovery, providing insights into the molecular pathogenesis of ZIKV-induced brain abnormalities.

Integrated CRISPR-Cas9 System-Mediated Knockout of IFN-γ and IFN-γ Receptor 1 in the Vero Cell Line Promotes Viral Susceptibility.

The current pandemic and the possible emergence of new viruses urgently require the rapid development of antiviral vaccines and therapeutics. However, some viruses or newly generated variants are difficult to culture in common cell types or exhibit low viral susceptibility in vivo, making it difficult to manufacture viral vector-based vaccines and understand host-virus interactions. To address these issues, we established new cell lines deficient in both type I and type II interferon responses, which are essential for host immunity and interference with virus replication. These cell lines were generated by developing an integrated CRISPR-Cas9 system that simultaneously expresses dual-guide RNA cassettes and Cas9 nuclease in a single plasmid. Using this highly efficient gene-editing system, we successfully established three cell lines starting from IFN-α/ß-deficient Vero cells, deleting the single interferon-gamma (IFNG) gene, the IFNG receptor 1 (IFNGR1) gene, or both genes. All cell lines clearly showed a decrease in IFN-γ-responsive antiviral gene expression and cytokine production. Moreover, production of IFN-γ-induced cytokines remained low, even after HSV-1 or HCoV-OC43 infection, while expression of the receptor responsible for viral entry increased. Ultimately, knockout of IFN-signaling genes in these cell lines promoted cytopathic effects and increased apoptosis after viral infection up to three-fold. These results indicate that our integrated CRISPR-Cas9-mediated IFNG- and IFNGR1-knockout cell lines promote virus replication and will be useful in viral studies used to design novel vaccines and therapies.

Study on suitable analysis method for HIV-1 non-catalytic integrase inhibitor.

BACKGROUND: Integrase (IN) is an essential protein for HIV replication that catalyzes insertion of the reverse-transcribed viral genome into the host chromosome during the early steps of viral infection. Highly active anti-retroviral therapy is a HIV/AIDS treatment method that combines three or more antiviral drugs often formulated from compounds that inhibit the activities of viral reverse transcriptase and protease enzymes. Early IN inhibitors (INIs) mainly serve as integrase strand transfer inhibitors (INSTI) that disrupt strand transfer by binding the catalytic core domain of IN. However, mutations of IN can confer resistance to INSTI. Therefore, non-catalytic integrase inhibitors (NCINI) have been developed as next-generation INIs. METHODS: In this study, we evaluated and compared the activity of INSTI and NCINI according to the analysis method. Antiviral activity was compared using p24 ELISA with MT2 cell and TZM-bl luciferase system with TZM-bl cell. Each drug was serially diluted and treated to MT2 and TZM-b1 cells, infected with HIV-1 AD8 strain and incubated for 5 and 2 days, respectively. Additionally, to analyze properties of INSTI and NCINI, transfer inhibition assay and 3'-processing inhibition assay were performed. RESULTS: During screening of INIs using the p24 ELISA and TZM-bl luciferase systems, we found an inconsistent result with INSTI and NCINI drugs. Following infection of MT2 and TZM-bl cells with T-tropic HIV-1 strain, both INSTI and NCINI treatments induced significant p24 reduction in MT2 cells. However, NCINI showed no antiviral activity in the TZM-bl luciferase system, indicating that this widely used and convenient antiretroviral assay is not suitable for screening of NCINI compounds that target the second round of HIV-1 replication. CONCLUSION: Accordingly, we recommend application of other assay procedures, such as p24 ELISA or reverse transcription activity, in lieu of the TZM-bl luciferase system for preliminary NCINI drug screening. Utilization of appropriate analytical methods based on underlying mechanisms is necessary for accurate assessment of drug efficacy.

Identification and characterization of a Toll-like receptor gene from Macrobrachium nipponense.

