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Jianhua Qin,1,2 Tingting Zhu,1,2 Weihua Wu,1,2 Huan Chen,3 Yi He4 1Department of Nephrology, Affiliated Hospital of Southwest Medical University, Luzhou 646000, Sichuan, People’s Republic of China; 2Sichuan Clinical Research Center for Nephropathy, Luzhou 646000, Sichuan, People’s Republic of China; 3Department of Pathogen Biology, Basic Medical College, Southwest Medical University, Luzhou 646000, Sichuan, People’s Republic of China; 4Department of Urology, Affiliated Hospital of Southwest Medical University, Luzhou 646000, Sichuan, People’s Republic of ChinaCorrespondence: Yi He Email jingyoufan46@163.comBackground: Long non-coding RNA (lncRNA) has been recognized as the new regulator and biomarker for cancers. However, in clear cell renal cell carcinoma (ccRCC), the functions of lncRNAs are not well characterized. This research aimed to probe the function of lncRNA PCED1B-AS1 in the progression of ccRCC.Materials and Methods: Quantitative real-time polymerase chain reaction (qRT-PCR) was conducted to detect the expression levels of PCED1B-AS1, microRNA-484 (miR-484), and zinc finger E-box binding homeobox 1 (ZEB1) in 40 pairs of human ccRCC tissues and corresponding adjacent kidney tissue samples. Chi-square test was employed to evaluate the association between PCED1B-AS1 expression level and clinicopathological characteristics. The effects of PCED1B-AS1, miR-484 and ZEB1 on the cell proliferation, migration and epithelial-mesenchymal transition (EMT) process of ccRCC cells were studied by CCK-8 assay, EdU cell proliferation assay, wound healing test and Western blotting. The regulatory relationships among PCED1B-AS1, miR-484, ZEB1 were examined by luciferase reporter gene assay and RNA immunoprecipitation assay.Results: PCED1B-AS1 was remarkably up-regulated in ccRCC tissues and cell lines. High expression of PCED1B-AS1 was associated with poor prognosis of the patients. Loss-of-function experiments showed that PCED1B-AS1 could regulate the proliferation, migration and EMT of ccRCC cells. PCED1B-AS1 sponged miR-484 to suppress its expression, and miR-484 targeted the 3ʹ-UTR of ZEB1 to repress the expression of ZEB1. MiR-484 counteracted the functions of PCED1B-AS1 in promoting the proliferation, migration and EMT of ccRCC cells, and PCED1B-AS1 promotes the expression of ZEB1 via repressing miR-484.Conclusion: PCED1B-AS1/miR-484/ZEB1 axis is involved in regulating the progression of ccRCC.Keywords: ccRCC, PCED1B-AS1, miR-484, ZEB1

The gamma-glutamyl transpeptidase (GGT) to platelet ratio (GPR) was proposed as a novel index for predicting liver inflammation in chronic hepatitis B (CHB) patients. We aimed to investigate GPR for predicting significant liver inflammation in CHB patients with normal (≤1×upper limit of normal, ULN) or mildly elevated (≤2×ULN) alanine transaminase (ALT).431 treatment-naïve CHB patients with normal or mildly elevated ALT who underwent liver biopsy were enrolled. Comparision of GPR and other parameters for significant liver inflammation (G≥3).For patients with ALT≤2×ULN, the receiver-operating characteristic curves (AUROCs) of GPR in predicting significant liver inflammation were 0.837 (95%CI 0.796 to 0.878), 0.860 (95%CI 0.809 to 0.910) and 0.809 (95%CI 0.739 to 0.878) in the entire patients, HBeAg positive and HBeAg negative CHB patients, respectively. The diagnostic performance of GPR was higher than ALT (P0.001, P0.001, respectively), aspartate transaminase (AST) (P=0.001, P=0.003, respectively) and GGT (P=0.002, P=0.002, respectively) in the entire and HBeAg positive patients, but was comparable with AST (P=0.096) and GGT (P=0.273) in the HBeAg negative CHB patients. For patients with ALT≤1×ULN, the diagnostic accuracy of GPR was significantly higher than ALT, AST and GGT in the entire (P0.001, P=0.008 and P=0.043, respectively) and HBeAg positive CHB patients (P0.001, P=0.009 and P=0.024, respectively), while was comparable to AST (P=0.209) and GGT(P=0.555) in the HBeAg negative CHB patients.GPR has a better diagnostic value than conventional parameters to predict significant liver inflammation in CHB patients with normal or mildly elevated ALT levels, especially for HBeAg positive CHB.

Noninvasive tests (NITs) for liver fibrosis are highly needed for chronic hepatitis B (CHB) patients. We aimed to investigate whether plateletcrit (PCT) could be used as a NIT in predicting liver fibrosis for CHB patients. Five hundred and sixty-seven treatment-naive CHB patients with available liver biopsies were included. Patients were randomly divided into a derivation cohort (n = 378) and a validation cohort (n = 189). The diagnostic accuracy of PCT was evaluated using receiver operating characteristic (ROC) curves. In the derivation cohort, PCT in CHB patients with S2-S4 (0.14%), S3-S4 (0.13%) and S4 (0.12%) was lower than patients with S0-S1 (0.17%, P < .001), S0-S2 (0.17%, P < .001) and S0-S3 (0.16%, P < .001), respectively. PCT was an independent predictor of significant fibrosis (≥S2), advanced fibrosis (≥S3) and cirrhosis (S4). The area under the ROC curve (AUROC) of PCT in predicting significant fibrosis, advanced fibrosis and cirrhosis was 0.645, 0.709 and 0.714, respectively. The AUROC of PCT was higher than the aspartate transaminase to platelet ratio index (APRI) in identifying advanced fibrosis and cirrhosis, while this was comparable with APRI in identifying significant fibrosis. The diagnostic value of PCT was comparable with fibrosis-4 score (FIB-4) in predicting significant fibrosis, advanced fibrosis and cirrhosis. In the validation cohort, PCT could also identify significant fibrosis, advanced fibrosis and cirrhosis with similar diagnostic accuracy as in the derivation cohort. PCT represents a simple and inexpensive indictor for liver fibrosis in CHB patients. PCT is just as good or better than other more complex tools for staging liver fibrosis in CHB patients.

Glycemic variability has been considered as one of the predictors of diabetes complications in patients with diabetes mellitus (DM). In this work, we evaluated whether glycemic variability induces cardiac fibrosis through regulating cardiac fibroblast activation and macrophage polarization. Moreover, we determined whether glucose transporter sodium-glucose cotransporter 1 (SGLT1) plays an important role in this process. Glycemic variability-induced mice were established using DM mice (GVDM mice) and intermittent high glucose (IHG) treatment was used to simulate glycemic variability in RAW264.7 macrophages and cardiac fibroblasts. The short hairpin RNA for SGLT1 was used to knock down SGLT1. The results showed that glycemic variability aggravated the cardiac fibrosis in GVDM mice. Additionally, glycemic variability promoted the expressions of fibrogenic cytokine and the extracellular matrix proteins in left ventricular tissues and cardiac fibroblasts. GVDM mice showed a higher incidence of macrophage infiltration and M1 polarization in left ventricular tissues. Moreover, IHG promoted RAW264.7 macrophages tended to differentiate to M1 phenotype. SGLT1 knockdown alleviated cardiac fibrosis in GVDM mice and inhibited activations of cardiac fibroblasts and macrophages M1 polarization. Our results indicated that glycemic variability aggravates cardiac fibrosis through activating cardiac fibroblast and macrophage M1 polarization, which could be partially inhibited by SGLT1 knockdown.

