Licochalcone A Induces Ferroptosis in Hepatocellular Carcinoma via Reactive Oxygen Species Activated by the SLC7A11/GPX4 Pathway.

Liver cancer is a common malignant tumor, and its incidence is increasing yearly. Millions of people suffer from liver cancer annually, which has a serious impact on global public health security. Licochalcone A (Lico A), an important component of the traditional Chinese herb licorice, is a natural small molecule drug with multiple pharmacological activities. In this study, we evaluated the inhibitory effects of Lico A on hepatocellular carcinoma cell lines (HepG2 and Huh-7), and explored the inhibitory mechanism of Lico A on hepatocellular carcinoma. First, we evaluated the inhibitory effects of Lico A on hepatocellular carcinoma, and showed that Lico A significantly inhibited and killed HepG2 and Huh-7 cells in vivo and in vitro. Transcriptomic analysis showed that Lico A inhibited the expression of solute carrier family 7 member 11 (SLC7A11), which induced ferroptosis. We confirmed through in vivo and in vitro experiments that Lico A promoted ferroptosis in hepatocellular carcinoma cells by downregulating SLC7A11 expression, thereby inhibiting the glutathione (GSH)-glutathione peroxidase 4 (GPX4) pathway and inducing activation of reactive oxygen species (ROS). In this study, we suggest that Lico A is a potential SLC7A11 inhibitor that induces ferroptotic death in hepatocellular carcinoma cells, thereby providing a theoretical basis for the development of natural small molecule drugs against hepatocellular carcinoma.

Immunogenicity and protective efficacy of a DNA vaccine inducing optimal expression of the SARS-CoV-2 S gene in hACE2 mice.

The wide spread of coronavirus disease 2019 (COVID-19) has significantly threatened public health. Human herd immunity induced by vaccination is essential to fight the epidemic. Therefore, highly immunogenic and safe vaccines are necessary to control SARS-CoV-2, whose S protein is the antigenic determinant responsible for eliciting antibodies that prevent viral entry and fusion. In this study, we developed a SARS-CoV-2 DNA vaccine expressing the S protein, named pVAX-S-OP, which was optimized according to the human-origin codon preference and using polyinosinic-polycytidylic acid as an adjuvant. pVAX-S-OP induced specific antibodies and neutralizing antibodies in BALB/c and hACE2 transgenic mice. Furthermore, we observed 1.43-fold higher antibody titers in mice receiving pVAX-S-OP plus adjuvant than in those receiving pVAX-S-OP alone. Interferon gamma production in the pVAX-S-OP-immunized group was 1.58 times (CD3+CD4+IFN-gamma+) and 2.29 times (CD3+CD8+IFN-gamma+) lower than that in the pVAX-S-OP plus adjuvant group but higher than that in the control group. The pVAX-S-OP vaccine was also observed to stimulate a Th1-type immune response. When, hACE2 transgenic mice were challenged with SARS-CoV-2, qPCR detection of N and E genes showed that the viral RNA loads in pVAX-S-OP-immunized mice lung tissues were 104 times and 106 times lower than those of the PBS control group, which shows that the vaccine could reduce the amount of live virus in the lungs of hACE2 mice. In addition, pathological sections showed less lung damage in the pVAX-S-OP-immunized group. Taken together, our results demonstrated that pVAX-S-OP has significant immunogenicity, which provides support for developing SARS-CoV-2 DNA candidate vaccines.

GII.P16-GII.2 Recombinant Norovirus VLPs Polarize Macrophages Into the M1 Phenotype for Th1 Immune Responses.

Norovirus (NoV) is a zoonotic virus that causes diarrhea in humans and animals. Outbreaks in nosocomial settings occur annually worldwide, endangering public health and causing serious social and economic burdens. The latter quarter of 2016 witnessed the emergence of the GII.P16-GII.2 recombinant norovirus throughout Asia. This genotype exhibits strong infectivity and replication characteristics, proposing its potential to initiate a pandemic. There is no vaccine against GII.P16-GII.2 recombinant norovirus, so it is necessary to design a preventive vaccine. In this study, GII.P16-GII.2 type norovirus virus-like particles (VLPs) were constructed using the baculovirus expression system and used to conduct immunizations in mice. After immunization of mice, mice were induced to produce memory T cells and specific antibodies, indicating that the VLPs induced specific cellular and humoral immune responses. Further experiments were then initiated to understand the underlying mechanisms involved in antigen presentation. Towards this, we established co-cultures between dendritic cells (DCs) or macrophages (Mø) and naïve CD4+T cells and simulated the antigen presentation process by incubation with VLPs. Thereafter, we detected changes in cell surface molecules, cytokines and related proteins. The results indicated that VLPs effectively promoted the phenotypic maturation of Mø but not DCs, as indicated by significant changes in the expression of MHC-II, costimulatory factors and related cytokines in Mø. Moreover, we found VLPs caused Mø to polarize to the M1 type and release inflammatory cytokines, thereby inducing naïve CD4+ T cells to perform Th1 immune responses. Therefore, this study reveals the mechanism of antigen presentation involving GII.P16-GII.2 recombinant norovirus VLPs, providing a theoretical basis for both understanding responses to norovirus infection as well as opportunities for vaccine development.

Construction and Immunogenicity of Recombinant Vaccinia Virus Vaccine Against Japanese Encephalitis and Chikungunya Viruses Infection in Mice.

