Dynamic detection and reversal of myocardial ischemia using an artificially intelligent bioelectronic medicine.

Myocardial ischemia is spontaneous, frequently asymptomatic, and contributes to fatal cardiovascular consequences. Importantly, myocardial sensory networks cannot reliably detect and correct myocardial ischemia on their own. Here, we demonstrate an artificially intelligent and responsive bioelectronic medicine, where an artificial neural network (ANN) supplements myocardial sensory networks, enabling reliable detection and correction of myocardial ischemia. ANNs were first trained to decode spontaneous cardiovascular stress and myocardial ischemia with an overall accuracy of ~92%. ANN-controlled vagus nerve stimulation (VNS) significantly mitigated major physiological features of myocardial ischemia, including ST depression and arrhythmias. In contrast, open-loop VNS or ANN-controlled VNS following a caudal vagotomy essentially failed to reverse cardiovascular pathophysiology. Last, variants of ANNs were used to meet clinically relevant needs, including interpretable visualizations and unsupervised detection of emerging cardiovascular stress. Overall, these preclinical results suggest that ANNs can potentially supplement deficient myocardial sensory networks via an artificially intelligent bioelectronic medicine system.

Variability in empathic fear response among 11 inbred strains of mice.

Empathy is an important emotional process that involves the ability to recognize and share emotions with others. We have previously developed an observational fear learning (OFL) behavioral assay to measure empathic fear in mice. In the OFL task, a mouse is conditioned for context-dependent fear when it observes a conspecific demonstrator receiving aversive stimuli. In the present study, by comparing 11 different inbred mouse strains that are commonly used in the laboratory, we found that empathic fear response was highly variable between different strains. Five strains--C57BL/6J, C57BL/6NTac, 129S1/SvImJ, 129S4/SvJae and BTBR T(+) Itpr3(tf) /J--showed observational fear (OF) responses, whereas AKR/J, BALB/cByJ, C3H/HeJ, DBA/2J, FVB/NJ and NOD/ShiLtJ mice exhibited low empathic fear response. Importantly, day 2 OF memory was significantly correlated with contextual memory in the classical fear conditioning among the 11 strains. Innate differences in anxiety, locomotor activity, sociability and preference for social novelty were not significantly correlated with OFL. Interestingly, early adolescent C57BL/6J mice exhibited an increase in acquisition of OF. The level of OFL in C57BL/6J strain was not affected by sex or strains of the demonstrator. Taken together, these data strongly suggest that there are naturally occurring OFL-specific genetic variations modulating empathic fear behaviors in mice. The identification of causal genes may uncover novel genetic pathways and underlying neural mechanisms that modulate empathic fear and, ultimately, provide new targets for therapeutic intervention in human mental disorders associated with impaired empathy.

The Effects of an Evidence-based Nursing Course Using Action Learning on Undergraduate Nursing Students / 한국간호교육학회지

PURPOSE: This study was conducted to evaluate the effectiveness of an evidence-based nursing (EBN) course using action learning-based team learning in undergraduate nursing students. METHODS: A quasi-experimental pretest-posttest control group design was employed. The participants who consented were 45 second-year nursing students (22 in the experimental, 23 in the control group) from a university in G-city, Korea. The intervention included lectures, practicals, team activities and reflection on overviewing EBN, formulating clinical questions, searching the evidence, and criticizing the research articles. At the beginning and the end of the 7-week EBN course, the participants completed self-reported questionnaires. Frequencies, chi2-test, t-test, and ANCOVA with the SPSS program 18.0, were used to analyze the data. RESULTS: The experimental group showed significantly higher scores on EBN competency (F=25.80, p<001), information literacy (F=13.75, p=.001), and proactivity in problem solving (F=5.32, p=.026) than the control group. CONCLUSION: This study provides evidence that an EBN course improves undergraduate nursing students' EBN competencies, information literacy, and proactivity in problem solving. Team learning in EBN education can be an effective teaching strategy.

SYNCRIP (synaptotagmin-binding, cytoplasmic RNA-interacting protein) is a host factor involved in hepatitis C virus RNA replication.

Hepatitis C virus (HCV) RNA replication requires viral nonstructural proteins as well as cellular factors. Recently, a cellular protein, synaptotagmin-binding, cytoplasmic RNA-interacting protein (SYNCRIP), also known as NSAP1, was found to bind HCV RNA and enhance HCV IRES-dependent translation. We investigate whether this protein is also involved in the HCV RNA replication. We found that SYNCRIP was associated with detergent-resistant membrane fractions and colocalized with newly-synthesized HCV RNA. Knock-down of SYNCRIP by siRNA significantly decreased the amount of HCV RNA in the cells containing a subgenomic replicon or a full-length viral RNA. Lastly, an in vitro replication assay after immunodepletion of SYNCRIP showed that SYNCRIP was directly involved in HCV RNA replication. These findings indicate that SYNCRIP has dual functions, participating in both RNA replication and translation in HCV life cycle.

