Heterologous vaccination utilizing viral vector and protein platforms confers complete protection against SFTSV.

Severe fever with thrombocytopenia syndrome virus was first discovered in 2009 as the causative agent of severe fever with thrombocytopenia syndrome. Despite its potential threat to public health, no prophylactic vaccine is yet available. This study developed a heterologous prime-boost strategy comprising priming with recombinant replication-deficient human adenovirus type 5 (rAd5) expressing the surface glycoprotein, Gn, and boosting with Gn protein. This vaccination regimen induced balanced Th1/Th2 immune responses and resulted in potent humoral and T cell-mediated responses in mice. It elicited high neutralizing antibody titers in both mice and non-human primates. Transcriptome analysis revealed that rAd5 and Gn proteins induced adaptive and innate immune pathways, respectively. This study provides immunological and mechanistic insight into this heterologous regimen and paves the way for future strategies against emerging infectious diseases.

Evaluation of glycoprotein E subunit and live attenuated varicella-zoster virus vaccines formulated with a single-strand RNA-based adjuvant.

INTRODUCTION: Varicella-zoster virus (VZV), a human alphaherpesvirus 3, elicits both chickenpox and shingles and/or postherpetic neuralgia. A live attenuated vaccine (LAV) and glycoprotein E (gE) subunit vaccine were developed to prevent VZV-induced diseases. We recently reported that single-strand RNA (ssRNA) based on the intergenic region of the internal ribosome entry site of cricket paralysis virus (CrPV) is an effective adjuvant for protein-based and virus-like particle-based vaccines. Here, Chinese hamster ovary expression system and an LAV from Oka/SK strains. METHODS: We appraised the adjuvant effect of the same CrPV ssRNA encoding the gE gene formulated in the two vaccines using VZV-primed C57BL/6 mice and guinea pigs. Humoral immunity and cell-mediated immunity were assessed by enzyme-linked immunosorbent assay (ELISA) and ELISPOT in gE subunit vaccine and by ELISA and fluorescent antibody to membrane antigen in LAV. RESULTS: The gE subunit vaccine-induced gE-specific antibodies and CD4+ T-cell responses (indicated by interferon-γ [IFN-γ] and interleukin-2 secretion) in the ssRNA-based adjuvant containing the VZV gE gene. Therefore, an ssRNA adjuvant combined with gE antigen can trigger the innate immune response and induce an adaptive immune response to ultimately activate humoral and cell-mediated responses. VZV LAV could also induce VZV-specific antibodies and IFN-γ stimulated by LAV, whereas the effect of ssRNA as a vaccine adjuvant could not be confirmed. However, the ssRNA adjuvant increased VZV-specific neutralizing antibody response. CONCLUSIONS: Taken together, these results highlight that the gE subunit vaccine and LAV developed in this study can be functional VZV vaccines, and ssRNAs appear to function better as adjuvants in a subunit vaccine than in an LAV.

Cricket paralysis virus internal ribosome entry site-derived RNA promotes conventional vaccine efficacy by enhancing a balanced Th1/Th2 response.

An ideal adjuvant should increase vaccine efficacy through balanced Th1/Th2 responses and be safe to use. Recombinant protein-based vaccines are usually formulated with aluminum (alum)-based adjuvants to ensure an adequate immune response. However, use of alum triggers a Th2-biased immune induction, and hence is not optimal. Although the adjuvanticity of RNA has been reported, a systematic and overall investigation on its efficacy is lacking. We found that single strand RNA (termed RNA adjuvant) derived from cricket paralysis virus intergenic region internal ribosome entry site induced the expression of various adjuvant-function-related genes, such as type 1 and 2 interferon (IFN) and toll-like receptor (TLR), T cell activation, and leukocyte chemotaxis in human peripheral blood mononuclear cells; furthermore, its innate and IFN transcriptome profile patterns were similar to those of a live-attenuated yellow fever vaccine. This suggests that protein-based vaccines formulated using RNA adjuvant function as live-attenuated vaccines. Application of the RNA adjuvant in mouse enhanced the efficacy of Middle East respiratory syndrome spike protein, a protein-subunit vaccine and human papillomavirus L1 protein, a virus-like particle vaccine, by activating innate immune response through TLR7 and enhancing pAPC chemotaxis, leading to a balanced Th1/Th2 responses. Moreover, the combination of alum and the RNA adjuvant synergistically induced humoral and cellular immune responses and endowed long-term immunity. Therefore, RNA adjuvants have broad applicability and can be used with all conventional vaccines to improve vaccine efficacy qualitatively and quantitively.

Epimedium koreanum Ameliorates Oxidative Stress-Mediated Liver Injury by Activating Nuclear Factor Erythroid 2-Related Factor 2.

