Search

Background/Aims: Infliximab (IFX) has proven effective as rescue therapy in steroid-refractory acute severe ulcerative colitis (ASUC), however, the long-term real-world data are scarce. Our study aimed to assess the long-term treatment outcomes of IFX in a real-life cohort.Methods: We established a multicenter retrospective cohort of hospitalized patients with ASUC, who met Truelove and Witt’s criteria and received intravenous corticosteroid (IVCS) or IFX during index hospitalization between 2006 and 2016 in 5 university hospitals in Korea. The cohort was systematically followed up until colectomy, death or last follow-up visit.Results: A total of 296 patients were followed up for a mean of 68.9 ± 44.0 months. During index hospitalization, 49 patients were treated with IFX; as rescue therapy for IVCS failure in 37 and as first-line medical therapy for ASUC in 12. All patients treated with IFX avoided colectomy during index hospitalization. The cumulative rates of rehospitalization and colectomy were 20.4% and 6.1% at 3 months and 39.6% and 18.8% at the end of follow-up, respectively. Patients treated with IFX presented with significantly shorter colectomy-free survival than IVCS responders (P= 0.04, log-rank test). Both cytomegalovirus colitis and Clostridioides difficile infection (CDI) were the significant predictors of colectomy in the overall study cohort (hazard ratios of 6.57 and 4.61, respectively). There were no fatalities.Conclusions: Our real-world cohort study demonstrated that IFX is an effective therapeutic option in Korean patients with ASUC, irrespective of IFX indication. Aggressive vigilance for cytomegalovirus colitis and CDI is warranted for hospitalized patients with ASUC.

Background/aims Irreversible electroporation (IRE) is a relatively new ablation method. However, the application of IRE ablation in the treatment of biliary disease has not been attempted. A minimally invasive approach using endoscopic retrograde cholangiopancreatography (ERCP) can be a novel therapeutic modality for IRE ablation. In this study, we aimed to investigate the feasibility of endoscopic IRE for the biliary tract using an animal model. Methods A new catheter-type electrode was developed for endoscopic IRE ablation of the biliary tract. We performed ERCP and endoscopic IRE ablations in the normal common bile duct of Yorkshire pigs. The experimental setting of IRE was 500 V/cm (50 pulses, 100-µs length). The animals were sacrificed after 24 hr, and the ablated bile duct was examined. Results Well-demarcated focal color changes were observed on the mucosa of the common bile duct. The depth of change after IRE was confined to the mucosal and submucosal layers. Apoptotic changes in the bile duct were observed only around the IRE ablation area. Immunohistochemistry assay showed cell death in the bile duct along the electrode. Conclusions Endoscopic IRE ablation using ERCP was successfully performed in the common bile duct. It can be a potential option for the treatment of biliary tumors.

Conventional straight-shaped laparoscopic surgical instruments have limitations that, unlike robotic surgery, the wrist joint cannot be used. This study aimed to analyze the short-term safety and feasibility of ArtiSential (Livsmed), a new articulating laparoscopic instrument, which obviates the limitations of conventional laparoscopic surgery and allows the wrist joint to be used freely over 360° as in robotic surgery.The study included patients who underwent conventional laparoscopy or laparoscopy with the ArtiSential instrument. Patients who underwent laparoscopic gastrectomy for primary gastric adenocarcinoma in our institution were retrospectively reviewed. The groups were propensity score matched in a 11 ratio. Primary endpoint was incidence of early postoperative complication (postoperative 30-day morbidity and mortality) and secondary endpoints were operative outcomes.A total of 327 patients (147 of the conventional group and 180 of the ArtiSential group) were propensity score matched. After propensity score matching was performed, each group comprised of 122 patients. Both groups were comparable with regard to operation time, estimated blood loss, number of retrieved lymph nodes, and length of hospital stay. The ArtiSential group had a faster time to a fluid diet (2.6 ± 1.3 days vs. 2.3 ± 0.6 days,The current study showed that the use of ArtiSential is a safe and feasible option without increasing operation time, length of hospital stay, and intraoperative bleeding.

Apoptosis inhibitor of macrophage (AIM) modulates the signaling in inflammatory responses, including infection, cancer, or other immune diseases. Recent studies suggest that like interleukin-10 (IL-10), AIM is involved in alternatively activated (M2) macrophage polarization. We aimed to understand whether and how AIM is involved in IL-10-induced inhibition of inflammasome activation and resolution of inflammation. First, we demonstrated that IL-10 induced increases in mRNA and protein expression of AIM in murine bone marrow-derived macrophages (BMDM). In addition, genetic and pharmacologic inhibition of STAT3 (signal transducer and activator of transcription 3) reduced IL-10-induced AIM expression. We also found that IL-10-induced STAT3 activity enhanced the AIM promoter activity by directly binding the promoter of the AIM gene. Additionally, reduction of LPS/adenosine triphosphate (ATP)-induced IL-1β production and caspase-1 activation by IL-10 was reversed in BMDM from AIM−/− mice. Treatment of BMDM from both wild type (WT) and IL-10−/− mice with recombinant AIM showed the inhibitory effects on IL-1β and IL-18 production and caspase-1 activation. Endogenous and exogenous AIM inhibited apoptosis-associated speck-like protein containing a caspase activation and recruitment domain (ASC) speck formation. In LPS-induced acute peritonitis, inhibition of IL-1β and IL-18 production in peritoneal lavage fluid (PLF) and serum, reduction of caspase-1 activation in peritoneal macrophages, and reduction of numbers of neutrophils and peritoneal macrophages in PLF by administration of IL-10 were not evident in AIM−/− mice. Our in vitro and in vivo data reveal a novel role of AIM in the inhibition of inflammasome-mediated caspase-1 activation and IL-1β and IL-18 production.

