N-Feruloyl Serotonin Attenuates Neuronal Oxidative Stress and Apoptosis in Aß25-35-Treated Human Neuroblastoma SH-SY5Y Cells.

Amyloid-beta (Aß) aggregation and deposition have been identified as a critical feature in the pathology of Alzheimer's disease (AD), with a series of functional alterations including neuronal oxidative stress and apoptosis. N-feruloyl serotonin (FS) is a plant-derived component that exerts antioxidant activity. This study investigated the protective effects of FS on Aß25-35-treated neuronal damage by regulation of oxidative stress and apoptosis in human neuroblastoma SH-SY5Y cells. The radical scavenging activities increased with the concentration of FS, exhibiting in vitro antioxidant activity. The Aß25-35-treated SH-SY5Y cells exerted neuronal cell injury by decreased cell viability and elevated reactive oxygen species, but that was recovered by FS treatment. In addition, treatment of FS increased anti-apoptotic factor B-cell lymphoma protein 2 (Bcl-2) and decreased the pro-apoptotic factor Bcl-2-associated X protein. The FS attenuated Aß-stimulated neuronal apoptosis by regulations of mitogen-activated protein kinase signaling pathways. Moreover, activated CREB-BDNF signaling was observed by the treatment of FS in Aß25-35-induced SH-SY5Y cells. These results demonstrate that FS shows potential neuroprotective effects on Aß25-35-induced neuronal damage by attenuation of oxidative stress and apoptosis, and suggest that FS may be considered a promising candidate for the treatment of AD.

Concept analysis of uncertainty in people with mental disabilities.

PURPOSE: This concept analysis clarified "uncertainty in people with mental disabilities". METHODS: The research was conducted using Walker & Avant's conceptual analysis methodology. RESULTS: Uncertainty in people with mental disabilities showed the defining attributes of "ambiguity," "unpredictability," "distrust," and "lack of insight into illness," of which "distrust" and "lack of insight" were unique to people with mental disabilities. Accordingly, it was defined as a state in which it is difficult to determine the treatment's timing because (1) the disease process is often too ambiguous to make a decision; (2) abnormal behavior can recur at any time while the patients are under treatment and in recovery; and (3) patients struggle to accept the illness, tend to distrust healthcare professionals and family, and show lack of knowledge, making them miss their treatment's timing and even making the expectation for recovery vague. CONCLUSION: The attributes of uncertainty in people with mental disabilities identified in this study confirmed that mental health professionals should proactively intervene to treat patients at an appropriate time and continuously manage them to prevent recurrence. PRACTICE IMPLICATION: The study findings can be utilized in mental health research and in developing interventions to reduce uncertainty in people with mental disabilities, helping them recover and integrate into the community.

Reelin protects against pathological α-synuclein accumulation and dopaminergic neurodegeneration after environmental enrichment in Parkinson's disease.

Two of the primary features of Parkinson's disease (PD) are the accumulation of α-synuclein (α-Syn) and the depletion of lysosomal-associated membrane protein 1 (LAMP1) in the brain. Beneficial effects of environmental enrichment (EE) have been reported on the activation of lysosomal function and the amelioration of PD symptoms. Furthermore, Reelin could be a novel therapeutic target in PD. Hence, in this study, we validated the effects of EE on the activation of LAMP1 via Reelin in PD. Heterogeneous α-Syn (A53T)-overexpressing transgenic mice (age 6 and 16 months) were exposed to EE for 8 weeks. After motor and cognitive tests, brain tissues were obtained from mice and subjected to immunohistochemistry and molecular analyses. EE ameliorated motor and non-motor symptoms, protected dopamine neurons, and reduced pathological α-Syn accumulation in the early stage of PD. Striatal Reelin levels were altered depending on the disease stage and regulated by EE in PD mice. To elucidate the underlying mechanism of the effect of EE on PD, we performed further molecular and cellular analyses using activated preformed fibril (PFF)-induced SH-SY5Y cells, an in vitro model of PD, which were treated with recombinant Reelin protein and a Reelin blocker, CR-50. The CR-50 increased pathological α-Syn accumulation and accelerated dopamine neuronal degeneration by decreasing LAMP1 in the PFF-induced PD model. Our results showed that Reelin increased LAMP1 after EE and decreased pathological α-Syn accumulation, thus protecting dopamine neurons from degeneration in the striatum and substantia nigra, and ameliorating neurobehavioral deficits. These results suggest that Reelin is a promising target in treating histopathological changes and improving behavioral symptoms associated with PD.

