A Pilot Study on Bioaccumulation and Tissue Distribution of Mercury in Barn Swallow (Hirundo rustica).

Although extensive research has been carried out on the occurrence of mercury (Hg) in biota, bioaccumulation and tissue distribution of Hg in songbirds have not been well characterized. In the present study, Hg was investigated in insects and barn swallows (Hirundo rustica) to explore the bioaccumulation characteristics of Hg. Hg in swallow feathers and tissues including muscle, liver, and bone was investigated to determine the tissue distribution of Hg. The concentrations of Hg were 1.39 ± 1.01 µg/g, 0.33 ± 0.09 µg/g, 0.47 ± 0.10 µg/g, and 0.23 ± 0.09 µg/g in feather, muscle, liver, and bone samples, respectively. The trophic magnification factor of Hg in swallows and insects was higher than 1. However, the Hg concentrations in swallow feathers were not significantly correlated with stable isotope values of carbon or nitrogen, which implies the complex food sources and exposure processes of Hg for swallows. Feathers had significantly higher concentrations of Hg than liver, muscle, and bone samples (p < 0.01 for all comparisons). Feather, muscle, bone, and other organs had fractions of 64.4 ± 11.9%, 6.07 ± 2.06%, 20.0 ± 8.19%, and 9.56 ± 2.96% in total body burden of Hg in swallows. Hg in feathers contributed more than half of Hg in the whole body for most swallow individuals. Swallows may efficiently eliminate Hg by molting, and the excretion flux of Hg and other contaminants via molting deserves more investigation.

Evaluation of the immunoprotective effects of porcine deltacoronavirus subunit vaccines.

Porcine deltacoronavirus (PDCoV), a new porcine enteric coronavirus, has seriously endangered the pig breeding industry and caused great economic losses. However, a PDCoV vaccine is not commercially available. Therefore, new and efficient PDCoV vaccines must be developed without delay. In this study, we used the ExpiCHO eukaryotic expression system to express and purify the following 3 structural proteins of PDCoV: S, N and M. Subsequently, the level of humoral and cellular immunity induced by the S protein (immunization with the S protein alone) and a protein mixture (immunization with a mixture of S, N and M proteins) were evaluated in mice and piglets, respectively, and the performances of the 2 immunizations in a challenge protection test were assessed in piglets. The results showed that both the S protein and the protein mixture induced the production of high levels of specific IgG antibodies and neutralizing antibodies and effectively neutralized PDCoV-infected LLC-PK cells in vitro. Furthermore, compared with the S protein, the N and M proteins in the protein mixture promoted the expression of CD8+ T cells and IFN-γ, induced a stronger cellular immune response, and effectively protected 4/5 of the piglets from PDCoV infection. In conclusion, the results of this study showed that the N and M proteins play important roles in inducing an immunoprotective response. Using N and M antigens as effective antigenic components in the development of PDCoV vaccines in the future will effectively increase the immune efficacy of the vaccines.

Adenovirus-vectored PDCoV vaccines induce potent humoral and cellular immune responses in mice.

Porcine deltacoronavirus (PDCoV) is a novel swine enteropathogenic coronavirus that causes severe watery diarrhea, vomiting, dehydration and high mortality in piglets, resulting in significant economic losses by the global pig industry. Recently, PDCoV has also shown the potential for cross-species transmission. However, there are currently few vaccine studies and no commercially available vaccines for PDCoV. Hence, here, two novel human adenovirus 5 (Ad5)-vectored vaccines expressing codon-optimized forms of the PDCoV spike (S) glycoprotein (Ad-PD-tPA-Sopt) and S1 glycoprotein (Ad-PD-oriSIP-S1opt) were constructed, and their effects were evaluated via intramuscular (IM) injection in BALB/c mice with different doses and times. Both vaccines elicited robust humoral and cellular immune responses; moreover, Ad-PD-tPA-Sopt-vaccinated mice after two IM injections with 108 infectious units (IFU)/mouse had significantly higher anti-PDCoV-specific neutralizing antibody titers. In contrast, the mice immunized with Ad-PD-tPA-Sopt via oral gavage (OG) did not generate robust systemic and mucosal immunity. Thus, IM Ad-PD-tPA-Sopt administration is a promising strategy against PDCoV and provides useful information for future animal vaccine development.

The GTPase activity and isoprenylation of Swine GBP1 are critical for inhibiting the production of Japanese Encephalitis Virus.

