Desmethylclomipramine triggers mitochondrial damage and death in TGF-ß-induced mesenchymal type of A549 cells.

Lung cancer is the leading cause of cancer deaths, where the metastasis often causes chemodrug resistance and leads to recurrence after treatment. Desmethylclomipramine (DCMI), a bioactive metabolite of clomipramine, shows the therapeutic efficacy with antidepressive agency as well as potential cytostatic effects on lung cancer cells. Here, we demonstrated that DCMI effectively caused transforming growth factor (TGF)-ß1-mediated mesenchymal type of A549 cells to undergo mitochondrial death via myeloid cell leukemia-1 (Mcl-1) suppression and activation of truncated Bid (tBid). TGF-ß1 induced epithelial mesenchymal transition in A549 cells with the increase of fibronectin and decrease of E-cadherin, the activation of Akt/glycogen synthase kinase-3ß (GSK-ß)/Mcl-1 axis, and the hypo-responsiveness to cisplatin. DCMI initiated a dose-dependent cytotoxicity on TGF-ß1-mediated mesenchymal type of A549 cells through inactivating Akt/GSK-ß/Mcl-1 axis, in which mitochondria instability and caspase-9/3 activation also occurred concurrently. Pharmacological inhibition of caspase-8 and cathepsin B partly reversed tBid expression and mitochondrial damage to further attenuate DCMI-mediated cytotoxicity. Additionally, DCMI presented partial therapeutic effects in treating mesenchymal type of A549 tumor bearing nude mice through an acceleration of cancer cell death. Taken together, DCMI exerts antitumor effects via initiating the mechanisms of Akt/GSK-ß/Mcl-1 inactivation and cathepsin B/caspase-8-regulated mitochondrial death, which suggests its potential role in mesenchymal type of cancer cell therapy.

Reduced Tolerogenic Program Death-Ligand 1-Expressing Conventional Type 1 Dendritic Cells Are Associated with Rapid Decline in Chronic Obstructive Pulmonary Disease.

BACKGROUND: Chronic obstructive pulmonary disease (COPD) is characterized, at least in part, by autoimmunity through amplified T helper 1 and 17 (Th1 and Th17) immune responses. The loss of immune tolerance controlled by programmed death-ligand 1 (PD-L1) may contribute to this. OBJECTIVES: We studied the tolerogenic role of PD-L1+ dendritic cells (DCs) and their subtypes in relation to specific T cell immunity and the clinical phenotypes of COPD. METHODS: We used flow cytometry to analyze PD-L1 expression by the DCs and their subtypes in the peripheral blood mononuclear cells (PBMCs) from normal participants and those with COPD. T cell proliferation and the signature cytokines of T cell subtypes stimulated with elastin as autoantigens were measured using flow cytometry and enzyme-linked immunosorbent assays (ELISA), respectively. MEASUREMENT AND MAIN RESULTS: A total of 83 participants were enrolled (normal, n = 29; COPD, n = 54). A reduced PD-L1+ conventional dendritic cell 1 (cDC1) ratio in the PBMCs of the patients with COPD was shown (13.7 ± 13.7%, p = 0.03). The decrease in the PD-L1+ cDC1 ratio was associated with a rapid decline in COPD (p = 0.02) and correlated with the CD4+ T cells (r = -0.33, p = 0.02). This is supported by the NCBI GEO database accession number GSE56766, the researchers of which found that the gene expressions of PD-L1 and CD4, but not CD8 were negatively correlated from PBMC in COPD patients (r = -0.43, p = 0.002). Functionally, the PD-L1 blockade enhanced CD4+ T cell proliferation stimulated by CD3/elastin (31.2 ± 22.3%, p = 0.04) and interleukin (IL)-17A production stimulated by both CD3 (156.3 ± 54.7, p = 0.03) and CD3/elastin (148 ± 64.9, p = 0.03) from the normal PBMCs. The PD-L1 blockade failed to increase IL-17A production in the cDC1-depleted PBMCs. By contrast, there was no significant change in interferon (IFN)-γ, IL-4, or IL-10 after the PD-L1 blockade. Again, these findings were supported by the NCBI GEO database accession number GSE56766, the researchers of which found that only the expression of RORC, a master transcription factor driving the Th17 cells, was significantly negatively correlated to PD-L1 (r = -0.33, p = 0.02). CONCLUSIONS: Circulating PD-L1+ cDC1 was reduced in the patients with COPD, and the tolerogenic role was suppressed with susceptibility to self-antigens and linked to rapid decline caused by Th17-skewed chronic inflammation.

