GPCR signaling contributes to immune characteristics of microenvironment and process of EBV-induced lymphomagenesis.

Epstein-Barr virus (EBV) is the oncogenic driver of multiple cancers. However, the underlying mechanism of virus-cancer immunological interaction during disease pathogenesis remains largely elusive. Here we reported the first comprehensive proteogenomic characterization of natural killer/T-cell lymphoma (NKTCL), a representative disease model to study EBV-induced lymphomagenesis, incorporating genomic, transcriptomic, and in-depth proteomic data. Our multi-omics analysis of NKTCL revealed that EBV gene pattern correlated with immune-related oncogenic signaling. Single-cell transcriptome further delineated the tumor microenvironment as immune-inflamed, -deficient, and -desert phenotypes, in association with different setpoints of cancer-immunity cycle. EBV interacted with transcriptional factors to provoke GPCR interactome (GPCRome) reprogramming. Enhanced expression of chemokine receptor-1 (CCR1) on malignant and immunosuppressive cells modulated virus-cancer interaction on microenvironment. Therapeutic targeting CCR1 showed promising efficacy with EBV eradication, T-cell activation, and lymphoma cell killing in NKTCL organoid. Collectively, our study identified a previously unknown GPCR-mediated malignant progression and translated sensors of viral molecules into EBV-specific anti-cancer therapeutics.

Prenatal diagnosis of total anomalous pulmonary venous connection using 2D and HDlive flow combined with spatiotemporal image correlation.

OBJECTIVES: The objective of this study is to examine the application value of two-dimensional (2D) and high-definition live (HDlive) flow combined with spatiotemporal image correlation (STIC) in diagnosing fetal total anomalous pulmonary venous connection (TAPVC). METHODS: Seventeen cases of fetal TAPVC were diagnosed using 2D and HDlive Flow combined with STIC. These cases were then retrospectively analyzed to examine the value of using 2D and HDlive Flow combined with STIC in the diagnosis of TAPVC. RESULTS: 2D and HDlive Flow combined with STIC detected 13 cases of supracardiac TAPVC (two isolated cases, seven cases with right atrial isomerism (RAI), four cases with other complex malformations), one case of isolated intra-cardiac TAPVC, and three cases of cardiac TAPVC (two isolated cases and one case with complex congenital heart anomaly). Small left atrium (LA), the absence of PVs drainage into the LA and the increased retroatrial distance between LA and the descending aorta (DAo) were significant signs that should raise the suspicion of fetal TAPVC. HDlive Flow combined with STIC can dynamically display the TAPVC which may assit the prenatal diagnosis of TAPVC. CONCLUSION: 2D and HDlive Flow combined with STIC can assit the diagnosis of fetal TAPVC abnormalities and has important clinical value.

Molecular characterization and pathogenesis of H9N2 avian influenza virus isolated from a racing pigeon.

H9N2 avian influenza viruses (AIVs) can cross species barriers and expand from birds tomammals and humans. It usually leads to economic loss for breeding farms and poses a serious threat to human health.This study investigated the molecular characteristics of H9N2 AIV isolated from a racing pigeon and its pathogenesis in BALB/c mice and pigeons. Phylogenetic analysis indicated that the H9N2 virus belonged to the Ck/BJ/94-like lineage, and acquired multiple specific amino acid substitutions that might contribute to viral transmission from birds to mammals and humans. A pathogenesis study showed that both mice and pigeons infected with H9N2 virus showed clinical signs and mortality. The H9N2 viruses efficiently replicated in mice and pigeons. In our study, high levels of viral shedding were detected in pigeons, but the infection was not transmitted to co-housed pigeons. Histopathological examination revealed the presence of inflammatory responses in the infected mice and pigeons. Immunohistochemical analysis showed the presence of H9N2 virus in multiple organs of the infected mice and pigeons. Moreover, the infected mice and pigeons demonstrated significant cytokine/chemokine production. Our results showed that the H9N2 virus can infect mice and pigeons, and can not be transmitted between pigeons through direct contact.

Lead toxicity induces autophagy to protect against cell death through mTORC1 pathway in cardiofibroblasts.

