Three IRF4 paralogs act as negative regulators of type â IFN responses in yellow catfish (Pelteobagrus fulvidraco).

IRF4 is a master member of the interferon regulatory factor (IRF) family playing vital regulatory roles in immune system development and function. Tetrapods have a single-copy IRF4 gene, while teleosts harbor duplicated IRF4 genes. This work describes three IRF4 paralogs from yellow catfish (Pelteobagrus fulvidraco), designated PfIRF4A, PfIRF4B and PfIRF4B-like. These genes all contain a typical IRF structural architecture. Phylogenic and synteny analyses indicate that they should arise from the teleost-specific whole-genome duplication. PfIRF4 genes are abundantly expressed in the immune-related tissues and upregulated by PolyI:C, LPS, and Edwardsiella ictaluri. Ectopic expression of these genes inhibits the activation of fish type Ⅰ IFN promoters and downregulates the transcription levels of IFN-responsive genes, thus allowing the efficient replication of a fish rhabdovirus, spring viremia of carp virus (SVCV). PfIRF4s possess a repressive effect on MyD88-mediated activation of IFN and NF-κB. Some differences are observed between each individual paralog. PfIRF4B is the main form expressed across the tissues and the most up-regulated one after pathogen induction. It exerts a stronger inhibitory effect on IFN antiviral response than the other two paralogs. PfIRF4A and PfIRF4B-like are primarily present in the nucleus, while PfIRF4B displays colocalization and direct associations with MyD88 in the cytoplasm. Overall, the data demonstrate that three PfIRF4 paralogs show shared and individual functional properties in the negative regulation of type Ⅰ IFN response.

CAR-based cell therapy: evaluation with bibliometrics and patent analysis.

Chimeric antigen receptors-based cell therapies have shown impressive preclinical and clinical success and revolutionized biomedicine. However, the link between science and invention, the impact of international cooperation, and the influence and prestige of CARs research have not been explored. This study analyzed the landscape of peer-reviewed articles and patents related to CARs. A total of 5,681 publications were analyzed using bibliometrics and machine learning-based text mining to assess publication metrics, subject areas, and research hotspots. 5,010 Inpadoc families were also analyzed for patent filing trends, priority countries, and applicant and inventor rankings. The results show that CARs research has the following distinctive features: high research prestige among research community; strong global geographical bias in both academic output and patenting patterns; strong links between science and invention, but significant differences among countries; and an inverse relationship between country size and international collaboration rates.

Red elemental selenium (Se0 ) improves the immunoactivities of EPC cells, crucian carp and zebrafish against spring viraemia of carp virus.

Selenium, as an essential trace element, interferes through selenoproteins in many physiological processes of plants and mammals. Its antiviral activity has recently attracted much attention because selenium improves the antiviral capacity of animal cells against a few viruses relevant to human diseases. In this study, the red elemental selenium was purified from the fermentative culture of Herbaspirillum camelliae WT00C and then used to culture epithelioma papulosum cyprinid (EPC) cells or feed crucian carp and zebrafish. Finally, its antiviral effects were investigated at the cell level and living fishes after spring viraemia of carp virus infection. At the cell level, 5, 10 and 20 µg ml-1 red elemental selenium significantly induced the expression of interferon (IFN) and ISG15 genes in EPC cells. The viral TCID50 (50% tissue culture infective dose) values in the EPC cells incubated with 5, 10 and 20 µg ml-1 red elemental selenium were significantly less than those of the control. More expression of IFN and ISG15 genes and less TCID50 values indicate that red elemental selenium indeed improves the antiviral capability of EPC cells. In the crucian carp fed with the food containing 5 and 10 µg g-1 red elemental selenium, IFN expressions showed 13- and 39-fold increases at the 16th day of post-injection, and its expression was dependent on selenium concentrations. Meanwhile, no fish death occurred in all the experimental groups. In the zebrafish fed with the red worm containing 5 µg g-1 red elemental selenium, IFN and Mx expressions and survival rate were significantly higher than those of the control. The results of this study show that red elemental selenium indeed improves the antiviral activity of fish. The antiviral effects of selenium mainly come from its immune regulation through its incorporation into selenoproteins. The optimum level of selenium contributes to improving fish immunity, whereas excess selenium causes excessive immune and inflammatory responses.

