Bat ASC2 suppresses inflammasomes and ameliorates inflammatory diseases.

Bats are special in their ability to live long and host many emerging viruses. Our previous studies showed that bats have altered inflammasomes, which are central players in aging and infection. However, the role of inflammasome signaling in combating inflammatory diseases remains poorly understood. Here, we report bat ASC2 as a potent negative regulator of inflammasomes. Bat ASC2 is highly expressed at both the mRNA and protein levels and is highly potent in inhibiting human and mouse inflammasomes. Transgenic expression of bat ASC2 in mice reduced the severity of peritonitis induced by gout crystals and ASC particles. Bat ASC2 also dampened inflammation induced by multiple viruses and reduced mortality of influenza A virus infection. Importantly, it also suppressed SARS-CoV-2-immune-complex-induced inflammasome activation. Four key residues were identified for the gain of function of bat ASC2. Our results demonstrate that bat ASC2 is an important negative regulator of inflammasomes with therapeutic potential in inflammatory diseases.

Discovery of a Novel HIV-1 Integrase/p75 Interacting Inhibitor by Docking Screening, Biochemical Assay, and in Vitro Studies.

Protein-protein interaction between lens epithelium-derived growth factor (LEDGF/p75) and HIV-1 integrase becomes an attractive target for anti-HIV drug development. The blockade of this interaction by small molecules could potentially inhibit HIV-1 replication. These small molecules are termed as LEDGINs; and several newly identified LEDGINs have been reported to significantly reduce HIV-1 replication. Through this project, we have finished the docking screening of the Maybridge database against the p75 binding site of HIV-1 integrase using both DOCK and Autodock Vina software. Finally, we have successfully identified a novel scaffold LEDGINs inhibitor DW-D-5. Its antiviral activities and anticatalytic activity of HIV-1 integrase are similar to other LEDGINs under development. We demonstrated that the combination of DW-D-5 and FDA approved anti-HIV drugs resulted in additive inhibitory effects on HIV-1 replication, indicating that DW-D-5 could be an important component of combination pills for clinic use in HIV treatment.

The Effect of an Online Health Literacy Promotion Program for Filipino Domestic Workers: A Mixed Method Study.

Introduction: Health literacy refers to acquiring and utilizing health information to make health-related actions and decisions. Filipino domestic workers with low health literacy are often vulnerable to health problems due to poor living and working conditions. Objective: This study examined the effect of an online health literacy promotion program on health literacy and health knowledge of Filipino domestic workers in Macao. Methods: The mixed method combined quantitative findings of a quasi-experimental study with qualitative results applied. The quasi-experiment included three parallel groups: a synchronous online education group (videoconference group), an asynchronous online education group (video group), and a control group. A total of 88 Filipino domestic workers were assigned to one of these groups, and eight respondents participated in two focus-group interviews respectively after the intervention. Results: For quantitative data, both synchronous and asynchronous online education interventions had positive effects. While comparing with the control group, participants in the videoconference group were more likely to have better health promotion health literacy after the intervention (ß = 5.36, p = .02), and participants in the video group were more likely to have better general health literacy (ß = 5.17, p = .01), disease prevention health literacy (ß = 5.31, p = .04), health promotion health literacy (ß = 5.97, p = .01). For qualitative data, three themes and eight subthemes were extracted after the online health literacy promotion program. After integrating the findings of this study, the study found that this program was essential and beneficial for Filipino domestic workers' health knowledge and health literacy. Conclusion: Overall, online health literacy promotion programe had positive impacts on participants revealed in this study. Asynchronous online education has made significant progress in overall health literacy, which may be more suitable as a widely promoted education method because of the characteristics and working conditions of this population.

Direct Intra-articular Antibiotic Administration for Acute Prosthetic Joint Infection in Knee Arthroplasty.

Prosthetic joint infection (PJI) remains a challenge to treat. We utilized intra-articular administration of antibiotics for the treatment of two infected total knee arthroplasties. The first patient developed an early post-operative infection with persistent wound drainage within a week after primary total knee arthroplasty (TKA). The second patient had an acute hematogenous infection, presenting with knee pain with a preceding history of leg cellulitis, one year after a primary TKA. Both patients were treated with surgical debridement, exchange of tibial insert with implant retention, and intra-articular administration of vancomycin for six weeks. Treatment was successful for both patients, with preservation of knee function and no recurrence of infection after one year. We reported two cases of PJI treated with direct intra-articular antibiotic administration following surgical debridement and implant retention.