Japanese encephalitis virus (JEV) is a flavivirus that cause severe neurological deficits. The guanylate-binding protein 1 (GBP1) gene is an interferon-stimulated gene and exerts antiviral functions on many RNA and DNA viruses via diverse mechanisms, however, the roles and the action modes of GBP1 in the antiviral effect on the production of JEV RNA and infectious virions remain to be clarified. In this study, we found that the RNA levels of swine GBP1 (sGBP1) in PK15 cells were up-regulated at the late stage of JEV infection. The overexpression of sGBP1 significantly inhibited the production of JEV while the knockdown of sGBP1 promoted the production of JEV. The GTPase activity and isoprenylation of sGBP1 both are critical for anti-JEV activity. The GTPase activity of sGBP1 is responsible for inhibiting the production of JEV genomic RNA. The isoprenylation of sGBP1 inhibited the expression and cleavage of JEV prM to decrease the yields of infectious virions, which may be associated with the interaction between sGBP1 and cellular proprotein convertase furin. Taken together, the study dissected the action modes of sGBP1with potent anti-JEV activity in more details.

Nicotinamide Efficiently Suppresses Porcine Epidemic Diarrhea Virus and Porcine Deltacoronavirus Replication.

Porcine epidemic diarrhea virus (PEDV) and porcine deltacoronavirus (PDCoV), members of the genus Coronavirus, mainly cause acute diarrhea, vomiting and dehydration in piglets, and thus lead to serious economic losses. In this study, we investigated the effects of nicotinamide (NAM) on PEDV and PDCoV replication and found that NAM treatment significantly inhibited PEDV and PDCoV reproduction. Moreover, NAM plays an important role in replication processes. NAM primarily inhibited PEDV and PDCoV RNA and protein synthesis rather than other processes. Furthermore, we discovered that NAM treatment likely inhibits the replication of PEDV and PDCoV by downregulating the expression of transcription factors through activation of the ERK1/2/MAPK pathway. Overall, this study is the first to suggest that NAM might be not only an important antiviral factor for swine intestinal coronavirus, but also a potential candidate to be evaluated in the context of other human and animal coronaviruses.

Recombinant human adenovirus type 5 based vaccine candidates against GIIa- and GIIb-genotype porcine epidemic diarrhea virus induce robust humoral and cellular response in mice.

Porcine epidemic diarrhea virus (PEDV) is a porcine enteropathogenic coronavirus causing severe watery diarrhea, vomiting, dehydration, and death in piglets. However, most commercial vaccines are developed based on the GI genotype strains, and have poor immune protection against the currently dominant GII genotype strains. Therefore, four novel replication-deficient human adenovirus 5-vectored vaccines expressing codon-optimized forms of the GIIa and GIIb strain spike and S1 glycoproteins were constructed, and their immunogenicity was evaluated in mice by intramuscular (IM) injection. All the recombinant adenoviruses generated robust immune responses, and the immunogenicity of recombinant adenoviruses against the GIIa strain was stronger than that of recombinant adenoviruses against the GIIb strain. Moreover, Ad-XT-tPA-Sopt-vaccinated mice elicited optimal immune effects. In contrast, mice immunized with Ad-XT-tPA-Sopt by oral gavage did not induce strong immune responses. Overall, IM administration of Ad-XT-tPA-Sopt is a promising strategy against PEDV, and this study provides useful information for developing viral vector-based vaccines.

Predictive role of arterial lactate in acute kidney injury associated with off-pump coronary artery bypass grafting.

Objectives: This observational study aims to explore the predictive role of postoperative arterial lactate in off-pump coronary artery bypass grafting (CABG)-associated acute kidney injury (AKI). Materials and methods: A total of 500 consecutive patients who underwent off-pump CABG from August 2020 to August 2021 at the Department of Cardiovascular Surgery, Qilu Hospital of Shandong University, were included. Logistic regression analysis was used to confirm the independent risk factors of off-pump CABG-associated AKI. Receiver operating characteristic (ROC) curve was performed to evaluate the discrimination ability and Hosmer-Lemeshow goodness of fit test was performed to evaluate the calibration ability. Results: The incidence of off-pump CABG-associated AKI was 20.6%. Female gender, preoperative albumin, baseline serum creatinine, 12 h postoperative arterial lactate and duration of mechanical ventilation were independent risk factors. The area under the ROC curve (AUC) of 12 h postoperative arterial lactate for predicting off-pump CABG-associated AKI was 0.756 and the cutoff value was 1.85. The prediction model that incorporated independent risk factors showed reliable predictive ability (AUC = 0.846). Total hospital stay, intensive care unit stay, occurrence of other postoperative complications, and 28-day mortality were all significantly higher in AKI group compared to non-AKI group. Conclusion: 12 h postoperative arterial lactate was a validated predictive biomarker for off-pump CABG-associated AKI. We constructed a predictive model that facilitates the early recognition and management of off-pump CABG-associated AKI.

