Pancreatic cancer is often resistant to chemotherapy. We previously showed the efficacy of combination treatment using gemcitabine and nafamostat mesilate (FUT-175) for patients with unresectable pancreatic cancer. However, the mechanisms that affect the sensitivity of FUT-175 are not fully understood. The purpose of the present study was to clarify the mechanism of the sensitivity to FUT-175, with a focus on the activity of glycogen synthase kinase-3β (GSK-3β). In vitro, we assessed sensitivity to FUT-175 in human pancreatic cancer cell lines (PANC-1 and MIAPaCa-2) and difference of signaling in these cells by cell proliferation assay, Western blot analysis and microarray. Next, we assessed cell viability, apoptotic signal and nuclear factor-kappa B (NF-κB) activity in response to treatment with FUT-175 alone and in combination with GSK-3 inhibitor or protein phosphatase 2A (PP2A) by cell proliferation assay, Western blot analysis and enzyme-linked immunosorbent assay. Phosphorylated GSK-3β level was significantly higher in MIAPaCa-2 (high sensitivity cell) than in PANC-1 (low sensitivity cell). Cell viability and NF-κB activity were significantly decreased by addition of GSK-3 inhibitor to FUT-175, and levels of cleaved caspase-8 were increased by inhibition of GSK-3. PP2A inhibitor increased the levels of phosphorylated GSK-3β and sensitized both cell lines to FUT-175 as measured by cell viability and apoptotic signal. The results indicate that GSK-3β activity plays a key role in the antitumor effect of FUT-175 in pancreatic cancer cells, and regulation of GSK-3β by PP2A inhibition could be a novel therapeutic approach for pancreatic cancer.

With the extension of life expectancy, cancer has been increasing in elderly populations. Postoperative pneumonia can negatively influence immediate mortality following gastrectomy for elderly patients, but its impact on long-term survival remains unclear. We retrospectively reviewed the cases of patients aged ≥75 years who underwent curative gastrectomy for gastric cancer from 2000 to 2014 to determine the long-term effects of postoperative pneumonia and to identify independent risk factors along with physical status and surgical procedure. Of 250 patients, 32 (12.8%) developed postoperative pneumonia. Patients with postoperative pneumonia had significantly worse overall survival (OS) than those without postoperative pneumonia (P<.001). A multivariate analysis identified postoperative pneumonia as a prognostic factor for OS (hazard ratio, 2.06; 95% CI, 1.05-3.75; P=.036). Significant risk factors for the development of postoperative pneumonia were male gender (P=.026) and D2 lymphadenectomy (P<.001). D2 lymphadenectomy was associated with poorer OS than D1 or D1+lymphadenectomy in patients with an American Society of Anesthesiologists (ASA) score 3 (P=.026), but did not influence OS negatively in patients with an ASA score ≤2. Limited lymphadenectomy did not affect the cancer-specific survival of elderly patients with ASA score 3. Postoperative pneumonia following gastrectomy has an adverse impact on the long-term survival of elderly gastric cancer patients. A limited lymphadenectomy during curative resection should be considered to prevent postoperative pneumonia in frail elderly patients with ASA score 3. Postoperative pneumonia following gastrectomy has an adverse impact on the long-term survival of elderly gastric cancer patients. Extent of lymph node dissection during curative resection should be limited to prevent postoperative pneumonia, based on the patient's frailty.

Sirtuin1 (Sirt1) and Sirtuin3 (Sirt3) are known to participate in regulating mitochondrial function. However, whether Total Salvianolic Acid Injection (TSI) protects against myocardial ischemia/reperfusion (I/R) injury through regulating Sirt1, Sirt3, and mitochondrial respiratory chain complexes is unclear. The aim of this study was to explore the effects of TSI on I/R-induced myocardial injury and the underlying mechanism. Male Sprague-Dawley rats were subjected to 30 min occlusion of the left anterior descending coronary artery followed by 90 min reperfusion with or without TSI treatment (8 mg/kg/h). The results demonstrated that TSI attenuated I/R-induced myocardial injury by the reduced infarct size, recovery of myocardial blood flow, and decreased cardiac apoptosis. Moreover, TSI protected heart from oxidative insults, such as elevation of myeloperoxidase, malondialdehyde, hydrogen peroxide, ROS, as well as attenuated I/R-elicited downregulation of Sirt1, Sirt3, NADH dehydrogenase [ubiquinone] 1 alpha subcomplex 10 (NDUFA10), succinate dehydrogenase complex, subunit A, flavoprotein variant (SDHA), and restoring mitochondrial respiratory chain complexes activity. The in vitro study in H9c2 cells using siRNA transfection further confirmed the critical role of Sirt1 and Sirt3 in the effect of TSI on the expression of NDUFA10 and SDHA. These results demonstrated that TSI attenuated I/R-induced myocardial injury via inhibition of oxidative stress, which was related to the activation of NDUFA10 and SDHA through the upregulation of Sirt1 and Sirt3.