Molecular characterization of yellow catfish (Pelteobagrus fulvidraco) IRF7 suggests involvement in innate immune response.

Interferon regulatory factor 7 (IRF7) serves as a critical mediator in the regulation of type Ι interferon (IFN) response to invading pathogens. Here, an ortholog of IRF7 was characterized in yellow catfish (Pelteobagrus fulvidraco). The full-length cDNA of PfIRF7 consisted of 1516 bp encoding a polypeptide of 425 amino acids. PfIRF7 protein comprised a typical IRF structural architecture, including a DNA binding domain (DBD), an IRF association domain (IAD) and a serine-rich domain (SRD). PfIRF7 was expressed predominantly in the immune-related tissues and transcriptionally upregulated by PolyI:C, LPS, and Edwardsiella ictaluri. Ectopic expression of PfIRF7 led to activation of fish type I IFN promoters and induction of IFN and Vig1, thereby conferring a strong antiviral effect against spring viremia of carp virus (SVCV). Overall, the present data suggest that PfIRF7 may play an essential role in type I IFN response of yellow catfish.

Membrane disruptive antimicrobial potential of NK-lysin from yellow catfish (Pelteobagrus fulvidraco).

NK-lysins, a type of broad-spectrum antimicrobial peptide (AMP), act as an essential effector of innate defense against microbial attack in higher vertebrates and so in fish. The present study delineates the structural and functional characterization of NK-lysin from yellow catfish (Pelteobagrus fulvidrac) (Pelteobagrus fulvidraco). PfNK-lysin encodes a 153-residue peptide, which displays the hallmark features of other known NK-lysins with the ordered array of six well-conserved cysteine residues and five-exon/four-intron structure. It was found to be ubiquitous in tissues, being detected most abundantly in gill and head kidney. In vivo exposure to stimuli (LPS, PolyI:C, and Edwardsiella ictaluri) induced PfNK-lysin expression in head kidney and spleen. Synthetic PfNK-lysin-derived peptide exhibited in vitro bactericidal potency against both Gram-positive and Gram-negative bacteria, with the highest inhibitory effect on pathogen Edwardsiella ictaluri. Fluorescence microscopy and scanning electron microscopy further confirmed its capacity to cause damage to the bacterial plasma membrane. Taken together, these data suggest that PfNK-lysin might participate in antimicrobial defense of yellow catfish by membrane-disruptive action.

Soluble expression and purification of high-bioactivity recombinant human bone morphogenetic protein-2 by codon optimisation in Escherichia coli.

We developed a simple method of preparing recombinant human bone morphogenetic protein-2 (rhBMP-2) with high biological activity. This rhBMP-2 was overproduced in Escherichia coli as a fusion protein with thioredoxin 6xHis-tag at its amino terminus. The cDNA fragment of human bone morphogenetic protein-2 (hBMP-2) fused to the secretion signal of alkaline phosphatase (PhoA) was expressed under T7 promoter in E. coli. After DNA sequence confirmation, the recombinant vector pETpho-bmp2 was transformed into E. coli BL21 (DE3). rhBMP-2 was produced by the recombinant strain pETpho-bmp2/BL21 (DE3) in a soluble form with an yield of 6.2 mg/L culture. Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS-PAGE) results showed that the molecular weight of the product was approximately 28 kD. Moreover, rhBMP-2 was secreted as a dimer with a natural structure. rhBMP-2, purified by Ni Nitrilotriacetic acid Agarose (Ni-NTA) affinity chromatography, was used to examine osteosarcoma MG-63 cells and assay the alkaline phosphatase (ALP) activity. Results showed that rhBMP-2 induced MG-63 cell differentiation. When the final concentration was 500 ng/mL, the effect was more remarkable and ALP activity reached 525% compared with that of the control group.

WITHDRAWN: Effects of cancer-associated point mutations on the structure, function, and stability of succinate dehydrogenase A.

This article has been withdrawn by the authors. Some of the SDHA enzyme activity data were flawed and were not performed and analyzed correctly. The withdrawing authors are in the process of correcting the data and re-evaluating them for resubmission.