Gas-Forming Psoas Abscess Secondary to Lumbar Spondylodiscitis.

Lower back pain is a common presentation in clinical practice. Although most are musculoskeletal in nature, occult spine infection such as spondylodiscitis is commonly missed due to its insidious onset and non-specific symptoms. We report a case of a 63-year-old diabetic woman who presented to our institution's emergency department with altered mental status, nausea, and vomiting. She had a fall one month prior with persistent lower back-pain of increasing intensity. Initial laboratory data revealed an elevated leukocyte count with neutrophil predominance. Plain radiographs showed extensive gas shadows over the chest wall, abdomen, left thigh, and left knee. CT scan revealed L2 compression fracture with spondylodiscitis at L1/L2, left psoas abscess, and extensive subcutaneous emphysema. Open abscess drainage with extensive wound debridement was performed. Intra-operative pus, as well as blood cultures, yielded Escherichia coli. Unfortunately, the patient succumbed to the infection on the seventh day of admission secondary to multi-organ failure.

Health Literacy among Filipino Domestic Workers in Macao.

Migrant worker is a global phenomenon that is associated with the health of individuals and populations. Filipino workers constitute the largest group of non-Chinese migrant workers in Macao, they are mainly employed as domestic workers. The purpose of this study is to investigate the status of health literacy (HL) and associated factors among Filipino domestic workers in Macao. The study is a cross-sectional study. Chi square's test and binary logistic regression models were used for data analyzing. Filipino who was employed by a family in Macao as a domestic worker within the valid contract period was eligible in the study. A total of 379 valid questionnaires were collected during December 2020 and March 2021. Health literacy was measured using the short-form Health Literacy Instrument (HLS-SF12). The results showed that only 37.4% of the respondents have sufficient health literacy. Age was an important factor that was associated with health literacy, with Filipino domestic workers younger than and equal to 30 years of age more likely to have inadequate health literacy. The results will help to make recommendations for further research and public health policy.

Synthesis, biological activity, molecular docking studies of a novel series of 3-Aryl-7H-thiazolo[3,2-b]-1,2,4-triazin-7-one derivatives as the acetylcholinesterase inhibitors.

The acetylcholinesterase inhibitors play a critical role in the drug therapy for Alzheimer's disease. In this study, twenty-nine novel 3-aryl-7H-thiazolo[3,2-b]-1,2,4-triazin-7-one derivatives were synthesized and assayed for their human acetylcholinesterase (hAChE) inhibitory activities. Inhibitory ratio values of seventeen compounds were above 55% with 4c having the highest value as 77.19%. The compounds with the halogen atoms in the aromatic ring, and N,N-diethylamino or N,N-dimethylamino groups in the side chains at C-3 positions exhibited good inhibitory activity. SAR study was carried out by means of molecular docking technique. According to molecular docking results, the common interacting site for all compounds were found to be peripheral anionic site whereas highly active compounds were interacting with the catalytic active site too. HIGHLIGHTSA novel series of 3-aryl-7H-thiazolo[3,2-b]-1,2,4-triazin-7-one derivatives were synthesized and assayed for their human acetylcholinesterase (hAChE) inhibitory activities.The SAR study of the target 3-aryl-7H-thiazolo[3,2-b]-1,2,4-triazin-7-one derivatives was summarized.The active sites in the acetylcholinesterase were analyzed by molecular docking technique.Communicated by Ramaswamy H. Sarma.

Development of an enzyme-linked immunosorbent assay for Keap1-Nrf2 interaction inhibitors identification.

Development of Keap1-Nrf2 interaction inhibitors is a promising strategy for the discovery of therapeutic agents against oxidative stress-mediated diseases. Two motifs of Nrf2, ETGE and DLG motif, are responsible for Keap1-Nrf2 binding. Previously, ETGE peptide or ETGE-derived peptide-based approaches were used to detect Keap1-Nrf2 interaction; however, these approaches are not able to monitor Keap1-DLG motif binding. We first report here a novel Enzyme-linked Immunosorbent Assay (ELISA) approach to detect the protein-protein interaction of full length Keap1 and Nrf2. In our assay, the test compounds can target either ETGE or DLG binding site, therefore facilitating the exploration of diverse Keap1-Nrf2 inhibitors. Three FDA-approved drugs, zafirlukast, dutasteride and ketoconazole, were found to inhibit the Keap1-Nrf2 interaction with IC50 of 5.87, 2.81 and 1.67 µM, respectively. Additionally, these three drugs also activated Nrf2 pathway in neuroblasts and lipopolysaccharide (LPS)-challenged mice. The results presented here indicate that the ELISA approach has the capacity to identify Keap1-Nrf2 inhibitors.

