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(Note: The title has been modified in peer-reviewed publication. Accepted manuscript will be available only after the one-year embargo period.)Objectives: In this study, damage at the composite-tooth interface was investigated by finite element analysis (FEA) using the cohesive zone model (CZM). Methods: Axisymmetric models of Class I restorations were created to illustrate the interfacial damage around composite restorations of different dimensions, with polymerization shrinkage modeled analogously to thermal shrinkage. The damage of the adhesive interface was determined using a CZM based on the fracture strength and fracture energy. For comparison with the conventional FEA approach, additional models were created using perfectly bonded composite restorations as controls to show the effects of damage.Results: The results indicated damage to the interface at the butt-joint cavosurface margin, dentinoenamel junction, and internal line angle. The percentage of damaged interfacial area was found to increase with the C-factor (the ratio of bonded area to unbonded surface area) for restorations of the same height. For a given diameter, the damage was more severe for restorations of greater depth. Comparisons between the CZM results and the FEA models for perfect adhesion showed stress relief and a changed deformation pattern around the damage.Significance: The fracture mechanics-based CZM is an essential refinement for the current FEA approach. The study yields indispensable insights for future studies by elucidating the links among interfacial damage, stress relief, and altered deformation. Furthermore, the open-source workflow provides a versatile framework to address clinically relevant issues in adhesive dentistry, proving that it is possible to perform these advanced analyses without expensive software.

Dicer expression involves in paclitaxel chemosensitization (S1); Dicer expression positively correlates with better survival outcomes in breast cancer patients (S2); E1A-induced chemosensitization of paclitaxel may through both Dicer- and E2Fs-mediated mechanisms (S3); AXL expression increases paclitaxel resistance of breast cancer cells, and positively correlates with invasive breast cancer carcinoma (S4); Dicer suppresses AXL expression and enhances doxorubicin, fluorouracil and cisplatin sensitization in MCF7 cells (S5); The mRNA and protein stability of Dicer in MDA-MB-231/vector and MDA-MB-231/E1A cells (S6); The association of C/EBPα expression in advanced breast cancer (S7); C/EBPα and MAPK14 expression positively correlates with better survival outcomes in breast cancer patients (S8).

Legend for Supplementary Figures S1-S8.

Description of additional methods and procedures used in the study. Also includes Supplementary Reference.

Association between stem cell markers and Dicer expression in breast cancer datasets (S1); Clinicopathological features, clinical outcomes and their association with Dicer expression in breast cancer datasets (S2); Association between stem cell markers and AXL expression in breast cancer datasets (S3); Association between Dicer, AXL, C/EBPα and MAPK14 expression in breast cancer datasets (S4); Association between stem cell markers and C/EBPα expression in breast cancer datasets (S5); Association between AXL, C/EBPα and MAPK14 expression in breast cancer datasets (S6); Association between C/EBPα and MAPK14 expression in breast cancer datasets (S7); Association between stem cell markers and MAPK14 expression in breast cancer datasets (S8); Sequences and information of PCR primers for microRNA (miR), ChIP and cloning (S9).