The total iron level in the brain increases with age, and excess iron is associated with neurodegenerative diseases; however, the mechanism of brain iron deposition is unknown. In peripheral cells, the expression of hepcidin, a master regulator of iron homeostasis, is regulated by estrogen. This study aimed to determine whether hepcidin was involved in iron deposition in the brain and brain endothelial cells of estrogen-deficient aged female mice. Aged mice showed increased levels of hepcidin and ferritin in the brain and brain microvessels compared with young mice, and these levels were reduced by estrogen replacement in ovariectomized aged mice. In the brain endothelial cell line bEnd.3, the lipopolysaccharide (10 ng/mL)-induced increases of hepcidin mRNA and protein levels, the number of Prussian blue–positive cells, and free radicals were reduced after estrogen treatment. These results suggest that estrogen deficiency with an increase of hepcidin is partly responsible for iron deposition in the brain and brain endothelial cells and that hepcidin can be a target to prevent brain aging and neurodegeneration in postmenopausal women.

Background Neuroinflammation plays a pivotal role in the pathogenesis of Parkinson’s disease (PD). Thus, the development of agents that can control neuroinflammation has been suggested as a promising therapeutic strategy for PD. In the present study, we investigated whether the phosphodiesterase (PDE) 10 inhibitor has anti-inflammatory and neuroprotective effects in neuroinflammation and PD mouse models. Methods Papaverine (PAP) was utilized as a selective inhibitor of PDE10. The effects of PAP on the expression of pro-inflammatory molecules were examined in lipopolysaccharide (LPS)–stimulated BV2 microglial cells by ELISA, RT-PCR, and Western blot analysis. The effects of PAP on transcription factors were analyzed by the electrophoretic mobility shift assay, the reporter gene assay, and Western blot analysis. Microglial activation and the expression of proinflammatory molecules were measured in the LPS- or MPTP-injected mouse brains by immunohistochemistry and RT-PCR analysis. The effect of PAP on dopaminergic neuronal cell death and neurotrophic factors were determined by immunohistochemistry and Western blot analysis. To assess mouse locomotor activity, rotarod and pole tests were performed in MPTP-injected mice. Results PAP inhibited the production of nitric oxide and proinflammatory cytokines in LPS-stimulated microglia by modulating various inflammatory signals. In addition, PAP elevated intracellular cAMP levels and CREB phosphorylation. Treatment with H89, a PKA inhibitor, reversed the anti-inflammatory effects of PAP, suggesting the critical role of PKA signaling in the anti-inflammatory effects of PAP. We verified the anti-inflammatory effects of PAP in the brains of mice with LPS-induced systemic inflammation. PAP suppressed microglial activation and proinflammatory gene expression in the brains of these mice, and these effects were reversed by H89 treatment. We further examined the effects of PAP on MPTP-injected PD model mice. MPTP-induced dopaminergic neuronal cell death and impaired locomotor activity were recovered by PAP. In addition, PAP suppressed microglial activation and proinflammatory mediators in the brains of MPTP-injected mice. Conclusions PAP has strong anti-inflammatory and neuroprotective effects and thus may be a potential candidate for treating neuroinflammatory disorders such as PD.

Background Sexual violence against adolescents is prevalent worldwide and results in significant physical and mental injuries as well as loss of economic and personal potential. Urban informal settlements such as those around Nairobi, Kenya have been shown to have especially high incidences of violence. Research has shown that empowerment interventions for female adolescents can reduce sexual assault. However, these interventions have had limited testing in urban informal settlements, with young adolescents, or in coordination with complementary programs for male adolescents. Methods/design This study was a two-arm, parallel, cluster-randomized trial testing a combination of a previously-tested girls’ intervention, IMPower, and a newly revised boys’ intervention, Source of Strength. Clusters were defined as schools within the informal settlements; participants were adolescent girls and boys in class 6, generally between the ages of 10–14 at baseline. Data collection began in January 2016 and continued through December 2018. The primary outcome was the change in incidence of self-reported sexual assault among girls from baseline, compared to a life skills standard of care intervention. Secondary outcomes included experiences of physical and emotional violence, as well as determining the effects of the intervention on self-efficacy, self-esteem, and gender attitudes and beliefs, and how those effects led to changes in experience of sexual assault. For the primary outcome and several of the secondary outcomes, we used an intention to treat estimand. Discussion This was the first randomized controlled trial with longitudinal follow-up of an empowerment self-defense approach to violence prevention for adolescents in informal settlements. The large size and rigorous design supported analysis to understand multiple subgroup experiences in the hypothesized reduction in sexual assault. The study was also unique in its focus on young (10–14 years of age) adolescents and in engaging both boys and girls in separate but coordinated curriculums. The focus on a highly vulnerable and understudied population will make it a significant contribution to the literature on violence prevention. Trial registration Clinical Trials.gov #NCT02771132. Version 3.1 registered May 2017, first participant enrolled January 2017. Retrospectively registered.

Breast cancer (BC) is one of the most common cancers in both women and female dogs. Methylation changes of LINE-1 have been reported in human cancers. The aim of this study was to determine the hypomethylation of canine LINE-1 in liquid biopsies for canine mammary tumors (CMT) and to assess its diagnostic performance in human plasma. BC associated LINE-1 methylation was measured by methylation sensitive (HpaII) and insensitive (MspI) restriction enzyme digestion followed by real-time PCR using the cfDNA isolated from 300 µl of plasma. The relative level of methylated canine LINE-1 was less than 0.4 in the benign and malignant CMTs (0.29 ± 0.061 and 0.39 ± 0.066, respectively) when it was 0.92 ± 0.067 in the healthy controls. The area under the ROC curve (AUC) was significantly high in both benign and malignant tumors (0.97 and 0.93). Furthermore, this approach was also successfully implemented in a set of 26 human BCs with 10 healthy controls (AUC = 0.78). Altogether, our data suggest that the comparative approach using a dog model might be helpful to rapidly develop a new diagnostic biomarker and that the methylation of LINE-1 in cfDNA may be a good target as a diagnostic marker of both human BC and CMT.