The responses of two maize (Zea mays L.) cultivars, ‘LY336’ (shade tolerant) and ‘LC803’ (shade sensitive), to shade stress in a pot experiment conducted in the 2015 and 2016 growing seasons were investigated. The impact of 50% shade stress treatment on shoot biomass, photosynthetic parameters, chlorophyll fluorescence, and malondialdehyde (MDA) content was evaluated. The shoot biomass of the two maize hybrids was decreased significantly by shade stress treatment, for shade stress 7 d, the LC803 and LY336 were reduced by 56.7% and 44.4% compared with natural light. Chlorophyll fluorescence parameters of LY336 were not significantly affected by shade stress, whereas those of LC803 were significantly affected, the Fo increased under shade stress; however Fm, FV/FM and ΦPSII were decreased under shade stress. Among photosynthetic parameters measured, net photosynthetic rate (Pn), stomatal conductance (Gs), and transpiration rate were significantly decreased compared with natural light, LY336 and LC803 reduction by 28.0%, 22.2%, 57.7% and 35.5%, 18.9%, 62.4%; however, intercellular CO2 concentration (Ci) was significantly increased, for the two cultivars. Under shade stress for different durations (1, 3, 5, 7 d), Pn, Gs, Ci, and MDA content differed significantly between the two cultivars. Results indicated that different maize genotypes showed different responses to shading. Shade-tolerant genotypes are only weakly affected by shade stress. RESUMO: As respostas de duas cultivares de milho (Zea mays L.), ‘LY336’ (tolerante à sombra) e ‘LC803’ (sensível à sombra), ao estresse de sombra em um experimento em vaso conduzido nas safras de 2015 e 2016 foram investigadas. O impacto do tratamento de estresse de sombra de 50% na biomassa da parte aérea, parâmetros fotossintéticos, fluorescência da clorofila e teor de malondialdeído (MDA) foi avaliado. A biomassa da parte aérea dos dois híbridos de milho foi reduzida significativamente pelo tratamento de estresse de sombra, para estresse de sombra 7 d, o LC803 e LY336 foram reduzidos em 56,7% e 44,4% em comparação com a luz natural. Os parâmetros de fluorescência da clorofila de LY336 não foram significativamente afetados pelo estresse de sombra, enquanto aqueles de LC803 foram significativamente afetados, o Fo aumentou sob estresse de sombra, porém Fm, FV / FM e ΦPSII diminuíram sob estresse de sombra. Entre os parâmetros fotossintéticos medidos, a taxa fotossintética líquida (Pn), a condutância estomática (Gs) e a taxa de transpiração diminuíram significativamente em comparação com a luz natural, redução de LY336 e LC803 em 28,0%, 22,2%, 57,7% e 35,5%, 18,9%, 62,4 %, porém a concentração intercelular de CO2 (Ci) aumentou significativamente para as duas cultivares. Sob estresse de sombra para diferentes durações (1, 3, 5, 7 d), os teores de Pn, Gs, Ci e MDA diferiram significativamente entre as duas cultivares. Os resultados indicam que diferentes genótipos de milho apresentam diferentes respostas ao sombreamento. Os genótipos tolerantes à sombra são apenas fracamente afetados pelo estresse de sombra.

F-box and WD repeat domain-containing protein 7 (FBW7) has been documented to be implicated in nuclear factor κB (NF-κB) signaling and inflammation, but its role in the pathogenesis of inflammatory bowel disease (IBD) remains unknown. FBW7 was increased both in colon tissues from IBD patients and trinitrobenzene sulphonic acid (TNBS)-induced colitis mice. Immunoprecipitation assay identified that FBW7 as a novel inhibitor of κBα (IκBα)-binding partner. FBW7 upregulation promoted IκBα ubiquitin-dependent degradation, NF-κB activation, and subsequent intestinal inflammation in intestinal epithelial cells, whereas inhibition of FBW7 produced the opposite effects. Computational analysis revealed that microRNA-129 (miR-129) directly targets at 3′ UTR of FBW7. The miR-129-suppressed proteasome pathway mediated the degradation of IκBα by negatively regulating FBW7. The in vivo study demonstrated that upregulation of miR-129 ameliorated intestinal inflammation in TNBS-induced colitis mice through inhibition of the NF-κB signaling pathway. In conclusion, FBW7 is a novel E3 ubiquitin ligase for IκBα and thereby leads to NF-κB activation and inflammation. miR-129 negatively regulates FBW7 expression, resulting in secondary inhibition of the NF-κB pathway and amelioration of intestinal inflammation. Our findings provide new insight into the development of therapeutic strategies for the treatment of IBD.

Objectives Severe fever with thrombocytopenia syndrome (SFTS) is an emerging infectious disease that is caused by the SFTS virus (SFTSV) and has a high fatality rate. SFTSV-specific antibody profiles among patients with different clinical outcomes are yet to be described. The nucleocapsid protein (NP) is the most immunogenic viral antigen of the SFTSV. This study, therefore, sought to determine NP-specific antibody responses among SFTS patients with different disease progressions. Methods In the present study, 43 patients with confirmed SFTS were enrolled in our cohort, and 9 of them deceased. The clinical presentations and key laboratory parameters associated with SFTS fatality were also recorded. Serum samples from each patient were collected every 2 days during their hospitalization. NP-specific IgM and IgG responses as well as Gn or Gc-specific IgM responses were examined by enzyme-linked immunosorbent assay (ELISA), whereas, the dynamic viral loads of SFTSV RNA were quantified via real-time reverse transcription polymerase chain reaction (RT-PCR). Results First, 77% of patients generated positive NP-specific IgM antibody responses within two weeks since illness onset, defined as ‘N-specific IgM-positive patients’, while the rest of the patients were termed as ‘N-specific IgM-delayed patients’. Only 17% of the patients generated NP-specific IgG responses. The absence of NP-specific humoral responses was strongly associated with a high risk of fatality and severity of SFTS. IgM-positive patients had significantly lower levels of viral loads, less disturbed coagulopathy, and hepatic and cardiac damage compared to IgM-delayed patients. Moreover, compared to severe or fatal SFTS patients, mild SFTS patients had significantly higher magnitudes of NP-specific IgM responses, but not NP-specific IgG, Gn-specific IgM, or Gc-specific IgM responses. The abundance of NP-specific IgM responses negatively correlated with viral loads, coagulation disturbances, and hepatic injuries among SFTS patients. Conclusions Our data highlight distinct humoral profiles of NP-specific IgM responses among SFTS patients with different disease progressions and clinical outcomes.

MicroRNAs (miRNAs) belong to a class of non-coding RNAs that suppress gene expression by complementary oligonucleotide binding to the sites in target messenger RNAs. Numerous studies have demonstrated that miRNAs play crucial role in virtually all cellular processes of both plants and animals, such as cell growth, cell division, differentiation, proliferation and apoptosis. The study of rice MIR168a has demonstrated for the first time that exogenous plant MIR168a influences cholesterol transport in mice by inhibiting low-density lipoprotein receptor adapter protein 1 expression. Inspired by this finding, the cross-kingdom regulation of plant-derived miRNAs has drawn a lot of attention because of its capability to provide novel therapeutic agents in the treatment of miRNA deregulation-related diseases. Notably, unlike mRNA, some plant miRNAs are robust because of their 3′ end modification, high G, C content, and the protection by microvesicles, miRNAs protein cofactors or plant ingredients. The stability of these small molecules guarantees the reliability of plant miRNAs in clinical application. Although the function of endogenous miRNAs has been widely investigated, the cross-kingdom regulation of plant-derived miRNAs is still in its infancy. Herein, this review summarizes the current knowledge regarding the anti-virus, anti-tumor, anti-inflammatory, anti-apoptosis, immune modulation, and intestinal function regulation effects of plant-derived miRNAs in mammals. It is expected that exploring the versatile role of plant-derived miRNAs may lay the foundation for further study and application of these newly recognized, non-toxic, and inexpensive plant active ingredients.

BackgroundWe aimed to investigate the feasibility of detecting epidermal growth factor receptor (EGFR) mutations in cell-free DNA (cfDNA) from cerebrospinal fluid (CSF) and plasma of advanced lung adenocarcinoma (LADC) with brain metastases (BMs) by droplet digital polymerase chain reaction (ddPCR).MethodsThirty advanced LADC patients with BMs were enrolled, and their matched CSF and plasma samples were collected. Droplet digital PCR was used to test cfDNA in CSF and plasma for EGFR mutation status. The clinical response and prognosis were evaluated.ResultsOut of 30 patients, there were 21 females and 9 males, aged 34-75 years. In all of the cases, CSF cytology were negative. In ddPCR assays, 10 patients (33.3%) had EGFR mutation in CSF, including 3 cases of EGFR T790M mutation, and 16 patients (53.3%) had EGFR mutation in plasma, including 6 cases of EGFR T790M mutation. Five patients with activating EGFR mutations in CSF achieved an intracranial partial response (iPR) after combination treatment with the first-generation EGFR-tyrosine kinase inhibitors. Three patients with EGFR T790M mutations in CSF achieved iPR after second-line osimertinib treatment. The median overall survival and intracranial progression-free survival were 17.0 months and 11.0 months, respectively.ConclusionIt was feasible to test EGFR mutation in cerebrospinal fluid and plasma. In LADC patients with brain metastasis, cerebrospinal fluid can be used as a liquid biopsy specimen to guide treatment strategy by monitoring EGFR mutation status.

Increasing evidence suggests that dysregulated immune responses are associated with the clinical outcome of coronavirus disease 2019 (COVID-19). Nucleocapsid protein (NP)-, spike (S)-, receptor binding domain (RBD)- specific immunoglobulin (Ig) isotypes, IgG subclasses and neutralizing antibody (NAb) were analyzed in 123 serum from 63 hospitalized patients with severe, moderate, mild or asymptomatic COVID-19. Mild to modest correlations were found between disease severity and antigen specific IgG subclasses in serum, of which IgG1 and IgG3 were negatively associated with viral load in nasopharyngeal swab. Multiple cytokines were significantly related with antigen-specific Ig isotypes and IgG subclasses, and IL-1β was positively correlated with most antibodies. Furthermore, the old patients (≥ 60 years old) had higher levels of chemokines, increased NAb activities and SARS-CoV-2 specific IgG1, and IgG3 responses and compromised T cell responses compared to the young patients (≤ 18 years old), which are related with more severe cases. Higher IgG1 and IgG3 were found in COVID-19 patients with comorbidities while biological sex had no effect on IgG subclasses. Overall, we have identified diseases severity was related to higher antibodies, of which IgG subclasses had weakly negative correlation with viral load, and cytokines were significantly associated with antibody response. Further, advancing age and comorbidities had obvious effect on IgG1 and IgG3.