Apigenin Ameliorates Scopolamine-Induced Cognitive Dysfunction and Neuronal Damage in Mice.

We investigated the protective effect and mechanisms of apigenin against cognitive impairments in a scopolamine-injected mouse model. Our results showed that intraperitoneal (i.p.) injection of scopolamine leads to learning and memory dysfunction, whereas the administration of apigenin (synthetic compound, 100 and 200 mg/kg/day) improved cognitive ability, which was confirmed by behavioral tests such as the T-maze test, novel objective recognition test, and Morris water maze test in mice. In addition, scopolamine-induced lipid peroxidation in the brain was attenuated by administration of apigenin. To further evaluate the protective mechanisms of apigenin on cognitive and memory function, Western blot analysis was carried out. Administration of apigenin decreased the B-cell lymphoma 2-associated X/B-cell lymphoma 2 (Bax/Bcl-2) ratio and suppressed caspase-3 and poly ADP ribose polymerase cleavage. Furthermore, apigenin down-regulated the ß-site amyloid precursor protein-cleaving enzyme, along with presenilin 1 (PS1) and PS2 protein levels. Apigenin-administered mice showed lower protein levels of a receptor for advanced glycation end-products, whereas insulin-degrading enzyme, brain-derived neurotrophic factor (BDNF), and tropomyosin receptor kinase B (TrkB) expression were promoted by treatment with apigenin. Therefore, this study demonstrated that apigenin is an active substance that can improve cognitive and memory functions by regulating apoptosis, amyloidogenesis, and BDNF/TrkB signaling pathways.

Chemoembolisation for hepatocellular carcinoma with bile duct invasion: is preprocedural biliary drainage mandatory?

PURPOSE: To determine the necessity of preprocedural biliary drainage prior to chemoembolisation for hepatocellular carcinoma (HCC) with bile duct invasion. MATERIALS AND METHODS: The study included 52 patients who received chemoembolisation for unresectable HCC invading bile duct and causing hyperbilirubinemia (>3 mg/dL). Patients were divided into three groups according to biliary drainage and its effect: effective drainage (n=21), ineffective drainage (n=17), and non-drainage (n=14). Thirty-day mortality, length of hospitalisation, adverse events recorded using Common Terminology Criteria for Adverse Events (CTCAE), survival, and tumour response was compared among three groups. RESULTS: Thirty-day mortality rates were 14.3% (n=3), 17.6% (n=3), and 7.1% (n=1) for effective, ineffective, and non-drainage groups, respectively, and did not differ significantly among groups (p=0.780). The mean length of hospitalisation was shorter in non-drainage group compared to ineffective drainage group (12.1±11.4 vs 34.1±29.6 days, p=0.012). Mean differences in CTCAE grade for laboratory parameters before and after chemoembolisation were not significantly different among three groups. Survival among three groups was not significantly different (p=0.239-0.825). The tumour response was also not significantly different among three groups (p=0.679). CONCLUSION: Biliary drainage may not be mandatory prior to chemoembolisation in patients with HCC invading the bile duct. KEY POINTS: • Chemoembolisation without biliary drainage can be performed for icteric HCC. • Chemoembolisation without biliary drainage is not accompanied by increased adverse events. • Preprocedural biliary drainage may not be mandatory for chemoembolisation for icteric HCC.

Tauroursodeoxycholic acid attenuates progression of steatohepatitis in mice fed a methionine-choline-deficient diet.

