Differential bicistronic gene translation mediated by the internal ribosome entry site element of encephalomyocarditis virus.

BACKGROUND: Internal ribosome entry sites (IRESs) allow the translation of a transcript independent of its cap structure. They are distributed in some viruses and cellular RNA. The element is applied in dual gene expression in a single vector. Although it appears the lower efficiency of IRES-mediated translation than that of cap-dependent translation, it is with the crucial needs to know the precise differences in translational efficacy between upstream cistrons (cap-dependent) and downstream cistrons (IRES-mediate, cap-independent) before applying the bicistronic vector in biomedical applications. METHODS: This study aimed to provide real examples and showed the precise differences for translational efficiency dependent upon target gene locations. We generated various bicistronic constructs with quantifiable reporter genes as upstream and downstream cistrons of the encephalomyocarditis virus (EMCV) IRES to precisely evaluate the efficacy of IRES-mediated translation in mammalian cells. RESULTS: There was no significant difference in protein production when the reporter gene was cloned as an upstream cistron. However, lower levels of protein production were obtained when the reporter gene was located downstream of the IRES. Moreover, in the presence of an upstream cistron, a markedly reduced level of protein production was observed. CONCLUSION: Our findings demonstrate the version of the EMCV IRES that is provided in many commercial vectors is relatively less efficient than cap-dependent translation and provide valuable information regarding the utilization of IRES to facilitate the expression of more than one protein from a transcript.

Locally Targeting the IL-17/IL-17RA Axis Reduced Tumor Growth in a Murine B16F10 Melanoma Model.

Interleukin (IL)-17 and the cells that produce it within the tumor microenvironment appear to promote tumor development and are associated with survival in cancer patients. Here we investigated the role of the IL-17/IL-17 receptor A (IL-17RA) axis in regulating melanoma progression and evaluated the therapeutic potential of blocking the IL-17/IL-17RA pathway. First, recombinant mouse IL-17 (γmIL-17) treatment significantly increased proliferation of mouse B16F10 cells and human A375 and A2058 cells. Silencing IL-17RA by small hairpin RNA (shRNA) in B16F10 cells reduced the γmIL-17-elicited cell proliferation, migration, and invasion, and significantly reduced vascular endothelial growth factor and matrix metalloproteinase production. Remarkably, knockdown of IL-17RA led to a significantly decreased capability of B16F10 cells to form tumors in vivo, similar to that in IL-17-deficient mice. Finally, local application of an adenovirus delivering a shRNA against IL-17RA mRNA not only significantly suppressed tumor development, but also enhanced antitumor immunity by increasing the interferon γ-expressing T cells and not T regulatory cells. Our results highlight the critical role of the IL-17/IL-17RA pathway in tumor progression and imply that targeting IL-17RA represents a promising therapeutic strategy.

Interleukin 17 and peripheral IL-17-expressing T cells are negatively correlated with the overall survival of head and neck cancer patients.

The presence and clinical significance of interleukin (IL)-17 and IL-17-expressing cells have recently been studied in several types of cancer, but their correlation to tumor development remains controversial. Additionally, the contribution of peripheral IL-17-expressing cells to head and neck cancer (HNC) progression is still poorly understood. We collected peripheral blood from healthy donors and HNC patients to isolate PBMCs. The percentages of IL-17-expressing cells and the production of inflammatory cytokines in PBMCs were measured to determine their association with clinical outcomes and overall survival in HNC. We evaluated the effect and potential mechanism of IL-17 on human oral squamous carcinomas in vitro using exogenous IL-17 stimulation. In comparison to healthy donors, the PBMCs of HNC patients have a significant accumulation of IL-17-expressing T cells and their frequencies were positively correlated with the disease stage. A significantly higher production of PBMC IL-17, TGF-ß and IL-21 and plasma VEGF-A were found in HNC patients. Importantly, the 5-years overall survival of HNC patients with a higher percentage of IL-17-expressing cells is significantly decreased. Furthermore, the addition of IL-17 appeared to promote human oral squamous carcinoma cell proliferation via the production of IL-6 and VEGF-A. Our findings suggest that IL-17 has the potential to mediate pro-tumor immunity in the HNC tumor microenvironment. Enhanced IL-17-expressing cells, including Th17 and Tc17 cells, in the peripheral blood could be a significant predictor of a poor prognosis for HNC patients.