Sphingomyelin (SM) is one major phospholipids on lipoproteins. It is enriched on apolipoprotein B-containing particles, including very low-density lipoprotein (VLDL) and its catabolites, low-density lipoprotein (LDL). SM is synthesized by sphingomyelin synthase 1 and 2 (SMS1 and SMS2) which utilizes ceramide and phosphatidylcholine, as two substrates, to produce SM and diacylglyceride. SMS1 and SMS2 activities are co-expressed in all tested tissues, including the liver where VLDL is produced. Thus, neither Sms1 gene knockout (KO) nor Sms2 KO approach is sufficient to evaluate the effect of SMS on VLDL metabolism. We prepared liver-specific Sms1 KO/ global Sms2 KO mice to evaluate the effect of hepatocyte SM biosynthesis in lipoprotein metabolism. We found that hepatocyte total SMS depletion significantly reduces cellular sphingomyelin levels. Also, we found that the deficiency induces cellular glycosphingolipid levels which is specifically related with SMS1 but not SMS2 deficiency. To our surprise, hepatocyte total SMS deficiency has marginal effect on hepatocyte ceramide, diacylglyceride, and phosphatidylcholine levels. Importantly, total SMS deficiency decreases plasma triglyceride but not apoB levels and reduces larger VLDL concentration. The reduction of triglyceride levels also was observed when the animals were on a high fat diet. Our results show that hepatocyte total SMS blocking can reduce VLDL-triglyceride production and plasma triglyceride levels. This phenomenon could be related with a reduction of atherogenicity.

Heat stroke can induce a systemic inflammatory response, which may lead to multi‑organ dysfunction including acute kidney injury (AKI) and electrolyte disturbances. To investigate the pathogenesis of heat stroke (HS)‑related AKI, a mouse model of HS was induced by increasing the animal's core temperature to 41˚C. Blood samples obtained from the tail vein were used to measure plasma glucose and creatinine levels. Micro‑positron emission tomography‑computed tomography (micro‑PET/CT), Hamp;E staining and transmission electron microscopy were conducted to examine metabolic and morphological changes in the mouse kidneys. Immunohistochemistry (IHC) and western blot analyses were performed to investigate the expression of apoptosis‑inducing factor mitochondria‑associated 2 (Aifm2), high‑mobility group box 1 (HMGB1) and receptor for advanced glycosylation end products (RAGE). Liquid chromatography‑mass spectrometry analysis was conducted to find differential metabolites and signaling pathways. The HS mouse model was built successfully, with significantly increased creatinine levels detected in the serum of HS mice compared with controls, whereas micro‑PET/CT revealed active metabolism in the whole body of HS mice. Hamp;E and TUNEL staining revealed that the kidneys of HS mice exhibited signs of hemorrhage and apoptosis. IHC and western blotting demonstrated significant upregulation of Aifm2, HMGB1 and RAGE in response to HS. Finally, 136 differential metabolites were screened out, and enrichment of the 'biosynthesis of unsaturated fatty acids' pathway was detected. HS‑associated AKI is the renal manifestation of systemic inflammatory response syndrome, and may be triggered by the HMGB1/RAGE pathway. Metabolomics indicated increased adrenic acid, docosahexaenoic acid and eicosapentaenoic acid may serve as metabolic biomarkers for AKI in HS. The findings suggested that a correlation between the HMGB1/RAGE pathway and biosynthesis of unsaturated fatty acids may contribute to the progression of HS‑related AKI.

Introduction As an emerging infectious disease, coronavirus disease 2019 (COVID‐19) has rapidly spread throughout worldwide. Health care workers (HCWs) on frontline directly participated in the diagnosis, treatment, and care of COVID‐19 patients are at high risk of getting infected with the highly infectious severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2), the novel coronavirus that causes COVID‐19. In Nanjing Drum Tower Hospital, a total of 222 medical staff went to Wuhan city for support. In this study, we aimed to determine any nosocomial infection among our cohort of HCWs who worked in Wuhan. Methods Throat swab samples were obtained for RNA testing on day 1 and 14 of their quarantine upon their return to Nanjing. Radiological assessments were performed by chest computed tomography (CT) on day 14 of their quarantine. The blood was collected from 191 HCWs between May 12 and May 15. Anti‐SARS‐CoV‐2 immunoglobulin M (IgM) and IgG antibody responses were determined by a chemiluminescence immunoassay. Results All the throat swab specimens were found negative for SARS‐CoV‐2. The radiological analysis revealed that there was no typical chest CT scan of COVID‐19 among 222 HCWs. Consistently, anti‐SARS‐CoV‐2 IgM or IgG was also found to be negative among 191 HCWs. Conclusions There was no nosocomial infection of SARS‐CoV‐2 among our cohort of the frontline HCWs, suggesting that zero occupational infection is an achievable goal with appropriate training, strict compliance, and psychological support for the frontline HCWs., Although coronavirus disease 2019 is a highly communicable disease, that zero occupational infection of severe acute respiratory syndrome coronavirus 2 is an achievable goal with appropriate training, strict compliance, and psychological support for frontline health care workers.

SummaryBackgroundManaging discharged COVID-19 (DC) patients with recurrent positive (RP) SARS-CoV-2 RNA test results is challenging. We aimed to comprehensively characterize the viral RNA level and serum antibody responses in RP-DC patients and evaluate their viral transmission risk.MethodsA population-based observational cohort study was performed on 479 DC patients discharged from February 1 to May 5, 2020 in Shenzhen, China. We conducted RT-qPCR, antibody assays, neutralisation assays, virus isolation, whole genome sequencing (WGS), and epidemiological investigation of close contacts.FindingsOf 479 DC patients, the 93 (19%) RP individuals, including 36 with multiple RP results, were characterised by young age (median age: 34 years, 95% confidence interval [CI]: 29–38 years). The median discharge-to-RP length was 8 days (95% CI: 7–14 days; maximum: 90 days). After readmission, RP-DC patients exhibited mild (28%) or absent (72%) symptoms, with no disease progression. The viral RNA level in RP-DC patients ranged from 1·9–5·7 log10copies/mL (median: 3·2, 95% CI: 3·1–3·5). At RP detection, the IgM, IgG, IgA, total antibody, and neutralising antibody (NAb) seropositivity rates in RP-DC patients were 38% (18/48), 98% (47/48), 63% (30/48), 100% (48/48), and 91% (39/43), respectively. Regarding antibody levels, there was no significant difference between RP-DC and non-RP-DC patients. The antibody level remained constant in RP-DC patients pre- and post-RP detection. Virus isolation of nine representative specimens returned negative results. WGS of six specimens yielded only genomic fragments. No clinical symptoms were exhibited by 96 close contacts of 23 RP-DC patients; their viral RNA (96/96) and antibody (20/20) test results were negative. After full recovery, 60% of patients (n=162, 78 no longer RP RP-DC and 84 non-RP-DC) had NAb titres of ≥1:32.InterpretationRP may occur in DC patients following intermittent and non-stable excretion of low viral RNA levels. RP-DC patients pose a low risk of transmitting SARS-CoV-2. An NAb titre of ≥ 1:32 may provide a reference indicator for evaluating humoral responses in COVID-19 vaccine clinical trials.FundingSanming Project of Medicine in Shenzhen, China National Science and Technology Major Projects Foundation, Special Foundation of Science and Technology Innovation Strategy of Guangdong Province of China, and Shenzhen Committee of Scientific and Technical Innovation grants.