Environmental factors can influence the house dust microbiota, which may impact health outcomes. Little is known about how farming exposures impact the indoor microbiota.

BACKGROUND Dynactin (DCTN) is a multi-subunit protein encoded by DCTN genes for 6 subunits. In different diseases the DCTN genes may have different roles; therefore, we investigated the prognostic potential of DCTN mRNA expression in cutaneous melanoma (CM). MATERIAL AND METHODS Data for DCTN mRNA expression in CM patients were obtained from the OncoLnc database, which contains updated gene expression data for 459 CM patients based on the Cancer Genome Atlas. Kaplan-Meier analysis and a Cox regression model were used to determine overall survival (OS) with calculation of hazard ratios (HRs) and 95% confidence intervals (CIs). RESULTS The multivariate survival analysis showed that individually low expression of DCTN1, DCTN2, and DCTN5 and high expression of DCTN6 were associated with favorable OS (adjusted P=0.008, HR=0.676, 95% CI=0.506-0.903; adjusted P=0.004, HR=0.648, 95% CI=0.485-0.867; adjusted P=0.011, HR=0.686, 95% CI=0.514-0.916; and adjusted P=0.018, HR=0.706, 95% CI=0.530-0.942, respectively). In a joint-effects analysis, combinations of low expression of DCTN1, DCTN2, and DCTN5 and high expression of DCTN6 were found to be more highly correlated with favorable OS (all P<0.05). CONCLUSIONS Our findings suggest that downregulated DCTN1, DCTN2, and DCTN5 and upregulated DCTN6 mRNA expression in CM are associated with favorable prognosis and may represent potential prognostic biomarkers. Moreover, use of the 4 genes in combination can improve the sensitivity for predicting OS in CM patients.

Optimization of DNA vaccine delivery improves the potency of the immune response and is crucial to clinical success. Here, we inquired how such optimization impacts the magnitude of the response, its specificity and type. BALB/c mice were DNA-immunized with two model immunogens, HIV-1 protease and reverse transcriptase by intramuscular or intradermal injections with electroporation. DNA immunogens were co-delivered with DNA encoding luciferase. Delivery and expression were monitored by in vivo bioluminescence imaging (BLI). The endpoint immune responses were assessed by IFN-γ/IL-2 FluoroSpot, multiparametric flow cytometry and antibody ELISA. Expression and immunogenicity were compared in relation to the delivery route. Regardless of the route, protease generated mainly IFN-γ, and reverse transcriptase, IL-2 and antibody response. BLI of mice immunized with protease- or reverse transcriptase/reporter plasmid mixtures, demonstrated significant loss of luminescence over time. The rate of decline of luminescence strongly correlated with the magnitude of immunogen-specific response, and depended on the immunogenicity profile and the immunization route. In vitro and in vivo BLI-based assays demonstrated that intradermal delivery strongly improved the immunogenicity of protease, and to a lesser extent, of reverse transcriptase. Immune response polarization and epitope hierarchy were not affected. Thus, by changing delivery/immunogen expression sites, it is possible to modulate the magnitude, but not the type or fine specificity of the induced immune response.