BACKGROUND AND AIM: Endoplasmic reticulum (ER) stress has been implicated in the development of nonalcoholic steatohepatitis. A methionine-choline-deficient (MCD) diet induces robust ER stress response and steatohepatitis, but the effects of ER stress modulation on the course of steatohepatitis remain uncertain. The present study evaluated whether reducing ER stress using the chemical chaperone tauroursodeoxycholic acid (TUDCA) could limit hepatocyte lipoapoptosis and progression of MCD diet-induced steatohepatitis. METHODS: HuH7 cells stably transfected with sodium taurocholate cotransporting polypeptide (HuH-Ntcp cells) and palmitate (PA) were used. Experimental steatohepatitis was induced in male C57BL/6 mice using an MCD diet, and three different doses of TUDCA (500, or 1,000 mg/kg, once daily; or 500 mg/kg twice daily) were administered by gavage from the start of the MCD diet regimen or after 4 weeks. RESULTS: TUDCA reduced PA-induced ER stress as manifested by decreased eIF2α phosphorylation, XBP1 splicing and expression of BiP, ATF4, and CHOP in HuH-Ntcp cells. TUDCA also decreased PA-induced JNK phosphorylation, Puma up-regulation and Bax activation, which in turn suppressed caspase-dependent hepatocyte lipoapoptosis. Mice given TUDCA did not show a significant decrease in the intrahepatic triglyceride contents and steatosis. However, TUDCA treatment significantly reduced hepatic damage compared to controls for both early and late treatment groups. TUDCA treatment reduced the expression of ER stress markers and pro-apoptotic proteins, leading to decreased apoptosis and oxidative stress. Finally, TUDCA reduced histological fibrosis along with the down-regulation of pro-fibrotic gene expression in both early and late treatment groups. CONCLUSIONS: These results show that TUDCA attenuates the progression of MCD diet-induced steatohepatitis by reducing ER stress.

Effect of annealing environments on self-organized TiO2 nanotubes for efficient photocatalytic applications.

In the present study, amorphous titanium dioxide (TiO2) nanotubes were synthesized by one-step anodization technique and subsequently annealed in different environments to investigate the effect of annealing atmospheres on the formation of different crystalline phases. X-ray Diffraction (XRD) patterns clearly showed the presence of anatase TiO2 phase with various crystallite sizes. The samples annealed in oxygen and air atmospheres at 500 degrees C showed a dominant anatase phase and a small amount of rutile phase, on the other hand, the samples annealed in nitrogen and argon atmospheres and in a vacuum at 500 degrees C contained the anatase phase only. XPS analysis of the samples showed a broadening in the binding energy curves with respect to variation in annealing atmosphere, confirming the variation in surface defects, which in turn affect photocatalytic degradation. The vacuum-annealed sample showed superior photocatalytic degradation efficiency as it had relatively higher pseudo-first order rate constants (k) of 0.009/min.

Synthesis of copper hydroxide and oxide nanostructures via anodization technique for efficient photocatalytic application.

We have demonstrated a facile protocol for synthesizing CuO and Cu2O mixed-phase nanostructures by anodization of copper hydroxide (Cu(OH)2) nanoneedles and their heat treatment in different atmospheres, which affect photocatalytic degradation efficiency. The oxygen annealed sample had relatively small (100 nm) lamellar, spherical nanoparticulate structures on the substrate surface, which showed better photocatalytic degradation of reactive black 5 dye resulting from the appropriate morphology and phase formation, compared to the samples annealed in different atmospheres and vacuum. The pseudo first-order rate constant (k) of the oxygen annealed sample was 0.0054/min, which was relatively high due to the formation of a CuO-Cu2O heterojunction with matching band potentials. Air, nitrogen, argon and vacuum annealing resulted in bigger particles and different morphologies, which led to pseudo first-order rate constants (k) of 0.0032/min (air-annealed); 0.0021/min (N2-annealed); 0.0033/min (Ar-annealed); and 0.0027/min (vacuum-annealed), which resulted in poor photocatalytic degradation of the reactive black 5 dye.

Herbal Mixture of Carthamus tinctorius L. Seed and Taraxacum coreanum Attenuates Amyloid Beta-Induced Cognitive Dysfunction In Vivo.