SARS-CoV-2 is a newly emerged coronavirus that was isolated from human infections in recent months. Since drugs and vaccines of Covid-19 are still being developed, accurate pathogen detection plays a crucial role in the current public health crisis. Quantitative real-time reverse transcriptase-polymerase chain reaction (RT-PCR) assay has been reliably used for the detection and confirmation of SARS-CoV-2 since the beginning of outbreak, whereas isothermal nucleic acid amplification based point of care automated assays has also been considered as a simpler and rapid alternative. However, since these assays have only been developed and applied for clinical use within a short timeframe, their analytical performance has not been adequately compared to-date. We describe a comparative study between a newly developed cross primer isothermal amplification (CPA) assay (Kit A) and five RT-PCR assays (Kits B to F), using clinical diagnosis as the reference standard to evaluate their sensitivity, specificity, predictive values and accuracy analysis. Clinical samples used (n=52) included throat swabs (n=30), nasal swabs (n=7), nasopharyngeal swabs (n=7) and sputum specimens (n=8), comprised of positive (n=26) and cleared cases (n=26) by clinical diagnosis. For the CPA assay (Kit A), the sensitivity, specificity, positive and negative predictive values and accuracy were 100%. Among the five RT-PCR kits, Kits B, C and F had good agreement with clinical diagnosis (Kappa≥0.75), Kits D and E were less congruent (0.4≤Kappa

The spread of influenza A/H3N2 variants possessing the hemagglutinin 121 K mutation and the unexpectedly high incidence of influenza in the 2017–2018 northern hemisphere influenza season have raised serious concerns about the next pandemic. We summarized the national surveillance data of seasonal influenza in China and identified marked differences in influenza epidemics between northern and southern China, particularly the predominating subtype and the presence of an additional summer peak in southern China. Notably, a minor spring peak of influenza caused by a different virus subtype was also observed. We also revealed that the 3C.2a lineage was dominant from the summer of 2015 to the end of the 2015–2016 peak season in China, after which the 3C.2a2 lineage predominated despite the importation and co-circulation of the 121 K variants of 3C.2a1 and 3C.2a3 lineages at the global level. Finally, an analysis based on genetic distances revealed a delay in A/H3N2 vaccine strain update. Overall, our results highlight the complicated circulation pattern of seasonal influenza in China and the necessity for a timely vaccine strain update worldwide.

In this study, we investigated the temporal association between carbapenems usage and antimicrobial resistance among major Gram-negative bacteria, using the data of quarterly carbapenems consumptions and percentages of antibiotic resistance for Gram-negative bacteria from inpatients from 2013 to 2017 in a tertiary hospital from Jiangsu Province, China. First, carbapenems consumption showed an increasing trend in the past 5 years, accompanied with the rising rates of A. baumannii and P. aeruginosa resistance against imipenem. In A. baumannii, we identified correlations between carbapenems consumption and antimicrobial resistance against piperacillin/tazobactam, ceftazidime, ciprofloxacin and imipenem, respectively. Additionally, close correlations were observed between carbapenems consumption and antimicrobial resistance against ceftazidime and ciprofloxacin in E. coli. Our data indicated that a significant positive correlation between the usage of carbapenems and the rate of antimicrobial resistance among A. baumannii and E. coli, respectively. Carbapenems should be cautiously prescribed to prevent antimicrobial resistance outbreak in A. baumannii and E. coli.

Avian influenza A (H7N9) virus has caused several epidemics and infection in both human and poultry. With mutation, the H7N9 virus gained its fifth endemic in China. Early diagnosis is crucial for the control of viral spread in poultry and prognosis of infected patients. In this study, we developed and evaluated a lateral flow dipstick recombinase polymerase amplification (LFD-RPA) assay for rapid detection of both hemagglutinin and neuraminidase gene of H7N9. Our H7-LFD-RPA and N9-LFD-RPA assay were able to detect 32 fg H7N9 nucleic acid which is more convenient and rapid than previous methods. Through detecting 50 influenza positive samples, cross-reaction was not found with other subtypes of influenza virus. The 100% analytical specificity and sufficient analytical sensitivity results agreed the real time RT-PCR assay. The results data demonstrated that our method performed well and could be applied to the detection of H7N9 virus. This LFD-RPA assay provides a candidate method for rapid point-of-care diagnosis of H7N9.

This study investigated the protective effects of myricetin (MYR) and myricetrin (MYRR) in dextran sodium sulfate (DSS) induced liver and colon damage in mice. The results showed that MYR and MYRR alleviate liver damage of colitis mice via tumor necrosis factor-α (TNF-α)/nuclear factor kappa-B (NF-κB) pathway. After dexamethasone acetate (DXMS) and MYR treatment, the expressions of myosin light chain kinase (MLCK) were increased in mice colon. The expression of occludin was increased after MYR treatment, while MYRR significantly increased tight junction protein 1 (ZO1) expression compared to colitis mice. MYR significantly reduced the abundance of intestinal Proteobacteria and Cyanobacteria to alleviate colitis. At the species level, treatment with MYR and MYRR reduced the abundance of Helicobacter-sp-MIT and Lachnospiraceae bacterium, and increased the abundance of Lactobacillus gasseri. The protective role of MYR and MYRR in DSS induced colitis mice may attribute to their intestinal microbiota and tight junction proteins modulation activities.

For all of types of applications in wireless sensor networks (WSNs), coverage is a fundamental and hot topic research issue. To monitor the interest field and obtain the valid data, the paper proposes a wireless sensor network coverage optimization model based on improved whale algorithm. The mathematic model of node coverage in wireless sensor networks is developed to achieve full coverage for the interest area. For the model, the idea of reverse learning is introduced into the original whale swarm optimization algorithm to optimize the initial distribution of the population. This method enhances the node search capability and speeds up the global search. The experiment shows that this algorithm can effectively improve the coverage of nodes in wireless sensor networks and optimize the network performance.

Recent studies have shown that blood glucose fluctuation is associated with complications of diabetes mellitus (DM). SGLT1 (sodium-dependent glucose cotransporter 1), is highly expressed in pathological conditions of heart, and is expressed in cardiomyocytes induced by high glucose. Herein, we constructed a diabetic mouse model with glucose fluctuation to investigate whether SGLT1 is involved in glucose fluctuation-induced cardiac injury. Echocardiography, histology examination, and TUNEL staining were performed to evaluate cardiac dysfunction and damage. To assess glucose fluctuation-induced oxidative stress, reactive oxygen species (ROS), malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT), and glutathione (GSH) levels were measured. To assess mitochondrial dysfunction, mitochondrial membrane potential (MMP), ATP content, mitochondrial respiratory chain complex activity, and expression of mitochondrial fusion and fission proteins were determined. The results indicated that diabetic mice with glucose fluctuation showed elevation of cardiac SGLT1 expression, left ventricular dysfunction, oxidative stress and mitochondrial dysfunction. Knockdown of SGLT1 could abrogate the effects of glucose fluctuation on cardiac injury. Thus, our study highlighted that SGLT1 plays an important role in glucose fluctuation induced cardiac injury through oxidative stress and mitochondrial dysfunction.

The interactions of triangular silver nanoprisms (TAgNPrs) with bovine serum albumin (BSA) were investigated using multiple spectroscopic techniques. A noticeable absorbance increase was noted in the peak ranges of 250 to 300 nm for BSA, and the intensity increased with the increasing concentration of TAgNPrs. Furthermore, a slight blue shift of the surface plasmon resonance band of TAgNPrs occurred, indicating that the protein absorbed on the TAgNPrs surface to form a bio-nano interface. Analysis of fluorescence quenching data using the Stern-Volmer method revealed that static quenching takes place with complex formation. Evaluation of thermodynamic parameter ΔG θ for the binding processes indicated that the binding reaction was exothermic. Furthermore, the values of binding constant K revealed that the size of nanoparticles can affect the binding degree. The order of binding affinity is 43.7 nm > 36.2 nm > 25.1 nm. The competitive experiments of site markers (flufenamic acid and phenylbutazone) suggested that the binding site of TAgNPrs on BSA was located in the region of subdomain IIIA (Sudlow site II). In addition, the conformational changes of BSA by TAgNPrs were analyzed by using synchronous fluorescence spectra, circular dichroism, and three-dimensional fluorescence spectra.

This study investigated whether the extracellular signal-regulated kinase 1/2 (ERK1/2) signal pathway affects cardiomyocyte apoptosis and the expression of tumor necrosis factor (TNF-α) at different glucose-lowering rates.Cardiomyocytes of Wistar neonate rats were maintained in a medium supplemented with 25 mmol/L glucosamine for 72 h. Then the medium was changed to different concentrations of glucosamine, and all cells were divided into five groups. The survival rate of cardiomyocyte was measured using the Cell Counting Kit-8; cardiomyocyte apoptosis was measured using the flow cytometry instrument and laser confocal microscope; TNF-α was measured using the enzyme-linked immunosorbent assay; and ERK1/2 protein and phosphorylation were measured using the Western blot. Cardiomyocyte apoptosis and TNF-α were measured again after adding U0126.As the glucose-lowering rate increased, the survival rate of cardiomyocytes increased in group B and decreased in groups C, D, and E. The TNF-α concentration increased in groups B, C, and D and decreased in group E. After 24 h, the apoptosis rate decreased in group B and increased in groups C, D, and E. The expression of p-ERK1/2 increased in groups B, D, and E, and was the lowest in group C. After adding U0126, the survival rate of cardiomyocyte in all groups increased and TNF-α concentration decreased.The influence of glucose-lowering rate on cardiomyocyte apoptosis and TNF-α was caused by the p-ERK1/2 pathway. During the glucose-lowering course, the p-ERK1/2 pathway promoted cardiomyocyte apoptosis, and TNF-α secretion was related to not only osmotic pressure but also ERK1/2 signal pathway activation.