Outbreaks of infectious disease in shrimp pose a serious threat to shrimp agriculture worldwide. Shrimp lack adaptive immunity and depend only on innate immunity as a defense system against infectious disease. Toll-like receptors (TLR) are reported to play a critical role in the innate immune system. In this study, we identified a Toll-like receptor gene of a species of freshwater shrimp, Macrobrachium nipponense, designated MnToll, for the first time. The sequence of MnToll encoded 935 residues arranged as 10 leucine-rich repeat (LRR) domains, a leucine-rich repeat C-terminal (LRR CT) domain and a Toll/interleukin-1 receptor (TIR) domain and displayed 90% amino acid similarity to previously identified TLRs (Toll 1 and 2) of Macrobrachium rosenbergii. We additionally evaluated mRNA expression of MnToll in various tissues, including heart, gills, stomach, digestive gland, ventral nerve cord, antennal gland and muscle. Following infection with a viral pathogen, white spot syndrome virus (WSSV), MnToll expression was significantly upregulated between 12 and 72 h. Our data collectively suggest that the newly identified MnToll gene belongs to the TLR family in shrimp and is potentially involved in innate host defense, especially against WSSV.

Porcine endogenous retrovirus envelope coated baculoviral DNA vaccine against porcine reproductive and respiratory syndrome virus.

PERV is a major virus concerning xenotransplantation study. However, the interesting part is that PERV is present in all kinds of pigs without pathogenicity and immune response. Furthermore, since pig cells have receptors for PERV, the gene delivery system using PERV envelope is highly likely to develop into an excellent viral vector in pigs. We developed a recombinant baculovirus with a modified surface for expressing the porcine endogenous retrovirus (PERV) envelope. Porcine reproductive and respiratory syndrome virus (PRRSV) infection is a severe concern in the porcine industry due to reproduction failure and respiratory symptoms. GP5 and M proteins are major immunogenic proteins of PRRSV. Using PERV-modified baculovirus (Ac mPERV) as a delivery vector, we constructed a dual antigen (GP5 and M)-encoding DNA vaccine system, Ac mPERV-C5/C6. Intramuscular immunization in mice and pigs, Ac mPERV-C5/C6 induced comparative high humoral and cellular immune responses. Our results support further development of Ac mPERV-C5/C6 as a potential PRRSV vaccine in the porcine industry. In addition, the Ac mPERV system may be applied to the generation of other effective DNA vaccines against porcine viral diseases.

Identifying Active Compounds and Targets of Fritillariae thunbergii against Influenza-Associated Inflammation by Network Pharmacology Analysis and Molecular Docking.

Complications due to influenza are often associated with inflammation with excessive release of cytokines. The bulbs of Fritillariae thunbergii (FT) have been traditionally used to control airway inflammatory diseases, such as bronchitis and pneumonia. To elucidate active compounds, the targets, and underlying mechanisms of FT for the treatment of influenza-induced inflammation, systems biology was employed. Active compounds of FT were identified through the TCMSP database according to oral bioavailability (OB) and drug-likeness (DL) criteria. Other pharmacokinetic parameters, Caco-2 permeability (Caco-2), and drug half-life (HL) were also identified. Biological targets of FT were retrieved from DrugBank and STITCH databases, and target genes associated with influenza, lung, and spleen inflammation were collected from DisGeNET and NCBI databases. Compound-disease-target (C-D-T) networks were constructed and merged using Cytoscape. Target genes retrieved from the C-D-T network were further analyzed with GO enrichment and KEGG pathway analysis. In our network, GO and KEGG results yielded two compounds (beta-sitosterol (BS) and pelargonidin (PG)), targets (PTGS1 (COX-1) and PTGS2 (COX-2)), and pathways (nitric oxide, TNF) were involved in the inhibitory effects of FT on influenza-associated inflammation. We retrieved the binding affinity of each ligand-target, and found that PG and COX-1 showed the strongest binding affinity among four binding results using a molecular docking method. We identified the potential compounds and targets of FT against influenza and suggest that FT is an immunomodulatory therapy for influenza-associated inflammation.

Construction of the safe neutralizing assay system using pseudotyped Nipah virus and G protein-specific monoclonal antibody.

