Development of the opioid self-management scale for advanced Cancer patients with pain and examination of its validity and reliability.

BACKGROUND: Approximately 60% of outpatients with advanced cancer experience pain; therefore, self-management of opioid use is important for appropriate pain relief. To date, no studies have clearly described the concept of opioid self-management or assessed the factors involved, including the improvement of self-management abilities. This study developed, and evaluated the validity and reliability of an opioid self-management scale for advanced cancer patients with pain (OSSA). Opioid self-management in advanced cancer patients with pain was defined as the management of opioid medication performed by patients with advanced cancer to relieve cancer pain on their own. METHODS: Three phases were required for validation and reliability of the OSSA: 1) testing content validity, 2) testing face validity, and 3) testing construct validity, concurrent validity and reliability. RESULTS: After a three-phase process, the OSSA consisted of 33 items on six subscales. The structural equation modeling was such that the χ2 value was 709.8 (p < 0.001, df = 467), goodness-of-fit index was 0.78, adjusted goodness-of-fit index was 0.73, root mean squares of approximation was 0.063, and comparative fit index was 0.92. The Pearson correlation coefficients between the total OSSA score and the 24-hour average pain or pain relief over 24 hours were - 0.21 (p < 0.05) and 0.26 (p < 0.01), respectively. Cronbach's α was 0.93. The intraclass correlation coefficient range was 0.59-0.90. CONCLUSION: The findings of this study show that the OSSA has acceptable validity and reliability, and that better self-management leads to greater pain relief. The OSSA can be considered effective for use in research, but shortened version should be prepared for realistic and practical clinical use.

The E3 ubiquitin ligase MIB2 enhances inflammation by degrading the deubiquitinating enzyme CYLD.

The tumor suppressor CYLD is a deubiquitinating enzyme that suppresses polyubiquitin-dependent signaling pathways, including the proinflammatory and cell growth-promoting NF-κB pathway. Missense mutations in the CYLD gene are present in individuals with syndromes such as multiple familial trichoepithelioma (MFT), but the pathogenic roles of these mutations remain unclear. Recent studies have shown that CYLD interacts with a RING finger domain protein, mind bomb homologue 2 (MIB2), in the regulation of NOTCH signaling. However, whether MIB2 is an E3 ubiquitin ligase that acts on CYLD is unknown. Here, using the cell-free-based AlphaScreen and pulldown assays to detect protein-protein interactions, along with immunofluorescence assays and murine Mib2 knockout cells and animals, we demonstrate that MIB2 promotes proteasomal degradation of CYLD and enhances NF-κB signaling. Of note, arthritic inflammation was suppressed in Mib2-deficient mice. We further observed that the ankyrin repeat in MIB2 interacts with the third CAP domain in CYLD and that MIB2 catalyzes Lys-48-linked polyubiquitination of CYLD at Lys-338 and Lys-530. MIB2-dependent CYLD degradation activated NF-κB signaling via tumor necrosis factor alpha (TNFα) stimulation and the linear ubiquitination assembly complex (LUBAC). Mib2-knockout mice had reduced serum interleukin-6 (IL-6) and exhibited suppressed inflammatory responses in the K/BxN serum-transfer arthritis model. Interestingly, MIB2 significantly enhanced the degradation of a CYLDP904L variant identified in an individual with MFT, although the molecular pathogenesis of the disease was not clarified here. Together, these results suggest that MIB2 enhances NF-κB signaling in inflammation by promoting the ubiquitin-dependent degradation of CYLD.

Characterization of RyDEN (C19orf66) as an Interferon-Stimulated Cellular Inhibitor against Dengue Virus Replication.

