Intervention factors associated with environmental stressors resulting from cross-provincial transfers by coal resource-based enterprises.

The environmental stressors associated with the cross-provincial transfer of coal resource-based enterprises (CREs) have become a critical concern for the green, sustainable, and high-quality development of resource-rich areas in central and western regions. This study referred to socioeconomic statistics and carried out an interview survey, literature review, and systematic analysis to clarify the mechanism underlying environmental stressors arising from the cross-provincial transfer of CREs. The intervention factors associated with such environmental stressors were identified, and the study conducted an empirical analysis of relevant data related to the coal-resources industry in three central and western provinces in China for the period 1997-2016. Research findings: (1) The intensity ranking of the influencing factors associated with environmental stressors caused by cross-provincial transfers of CREs has certain rules. The 'level of the enterprise's investment in environmental protection' is the weakest, the 'enterprise's development mode level' is slightly stronger, the 'enterprise scale' is stronger, and 'environmental regulation' is the strongest. (2) Stricter endogenous and exogenous policy regulations for environmental governance in rich coal resource-based regions are associated with weaker negative externalities in respect of resource development and the intensity of stressors. (3) Larger CREs are associated with a better green mining capacity, environmental repair cost advantages, social constraints, self-discipline, and thus, a weaker stress effect. (4) CREs that adopt more superior modes of development that focus on the utilization of the 'three wastes' are associated with a weaker stress effect. (5) The higher the level of investment by CREs in environmental protection technology, facilities, and equipment, the weaker the stress effect. The conclusions of the study can provide a theoretical basis to assist the Chinese government to develop relevant regulations to control inter-provincial transfers by CREs and to thereby diminish environmental stressor effects.

U-Shaped Mobility Analyzer: A Compact and High-Resolution Counter-Flow Ion Mobility Spectrometer.

In recent years, there has been a rapid increase in the use of counter-flow-type ion mobility spectrometers (IMS) due to their improved resolution and functionality. In this study, we developed a new type of counter-flow ion mobility device named the U-shaped mobility analyzer (UMA) for coupling with a mass spectrometer, where the analyte ions could travel along a gas flow in the first channel of the UMA device and then against a gas flow in the second channel of the device. Hence, a mobility band-pass filter was formed by setting different electric fields in the two channels, which enables high-resolution mobility selection of analyte ions. A resolution of ca. 180 was achieved for singly charged small organic molecules, and a resolution of up to ca. 370 was achieved for multiply charged +15 myoglobin. It was thus demonstrated that this filtering function can greatly enhance the dynamic range of an IMS-MS instrument, particularly favoring targeted analysis in complex matrices. Alternatively, the analyte ions could be operated in a so-called trap-scan mode in which ions were trapped first in one of the channels and released sequentially for mobility analysis with an even higher resolution (ca. 210 for singly charged small organic molecules and ca. 590 for +15 myoglobin). Overall, this new UMA device would enable many new applications in omics studies with its high resolution and dynamic range, especially when using the filter-scan mode for scrutinizing analytes with very low concentrations under high chemical backgrounds.

EYA4 Promotes Cell Proliferation Through Downregulation of p27Kip1 in Glioma.

BACKGROUND/AIMS: Accumulating evidence suggests that Eyes Absent Homologue 4 (EYA4) plays an important role in tumorigenesis and progression of various cancers. However, the role of EYA4 in glioma development is still unclear. METHODS: The expression of EYA4 was examined in glioma tissues by immunohistochemistry. Cell viability and apoptosis were analyzed by CCK-8, BrdU assay, and flow cytometry. RESULTS: We found that EYA4 was upregulated in glioma, and its expression was positively correlated with advanced tumor stage. Moreover, higher expression of EYA4 predicted a worse overall survival in patients with glioma. Forced overexpression of EYA4 enhanced glioma cell proliferation, and EYA4 suppressed the expression of p27Kip1 directly in these cells. Furthermore, Six1 was required for EYA4 to suppress the expression of p27Kip1 in glioma. CONCLUSION: Together, we demonstrate that EYA4 promotes cell proliferation by directly suppressing the expression of p27Kip1 in glioma.

SNHG6 Promotes Tumor Growth via Repression of P21 in Colorectal Cancer.