Nipah virus (NiV) is a recently emerged paramyxovirus that causes acute respiratory illness and fatal encephalitis in a broad spectrum of vertebrates, including humans. Due to its high pathogenicity and mortality rates, NiV requires handling in biosafety level-4 (BSL-4) containment facilities and no effective vaccines or therapeutic agents are currently available. Since current diagnostic tests for detecting serum neutralizing antibodies against NiV mainly employ live viruses, establishment of more safe and robust alternative diagnostic methods is an essential medical requirement. Here, we have developed a pseudotyped NiV and closely related Hendra virus (HeV) expressing envelope attachment (G) and fusion (F) glycoproteins using the Moloney murine leukemia virus (MuLV) packaging system. We additionally generated polyclonal antibodies (pAbs) against NiV-G and HeV-G and assessed their neutralizing activities for potential utilization in the pseudovirus-based neutralization assay and further application in the serum diagnostic test. To enhance the specificity of neutralizing antibody and sensitivity of the serological diagnostic test, monoclonal antibodies (mAbs) against NiV-G were generated, and among which four out of six mAb clones showed significant reactivity. Specifically, the 7G9 clone displayed the highest sensitivity. The selected mAb clones showed no cross-reactivity with HeV-G and efficient neutralizing activities against pseudotyped NiV. These results validate the safety and specificity of neutralization assays against NiV and HeV and present a useful tool to design effective vaccines and serological diagnosis.

Identification of peptide based B-cell epitopes in Zika virus NS1.

Zika virus (ZIKV), a mosquito-borne flavivirus that has recently emerged globally, poses a major threat to public health. To control this emerging disease, accurate diagnostics are required for monitoring current ZIKV outbreaks. Owing to the high nucleotide sequence similarity and cross-reactivity of ZIKV with other members of the Flaviviridae family, discrimination from other flavivirus infections is often difficult in endemic areas. ZIKV NS1 induces major virus-specific antibodies and is therefore utilized as a serological marker for ZIKV diagnosis. To identify ZIKV specific epitopes for clinical application, 33 NS1 peptides that are 15-30 amino acid in length covering whole NS1 were synthesized and analyzed linear B-cell epitopes with 38 human serum samples (20 ZIKV-positive and 18 ZIKV-negative). As a result of screening, eight epitope regions were identified. In particular, the Z8 and Z14 peptides located in the ß-ladder surface region showed higher levels of binding activity in ZIKV-positive sera without cross-reactivity to other flaviviruses. These identified sensitive and specific epitopes provide a tool for design of diagnostics and structure-based vaccine antigens for ZIKV infection.

Fusion of flagellin 2 with bivalent white spot syndrome virus vaccine increases survival in freshwater shrimp.

Despite large economic losses attributable to white spot syndrome virus (WSSV), an infectious pathogen of penaeid shrimp and other crustaceans worldwide, no efficient vaccines or antiviral agents to control the virus are available at present. Here, we designed and constructed baculovirus-based vaccines delivering genes encoding the WSSV envelope proteins, VP28 and VP19. To enhance the immunogenicity of the baculovirus-based vaccine, we fused a Salmonella typhimurium flagellin 2 (FL2) gene with VP28 or VP19 gene. Both vaccine constructs elicited similar high titlers of anti-WSSV IgG after oral immunization in mice. The protective effect of oral vaccines upon WSSV challenge was observed in Macrobrachium nipponense. Bivalent vaccine displaying WSSV envelope proteins, VP19 and VP28, led to enhanced more than 10% survival protection against WSSV infection, compared to monovalent vaccine containing WSSV envelope protein, VP19 or VP28. Furthermore, a baculovirus-based WSSV vaccine fused with FL2 gene, Ac-VP28-ie1VP19FL2, efficiently protected mice against WSSV challenge (89.5% survival rate). In support of the efficacy of FL2 in our vaccine, we verified FL2 enhanced survival rate and induced the NF-κB gene in Palaemon paucidens. The collective results strongly suggest that our recombinant baculoviral system displaying WSSV envelope protein and delivering FL2-fused WSSV envelope gene effectively induced protective responses, supporting the utility of a potential new oral DNA vaccine against WSSV.

Development and evaluation of a semifield test for repellent efficacy testing.

Estimation of the efficacy of mosquito repellents requires both laboratory and field tests. The results of field tests are more meaningful, but the safety of volunteers in such tests may be a significant concern. In the current study, we compared tests of mosquito repellent efficacy under semifield conditions in an outdoor enclosure with those under laboratory and field conditions. In this study, we assessed the efficacy of N,N-diethyl-meta-toluamide under laboratory conditions with human volunteers and under semifield and field conditions with Centers for Disease Control and Prevention traps and experimental mice. A semifield test may be a suitable replacement for the more difficult field test for assessment of mosquito repellent efficacy. Semifield tests should be considered when developing new guidelines for testing.

Comparative Biodistribution Study of Baculoviral and Adenoviral Vector Vaccines against SARS-CoV-2.