Dengue virus (DENV) is one of the most important arthropod-borne pathogens that cause life-threatening diseases in humans. However, no vaccine or specific antiviral is available for dengue. As seen in other RNA viruses, the innate immune system plays a key role in controlling DENV infection and disease outcome. Although the interferon (IFN) response, which is central to host protective immunity, has been reported to limit DENV replication, the molecular details of how DENV infection is modulated by IFN treatment are elusive. In this study, by employing a gain-of-function screen using a type I IFN-treated cell-derived cDNA library, we identified a previously uncharacterized gene, C19orf66, as an IFN-stimulated gene (ISG) that inhibits DENV replication, which we named Repressor of yield of DENV (RyDEN). Overexpression and gene knockdown experiments revealed that expression of RyDEN confers resistance to all serotypes of DENV in human cells. RyDEN expression also limited the replication of hepatitis C virus, Kunjin virus, Chikungunya virus, herpes simplex virus type 1, and human adenovirus. Importantly, RyDEN was considered to be a crucial effector molecule in the IFN-mediated anti-DENV response. When affinity purification-mass spectrometry analysis was performed, RyDEN was revealed to form a complex with cellular mRNA-binding proteins, poly(A)-binding protein cytoplasmic 1 (PABPC1), and La motif-related protein 1 (LARP1). Interestingly, PABPC1 and LARP1 were found to be positive modulators of DENV replication. Since RyDEN influenced intracellular events on DENV replication and, suppression of protein synthesis from DENV-based reporter construct RNA was also observed in RyDEN-expressing cells, our data suggest that RyDEN is likely to interfere with the translation of DENV via interaction with viral RNA and cellular mRNA-binding proteins, resulting in the inhibition of virus replication in infected cells.

Identification of RFPL3 protein as a novel E3 ubiquitin ligase modulating the integration activity of human immunodeficiency virus, type 1 preintegration complex using a microtiter plate-based assay.

Integration, one of the hallmarks of retrovirus replication, is mediated by a nucleoprotein complex called the preintegration complex (PIC), in which viral DNA is associated with many protein components that are required for completion of the early phase of infection. A striking feature of the PIC is its powerful integration activity in vitro. The PICs from a freshly isolated cytoplasmic extract of infected cells are able to insert viral DNA into exogenously added target DNA in vitro. Therefore, a PIC-based in vitro assay is a reliable system for assessing protein factors influencing retroviral integration. In this study, we applied a microtiter plate-based in vitro assay to a screening study using a protein library that was produced by the wheat germ cell-free protein synthesis system. Using a library of human E3 ubiquitin ligases, we identified RFPL3 as a potential stimulator of human immunodeficiency virus, type 1 (HIV-1) PIC integration activity in vitro. This enhancement of PIC activity by RFPL3 was likely to be attributed to its N-terminal RING domain. To further understand the functional role of RFPL3 in HIV infection, we created a human cell line overexpressing RFPL3. Immunoprecipitation analysis revealed that RFPL3 was associated with the human immunodeficiency virus, type 1 PICs in infected cells. More importantly, single-round HIV-1 infection was enhanced significantly by RFPL3 expression. Our proteomic approach displays an advantage in the identification of new cellular proteins affecting the integration activity of the PIC and, therefore, contributes to the understanding of functional interaction between retroviral integration complexes and host factors.

Effectiveness of a telenursing intervention program in reducing exacerbations in patients with chronic respiratory failure receiving noninvasive positive pressure ventilation: A randomized controlled trial.

Telenursing for patients with chronic respiratory failure receiving noninvasive positive pressure ventilation (NPPV) is an important aid in reducing exacerbations; however, there is insufficient evidence. This randomized controlled trial investigated the effectiveness of a telenursing intervention program in reducing exacerbations in patients with chronic respiratory failure receiving NPPV at home. We included patients receiving NPPV at home who could handle a tablet device. The intervention group (n = 15) underwent an information and communications technology-based telenursing intervention program in addition to usual care; the control group (n = 16) received the usual care only. The telenursing intervention program comprised telemonitoring and health counseling sessions via videophone. The intervention was evaluated once at enrollment and after 3 months. The primary endpoints were the number of unscheduled outpatient visits, hospitalizations, and hospital days. The secondary endpoints included the St. George's Respiratory Questionnaire (SGRQ) score, Euro QOL 5 Dimension score, Self-Care Agency Questionnaire (SCAQ) score, pulmonary function tests, and 6-min walking distance. We used the Mann-Whitney U test for our analysis. We found no significant differences between the intervention and control groups at enrollment. Then, the differences between the endpoints at baseline and 3 months after enrollment were calculated and used to compare both groups. At follow-up, the number of routine outpatient visits for acute exacerbations (p = .045), the number of hospitalizations (p = .037), the number of hospital days (p = .031), SGRQ (p = .039) score, and SCAQ (p = .034) score were significantly different. The increase in the number of unscheduled outpatient visits in the intervention group during follow-up was attributed to acute exacerbations and a significant decrease in the number of hospitalizations and hospital days. Hence, the telenursing intervention program may be effective in reducing exacerbations in patients with chronic respiratory failure receiving NPPV at home. Trial registration: UMIN Clinical Trials Registry (UMIN-CTR) UMIN000027657.