BACKGROUND/AIMS: SNHG6 (Small Nucleolar RNA Host Gene 6) is a novel non-coding RNA (ncRNA) and its cellular function is largely unknown. METHODS: Cell Counting Kit-8 (CCK-8) cell growth assay, colony formation and flow cytometry were used to determine colorectal cancer cell proliferation, cell cycle progression and apoptosis in vitro. The xenograft tumor formation assay in nude mice was established to evaluate tumor growth in vivo. RNA immunopreciptation (RIP) analysis was performed to examine whether SNHG6 could bind to EZH2 (enhancer of zeste 2 polycomb repressive complex 2 subunit), and chromatin immunoprecipitation (ChIP) assay was conducted to examine whether SNHG6 could repress p21 transcription by recruiting EZH2 to the p21 promoter. RESULTS: Here we found that SNHG6 was upregulated and its expression levels were positively correlated with advanced tumor stage in colorectal cancer. Survival analysis suggested that higher expression of SNHG6 predicted poor prognosis in patients with colorectal cancer. Functional studies indicated that SNHG6 could promote cell proliferation via a direct suppression of p21 expression in colorectal cancer cells. Moreover, SNHG6 repressed p21 transcription through recruiting EZH2 to the p21 promoter in colorectal cancer cells. CONCLUSION: Taken together, our study demonstrates that SNHG6 promotes tumor growth via repression of p21 in colorectal cancer, which may provide a promising target for novel anticancer therapeutics.

Improved hydrolase activity in barley and reduced malting time by adding phytase as an activator during malting steeping.

OBJECTIVES: Exogenous phytase improved the activity of hydrolases to decrease the malting time. RESULTS: Treatment with phytase during barley steeping increased activity of hydrolases (α-/ß-amylase, proteinase, ß-glucanase and xylanase) in green malt. Maximal activity was observed for α-/ß-amylase, ß-glucanase and xylanase with 0.8 U phytase/g and proteinase with 1.2 U phytase/g. Phytase promoted acrospire growth of green malt and reduced malting process with 0.8 U phytase/g in 96 h, which is 24 h less than the control. No significant variation of malt quality was found between control malt and malt treated with 0.8 U/g or 1.2 U phytase/g (P > 0.05), which makes application of exogenous phytase during steeping process as a good option for reducing malting time. CONCLUSION: Adding phytase during steeping process increases the activity of hydrolases, which reduces malting time without impacting on malt quality.

Isomer differentiation through supramolecular self-assembly in microdroplets of milliseconds life-time.

Supramolecular recognition of thymine (or its analogs) with various central cations can form magic number clusters. Dual nano-ESI via theta tip emitters was used to on-line synthesize clusters. Even thermodynamically unstable clusters can be detected by MS thanks to the very short life-time (∼ms) of the generated microdroplets. By recording characteristic cluster distributions, isomers can be clearly differentiated in a novel "bottom-up" way. Theoretical calculations were performed to explain the MS results.

Reaction and detection click in high-voltage assisted laser desorption ionization mass spectrometry.

A click reaction (copper-catalyzed azide-alkyne cycloaddition) catalyzed by "naked" copper ions (without ligands) generated in situ from a copper substrate by laser ablation in a high-voltage assisted laser desorption/ionization (HALDI) ion source is demonstrated.

Preliminary research on the flavor substance and antioxidant capacity of beers produced with baking Qingke.

The addition of baked Qingke improves the flavor profile of beer. In this study, beer was brewed using Qingke baked at various temperatures. The beer produced with Qingke baked at 180 °C achieved the highest sensory score (40/50), an alcohol content of 6.92% (v/v), a total phenolic content of 446.42 mg/L, melanoidin concentration of 98.22 g/L, a color value of 10.88 EBC, and exhibited satisfactory antioxidant activity. Analysis of volatile compounds using HS-SPME-GC-MS revealed 48 compounds, of which esters accounted for 63% and alcohols accounted for 27% of the total content. The flavor profile of the beer varied across different baking temperatures. Pyrazines and aldehydes were predominantly present in samples baked at higher temperatures (T3, T4, and T5). Correlation analysis showed that the baking flavor in the beer was primarily correlated with 2, 5-dimethyl-pyrazine, trimethyl-pyrazine, phenylacetaldehyde, and ethyl 9-decenoate (R > 0.9).

Effects of tea addition on antioxidant capacity, volatiles, and sensory quality of beer.