Background Sexual violence against adolescents is prevalent worldwide and results in significant physical and mental injuries as well as loss of economic and personal potential. Urban informal settlements such as those around Nairobi, Kenya have been shown to have especially high incidences of violence. Research has shown that empowerment interventions for female adolescents can reduce sexual assault. However, these interventions have had limited testing in urban informal settlements, with young adolescents, or in coordination with complementary programs for male adolescents. Methods/design This study was a two-arm, parallel, cluster-randomized trial testing a combination of a previously-tested girls’ intervention, IMPower, and a newly revised boys’ intervention, Source of Strength. Clusters were defined as schools within the informal settlements; participants were adolescent girls and boys in class 6, generally between the ages of 10–14 at baseline. Data collection began in January 2016 and continued through December 2018. The primary outcome was the change in incidence of self-reported sexual assault among girls from baseline, compared to a life skills standard of care intervention. Secondary outcomes included experiences of physical and emotional violence, as well as determining the effects of the intervention on self-efficacy, self-esteem, and gender attitudes and beliefs, and how those effects led to changes in experience of sexual assault. For the primary outcome and several of the secondary outcomes, we used an intention to treat estimand. Discussion This was the first randomized controlled trial with longitudinal follow-up of an empowerment self-defense approach to violence prevention for adolescents in informal settlements. The large size and rigorous design supported analysis to understand multiple subgroup experiences in the hypothesized reduction in sexual assault. The study was also unique in its focus on young (10–14 years of age) adolescents and in engaging both boys and girls in separate but coordinated curriculums. The focus on a highly vulnerable and understudied population will make it a significant contribution to the literature on violence prevention. Trial registration Clinical Trials.gov #NCT02771132. Version 3.1 registered May 2017, first participant enrolled January 2017. Retrospectively registered.

Fibroblasts in the tumor microenvironment, known as cancer-associated fibroblasts (CAFs), promote the migration, invasion, and metastasis of cancer cells when they are activated through diverse processes, including post-transcriptional regulation by microRNAs (miRNAs). To identify the miRNAs that regulate CAF activation, we used NanoString to profile miRNA expression within normal mouse lung fibroblasts (LFs) and CAFs. Based on NanoString profiling, miR-196a was selected as a candidate that was up-regulated in CAFs. miR-196a-overexpressed LFs (LF-196a) promoted the migration and invasion of lung cancer cells in co-culture systems (Transwell migration and spheroid invasion assays). ANXA1 was confirmed as a direct target of miR-196a, and adding back ANXA1 to LF-196a restored the cancer cell invasion promoted by miR-196a. miR-196a increased CCL2 secretion in fibroblasts, and that was suppressed by ANXA1. Furthermore, blocking CCL2 impeded cancer spheroid invasion. In lung adenocarcinoma patients, high miR-196a expression was associated with poor prognosis. Collectively, our results suggest that CAF-specific miR-196a promotes lung cancer progression in the tumor microenvironment via ANXA1 and CCL2 and that miR-196a will be a good therapeutic target or biomarker in lung adenocarcinoma.

Parkinson’s disease (PD) is a neurodegenerative disorder characterized by motor impairments. Most PD drugs act by improving motor impairments, whereas very few drugs that efficiently recover PD-related neuropathological features, particularly α-synuclein-related toxicity, have been developed. In this study, we found that papaverine (PAP) attenuated behavioral deficits and protected against nigrostriatal dopaminergic degeneration in the subacute 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/probenecid (MPTP/P) mouse model of PD. Histological analysis of tissue dissected from mice sacrificed nearly 3 weeks after the completion of treatment revealed that PAP significantly ameliorated microglia/astrocyte activation in the striatum and substantia nigra of MPTP/P-treated mice. In addition, PAP diminished α-synuclein expression and aggregation in this model. Furthermore, PAP inhibited the phosphorylation of α-synuclein at serine 129, which may underlie the observed reduction in α-synuclein aggregation. PAP also reduced the expression of matrix metalloproteinase-3 (MMP-3), and the MMP3-positive area co-labeled with thioflavin-S. Taken together, our data suggest that PAP inhibits dopaminergic neuronal cell death and α-synuclein aggregation by suppressing neuroinflammation and MMP-3 expression in the subacute MPTP/P mouse model of PD. Accordingly, PAP may be a promising drug for the treatment of PD.