Erratum: E3 Ubiquitin ligase ZNRF4 negatively regulates NOD2 signalling and induces tolerance to MDP.

This corrects the article DOI: 10.1038/ncomms15865.

E3 Ubiquitin ligase ZNRF4 negatively regulates NOD2 signalling and induces tolerance to MDP.

Optimal regulation of the innate immune receptor nucleotide-binding oligomerization domain-containing protein 2 (NOD2) is essential for controlling bacterial infections and inflammatory disorders. Chronic NOD2 stimulation induces non-responsiveness to restimulation, termed NOD2-induced tolerance. Although the levels of the NOD2 adaptor, RIP2, are reported to regulate both acute and chronic NOD2 signalling, how RIP2 levels are modulated is unclear. Here we show that ZNRF4 induces K48-linked ubiquitination of RIP2 and promotes RIP2 degradation. A fraction of RIP2 localizes to the endoplasmic reticulum (ER), where it interacts with ZNRF4 under either 55 unstimulated and muramyl dipeptide-stimulated conditions. Znrf4 knockdown monocytes have sustained nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) activation, and Znrf4 knockdown mice have reduced NOD2-induced tolerance and more effective control of Listeria monocytogenes infection. Our results thus demonstrate E3-ubiquitin ligase ZNRF4-mediated RIP2 degradation as a negative regulatory mechanism of NOD2-induced NF-κB, cytokine and anti-bacterial responses in vitro and in vivo, and identify a ZNRF4-RIP2 axis of fine-tuning NOD2 signalling to promote protective host immunity.

Nursing students' perception of high-fidelity simulation activity instead of clinical placement: A qualitative study.

BACKGROUND: The high-fidelity simulation (HFS) has been utilized in nursing education for more than 20 years. Advantages of the use of high-fidelity simulation in nursing education have been documented in the literature. Based on the advantages, it has been arranged as a part of the clinical study course of the first year baccalaureate nursing program in one of the nursing colleges in Macau recently. OBJECTIVE: The aim of this study is to explore undergraduate nursing students' perception of using high-fidelity simulation as part of their clinical study course in Macau. DESIGN: This is a qualitative study using open-ended questionnaire. SETTING: This study was implemented at the nursing laboratory between 1 April and 17 April 2015, which was the period of preliminary clinical study course of year one nursing students. PARTICIPANTS: A purposive sample was sought from the voluntary year one undergraduate nursing students who participated in the clinical study course. METHODS: Students received two high-fidelity simulation sections during the course, while a self-administered open-ended questionnaire was allocated afterward. Qualitative content analysis was performed after data collection. RESULTS: Two themes emerged in this study, which included "appreciation" and "misunderstanding". They were further divided into five categories; as "positive feelings", "gaining a suitable atmosphere for learning", "assist of adequate emergency preparation: resourceful ability", "contempt", and "rote learning". CONCLUSION: This was the first time to utilize HFS activities as a part of the clinical study course in one nursing college in Macau. These HFS activities instead of a part of real clinical placement were appreciated by nursing students. And it mainly contributed to the resourceful ability in students' view. During the HFS activities, nursing educators should consider the misunderstanding of HFS activities of students that a few nursing students despised simulator's life and got rote learning method.

Hepatitis B virus-like particles access major histocompatibility class I and II antigen presentation pathways in primary dendritic cells.