Various types of vaccines have been developed against COVID-19, including vector vaccines. Among the COVID-19 vaccines, AstraZeneca's chimpanzee adenoviral vaccine was the first to be commercialized. For viral vector vaccines, biodistribution studies are critical to vaccine safety, gene delivery, and efficacy. This study compared the biodistribution of the baculoviral vector vaccine (AcHERV-COVID19) and the adenoviral vector vaccine (Ad-COVID19). Both vaccines were administered intramuscularly to mice, and the distribution of the SARS-CoV-2 S gene in each tissue was evaluated for up to 30 days. After vaccination, serum and various tissue samples were collected from the mice at each time point, and IgG levels and DNA copy numbers were measured using an enzyme-linked immunosorbent assay and a quantitative real-time polymerase chain reaction. AcHERV-COVID19 and Ad-COVID19 distribution showed that the SARS-CoV-2 spike gene remained predominantly at the injection site in the mouse muscle. In kidney, liver, and spleen tissues, the AcHERV-COVID19 group showed about 2-4 times higher persistence of the SARS-CoV-2 spike gene than the Ad-COVID19 group. The distribution patterns of AcHERV-COVID19 and Ad-COVID19 within various organs highlight their contrasting biodistribution profiles, with AcHERV-COVID19 exhibiting a broader and prolonged presence in the body compared to Ad-COVID19. Understanding the biodistribution profile of AcHERV-COVID19 and Ad-COVID19 could help select viral vectors for future vaccine development.

Delivery of SARS-CoV-2 spike and membrane genes in a single Baculoviral vector enhance the immune breadth against SARS-CoV-2 variants of concern.

Although the coronavirus pandemic has ended, new variants of concern (VOCs) continue to emerge. Therefore, novel vaccines targeting VOCs are highly warranted. We initially constructed three recombinant baculovirus-vectored vaccines (AcHERV-COVID19S) carrying the spike genes of the SARS-CoV-2 prototype, Delta, and Omicron BA.1 variants. However, the SARS-CoV-2 spike gene alone could not provide protection against multiple VOCs. To develop a universal vaccine, we constructed a recombinant baculovirus-vectored vaccine (AcHERV-COVID19 OmiM) by introducing the M gene, which is conserved among VOCs, as a secondary cellular immune antigen in addition to the S gene. AcHERV-COVID19 OmiM could provide higher protection against SARS-CoV-2 variants (prototype, Delta, BA.5 and XBB.1) compared with that of AcHERV-COVID19S. The membrane protein of SARS-CoV-2 synergizes with the S gene, thereby enhancing both humoral and cellular immunity against VOCs. Although AcHERV-COVID19 OmiM may not provide sterile protection against new variants, it may help reduce symptoms and curb viral transmission.

Generation of rescued Japanese encephalitis virus genotype 1 from infectious full-size clone using reverse genetics.

Japanese encephalitis virus (JEV) is a pathogen responsible for high mortality and morbidity rates among children with encephalitis. Since JEV genotype 1 (GI) is the most prevalent strain in South Korea these days, corresponding research and vaccine development is urgently required. Molecular genetic studies on JEV vaccines can be boosted by obtaining genetically stable full-length infectious JEV complementary DNA (cDNA) clones. Furthermore, the significance of the reverse genetics system in facilitating molecular biological analyses of JEV properties has been demonstrated. This study constructed a recombinant JEV-GI strain using a reverse genetics system based on a Korean wild-type GI isolate (K05GS). RNA extracted from JEV-GI was used to synthesize cDNA, a recombinant full-length JEV clone, pTRE-JEVGI, was generated from the DNA fragment, and the virus was rescued. We performed in vitro and in vivo experiments to analyze the rescued JEV-GI virus. The rescued JEV-GI exhibited similar characteristics to wild-type JEV. These results suggest that our reverse genetics system can generate full-length infectious clones that can be used to analyze molecular biological factors that influence viral properties and immunogenicity. Additionally, it may be useful as a heterologous gene expression vector and help develop new strains for JEV vaccines.

Baculovirus Vector-Based Varicella-Zoster Virus Vaccine as a Promising Alternative with Enhanced Safety and Therapeutic Functions.