Background/Aims: Glioblastoma multiforme (GBM) is the most common primary brain tumor in adults. The defining characteristics of GBM are diffuse infiltration of tumor cells into normal brain parenchyma, rapid growth, a high degree of infiltration of microglia and macrophages, and the presence of necrosis. Microglia/macrophages are frequently found in gliomas and they extensively infiltrate GBM tissue, up to 30% of total tumor mass. However, little is known about the effect of necrotic cells (NCs) on microglia infiltration in GBM and the tumor-infiltrating microglia-induced factors in GBMs. Methods: In this study, to address whether necrosis or necrosis-exposed GBM cells affect the degree of microglia/macrophage infiltration, migration and invasion/infiltration assays were performed. Culture supernatants and nuclear extracts of CRT-MG cells treated or untreated with necrotic cells were analyzed using a chemokine array and electrophoretic mobility shift assay, respectively. Results: The presence of NCs promoted the migration/infiltration of microglia, and GBM cell line CRT-MG cells exposed to NCs further enhanced the migration and infiltration of HMO6 microglial cells. Treatment with NCs induced mRNA and protein expression of chemokines such as Monocyte Chemoattractant Protein-1 (CCL2/MCP-1) and Macrophage Inflammatory Protein-3α (CCL20/MIP-3α) in CRT-MG cells. In particular, CCL2/MCP-1 and CCL20/MIP-3α were significantly increased in NC-treated CRT-MG cells. NCs induced DNA binding of the transcription factors Nuclear Factor (NF)-κB and Activator Protein 1 (AP-1) to the CCL2/MCP-1 and CCL20/MIP-3α promoters, leading to increased CCL2/MCP-1 and CCL20/MIP-3α mRNA and protein expression in CRT-MG cells. Conclusion: These results provide evidence that NCs induce the expression of CCL2/MCP-1 and CCL20/MIP-3α in glioblastoma cells through activation of NF-κB and AP-1 and facilitate the infiltration of microglia into tumor tissues.

We previously reported that the bone morphogenetic protein (BMP) antagonist, noggin, improved the repair process with an increase in the reactive microglia/macrophage population in the ischemic brain. Since BMP plays a role in intracellular iron homeostasis via the hepcidin/ferroportin axis, and iron is required for myelination, this study was aimed to determine whether noggin affected iron status and remyelination in the brain following ischemic stroke. We further examined the effect of blocking the BMP/hepcidin pathway on reactive microglia (BV2) and myelination of oligodendroglial cells (MO3.13) to define the link between microglial iron status and myelin formation. Following the noggin infusion into the ischemic brain of mice, the induction of hepcidin and ferritin protein levels decreased, and the number of myelinated axons and myelin thickness increased at 8 weeks after ischemic stroke. Noggin repressed the increase in hepcidin and ferritin levels in BV2 exposed to lipopolysaccharide (LPS) and oxygen/glucose deprivation and reperfusion (OGD/R). When MO3.13 were exposed to the conditioned media from noggin-treated BV2 (noggin CM) during reperfusion, OGD/R-induced MO3.13 cell death was reduced. Under normal conditions, noggin CM induced myelin production with an increase in ferritin levels in MO3.13, which was reversed by the iron chelator, deferoxamine. These results indicated that noggin altered the iron status in reactive microglia from the iron-storing to the iron-releasing phenotype, which contributed to myelin synthesis by providing iron. We suggest that the BMP/hepcidin pathway can be a target for the regulation of the iron status in microglia to enhance remyelination in the ischemic brain.

Background/Aims: The signal transducer and activator of transcription 6 (STAT6) transcription factor mediates PPARγ-regulated gene expression in macrophages. However, it remains largely unknown how proximal membrane signaling events initiated by apoptotic cell recognition upregulate PPARγ expression and activate the lung homeostatic program. Methods: The STAT6 inhibitor AS1517499 was used to determine the role of STAT6 in mediating PPARγ activity, anti-inflammatory effects, and anti-fibrotic effects induced by apoptotic cell instillation after bleomycin treatment into C57BL/6 mice. Bronchoalveolar lavage fluid, alveolar macrophages and lungs were harvested at days 2, 7, and 14 and then analyzed by real-time PCR, immunoblotting, ELISA, immunocytochemistry and immunohistochemistry assays. Results: Our data demonstrate that apoptotic cell instillation after bleomycin results in prolonged enhancement of STAT6 phosphorylation in alveolar macrophages and lung. Co-administration of the STAT6 inhibitor, AS1517499, reversed the enhanced PPARγ expression and activity induced by apoptotic cell instillation after bleomycin treatment. By reducing the expression of PPARγ target genes, including CD36, macrophage mannose receptor, and arginase 1, AS1517499 inhibited efferocytosis and restored pro-inflammatory cytokine expression, neutrophil recruitment, protein levels, hydroxyproline content, and expression of fibrosis markers, including type 1 collagen α2, fibronectin, and α-smooth muscle actin. STAT6 inhibition reversed the expression profile of hepatocyte growth factor and interleukin-10. Conclusion: These results indicate that prolonged STAT6 activation following one-time apoptotic cell instillation facilitates continuous PPARγ activation, resulting in the resolution of bleomycin-induced lung inflammation and fibrosis.

The invasion of activated fibroblasts is a key mechanism of tissue fibrosis pathology. The recognition and uptake of apoptotic cells can induce the anti-fibrogenic programming of macrophages. We demonstrate that after interacting with apoptotic cells, macrophages secrete bioactive molecules that antagonize TGF-β1-induced increases in myofibroblast (fibroproliferative) phenotypic markers and reduce the enhanced invasive capacity of TGF-β1- or EGF-treated mouse lung fibroblasts (MLg). Furthermore, numerous treatment strategies prevented the anti-fibrotic effects of conditioned media, including transfection of macrophages with COX-2 or RhoA siRNAs or treatment of MLg cells with receptor antagonists for prostaglandin E2 (PGE2), PGD2, or hepatocyte growth factor (HGF). Additionally, administration of apoptotic cells in vivo inhibited the bleomycin-mediated invasive capacity of primary fibroblasts, as well as adhesion and extracellular matrix protein mRNA expression. These data suggest that the anti-fibrogenic programming of macrophages by apoptotic cells can be used as a novel tool to control the progressive fibrotic reaction.