Managing recovered COVID-19 patients with recurrent-positive SARS-CoV-2 RNA test results is challenging. We performed a population-based observational study to characterize the viral RNA level and serum antibody responses in recurrent-positive patients and evaluate their viral transmission risk. Of 479 recovered COVID-19 patients, 93 (19%) recurrent-positive patients were identified, characterized by younger age, with a median discharge-to-recurrent-positive length of 8 days. After readmission, recurrent-positive patients exhibited mild (28%) or absent (72%) symptoms, with no disease progression. The viral RNA level in recurrent-positive patients ranged from 1.8 to 5.7 log10 copies/mL (median: 3.2), which was significantly lower than the corresponding values at disease onset. There are generally no significant differences in antibody levels between recurrent-positive and non-recurrent-positive patients, or in recurrent-positive patients over time (before, during, or after recurrent-positive detection). Virus isolation of nine representative specimens returned negative results. Whole genome sequencing of six specimens yielded only genomic fragments. 96 close contacts and 1,200 candidate contacts of 23 recurrent-positive patients showed no clinical symptoms; their viral RNA (1,296/1,296) and antibody (20/20) tests were negative. After full recovery (no longer/never recurrent-positive), 60% (98/162) patients had neutralizing antibody titers of ≥1:32. Our findings suggested that an intermittent, non-stable excretion of low-level viral RNA may result in recurrent-positive occurrence, rather than re-infection. Recurrent-positive patients pose a low transmission risk, a relatively relaxed management of recovered COVID-19 patients is recommended.

Wound healing is a multistep phenomenon that relies on complex interactions between various cell types. Calpains are a well-known family of calcium-dependent cysteine proteases that regulate several processes, including cellular adhesion, proliferation, and migration, as well as inflammation and angiogenesis. CAPNS1, the common regulatory subunit of Calpain-1 and 2, is indispensable for catalytic subunit stabilization and activity. Calpain inhibition has been shown to reduce organ damage in various disease models. Here, we report that endothelial calpain-1/2 is crucially involved in skin wound healing. Using a mouse genetic model where Capns1 is deleted only in endothelial cells, we showed that calpain-1/2 disruption is associated with reduced injury-activated inflammation, reduced CD31+ blood vessel density, and delayed wound healing. Moreover, in cultured HUVECs, inhibition of calpain reduced TNF-α-induced proliferation, migration, and tube formation. Deletion of Capns1 was associated with elevated levels of IκB and downregulation of β-catenin expression in endothelial cells. These observations delineate a novel mechanistic role for calpain in the crosstalk between inflammation and angiogenesis during skin repair.

The family of flaviviruses is one of the most medically important groups of emerging arthropod-borne viruses. Host cell cytoskeletons have been reported to have close contact with flaviviruses during virus entry, intracellular transport, replication, and egress process, although many detailed mechanisms are still unclear. This article provides a brief overview of the function of the most prominent flaviviruses-induced or -hijacked cytoskeletal structures including actin, microtubules and intermediate filaments, mainly focus on infection by dengue virus, Zika virus and West Nile virus. We suggest that virus interaction with host cytoskeleton to be an interesting area of future research.

Schistosomiasis, an infectious disease caused by the Schistosoma parasitic worm, presents a serious public health issue. To date, investigation of anti-Schistosomiasis drug mechanisms through fluorescence imaging remains challenging due to the lack of appropriate dyes as fluorescent probes. Phosphorescent Ir(iii) complexes have been attracting substantial attention among various classes of fluorophores given their excellent photophysical properties. Herein, four phosphorescent Ir(iii) complexes were synthesized, two of which contained a triethylene glycol (TEG) hydrophilic group. The phosphorescent emission range of the four complexes lay between 500 and 750 nm, and their quantum yields ranged from 0.031 to 0.146. Furthermore, under the experimental concentration conditions, the TEG-modified complexes had low cytotoxicity. Cell fluorescence labeling experiments indicated that the TEG-modified complexes had good membrane permeability. Finally, the TEG-modified complexes showed remarkable labeling effects in adult Schistosoma fluorescence imaging. Thus, TEG-modified Ir(iii) complexes could be used as a new class of bilharzial fluorescent probes.

The feasibility of converting algal protein to mixed alcohols has recently been demonstrated with an engineered E. coli strain, enabling comprehensive utilization of the biomass for biofuel applications. However, the yield and titers of mixed alcohol production must be improved for market adoption. A major limiting factor for achieving the necessary yield and titer improvements is cofactor imbalance during the fermentation of algal protein. To resolve this problem, a directed evolution approach was applied to modify the cofactor specificity of two key enzymes (IlvC and YqhD) from NADPH to NADH in the mixed alcohol metabolic pathway. Using high throughput screening, more than 20 YqhD mutants were identified to show activity on NADH as a cofactor. Of these 20 mutants, the four highest activity YqhD mutants were selected for combination with two IlvC mutants, both accepting NADH as a redox cofactor, for modification of the protein conversion strain. The combination of the IlvC and YqhD mutants yielded a refined E. coli strain, subtype AY3, with increased fusel alcohol yield of ~ 60% compared to wild type under anaerobic fermentation on amino acid mixtures. When applied to real algal protein hydrolysates, the strain AY3 produced 100% and 38% more total mixed alcohols than the wild type strain on two different algal hydrolysates, respectively. The results indicate that cofactor engineering is a promising approach to improve the feasibility of bioconversion of algal protein into mixed alcohols as advanced biofuels.

The untranslated regions within viral segments are the essential promoter elements required for the initiation of viral replication and transcription. The end of the UTR sequence and part of the ORF sequence constitute the packaging signal for progeny viruses. To explore the influence of single-point and multi-site joint mutations in the UTR of the NA gene on the viral expression, we select clones with upregulated expression of the reporter gene and analyze their sequence characteristics. Bioinformatics methods were used to analyze polymorphisms in the untranslated region (UTR) of the neuraminidase gene of the H9N2 influenza A virus. Using the RNA polymerase I reporting system with enhanced green fluorescence protein (EGFP) gene as the reporter gene, libraries containing random mutations at sites within the N2 UTR were constructed using random mutagenesis. The mutants were selected from the randomized mutagenesis libraries for the N2-UTR. The N2-UTR-RNA polymerase I fluorescence reporter system was identified by sequencing and transfected into infected MDCK cells. The expression of the reporter EGFP was observed using fluorescence microscopy, and the relative fluorescence intensity was measured using a multifunctional microplate reader to analyze the expression of the reporter gene (EGFP) qualitatively and quantitatively. Herein, an RNA polymerase reporter system was constructed to rescue the mutated viruses and measure their tissue culture infective dose (TCID50). The results showed that the U13 → C13 mutation in the 3′end of the NA gene promoted the expression of viral RNA and protein, and mutation of other sites within the UTR could differentially regulate viral genomic transcription and translation. These data showed that the U13 → C13 mutation within the variable region of the 3′UTR of the NA gene in the H9N2 influenza virus promotes viral genomic expression and infection.

It is found that catechol inhibits the oxidase-mimicking activity of chitosan-protected platinum nanoparticles (Chit-PtNPs) by competing with the substrate for the active site of the Ch-PtNPs. The inhibition mechanism of catechol is different from that of ascorbic acid in that it neither reacts with O2•- nor reduces the oxidized 3,3′,5,5′-tetramethylbenzidine (TMB). Tyrosinase (TYRase) catalyzes the oxidation of catechol, thus restoring the activity of oxidase-mimicking Chit-PtNPs. By combining the Chit-PtNP, catechol, and TYRase interactions with the oxidation of TMB to form a yellow diamine (maximal absorbance at 450 nm), a colorimetric analytical method was developed for TYRase determination and inhibitor screening. The assay works in the 0.5 to 2.5 U·mL−1 TYRase activity range, and the limit of detection is 0.5 U·mL−1. In our perception, this new assay represents a powerful approach for determination of TYRase activity in biological samples.