Effectiveness and safety of CD22 and CD19 dual-targeting chimeric antigen receptor T-cell therapy in patients with relapsed or refractory B-cell malignancies: A meta-analysis.

BACKGROUND: The efficacy of CD22 or CD19 chimeric antigen receptor T (CAR-T) cells in the management of acute lymphoblastic leukemia (ALL) and non-Hodgkin lymphoma (NHL) was observed. Because antigen loss and lack of CAR-T-cell persistence are the leading causes of progressive disease following single-antigen targeting, we evaluated CD22/CD19 dual-targeting CAR-T-cell therapy efficacy and safety in relapsed/refractory B-cell malignancies. METHODS: The Web of Science, PubMed, Cochrane, and Embase databases were searched until July 2022. Patients confirmed with any relapsed/refractory B-cell hematological malignancies were included regardless of age, gender, or ethnicity, receiving CD22 and CD19-dual-targeting CAR-T-cell therapy. The studies conducted on patients with coexisting other cancer were excluded. We used random-effect models to explore the outcome, and heterogeneity was investigated by subgroup analysis. RESULTS: Fourteen studies (405 patients) were included. The pooled overall response (OR) and complete remission (CR) were 97% and 93%, respectively, for ALL patients. The 1-year proportions of overall survival (OS) and progression-free survival (PFS) were 70% and 49%, respectively. For NHL, OR occurred in 85% of patients, and 57% experienced CR. The results illustrated that the 1-year OS and 1-year PFS were 77% and 65%, respectively. The subgroup analysis showed that the dual-targeting modality achieved higher CR in the following cases: coadministration of CD22/CD19-CAR-T cells and third-generation CAR-T cells combined with ASCT and BEAM pretreatment. The ALL and NHL groups seemed similar in treatment-related toxicity: all grade cytokine release syndrome (CRS), severe CRS, and neurotoxicity occurred in 86%, 7%, and 12% of patients, respectively. CONCLUSIONS: Our meta-analysis demonstrated that the CD22/CD19 dual-targeting CAR-T-cell strategy has high efficiency with tolerable adverse effects in B-cell malignancies.

Efficacy and safety of add-on anti-CD20 monoclonal antibody to Bruton tyrosine kinase inhibitor treatment for chronic lymphocytic leukemia: a meta-analysis.

The efficacy of Bruton tyrosine kinase inhibitors (BTKi) remains suboptimal in chronic lymphocytic leukemia (CLL) treatment. A systematic review and meta-analysis were conducted to compare the outcomes of combining anti-CD20 monoclonal antibodies (mAb) with BTKi therapy versus BTKi monotherapy for patients with CLL. We searched for relevant studies in the Pubmed, Medline, Embase, and Cochrane databases until December 2022. We estimated the effective results using a hazard ratio (HR) for survival outcomes and relative risk (RR) for response outcomes and safety. Four randomized controlled trials (including 1056 patients) were found until November 2022 and fulfilled the inclusion criteria. Progression-free survival was significantly improved with the addition of anti-CD20 mAb to BTKi over BTKi (HR 0.70, 95% confidence interval (CI) 0.51-0.97), whereas pooled analysis of overall survival did not favor combination therapy compared to BTKi monotherapy (HR 0.72, 95% CI 0.50-1.04). Combination therapy was related to a statistically better complete response (RR, 2.03; 95% CI 1.01 to 4.06) and an undetectable minimal residual disease rate (RR, 6.43; 95% CI 3.54 to 11.67). The risk of grade ≥ 3 adverse events was comparable between the two groups (RR, 1.08; (95% CI 0.80 to 1.45). Overall, adding anti-CD20 mAb to BTKi revealed superior efficacy than BTKi alone in untreated or previously treated CLL patients without affecting the safety of single-agent BTKi. Conducting further randomized studies to confirm our results and determine the optimal therapy for managing patients with CLL is essential.