Rahmat Dani Satria,1– 4,&ast; Ming-Kai Jhan,4,5,&ast; Chia-Ling Chen,6 Po-Chun Tseng,4,7 Yung-Ting Wang,4 Chiou-Feng Lin4,5,7 1International Ph.D. Program in Medicine, College of Medicine, Taipei Medical University, Taipei, 110, Taiwan; 2Department of Clinical Pathology and Laboratory Medicine, Faculty of Medicine, Public Health and Nursing, Universitas Gadjah Mada, Yogyakarta, 55281, Indonesia; 3Clinical Laboratory Installation, Dr. Sardjito Central General Hospital, Yogyakarta, 55281, Indonesia; 4Department of Microbiology and Immunology, School of Medicine, College of Medicine, Taipei Medical University, Taipei, 110, Taiwan; 5Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, 110, Taiwan; 6School of Respiratory Therapy, College of Medicine, Taipei Medical University, Taipei, 110, Taiwan; 7Core Laboratory of Immune Monitoring, Office of Research & Development, Taipei Medical University, Taipei, 110, Taiwan&ast;These authors contributed equally to this workCorrespondence: Chiou-Feng Lin, Department of Microbiology and Immunology, School of Medicine, College of Medicine, Taipei Medical University, Taipei, 110, Taiwan, Tel +886 2 27361661 ext. 7156, Email cflin2014@tmu.edu.twBackground: Infection with dengue virus (DENV) causes hematological complications in dengue diseases characterized by thrombocytopenia accompanied by macrophage activation syndrome and hemophagocytosis in fatal patients.Methods: In this study, we investigate the undefined mechanisms underlying the progression of thrombocytopenia caused by thrombophagocytosis based on an ex vivo whole-blood co-culture model of DENV infection for mimicking the acute febrile phase of infection.Results: In this model, complete blood count test showed a decrease in monocytes (p < 0.01), but not neutrophils nor other white blood cells, accompanied by a low thrombocyte count (p < 0.01) in DENV infection with a positive correlation (r = 0.636, p < 0.05). Furthermore, DENV exposure caused significant thrombophagocytosis in mononuclear cells (p < 0.05). Abnormal production of tumor necrosis factor (TNF)-α was highly associated with induction of thrombophagocytosis (r = 0.758, p < 0.01), decreased monocytes (r = − 0.758, p < 0.01), and decreased thrombocyte (r = − 0.728, p < 0.01). Neutralizing TNF-α considerably (p < 0.05) reversed such DENV-induced effects and was further validated by immunostaining-based flow cytometry analysis on mononuclear CD14 positive monocytes. Exogenous administration of TNF-α effectively caused thrombophagocytosis accompanied by decreased monocytes and thrombocytes, probably causing monocyte activation.Conclusion: These results demonstrate the potential pathogenesis of thrombocytopenia caused by TNF-α-induced thrombophagocytosis in monocytes during DENV infection.Graphical Abstract: Keywords: dengue virus, monocyte, thrombophagocytosis, ex vivo, TNF-&#x03B1

Bệnh xuất huyết do vi khuẩn Streptococcus sp là mầm bệnh truyền nhiễm chính gây thiệt hại đáng kể đến sản lượng cá rô phi toàn cầu. Hepcidin/HAMP ở cá đã được báo cáo có liên quan đến miễn dịch bẩm sinh chống lại các mầm bệnh vi khuẩn. Trong nghiên cứu này, chúng tôi tiến hành phân tích mối quan hệ giữa tính đa hình microsatellites/SSRs liên kết với gen hepcidin/HAMP và khả năng kháng bệnh do Streptococcus iniae trên cá rô phi vằn dòng NT1 (Đài Loan). 17 chỉ thị SSRs và cặp mồi đặc hiệu đã được thiết kế dựa trên WebSat. Kết quả đánh giá trên 95 cá thể cho thấy 9/17 chỉ thị SSRs có tính đa hình cao và tuân theo định luật Hardy-Weinberg. Các chỉ thị này sẽ được sử dụng để đánh giá khả năng kháng vi khuẩn S. iniae. 29 cá rô phi NT1 thế hệ thứ nhất (khối lượng 23,59 ± 5,388 g/con) đã được cảm nhiễm với vi khuẩn S. iniae 89353 bằng phương pháp tiêm, với liều tiêm LD50 là 1,3x105 cfu/mL. Kết quả phân tích cho thấy, có sự sai khác có ý nghĩa thống kê về kiểu gen và tần số alen giữa nhóm sống và nhóm chết sau cảm nhiễm vi khuẩn ở 3 chỉ thị SSRs (SSR7, SSR9 và SSR16) (p

Diabetes mellitus (DM) is highly comorbid with severe dengue diseases; however, the underlying mechanisms are unclear. Patients with DM have a 1.61-fold increased risk of developing dengue hemorrhagic fever. In search of host factors involved in dengue virus (DENV) infection, we used high-glucose (HG) treatment and showed that HG increased viral protein expression and virion release but had no effects on the early stages of viral infection. After HG stimulation, DENV-firefly luciferase-transfected assay and cellular replicon-based assay indicated increased viral translation, whereas using the glucose uptake inhibitor phloretin blocked this effect. HG treatment increased the translational factor poly(A)-binding protein (PABP) in a glucose transporter-associated, PI3K/AKT-regulated manner. Silencing PABP significantly decreased HG-prompted virion production. HG enhanced the formation of the PABP-eukaryotic translation initiation factor 4G complex, which is regulated by protein-disulfide isomerase. Hyperglycemia increased PABP expression, mortality rate, viral protein expression, and viral loads in streptozotocin-induced DM mice. Overall, hyperglycemic stress facilitates DENV infection by strengthening PABP-mediated viral translation.