Green tea has great potential to enhance the quality of beer. In this study, green tea was added at different stages of beer brewing, and evaluated the antioxidant capacity, volatile components, as well as sensory quality. The results showed that the addition of green tea during the start of boiling has great potential for application, and the green tea beer (GTB) had remarkable antioxidant properties (ABTS radical scavenging ability, 8.67 mmol TE/L; DPPH radical scavenging ability, 3.97 mmol TE/L; reducing power, 3.28 mmol TE/L), and an excellent sensory quality (acceptance, 6.09/9). HPLC analysis indicated that the principal phenolics in GTB were catechin and caffeic acid, in addition, the relative amounts of ferulic acid, gallic acid can be used to differentiate between GTB and beer. HS-SPME-GC-MS analyses showed that ethyl caprylate, ethyl nonanoate, ethyl caprate, linalool, and phenethyl alcohol were potentially significant for the aroma profile of GTB.

Total flavonoids of litchi Seed alleviates schistosomiasis liver fibrosis in mice by suppressing hepatic stellate cells activation and modulating the gut microbiomes.

Infection with Schistosoma japonicum (S. japonicum) is an important zoonotic parasitic disease that causes liver fibrosis in both human and domestic animals. The activation of hepatic stellate cells (HSCs) is a crucial phase in the development of liver fibrosis, and inhibiting their activation can alleviate this progression. Total flavonoids of litchi seed (TFL) is a naturally extracted drug, and modern pharmacological studies have shown its anti-fibrotic and liver-protective effects. However, the role of TFL in schistosomiasis liver fibrosis is still unclear. This study investigated the therapeutic effects of TFL on liver fibrosis in S. japonicum infected mice and explored its potential mechanisms. Animal study results showed that TFL significantly reduced the levels of Interleukin-1ß (IL-1ß), Tumor Necrosis Factor-α (TNF-α), Interleukin-4 (IL-4), and Interleukin-6 (IL-6) in the serum of S. japonicum infected mice. TFL reduced the spleen index of mice and markedly improved the pathological changes in liver tissues induced by S. japonicum infection, decreasing the expression of alpha-smooth muscle actin (α-SMA), Collagen I and Collagen III protein in liver tissues. In vitro studies indicated that TFL also inhibited the activation of HCSs induced by Transforming Growth Factor-ß1 (TGF-ß1) and reduced the levels of α-SMA. Gut microbes metagenomics study revealed that the composition, abundance, and functions of the mice gut microbiomes changed significantly after S. japonicum infection, and TLF treatment reversed these changes. Therefore, our study indicated that TFL alleviated granulomatous lesions and improved S. japonicum induced liver fibrosis in mice by inhibiting the activation of HSCs and by improving the gut microbiomes.

Enhanced Downstream Processing for a Cell-Based Avian Influenza (H5N1) Vaccine.

H5N1 highly pathogenic avian influenza virus (HPAIV) infections pose a significant threat to human health, with a mortality rate of around 50%. Limited global approval of H5N1 HPAIV vaccines, excluding China, prompted the need to address safety concerns related to MDCK cell tumorigenicity. Our objective was to improve vaccine safety by minimizing residual DNA and host cell protein (HCP). We developed a downstream processing method for the cell-based H5N1 HPAIV vaccine, employing CaptoTM Core 700, a multimodal resin, for polishing. Hydrophobic-interaction chromatography (HIC) with polypropylene glycol as a functional group facilitated the reversible binding of virus particles for capture. Following the two-step chromatographic process, virus recovery reached 68.16%. Additionally, HCP and DNA levels were reduced to 2112.60 ng/mL and 6.4 ng/mL, respectively. Western blot, high-performance liquid chromatography (HPLC), and transmission electron microscopy (TEM) confirmed the presence of the required antigen with a spherical shape and appropriate particle size. Overall, our presented two-step downstream process demonstrates potential as an efficient and cost-effective platform technology for cell-based influenza (H5N1 HPAIV) vaccines.

Engineering Phage Nanocarriers Integrated with Bio-Intelligent Plasmids for Personalized and Tunable Enzyme Delivery to Enhance Chemodynamic Therapy.