Differences in the pathogenicity of Chinese virulent genotype GIIa and GIIb porcine epidemic diarrhea virus strains and the humoral immune status of one- and two-month-old weaned pigs infected with these strains.

We evaluated differences in the pathology and humoral immune status in one- and two-month-old weaned pigs infected with virulent Chinese genotype GIIa and GIIb strains of porcine epidemic diarrhea virus (PEDV). All pigs infected with the GIIa strain developed severe diarrhea (100%), while the morbidity of the GIIb strain in one- and two-month-old weaned pigs was 80% (4/5) and 40% (2/5), respectively. There was no significant difference in IgA, IgG, or virus-neutralizing (VN) antibody levels associated with GIIa and GIIb in one-month-old weaned pigs (P > 0.05), but in two-month-old weaned pigs, the IgA, IgG, and VN antibody levels associated with GIIa were significantly higher than those associated with GIIb (P < 0.05).

Change in left ventricular diastolic function after pioglitazone treatment in patients with type 2 diabetes mellitus: A protocol for systematic review and meta-analysis.

BACKGROUND: Pioglitazone is currently used as an anti-diabetic agent and can reduce cardiovascular events in in patients with type 2 diabetes mellitus (T2DM). Left ventricular diastolic dysfunction has been recognized as an early manifestation of myocardial dysfunction in T2DM patients. This systematic review and meta-analysis aimed to investigate changes in the left ventricular diastolic function after the treatment of pioglitazone. METHODS: A systematic literature search of PubMed, Embase, and the Cochrane Library until May 2021 with keywords pioglitazone and left ventricular diastolic function was performed in accordance with the meta-analysis of observational studies in epidemiology guidelines and preferred reporting items for systematic reviews and meta-analyses statement. Three reviewers independently selected the studies and extracted data. Quality assessment of the included studies was undergone. A fixed effects model was used to calculate overall effect sizes. Subgroup analyses were subsequently performed. A fixed effects model was used to calculate the overall effect size. Subgroup analyses were then performed. RESULTS: Seven studies with 233 patients were investigated. We found pioglitazone significantly improved hemoglobin A1c (%) in patients with T2DM and left ventricular diastolic function had an improvement tendency (weighted mean difference [WMD], 0.03; 95% confidence interval [CI], 0.01-0.05, P < .01) despite moderate heterogeneity (I2 = 66%). Subsequent subgroup analysis indicated that left ventricular diastolic function were significantly improved (WMD, 0.20; 95% CI, 0.12-0.29, P < .001) in T2DM patients whose average age < 55 after receiving pioglitazone treatment. However, in T2DM patients with mean age ≥ 55 years, there was no significant improvement of left ventricular diastolic function (WMD, 0.02; 95% CI, 0-0.04, P = .04). CONCLUSION: Pioglitazone treatment significantly improved left ventricular diastolic function in type 2 diabetic patients with a mean age of < 55 years, but did not improve left ventricular diastolic function in patients with a mean age of ≥ 55 years.

Transcriptome Analysis of LLC-PK Cells Single or Coinfected with Porcine Epidemic Diarrhea Virus and Porcine Deltacoronavirus.

Porcine epidemic diarrhea virus (PEDV) and porcine deltacoronavirus (PDCoV) are the two most prevalent swine enteric coronaviruses worldwide. They commonly cause natural coinfections, which worsen as the disease progresses and cause increased mortality in piglets. To better understand the transcriptomic changes after PEDV and PDCoV coinfection, we compared LLC porcine kidney (LLC-PK) cells infected with PEDV and/or PDCoV and evaluated the differential expression of genes by transcriptomic analysis and real-time qPCR. The antiviral efficacy of interferon-stimulated gene 20 (ISG20) against PDCoV and PEDV infections was also assessed. Differentially expressed genes (DEGs) were detected in PEDV-, PDCoV-, and PEDV + PDCoV-infected cells at 6, 12, and 24 h post-infection (hpi), and at 24 hpi, the number of DEGs was the highest. Furthermore, changes in the expression of interferons, which are mainly related to apoptosis and activation of the host innate immune pathway, were found in the PEDV and PDCoV infection and coinfection groups. Additionally, 43 ISGs, including GBP2, IRF1, ISG20, and IFIT2, were upregulated during PEDV or PDCoV infection. Furthermore, we found that ISG20 significantly inhibited PEDV and PDCoV infection in LLC-PK cells. The transcriptomic profiles of cells coinfected with PEDV and PDCoV were reported, providing reference data for understanding the host response to PEDV and PDCoV coinfection.