Omicron variants of SARS-CoV-2 has been dominantly distributed worldwide since December 2021. Sequence analysis has revealed a 9 base pair (bp) nucleotide deletion located in N gene was found in Omicron strains but not in other prevalent variants. Based on this feature, we have developed a highly specific and sensitive real-time reverse-transcriptase polymerase chain reaction (RT-PCR) assay for differentiating Omicron lineages from other SARS-CoV-2. With this unique design of primers and probes, we consider this approach could be applied to identify particular genome feature for future SARS-CoV-2 variant characterization.

History A 55-year-old woman without systemic underlying disease, such as diabetes mellitus, inflammatory bowel disease, autoimmune disease, or chronic kidney disease, presented with generalized dull abdominal pain of 1-week duration. She had ingested herbal medicine for physical conditioning for several years. Laboratory findings, including biochemistry, electrolyte levels, and complete blood count, were all within normal limits, except for elevated serum C-reactive protein level (7.719 mg/dL; normal range

Chun-Yi Lai,1,2,&ast; Po-Chun Tseng,1,2,&ast; Chia-Ling Chen,3 Rahmat Dani Satria,2,4– 6 Yung-Ting Wang,2 Chiou-Feng Lin1,2,7,8 1Core Laboratory of Immune Monitoring, Office of Research & Development, Taipei Medical University, Taipei, 110, Taiwan; 2Department of Microbiology and Immunology, School of Medicine, College of Medicine, Taipei Medical University, Taipei, 110, Taiwan; 3School of Respiratory Therapy, College of Medicine, Taipei Medical University, Taipei, 110, Taiwan; 4International Ph.D. Program in Medicine, College of Medicine, Taipei Medical University, Taipei, 110, Taiwan; 5Department of Clinical Pathology and Laboratory Medicine, Faculty of Medicine, Public Health and Nursing, Universitas Gadjah Mada, Yogyakarta, 55281, Indonesia; 6Clinical Laboratory Installation, Dr. Sardjito Central General Hospital, Yogyakarta, 55281, Indonesia; 7Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, 110, Taiwan; 8International Ph.D. Program in Cell Therapy and Regenerative Medicine, College of Medicine, Taipei Medical University, Taipei, 110, Taiwan&ast;These authors contributed equally to this workCorrespondence: Chiou-Feng LinDepartment of Microbiology and Immunology, School of Medicine, College of Medicine, Taipei Medical University, Taipei, 110, TaiwanTel +886 2 27361661 ext. 7156Email cflin2014@tmu.edu.twBackground: Phorbol 12-myristate 13-acetate (PMA)-induced differentiation of human monocytic THP-1 cells is an experimental model for preparing resting macrophages (M0) for cell polarization toward the different functional specializations of macrophages.Methods: In this study, we examined the expression of immune checkpoints by using flow cytometry following multicolor staining. The blockade of immune checkpoint by using neutralizing antibodies was performed to assess their role in PMA-induced THP-1-differentiated macrophages.Results: Upon the inducible macrophage differentiation caused by PMA, increased expression levels of CD11b and CD68 were measured and characterized according to their adherent phenotype accompanied by the generation of cellular complexity. While the cell growth rate was abolished post-differentiation, some cells underwent cell death. Notably, we found increases in the expression of programmed cell death protein 1, also known as PD-1 (CD279), and its ligand PD-L1 (CD274), mainly in differentiated M0 (CD68+CD11b+) macrophages. However, neutralizing PD-L1/PD-1 neither blocked THP-1 cell differentiation toward macrophages nor inhibited macrophage polarization in M1 and M2. In specializing macrophages, a decrease both in CD274 and CD279 was found in M2.Conclusion: These results revealed the inducible expression of PD-L1/PD-1 in PMA-induced THP-1-differentiated M0 macrophages followed by a decrease in M2 macrophages.Keywords: PMA, THP-1, macrophage, differentiation, PD-L1, PD-1