Customizable and number-tunable enzyme delivery nanocarriers will be useful in tumor therapy. Herein, a phage vehicle, T4-Lox-DNA-Fe (TLDF), which adeptly modulates enzyme numbers using phage display technology to remodel the tumor microenvironment (TME) is presented. Regarding the demand for lactic acid in tumors, each phage is engineered to display 720 lactate oxidase (Lox), contributing to the depletion of lactic acid to restructure the tumor's energy metabolism. The phage vehicle incorporated dextran iron (Fe) with Fenton reaction capabilities. H2O2 is generated through the Lox catalytic reaction, amplifying the H2O2 supply for dextran iron-based chemodynamic therapy (CDT). Drawing inspiration from the erythropoietin (EPO) biosynthetic process, an EPO enhancer is constructed to impart the EPO-Keap1 plasmid (DNA) with tumor hypoxia-activated functionality, disrupting the redox homeostasis of the TME. Lox consumes local oxygen, and positive feedback between the Lox and the plasmid promotes the expression of kelch ECH Associated Protein 1 (Keap1). Consequently, the downregulation of the antioxidant transcription factor Nrf2, in synergy with CDT, amplifies the oxidative killing effect, leading to tumor suppression of up to 78%. This study seamlessly integrates adaptable T4 phage vehicles with bio-intelligent plasmids, presenting a promising approach for tumor therapy.

Effect of early peri-operative arterial lactate concentration level ratios on post-hepatectomy liver failure.

BACKGROUND: Post-hepatectomy liver failure (PHLF) is a serious complication after hepatectomy and a major cause of death. The current criteria for PHLF diagnosis (ISGLS consensus) require laboratory data of elevated INR level and hyperbilirubinemia on or after postoperative day 5. This study aims to propose a new indicator for the early clinical prediction of PHLF. METHODS: The peri-operative arterial lactate concentration level ratios were derived from time points within the 3 days before surgery and within POD1, the patients were divided into two groups: high lactate ratio group (≥ 1) and low lactate ratio group (< 1). We compared the differences in morbidity rates between the two groups. Utilized logistic regression analysis to identify the risk factors associated with PHLF development and ROC curves to compare the predictive value of lactate ratio and other liver function indicators for PHLF. RESULTS: A total of 203 patients were enrolled in the study. Overall morbidity and severe morbidity occurred in 64.5 and 12.8 per cent of patients respectively. 39 patients (19.2%) met the criteria for PHLF, including 15 patients (7.4%) with clinically relevant Post-hepatectomy liver failure (CR-PHLF). With a significantly higher incidence of PHLF observed in the lactate ratio ≥ 1 group compared to the lactate ratio < 1 group (n = 34, 26.8% vs. n = 5, 6.6%, P < 0.001). Multivariable logistic regression analysis revealed that a lactate ratio ≥ 1 was an independent predictor for PHLF (OR: 3.239, 95% CI 1.097-9.565, P = 0.033). Additionally, lactate ratio demonstrated good predictive efficacy for PHLF (AUC = 0.792). CONCLUSIONS: Early assessment of peri-operative arterial lactate concentration level ratios may provide experience in early intervention of complications in patients with hepatocellular carcinoma, which can reduce the likelihood of PHLF occurrence and improve patient prognosis.

Optimization of the Fermentation Conditions of Huaniu Apple Cider and Quantification of Volatile Compounds Using HS-SPME-GC/MS.

The fermentation process and composition of volatile compounds play a crucial role in the production of Huaniu apple cider. This study aimed to optimize the fermentation conditions of Huaniu apple cider and quantify its volatile compounds using headspace solid-phase microextraction-gas chromatography-mass spectrometry (HS-SPME-GC/MS). The optimal fermentation parameters were determined using response surface methodology (RSM). The optimal fermentation temperature was 25.48 °C, initial soluble solids were 18.90 degrees Brix, inoculation amount was 8.23%, and initial pH was 3.93. The fermentation rate was determined to be 3.0, and the predicted value from the verification test was 3.014. This finding demonstrated the excellent predictability of a RSM-optimized fermentation test for Huaniu apple cider, indicating the reliability of the process conditions. Moreover, the analysis of volatile compounds in the optimized Huaniu cider identified 72 different ingredients, including 41 esters, 16 alcohols, 6 acids, and 9 other substances. Notably, the esters exhibited high levels of ethyl acetate, ethyl octanoate, and ethyl capricate. Similarly, the alcohols demonstrated higher levels of 3-methyl-1-butanol, phenethylethanol, and 2-methyl-1-propanol, while the acids displayed increased concentrations of acetic acid, caproic acid, and caprylic acid. This study provides the essential technical parameters required for the preparation of Huaniu apple cider while also serving as a valuable reference for investigating its distinct flavor profile.

Development of MDCK-based quadrivalent split seasonal influenza virus vaccine with high safety and immunoprotection: A preclinical study.