Background Two health concerns primarily related to triatomine bugs are transmission of Trypanosoma cruzi through infective feces, and allergic reactions induced by triatomine bites. In the Southwestern United States, reduviid bugs bites commonly cause insect allergy. In South China, four cases of anaphylactic shock have been reported after this bite exposure. To further classify the species of these bugs and confirm the sensitization of the triatomine saliva, we caught triatomine bugs from the region where the bites occurred and performed phylogenetic and immunohistochemical (IHC) analysis. Methods Triatomine bugs were collected in Donghai Island of Zhanjiang City in South China. The genomic DNA was extracted from three legs of the bugs. The fragments of mitochondrial 16S rRNA, cytochrome c oxidase subunit I (COI) gene and nuclear ribosomal 18S and 28S rRNA genes were obtained by PCR and sequenced. A phylogenetic tree was constructed based on the sequence of 16S rRNA gene using a maximum likelihood method with MEGA 7.0 software. Trypanosomal specific fragments and vertebrate COI genes were amplified from the fecal DNA to detect the infection of trypanosomes and analyze the blood feeding patterns, respectively. Paraffin-embedded sections were then prepared from adult triatomines and sent for IHC staining. Results We collected two adult triatomine bugs in Donghai Island. Morphological and molecular analyses indicated that the triatomines were Triatoma rubrofasciata. No fragments of T. cruzi or other trypanosomes were detected from the fecal DNA. Mitochondrial gene segments of Homo sapiens and Mus musculus were successfully amplified. The allergens which induced specific IgE antibodies in human serum were localized in the triatomine saliva by IHC assay. Conclusions The two triatomine bugs from Donghai Island were T. rubrofasciata. They had bitten humans and mice. Their saliva should contain the allergens related to the allergic symptoms and even anaphylactic shock of exposed residents. Great consideration should be given to this triatomine bugs due to their considerable distribution and potential threat to public health in South China.

The interaction between graphene oxide-sliver nanocomposites (GO-AgNCPs) and bovine serum albumin (BSA) in aqueous buffer solution was investigated by using several spectroscopic and imaging techniques. The visible absorbance intensity of GO-AgNCPs increased with increasing concentrations of BSA, and a slight redshift of the surface plasmon resonance band (SPR) occurred due to the absorption of BSA on the surface of GO-AgNCPs. Fluorescence data revealed a static quenching process of BSA caused by GO-AgNCPs. Thermodynamic parameters of the absorption process, including adsorption equilibrium constants, changes in Gibbs free energy (ΔG), enthalpy (ΔH) and entropy (ΔS), were evaluated at different temperatures. Negative values of ΔG showed that this process was spontaneous and the BSA-GO-AgNCPs complex might form in aqueous solution. Negative values of ΔH and ΔS suggested that the binding was mainly an enthalpy-driven process, and van der Waals forces and hydrogen bonding were the major force in the formation of the nanoparticle-protein corona. Analysis of synchronous, three dimensional (3D) fluorescence and circular dichroism (CD) spectra demonstrated that the conformation of BSA was slightly altered in the presence of GO-AgNCPs. The protein corona formed on the surface of GO-AgNCPs was directly observed by scanning probe microscopy (SPM).

Vascular calcification is a main cause of increased cardiovascular morbidity and mortality in chronic kidney disease (CKD) patients. This study aimed to investigate the role of the bone morphogenetic protein (BMP) signaling pathway in the early development of vascular calcification in CKD. A CKD vascular calcification rat model was established by providing rats with a 1.8% high-phosphorus diet and an intragastric administration of 2.5% adenine suspension. The kidney and aortic pathologies were analyzed. Blood biochemical indicators, serum BMP-2 and BMP-4 levels, and aortic calcium content were determined. The expression levels of BMP-2, BMP-4, bone morphogenetic protein receptor-IA (BMPR-IA), and matrix Gla protein (MGP) in aorta were examined by quantitative real-time polymerase chain reaction and immunohistochemistry. Compared with the normal control (Nor) rats, the CKD rats exhibited a significantly decreased body weight and an increased kidney weight as well as abnormal renal function and calcium-phosphorus metabolism. Aortic von Kossa and Alizarin red staining showed massive granular deposition and formation of calcified nodules in aorta at 8 weeks. The aortic calcium content was significantly increased, which was positively correlated with the serum BMP-2 (r=0.929; P<0.01) and serum BMP-4 (r=0.702; P<0.01) levels in CKD rats. The rat aortic BMP-2 mRNA level in the CKD rats was persistently increased, and the BMP-4 mRNA level was prominently increased at the 4th week, declining thereafter. Strong staining of BMP-2, BMP-4, BMPR-IA, and MGP proteins was observed in the tunica media of the aorta from the 4th week after model induction. In conclusion, activation of the BMP signaling pathway is involved in the early development of vascular calcification in CKD. Therefore, elevated serum BMP-2 and BMP-4 levels may serve as serum markers for CKD vascular calcification.

Background First generation bioethanol production utilizes the starch fraction of maize, which accounts for approximately 60% of the ash-free dry weight of the grain. Scale-up of this technology for fuels applications has resulted in a massive supply of distillers’ grains with solubles (DGS) coproduct, which is rich in cellulosic polysaccharides and protein. It was surmised that DGS would be rapidly adopted for animal feed applications, however, this has not been observed based on inconsistency of the product stream and other logistics-related risks, especially toxigenic contaminants. Therefore, efficient valorization of DGS for production of petroleum displacing products will significantly improve the techno-economic feasibility and net energy return of the established starch bioethanol process. In this study, we demonstrate ‘one-pot’ bioconversion of the protein and carbohydrate fractions of a DGS hydrolysate into C4 and C5 fusel alcohols through development of a microbial consortium incorporating two engineered Escherichia coli biocatalyst strains. Results The carbohydrate conversion strain E. coli BLF2 was constructed from the wild type E. coli strain B and showed improved capability to produce fusel alcohols from hexose and pentose sugars. Up to 12 g/L fusel alcohols was produced from glucose or xylose synthetic medium by E. coli BLF2. The second strain, E. coli AY3, was dedicated for utilization of proteins in the hydrolysates to produce mixed C4 and C5 alcohols. To maximize conversion yield by the co-culture, the inoculation ratio between the two strains was optimized. The co-culture with an inoculation ratio of 1:1.5 of E. coli BLF2 and AY3 achieved the highest total fusel alcohol titer of up to 10.3 g/L from DGS hydrolysates. The engineered E. coli co-culture system was shown to be similarly applicable for biofuel production from other biomass sources, including algae hydrolysates. Furthermore, the co-culture population dynamics revealed by quantitative PCR analysis indicated that despite the growth rate difference between the two strains, co-culturing didn’t compromise the growth of each strain. The q-PCR analysis also demonstrated that fermentation with an appropriate initial inoculation ratio of the two strains was important to achieve a balanced co-culture population which resulted in higher total fuel titer. Conclusions The efficient conversion of DGS hydrolysates into fusel alcohols will significantly improve the feasibility of the first generation bioethanol process. The integrated carbohydrate and protein conversion platform developed here is applicable for the bioconversion of a variety of biomass feedstocks rich in sugars and proteins. Electronic supplementary material The online version of this article (10.1186/s12934-017-0804-8) contains supplementary material, which is available to authorized users.

In pulmonary hypertension (PH), pulmonary arterial smooth muscle cells (PASMCs) are dedifferentiated, undergoing a contractile-to-synthetic phenotypic switching. Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) plays diverse roles in the cardiovascular system, but its contribution to PH remains to be fully defined. The present study was undertaken to explore the role of LOX-1 in PASMCs dedifferentiation in hypoxia-induced pulmonary vascular remodeling and PH. In a rat model of hypoxic PH, pulmonary vascular remodeling was accompanied by increased expression of LOX-1 in pulmonary arteries. In primary rat PASMCs, hypoxia-induced PASMCs dedifferentiation occurred concomitantly with LOX-1 upregulation. Inhibition of LOX-1 by either siRNA knockdown or neutralizing antibody significantly ameliorated PASMCs dedifferentiation. Mechanistically, LOX-1 promotes PASMCs dedifferentiation under hypoxic conditions via ERK1/2-Elk-1/MRTF-A/SRF signaling pathway. In conclusion, our data uncovers an important role of LOX-1 in the maintenance of PASMCs phenotype. Therapeutic targeting of LOX-1/ERK1/2-Elk-1/MRTF-A/SRF signaling axis would be exploited to treat hypoxic PH.

International audience; Recently, several endophytic fungi have been demonstrated to produce volatile organic compounds (VOCs) with properties similar to fossil fuels, called "mycodiesel," while growing on lignocellulosic plant and agricultural residues. The fact that endophytes are plant symbionts suggests that some may be able to produce lignocellulolytic enzymes, making them capable of both deconstructing lignocellulose and converting it into mycodiesel, two properties that indicate that these strains may be useful consolidated bioprocessing (CBP) hosts for the biofuel production. In this study, four endophytes Hypoxylon sp. CI4A, Hypoxylon sp. EC38, Hypoxylon sp. CO27, and Daldinia eschscholzii EC12 were selected and evaluated for their CBP potential. Analysis of their genomes indicates that these endophytes have a rich reservoir of biomass-deconstructing carbohydrate-active enzymes (CAZys), which includes enzymes active on both polysaccharides and lignin, as well as terpene synthases (TPSs), enzymes that may produce fuel-like molecules, suggesting that they do indeed have CBP potential. GC-MS analyses of their VOCs when grown on four representative lignocellulosic feedstocks revealed that these endophytes produce a wide spectrum of hydrocarbons, the majority of which are monoterpenes and sesquiterpenes, including some known biofuel candidates. Analysis of their cellulase activity when grown under the same conditions revealed that these endophytes actively produce endoglucanases, exoglucanases, and β-glucosidases. The richness of CAZymes as well as terpene synthases identified in these four endophytic fungi suggests that they are great candidates to pursue for development into platform CBP organisms.