Background. Dengue virus (DENV) infection is the most common arboviral disease that affects tropical and subtropical regions. Based on the clinical hallmarks, the different severities of patients range from mild dengue fever (MDF) to severe dengue diseases (SDDs) and include dengue hemorrhagic fever or dengue shock syndrome. These are commonly associated with cytokine release syndrome (CRS). The types and levels of cytokines/chemokines, which are suppressed or enhanced, are varied, indicating CRS’s pathogenic and host defensive effects. Principal Finding. In this study, we created an integrated and precise multiplex panel of cytokine/chemokine assays based on our literature analysis to monitor dengue CRS. A 24-plex panel of cytokines/chemokines was evaluated to measure the plasma levels of targeting factors in dengue patients with an MDF and SDD diagnosis without or with comorbidities. As identified in sixteen kinds of cytokines/chemokines, ten were significantly (P<0.05) (10/16) increased, one was significantly (P<0.01) (1/16) decreased, and five were potentially (5/16) altered in all dengue patients (n=30) in the acute phase of disease onset. Compared to MDF, the levels of IL-8 (CXCL-8) and IL-18 in SDD were markedly (P<0.05) increased, accompanied by positively increased IL-6 and TNF-α and decreased IFN-γ and RANTES. With comorbidities, SDD significantly (P<0.01) portrayed elevated IL-18 accompanied by increased IL-6 and decreased IFN-α2 and IL-12. In addition, decreased platelets were significantly (P<0.05) associated with increased IL-18. Significance. These results demonstrate an efficient panel of dengue cytokine/chemokine assays used to explore the possible level of CRS during the acute phase of disease onset; also, we are the first to report the increase of IL-18 in severe dengue with comorbidity compared to severe dengue without comorbidity and mild dengue.

The adverse effect of cisplatin administration causes acute kidney injury (AKI) following renal inflammation and nephrotoxicity, characterized by proximal tubular cell apoptosis and necrosis. Pro-apoptotic and pro-inflammatory roles of glycogen synthase kinase (GSK)-3β have been reported. This study investigated the therapeutic blockade of GSK-3β in cisplatin-induced AKI. A renal cisplatin nephrotoxicity model showed activation of GSK-3β in vivo, particularly in proximal tubular epithelial cells. Pharmacologically inhibiting GSK-3β abolished cisplatin nephrotoxicity, including proximal tubular injury, cell cytotoxicity, and biochemical dysfunction. Additionally, GSK-3β inhibitor treatment ameliorated renal inflammation by reducing immune cell infiltration, cell adhesion molecule expression, and pro-inflammatory cytokine/chemokine production. Cisplatin treatment caused GSK-3β activation in vitro in the human renal proximal tubular epithelial cell line HK-2, whereas either pharmacological administration of GSK-3β inhibitors or genetic transduction of GSK-3β short-hairpin RNA impeded cisplatin-induced cytotoxicity. These results indicate that cisplatin activates GSK-3β followed by GSK-3β-mediated renal inflammation and nephrotoxicity, contributing to AKI.

The epithelial-to-mesenchymal transition (EMT) is involved in cancer metastasis; nevertheless, interferon (IFN)-γ induces anticancer activities by causing cell growth suppression, cytotoxicity, and migration inhibition. Regarding the poor response to exogenously administered IFN-γ as anticancer therapy, it was hypothesized that malignant cells may acquire a means of escaping from IFN-γ immunosurveillance, likely through an EMT-related process. A genomic analysis of human lung cancers revealed a negative link between the EMT and IFN-γ signaling, while compared to human lung adenocarcinoma A549 cells, IFN-γ-hyporesponsive AS2 cells exhibited mesenchymal characteristics. Chemically, physically, and genetically engineered EMT attenuated IFN-γ-induced IFN regulatory factor 1 transactivation. Poststimulation of transforming growth factor-β induced the EMT and also selectively retarded IFN-γ-responsive gene expression as well as IFN-γ-induced signal transducer and activator of transcription 1 activation, major histocompatibility complex I, and CD54 expression, cell migration/invasion inhibition, and direct/indirect cytotoxicity. Without changes in IFN-γ receptors, excessive oxidative activation of Src homology-2 containing phosphatase 2 (SHP2) in cells undergoing the EMT primarily caused cellular hyporesponsiveness to IFN-γ signaling and cytotoxicity, while combining an SHP2 inhibitor or antioxidant sensitized EMT-associated AS2 and mesenchymal A549 cells to IFN-γ-induced priming effects on tumor necrosis factor-related apoptosis-inducing ligand cytotoxicity. In cell line-derived xenograft models, combined treatment with IFN-γ and an SHP2 inhibitor induced enhanced anticancer activities. These results imply that EMT-associated SHP2 activation inhibits IFN-γ signaling, facilitating lung cancer cell escape from IFN-γ immunosurveillance.