Deposition of amyloid-beta (Aß) in the aging brain has been often observed and is thought to be a pathological feature of Alzheimer's disease. The use of natural products for disease prevention and treatment is gaining attention worldwide. Carthamus tinctorius L. seed and Taraxacum coreanum have been used as traditional medicines in Asian countries, where they have been reported to exert anti-inflammatory and anti-oxidative effects. It has been demonstrated that the combination of C. tinctorius L. seed and T. coreanum has an effect on cognitive enhancement, indicating a ratio of 5:5 synergistically enhancing learning and memory abilities in comparison with a single treatment. Here, we aimed to investigate the protective effect of C. tinctorius L. seed and T. coreanum mixture (CT) at different concentrations on cognition in Aß25-35-infused mice. CT-administered mice showed significant cognitive improvement in the T-maze, novel object recognition, and Morris water maze tests. Moreover, amyloidogenesis-related proteins, such as ß-secretase and γ-secretase, were detected and their protein levels decreased after treatment with CT. Our study shows that CT attenuates cognitive dysfunction by improving learning and memory capability and regulating Aß-related proteins in Aß25-35-injected mice. These findings suggest that CT might be a candidate for functional food on cognitive improvement.

Combined Effects of Chronic Kidney Disease and Nonalcoholic Fatty Liver Disease on the Risk of Cardiovascular Disease in Patients with Diabetes.

BACKGROUND: We investigated the combined effect of chronic kidney disease (CKD) and nonalcoholic fatty liver disease (NAFLD) on the risk of cardiovascular disease (CVD) in patients with type 2 diabetes. METHODS: Data were obtained from the Korean National Health Insurance Service. Patients with diabetes who participated in health screenings from 2009 to 2011 were included. The fatty liver index (FLI) was used as a surrogate marker for NAFLD. RESULTS: During a mean follow-up of 6.9 years, 40,863 incidents of myocardial infarction (MI), 58,427 strokes, and 116,977 deaths were reported in 1,607,232 patients with type 2 diabetes. After adjusting for conventional risk factors, patients with CKD and NAFLD showed the highest risk of MI and stroke (hazard ratio (HR) = 1.49; 95% confidence interval (CI): 1.42-1.57 and stroke, HR = 1.48; 95% CI: 1.41-1.54, respectively) compared with those without either CKD or NAFLD. Both overall and cardiovascular mortality were highest in the CKD/NAFLD group compared with other groups (HR = 2.00; 95% CI: 1.94-2.06, and HR = 2.20; 95% CI: 2.07-2.35, respectively). Advanced liver fibrosis was significantly associated with an increased risk of CVD in patients with NAFLD. Proteinuria was significantly associated with incidence of CVD events in patients with CKD. CONCLUSIONS: The combination of CKD and NAFLD was associated with an increased risk of CVD and mortality in patients with type 2 diabetes. Close monitoring and appropriate management of CKD and NAFLD may be warranted to prevent CVD in these patients.

Inhibition of hypoxia-inducible carbonic anhydrase-IX enhances hexokinase II inhibitor-induced hepatocellular carcinoma cell apoptosis.

AIM: The hypoxic condition within large or infiltrative hypovascular tumors produces intracellular acidification, which could activate many signaling pathways and augment cancer cell growth and invasion. Carbonic anhydrase-IX (CA-IX) is an enzyme lowering pH. This study is to examine whether hypoxia induces CA-IX in hepatocellular carcinoma (HCC) cells, and to evaluate its clinical implication in HCC patients. METHODS: Human HCC cell lines (Huh-7 and HepG2 cells) were used, and cell growth was assessed using MTS assay. CA-IX expression and apoptotic/kinase signaling were evaluated using immunoblotting. The cells were transfected with CA-IX-specific siRNA, or treated with its inhibitor 4-(2-aminoethyl) benzenesulfonamide (CAI#1), and/or the hexokinase II inhibitor, 3-bromopyruvate (3-BP). A clinic pathological analysis of 69 patients who underwent an HCC resection was performed using a tissue array. RESULTS: Incubation of HCC cells under hypoxia (1% O2, 5% CO2, 94% N2) for 36 h significantly increased CA-IX expression level. CAI#1 (400 µmol/L) or CA-IX siRNA (100 µmol/L) did not influence HCC cell growth and induce apoptosis. However, CAI#1 or CA-IX siRNA at these concentrations enhanced the apoptosis induced by 3-BP (100 µmol/L). This enhancement was attributed to increased ER stress and JNK activation, as compared with 3-BP alone. Furthermore, a clinic pathological analysis of 69 HCC patients revealed that tumor CA-IX intensity was inversely related to E-cadherin intensity. CONCLUSION: Inhibition of hypoxia-induced CA-IX enhances hexokinase II inhibitor-induced HCC apoptosis. Furthermore, CA-IX expression profiles may have prognostic implications in HCC patients. Thus, the inhibition of CA-IX, in combination with a hexokinase II inhibitor, may be therapeutically useful in patients with HCCs that are aggressively growing in a hypoxic environment.