Efficacy and Safety of Bruton Tyrosine Kinase Inhibitor Plus Anti-CD20 Antibody Therapy Compared With Chemoimmunotherapy as Front-line Treatment for Chronic Lymphocytic Leukemia: A Systematic Review and Meta-analysis of Randomized Controlled Trials.

Treatment with chemoimmunotherapy (CIT) is considered an appropriate front-line treatment option for chronic lymphocytic leukemia (CLL). However, outcomes remain suboptimal. Bruton tyrosine kinase inhibitor (BTKi) combined with anti-CD20 antibody is an effective treatment for treatment-naïve, relapsed/refractory CLL patients. A systematic review and meta-analysis of randomized controlled trials was performed to compare the efficacy and safety of CIT versus BTKi + anti-CD20 antibody as front-line treatment for CLL patients. The endpoints of interest included progression-free survival (PFS), overall survival (OS), overall response rate (ORR), complete response (CR) rate, and safety. Four trials (including 1479 patients) were available as of December 2022 and fulfilled the eligibility criteria. BTKi + anti-CD20 antibody treatment significantly prolonged PFS [hazard ratio (HR), 0.25; 95% confidence interval (CI), 0.15-0.42] compared with CIT, while the combination therapy did not significantly improve OS compared with CIT (HR, 0.73; 95% CI, 0.50-1.06). We observed consistent benefits for PFS among patients with unfavorable features. Although pooled analysis indicated that the addition of BTKi to anti-CD20 antibody led to a higher ORR than CIT [risk ratio (RR), 1.16; 95% CI, 1.13-1.20], there was no difference in CR between the two arms (RR, 1.10; 95% CI, 0.27-4.55). The risk of grade ≥3 adverse effects (AE) was comparable between the two groups (RR, 1.04; 95% CI, 0.92-1.17). The BTKi + anti-CD20 antibody therapy has superior outcomes compared with CIT among patients with treatment-naïve CLL, without excess of toxicity. Future studies should compare next-generation targeted agent combinations versus CIT to determine the optimal management of CLL patients.

Efficacy and Safety of Bruton Tyrosine Kinase Inhibitor Monotherapy Compared with Combination Therapy for Chronic Lymphocytic Leukemia and Small Lymphocytic Lymphoma: A Systematic Review and Meta-Analysis.

The effectiveness and safety of combination treatments such as chemoimmunotherapies in chronic lymphocytic leukemia and small lymphocytic lymphoma (CLL/SLL) remain controversial. Bruton tyrosine kinase inhibitors (BTKis) are an effective therapy for CLL/SLL patients. This meta-analysis aimed to compare the efficacy and safety of BTKis versus combination therapy in CLL/SLL patients. We searched the PubMed, Cochrane, Medline, and Embase databases through February 2023 for relevant randomized controlled trials (RCTs). Four RCTs (including 1510 patients) were found and met the inclusion criteria. Progression-free survival (PFS) was significantly improved with BTKis when compared to the combination therapy (hazard ratio (HR), 0.30; 95% confidence interval (CI), 0.22-0.40), while a pooled analysis of overall survival did not favor single-agent BTKis over the combination therapy (HR, 0.87; 95% CI, 0.67-1.15). We observed consistent benefits for PFS among patients with high-risk disease characteristics. Although there was no difference in complete response between the two arms (risk ratio (RR), 0.54; 95% CI, 0.20-1.46), BTKi use was related to a better overall response rate (RR, 1.10; 95% CI, 1.04-1.16). The risk of grade ≥3 adverse events (AEs) was comparable between the two arms (RR, 0.82; 95% CI, 0.55-1.23). However, the risk of grade ≥3 AEs was significantly lower in the second-generation BTKi group than in the combination therapy group (RR, 0.73; 95% CI, 0.54-0.98). Overall, BTKis have superior efficacy compared to the combination regimens in patients with untreated or treated CLL/SLL without excess toxicity. Further studies are needed to confirm these results and determine the optimal therapy for managing patients with CLL/SLL.

Negative regulation of type I interferon signaling by integrin-linked kinase permits dengue virus replication.