Patients receiving hemodialysis (HD) are at risk of TB development. IGRA-positive patients showed significant decrease in quantitative IGRA result with alterations in CD3+CD4+CD45RO+, NK cell, and monocyte subsets immediately upon HD procedure. Our result suggested that the timing of IGRA testing is crucial in end-stage renal disease population.

Propofol, 2,6-diisopropylphenol, is a short-acting intravenous sedative agent used in adults and children. Current studies show its various antimicrobial as well as anti-inflammatory effects. Dengue virus (DENV) is an emerging infectious pathogen transmitted by mosquitoes that causes mild dengue fever and progressive severe dengue diseases. In the absence of safe vaccines and antiviral agents, adjuvant treatments and supportive care are generally administered. This study investigated the antiviral effects of propofol against DENV infection and cellular inflammation by using an in vitro cell model. Treatment with propofol significantly inhibited DENV release 24 h postinfection in BHK-21 cells. Furthermore, it also blocked viral protein expression independent of the translational blockade. Propofol neither caused inhibitory effects on endosomal acidification nor prevented dsRNA replication. Either the proinflammatory TNF-α or the antiviral STAT1 signaling was reduced by propofol treatment. These results provide evidence to show the potential antiviral effects of the sedative propofol against DENV infection and cellular inflammation.

Proinflammatory TNF-α facilitates dengue virus (DENV) infection in endovascular dysfunction and neurotoxicity. The introduction of TNF-α blocking therapy with Abs is performed to test its therapeutic effect in this study. In DENV-infected mice, TNF-α production in the brain accompanied the progression of neurotoxicity and encephalitis. DENV infection caused the loss of hippocampal neurons with TNF-α expression around damaged regions, and immunostaining showed the induction of apoptosis in hippocampal neurons. TNF-α was expressed in active microglia and astrocytes in DENV-infected mice. TNF-α facilitated DENV-induced neurotoxicity in vitro in murine Neuro-2a cells. Using a currently established encephalitic mouse model in which DENV infection causes progressive hunchback posture, limbic seizures, limbic weakness, paralysis, and lethality 7 days postinfection, we showed that TNF-α transgenic mice represented the progressive disease development and administration of neutralizing TNF-α Ab reduced dengue encephalitis and mortality. These results demonstrate an immunopathogenesis of TNF-α for mediating DENV-induced encephalitis-associated neurotoxicity and that targeting TNF-α can be used as a strategy against dengue encephalitis. Dengue virus infection induces TNF-α expression around the damaged neuronal cells in the brain while TNF-α blockade ameliorates dengue neurotoxicity and encephalitis.

Dengue virus (DENV), an arthropod-borne virus transmitted by mosquitoes, may cause the severe disease known as dengue hemorrhagic fever, which is characterized by lethal complications due to plasma leakage, ascites, pleural effusion, respiratory distress, severe bleeding, and organ impairment. A few cases of DENV infection present neurological manifestations; however, studies have not explored DENV-induced neuropathogenesis further. In this study, we present a protocol to use an immunocompetent outbred ICR (Institute of Cancer Research) mouse for investigating the induction of central nervous system (CNS) infection with DENV, followed by the progression of acute viral encephalitis-like disease.