Activated tyrosine kinases in gastrointestinal stromal tumor with loss of KIT oncoprotein expression.

Oncogenic KIT or PDGFRA receptor tyrosine kinase (TK) mutations are compelling therapeutic targets in gastrointestinal stromal tumors (GISTs), and the KIT/PDGFRA kinase inhibitor, imatinib, is the standard of care for patients with metastatic GIST. However, approximately 10% of KIT-positive GIST metastases lose KIT expression at the time of clinical progression during imatinib therapy. In the present report, we performed TK-activation screens, using phosphotyrosine-TK double immunoaffinity purification and mass spectrometry, in GIST in vitro models lacking KIT expression. These studies demonstrated tyrosine-phosphorylated EGFR, AXL, and EPHA2 in four of six KIT-negative GIST lines (GIST62, GIST522, GIST54, GIST226, GIST48B, and GIST430B), and tyrosine-phosphorylated focal adhesion kinase (FAK) in each of the six KIT-negative lines. AXL expression was strong in KIT-negative or -weak clinical GIST samples that were obtained from progressing metastases during imatinib therapy. AXL knockdown inhibited viability in three KIT-negative GIST cell lines (GIST62, GIST54, and GIST522), but not in an AXL-negative, KIT-positive GIST control cell line (GIST430). AXL inhibition by R428, a specific AXL kinase inhibitor, reduced viability in AXL-activated GIST54. AXL knockdown in GIST62, GIST522, and GIST54 was accompanied by an increase in p21, p27, and p53 expression. By contrast, gefitinib-mediated EGFR inhibition, PF562271-mediated FAK inactivation, and shRNA-mediated knockdowns of EPHA2 and FAK had no effect on viability or colony formation of the KIT-negative GISTs. These findings highlight the potential relevance of AXL/p53 signaling as a therapeutic target in a subset of GISTs that have lost KIT oncoprotein expression.

Epidemiology of schizophrenia and risk factors of schizophrenia-associated aggression from 2011 to 2015.

Objective To investigate the risk factors associated with aggression in patients with schizophrenia. Methods Patient clinical, behavioural, and demographic information was collected and reported online to the Beijing Mental Health Information Management System by psychiatrists. We used chi-square tests to analyse information between 2011 and 2015 to determine the prevalence and incidence of schizophrenia and the rate of aggression. We used univariate and binary logistic regression to analyse risk factors of aggressive behaviours. Results The prevalence and incidence of schizophrenia, and the proportion of cases displaying aggressive behaviour, increased considerably from 2011 to 2015. Risk of aggression was associated with non-adherence to medication (odds ratio [OR]: 2.92; 95% confidence intervals [CI]: 2.08-4.11), being unmarried (OR: 1.62; 95% CI: 1.03-2.55), having physical disease (OR: 3.26; 95% CI: 2.28-4.66), and higher positive symptom scores (OR: 2.01; 95% CI: 1.06-3.81). Physical disease was a risk factor associated with committing more than one type of aggression. Conclusion We confirmed that demographic factors, treatment-related factors, and clinical symptoms were associated with aggression in patients with schizophrenia in Beijing. A focus on improving controllable factors, including medication adherence and physical health status, might help to prevent aggressive behaviour.

Dual Targeting of Insulin Receptor and KIT in Imatinib-Resistant Gastrointestinal Stromal Tumors.