Dengue virus (DENV) infection can induce life-threatening dengue hemorrhagic fever/dengue shock syndrome in infected patients. DENV is a threat to global health due to its growing numbers and incidence of infection in the last 50 years. During infection, DENV expresses ten structural and nonstructural proteins modulating cell responses to benefit viral replication. However, the lack of knowledge regarding the cellular proteins and their functions in enhancing DENV pathogenesis impedes the development of antiviral drugs and therapies against fatal DENV infection. Here, we identified that integrin-linked kinase (ILK) is a novel enhancing factor for DENV infection by suppressing type I interferon (IFN) responses. Mechanistically, ILK binds DENV NS1 and NS3, activates Akt and Erk, and induces NF-κB-driven suppressor of cytokine signaling 3 (SOCS3) expression. Elevated SOCS3 in DENV-infected cells inhibits phosphorylation of STAT1/2 and expression of interferon-stimulated genes (ISGs). Inhibiting ILK, Akt, or Erk activation abrogates SOCS3 expression. In DENV-infected mice, the treatment of an ILK inhibitor significantly reduces viral loads in the brains, disease severity, and mortality rate. Collectively, our results show that ILK is a potential therapeutic target against DENV infection.

Polypeptide antibiotic actinomycin D induces Mcl-1 uncanonical downregulation in lung cancer cell apoptosis.

AIMS: Actinomycin (Act) D, a polypeptide antibiotic, is used clinically to inhibit the growth of malignant tumors. Act D binds to DNA at the transcription initiation complex to prevent the elongation of RNA. Act D causes DNA damage, growth inhibition, and cell death. Myeloid cell leukemia (Mcl-1) is an anti-apoptotic Bcl-2 family member protein, and the present study explored the effects and molecular mechanism of Act D-induced Mcl-1 downregulation. MAIN METHODS: Human adenocarcinoma A549 cells were used to check the cytotoxic signaling pathways of Act D, particularly in apoptotic mechanism, in a cell-based study approach. Specific blockers targeting the apoptotic factors were examined for their possible roles. KEY FINDINGS: We found that Act D caused cell growth inhibition and apoptosis. Propidium iodide-based flow cytometric analysis and immunostaining confirmed cell apoptosis. Treatment with Act D caused DNA damage, followed by p53-independent cell death. Western blotting showed a significant decrease in Mcl-1 expression, mitochondrial transmembrane potential loss, and caspase-9/caspase-3 cascade activation. The proteasome inhibitor MG132 reversed Act D-induced Mcl-1 downregulation. However, pharmacological inhibition of glycogen synthase kinase-3, p53 expression, ER stress, autophagy, and vesicle acidification, which are Mcl-1-regulating signaling pathways, did not rescue these effects. Notably, Cullin-Ring E3 ligase partially mediated Mcl-1 downregulation. Administration of transforming growth factor-ß induced mesenchymal cell differentiation, but Act D still decreased Mcl-1 and caused cell apoptosis. SIGNIFICANCE: All of these data show a potential pro-apoptotic effect for Act D by facilitating Mcl-1 uncanonical downregulation.

Measurement of lipid droplets in peripheral immune cells shows an immunomodulatory effect on monocyte polarization in experimental dyslipidaemia.

Intracellular lipid droplet (LD) generation is the primary site of energy storage, which is necessary for physiological homeostasis but is related to pathological metabolic disorders. Lipid metabolism is critical for maintaining innate and adaptive immunity; however, it is mainly undefined in peripheral immune cells. Flow cytometry-based immune profiling in healthy peripheral blood cells showed significant original generation of LDs in dendritic cells (DCs, CD3-CD19-CD56-CD11+), monocytes (CD3-CD19-CD56-CD14+), natural killer cells (NK, CD3-CD19-CD56+), and B cells (CD3-CD19+). CD36, a common scavenger receptor of lipids, was also highly expressed in LD-accumulated DCs and monocytes. Following short-term treatment with oxidized LDL (oxLDL) in an experimental ex vivo model, CD14+ monocytes showed an effective increase in LD generation, but there were no alterations in the immune cell populations. Furthermore, oxLDL-treated CD14+ monocytes displayed CD36 expression. However, oxLDL-primed CD14+ monocytes showed a blockade in the uptake of extra oxLDL, even while expressing increased CD36, indicating a defect in lipid clearance. Exogenous treatment with oxLDL caused monocyte type 1 polarization accompanied by increased LD accumulation and CD36 expression. This study describes a method to monitor LD generation and CD36 expression in peripheral immune cells and identified an immunomodulatory effect of oxLDL on monocytes by tilting them towards type 1 polarization.