The Macrophage Migration Inhibitory Factor (MIF) is an inflammatory cytokine that is overexpressed in a number of cancer types, with increased MIF expression often correlating with tumor aggressiveness and poor patient outcomes. In this study, we aimed to better understand the link between primary tumor expression of MIF and increased tumor growth. Using the MMTV-PyMT murine model of breast cancer, we observed that elevated MIF expression promoted tumor appearance and growth. Supporting this, we confirmed our previous observation that higher MIF expression supported tumor growth in the 4T1 murine model of breast cancer. We subsequently discovered that loss of MIF expression in 4T1 cells led to decreased cell numbers and increased apoptosis in vitro under reduced serum culture conditions. We hypothesized that this increase in cell death would promote detection by the host immune system in vivo, which could explain the observed impairment in tumor growth. Supporting this, we demonstrated that loss of MIF expression in the primary tumor led to an increased abundance of intra-tumoral IFNgamma-producing CD4+ and CD8+ T cells, and that depletion of T cells from mice bearing MIF-deficient tumors restored growth to the level of MIF-expressing tumors. Furthermore, we found that MIF depletion from the tumor cells resulted in greater numbers of activated intra-tumoral dendritic cells (DCs). Lastly, we demonstrated that loss of MIF expression led to a robust induction of a specialized form of cell death, immunogenic cell death (ICD), in vitro. Together, our data suggests a model in which MIF expression in the primary tumor dampens the anti-tumor immune response, promoting tumor growth.

Preterm children often have language problems. This atypical language development is probably due to atypical brain development. We conducted a systematic review to provide an overview of the extensive and diverse scientific literature on the relations between language outcome and underlying brain structures in school-aged preterm-born children. Embase, Medline Ovid, Web of Science, Cochrane central and Google scholar were searched for relevant studies. Inclusion criteria were: cases are school-aged preterm children; structural MRI (T1- and T2-weighted sequences) or DTI used in combination with a neurocognitive language test; publication in an English-language peer-reviewed journal. Correlational measures between language scores and brain volume or fractional anisotropy of a brain structure were extracted. 23 studies were included. The relations between oral language, verbal fluency and/or written language and MRI/DTI measurements of white matter, gray matter, cerebellum, corpus callosum and/or the fasciculi are presented. Oral language skills and verbal fluency appear to be related to the corpus callosum. Oral language skills are also related to the uncinate fasciculus. There seems to be no clear relation between cerebellar development and verbal fluency skills. Not one single brain area is responsible for atypical language development, but several brain areas and their connections are essential. For future research it is recommended to relate brain areas to oral language skills on a microstructural level in preterm children. We also recommend to use language tests in which it is possible to distinguish between several language domains, such as perceptive and expressive language.

The genus Hypericum is one of the most popular genera in both traditional medicine and scientific platform. This study is designed to provide conceptual insights on the biological potential and chemical characterization of H. salsugineum, which is endemic to Turkey. The qualitative and quantitative phenolic content of the extracts was characterized by HPLC-ESI-MSn. Biological efficiency was investigated by enzyme inhibitory assays (cholinesterases, tyrosinase, amylase, and glucosidase) and anti-cancer efficacy tests (anti-proliferative activities with the iCELLigence technology, colony formation and wound healing scratch assays). Phenolic acids (3-O-caffeoylquinic, 5-O-caffeoylquinic, and 4-O-caffeoylquinic acids) were the predominant group in the studied extracts, although several flavonoids were also detected and quantified. The extracts exhibited good inhibitory effects on tyrosinase and glucosidase, while they had weak ability against cholinesterases and amylase. Computational studies were also performed to explain the interactions between the major phenolics and these enzymes. The extracts displayed significant anti-cancer effects on breast carcinoma cell lines. Our findings suggest that Hypericum salsugineum could be valued as a potential source of biologically-active compounds for designing novel products.

Glioblastoma is a primary brain tumor with a poor prognosis despite of many treatment regimens. Radiotherapy significantly prolongs patient survival and remains the most common treatment. Slit2 and Robo1 are evolutionarily conserved proteins involved in axon guidance, migration, and branching of neuronal cells. New studies have shown that Slit2 and Robo1 could play important roles in leukocyte chemotaxis and glioblastoma cell migration. Therefore, we investigated whether the Slit2/Robo1 complex has an impact on the motility of glioblastoma cells and whether irradiation with therapeutic doses modulates this effect. Our results indicate that photon irradiation increases the migration of glioblastoma cells in vitro. qPCR and immunoblotting experiments in two different glioblastoma cell lines (U-373 MG and U-87 MG) with different malignancy revealed that both Slit2 and Robo1 are significantly lower expressed in the cell populations with the highest motility and that the expression was reduced after irradiation. Overexpression of Robo1 significantly decreased the motility of glioblastoma cells and inhibited the accelerated migration of wild-type cells after irradiation. Immunoblotting analysis of migration-associated proteins (fascin and focal adhesion kinase) and of the epithelial-mesenchymal-transition-related protein vimentin showed that irradiation affected the migration of glioblastoma cells by increasing vimentin expression, which can be reversed by the overexpression of Slit2 and Robo1. Our findings suggest that Robo1 expression might counteract migration and also radiation-induced migration of glioblastoma cells, a process that might be connected to mesenchymal-epithelial transition.