Oncogenic KIT or PDGFRA receptor tyrosine kinase (RTK) mutations are compelling therapeutic targets in gastrointestinal stromal tumors (GIST), and treatment with the KIT/PDGFRA inhibitor imatinib is the standard of care for patients with metastatic GIST. Most GISTs eventually acquire imatinib resistance due to secondary mutations in the KIT kinase domain, but it is unclear whether these genomic resistance mechanisms require other cellular adaptations to create a clinically meaningful imatinib-resistant state. Using phospho-RTK and immunoblot assays, we demonstrate activation of KIT and insulin receptor (IR) in imatinib-resistant GIST cell lines (GIST430 and GIST48) and biopsies with acquisition of KIT secondary mutations, but not in imatinib-sensitive GIST cells (GIST882 and GIST-T1). Treatment with linsitinib, a specific IR inhibitor, inhibited IR and downstream intermediates AKT, MAPK, and S6 in GIST430 and GIST48, but not in GIST882, exerting minimal effect on KIT phosphorylation in these cell lines. Additive effects showing increased apoptosis, antiproliferative effects, cell-cycle arrest, and decreased pAKT and pS6 expression, tumor growth, migration, and invasiveness were observed in imatinib-resistant GIST cells with IR activation after coordinated inhibition of IR and KIT by linsitinib (or IR shRNA) and imatinib, respectively, compared with either intervention alone. IGF2 overexpression was responsible for IR activation in imatinib-resistant GIST cells, whereas IR activation did not result from IR amplification, IR mutation, or KIT phosphorylation. Our findings suggest that combinatorial inhibition of IR and KIT warrants clinical evaluation as a novel therapeutic strategy in imatinib-resistant GISTs. Cancer Res; 77(18); 5107-17. ©2017 AACR.

Relationship preferences and experience of primary care patients in continuity of care: a case study in Beijing, China.

BACKGROUND: Continuity of care can bring a wide range of benefits to consumers, providers and health care systems. This study aimed to understand the relationship preferences of primary care patients and their associations with patient experience of continuity of care. METHODS: A questionnaire survey was conducted on 700 patients who sought medical care from a community health organisation in Beijing. The survey contained four items examining the relationship preferences of the respondents, and a modified Questionnaire of Continuity between Care Levels (CCAENA) measuring patient experience of continuity of care based on a three dimensional (relational, informational and managerial) model. The associations between the relationship preferences and the experience of respondents in continuity of care was tested using a linear regression model controlling for age, sex, education, medical insurance, personal income and servicing facilities. RESULTS: The respondents experienced relatively lower levels of informational and managerial continuity compared with relational continuity of care. More than 80% of respondents preferred free choice and a continuing relationship with doctors, compared with 59% who endorsed community facility control over hospital appointments. A preference for a continuing relationship with doctors was associated with all aspects of continuity of care. A preference in favour of community facility control over hospital appointments was a strong predictor of managerial continuity (ß = 0.333, p < 0.001) and informational continuity (ß = 0.256, p < 0.001). Patient preference for free choice of doctors was positively associated with relational continuity with specialists (p < 0.001), but not with primary care providers (p > 0.08). Perceived importance of information exchange was associated with relational and managerial continuity (p < 0.05), but not with informational continuity (p = 0.34). CONCLUSIONS: Patients prefer a high level of freedom of choice and sustained individual relationship with doctors. Relationship preferences of patients are associated with their experience of continuity of care. But patient strong preference for free choice of doctors is not aligned with relational continuity with primary care, a desirable feature of cost-effective healthcare systems.

JAK-STAT signaling mediates the senescence of bone marrow-mesenchymal stem cells from systemic lupus erythematosus patients.

Previous studies have revealed that bone marrow-mesenchymal stem cells (BM-MSCs) from systemic lupus erythematosus (SLE) patients exhibited early signs of senescence, which may participate in the development of SLE. However, the molecular mechanisms about this phenomenon have not been fully elucidated. In the current study, we aimed to investigate whether Janus kinase (JAK)-signaling transducers and activators of transcription (STAT) signaling mediated the senescence of BM-MSCs from SLE patients. Twelve female SLE patients and healthy subjects were enrolled in the study. All BM-MSCs were isolated by density gradient centrifugation. Western blot analysis was used to test the expression of JAK-STAT signaling molecules. We observed the activity of ß-gal of cells, the changes of cytoskeletal structure by F-actin staining, and the distribution of cell cycle by flow cytometry. BM-MSCs from SLE patients showed prominent features of senescence, and abnormal activation of JAK-STAT signaling transduction, high level of phosphorylated JAK2, and STAT3. After stimulation of IFN-γ in normal MSCs, JAK-STAT signaling was activated. The cell volume and the number of senescence-associated ß-galactosidase (SA-ß-gal) positive in SLE BM-MSCs were increased. The organization of cytoskeleton was nearly disordered. The rate of cell proliferation was decreased. AG490, the inhibitor of JAK2, and knockdown of STAT3 in BM-MSCs, could significantly reverse the senescence. In summary, our study indicated that JAK-STAT signaling pathway may play a critical role in the senescence of SLE BM-MSCs.