Predictive role of the neutrophil: lymphocyte ratio in acute kidney injury associated with off-pump coronary artery bypass grafting.

Objectives: This study aims to investigate whether the ratios of cell types in peripheral blood could be used as reliable predictors of off-pump coronary artery bypass grafting (CABG)-associated acute kidney injury (AKI). Materials and methods: We retrospectively reviewed patients (n = 420) undergoing off-pump CABG from January 1, 2021 to January 1, 2022 in Qilu Hospital of Shandong University. We used logistic regression analysis to identify the potential predictors of off-pump CABG-associated AKI and construct a predictive model. Receiver operating characteristic (ROC) curve analysis was used to evaluate the predictive ability of predictors and prediction models. Results: The prevalence of AKI associated with off-pump CABG was 20.95%. Patients in the AKI group had significantly higher ratios of peripheral blood cells on postoperative day (POD)1 than patients in the non-AKI group (P < 0.01). The area under the ROC curve (AUC) of the neutrophil:lymphocyte ratio (NLR) on POD1 for predicting off-pump CABG-associated AKI was 0.780 and the cutoff value was 20.07. Patients with high NLR on POD1 had a poor short-term prognosis. The AUC of the predictive model constructed by logistic regression analysis was 0.882. The sensitivity was 68.2% and the specificity was 93.1%. Conclusion: The NLR on POD1 was a reliable predictive biomarker of off-pump CABG-associated AKI. And we successfully construct a prediction model, which contribute to the early recognition and management of off-pump CABG-associated AKI.

[Using mouse model to evaluate the immune effect of DNA prime-protein boost strategies targeting Japanese encephalitis virus].

In order to evaluate the immune effect of the genotype Ⅰ Japanese encephalitis virus prM-E DNA vaccine and the prM-EⅢ fusion protein subunit vaccine on mice using DNA prime-protein boost strategy, the prM-E gene was inserted into the pVAX1 eukaryotic expression vector. The recombinant expression vector prM-E-pVAX1 was constructed as a DNA vaccine for initial immunity, and the recombinant prM-EⅢ fusion protein was obtained using a prokaryotic expression system as a subunit vaccine for enhanced immunity. Thirty two female BALB/c mice aged 4-6 weeks were randomly divided into four groups, and a prM-E-pVAX1 DNA vaccine group, a DNA prime-protein boost immune group, a prM-EⅢ subunit vaccine group, and a pVAX1 vector control group were set up. The specific antibody level in serum was monitored by ELISA, the neutralizing antibody titer was detected by plaque reduction neutralization, and the cellular immune responses induced by different vaccine immune groups were analyzed by cytokine expression abundance and lymphocyte proliferation experiments. The results showed that the neutralizing antibody titers induced by mice immunized with the DNA prime-protein boost strategy were close to that of the group immunized with the single prM-EⅢ subunit vaccine, but significantly higher than that of the group immunized with the single prM-E-pVAX1 DNA vaccine. DNA prime-protein boost strategies induced effective Th1/Th2 immune responses in mouse models, in particular the Th1 cell-mediated immune responses. This study provides a new immune strategy that may facilitate the prevention of Japanese encephalitis.

[Comparson of the immunogenicity of genotypeâ Japanese encephalitis virus subunit vaccine candidate antigens].