To investigate the role of antithymocyte globulin (ATG)-containing reduced-intensity conditioning (RIC) in hematopoietic cell transplantation (HCT) from unrelated (UD) or haploidentical family donors (HFD), we conducted a phase 2 trial of 237 patients (age range, 16 to 69 years) with acute myeloid leukemia (AML) in remission. Patients undergoing UD-HCT (n = 93) or HFD-HCT (n = 59) received RIC comprising busulfan, fludarabine, and ATG, 9 mg/kg, whereas those undergoing HCT from matched sibling donors (MSD, n = 85) received myeloablative busulfan and cyclophosphamide conditioning or aforementioned RIC with ATG, 4.5 mg/kg. For graft-versus-host disease (GVHD) prophylaxis, cyclosporine and methotrexate were administered. The median follow-up period was 44.7 months after HCT for 161 survivors. For UD-HCT versus HFD-HCT, there were no significant differences in leukemia recurrence, nonrelapse mortality, relapse-free survival, grades 2 to 4 acute GVHD, and moderate-to-severe chronic GVHD. Furthermore, when the outcomes of UD-HCT and HFD-HCT were combined and compared with those of MSD-HCT, there were no significant differences in leukemia recurrence (3-year cumulative incidence, 30% versus 29%), nonrelapse mortality (3-year cumulative incidence, 7% versus 8%), relapse-free survival (3-year estimate, 63% versus 63%), and grades 2 to 4 acute GVHD (120-day cumulative incidence, 16% versus 13%). Moderate-to-severe chronic GVHD, however, occurred less frequently in UD/HFD-HCT (2-year cumulative incidence, 22% versus 40%; P = .006). The addition of ATG to conditioning regimen was a significant predictor for less chronic GVHD (subdistribution hazard ratio, .59). In AML in remission, UD/HFD-HCT after ATG-containing RIC achieved leukemia control equivalent to that of MSD-HCT. Despite HLA disparity in UD/HFD-HCT, chronic GVHD occurred less frequently after ATG-containing RIC, suggesting a strong GVHD-modulating effect of ATG.

Background/Aims: CD4+ T cells are a critical component of the adaptive immune response. While the mechanisms controlling the differentiation of the Th1, Th17, and regulatory T cell subsets from naive CD4+ T cells are well described, the factors that induce Th2 differentiation are still largely unknown. Methods: The effects of treatment with exogenous H 2 O 2 on STAT-6 phosphorylation and activation in T cells were examined by immunoblotting, immunofluorescence and gel shift assay. Anti-CD3 antibody and methyl-β-cyclodextrin were utilized to induce lipid raft assembly and to investigate the involvement of lipid rafts, respectively. Results: Jurkat and EL-4 T cells that were exposed to H 2 O 2 showed rapid and strong STAT-6 phosphorylation, and the extent of STAT-6 phosphorylation was enhanced by co-treatment with anti-CD3 antibody. The effect of H 2 O 2 on STAT-6 phosphorylation and translocation was inhibited by disruption of lipid rafts. STAT-6 activation in response to H 2 O 2 treatment regulated IL-4 gene expression, and this response was strengthened by treatment with anti-CD3. Conclusion: Our results indicate that reactive oxygen species such as H 2 O 2 can act on upstream and initiating factors for activation of STAT-6 in T cells and contribute to formation of a positive feedback loop between STAT-6 and IL-4 in the Th2 differentiation process.

Prostate cancer (PC) is the second most common cancer in men and is the fifth leading cause of cancer-related deaths worldwide. Additionally, there is concern for overdiagnosis and overtreatment of PC. Thus, selection of an appropriate candidate for active surveillance as well as more accurate and less invasive tools for monitoring advanced PC is required. Circulating tumor cells (CTCs) have emerged as a liquid biopsy tool; there have been several reports on its role, technologies, and applications to various cancers, including PC. Liquid biopsy using CTCs has been gaining attention as a minimal invasive tool for investigation of biomarkers and for prognosis and assessment of response to therapies in patients with PC. Because of the lower invasiveness of liquid biopsy using CTCs, it can be performed more frequently; accordingly, personalized disease status can be successively determined at serial time points. CTC analysis enables detection of genomic alterations, which is drug-targetable, and it is a potential tool for monitoring response to therapeutic agents in patients with PC. This review focuses on the characteristics, technologies for analysis, and advantages and disadvantages of CTCs as a liquid biopsy tool and their application in PC. Finally, we propose future directions of CTCs.

Excessive exercise causes the loss of inherent physiological function of the muscles through skeletal muscle lesioning. This damage can induce various...

Circulating tumor cells (CTCs) are cancer cells that have been shed from a primary tumor and circulate in the bloodstream during progression of cancer. They may thus serve as circulating biomarkers that can predict, diagnose and guide therapy. Moreover, phenotypic and genotypic analysis of CTCs can facilitate prospective assessment of mutations and enable personalized treatment. A number of methodologies based on biological and physical differences between circulating tumor and non-tumor cells have been developed over the past few years. However, these methods did not have sufficient sensitivity or specificity. In this work, a remote analysis protocol was designed using motion microscopy that amplifies cellular micro motions in a captured video by re-rendering small motions to generate extreme magnified visuals to detect dynamic motions that are not otherwise visible by naked eye. Intriguingly, motion microscopy demonstrated fluctuations around breast tumor cells, which we referred to herein as cellular trail. Phenomena of cellular trail mostly emerged between 0.5 and 1.5 Hz on amplified video images. Interestingly, cellular trails were associated with cell surface proteins and flow rates rather than mitochondrial activity. Moreover, cellular trails were present only around circulating tumor cells from individuals with breast cancer under conditions of 20–30 μm/s and 0.5–1.5 Hz. Thus, motion microscopy based CTC detection method can offer a valuable supplementary diagnostic tool for assessment of drug efficacy and identifying physical characteristics of tumor cells for further research.