Oncolytic viruses against cancer stem cells: A promising approach for gastrointestinal cancer.

Gastrointestinal cancer has been one of the five most commonly diagnosed and leading causes of cancer mortality over the past few decades. Great progress in traditional therapies has been made, which prolonged survival in patients with early cancer, yet tumor relapse and drug resistance still occurred, which is explained by the cancer stem cell (CSC) theory. Oncolytic virotherapy has attracted increasing interest in cancer because of its ability to infect and lyse CSCs. This paper reviews the basic knowledge, CSC markers and therapeutics of gastrointestinal cancer (liver, gastric, colon and pancreatic cancer), as well as research advances and possible molecular mechanisms of various oncolytic viruses against gastrointestinal CSCs. This paper also summarizes the existing obstacles to oncolytic virotherapy and proposes several alternative suggestions to overcome the therapeutic limitations.

Effect of Community-Based Social Skills Training and Tai-Chi Exercise on Outcomes in Patients with Chronic Schizophrenia: A Randomized, One-Year Study.

BACKGROUND: Antipsychotic drugs are limited in their ability to improve negative symptoms, quality of life, and medication adherence in patients with schizophrenia. The addition of nonpharmacological interventions like social skills training has a positive effect on medication adherence and decreases rehospitalization rates but is limited in improving patients' symptoms, aggressive behaviors, and quality of life. Aerobic exercise, especially Tai-chi, can potentially reduce psychopathological and negative symptoms, decrease aggressive behaviors, and improve quality of life. It is an ideal rehabilitation intervention for patients with schizophrenia. However, no study has investigated the effects of social skills training plus Tai-chi on outcomes among outpatients with schizophrenia. This study analyzes the effect of antipsychotics combined with community-based integrated interventions on outcomes of schizophrenia. METHODS: In this study, a 24-session social skills training plus Tai-chi was used in community settings among patients with schizophrenia. A total of 244 patients were randomly assigned to medication treatment alone (MTA group) or community-based integrated intervention (CBII group), which accepted social skills training plus Tai-chi in addition to medication treatment. Generalized linear mixed models were used to evaluate the intervention effect (group effect), intervention effect over time (time effect), and interaction effect (group × time effect). t tests were used to evaluate between-group differences on clinical variables. Multiple linear regression analysis was used to analyze the differences between the intervention at 12 months and baseline for the Positive and Negative Syndrome Scale (PANSS) negative symptoms and quality of life-social domain. RESULTS: Compared with the MTA group, the CBII group had lower scores on PANSS (F = 17.312, p < 0.001) and negative symptoms (F = 44.909, p < 0.001), a lower risk for aggressive behavior (F = 12.382, p < 0.001), and a greater improvement in adherence to medication (F = 12.391, p < 0.001) after 1 year of intervention. The changes in PANSS total scores, negative scores, and social domain of the World Health Organization Quality of Life Scale-Brief version (WHOQOL-BREF) from baseline to 12 months were significant between the two groups (PANSS total score: t = 4.839, p < 0.001; negative symptoms: t = 8.250, p < 0.001, and quality of life-social domain: t = -2.171, p = 0.031). Multiple linear regression analysis also showed that the intervention was significantly effective for changes from baseline to 12 months on PANSS total score (B = 0.804, p < 0.001), negative score (B = 0.709, p < 0.001), and social domain of quality of life (B = -0.673, p = 0.044). CONCLUSIONS: This study suggested that the community-based integrated intervention such as social skills training plus Tai-chi should be part of a rehabilitation effort for patients with schizophrenia in order to improve clinical symptoms, quality of life, and medication adherence.

Downregulation of cyclin D1 sensitizes cancer cells to MDM2 antagonist Nutlin-3.