Virus-like particles (VLPs) represent high density displays of viral proteins that efficiently trigger immunity. VLPs composed of the small hepatitis B virus envelope protein (HBsAgS) are useful vaccine platforms that induce humoral and cellular immune responses. Notably, however, some studies suggest HBsAgS VLPs impair dendritic cell (DC) function. Here we investigated HBsAgS VLP interaction with DC subsets and antigen access to major histocompatibility complex (MHC) class I and II antigen presentation pathways in primary DCs. HBsAgS VLPs impaired plasmacytoid DC (pDC) interferon alpha (IFNα) production in response to CpG in vitro, but did not alter conventional DC (cDC) or pDC phenotype when administered in vivo. To assess cellular immune responses, HBsAgS VLPs were generated containing the ovalbumin (OVA) model epitopes OVA(257-264) and OVA(323-339) to access MHCI and MHCII antigen presentation pathways, respectively; both in vitro and following immunisation in vivo. HBsAgS VLP-OVA(257-264) elicited CTL responses in vivo that were not enhanced by inclusion of an additional MHCII helper epitope. HBsAgS VLP-OVA(257-264) administered in vivo was cross-presented by CD8(+) DCs, but not CD8(-) DCs. Therefore, HBsAgS VLPs can deliver antigen to both MHCI and MHCII antigen presentation pathways in primary DCs and promote cytotoxic and helper T cell priming despite their suppressive effect on pDCs.

Acetylshikonin, a Novel AChE Inhibitor, Inhibits Apoptosis via Upregulation of Heme Oxygenase-1 Expression in SH-SY5Y Cells.

Acetylcholinesterase inhibitors are prominent alternative in current clinical treatment for AD patients. Therefore, there is a continued need to search for novel AChEIs with good clinical efficacy and less side effects. By using our in-house natural product database and AutoDock Vina as a tool in docking study, we have identified twelve phytochemicals (emodin, aloe-emodin, chrysophanol, and rhein in Rhei Radix Et Rhizoma; xanthotoxin, phellopterin, alloisoimperatorin, and imperatorin in Angelicae dahuricae Radix; shikonin, acetylshikonin, isovalerylshikonin, and ß,ß-dimethylacrylshikonin in Arnebiae Radix) as candidates of AChEIs that were not previously reported in the literature. In addition to AChEI activity, a series of cell-based experiments were conducted for the investigation of their neuroprotective activities. We found that acetylshikonin and its derivatives prevented apoptotic cell death induced by hydrogen peroxide in human and rat neuronal SH-SY5Y and PC12 cells at 10 µM. We showed that acetylshikonin exhibited the most potent antiapoptosis activity through the inhibition of the generation of reactive oxygen species as well as protection of the loss of mitochondria membrane potential. Furthermore, we identified for the first time that the upregulation of heme oxygenase 1 by acetylshikonin is a key step mediating its antiapoptotic activity from oxidative stress in SH-SY5Y cells.

Modulation of the immunogenicity of virus-like particles composed of mutant hepatitis B virus envelope subunits.

Virus-like particles (VLPs) are non-infectious subviral protein complexes, which possess structural features identical or closely related to infectious virions. They are utilized as delivery tools for immunologically relevant antigenic sequences. In order to investigate whether mutant subunits can modulate the VLP immunogenicity, comparative immunization studies with wild-type and non-native VLPs were performed. To determine whether disulfide bonding impacts on the immunogenicity of hepatitis B virus envelope proteins (HBsAg), mutant HBsAg subunits with single, double and triple cysteine residue substitutions were generated. The mutant proteins were expressed in cell culture, secretion competent non-native VLPs generated, followed by immunization studies in mice to measure the cellular immune response. The reduced ability of mutant HBsAg proteins to form disulfide bonds does not interfere with their ability to assemble into secretion competent VLPs. Depending on specific cysteine to alanine changes, VLPs could be generated with or without an increased ratio of monomeric versus dimeric/oligomeric subunits compared to wild-type VLPs. The utilization of non-native VLPs resulted in enhanced cellular immune responses and does not seem to depend on the ratio between monomeric or dimeric/oligomeric subunits. Comparative immunization studies strongly indicate that changes in the disulfide bonding modulate the VLP immunogenicity most likely due to structural changes. We hypothesize that structural features have evolved with reduced immunogenicity to evade the constraints imposed by the immune system. Altering VLP conformation may represent an attractive strategy to modulate antigen processing resulting in an enhanced immune response and/or a changed hierarchy of epitope presentation.

Delivery of a foreign epitope by sharing amino acid residues with the carrier matrix.