Predictors of HBsAg seroclearance in HBeAg-negative chronic hepatitis B patients.

BACKGROUND AND AIMS: In chronic hepatitis B virus (HBV) infections, hepatitis B surface antigen (HBsAg) seroclearance reportedly occurs at a rate of 0.50-2.26%. Several factors were suggested to be associated with seroclearance including age and HBeAg negativity. However, there are few studies evaluating whether HBV DNA levels are an independent predictor of HBsAg seroclearance. The aim of this study was to evaluate the annual HBsAg seroclearance rate and its predictors including serum HBV DNA levels in HBeAg-negative chronic hepatitis B patients. METHODS: We included 880 HBeAg-negative chronic hepatitis B patients who underwent an evaluation of baseline serum HBV DNA levels. We reviewed the electronic charts for baseline clinical, biochemical, and virological factors. RESULTS: The median age was 50 years, and the follow-up duration was 31 months. The annual rate for the HBsAg seroclearance was 1.8%. In multivariable analysis, the probability of HBsAg seroclearance was positively associated with old age (p = 0.046), with a sustained inactive phase rather than the occurrence of HBeAg-negative hepatitis (p = 0.041), and with an initial HBV DNA level <2,000 IU/ml (p = 0.001). CONCLUSION: The annual rate for the HBsAg seroclearance in HBeAg-negative chronic hepatitis B patients was 1.8%. Old age, sustained inactive phase, and low levels of HBV DNA were independent predictors of HBsAg seroclearance.

Factors Influencing Compassion Fatigue among Hospice and Palliative Care Unit Nurses.

Purpose: This descriptive study aimed to explore nursing workplace spirituality, end-of-life care stress, and resilience as factors influencing compassion fatigue among nurses working in hospice and palliative care units. Methods: Data were collected using a self-report questionnaire completed by 146 nurses at 14 hospice and palliative care institutions across South Korea who had worked in a hospice and palliative care institution for at least 6 months and had experience providing end-of-life care. Data were collected from February 25, 2019 to April 12, 2019, and analyzed using SPSS for Windows version 18.0. As appropriate, descriptive statistics, the t-test, analysis of variance, the Scheffé test, Pearson correlation coefficients, and stepwise multiple regression were used. Results: The survey results showed that factors influencing compassion fatigue were resilience, subjective health status, current satisfaction with the hospice ward, and end-of-life care stress. Higher levels of resilience, a subjective health status of "healthy", high levels of current satisfaction with the hospice ward, and lower levels of end-of-life care stress were associated with lower levels of compassion fatigue, explaining 42.9% of the total variance. Conclusion: The results of this study suggest that resilience is an important factor mitigating compassion fatigue among nurses at hospice and palliative care institutions. Therefore, intervention programs should be developed to reduce compassion fatigue.

Reelin Alleviates Mesenchymal Stem Cell Senescence and Reduces Pathological α-Synuclein Expression in an In Vitro Model of Parkinson's Disease.