To screen the best genotypeⅠJapanese encephalitis virus subunit vaccine candidate antigens, the prMEIII gene, the polytope gene and the prMEIII-polytope fusion gene of the GenotypeⅠJapanese encephalitis virus GS strain were cloned into prokaryotic expression vector pET-30a. The recombinant proteins were obtained after the induction and purification. The prepared recombinant proteins were immunized to mice, and the immunogenicity of the subunit vaccine candidate antigens was evaluated through monitoring the humoral immune response by ELISA, detecting the neutralizing antibody titer by plaque reduction neutralization test, and testing the cell-mediated immune response by lymphocyte proliferation assay and cytokine profiling. The recombinant proteins with the molecular weights of 35 (prMEIII), 28 (polytope antigen) and 57 kDa (prMEIII-polytope) induced strong humoral and cellular immune responses in mice. Compared with prMEIII-polytope and polytope proteins, the prMEIII protein induced a significant expression of IL-2 and IFN-γ (P<0.05) and the significant lymphoproliferation of splenocytes (P<0.05). The neutralizing antibody titer induced by the prMEIII protein was close to that induced by the commercial attenuated vaccine SA14-14-2 (P>0.05). The study suggests that the prMEIII protein can be used for the development of the Japanese encephalitis virus subunit vaccine.

Nitrogen-doped fluorescent carbon dots used for the imaging and tracing of different cancer cells.

Here, we report the synthesis of nitrogen-doped fluorescent carbon (C) dots using a one-pot hydrothermal method. Transmission electron microscopy results reveal that the particle size of the nitrogen-doped C-dots is very small, with an average diameter of 4.6 nm. After being kept in water for 10 days, the nitrogen-doped C-dots can still dissolve well in the water, showing good stability and compatibility in aqueous solution. The fluorescence spectra show that the nitrogen-doped C-dots exhibit emission-tunable color from blue to green upon excitation from 230 to 520 nm. Cell tests show that the C-dots are low in cytotoxicity and can be used for imaging, detecting and tracing between hepatoma and HeLa cells, because hepatoma and HeLa cells show different sensitivity to different fluorescent colors pumped at different excitation wavelengths.

Development of an indirect ELISA based on the recombinant Spm2 protein for detection of tropical theileriosis.

Tropical theileriosis, caused by Theileria annulata, is distributed worldwide and causes great economic losses in dairy. The reliable diagnostic method is critical for prevention and control of the disease. In this study, a sporozoite and macroschizont gene 2 (spm2) protein from T. annulata was used to develop an indirect ELISA for tropical theileriosis. Specificity test showed that there were no cross-reactions with antibodies raised against other bovine piroplasm species using ELISA or western blotting. The specificity and sensitivity were 98.4% and 98.7%, respectively, with a threshold of 35.5% of the specific mean antibody rate (AbR). Furthermore, a total of 196 field sera samples collected from Xinjiang and Gansu provinces were detected by the spm2 ELISA and IFA. The results obtained with the spm2 ELISA and IFA in this study had the moderate agreement. The average positive rates of T. annulata sera samples detected in the present study were close to the prevalence of previous reports in these endemic areas. This indicated that the Spm2 ELISA could be used as a reliable diagnostic tool for serological survey of T. annulata infection in areas where Theileria parva is not present.

Long non-coding RNA UCA1 regulates the expression of Snail2 by miR-203 to promote hepatocellular carcinoma progression.

PURPOSE: Long non-coding RNA (LncRNA) urothelial carcinoma-associated 1 (UCA1) is reported to be dysregulated in hepatocellular carcinoma (HCC) progression. However, the functions of UCA1 in HCC still need further study. The aim is to detect the role of UCA1 involving in HCC cells proliferation and invasion, and epithelial-mesenchymal transition (EMT). METHODS: The quantitative real-time PCR was used to detect the UCA1 and miR-203 expression levels in 60 cases' HCC tissues and adjacent normal tissues. Western blotting analysis was performed to detect the EMT markers E-cadherin, Vimentin and transcription factor Snail1, Snail2 expression. Luciferase reporter assay, RNA immunoprecipitation (RIP) and pull-down assays were used to evaluate whether miR-203 was a target of UCA1. RESULTS: Our results showed that UCA1 was markedly upregulated in HCC tissues and higher UCA1 expression in HCC was positively associated with tumor size, vascular invasion and American Joint Committee on Cancer (AJCC) stage (P < 0.05). Furthermore, gain-of-function and loss-of-function analysis showed that UCA1 knockdown inhibited HCC cells proliferation and invasion in vitro and xenograft tumour growth in vivo. Moreover, UCA1 overexpression promoted cell epithelial-mesenchymal transition (EMT) in HCC via effectively sponging to miR-203 and thereby activating the expression of transcription factor Snail2. CONCLUSIONS: Our results identified that UCA1/miR-203/Snail2 pathway might involve in HCC progression. Inhibition of UCA1 acted as a promising therapeutic target for HCC patients.