Varicella-zoster virus (VZV) poses lifelong risks, causing varicella and herpes zoster (HZ, shingles). Currently, varicella and HZ vaccines are predominantly live attenuated vaccines or adjuvanted subunit vaccines utilizing VZV glycoprotein E (gE). Here, we propose our vaccine candidates involving a comparative analysis between recombinant baculoviral vector vaccines (AcHERV) and a live attenuated vaccine strain, vOka. AcHERV vaccine candidates were categorized into groups encoding gE only, VZV glycoprotein B (gB) only, or both gE and gB (gE-gB) as AcHERV-gE, AcHERV-gB, and AcHERV-gE-gB, respectively. Humoral immune responses were evaluated by analyzing total IgG, IgG1, IgG2a, and neutralizing antibodies. Cell-mediated immunity (CMI) responses were evaluated by enzyme-linked immunospot (ELISPOT) assay and Th1/Th2/Th17 cytokine profiling. In the mouse model, AcHERV-gE-gB elicited similar or higher total IgG, IgG2a, and neutralizing antibody levels than vOka and showed robust VZV-specific CMI responses. From the perspective of antigens encoded in vaccines and their relationship with CMI response, both AcHERV-gB and AcHERV-gE-gB demonstrated results equal to or superior to AcHERV-gE, encoding only gE. Taken together, these results suggest that AcHERV-gE-gB can be a novel candidate for alleviating risks of live attenuated vaccine-induced latency and effectively preventing varicella during early stages of life while providing strong CMI for effective resistance against HZ and therapeutic potential in later stages of life.

METTL18 functions as a Phenotypic Regulator in Src-Dependent Oncogenic Responses of HER2-Negative Breast Cancer.

Methyltransferase-like (METTL)18 has histidine methyltransferase activity on the RPL3 protein and is involved in ribosome biosynthesis and translation elongations. Several studies have reported that actin polymerization serves as a Src regulator, and HSP90 is involved in forming polymerized actin bundles. To understand the role of METTL18 in breast cancer and to demonstrate the importance of METTL18 in HER-2 negative breast cancer metastasis, we used biochemical, molecular biological, and immunological approaches in vitro (breast tumor cell lines), in vivo (tumor xenograft model), and in samples of human breast tumors. A gene expression comparison of 31 METTL series genes and 22 methyltransferases in breast cancer patients revealed that METTL18 is highly amplified in human HER2-negative breast cancer. In addition, elevated levels of METTL18 expression in patients with HER2-negative breast cancer are associated with poor prognosis. Loss of METTL18 significantly reduced the metastatic responses of breast tumor cells in vitro and in vivo. Mechanistically, METTL18 indirectly regulates the phosphorylation of the proto-oncogene tyrosine-protein kinase Src and its downstream molecules in MDA-MB-231 cells via METTL18-mediated RPL3 methylation, which is also involved in determining HSP90 integrity and protein levels. In confocal microscopy and F/G-actin assays, METTL18 was found to induce actin polymerization via HSP90. Molecular events involving METTL18, RPL3, HSP90, and actin polymerization yielded Src phosphorylated at both tyrosine 419 and tyrosine 530 with kinase activity and oncogenic functions. Therefore, it is suggested that the METTL18-HSP90-Actin-Src regulatory axis plays critical oncogenic roles in the metastatic responses of HER2-negative breast cancer and could be a promising therapeutic target.

Timing of maternal vaccination against COVID-19 for effective protection of neonates: cohort study.

Introduction: Although the safety and effectiveness of COVID-19 vaccination during pregnancy have been proven, there is still little data explaining neonatal outcomes of maternal pre-pregnancy vaccination. Methods: Here, we investigated the impact of vaccination and SARS-CoV-2 infection on maternal-neonate immune response in a cohort study involving 141 pregnant individuals, and defined the importance of maternal COVID-19 vaccination timing for its effectiveness. Results and discussion: Our data indicate that vertically transferred maternal hybrid immunity provides significantly better antiviral protection for a neonate than either maternal post-infection or post-vaccination immunity alone. Higher neutralization potency among mothers immunized before pregnancy and their newborns highlights the promising role of pre-pregnancy vaccination in neonatal protection. A comparison of neutralizing antibody titers calculated for each dyad suggests that infection and pre-/during-pregnancy vaccination all support transplacental transfer, providing the offspring with strong passive immunity against SARS-CoV-2. Analysis of neutralizing antibody levels in maternal sera collected during pregnancy and later during delivery shows that immunization may exert a positive effect on maternal protection.