Heavy metals, such as lead (Pb(2+)), are usually accumulated in human bodies and impair human's health. Lead is a metal with many recognized adverse health side effects and yet the molecular processes underlying lead toxicity are still poorly understood. In the present study, we proposed to investigate the effects of lead toxicity in cultured cardiofibroblasts. After lead treatment, cultured cardiofibroblasts showed severe endoplasmic reticulum (ER) stress. However, the lead-treated cardiofibroblasts were not dramatically apoptotic. Further, we found that these cells determined to undergo autophagy through inhibiting mammalian target of rapamycin complex 1 (mTORC1) pathway. Moreover, inhibition of autophagy by 3-methyladenine (3-MA) may dramatically enhance lead toxicity in cardiofibroblasts and cause cell death. Our data establish that lead toxicity induces cell stress in cardiofibroblasts and protective autophagy is activated by inhibition of mTORC1 pathway. These findings describe a mechanism by which lead toxicity may promote the autophagy of cardiofibroblasts cells, which protects cells from cell stress. Our findings provide evidence that autophagy may help cells to survive under ER stress conditions in cardiofibroblasts and may set up an effective therapeutic strategy for heavy metal toxicity.

Nitrifying biocathode enables effective electricity generation and sustainable wastewater treatment with microbial fuel cell.

Simultaneous organics removal and nitrification using a novel nitrifying biocathode microbial fuel cell (MFC) reactor were investigated in this study. Remarkably, the introduction of nitrifying biomass into the cathode chamber caused higher voltage outputs than that of MFC operated with the abiotic cathode. Results showed the maximum power density increased 18% when cathode was run under the biotic condition and fed by nitrifying medium with alkalinity/NH4+-N ratio of 8 (26 against 22 mW/m2). The voltage output was not differentiated when NH4+-N concentration was increased from 50 to 100 mg/L under such alkalinity/NH4+-N ratio. However, interestingly, the cell voltage rose significantly when the alkalinity/NH4+-N ratio was decreased to 6. Consequently, the maximum power density increased 68% in compared with the abiotic cathode MFC (37 against 22 mW/m2). Polarization curves demonstrated that both activation and concentration losses were lowered during the period of nitrifying biocathode operation. Ammonium was totally nitrified and mostly converted to nitrate in all cases of the biotic cathode conditions. High COD removal efficiency (98%) was achieved. In light of the results presented here, the application of nitrifying biocathode is not only able to integrate the nitrogen and carbon removal but also to enhance the power generation in MFC system. Our system can be suggested to open up a new feasible way for upgrading and retrofitting the existing wastewater treatment plant by the use of MFC-based technologies.

Estimation of allele frequency in pooled DNA by using PCR-RFLP combined with microchip electrophoresis.

Biallelic marker, most commonly single nucleotide polymorphism (SNP), is widely utilized in genetic association analysis, which can be speeded up by estimating allele frequency in pooled DNA instead of individual genotyping. Several methods have shown high accuracy and precision for allele frequency estimation in pools. Here, we explored PCR restriction fragment length polymorphism (PCR-RFLP) combined with microchip electrophoresis as a possible strategy for allele frequency estimation in DNA pools. We have used the commercial available Agilent 2100 microchip electrophoresis analysis system for quantifying the enzymatically digested DNA fragments and the fluorescence intensities to estimate the allele frequencies in the DNA pools. In this study, we have estimated the allele frequencies of five SNPs in a DNA pool composed of 141 previously genotyped health controls and a DNA pool composed of 96 previously genotyped gastric cancer patients with a frequency representation of 10-90% for the variant allele. Our studies show that accurate, quantitative data on allele frequencies, suitable for investigating the association of SNPs with complex disorders, can be estimated from pooled DNA samples by using this assay. This approach, being independent of the number of samples, promises to drastically reduce the labor and cost of genotyping in the initial association analysis.

[Effects of different delivery routes of CD34+ stem cells on cardiac function in the ischemic cardiomyopathy of rats].