Japanese encephalitis virus (JEV) is recognized as a public health risk by the World Health Organization. In Asia, each year, ∼70,000 people become infected with JEV, which results in ∼10,000 deaths. Chikungunya virus (CHIKV) is an RNA virus, whose infection mainly causes fever, myalgia, and skin rash. Although the mortality rate is low, it seriously affects daily life. JEV and CHIKV infect humans through mosquitoes; therefore, a recombinant vaccinia virus coexpressing JEV E and CHIKV E1 proteins was constructed to prevent their concurrent infection. In this study, after mice first immunization, booster immunization was performed at 21 days postimmunization (dpi). At 35 dpi, mice were challenged with JEV and CHIKV. Specific antibodies significantly increased in the rVTT-CE1-JE-EGFP group, which were significantly (p < 0.01) higher than those of the control groups at 35 dpi. The plaque reduction neutralization tests (JEV) of rVTT-CE1-JE-EGFP group was 1:320 at 35 dpi. Furthermore, cytokine levels and the percentage of CD3+CD4+ and CD3+CD8+ T-lymphocytes in the rVTT-CE1-JE-EGFP group were significantly (p < 0.01) higher than those in the control groups at 35 dpi. After challenge, mice body weights in rVTT-CE1-JE-EGFP group were not significantly altered, and the survival rate was 100%. These results showed the rVTT-CE1-JE-EGFP group elicited significant humoral and cellular immune responses, thus indicating that the recombinant vaccine may serve as a candidate for effective prevention of CHIKV and JEV infection.

Pathogenicity of porcine reproductive and respiratory syndrome virus (ORF5 RFLP 1-7-4 viruses) in China.

Porcine reproductive and respiratory syndrome virus (PRRSV) is an RNA virus that causes reproductive failure in sows and respiratory problems in piglets. PRRSV infection leads to substantial pig mortality and causing huge economic losses so that disease outbreaks caused by the new PRRSV strain from other regions have caused great concern in China. In this study, we analysed the pathogenicity of the novel ORF5 RFLP 1-7-4-like PRRSV strain, named PRRSV-ZDXYL-China-2018-1 in pigs. The viral challenge test showed that PRRSV-ZDXYL-China-2018-1 infection can cause persistent fever, moderate dyspnoea, serum viraemia and interstitial pneumonia in piglets. The levels of viral loads in serum and PRRSV-specific antigen were also detected in lung tissues were used one-step Taq-Man RT-qPCR and Immunohistochemistry, respectively. At 28dpi, the level of specific antibodies was increased among infected piglets. Importantly, the new virus appeared be a moderately virulent isolate with pathogenicity compared to HP-PRRSV strain LQ (JXA1-like strain). Histological examination revealed severe monocyte haemorrhage and interstitial pneumonia associated with monocyte infiltration in the lung tissue of pigs infected with PRRSV-ZDXYL-China-2018-1 and LQ-JXA1 strains. Immunohistochemistry (IHC) results showed positive brown-red epithelial cells and macrophages in pig lungs. Therefore, it is critical to establish an effective strategy to control the spread of PRRSV in China.

First detection and genomic characterization of porcine circovirus 3 in mosquitoes from pig farms in China.

The porcine circovirus type 3 (PCV3) becomes an important causative agent of swine disease since its discovery in 2016. PCV3 infection exhibits a wide range of clinical syndromes causing substantial economic losses in swine industry. Previous studies have reported the detection of numerous known viruses including circovirus in mosquitoes. However, the transmission of PCV3 in field-caught mosquitoes remains largely unknown. This study aims to detect PCV3 infection in mosquitoes and analyze its genomic characteristics. Here, we performed a PCR to detect the PCV3 in 269 mosquito samples collected from pig farms located in Heilongjiang, Jilin, and Yunnan provinces. The proportion of PCV3-positive mosquitoes was 32.0 % (86/269), ranging from 21.4%-42.5% at farm level, which may imply that mosquito serves as a route of transmission for PCV3. To determine the possible origin of PCV3 in mosquitoes, 80 pig serum samples were collected from the pig farms where mosquito sampling was also performed. The proportion of PCV3-positive farms ranged from 15.0%-30.0 % in which infection of positive pigs positively correlated with mosquitoes carrying the virus. Additionally, we sequenced the entire genome of 6 strains of PCV3 in mosquitoes and 2 strains of PCV3 in pigs. Sequence analysis indicated a 100 % nucleotide similarity between mosquito and pig viral isolates that were all collected from similar farms. Phylogenetic analysis showed that PCV3 could be divided into two clades, PCV3a and PCV3b, and the PCV3 strains isolated in mosquitoes were distributed on the two clades. Our results demonstrate that mosquitoes may serve as a potential transmission vector in the life-cycle of PCV3, revealing possible transmission routes of PCV3.

Newcastle Disease Virus Inhibits the Proliferation of T Cells Induced by Dendritic Cells In Vitro and In Vivo.

Newcastle disease virus (NDV) infects poultry and antagonizes host immunity via several mechanisms. Dendritic cells (DCs) are characterized as specialized antigen presenting cells, bridging innate and adaptive immunity and regulating host resistance to viral invasion. However, there is little specific knowledge of the role of DCs in NDV infection. In this study, the representative NDV lentogenic strain LaSota was used to explore whether murine bone marrow derived DCs mature following infection. We examined surface molecule expression and cytokine release from DCs as well as proliferation and activation of T cells in vivo and in vitro in the context of NDV. The results demonstrated that infection with lentogenic strain LaSota induced a phenotypic maturation of immature DCs (imDCs), which actually led to curtailed T cell responses. Upon infection, the phenotypic maturation of DCs was reflected by markedly enhanced MHC and costimulatory molecule expression and secretion of proinflammatory cytokines. Nevertheless, NDV-infected DCs produced the anti-inflammatory cytokine IL-10 and attenuated T cell proliferation, inducing Th2-biased responses. Therefore, our study reveals a novel understanding that DCs are phenotypically mature but dysfunctional in priming T cell responses during NDV infection.