Development of the quality of life of family caregivers of advanced cancer patients scale in Japan: Psychometric properties.

Objective: We aimed to develop a new scale for use in Japan, called the "Quality of Life of Family Caregivers of Advanced Cancer Patients Scale (QFCS)" and to examine its psychometric properties. Methods: A draft scale was extracted based on qualitative inductive and deductive analyses, and its content validity and surface validity were investigated. Its psychometric properties were examined. Results: The QFCS consists of 30 items comprising four factors. Cronbach's α was 0.92 and the intraclass correlation coefficient was 0.90. Correlation coefficients between the total QFCS score and eight subscale scores of the revised Medical Outcomes Study 12-Item Short Form Survey Instrument were rs â€‹= â€‹0.22-0.65 (P â€‹< â€‹0.01-0.05). The Physical Component Summary was r â€‹= â€‹0.29 (P â€‹< â€‹0.01), and the Mental Component Summary was r â€‹= â€‹0.67 (P â€‹< â€‹0.01). Correlation coefficients between the QFCS total score and four subscale scores of the Caregiver Quality of Life Index-Cancer (CQOLC) were r â€‹= â€‹0.27-0.59 (P â€‹<0 â€‹.01) and the CQOLC total score was r â€‹= â€‹0.65 (P â€‹<0 â€‹.01). Conclusions: Our results suggest that the QFCS exhibited acceptable psychometric properties in measuring the quality of life of family caregivers of patients with advanced cancer. Future research is needed to evaluate the effectiveness and quality of family support using the QFCS.

N95 masks worn to protect against COVID-19 prevented tuberculosis exposure in healthcare workers.

Since the start of the COVID-19 pandemic, the healthcare workers in our institution have been equipped with N95 masks when performing aerosol-generating procedures, as these are associated with an increased risk of infection. We present a case in which using an N95 mask prevented tuberculosis (TB) exposure among healthcare workers administering prehospital care in rapid response vehicles. Even after the resolution of the COVID-19 pandemic in the future, wearing N95 masks among healthcare workers is recommended to protect against pathogens, including TB, when performing aerosol-producing procedures or prehospital activities for patients suspected of having respiratory diseases.

Development of an automated system to measure ion channel currents using a surface-modified gold probe.

Artificial lipid bilayer single-channel recording technique has been employed to determine the biophysical and pharmacological properties of various ion channels. However, its measurement efficiency is very low, as it requires two time-consuming processes: preparation of lipid bilayer membranes and incorporation of ion channels into the membranes. In order to address these problems, we previously developed a technique based on hydrophilically modified gold probes on which are immobilized ion channels that can be promptly incorporated into the bilayer membrane at the same time as the membrane is formed on the probes' hydrophilic area. Here, we improved further this technique by optimizing the gold probe and developed an automated channel current measurement system. We found that use of probes with rounded tips enhanced the efficiency of channel current measurements, and introducing a hydrophobic area on the probe surface, beside the hydrophilic one, further increased measurement efficiency by boosting membrane stability. Moreover, we developed an automated measurement system using the optimized probes; it enabled us to automatically measure channel currents and analyze the effects of a blocker on channel activity. Our study will contribute to the development of high-throughput devices to identify drug candidates affecting ion channel activity.