Importance Ocular symptoms represent approximately 2% to 3% of all emergency department (ED) visits. These disease processes may progress to permanent vision loss if not diagnosed and treated quickly. Use of ocular point-of-care ultrasonography (POCUS) may be effective for early and accurate detection of ocular disease. Objective To perform a large-scale, multicenter study to determine the utility of POCUS for diagnosing retinal detachment, vitreous hemorrhage, and vitreous detachment in the ED. Design, Setting, and Participants A prospective diagnostic study was conducted at 2 academic EDs and 2 county hospital EDs from February 3, 2016, to April 30, 2018. Patients who were eligible for inclusion were older than 18 years; were English- or Spanish-speaking; presented to the ED with ocular symptoms with concern for retinal detachment, vitreous hemorrhage, or vitreous detachment; and underwent an ophthalmologic consultation that included POCUS. Patients with ocular trauma or suspicion for globe rupture were excluded. The accuracy of the ultrasonographic diagnosis was compared with the criterion standard of the final diagnosis of an ophthalmologist who was masked to the POCUS findings. Seventy-five unique emergency medicine attending physicians, resident physicians, and physician assistants performed ocular ultrasonography. Exposure Point-of-care ultrasonography performed by an emergency medicine attending physician, resident physician, or physician assistant. Main Outcomes and Measures Sensitivity and specificity of POCUS in identifying retinal detachment, vitreous hemorrhage, and vitreous detachment in patients presenting to the ED with ocular symptoms. Results Two hundred twenty-five patients were enrolled. Of these, the mean age was 51 years (range, 18-91 years) and 135 (60.0%) were men; ophthalmologists diagnosed 47 (20.8%) with retinal detachment, 54 (24.0%) with vitreous hemorrhage, and 34 (15.1%) with vitreous detachment. Point-of-care ultrasonography had an overall sensitivity of 96.9% (95% CI, 80.6%-99.6%) and specificity of 88.1% (95% CI, 81.8%-92.4%) for diagnosis of retinal detachment. For diagnosis of vitreous hemorrhage, the sensitivity of POCUS was 81.9% (95% CI, 63.0%-92.4%) and specificity was 82.3% (95% CI, 75.4%-87.5%). For vitreous detachment, the sensitivity was 42.5% (95% CI, 24.7%-62.4%) and specificity was 96.0% (95% CI, 91.2%-98.2%). Conclusions and Relevance These findings suggest that emergency medicine practitioners can use POCUS to accurately identify retinal detachment, vitreous hemorrhage, and vitreous detachment. Point-of-care ultrasonography is not intended to replace the role of the ophthalmologist for definitive diagnosis of these conditions, but it may serve as an adjunct to help emergency medicine practitioners improve care for patients with ocular symptoms.

We aimed to improve the tumor localization system using radiofrequency identification (RFID) technology used during laparoscopic surgery for gastric and colorectal cancer. To this end, we developed a detection algorithm and designed improvement for the RFID clip. To evaluate the proposed system, a swine-based animal study was conducted, followed by experiments on porcine stomachs and colons using the EASIE-R simulator. The success rates of endoscopic clipping, detection time, and detection accuracy, which is the distance between the detection point and RFID tag, were measured. Results of the in vivo swine animal study showed success in all three clippings and detections of the RFID clips. Results of the 60 RFID endoclip attempts using the EASIE-R simulator showed a total clipping success rate of 85.0% (n = 51/60; stomach, 83.3%, n = 25/30; colon, 86.7%, n = 26/30). The median detection times were 29.2 s for the stomach and 25.5 s for the colon. The median detection accuracy was 4.0 mm for the stomach and 4.5 mm for the colon. We confirmed that the proposed RFID-based system showed improvements over the system of a previous study. This RFID-based system is effective at localizing gastric and colorectal tumors.

Background Recent evidence suggests that reactive astrocytes play an important role in neuroinflammation and neurodegenerative diseases. Thus, controlling astrocyte reactivity has been suggested as a promising strategy for treating neurodegenerative diseases. In the present study, we investigated whether a matrix metalloproteinase (MMP)-8 inhibitor, M8I, could control neuroinflammation in lipoteichoic acid (LTA)-stimulated rat primary astrocytes. Methods The effects of M8I on the expression of inducible nitric oxide synthase, cytokines, and MMPs were examined in LTA-stimulated rat primary astrocytes by ELISA, RT-PCR, and Western blot analysis. The effects of M8I on reactive oxygen species (ROS) generation and phase II antioxidant enzyme expression were examined by the DCF-DA assay, RT-PCR, and Western blot analysis. The detailed molecular mechanisms underlying the anti-inflammatory and antioxidant effects of M8I were analyzed by the electrophoretic mobility shift assay, the reporter gene assay, Western blot, and RT-PCR analysis. Results Treatment with LTA, a major cell wall component of Gram-positive bacteria, led to astrocyte activation and induced the expression of inflammatory molecules such as iNOS, COX-2, and pro-inflammatory cytokines. In addition, LTA induced the expression of MMPs such as MMP-1, MMP-3, MMP-8, MMP-9, and MMP-13 in rat primary astrocytes. Based on previous reports showing that MMP-8 plays a role as a proinflammatory mediator in microglia, we investigated whether MMP-8 is also involved in inflammatory reactions of reactive astrocytes. We found that treatment of astrocytes with M8I significantly inhibited LTA-induced expression of iNOS, TNF-α, IL-1β, IL-6, and TLR-2. In addition, M8I inhibited LTA-induced NF-κB, MAP kinase, and Akt activities, while it increased the anti-inflammatory PPAR-γ activities. Moreover, M8I showed antioxidant effects by suppressing ROS production in LTA- or H2O2-stimulated astrocytes. Interestingly, M8I increased the expression of phase II antioxidant enzymes such as hemeoxygenase-1, NQO1, catalase, and MnSOD by modulating the Nrf2/ARE signaling pathway. Conclusions The data collectively suggest the therapeutic potential of an MMP-8 inhibitor in neuroinflammatory disorders that are associated with astrocyte reactivity. Electronic supplementary material The online version of this article (10.1186/s12974-018-1363-6) contains supplementary material, which is available to authorized users.