Parkinson's disease (PD) is one of the most common neurodegenerative diseases. The mechanisms underlying PD remain to be fully elucidated, and research into treatments for this condition is ongoing. Recent advances in genetic research have shed light on the mechanisms underlying PD. In this study, we used PD and control mesenchymal stem cells (MSCs) obtained from adipose tissues to confirm the differences between groups at the cellular and molecular levels. The results revealed that in PD MSCs, cell viability was clearly lower, and the rate of cell senescence was higher compared to the controls. Next, to compare the gene expression in PD and control cells, transcriptome analysis was performed. Genes in pathways, including extracellular matrix (ECM) receptor interaction, P53 signaling, and focal adhesion, were down-regulated in PD. Among genes related to ECM receptor interaction, RELN gene expression was markedly decreased in PD cells; however, after being treated with recombinant Reelin protein, a significant increase in cell viability and a decrease in α-Synuclein aggregation and cell senescence were observed. In conclusion, Reelin affects PD by positively influencing the cell characteristics. Our findings will facilitate research into new treatments for PD.

Fatty Liver Index for Predicting Nonalcoholic Fatty Liver Disease in an Asymptomatic Korean Population.

Nonalcoholic fatty liver disease (NAFLD) is increasing worldwide, highlighting the importance of early and accurate detection and the appropriate management of NAFLD. However, ultrasonography (US) is not included in many mass screening programs, and people have limited access to it. The aim of this study is to validate the fatty liver index (FLI) and investigate the optimal cutoff value for predicting NAFLD in an asymptomatic population. We conducted a retrospective cohort study in Korea. All subjects who underwent health checkup exams, including abdominal US, controlled attenuation parameter (CAP) and blood testing, were enrolled. Analyses of the area under the receiver operating characteristic curve (AUROC) were used to evaluate the diagnostic accuracy and to calculate the optimal FLI cutoff for US-NAFLD. Among the 4009 subjects (mean age 54.9 years, 83.5% male), the prevalence of US-diagnosed NAFLD and CAP-defined hepatic steatosis was 61.4% and 55.4%. The previously used cutoff of FLI = 60 showed poor performance in predicting US-diagnosed NAFLD, with an AUROC of 0.63 (0.62-0.64), and CAP-defined NAFLD, with an AUROC 0.63 (0.62-0.64). The optimal FLI cutoff values to discriminate fatty liver detected by US were 29 for the entire population, with an AUROC of 0.82 (0.81-0.84). The sex-specific values were 31 for males and 18 for females (sensitivity 72.8% and 73.4%; specificity 74.2% and 85.0%, respectively). The FLI cutoff for US-diagnosed NAFLD can be set as 29 for the entire Korean population. Considering the sex dimorphism in NAFLD, different cutoff values are suggested to predict US-diagnosed NAFLD. These results may be helpful in the accurate non-invasive diagnosis of NAFLD.

The Change in Metabolic Syndrome Status and the Risk of Nonviral Liver Cirrhosis.

BACKGROUND: Nonalcoholic fatty liver disease is considered to be the hepatic component of metabolic syndrome (MetS). However, the association between changes in MetS status and the risk of liver cirrhosis (LC) has not been investigated to date. This study assessed the association between changes in MetS and subsequent nonviral LC development. METHODS: Data were obtained from the Korean National Health Insurance Service. Individuals who participated in health screenings from both 2009 to 2010 and 2011 to 2012 were included. The primary outcome was LC development according to the static and dynamic MetS status. Subjects were stratified into four groups according to the change in MetS status observed from the two-year interval screening (2009-2011). Cox regression analysis was used to examine the hazard ratios of LC. RESULTS: During a median of 7.3 years of follow-up, 24,923 incident LC cases developed among 5,975,308 individuals. After adjusting for age, sex, smoking, alcohol, regular exercise, and body mass index, the adjusted hazard ratios (95% confidence intervals) for LC development were 1.39 (1.33-1.44) for the MetS-Developed group, 1.32 (1.26-1.37) for the MetS-Recovered group, and 1.51 (1.45-1.56) for the MetS-Sustained group, relative to the MetS-Free group. Stratified analyses according to age, sex, smoking, alcohol intake, exercise, diabetes mellitus, hypertension, dyslipidemia, and chronic kidney disease showed similar results. CONCLUSIONS: Both static and dynamic MetS status are independent risk factors for LC development. The risk of LC was the highest in people with sustained MetS and was lower in the MetS-Recovered group than in the MetS-Sustained group. These results suggest that improving a person's MetS status may be helpful in preventing LC.