A broad range of structural viral proteins has the ability to assemble into virus-like particles (VLPs). Under the condition that modified subunits are still competent to assemble into VLPs, they are epitope delivery platforms suitable for vaccination purposes. The insertion of foreign sequences can be detrimental for the formation of chimeric VLPs as a result of misfolded subunit proteins. Hence, a strategy was adopted to screen for locations allowing the use of shared residues between the wildtype subunit sequence and the foreign insert. The insertion of a cysteine-containing sequence of hepatitis C virus (HCV) envelope protein 2 (E2) without adding an additional cysteine residue retained the ability of recombinant small hepatitis B surface antigen (HBsAg-S) to form secretion competent VLPs. A cysteine residue shared by the insert and the template protein avoided the formation of non-native disulfide bonds, and allowed the formation of VLPs. The chimeric HBsAg-S VLPs were similar to wildtype VLPs in density exposing the inserted foreign epitope and being immunogenic. Overall, the use of shared sequences between the insert and the subunit will facilitate the design of chimeric VLPs carrying multiple epitopes.

Chimeric virus-like particles for the delivery of an inserted conserved influenza A-specific CTL epitope.

The small hepatitis B virus surface antigens (HBsAg-S) have the ability to self-assemble with host-derived lipids into empty non-infectious virus-like particles (VLPs). HBsAg-S VLPs are the sole component of the licensed hepatitis B vaccine, and they are a useful delivery platform for foreign epitopes. To develop VLPs capable of transporting foreign cytotoxic T lymphocyte (CTL) epitopes, HBsAg-S specific CTL epitopes at various sites were substituted with a conserved CTL epitope derived from the influenza matrix protein. Depending on the insertion site, the introduction of the MHC class I A2.1-restricted influenza epitope was compatible with the secretion competence of HBsAg-S indicating that chimeric VLPs were assembled. Immunizations of transgenic HHDII mice with chimeric VLPs induced anti-influenza CTL responses proving that the inserted foreign epitope can be correctly processed and cross-presented. Chimeric VLPs in the absence of adjuvant were able to induce memory T cell responses, which could be recalled by influenza virus infections in the mouse model system. The ability of chimeric HBsAg-S VLPs to induce anti-foreign CTL responses and also with the proven ability to induce humoral immune responses constitute a highly versatile platform for the delivery of selected multiple epitopes to target disease associated infectious agents.

Wikstroelide M potently inhibits HIV replication by targeting reverse transcriptase and integrase nuclear translocation / 中国天然药物

AIM@#To evaluate the anti-HIV activity and mechanism of action of wikstroelide M, a daphnane diterpene from Daphne acutiloba Rehder (Thymelaeaceae).@*METHODS@#The anti-HIV activities of wikstroelide M against different HIV strains were evaluated by cytopathic effect assay and p24 quantification assay with ELISA. The inhibitory effect of wikstroelide M on HIV reverse transcription was analyzed by real-time PCR and ELISA. The effect of wikstroelide M on HIV-1 integrase nuclear translocation was observed with a cell-based imaging assay. The effect of wikstroelide M on LEDGF/p75-IN interaction was assayed by molecular docking.@*RESULTS@#Wikstroelide M potently inhibited different HIV-1 strains, including HIV-1IIIB, HIV-1A17, and HIV-19495, induced a cytopathic effect, with EC50 values ranging from 3.81 to 15.65 ng·mL⁻¹. Wikstroelide M also had high inhibitory activities against HIV-2ROD and HIV-2CBL-20-induced cytopathic effects with EC50 values of 18.88 and 31.90 ng·mL⁻¹. The inhibitory activities of wikstroelide M on the three HIV-1 strains were further confirmed by p24 quantification assay, with EC50 values ranging from 15.16 to 35.57 ng·mL⁻¹. Wikstroelide M also potently inhibited HIV-1IIIB induced cytolysis in MT-4 cells, with an EC50 value of 9.60 ng·mL⁻¹. The mechanistic assay showed that wikstroelide M targeted HIV-1 reverse transcriptase and nuclear translocation of integrase through disrupting the interaction between integrase and LEDGF/p75.@*CONCLUSION@#Wikstroelide M may be a potent HIV-1 and HIV-2 inhibitor, the mechanisms of action may include inhibition of reverse trascriptase activity and inhibition of integrase nuclear translocation through disrupting the interaction between integrase and LEDGF/p75.