Trypanosomes, protozoan parasites of medical importance, essentially rely on post-transcriptional mechanisms to regulate gene expression in insect vectors and vertebrate hosts. RNA binding proteins (RBPs) that associate to the 3'-UTR of mature mRNAs are thought to orchestrate master developmental programs for these processes to happen. Yet, the molecular mechanisms by which differentiation occurs remain largely unexplored in these human pathogens. Here, we show that ectopic inducible expression of the RBP TcUBP1 promotes the beginning of the differentiation process from non-infective epimastigotes to infective metacyclic trypomastigotes in Trypanosoma cruzi. In early-log epimastigotes TcUBP1 promoted a drop-like phenotype, which is characterized by the presence of metacyclogenesis hallmarks, namely repositioning of the kinetoplast, the expression of an infective-stage virulence factor such as trans-sialidase, increased resistance to lysis by human complement and growth arrest. Furthermore, TcUBP1-ectopic expression in non-infective late-log epimastigotes promoted full development into metacyclic trypomastigotes. TcUBP1-derived metacyclic trypomastigotes were infective in cultured cells, and developed normally into amastigotes in the cytoplasm. By artificial in vivo tethering of TcUBP1 to the 3' untranslated region of a reporter mRNA we were able to determine that translation of the reporter was reduced by 8-fold, while its mRNA abundance was not significantly compromised. Inducible ectopic expression of TcUBP1 confirmed its role as a translational repressor, revealing significant reduction in the translation rate of multiple proteins, a reduction of polysomes, and promoting the formation of mRNA granules. Expression of TcUBP1 truncated forms revealed the requirement of both N and C-terminal glutamine-rich low complexity sequences for the development of the drop-like phenotype in early-log epimastigotes. We propose that a rise in TcUBP1 levels, in synchrony with nutritional deficiency, can promote the differentiation of T. cruzi epimastigotes into infective metacyclic trypomastigotes.

Mortality information of populations is aggregated in life tables that serve as a basis for calculation of life expectancy and various life disparity measures. Conventional life-table methods address right-censoring inadequately by assuming a constant hazard in the last open-ended age group. As a result, life expectancy can be substantially distorted, especially in the case when the last age group in a life table contains a large proportion of the population. Previous research suggests addressing censoring in a gamma-Gompertz-Makeham model setting as this framework incorporates all major features of adult mortality. In this article, we quantify the difference between gamma-Gompertz-Makeham life expectancy values and those published in the largest publicly available high-quality life-table databases for human populations, drawing attention to populations for which life expectancy values should be reconsidered. We also advocate the use of gamma-Gompertz-Makeham life expectancy for three reasons. First, model-based life-expectancy calculation successfully handles the problem of data quality or availability, resulting in severe censoring due to the unification of a substantial number of deaths in the last open-end age group. Second, model-based life expectancies are preferable in the case of data scarcity, i.e. when data contain numerous age groups with zero death counts: here, we provide an example of hunter-gatherer populations. Third, gamma-Gompertz-Makeham-based life expectancy values are almost identical to the ones provided by the major high-quality human mortality databases that use more complicated procedures. Applying a gamma-Gompertz-Makeham model to adult mortality data can be used to revise life-expectancy trends for historical populations that usually serve as input for mortality forecasts.