Soft tissue masses are common in the pediatric population and may represent a broad range of conditions. Point-of-care ultrasound can be used for rapid visualization and assessment of soft tissue masses in the emergency setting. We report a case of a pediatric head and neck mass in which point-of-care ultrasound was used to identify an infantile parotid hemangioma.

Apoptotic cell clearance by phagocytes is essential in tissue homeostasis. We demonstrated that conditioned medium (CM) from macrophages exposed to apoptotic cancer cells inhibits the TGFβ1-induced epithelial–mesenchymal transition (EMT), migration, and invasion of cancer cells. Apoptotic 344SQ (ApoSQ) cell-induced PPARγ activity in macrophages increased the levels of PTEN, which was secreted in exosomes. Exosomal PTEN was taken up by recipient lung cancer cells. ApoSQ-exposed CM from PTEN knockdown cells failed to enhance PTEN in 344SQ cells, restore cellular polarity, or exert anti-EMT and anti-invasive effects. The CM that was deficient in PPARγ ligands, including 15-HETE, lipoxin A4, and 15d-PGJ2, could not reverse the suppression of PPARγ activity or the PTEN increase in 344SQ cells and consequently failed to prevent the EMT process. Moreover, a single injection of ApoSQ cells inhibited lung metastasis in syngeneic immunocompetent mice with enhanced PPARγ/PTEN signaling both in tumor-associated macrophages and in tumor cells. PPARγ antagonist GW9662 reversed the signaling by PPARγ/PTEN; the reduction in EMT-activating transcription factors, such as Snai1 and Zeb1; and the antimetastatic effect of the ApoSQ injection. Thus, the injection of apoptotic lung cancer cells may offer a new strategy for the prevention of lung metastasis.

Previously, we demonstrated that growth arrest-specific protein 6 (Gas6)/Axl or Mer signaling inhibited the transforming growth factor (TGF)-&beta, 1-induced epithelial&ndash, mesenchymal transition (EMT) in lung epithelial cells. Hepatocyte growth factor (HGF) has also been shown to inhibit TGF-&beta, 1-induced changes in EMT markers. Here, we examined whether Gas6 signaling can induce the production of HGF and c-Met in lung alveolar epithelial cells to mediate the inhibition of EMT and to inhibit the migration and invasion of epithelial cells. The inhibition of the RhoA/Rho kinase pathway, using either a RhoA-targeted small interfering RNA (siRNA) or the Rho kinase pharmacologic inhibitor Y27362, prevented the inhibition of TGF-&beta, 1-induced EMT in LA-4 cells and primary alveolar type II (AT II) epithelial cells. The c-Met antagonist PHA-665752 also blocked the anti-EMT effects associated with Gas6. Moreover, treatment with Y27362 or PHA-665752 prevented the Gas6-mediated inhibition of TGF-&beta, 1-induced migration and invasion. Our data provided evidence that the RhoA-dependent production of HGF and c-Met mediated the Gas6-induced inhibition of EMT, migration and invasion in lung alveolar epithelial cells. Thus, Gas6/Axl and Mer/RhoA signaling may be necessary for the maintenance of homeostasis in the alveolar epithelium, via HGF and c-Met.

Background Chronic obstructive pulmonary disease (COPD) is a debilitating lung disease. To date, a large number of clinical studies have been conducted to investigate the association between genetic variations and COPD. However, little is known regarding the genetic susceptibility of Koreans to this disease. MER receptor tyrosine kinase (MERTK) plays important roles in the inhibition of inflammation and in the clearance of apoptotic cells. Here, we investigated the association between genetic variations in MERTK and the development of COPD in Koreans. Methods We conducted genetic analysis of MERTK using genomic DNA samples from 87 patients with COPD and 88 healthy controls and compared the frequency of each variation or haplotype between the patient and control groups. Subsequently, the effect of each variation was evaluated using in vitro assays. Results Ten variations were identified in this study, four of them for the first time. In addition, we found that the frequency of each variation or haplotype was comparable between the patient and control groups. However, we observed that the frequency for the wild-type haplotype was higher in the control group, compared to that in the group of patients with COPD, in the subgroup analysis of current smokers, although the difference was not statistically significant (P = 0.080). In in vitro assays, we observed that none of the variations affected the activity of the promoter or the expression of MERTK. Conclusion Our findings indicate that the susceptibility to COPD is not related to the genetic variations or haplotypes of MERTK in Koreans., Graphical Abstract