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging epidemic infectious disease caused by the SFTS bunyavirus (SFTSV) with an estimated high case-fatality rate of 12.7% to 32.6%. Currently, the disease has been reported in mainland China, Japan, Korea, and the United States. At present, there is no specific antiviral therapy for SFTSV infection. Considering the higher mortality rate and rapid clinical progress of SFTS, supporting the appropriate treatment in time to SFTS patients is critical. Therefore, it is very important for clinicians to predict these SFTS cases who are more likely to have a poor prognosis or even more likely to decease. In the present study, we established a simple and feasible model for assessing the severity and predicting the prognosis of SFTS patients with high sensitivity and specificity. This model may aid the physicians to immediately initiate prompt treatment to block the rapid development of the illness and reduce the fatality of SFTS patients., Author summary Severe fever with thrombocytopenia syndrome (SFTS) is an emerging infectious disease caused by a novel SFTS bunyavirus (SFTSV) with an estimated high case-fatality rate. However, there is no specific antiviral therapy for SFTSV infection. Symptomatic treatment and supportive therapy are the most essential part of case management. It is very important for clinicians to identify critical patients at admission. In this study, we established a simple and feasible scoring system for assessing the severity and predicting the prognosis of SFTS patients with objective parameters. This model may help the physicians to perform intervention measures in advance, control the disease progression and improve the prognosis.

In order to investigate the lesions and proteins with differential expression in cells infected with the 2009 A (H1N1) virus and to determine the specific proteins involved in cell damage, the present study has been performed. BEAS-2B cells were infected with the 2009 A (H1N1) influenza virus or the seasonal H1N1 influenza virus for 12, 24, 48, and 72 h, and cell cycle and apoptosis were analyzed with flow cytometry. Total cellular proteins were extracted and underwent two-dimensional gel electrophoresis. The differentially expressed proteins underwent mass spectrometry for identification. The results showed that after 12 h, cells infected with the virus strain sourced from severe cases had the highest apoptosis rate (P < 0.05). After 48 h, cells infected with the virus strain sourced from fatal cases and severe cases had the highest apoptosis rate (P < 0.05), and after 72 h, cells infected with virus strains from fatal cases and ordinary cases had the highest apoptosis rate (P < 0.05). All the four influenza virus strains induced cell cycle arrest mainly at the G0/G1 phase. Eighteen differentially expressed proteins were identified, including galectin-1, cofilin-1, protein DJ-1, proteasome subunit α type-5, macrophage migration inhibitory factor, translationally controlled tumor protein, profilin 1, and interferon α-2. Galectin-1 was specifically observed in BEAS-2B infected with 2009 A (H1N1) influenza viruses, and cofilin-1 was specifically observed in BEAS-2B cells in the late stage of 2009 A (H1N1) influenza virus infection. In conclusion, differential effects of the 2009 A (H1N1) influenza virus and seasonal H1N1 influenza virus were identified on the cell cycle and apoptosis, and galectin-1 may play a role in cell apoptosis induced by 2009 A (H1N1) influenza virus.

This study investigated the cellular location and the contribution of individual β-glucosidase (BGL) to total BGL activity in Neurospora crassa. Among the seven bgl genes, bgl3, bgl5, and bgl7 were transcribed at basal levels, whereas bgl1, bgl2, bgl4, and bgl6 were significantly up-regulated when the wild-type strain was induced with cellulose (Avicel). BGL1 and BGL4 were found to be contributors to intracellular BGL activity, whereas the activities of BGL2 and BGL6 were mainly extracellular. Sextuple bgl deletion strains expressing one of the three basally transcribed bgls did not produce any detectable BGL activity when they were grown on Avicel. BGL6 is the major contributor to overall BGL activity, and most of its activity resides cell-bound. The sextuple bgl deletion strain containing only bgl6 utilized cellobiose at a rate similar to that of the wild type, while the strain with only bgl6 deleted utilized cellobiose much slower than that of the wild type.

Cellobiose dehydrogenase production by Neurospora crassa was investigated in this study. N. crassa has two putative cellobiose dehydrogenase (CDH) genes (cdh) in its genome. CDH was produced only under cellulolytic conditions. Deletion of nc-cdh1 eliminated almost all of the strain’s CDH activity, whereas the deletion of nc-cdh2 had little effect on total extracellular CDH activity, which indicates that NC-CDH1 is a major contributor to overall CDH activity. The homologous expression of nc-cdh1 and nc-cdh2 under the control of the constitutive D-glyceraldehyde-3-phosphate dehydrogenase (gpdA) promoter enabled recombinant CDH production under non-cellulolytic conditions. Both NC-CDH1 and NC-CDH2 produced by N. crassa were successfully purified and characterized for the first time. NC-CDH1 and NC-CDH2 have molecular weights of 100 kDa and 130 kDa, respectively. When their N-linked glycans were removed by N-glycosidase F treatment, both enzymes showed a molecular weight of 95 kDa. Although NC-CDH2 lacks the cellulose-binding module and contributed marginally to total CDH activity in N. crassa, NC-CDH2 has specific activity similar to that of NC-CDH1 (7.93 vs. 8.89 IU mg−1), and it has a much lower Km value than that of NC-CDH1 (5.79 vs. 25.72 μM). The lower activity contribution of NC-CDH2 in the wild-type strain may results from its lower enzyme production.

Altered metabolism is one of the hallmarks of cancer cells. The best-known metabolic abnormality in cancer cells is the Warburg effect, which demonstrates an increased glycolysis even in the presence of oxygen. However, tumor-related metabolic abnormalities are not limited to altered balance between glucose fermentation and oxidative phosphorylation. Key tumor genes such as p53 and c-myc are found to be master regulators of metabolism. Metabolic enzymes such as succinate dehydrogenase, fumarate hydratase, pyruvate kinase, and isocitrate dehydrogenase mutations or expressing level alterations are all linked to tumorigenesis. In this review, we introduce some of the cancer-associated metabolic disorders and current understanding of their molecular tumorigenic mechanisms.

Sphingobium sp. SYK-6 is a soil bacterium boasting a well-studied ligninolytic pathway and the potential for development into a microbial chassis for lignin valorization. An improved understanding of its metabolism will help researchers in the engineering of SYK-6 for the production of value-added chemicals through lignin valorization. We used 13C-fingerprinting, 13C metabolic flux analysis (13C-MFA), and RNA-sequencing differential expression analysis to uncover the following metabolic traits: (i) SYK-6 prefers alkaline conditions, making it an efficient host for the consolidated bioprocessing of lignin, and it also lacks the ability to metabolize sugars or organic acids; (ii) the CO2 release (i.e., carbon loss) from the ligninolysis-based metabolism of SYK-6 is significantly greater than the CO2 release from the sugar-based metabolism of Escherichia coli; (iii) the vanillin catabolic pathway (which is the converging point of majority of the lignin catabolic pathways) is coupled with the tetrahydrofolate-dependent C1 pathway that is essential for the biosynthesis of serine, histidine, and methionine; (iv) catabolic end products of lignin (pyruvate and oxaloacetate) must enter the tricarboxylic acid (TCA) cycle first and then use phosphoenolpyruvate carboxykinase to initiate gluconeogenesis; and (v) 13C-MFA together with RNA-sequencing differential expression analysis establishes the vanillin catabolic pathway as the major contributor of NAD(P)H synthesis. Therefore, the vanillin catabolic pathway is essential for SYK-6 to obtain sufficient reducing equivalents for its healthy growth; cosubstrate experiments support this finding. This unique energy feature of SYK-6 is particularly interesting because most heterotrophs rely on the transhydrogenase, the TCA cycle, and the oxidative pentose phosphate pathway to obtain NADPH.