Noncatalytic carbohydrate binding modules (CBMs) have been demonstrated to play various roles with cognate catalytic domains. However, for polysaccharide lyases (PLs), the roles of CBMs remain mostly unknown. AlyB is a multidomain alginate lyase that contains CBM32 and a PL7 catalytic domain. The AlyB structure determined herein reveals a noncanonical alpha helix linker between CBM32 and the catalytic domain. More interestingly, CBM32 and the linker does not significantly enhance the catalytic activity but rather specifies that trisaccharides are predominant in the degradation products. Detailed mutagenesis, biochemical and cocrystallization analyses show "weak but important" CBM32 interactions with alginate oligosaccharides. In combination with molecular modeling, we propose that the CBM32 domain serves as a "pivot point" during the trisaccharide release process. Collectively, this work demonstrates a novel role of CBMs in the activity of the appended PL domain and provides a new avenue for the well-defined generation of alginate oligosaccharides by taking advantage of associated CBMs.

Chronic stress with lack of reward presumably may impair brain reward circuit, leading to major depressive disorder (MDD). Most individuals experiencing chronic stress do not suffer from MDD, i.e., resilience, implying the presence of endogenous anti-depression in the brain. Molecular mechanisms underlying stress-induced depression versus resilience were investigated. Mice were treated by chronic unpredictable mild stress (CUMS) for four weeks. Their mood state was assessed by behavioral tasks, such as sucrose preference, Y-maze and forced swimming testes. To reveal comprehensive molecular profiles of major depression versus resilience, mRNA and microRNA profiles were analyzed by high-throughput sequencing in the ventral tegmental area (VTA) harvested from control, CUMS-susceptible and CUMS-resilience mice. In data analyses of control versus CUMS-susceptible mice as well as control versus CUMS-resilience mice, 1.5 fold ratio in reads per kilo-base per million reads was set as the threshold to judge the involvement of mRNAs and microRNAs in the CUMS, depression or resilience. The downregulation of synaptic vesicle cycle, neurotrophin, GABAergic synapse and morphine addiction as well as the upregulation of transmitter release, calcium signal and cAMP-dependent response element binding are associated to CUMS-susceptibility. The downregulation of tyrosine metabolism and protein process in endoplasmic reticulum as well as the upregulation of amino acid biosynthesis, neuroactive ligand-receptor interaction and dopaminergic synapse are associated to CUMS-resilience. Therefore, the impairment of neurons and GABA/dopaminergic synapses in the VTA is associated with major depression. The upregulation of these entities is associated with resilience. Consistent results obtained from analyzing mRNAs and microRNAs as well as using different approaches strengthen our finding and conclusion.

Four molecular weights of poly-l-lysine (PLL) [1000-5000 Da, 1500-8000 Da, 4000-15,000 Da and 15,000-30,000 Da] and three molecular weights of polyethyleneimine (PEI) [800 Da, 2,000 Da, and 25,000 Da] were used to systematically study the effect of calcium (Ca2+) to improve transfection efficiency of polyelectrolyte complexes. Complexes made using different molecular weights of PLL or PEI polymer showed clear differences in the levels of gene expression in the presence and absence of calcium chloride when tested using A549 human lung carcinoma cells. Complexes formed from PLL or PEI 800 Da were exhibited negligible expression of pDNA according to a luciferase reporter assay. Low molecular weight PLL and PEI 800 Da, however, became highly efficient gene delivery vehicles when calcium was added to the nascent complexes while maintaining the low cytotoxicity of low molecular weight polyamines. Additional analyses indicated that the most effective formulations utilized a threshold level of calcium, which created small, stable particles, but also facilitated unpackaging of the gene complexes and release of pDNA.

The excess from fecal samples submitted to a centralized laboratory in Roanoke, Virginia for routine C. difficile testing was used for this research study. We tested all samples, including any formed samples usually not assayed in diagnostic laboratories. Our first aim was to rank ribotypes by their frequency. Between 2007 and 2013, fluoroquinolone resistant 027 (027FQR), a multi-drug resistant ribotype, was 32% of 3118 Clostridium difficile isolates and the most common of 128 ribotypes. 027FQR was in 45% of cytotoxin positive but only 17% of cytotoxin negative fecal samples (p = 0.001) and 34% of unformed but only 21% of formed stool samples (p = 0.001), strong associations with features of symptomatic infection. Conversely, 014/020 (10% of isolates, third most common ribotype) was more often in unformed than formed stools (14% versus 9%; p = 0.002) and in cytotoxin negative than cytotoxin positive samples (11% versus 8%, p = 0.01). Fecal lactoferrin levels, an indication of intestinal inflammation, were significantly higher with 027FQR than with 014/020 infections (median 308 versus 26 ng/mL, p = 0.02). 027FQR fecal bioburdens and toxin levels were significantly higher than their 014/020 equivalents (median 104.1 versus 103.2/g feces, p = 0.01; median TcdA 58.7 versus 1.3 ng/g feces, p = 0.04; median TcdB 43.4 versus 0.3 ng/g feces, p = 0.001). Binary toxin was present in 40% of 027FQR positive samples but none of the 014/020 or non-toxigenic C. difficile positive samples. 027FQR made no more TcdA/cell than did 014/020 (p = 0.7) but did make close to significantly more TcdB/cell (p = 0.08).