Vaccination remains the best prevention strategy against influenza. The MDCK-based influenza vaccine prompted the development of innovative cell culture manufacturing processes. In the present study, we report the effects of multiple administrations of a candidate, seasonal, MDCK-based, quadrivalent split influenza virus vaccine MDCK-QIV in Sprague-Dawley (SD) rats. Moreover, the effects of the vaccine were evaluated in terms of fertility and early embryonic development, embryo-fetal development, and perinatal toxicity in the SD rats and immunogenicity in Wistar rats and BALB/c mice. Regarding the safety profile, MDCK-QIV demonstrated tolerance in local stimulation with repeated dose administration and presented no significant effect on the development, growth, behavior, fertility, and reproductive performance of the adult male rats, maternal rats, and their offspring. MDCK-QIV elicited strong hemagglutination inhibition neutralizing antibody response and protection against the influenza virus in the mouse model. Thus, data supported that MDCK-QIV could be further evaluated in human clinical trial, which is currently underway.

Pulling Off Stable Economic System Adhering Carbon Emissions, Urban Development and Sustainable Development Values.

In today's world, sustainability has become a major concern. The current study attempts to look into the impact of six sustainable development values on economic growth and emissions of carbon footprints: freedom, equality, solidarity, tolerance, respect for nature, and shared mobility. The moderating importance of urban development in regulating the relationship of economic growth and carbon footprints has also been investigated in this study. The structural equation modeling (SEM) analysis technique was utilized to confirm pathways and verify path modeling in this work. Overall, 302 responses were selected for data analysis, and data were collected using a self-administered questionnaire with a random sampling technique. The study found that two of the six sustainable development values (tolerance and solidarity) have a beneficial impact on a country's economic growth. However, freedom, equality, respect for nature, and shared mobility are not indicators of a country's economic progress. Similarly, economic expansion helps to mediate the relationship between tolerance, solidarity, and carbon emissions. Although urban growth did not play a moderating role, it has a positive impact on carbon emissions. The current study suggests various implications for long-term development to improve an economy's or country's economic performance. Furthermore, complete emphasis must be paid to sustainable development values, which are more closely linked to economic growth but also help cut carbon emissions. Future research should look into the current model in other countries.

Tumor-Associated Macrophages: Key Players in Triple-Negative Breast Cancer.

Triple negative breast cancer (TNBC) refers to the subtype of breast cancer which is negative for ER, PR, and HER-2 receptors. Tumor-associated macrophages (TAMs) refer to the leukocyte infiltrating tumor, derived from circulating blood mononuclear cells and differentiating into macrophages after exuding tissues. TAMs are divided into typical activated M1 subtype and alternately activated M2 subtype, which have different expressions of receptors, cytokines and chemokines. M1 is characterized by expressing a large amount of inducible nitric oxide synthase and TNF-α, and exert anti-tumor activity by promoting pro-inflammatory and immune responses. M2 usually expresses Arginase 1 and high levels of cytokines, growth factors and proteases to support their carcinogenic function. Recent studies demonstrate that TAMs participate in the process of TNBC from occurrence to metastasis, and might serve as potential biomarkers for prognosis prediction.

[Effect of "joint treatment of lung and intestine" with moxibustion on lung function and airway inflammation in asthmatic rats].