The modulation of Th2 immune pathway in the immunosuppressive effect of human umbilical cord mesenchymal stem cells in a murine asthmatic model.

BACKGROUND: Asthma is a chronic airway inflammatory disease that has a high prevalence nowadays, and seeking the means of relieving asthmatic symptoms is now an issue with increased importance. While mesenchymal stem cells have been demonstrated to display immunomodulatory effects, the effect of fetus-type mesenchymal stem cells (MSCs) on asthmatic symptoms in vivo have not been reported to date. METHODS: Female BALB/c mice at 8 weeks of age were sensitized by ovalbumin, and MSCs derived from Wharton's jelly of human umbilical cord mesenchymal stem cells (hUCMSCs) were injected into the asthmatic mice. Airway hyper-responsiveness, lung eosinophil infiltration, cytokine level in splenocyte cultures and serum immunoglobulin level were measured. Enzyme-linked immunosorbent assay was used to determine cytokine and immunoglobulin levels. RESULTS: This current study demonstrated that hUCMSCs attenuated both lung lymphocyte and eosinophil infiltration, and significantly decreased the concentration of Th2 cytokines interleukin-5 in splenocyte cultures. CONCLUSIONS: Human umbilical cord mesenchymal stem cells have the advantage of being easily harvested non-invasively and are capable of rapid proliferation, therefore an ideal material for stem cell-based immune therapies. The current study showed that fetal-type MSCs were able to suppress asthmatic symptoms efficiently, and its immunomodulatory effect resulted primarily from suppressing the Th2 pathway in the animal model. This study suggested that hUCMSCs could be an ideal candidate for cell-based therapies of asthma.

The Correlation between Chitin and Acidic Mammalian Chitinase in Animal Models of Allergic Asthma.

Asthma is the result of chronic inflammation of the airways which subsequently results in airway hyper-responsiveness and airflow obstruction. It has been shown that an elicited expression of acidic mammalian chitinase (AMCase) may be involved in the pathogenesis of asthma. Our recent study has demonstrated that the specific suppression of elevated AMCase leads to reduced eosinophilia and Th2-mediated immune responses in an ovalbumin (OVA)-sensitized mouse model of allergic asthma. In the current study, we show that the elicited expression of AMCase in the lung tissues of both ovalbumin- and Der P2-induced allergic asthma mouse models. The effects of allergic mediated molecules on AMCase expression were evaluated by utilizing promoter assay in the lung cells. In fact, the exposure of chitin, a polymerized sugar and the fundamental component of the major allergen mite and several of the inflammatory mediators, showed significant enhancement on AMCase expression. Such obtained results contribute to the basis of developing a promising therapeutic strategy for asthma by silencing AMCase expression.

Immune checkpoint blockade therapy: the 2014 Tang Prize in Biopharmaceutical Science.

The first Tang Prize for Biopharmaceutical Science has been awarded to Prof. James P. Allison and Prof. Tasuku Honjo for their contributions leading to an entirely new way to treat cancer by blocking the molecules cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) and programmed cell death protein 1 (PD-1) that turn off immune response. The treatment, called "immune checkpoint blockade therapy," has opened a new therapeutic era. Here the discoveries of the immune checkpoints and how they contribute to the maintenance of self-tolerance, as well as how to protect tissues from the excess immune responses causing damage are reviewed. The efforts made by Prof. Allison and Prof. Honjo for developing the most promising approaches to activate therapeutic antitumor immunity are also summarized. Since these certain immune checkpoint pathways appear to be one of the major mechanisms resulting in immune escape of tumors, the presence of anti-CTLA-4 and/or anti-PD-1 should contribute to removal of the inhibition signals for T cell activation. Subsequently, it will enhance specific T cell activation and, therefore, strengthen antitumor immunity.

Assessing schizophrenia-relevant cognitive and social deficits in mice: a selection of mouse behavioral tasks and potential therapeutic compounds.