Anticancer polypyrrole-polyethylenimine drug-free nanozyme for precise B-cell lymphoma therapy.

As an alternative strategy for cancer treatment, the combination of cancer nanomedicine and immunotherapy is promising with regard to efficacy and safety; however, precise modulation of the activation of antitumor immunity remains challenging. Therefore, the aim of the present study was to describe an intelligent nanocomposite polymer immunomodulator, drug-free polypyrrole-polyethyleneimine nanozyme (PPY-PEI NZ), which responds to the B-cell lymphoma tumor microenvironment, for precision cancer immunotherapy. Earlier engulfment of PPY-PEI NZs in an endocytosis-dependent manner resulted in rapid binding in four different types of B-cell lymphoma cells. The PPY-PEI NZ effectively suppressed B cell colony-like growth in vitro accompanied by cytotoxicity via apoptosis induction. During PPY-PEI NZ-induced cell death, mitochondrial swelling, loss of mitochondrial transmembrane potential (MTP), downregulation of antiapoptotic proteins, and caspase-dependent apoptosis were observed. Deregulated AKT and ERK signaling contributed to glycogen synthase kinase-3-regulated cell apoptosis following deregulation of Mcl-1 and MTP loss. Additionally, PPY-PEI NZs induced lysosomal membrane permeabilization while inhibiting endosomal acidification, partly protecting cells from lysosomal apoptosis. PPY-PEI NZs selectively bound and eliminated exogenous malignant B cells in a mixed culture system with healthy leukocytes ex vivo. While PPY-PEI NZs showed no cytotoxicity in wild-type mice, they provided long-term and efficient inhibition of the growth of B-cell lymphoma-driven nodules in a subcutaneous xenograft model. This study explores a potential PPY-PEI NZ-based anticancer agent against B-cell lymphoma.

Zika virus cleaves GSDMD to disseminate prognosticable and controllable oncolysis in a human glioblastoma cell model.

Glioblastoma (GBM) is the most common aggressive malignant brain cancer and is chemo- and radioresistant, with poor therapeutic outcomes. The "double-edged sword" of virus-induced cell death could be a potential solution if the oncolytic virus specifically kills cancer cells but spares normal ones. Zika virus (ZIKV) has been defined as a prospective oncolytic virus by selectively targeting GBM cells, but unclear understanding of how ZIKV kills GBM and the consequences hinders its application. Here, we found that the cellular gasdermin D (GSDMD) is required for the efficient death of a human GBM cell line caused by ZIKV infection. The ZIKV protease specifically cleaves human GSDMD to activate caspase-independent pyroptosis, harming both viral protease-harboring and naive neighboring cells. Analyzing human GSDMD variants showed that most people were susceptible to ZIKV-induced cytotoxicity, except for those with variants that resisted ZIKV cleavage or were defective in oligomerizing the N terminus GSDMD cleavage product. Consistently, ZIKV-induced secretion of the pro-inflammatory cytokine interleukin-1ß and cytolytic activity were both stopped by a small-molecule inhibitor targeting GSDMD oligomerization. Thus, potential ZIKV oncolytic therapy for GBM would depend on the patient's GSDMD genetic background and could be abolished by GSDMD inhibitors if required.

Hyperglycemia exacerbates dengue virus infection by facilitating poly(A)-binding protein-mediated viral translation.

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.

Monocyte Distribution Width in Children With Systemic Inflammatory Response: Retrospective Cohort Examining Association With Early Sepsis.