OBJECTIVE: To compare the effects of CD34-positive stem cells delivered by different routes on cardiac function in rats with ischemic cardiomyopathy, and to evaluate the efficacy and safety of stem cell transplantation. METHODS: Sixty-four male Wistar rats were randomized into cell infusion group (n=30), acute myocardial infarction (AMI) model group (n=20) and sham operation group (n=14). AMI model was reproduced by ligation of left anterior descending coronary artery. CD34+ stem cells mobilized with granulocyte-colony stimulating factor (G-CSF) were purified by immunomagnetic beads in 30 donor rats. Seven days after the injury about (7-9)x10(7) CD34+ stem cells were infused through the femoral vein or through epicardium of recipient rats respectively. Cardiac function was evaluated before AMI, 1, 2 and 4 weeks after cell delivery. Hemodynamic parameters were determined 4 weeks after cell infusion. RESULTS: Compared with model group, left ventricular ejection fraction(LVEF), fractional shortening (DeltaFS), left ventricular end-systolic pressure (LVESP) and maximal positive change in filling pressure versus time (+dp/dt max) were improved significantly (all P<0.01), whereas left ventricular end-systolic dimension (LVESD), left ventricular end-diastolic pressure (LVEDP), maximal negative change in filling pressure versus time (-dp/dt max), time constant of left ventricular relaxation (Tc) were lowered in cell infusion groups (all P<0.01). There were no significant differences in cardiac function indexes between intravenous infusion and trans-epicardial injection group (all P>0.05). CONCLUSION: Intravenous and trans-epicardial delivery of hematopoietic stem cells (HSC) can significantly improve cardiac function, and both methods may be safe and effective for the treatment of AMI.

Simultaneous organics removal and bio-electrochemical denitrification in microbial fuel cells.

Simultaneous organics removal and bio-electrochemical denitrification using a microbial fuel cell (MFC) reactor were investigated in this study. The electrons produced as a result of the microbial oxidation of glucose in the anodic chamber were transferred to the anode, which then flowed to the cathode in the cathodic chamber through a wire, where microorganisms used the transferred electrons to reduce the nitrate. The highest power output obtained on the MFCs was 1.7 mW/m(2) at a current density of 15 mA/m(2). The maximum volumetric nitrate removal rate was 0.084 mg NO(3)(-)-N cm(-2) (electrode surface area) day(-1). The coulombic efficiency was about 7%, which demonstrated that a substantial fraction of substrate was lost without current generation.

Poly[µ-aqua-diaqua-(µ(3)-N'-carboxy-methyl-ethylenediamine-N,N,N'-tri-acetato)oxidopotassium(I)vanadium(IV)].

In the crystal structure of the title compound, [KV(C(10)H(13)N(2)O(8))O(H(2)O)(3)](n), the V(IV) ion adopts a distorted octa-hedral geometry, coordinated by one oxide group, two N and three carboxylate O atoms from the same N'-carboxy-methyl-ethyl-ene-diamine-N,N,N'-triacetate (HEDTA) ligand. The potassium ion is hepta-coordinated by two water mol-ecules, two bridging water mol-ecules and three carboxylate O atoms from three neighbouring HEDTA ligands. The HEDTA ligands and some of the water mol-ecules act as bridges, linking the compound into a three-dimensional architecture via 2(1) screw, c-glide, translation and inversion symmetry operators. Meanwhile, three types of O-H⋯O hydrogen bonds provide an additional stabilization of the three-dimensional architecture.

[Effects of adenovirus-mediated gene transfer of ICOSIg fusion protein on experimental autoimmune myocarditis in Lewis rats].

OBJECTIVE: To explore the effects of adenovirus vector-mediated gene transfer of ICOSIg fusion protein on experimental autoimmune myocarditis (EAM) in Lewis rats. METHODS: Expression vector containing ICOSIg (p-Adeno-ICOSIg) was constructed by fusion of human ICOS and IgGFc segment. Adenovirus vector was digested by PacI enzyme and transfected into HEK 293 cells. Adenovirus expressing ICOSIg was produced. EGFP was constructed into adenovirus vector and used as control. EAM was induced in Lewis rats by injection of porcine cardiac myosin. All immunized Lewis rats were divided into 4 groups. Group A (n = 15) and B (n = 15) received adenovirus containing ICOSIg on day 0 and day 14 respectively to study the effects of costimulatory molecules gene therapy on T cell activation and inflammation; group C (n = 10) and group D (n = 10) received adenovirus containing EGFP on day 0 and day 14 respectively as controls. Group E (n = 10) was normal controls that did not receive immunization. On day 28, all rats were killed after echocardiography examination. Histopathological examination was performed to observe myocardial inflammation. Protein levels of ICOS, ICOSL, B7-1 and B7-2 were detected by Western blot. INF-gamma, IL-2 and IL-4 mRNA were determined by realtime RT-PCR. RESULTS: On day 28, cardiac function was significantly improved and myocardial inflammation significantly attenuated in group B compared to group A, C and D (all P < 0.05). B7-1 expression at protein level was significantly lower in group B than that of group C (P < 0.05). ICOS and ICOSL expressions at protein level were significantly decreased in both group A and B compared with group C and D (P < 0.05). IFN-gamma mRNA level significantly decreased and IL-4 mRNA significantly increased in group A and B compared to group C and D (P < 0.05). CONCLUSIONS: Blockade of costimulatory pathway with gene therapy of ICOSIg alleviated autoimmune inflammatory damage and improved cardiac function in Lewis rats with EAM. Down-regulated costimulatory molecules in the myocardium and reduced inflammatory cytokine secretion might be responsible for the beneficial effects of ICOSIg in this model.