Schizophrenia and other psychiatric disorders are generally diagnosed based on a collection of symptoms defined by a combination of an individual's feelings, perceptions, and behaviors. Many of these disorders are characterized by specific cognitive and social deficits. Although it is nearly impossible to recapitulate the full phenotypic spectrum of schizophrenia in mice, mouse models play an indispensable role in understanding the pathogenesis of this disorder and the development of new therapeutics. Genetic mouse models of schizophrenia and mouse behavioral tests provide a feasible approach for elucidating causal relationships between susceptibility gene(s) and schizophrenia-related symptoms. There has been a proliferation of studies characterizing basic behavioral phenotypes in mice. Since there is no way to completely model human psychiatric symptoms in mice, the major role of behavioral tests is to provide insights into underlying affected circuitry and pathophysiology. Given that the recovery of cognitive and social abilities significantly benefits functional outcomes, there has been an increasing interest in characterizing cognitive and social functions in mutant mice; however, these functions are not easy to measure. In this review, a selection of conventional behavioral tasks was briefly described and three specific behavioral tasks aimed at characterizing social communication, attentional function, and choice behavior in mice were highlighted. The choice of specific behavioral tasks during experimental planning should take into consideration a variety of factors, including their validity, reliability, sensitivity, utility, and specificity. Based upon the hypothetical hypofunction of N-methyl-D-aspartate receptor (NMDAR)-mediated signaling pathways in the involvement of cognitive and social impairments in schizophrenia, three NMDAR-related compounds/drugs, D-serine, sarcosine, and D-cycloserine, are discussed as an example.

Blockade of phospholipid scramblase 1 with its N-terminal domain antibody reduces tumorigenesis of colorectal carcinomas in vitro and in vivo.

BACKGROUND: Membrane-bound phospholipid scramblase 1 (PLSCR1) is involved in both lipid trafficking and cell signaling. Previously, we showed that PLSCR1 is overexpressed in many colorectal carcinomas (CRCs). In the present study, we investigated the tumorigenic role of PLSCR1 in CRC and suggest that it is a potential therapeutic target. METHODS: To identify PLSCR1 as a therapeutic target, we studied the tumorigenic properties of CRC cell lines treated with a monoclonal antibody (NP1) against the N-terminus of PLSCR1 in vitro and in vivo. We also investigated cell cycle status and epidermal growth factor receptor-related pathways and downstream effectors of PLSCR1 after blocking its function with NP1. RESULTS: Treating CRC cells with NP1 in vitro and in vivo decreased cell proliferation, anchorage-independent growth, migration, and invasion. Adding NP1 to the CRC cell line HT29 caused arrest at G1/S. Treating HT29 cells with NP1 significantly decreased the expression of cyclin D1 and phosphorylation levels of Src, the adaptor protein Shc, and Erks. The reduced level of cyclin D1 led to an increase in the activated form of the tumor suppressor retinoblastoma protein via dephosphorylation. These actions led to attenuation of tumorigenesis. CONCLUSIONS: Therefore, PLSCR1 may serve as a potential therapeutic target for CRC.

Evaluating the effects of an Internet education programme on newborn care in Taiwan.

AIM: The objectives of this study were to evaluate an Internet education programme provided to primigravida in the third trimester of pregnancy with the aim of enhancing mothers' knowledge about newborn care and increasing their maternal confidence. BACKGROUND: Shorter hospital stays have had an impact on the traditional role of mother-baby nurses in providing education about parenting to their parturient women. Internet education is an efficient way to provide nursing instruction. DESIGN: A randomised controlled trial was used. A total of 118 women receiving prenatal care in a hospital clinic who met study criteria and who consented were assigned randomly to intervention and control groups. The study was conducted at a hospital in Taiwan. METHODS: The target population was women at 32-34 weeks gestation, using the Internet on a regular basis. The primigravida were randomly assigned to either the control group (n = 57) or the experimental group (n = 61). Two primary outcome measures were newborn-care knowledge and maternal confidence. RESULTS: The changes in newborn-care knowledge were 7.21 for the experimental group, compared with 1.95 for the control group; the difference between the least-squares means computed by ancova was 5.73 and statistically significant (p < 0.001). The changes in maternal confidence were 8.46 for the experimental group and 3.05 for the control group; the difference between the least-squares means computed by ancova was 5.94 and statistically significant (p < 0.001). CONCLUSION: Results suggest that Internet education about newborn care may contribute to greater care knowledge and maternal confidence. Relevance to clinical practice. Internet newborn-care education programmes can achieve success in promoting newborn care and provide health professionals with evidence-based intervention.