Low forced expiratory volume in one second is associated with the history of acute coronary syndrome in patients with organic coronary stenosis.

BACKGROUND: Chronic obstructive pulmonary disease often coexists with cardiovascular diseases and airflow limitation has been known as a risk of cardiovascular death. However, the association between airflow limitation and the history of acute coronary syndrome (ACS) in patients with coronary stenosis remains to be determined. METHODS: Study subjects were 271 consecutive patients (age: 70.6±9.5 years, sex: 200 males) who underwent coronary angiography and in whom organic coronary stenosis was detected. We collected spirometric data from those patients and investigated the association of the pulmonary function and the history of ACS. We also compared the prevalence of airflow limitation of the present subjects with Japanese epidemiological data that had been previously published. RESULTS: Multivariate analysis with multiple logistic regression analysis showed that the reduced forced expiratory volume in one second (FEV1.0) less than 80% of predicted value was significantly associated with a history of ACS (odds ratio: 2.81, 95% CI: 1.27-6.20, p<0.02) independently of age, sex, body mass index, and classic coronary risk factors including smoking habit, diabetes mellitus, hypertension, and dyslipidemia. Furthermore, the airflow limitation was more prevalent in the present subjects than in the Japanese general population (25.8% vs. 10.9%, p<0.05). CONCLUSIONS: Reduced FEV1.0 is associated with a history of ACS in patients with coronary arterial stenosis irrespective of any coronary risk factors. Airflow limitation is more prevalent in patients with coronary stenosis than in the general population.

Establishment of a Wheat Cell-Free Synthesized Protein Array Containing 250 Human and Mouse E3 Ubiquitin Ligases to Identify Novel Interaction between E3 Ligases and Substrate Proteins.

Ubiquitination is a key post-translational modification in the regulation of numerous biological processes in eukaryotes. The primary roles of ubiquitination are thought to be the triggering of protein degradation and the regulation of signal transduction. During protein ubiquitination, substrate specificity is mainly determined by E3 ubiquitin ligase (E3). Although more than 600 genes in the human genome encode E3, the E3s of many target proteins remain unidentified owing to E3 diversity and the instability of ubiquitinated proteins in cell. We demonstrate herein a novel biochemical analysis for the identification of E3s targeting specific proteins. Using wheat cell-free protein synthesis system, a protein array containing 227 human and 23 mouse recombinant E3s was synthesized. To establish the high-throughput binding assay using AlphaScreen technology, we selected MDM2 and p53 as the model combination of E3 and its target protein. The AlphaScreen assay specifically detected the binding of p53 and MDM2 in a crude translation mixture. Then, a comprehensive binding assay using the E3 protein array was performed. Eleven of the E3s showed high binding activity, including four previously reported E3s (e.g., MDM2, MDM4, and WWP1) targeting p53. This result demonstrated the reliability of the assay. Another interactors, RNF6 and DZIP3-which there have been no report to bind p53-were found to ubiquitinate p53 in vitro. Further analysis showed that RNF6 decreased the amount of p53 in H1299 cells in E3 activity-dependent manner. These results suggest the possibility that the RNF6 ubiquitinates and degrades p53 in cells. The novel in vitro screening system established herein is a powerful tool for finding novel E3s of a target protein.

Should increasing the dose or adding an AT1 receptor blocker follow a relatively low dose of ACE inhibitor initiated in acute myocardial infarction?