[IL-10 gene modification on immature dendritic cells induces antigen-specific tolerance in experimental autoimmune myocarditis].

OBJECTIVE: To investigate whether IL-10 gene modification on immature dendritic cells (iDC) could induce autoimmune tolerance in rat experimental autoimmune myocarditis (EAM). METHODS: EAM was induced by cardiac myosin immunization on day 0 and day 7 in rats. A total of 2 x 10(6) mature DC (mDC), iDC, pcDNA3 transfected iDC, pcDNA3-IL-10 transfected iDC or PBS were injected intravenously at 5th immunization day. Three weeks later, echocardiography and HE staining were performed to observe the cardiac function and myocardial inflammation. Th1/Th2 cytokines were detected by ELISA and MHC-II molecules, costimulatory molecules were identified by flow cytometry. In vitro T lymphocyte proliferation assay and adoptive transfer of DCs were performed to determine the antigen specific tolerance induced by IL-10 gene modification on iDCs. RESULTS: EAM rats treated with pcDNA3-IL-10 transfected iDC showed improved cardiac function and reduced inflammatory cells infiltration into myocardium. Moreover, lower Th1 and higher Th2-type response was induced, MHC-II and costimulatory molecules down-regulated and antigen specific immunological responses towards cardiac myosin inhibited in pcDNA3-IL-10-iDC treated EAM rats. CONCLUSION: Treatment with IL-10 gene modified iDCs could ameliorates EAM by inducing Th2 polarization and down-regulation of MHC-II molecules and costimulatory molecule expressions.

Antigen-specific tolerance induced by IL-10 gene modified immature dendritic cells in experimental autoimmune myocarditis in rats.

BACKGROUND: Experimental autoimmune myocarditis (EAM) in rats is a T-cell-mediated disorder. The initiation and maintenance of autoimmune responses in EAM depend on the maturation state of dendritic cells. IL-10 is a pleiotrophic immunomodulatory cytokine that functions at different levels of the immune response, so it has emerged as a promising therapeutic factor for the treatment of autoimmune/inflammatory diseases. This study was designed to test the hypothesis that IL-10 gene modified bone marrow-derived immature dendritic cells (iDCs) ameliorate EAM and to explore the underlying mechanisms. METHODS: EAM was induced using the methods of cardiac myosin immunization on day 0 and day 7. Immature and mature bone marrow-derived dendritic cells (BMDCs) were generated without or with the stimulation by lipopolysaccharide (LPS) and the phenotype was analyzed by flow cytometry. Some of the iDCs were transfected by pcDNA3-IL-10 plasmid. 2 x 10(6)/per rat mature DC (mDC), immature DC (iDC), pcDNA3 transfected iDC, pcDNA3-IL-10 transfected iDC or phosphate buffered saline (PBS) were injected intravenously for treatment 5 days after the first immunization. On day 21, HE staining was performed to detect the myocardial inflammation and T lymphocyte proliferation assay was used to determine the effects of IL-10 gene transfected iDC on autoreactive T cell proliferation. Expression of IkappaB, the inhibitor of NF-kappaB pathway, was determined by Western blot. RESULTS: BMDCs generated in a medium supplemented with granulocyte-macrophage-colony-stimulating factor (GM-CSF) were relatively immature, as determined by flow cytometry. However, stimulation with LPS induced these cells to become mature (m) DCs with higher levels of surface major histocompatibility complex (MHC)-II and costimulatory molecules. Intravenous administration of iDCs, especially pcDNA3-IL-10 transfected iDC, ameliorated the histopathological severity of the myosin induced-EAM, and the effect was lost after the DCs underwent maturation induced by in vitro exposure to LPS. IL-10 gene modified iDC inhibited the antigen specific T cell responses towards cardiac myosin. IkappaB protein was up-regulated significantly in the IL-10 gene modified iDC group. CONCLUSIONS: IL-10 gene modified iDC induced antigen-specific tolerance in EAM. The underlying mechanisms may be related to costimulatory molecules down-regulation and NF-kappaB pathway inhibition.