OBJECTIVE: To investigate the role of moxibustion of "Feishu" (BL13)，"Tianshu" (ST25) for asthma by simultaneously treating lung and intestine (i.e., treating both lung and intestine at the same time) in asthmatic rats. METHODS: A total of 48 SD rats were randomly divided into normal, model, lung treatment and joint-treatment of lung and intestine (joint-treatment) groups, with 12 rats in each. The asthma model was established by subcutaneous (bilateral back and inguinal regions) and intraperitoneal injection of mixture solution of albumin and Aluminium Hydroxide gel (on day 1st, and 9th) and followed by inhalation of atomized 1% ovalbumin (on day 15th, 20 min each time, once daily for 1 week). Moxibustion was applied to bilateral BL13 for rats of the lung treatment group or bilateral BL13 and ST25 for rats of the joint-treatment group. One hour after the intervention, the rats in the later three groups were separately given nebulized 1% ovalbumin solution inhalation for 20 min. The treatments were conducted once daily for 14 consecutive days. After intervention, the lung functions including the forced expiratory flow 25% (FEF 25%), maximal mid-expiratory flow (MMEF), dynamic lung compliance (Cdyn), forced expiratory volume/ forced vital capacity (FEV/FVC), peak expiratory flow (PEF), and lung resistance (RL) were measured by using a small animal lung function detector, and pathological changes and collagen deposition in the lung tissues were observed by H.E. and Masson staining, separately. The levels of interleukin (IL)-17, IL-4, IL-13, IL-33, IL-5, leukotriene (LT) and thymic stromal lymphocyte (TSLP) in the lung tissue were measured by ELISA. RESULTS: Compared with the normal group, the FEF 25%, MMEF， Cdyn， FEV/FVC and PEF were significantly decreased (P<0.05, P<0.01)， and the pulmonary RL, collagen deposition, and contents of IL-17, IL-4, IL-13, IL-33, IL-5, TSLP and LT were notably increased (P<0.05, P<0.01) in the model group. After intervention, the MMEF and Cdyn in the lung treatment group, PEF, MMEF, Cdyn, FEV/FVC, FEF 25% in the joint-treatment group, were markedly increased (P<0.05, P<0.01), whereas the collagen deposition, IL-17, IL-4 and TSLP in both the lung treatment and joint-treatment groups, RL, IL-13, IL-33, IL-5 and LT in the joint-treatment group were considerably down-regulated (P<0.05, P<0.01). The effects of the joint treatment were apparently superior to those of lung treatment in down-regulating the contents of TSLP and LT (P<0.05, P<0.01). H.E. staining showed thickened alveolar wall, infiltration of a large number of inflammatory cells in the bronchus of the model group, which was relatively milder in the joint-treatment group. CONCLUSION: "Joint treatment of lung and intestine" with moxibustion is superior to "lung treatment" alone in ameliorating the lung function and mitigating airway inflammation in rats with asthma.

[Corrigendum] TP53 and RET may serve as biomarkers of prognostic evaluation and targeted therapy in hepatocellular carcinoma.

Subsequently to the publication of the above article, the authors have realized that a couple of clerical errors were made when writing the article, and wish to correct these errors in a corrigendum statement. First, in the Materials and methods section on p. 2216, the final sentence of the 'Immunohistochemistry and tissue microarray' subsection, the authors wish to add a further definition, so that the text reads as follows (changes highlighted in bold): 'The positive expression of RET was defined as ≥5% staining of a tumor section; high expression was defined as ≥40% staining of a tumor section, and low expression as <40%'. Secondly, in the Results section, 'Mutation frequency of each gene distributed in 4 biological categories' subsection, p. 2220, right­hand column, second paragraph, line 17, the sentence written here should have read as follows: 'The group with the positive expression of RET included 28.9% (26/90) of the patients, and 4 of these patients were defined as high expression'. The authors are grateful to the Editor of Oncology Reports for allowing them this opportunity to publish a corrigendum, and apologize to the readership of the journal for any inconvenience caused. [Oncology Reports 37: 2215­2226, 2017; DOI: 10.3892/or.2017.5494].

Preclinical immunogenicity assessment of a cell-based inactivated whole-virion H5N1 influenza vaccine.

In influenza vaccine development, Madin-Darby canine kidney (MDCK) cells provide multiple advantages, including large-scale production and egg independence. Several cell-based influenza vaccines have been approved worldwide. We cultured H5N1 virus in a serum-free MDCK cell suspension. The harvested virus was manufactured into vaccines after inactivation and purification. The vaccine effectiveness was assessed in the Wuhan Institute of Biological Products BSL2 facility. The pre- and postvaccination mouse serum titers were determined using the microneutralization and hemagglutination inhibition tests. The immunological responses induced by vaccine were investigated using immunological cell classification, cytokine expression quantification, and immunoglobulin G (IgG) subtype classification. The protective effect of the vaccine in mice was evaluated using challenge test. Antibodies against H5N1 in rats lasted up to 8 months after the first dose. Compared with those of the placebo group, the serum titer of vaccinated mice increased significantly, Th1 and Th2 cells were activated, and CD8+ T cells were activated in two dose groups. Furthermore, the challenge test showed that vaccination reduced the clinical symptoms and virus titer in the lungs of mice after challenge, indicating a superior immunological response. Notably, early after vaccination, considerably increased interferon-inducible protein-10 (IP-10) levels were found, indicating improved vaccine-induced innate immunity. However, IP-10 is an adverse event marker, which is a cause for concern. Overall, in the case of an outbreak, the whole-virion H5N1 vaccine should provide protection.