Oxidative stress induced by reactive oxygen species is the main cause of various liver diseases. This study investigated the hepatoprotective effect of Epimedium koreanum Nakai water extract (EKE) against arachidonic acid (AA)[Formula: see text][Formula: see text][Formula: see text]iron-mediated cytotoxicity in HepG2 cells and carbon tetrachloride (CCl4-)-mediated acute liver injury in mice. Pretreatment with EKE (30 and 100[Formula: see text][Formula: see text]g/mL) significantly inhibited AA[Formula: see text][Formula: see text][Formula: see text]iron-mediated cytotoxicity in HepG2 cells by preventing changes in the expression of cleaved caspase-3 and poly(ADP-ribose) polymerase. EKE attenuated hydrogen peroxide production, glutathione depletion, and mitochondrial membrane dysfunction. EKE also increased the nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf2), transactivated anti-oxidant response element harboring luciferase activity, and induced the expression of anti-oxidant genes. Furthermore, the cytoprotective effect of EKE against AA[Formula: see text][Formula: see text][Formula: see text]iron was blocked in Nrf2 knockout cells. Ultra-performance liquid chromatography analysis showed that EKE contained icariin, icaritin, and quercetin; icaritin and quercetin were both found to protect HepG2 cells from AA[Formula: see text][Formula: see text][Formula: see text]iron via Nrf2 activation. In a CCl4-induced mouse model of liver injury, pretreatment with EKE (300[Formula: see text]mg/kg) for four consecutive days ameliorated CCl4-mediated increases in serum aspartate aminotransferase activity, histological activity index, hepatic parenchyma degeneration, and inflammatory cell infiltration. EKE also decreased the number of nitrotyrosine-, 4-hydroxynonenal-, cleaved caspase-3-, and cleaved poly(ADP-ribose) polymerase-positive cells in hepatic tissues. These results suggest EKE is a promising candidate for the prevention or treatment of oxidative stress-related liver diseases via Nrf2 activation.

Tryptanthrin prevents oxidative stress-mediated apoptosis through AMP-activated protein kinase-dependent p38 mitogen-activated protein kinase activation.

Tryptanthrin (6,12-dihydro-6,12-dioxoindolo-(2,1-b)-quinazoline) has been reported to have a variety of pharmacological activities. Present study investigated the cytoprotective effects of tryptanthrin on arachidonic acid (AA) + iron-mediated oxidative stress and the molecular mechanisms responsible. In HepG2 cells, pretreatment with tryptanthrin inhibited the cytotoxic effect of AA + iron in a concentration-dependent manner. In addition, tryptanthrin prevented the changes in the levels of apoptosis-related proteins, and attenuated reactive oxygen species production, glutathione depletion, and mitochondrial membrane impairment induced by AA + iron. Mechanistic investigations showed that tryptanthrin increased the phosphorylations of AMP-activated protein kinase (AMPK) and of p38 mitogen-activated protein kinase (p38). Furthermore, inhibition of AMPK or p38 reduced the ability of tryptanthrin to prevent AA + iron-induced cell death and mitochondrial dysfunction. Transfection experiments using AMPK mutants indicated that p38 phosphorylation by tryptanthrin was dependent on AMPK activation. In a phenylhydrazine-induced acute liver injury model, tryptanthrin decreased serum levels of alanine aminotransferase, aspartate aminotransferase, and bilirubin in mice. Additionally, tryptanthrin reduced numbers of degenerating hepatocytes, infiltrating inflammatory cells, 4-hydroxynonenal-, and nitrotyrosine-positive cells in hepatic tissues. Thus, these results suggest tryptanthrin has therapeutic potential to protect cells from oxidative injury via AMPK-dependent p38 activation.

Eupatilin induces Sestrin2-dependent autophagy to prevent oxidative stress.

Eupatilin (5,7-dihydroxy-3,4,6-trimethoxyflavone) has many pharmacological activities including anti-inflammation, anti-oxidant and anti-cancer effects. Autophagy is the basic cellular machinery involving the digestion of damaged cellular components. In the present study, we investigated the protection effects of eupatilin against arachidonic acid (AA) and iron-induced oxidative stress in HepG2 cells and tried to elucidate the molecular mechanisms responsible. Eupatilin increased cell viability against AA + iron in a concentration-dependent manner and prevented mitochondrial dysfunction and reactive oxygen species (ROS) production. In addition, AA + iron increased the levels of pro-apoptotic proteins and these changes were prevented by eupatilin. Eupatilin also induced autophagy, as evidenced by the accumulation of microtubule-associated protein 1 light chain3-II and the detection of autophagic vacuoles. Furthermore, the protective effects of eupatilin on mitochondrial dysfunction and ROS production were significantly abolished by autophagy inhibitors. Eupatilin also increased the mRNA level of sestrin-2 and its promoter-driven reporter gene activity, which resulted in the up-regulation of sestrin-2 protein. Finally, gene silencing using sestrin-2 siRNA and the ectopic expression of recombinant adenoviral sestrin-2 indicated that sestrin-2 induction by eupatilin was required for autophagy-mediated cytoprotection against AA + iron. Our results suggest that eupatilin activates sestrin-2-dependent autophagy, thereby preventing oxidative stress induced by AA + iron.