Initial preclinical safety of non-replicating human endogenous retrovirus envelope protein-coated baculovirus vector-based vaccines against human papillomavirus.

Human endogenous retrovirus (HERV) envelope protein-coated, baculovirus vector-based HPV 16 L1 (AcHERV-HPV16L1) is a non-replicating recombinant baculoviral vaccine. Here, we report an initial evaluation of the preclinical safety of AcHERV-HPV16L1 vaccine. In an acute toxicity study, a single administration of AcHERV-HPV16L1 DNA vaccine given intramuscularly (i.m.) to mice at a dose of 1 × 10(8) plaque-forming units (PFU) did not cause significant changes in body weight compared with vehicle-treated controls. It did cause a brief increase in the weights of some organs on day 15 post-treatment, but by day 30, all organ weights were not significantly different from those in the vehicle-treated control group. No hematological changes were observed on day 30 post-treatment. In a range-finding toxicity study with three doses of 1 × 10(7) , 2 × 10(7) and 5 × 10(7) PFU once daily for 5 days, the group treated with 5 × 10(7) PFU showed a transient decrease in the body weights from day 5 to day 15 post-treatment, but recovery to the levels similar to those in the vehicle-treated control group by post-treatment day 20. Organ weights were slightly higher for lymph nodes, spleen, thymus and liver after repeated dosing with 5 × 10(7) PFU on day 15, but had normalized by day 30. Moreover, repeated administration of AcHERV-HPV16L1 did not induce myosin-specific autoantibody in serum, and did not cause immune complex deposition or tissue damage at injection sites. Taken together, these results provide preliminary evidence of the preclinical safety of AcHERV-based HPV16L1 DNA vaccines in mice.

Elucidating anti-aging and antioxidant activity of Hydnocarpus anthelmintica on skin aging and cosmeceutical potentials: in silico and in vitro study.

INTRODUCTION: Hydnocarpus anthelmintica (HA) has been traditionally used for treating leprosy and is known for its antioxidant and anti-inflammatory activities. The aim of this study was to investigate the active compounds and targets of HA extracts, involved in oxidative stress and skin aging. The active compounds and targets of HA extracts were identified using network pharmacology. METHOD: The pathway study was conducted using the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. HA semen was measured for its in-vitro antioxidant activity using 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay and anti-aging activities using collagenase, elastase, and tyrosinase assays. A total of 21 intersecting core targets were identified from 8 compounds, 51 action targets, and 1810 skin aging and oxidative stress-associated target genes. RESULT: A compound-target network was constructed, and 3 compounds (luteolin, beta-carotene and genkwanin), and 4 hub genes (TP53, HSP90AA1, JUN, and MAPK1) were identified. The KEGG pathway study revealed that the compounds were correlated with PI3K-Akt, p53, HIF-1, and MAPK signaling. CONCLUSION: The results of in-vitro experiments showed the effect of HA extract on oxidative stress reduction and collagenase inhibition. We discovered two main active compounds, luteolin and ß-carotene, that may be involved in p53 and MAPK signaling, and showed HA extract activity against oxidative stress and collagenase.

Attenuated Chimeric GI/GIII Vaccine Candidate against Japanese Encephalitis Virus.

Japanese encephalitis (JE) is a very severe disease characterized by high fatality rates and the development of permanent behavioral, psychiatric, and neurological sequelae among survivors. Japanese encephalitis virus (JEV), a flavivirus, is responsible for JE. In Asia, Genotype I (GI) has emerged as the dominant strain, replacing Genotype III (GIII). However, no clinically approved drug is available to treat JEV infection, and currently available commercial vaccines derived from JEV GIII strains provide only partial protection against GI. Utilizing a reverse genetics system, this study attempted to produce a novel chimeric JEV strain with high efficacy against JEV GI. Accordingly, a GI/GIII intertypic recombinant strain, namely SA14-GI env, was generated by substituting the E region of the GIII SA14-14-2 strain with that of the GI strain, K05GS. The neurovirulence of the mutant virus was significantly reduced in mice. Analysis of the immunogenicity of the chimeric virus revealed that it induced neutralizing antibodies against JEV GI in mice, and the protective efficacy of SA14-GI env was higher than that of SA14-14-2. These findings suggest that SA14-GI env may be a safe and effective live-attenuated vaccine candidate against JEV GI.