The aims of this study was to elucidate whether the expression of E-cadherin can be affected by the recombinant has-mir-373 eukaryotic expression plasmid vector through tests in vitro, and to analyze the relationship between the expression of E-cadherin and tumor growth. According to the has-mir-373 sequence in miRBase database, two template DNA sequences were designed. The has-mir-373 sequence and a control sequence were synthesized and cloned into pGenesil-1 eukaryotic expression plasmid vector. The recombinant plasmids were transfected into human lung cancer A549 cells by liposome-mediated method. The mir-373 expression in A549 cells was detected by using real-time quantitative polymerase chain reaction (real-time PCR). MTT (methyl thiazolyl tetrazolium) was used to analyze the growth of cancer cell cycle. RT-PCR and Western blotting were used to evaluate the levels of E-cadherin mRNA and protein expression, respectively. The expression of E-cadherin in cells was determined by immunocytochemistry. The mobility capability of transfected cells were evaluated by using wound healing assay in vitro. The fluorescent light was observed under fluorescent microscope. RT-PCR indicated that the mRNA of E-cadherin increased, and the Western blotting results also displayed that mir-373 promoted the expression of the E-cadherin protein. Compared with the control groups, MTT method and wound healing assay demonstrated that both the growth rate and migration of A549 cells transfected with the recombinant has-mir-373 eukaryotic expression plasmid was also decreased significantly (p0.001). The differences between the other two control groups were not significant (p0.05). The immunocytochemistry demonstrated a significant increase of E-cadherin protein levels in the cells transfected with mir-373, but not in the cells of the control group. Mir-373 could increase the expression levels of the E-cadherin and decrease the migration ability of human lung cancer A549 cells in vitro.

Monoamine oxidase (MAO)-A is a key enzyme for the degradation of brain serotonin (5-hydroxytryptamine, 5-HT) and norepinephrine (NE). In humans and mice, total MAO-A deficiency results in high 5-HT and NE levels, as well as elevated reactive aggression. Here we report the generation of MAO-A(Neo) mice, a novel line of hypomorphic MAO-A mutants featuring the insertion of a floxed neomycin-resistance cassette in intron-12 of the Maoa gene. This construct resulted in a chimeric, non-functional variant of the Maoa-Neo transcript, with a truncated C-terminus, likely due to aberrant splicing; these deficits notwithstanding, small amounts of functional Maoa transcript were found in the brain of MAO-A(Neo) mice. In the prefrontal cortex and amygdala, MAO-A(Neo) mice showed low, yet detectable, MAO-A catalytic activity, as well as 5-HT levels equivalent to WT littermates; conversely, the hippocampus and midbrain of MAO-A(Neo) mice featured a neurochemical profile akin to MAO-A-knockout (KO) mice, with undetectable MAO-A activity and high 5-HT concentrations. MAO-A(Neo) mice showed significant increases in dendritic length in the pyramidal neurons of orbitofrontal cortex, but not basolateral amygdala, in comparison with WT littermates; by contrast, the orbitofrontal cortex of MAO-A KO mice showed significant reductions in basilar dendritic length, as well as a profound increase in apical dendritic length. MAO-A(Neo) mice showed a unique set of behavioral abnormalities, encompassing reduced open-field locomotion, perseverative responses, such as marble burying and water mist-induced grooming, and a lack of anxiety-like behaviors in the elevated plus-maze and light-dark box paradigms. Notably, whereas MAO-A(Neo) and KO mice showed significant reductions in social interaction, only the latter genotype showed increases in resident-intruder aggression. Taken together, our findings indicate that MAO A hypomorphism results in behavioral and morphological alterations distinct from those featured by MAO-A KO mice.

The Arabidopsis (Arabidopsis thaliana) genome encodes nine Salt Overly Sensitive3 (SOS3)-like calcium-binding proteins (SCaBPs; also named calcineurin B-like protein [CBL]) and 24 SOS2-like protein kinases (PKSs; also named as CBL-interacting protein kinases [CIPKs]). A general regulatory mechanism between these two families is that SCaBP calcium sensors activate PKS kinases by interacting with their FISL motif. In this study, we demonstrated that phosphorylation of SCaBPs by their functional interacting PKSs is another common regulatory mechanism. The phosphorylation site serine-216 at the C terminus of SCaBP1 by PKS24 was identified by liquid chromatography-quadrupole mass spectrometry analysis. This serine residue is conserved within the PFPF motif at the C terminus of SCaBP proteins. Phosphorylation of this site of SCaBP8 by SOS2 has been determined previously. We further showed that CIPK23/PKS17 phosphorylated CBL1/SCaBP5 and CBL9/SCaBP7 and PKS5 phosphorylated SCaBP1 at the same site in vitro and in vivo. Furthermore, the phosphorylation stabilized the interaction between SCaBP and PKS proteins. This tight interaction neutralized the inhibitory effect of PKS5 on plasma membrane H+-ATPase activity. These data indicate that SCaBP phosphorylation by their interacting PKS kinases is a critical component of the SCaBP-PKS regulatory pathway in Arabidopsis.

The analysis of previous reported results envisioned that residues Asp195, Tyr196 and Gln197 of formate dehydrogenase from Candida bodinii (CboFDH) might play critical roles in determining the enzyme's cofactor specificity. With the aim to develop novel NADP(+)-dependent formate dehydrogenase enzymes, simultaneous site-saturation mutagenesis of residues Asp195 and Tyr196 of CboFDH coupled with screening resulted in two mutant enzymes, D195Q/Y196R and D195S/Y196P, which showed significant NADP(+) specificity. The overall catalytic efficiencies (k(cat)/K-m) toward NADP(+) were 1.14 x 10(4) and 2.9 x 10(3) M-1 s(-1), respectively, which are higher than the reported mutant CboFDHs obtained by sequential mtltagenesis. The ratio of catalytic efficiencies (k(cat)/K-m)(NADP+)/(k(cat)/K-m)(NAD+) of D195Q/Y196R and D195S/Y196P were 2.1 and 0.2, respectively. Mutation of residue Gln197 of D195Q/Y196R to Asn further increased the enzyme's overall catalytic efficiencies (k(cat)/K-m) toward NADP(+) to 29.1 x 10(3) M-1 s(-1), with (k(cat)/K-m)(NADP+)/(k(cat)/K-m)(NAD+) being 17.1, which are much higher than the reported data for a mutant enzyme of formate dehydrogenase from Pseudomanas sp. 101 (PSeFDH). This study demonstrates that residues 195. 196 and 197 really play critical roles in determining the enzyme's cofactor specificity. (C) 2009 Elsevier B.V. All rights reserved.

© 2016 Wu et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in anymedium, provided the original author and source are credited. Endophytic fungi are ubiquitous plant endosymbionts that establish complex and poorly understood relationships with their host organisms. Many endophytic fungi are known to produce a wide spectrum of volatile organic compounds (VOCs) with potential energy applications, which have been described as "mycodiesel". Many of these mycodiesel hydrocarbons are terpenes, a chemically diverse class of compounds produced by many plants, fungi, and bacteria. Due to their high energy densities, terpenes, such as pinene and bisabolene, are actively being investigated as potential "drop-in" biofuels for replacing diesel and aviation fuel. In this study, we rapidly discovered and characterized 26 terpene synthases (TPSs) derived from four endophytic fungi known to produce mycodiesel hydrocarbons. The TPS genes were expressed in an E. coli strain harboring a heterologous mevalonate pathway designed to enhance terpene production, and their product profiles were determined using Solid Phase Micro-Extraction (SPME) and GC-MS. Out of the 26 TPS's profiled, 12 TPS's were functional, with the majority of them exhibiting both monoterpene and sesquiterpene synthase activity.

A newly emerged H7N9 influenza virus poses high risk to human beings. However, the pathogenic mechanism of the virus remains unclear. The temporal response of primary human alveolar adenocarcinoma epithelial cells (A549) infected with H7N9 influenza virus and H1N1 influenza A virus (H1N1, pdm09) were evaluated using the proteomics approaches (2D-DIGE combined with MALDI-TOF-MS/MS) at 24, 48 and 72 hours post of the infection (hpi). There were 11, 12 and 33 proteins with significant different expressions (P

Previous studies have established that abrogation of monoamine oxidase (MAO) A expression leads to a neurochemical, morphological, and behavioral specific phenotype with increased levels of serotonin (5-HT), norepinephrine, and dopamine, loss of barrel field structure in mouse somatosensory cortex, and an association with increased aggression in adults. Forebrain-specific MAO A transgenic mice were generated from MAO A knock-out (KO) mice by using the promoter of calcium-dependent kinase IIalpha (CaMKIIalpha). The presence of human MAO A transgene and its expression were verified by PCR of genomic DNA and reverse transcription-PCR of mRNA and Western blot, respectively. Significant MAO A catalytic activity, autoradiographic labeling of 5-HT, and immunocytochemistry of MAO A were found in the frontal cortex, striatum, and hippocampus but not in the cerebellum of the forebrain transgenic mice. Also, compared with MAO A KO mice, lower levels of 5-HT, norepinephrine, and DA and higher levels of MAO A metabolite 5-hydroxyindoleacetic acid were found in the forebrain regions but not in the cerebellum of the transgenic mice. These results suggest that MAO A is specifically expressed in the forebrain regions of transgenic mice. This forebrain-specific differential expression resulted in abrogation of the aggressive phenotype. Furthermore, the disorganization of the somatosensory cortex barrel field structure associated with MAO A KO mice was restored and became morphologically similar to wild type. Thus, the lack of MAO A in the forebrain of MAO A KO mice may underlie their phenotypes.