OBJECTIVES: To investigate the association between increased monocyte distribution width (MDW) and pediatric sepsis in the emergency department (ED). DESIGN: Retrospective cohort study. SETTING: A single academic hospital study. PATIENTS: Patients from birth to the age of 18 years who presented at the ED of an academic hospital with systemic inflammatory response syndrome (SIRS) were consecutively enrolled. Sepsis was diagnosed using the International Pediatric Surviving Sepsis Campaign criteria. INTERVENTIONS: Antibiotic treatment was administrated once infection was suspected. MEASUREMENTS AND MAIN RESULTS: Routine complete blood cell count, neutrophil-to-lymphocyte ratio (NLR), and MDW, a new inflammatory biomarker, were evaluated in the ED. Logistic regression models were used to explore associations with early pediatric sepsis. We included 201 patients with sepsis and 1,050 without sepsis. In the multivariable model, MDW greater than 23 U (odds ratio [OR], 4.97; 95% CI, 3.42-7.22; p < 0.0001), NLR greater than 6 (OR, 2.06; 95% CI, 1.43-2.94; p = 0.0001), WBC greater than 11,000 cells/µL (OR, 6.52; 95% CI, 4.45-9.53; p < 0.0001), and the SIRS score (OR, 3.42; 95% CI, 2.57-4.55; p < 0.0001) were associated with pediatric sepsis. In subgroup analysis, MDW greater than 23 U remained significantly associated with sepsis for children 6-12 years old (OR, 6.76; 95% CI, 2.60-17.57; p = 0.0001) and 13-18 years (OR, 17.49; 95% CI, 7.69-39.76; p = 0.0001) with an area under the receiver operating curve of 0.8-0.9. CONCLUSIONS: MDW greater than 23 U at presentation is associated with the early diagnosis of sepsis in children greater than or equal to 6 years old. This parameter should be considered as a stratification variable in studies of pediatric sepsis.

Role of Glycogen Synthase Kinase-3 in Interferon-γ-Mediated Immune Hepatitis.

Glycogen synthase kinase-3 (GSK-3), a serine/threonine kinase, is a vital glycogen synthase regulator controlling glycogen synthesis, glucose metabolism, and insulin signaling. GSK-3 is widely expressed in different types of cells, and its abundant roles in cellular bioregulation have been speculated. Abnormal GSK-3 activation and inactivation may affect its original bioactivity. Moreover, active and inactive GSK-3 can regulate several cytosolic factors and modulate their diverse cellular functional roles. Studies in experimental liver disease models have illustrated the possible pathological role of GSK-3 in facilitating acute hepatic injury. Pharmacologically targeting GSK-3 is therefore suggested as a therapeutic strategy for liver protection. Furthermore, while the signaling transduction of GSK-3 facilitates proinflammatory interferon (IFN)-γ in vitro and in vivo, the blockade of GSK-3 can be protective, as shown by an IFN-γ-induced immune hepatitis model. In this study, we explored the possible regulation of GSK-3 and the potential relevance of GSK-3 blockade in IFN-γ-mediated immune hepatitis.

Epithelial-to-mesenchymal transition hinders interferon-γ-dependent immunosurveillance in lung cancer cells.

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-ß 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.

Proteomic networks associated with tumor-educated macrophage polarization and cytotoxicity potentiated by heat-killed tuberculosis.

Local administration of attenuated mycobacterium has been used as a cancer treatment adjuvant to re-boost patient immune responses with variable clinical outcomes. We aimed to clarify the impact of attenuated heat-killed tuberculosis (HKTB) on tumor-associated macrophages which play critical roles in shaping immunological regulation in the tumor microenvironment. Upon HKTB stimulation, both primary macrophages derived from the peripheral blood of healthy subjects and from lung cancer patients as well as THP1-derived classically activated macrophages (Ms) and tumor-educated macrophages (TEMs) were polarized into the proinflammatory phenotype, as characterized by increased expression cluster of differentiation 86. A quantitative proteomic analysis revealed that stimulated TEMs were unable to activate the toll-like receptor 2, signal transducer and activator of transcription 1, or nuclear factor-κB signaling. Instead, they showed distinct intercellular adhesion molecule 1 signaling, impaired cell adhesion, and mitochondrial dysfunction. These molecular mechanisms might contribute to lower cytotoxicity of HKTB-stimulated TEMs against A549 cells via the release of distinct inflammatory cytokines compared to HKTB-stimulated Ms. Our study provides an unbiased and systematic interpretation of cellular and molecular alterations of HKTB-reeducated macrophages which should help illuminate potential strategies of HKTB-stimulated macrophage-based combination therapy for cancer treatment.

Elevated TNF-α Induces Thrombophagocytosis by Mononuclear Cells in ex vivo Whole-Blood Co-Culture with Dengue Virus.

Background: 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.