Interferon (IFN)-γ is crucial for normal immune surveillance and exhibits immunomodulatory, antimicrobial, and anticancer activity. Patients with nontuberculous mycobacteria (NTM) infection commonly express high levels of anti-IFN-γ autoantibodies (autoAbs) and suffer from recurrent infections due to adult-onset immunodeficiency with defects in IFN-γ immune surveillance. In this study, we developed the methods for determination of anti-IFN-γ autoAbs and then characterized their neutralizing activity in patients with NTM infection. A modified sandwich ELISA-based colorimetric assay followed by immunoblot analysis detected the presence of autoAbs in three out of five serum samples. Serum levels of IFN-γ were decreased. Synthetic peptide binding assay showed variable patterns of epitope recognition in patients positive for anti-IFN-γ autoAbs. Functional tests confirmed that patient serum blocked IFN-γ-activated STAT1 activation and IRF1 transactivation. Furthermore, IFN-γ-regulated inflammation, chemokine production and cytokine production were also blocked. These results provide potentially useful methods to assay anti-IFN-γ autoAbs and to characterize the effects of neutralizing autoAbs on IFN-γ signaling and bioactivity.

The interferon- (IFN-) γ expression is elicited in response to microbial infections and activates immune surveillance by antimicrobial immune elements to induce microbial killing. Patients with adult-onset immunodeficiency who suffer from recurrent infections with microbes, particularly nontuberculous mycobacteria (NTM), commonly display genetic defects in IFN-γ signaling as well as the generation of anti-IFN-γ autoantibodies (autoAbs). Because IFN-γ is an activator of macrophage differentiation and a proinflammatory activator of innate immunity, the blockade effects of the autoAbs present in NTM patient serum on IFN-γ are hypothesized to regulate the antimicrobial function of macrophages. In the presence of patient serum, IFN-γ-induced type 1 macrophage (M1) differentiation was inhibited in PMA-stimulated human monocytic THP-1 cells. Treatment with patient serum significantly blocked the production of proinflammatory factors, including cytokines/chemokines and reactive oxygen/nitrogen species, by M1 macrophages. Importantly, IFN-γ-facilitated phagocytosis and degradation of heat-killed mycobacterium were decreased by cotreatment with patient serum. These results show the blockade activity of anti-IFN-γ autoAbs on IFN-γ-mediated antimicrobial immunity in macrophages.

The heterogeneity of immune response to COVID-19 has been reported to correlate with disease severity and prognosis. While so, how the immune response progress along the period of viral RNA-shedding (VRS), which determines the infectiousness of disease, is yet to be elucidated. We aim to exhaustively evaluate the peripheral immune cells to expose the interplay of the immune system in uncomplicated COVID-19 cases with different VRS periods and dynamic changes of the immune cell profile in the prolonged cases. We prospectively recruited four uncomplicated COVID-19 patients and four healthy controls (HCs) and evaluated the immune cell profile throughout the disease course. Peripheral blood mononuclear cells (PBMCs) were collected and submitted to a multi-panel flowcytometric assay. CD19+-B cells were upregulated, while CD4, CD8, and NK cells were downregulated in prolonged VRS patients. Additionally, the pro-inflammatory-Th1 population showed downregulation, followed by improvement along the disease course, while the immunoregulatory cells showed upregulation with subsequent decline. COVID-19 patients with longer VRS expressed an immune profile comparable to those with severe disease, although they remained clinically stable. Further studies of immune signature in a larger cohort are warranted.

Paclitaxel is a standard-of-care chemotherapy for breast cancer, despite the increasing recognition of its poor effectiveness in the treatment of patients with advanced disease. Here, we report that adenovirus-type 5 E1A-mediated elevation of the miRNA-processing enzyme Dicer is sufficient to enhance paclitaxel sensitization and reduce cancer stem-like cell properties in this setting. Elevating Dicer expression increased levels of the AXL kinase targeting miRNA miR-494, thereby repressing AXL expression to increase paclitaxel sensitivity. We found that Dicer expression was regulated at the transcription level by E1A, through activation of an MAPK14/CEBPα pathway. Our findings define a mechanism of E1A-mediated chemosensitization for paclitaxel, which is based upon the suppression of breast cancer stem-like cells, with potential implications for the diagnosis and treatment of breast cancer patients. Cancer Res; 76(13); 3916–28. ©2016 AACR.