Polypyrimidine-tract-binding protein is a component of the HCV RNA replication complex and necessary for RNA synthesis.

The machinery for hepatitis C virus (HCV) RNA replication is still poorly characterized. The relationship between HCV RNA replication and translation is also not clear. We have previously shown that a cellular protein polypyrimidine-tract-binding protein (PTB) binds to HCV RNA at several different sites and modulates HCV translation in several ways. Here we show that PTB also participates in RNA replication. By bromouridine triphosphate (BrUTP) labeling and confocal microscopy of cells harboring an HCV replicon, we showed that the newly synthesized HCV RNA was localized to distinct structures in the cytoplasm, which also contain PTB. Membrane flotation analysis demonstrated that a fraction of cytoplasmic PTB was associated with a detergent-resistant membrane (DRM) structure consisting of lipid rafts, which also contained HCV nonstructural proteins and the human vesicle-associated membrane protein-associated protein (hVAP-33). PTB in the DRM was resistant to protease digestion, but became sensitive after treatment with the raft-disrupting agents. PTB in the DRM consisted of multiple isoforms and the brain-specific paralog. By using small interfering RNA (siRNA) of PTB, we showed that silencing of the endogenous PTB reduced the replication of HCV RNA replicon. In a cell-free, de novo HCV RNA synthesis system, HCV RNA synthesis was inhibited by anti-PTB antibody. These studies together indicated that PTB is a part of the HCV RNA replication complex and participates in viral RNA synthesis. Thus, PTB has dual functions in HCV life cycle, including translation and RNA replication.

Murine coronavirus requires lipid rafts for virus entry and cell-cell fusion but not for virus release.

Thorp and Gallagher first reported that depletion of cholesterol inhibited virus entry and cell-cell fusion of mouse hepatitis virus (MHV), suggesting the importance of lipid rafts in MHV replication (E. B. Thorp and T. M. Gallagher, J. Virol. 78:2682-2692, 2004). However, the MHV receptor is not present in lipid rafts, and anchoring of the MHV receptor to lipid rafts did not enhance MHV infection; thus, the mechanism of lipid rafts involvement is not clear. In this study, we defined the mechanism and extent of lipid raft involvement in MHV replication. We showed that cholesterol depletion by methyl beta-cyclodextrin or filipin did not affect virus binding but reduced virus entry. Furthermore, MHV spike protein bound to nonraftraft membrane at 4 degrees C but shifted to lipid rafts at 37 degrees C, indicating a redistribution of membrane following virus binding. Thus, the lipid raft involvement in MHV entry occurs at a step following virus binding. We also found that the viral spike protein in the plasma membrane of the infected cells was associated with lipid rafts, whereas that in the Golgi membrane, where MHV matures, was not. Moreover, the buoyant density of the virion was not changed when MHV was produced from the cholesterol-depleted cells, suggesting that MHV does not incorporate lipid rafts into the virion. These results indicate that MHV release does not involve lipid rafts. However, MHV spike protein has an inherent ability to associate with lipid rafts. Correspondingly, cell-cell fusion induced by MHV was retarded by cholesterol depletion, consistent with the association of the spike protein with lipid rafts in the plasma membrane. These findings suggest that MHV entry requires specific interactions between the spike protein and lipid rafts, probably during the virus internalization step.

Effects of a Web-based teaching method on undergraduate nursing students' learning of electrocardiography.

This study examined the effects of a Web-based teaching method (versus a traditional lecture method) on undergraduate nursing students' learning of electrocardiography (ECG). The Web-based learning program was developed by the authors and implemented for 4 weeks. The study used a pretest-posttest experimental design. A total of 105 senior nursing students were recruited at a university in Korea. Fifty-four students were assigned to an experimental group in 2002, and 51 were assigned to a control group in 2003. Knowledge about ECG among students in the Web-based group was significantly lower than that of students in the control group (p < .01). Conversely, the ability to interpret ECG recordings was significantly higher among students in the Web-based group (p < .05). No significant differences were found between the two groups in level of motivation or satisfaction with learning. The self-directed, Web-based ECG learning program appears to be effective in helping nursing students to interpret ECG recordings.