OBJECTIVE: Based on currently available clinical evidence, we should use high-dose angiotensin converting enzyme inhibitor (ACE-I) for patients with acute myocardial infarction (MI), initiating it at incremental doses to avoid excessive hypotension. Recent animal studies with acute MI models failed to demonstrate the superiority of the combination therapy of ACE-I and angiotensin receptor blocker (ARB) to high-dose ACE-I treatment with comparable blood pressure reductions, which however might be attributed to the initiation of the targeted doses from the beginning. The aim of this study was to compare the effect of increasing the dose of ACE-I with that of adding ARB following a relatively low dose of ACE-I on the survival and left ventricular (LV) remodeling after MI. METHODS: Rats underwent left coronary artery ligation and were treated with either ACE-I temocapril (5 mg/kg/day) or vehicle for 2 weeks, which was initiated 3 days after the surgery. The rats treated with temocapril were further randomly assigned to receive either high-dose temocapril (10 mg/kg/day) or combination therapy (temocapril 5 mg/kg/day+olmesartan 2.5 mg/kg/day), which was continued for another 6 weeks. RESULTS: Both treatments similarly reduced the blood pressure, improved survival and ameliorated LV enlargement. In contrast, several parameters of LV function were significantly ameliorated only by the high-dose ACE-I but not by the combination therapy. CONCLUSIONS: After the initiation of a relatively low dose of ACE-I in acute MI, increasing the dose of ACE-I or adding ARB may equally improve survival and LV remodeling in the setting of an equal hypotensive effect. Further study with a longer treatment protocol is required to determine whether the several favorable effects on LV function elicited only by the high-dose ACE-I treatment provide further beneficial effects on survival and LV remodeling compared with the combination therapy.

Establishment of a robust dengue virus NS3-NS5 binding assay for identification of protein-protein interaction inhibitors.

Whereas the dengue virus (DENV) non-structural (NS) proteins NS3 and NS5 have been shown to interact in vitro and in vivo, the biological relevance of this interaction in viral replication has not been fully clarified. Here, we first applied a simple and robust in vitro assay based on AlphaScreen technology in combination with the wheat-germ cell-free protein production system to detect the DENV-2 NS3-NS5 interaction in a 384-well plate. The cell-free-synthesized NS3 and NS5 recombinant proteins were soluble and in possession of their respective enzymatic activities in vitro. In addition, AlphaScreen assays using the recombinant proteins detected a specific interaction between NS3 and NS5 with a robust Z' factor of 0.71. By employing the AlphaScreen assay, we found that both the N-terminal protease and C-terminal helicase domains of NS3 are required for its association with NS5. Furthermore, a competition assay revealed that the binding of full-length NS3 to NS5 was significantly inhibited by the addition of an excess of NS3 protease or helicase domains. Our results demonstrate that the AlphaScreen assay can be used to discover novel antiviral agents targeting the interactions between DENV NS proteins.

Method for detection of extraneous active bovine viral diarrhoea virus and classical swine fever virus in animal viral vaccines by RT-PCR, which amplify negative-strand viral RNA in infected cells.

An oligonucleotide sense primer, Pst324alpha, was designed and used for synthesizing cDNA from negative-strand viral RNA in infected cells and used for rapid detection of active extraneous bovine viral diarrhoea virus (BVDV) and classical swine fever virus (CSFV) in animal viral vaccines by culturing a sample in cells followed by reverse transcriptase-polymerase chain reaction (RT-PCR). Active and inactivated viruses of BVDV No. 12-43 strain and CSFV GPE(-)strain were inoculated to bovine testicle and swine testicle cells for incubation. After the complete extraction of RNA from these cells, cDNA was synthesized using Pst324alpha, and PCR was carried out using primers 324 and 326 (novel RT-PCR). Amplification of novel RT-PCR products was observed in cells infected with active viruses but not in cells inoculated with inactivated viruses, inoculums and cultured media after incubation. This novel RT-PCR was able to amplify viral sequences from cells infected with only a small number of infectious particles (less than 10 TCID50) at three days postinoculation and was as sensitive as the general RT-PCR using a random primer and the interference and immunofluorescent antibody (FA) methods. The results of experiments on detection of BVDV RNA from vaccines contaminated with active and inactivated BVDV showed that the sensitivity of the novel RT-PCR was almost the same as the sensitivities of the interference and FA methods. These results suggest that the novel RT-PCR is easier and more rapid than the interference method for detection of active BVDV and that the novel RT-PCR is a reliable means for detection of active extraneous BVDV for quality control of animal vaccines.