The MDM2-p53 pathway has a prominent oncogenic function in the pathogenesis of various cancers. Nutlin-3, a small-molecule antagonist of MDM2-p53 interaction, inhibits proliferation in cancer cells with wild-type p53. Herein, we evaluate the expression of MDM2, both the full length and a splicing variant MDM2-A, and the sensitivity of Nutlin-3 in different cancer cell lines. Included are seven cell lines with wild-type p53 (four mesothelioma, one breast cancer, one chondrosarcoma, and one leiomyosarcoma), two liposarcoma cell lines harboring MDM2 amplification and wild-type p53, and one mesothelioma cell line harboring a p53 point mutation. Nutlin-3 treatment increased expression of cyclin D1, MDM2, and p53 in cell lines with wild-type p53. Additive effects were observed in cells containing wild-type p53 through coordinated attack on MDM2-p53 binding and cyclin D1 by lentivirual shRNA knockdown or small molecule inhibition, as demonstrated by immunoblots and cell viability analyses. Further results demonstrate that MDM2 binds to cyclin D1, and that an increase in cyclin D1 expression after Nutlin-3 treatment is correlated with expression and ubiquitin E3-ligase activity of MDM2. MDM2 and p53 knockdown experiments demonstrated inhibition of cyclin D1 by MDM2 but not p53. These results indicate that combination inhibition of cyclin D1 and MDM2-p53 binding warrants clinical evaluation as a novel therapeutic strategy in cancer cells harboring wild-type p53.

[Screening of anti-lung cancer bioactive compounds from Curcuma longa by target cell extraction and UHPLC/LTQ Orbitrap MS].

A target cell extraction-chemical profiling method based on human alveolar adenocarcinoma cell line (A549 cells) and UHPLC/LTQ Orbitrap MS for screening the anti-lung cancer bioactive compounds from Curcuma longa has been developed in this paper. According to the hypothesis that when cells are incubated together with the extract of Curcuma longa, the potential bioactive compounds in the extract should selectively combine with the cells, then the cell-binding compounds could be separated and analyzed by LC-MS. The bioactive compounds in C. longa are lipophilic components. They intend to be absorbed on the inner wall of cell culture flask when they were incubated with A549 cells, which will produce interference in the blank solution. In this paper, by using cells digestion and multi-step centrifugation and transfer strategy, the interference problem has been solved. Finally, using the developed method, three cell-binding compounds were screened out and were identified as bisdemethoxycurcumin, demethoxycurcumin, and curcumin. These compounds are the main bioactive compounds with anti-lung cancer bioactivity in C. longa. The improved method developed in this paper could avoid the false positive results due to the absorption of lipophilic compounds on the inner wall of cell culture flask, which will to be an effective complementary method for current target cell extraction-chemical profiling technology.

Comparison of paroxetine and agomelatine in depressed type 2 diabetes mellitus patients: a double-blind, randomized, clinical trial.

BACKGROUND: Comorbid depression/anxiety in type 2 diabetes mellitus (DM) patients is highly prevalent, affecting both diabetes control and quality of life. However, the best treating method for depression/anxiety in type 2 DM patients is still unclear. This study was conducted to compare the efficacy of paroxetine and agomelatine on depression/anxiety and metabolic control of type 2 DM patients. METHODS: A total of 116 depressed, type 2 DM patients were recruited for 12 weeks treatment. Patients were randomly assigned to receive either paroxetine or agomelatine. Hamilton Depression Rating Scale and Hamilton Anxiety Rating Scale were used to assess depression and anxiety, respectively. Hemoglobin A1c, fasting plasma glucose, and body mass index were assessed at baseline and at the end of the trial. RESULTS: At the end of the trial, there were 34 (60.7%) responders and 22 (39.3%) remissions in paroxetine group; and 38 (63.3%) responders and 26 (43.3%) remissions in agomelatine group. Compared to paroxetine group, lower depression scores were observed in agomelatine group. Fasting plasma glucose and body mass index were not significantly different after 12 weeks treatment between the two groups, but agomelatine group had a significantly lower final hemoglobin A1c level compared to paroxetine group. The two antidepressants had comparable acceptability. CONCLUSION: These results showed that compared to paroxetine, agomelatine might have some advantages in treating symptoms of depression/anxiety and glycemic control in depressed type 2 DM patients. The clinical applicability of agomelatine shows greater promise and should be explored further. Limited by the relatively small samples, future studies are needed to verify and support our findings.