SYNCRIP, a member of the heterogeneous nuclear ribonucleoprotein family, is involved in mouse hepatitis virus RNA synthesis.

Several cellular proteins, including several heterogeneous nuclear ribonucleoproteins (hnRNPs), have been shown to function as regulatory factors for mouse hepatitis virus (MHV) RNA synthesis as a result of their binding to the 5' and 3' untranslated regions (UTRs) of the viral RNA. Here, we identified another cellular protein, p70, which has been shown by UV cross-linking to bind both the positive- and negative-strand UTRs of MHV RNA specifically. We purified p70 with a a one-step RNA affinity purification procedure with the biotin-labeled 5'-UTR. Matrix-assisted laser desorption ionization (MALDI)-mass spectrometry identified it as synaptotagmin-binding cytoplasmic RNA-interacting protein (SYNCRIP). SYNCRIP is a member of the hnRNP family and localizes largely in the cytoplasm. The p70 was cross-linked to the MHV positive- or negative-strand UTR in vitro and in vivo. The bacterially expressed SYNCRIP was also able to bind to the 5'-UTR of both strands. The SYNCRIP-binding site was mapped to the leader sequence of the 5'-UTR, requiring the UCUAA repeat sequence. To investigate the functional significance of SYNCRIP in MHV replication, we expressed a full-length or a C-terminally truncated form of SYNCRIP in mammalian cells expressing the MHV receptor. The overexpression of either form of SYNCRIP inhibited syncytium formation induced by MHV infection. Furthermore, downregulation of the endogenous SYNCRIP with a specific short interfering RNA delayed MHV RNA synthesis; in contrast, overexpression or downregulation of SYNCRIP did not affect MHV translation. These results suggest that SYNCRIP may be directly involved in MHV RNA replication as a positive regulator. This study identified an additional cellular hnRNP as an MHV RNA-binding protein potentially involved in viral RNA synthesis.

Polypyrimidine-tract-binding protein affects transcription but not translation of mouse hepatitis virus RNA.

Polypyrimidine-tract-binding protein (PTB) has been shown to bind specifically to the 5' ends of mouse hepatitis virus (MHV) RNA and its complementary strand. To further characterize the function of PTB in MHV replication, we generated dominant-negative mutant cell lines that express a full-length PTB or a truncated form of PTB, which includes only the N-terminal half of the protein, retaining its protein-dimerization domain. The truncated form of PTB was localized in the cytoplasm, whereas the full-length PTB was present mainly in the nucleus. The truncated form can interact with the full-length PTB in vitro. We observed that both the full-length and the truncated PTB, when overexpressed, functioned in a dominant-negative manner in MHV replication. However, the truncated form exhibited more severe effects on syncytia formation, virus production, and synthesis of viral RNA and viral proteins. To clarify the precise function of PTB in MHV replication, we dissociated the processes of viral transcription from translation by transfecting different types of MHV defective-interfering (DI) RNA that contain various reporter genes into these stable cell lines. Transcription of the DI RNA during MHV infection was greatly inhibited in these cell lines, indicating that PTB modulates MHV transcription. In contrast, translation of the DI RNA was not affected by PTB depletion in in vitro translation in rabbit reticulocyte lysate or by PTB overexpression in in vivo translation experiments in MHV-infected cells. Given that PTB interacts with the viral N protein, which is one of the components of the MHV replication complex, PTB may exert its function on viral replication/transcription by association with viral RNA as well as other viral and cellular factors in the replication complex.

Inhibition of prostalglandin E2 production by 2'-hydoxychalcone derivatives and the mechanism of action.

The effects of 14 synthetic 2'-hydroxychalcone derivatives on prostaglandin E2 (PGE2) production in rat peritoneal macrophages stimulated by the protein kinase C activator, 12-O-tetradecanoylphorbol 13-acetate (TPA), were examined to clarify the structure-activity relationship. 2',4-Dihydroxy-4'-methoxychalcone (compound 3), 2',4-dihydroxy-6'-methoxychalcone (compound 8) and 2'-hydroxy-4'-methoxychalcone (compound 9) suppressed PGE2 production more potently than the other compounds. The IC50 (50% Inhibitory concentration) value for compounds 3, 8 and 9 was calculated to be 3 microM. The activity of cyclooxygenase (COX)-1 was inhibited slightly by compound 9, but that of COX-2 was not inhibited. At concentrations that inhibited the production of PGE2, compound 9 had no effect on the release of radioactivity from [3H]arachidonic acid-labelled macrophages stimulated by TPA. Western-blot analysis revealed that the induction of COX-2 protein by TPA was inhibited by compound 9 in parallel with the inhibition of PGE2 production. Compounds 3 and 8 had similar effects. These findings suggest that 4'-methoxyl and 6'-methoxyl groups are required for the expression of more potent inhibitory activity against PGE2 production, and that the inhibition of PGE2 production by these 2'-hydroxychalcone derivatives is due to the inhibition of TPA-induced COX-2 protein expression.