Biosensor system-on-a-chip including CMOS-based signal processing circuits and 64 carbon nanotube-based sensors for the detection of a neurotransmitter.

We developed a carbon nanotube (CNT)-based biosensor system-on-a-chip (SoC) for the detection of a neurotransmitter. Here, 64 CNT-based sensors were integrated with silicon-based signal processing circuits in a single chip, which was made possible by combining several technological breakthroughs such as efficient signal processing, uniform CNT networks, and biocompatible functionalization of CNT-based sensors. The chip was utilized to detect glutamate, a neurotransmitter, where ammonia, a byproduct of the enzymatic reaction of glutamate and glutamate oxidase on CNT-based sensors, modulated the conductance signals to the CNT-based sensors. This is a major technological advancement in the integration of CNT-based sensors with microelectronics, and this chip can be readily integrated with larger scale lab-on-a-chip (LoC) systems for various applications such as LoC systems for neural networks.

Purification of an Intact Human Protein Overexpressed from Its Endogenous Locus via Direct Genome Engineering.

The overproduction and purification of human proteins is a requisite of both basic and medical research. Although many recombinant human proteins have been purified, current protein production methods have several limitations; recombinant proteins are frequently truncated, fail to fold properly, and/or lack appropriate post-translational modifications. In addition, such methods require subcloning of the target gene into relevant plasmids, which can be difficult for long proteins with repeated domains. Here we devised a novel method for target protein production by introduction of a strong promoter for overexpression and an epitope tag for purification in front of the endogenous human gene, in a sense performing molecular cloning directly in the human genome, which does not require cloning of the target gene. As a proof of concept, we successfully purified intact human Reelin protein, which is lengthy (3460 amino acids) and contains repeating domains, and confirmed that it was biologically functional.

Antiviral activity of ovine interferon tau 4 against foot-and-mouth disease virus.

Foot-and-mouth disease (FMD) is an economically important disease in most parts of the world and new therapeutic agents are needed to protect the animals before vaccination can trigger the host immune response. Although several interferons have been used for their antiviral activities against Foot-and-mouth disease virus (FMDV), ovine interferon tau 4 (OvIFN-τ4), with a broad-spectrum of action, cross-species antiviral activity, and lower incidence of toxicity in comparison to other type І interferons, has not yet been evaluated for this indication. This is the first study to evaluate the antiviral activity of OvIFN-τ4 against various strains of FMDV. The effective anti-cytopathic concentration of OvIFN-τ4 and its effectiveness pre- and post-infection with FMDV were tested in vitro in LFBK cells. In vivo activity of OvIFN-τ4 was then confirmed in a mouse model of infection. OvIFN-τ4 at a concentration of 500 ng, protected mice until 5days post-FMDV challenge and provided 90% protection for 10 days following FMDV challenge. These results suggest that OvIFN-τ4 could be used as an alternative to other interferons or antiviral agents at the time of FMD outbreak.

Structural Insights into Modulation of Neurexin-Neuroligin Trans-synaptic Adhesion by MDGA1/Neuroligin-2 Complex.

Membrane-associated mucin domain-containing glycosylphosphatidylinositol anchor proteins (MDGAs) bind directly to neuroligin-1 (NL1) and neuroligin-2 (NL2), thereby respectively regulating excitatory and inhibitory synapse development. However, the mechanisms by which MDGAs modulate NL activity to specify development of the two synapse types remain unclear. Here, we determined the crystal structures of human NL2/MDGA1 Ig1-3 complex, revealing their stable 2:2 arrangement with three interaction interfaces. Cell-based assays using structure-guided, site-directed MDGA1 mutants showed that all three contact patches were required for the MDGA's negative regulation of NL2-mediated synaptogenic activity. Furthermore, MDGA1 competed with neurexins for NL2 via its Ig1 domain. The binding affinities of both MDGA1 and MDGA2 for NL1 and NL2 were similar, consistent with the structural prediction of similar binding interfaces. However, MDGA1 selectively associated with NL2, but not NL1, in vivo. These findings collectively provide structural insights into the mechanism by which MDGAs negatively modulate synapse development governed by NLs/neurexins.