Platinum-based chemotherapeutic drugs, especial cisplatin, are the most common and effective anticancer drugs to treat the non-small cell lung cancer (NSCLC), but the major obstacle of this treatment is the resistance to chemotherapeutic drugs due to the anti-apoptosis of cancer cells. In our study, we found that Canopy homolog 2 (CNPY2) is increased in NSCLC tissues compared to the normal lung tissues, and the upregulation of CNPY2 is correlated with poor survival. Next, colony formation, annexin V-binding and TUNEL assays revealed that overexpression of CNPY2 inhibits the apoptosis of NSCLC cells induced by cisplatin. Further assays demonstrated that the anti-apoptosis may be aroused by the hyperactivation of NF-κB signaling pathway, and blocking the NF-κB pathway promotes the apoptosis of CNPY2-upregulating cells. The above results suggest that CNPY2 can serves as a therapeutic target to promote the effect of chemotherapy in NSCLC.

Targeting protein for Xenopus kinesin-like protein 2 (TPX2) is a microtubule-associated proteinrequired for mitosis and spindle assembly. It has been revealed that TPX2 is overexpressedin various human cancers and promotes cancer progression.

Platinum nanoparticles (PtNPs) attract great attention due to their efficient catalysis and good degree of cytocompatibility, but information about their effects on the human immune system is still missing. Monocytes are key cells of the innate immune system and the understanding of their reactions to PtNPs is crucial in view of any feasible application to human pathologies. Here, we evaluate the internalization of citrate-coated PtNPs into THP-1 monocytes and its consequences on immune cell responses. We found that the presence of intracellular PtNPs efficiently reduce reactive oxygen species (ROS) without affecting cell viability. The physiological expression of the immune receptors Cluster of Differentiation 14 (CD14), CD11b, CC-Chemokine Receptor 2 (CCR2) and CCR5 and the expression of cytokines and chemokines are not compromised by the presence of PtNPs within THP-1 cells. On the other hand, the treatment with PtNPs modulates the transcription of sixty genes, some of them involved in lipopolysaccharide (LPS) signaling in different cells. However, the treatment with PtNPs of monocytes does not compromise the LPS-induce increase of cytokines in THP-1 monocytes in vitro. Our results demonstrate that citrate-coated PtNPs are non-toxic, perform efficient intracellular reactive oxygen species (ROS) scavenging activity and possess good immune-compatibility, suggesting them as feasible synthetic enzymes for applications in nanomedicine.

Treatment variation in medicine may be driven by evidence gaps, clinician factors, and patient preferences. Although well-documented in human medicine, variation in clinical management is relatively unexplored in veterinary practice. Clinical vignette questionnaires were administered to a cross section of general practitioners (GPs) and veterinarians with postgraduate training in ophthalmology (PGs) to survey recommended management of canine prolapsed nictitans gland ("cherry eye", PNG) and feline herpesvirus (FHV-1) keratitis. The majority of veterinarians (96.2%) suggested surgical replacement of cherry eye, with a pocketing technique being the most frequently nominated procedure. GPs were more likely to suggest gland excision in the event of surgical failure, while PGs more frequently nominated techniques incorporating a periosteal anchor for salvage repair. Most respondents managed FHV-1 keratitis with topical antibiotics (76.4%), with a minority suggesting topical antivirals (32.2%). GPs favoured topical acyclovir whilst PGs more frequently recommended topical trifluorothymidine. A significantly larger proportion of PGs nominated systemic famciclovir and lysine supplement for FHV-1 keratitis. This survey revealed moderate treatment variation for these conditions, both between and within practitioner groups. Additional research is needed to assess the reasons for this variation, particularly for conditions in which high quality evidence is scant.