Lack of the burst firing of thalamocortical relay neurons and resistance to absence seizures in mice lacking alpha(1G) T-type Ca(2+) channels.

T-type Ca(2+) currents have been proposed to be involved in the genesis of spike-and-wave discharges, a sign of absence seizures, but direct evidence in vivo to support this hypothesis has been lacking. To address this question, we generated a null mutation of the alpha(1G) subunit of T-type Ca(2+) channels. The thalamocortical relay neurons of the alpha(1G)-deficient mice lacked the burst mode firing of action potentials, whereas they showed the normal pattern of tonic mode firing. The alpha(1G)-deficient thalamus was specifically resistant to the generation of spike-and-wave discharges in response to GABA(B) receptor activation. Thus, the modulation of the intrinsic firing pattern mediated by alpha(1G) T-type Ca(2+) channels plays a critical role in the genesis of absence seizures in the thalamocortical pathway.

Synthesis of flavonoids and their effects on aldose reductase and sorbitol accumulation in streptozotocin-induced diabetic rat tissues.

Aldose reductase, the key enzyme of the polyol pathway, and oxidative stress are known to play important roles in the complications of diabetes. A drug with potent inhibition of aldose reductase and oxidative stress, therefore, would be a most promising drug for the prevention of diabetic complications. The purpose of this study was to develop new compounds with these dual-effects through synthesis of chalcone derivatives and by examining the structure-activity relationships on the inhibition of rat lens aldose reductase as well as on antioxidant effects. A series of 35 flavonoid derivatives were synthesized by Winget's condensation, oxidation, and reduction of appropriate acetophenones with appropriate benzaldehydes. The inhibitory activity of these derivatives on rat lens aldose reductase and their antioxidant effects, measured using Cu2+ chelation and radical scavenging activities on 1,1-diphenyl-picrylhydrazyl in-vitro, were evaluated. Their effect on sorbitol accumulation in the red blood cells, lenses and sciatic nerves of streptozotocin-induced diabetic rats was also estimated. Among the new flavonoid derivatives synthesized, those with the 2',4'-dihydroxyl groups in the A ring such as 2,4,2',4'-tetrahydroxychalcone (22), 2,2',4'-trihydroxychalcone (11), 2',4'-dihydroxy-2,4-dimethylchalcone (21) and 3,4,2',4'-tetrahydroxychalcone (18) were found to possess the highest rat lens aldose reductase inhibitory activity in-vitro, their IC50 values (concentration of inhibitors giving 50% inhibition of enzyme activity) being 1.6 x 10(-7), 3.8 x 10(-7), 4.0 x 10(-7) and 4.6 x 10(-7) M, respectively. All of the chalcones tested except 3, 18, 23 with o-dihydroxy or hydroquinone moiety showed a weak free radical scavenging activity. In the in-vivo experiments, however, compound 18 with o-dihydroxy moiety in the B ring showed the strongest inhibitory activity in the accumulation of sorbitol in the tissues. It also showed the strongest activity in transition metal chelation and free radical scavenging activity. Of the 35 4,2'-dihydroxyl and 2',4'-dihydroxyl derivatives of flavonoid synthesized, including chalcone, flavone, flavanone, flavonol and dihydrochalcone, some chalcone derivatives synthesized were found to possess aldose reductase inhibition and antioxidant activities in-vitro as well as inhibition in the accumulation of sorbitol in the tissues in-vivo. 3,4,2',4'-Tetrahydroxychalcone (18, butein) was the most promising compound for the prevention or treatment of diabetic complications.

Inhibitory effects of 2'-hydroxychalcones on rat lens aldose reductase and rat platelet aggregation.

Inhibitory effects of synthetic 2'-hydroxychalcone derivatives on rat lens aldose reductase (RLAR) and on platelet aggregation were investigated for the prevention or the treatment of chronic diabetic complications. 5'-chloro-4,2'-dihydroxychalcone (8) and 5'-chloro-3,2'-dihydroxychalcone (27) exhibited a potent inhibitory effect on rat platelet aggregation induced by ADP (IC50=0.10 and 0.06 mg/ml, respectively) and collagen (IC50=44 and 16 microg/ml, respectively) but showed relatively weak inhibitory activities on RLAR.