Ginsenoside Rh3 is a bacterial metabolite of Rg5, which is the main constituent of heat-processed ginseng. The present study was undertaken to examine the anti-inflammatory effect of ginsenoside Rh3 in lipopolysaccharide (LPS)-stimulated microglia. Rh3 inhibits the expressions of inducible nitric oxide synthase (iNOS) and proinflammatory cytokines, such as tumor necrosis factor (TNF)-α and interleukin (IL)-6, at mRNA and protein levels, while Rh3 enhanced anti-inflammatory hemeoxygenase-1 expression. Moreover, Rh3 inhibited nuclear factor-κB (NF-κB) by upregulation of sirtuin 1 (SIRT1) and enhanced Nrf2 DNA-binding activities. Analysis of signaling pathways revealed that Rh3 enhanced the phosphorylation of 5'-adenosine monophosphate-activated protein kinase (AMPK) and inhibited Akt and janus kinase 1 (JAK1)/signal transducer and activator of transcription 1 (STAT1) induced by LPS. By treatment of BV2 cells with AICAR (a pharmacological activator of AMPK), we found that AMPK is an upstream regulator of phosphatidylinositol 3-kinase (PI3K)/Akt and JAK1/STAT1. Furthermore, AMPK knockdown experiments demonstrated the anti-inflammatory role of AMPK in LPS/Rh3-treated BV2 microglia. Our data collectively suggest that Rh3 exerts an anti-inflammatory effect in microglia by modulating AMPK and its downstream signaling pathways.

In this experiment, agarose degradation was conducted under identical hydrolysis conditions, but different substrate and enzyme concentrations. Hydrolysis products i.e., agaro-oligosaccharide were monitored using viscometric and reducing end assay. Hydrolysis yields were found to range between 3.73 and 20.51%. Thereafter, agaro-oligosaccharides with different degree hydrolysis were subjected to various physicochemical analyses and in vitro antioxidant assays. Agaro-oligosaccharides were shown to exhibit high solubility percentages (59.84–86.78%), water absorption capacities (1.5–3.0 g/g) and oil absorption capacities (1.8–4.6 g/g). Agaro-oligasaccharides were also shown to exhibit considerable DPPH and ABTS+ radical scavenging and ferric reducing antioxidant activities; however, this effect depended on the degree hydrolysis.

Extracellular matrix (ECM) plays a critical role in the provision of the necessary microenvironment for the proper regeneration of the cardiac tissue. However, specific mechanisms that lead to ECM-mediated cardiac regeneration are not well understood. To elucidate the potential mechanisms, we investigated ultra-structures of the cardiac ECM using electron microscopy. Intriguingly, we observed large quantities of micro-vesicles from decellularized right atria. RNA and protein analyses revealed that these contained exosomal proteins and microRNAs (miRNAs), which we referred to herein as ECM-derived extracellular vesicles (ECM-EVs). One particular miRNA from ECM-EVs, miR-199a-3p, promoted cell growth of isolated neonatal cardiomyocytes and sinus nodal cells by repressing homeodomain-only protein (HOPX) expression and increasing GATA-binding 4 (Gata4) acetylation. To determine the mechanisms, we knocked down Gata4 and showed that miR-199a-3p actions required Gata4 for cell proliferation in isolated neonatal cardiomyocytes and sinus nodal cells. To further explore the role of this miRNA, we isolated neonatal cardiac cells and recellularized into atrial ECM, referred here has engineered atria. Remarkably, miR-199a-3p mediated the enrichment of cardiomyocyte and sinus nodal cell population, and enhanced electrocardiographic signal activity of sinus nodal cells in the engineered atria. Importantly, antisense of miRNA (antagomir) against miR-199a-3p was capable of abolishing these actions of miR-199a-3p in the engineered atria. We further showed in Ang II-infused animal model of sinus nodal dysfunction that miR-199-3p-treated cardiac cells remarkably ameliorated and restored the electrical activity as shown by normalization of the ECG, in contrast to untreated cells, which did not show electrical recovery. In conclusion, these results provide clear evidence of the critical role of ECM, in not only providing a scaffold for cardiac tissue growth, but also in promoting atrial electrical function through ECM-derived miR-199a-3p.

We previously reported that bone morphogenetic proteins (BMPs) and their endogenous antagonist noggin are expressed in the brain weeks after an ischemic insult. Here, to define their roles in ischemic brain tissue repair and remodeling, we infused recombinant BMP7 or noggin into the ipsilateral ventricle of mice for 2weeks starting 2weeks after transient middle cerebral artery occlusion (MCAO). Four weeks after MCAO, we measured ischemic brain volume, functional recovery, and molecules related to neurogenesis and angiogenesis such as synaptophysin, GAP-43, and VEGF. Noggin-treated mice but not BMP7-treated mice showed preserved ipsilateral brain volume and reduced neurological deficits compared with artificial cerebrospinal fluids (aCSF)-treated mice. Noggin treatment also decreased glial scar thickness, increased levels of GAP-43 and VEGF protein, and increased the number of Iba1-positive activated microglia in the ipsilateral brain. Furthermore, noggin treatment decreased M1 markers (IL-1β, TNF-α, IL-12, CCL2 and CD86) and increased M2 markers (IL-1ra, IL-10, arginase 1, CD206 and Ym1) of activated microglia, suggesting a shift from M1 to M2 phenotypes. These results suggest that noggin improves functional recovery from ischemic stroke and enhances alternatively activated microglia, thereby promoting tissue repair and remodeling.

Recognition of apoptotic cells by macrophages is crucial for resolution of inflammation, immune tolerance, and tissue repair. Cyclooxygenase-2 (COX-2)/prostaglandin E2 (PGE2) and hepatocyte growth factor (HGF) play important roles in the tissue repair process. We investigated the characteristics of macrophage COX-2 and PGE2expression mediated by apoptotic cells and then determined how macrophages exposed to apoptotic cellsin vitroandin vivoorchestrate the interaction between COX-2/PGE2and HGF signaling pathways. Exposure of RAW 264.7 cells and primary peritoneal macrophages to apoptotic cells resulted in induction of COX-2 and PGE2. The COX-2 inhibitor NS-398 suppressed apoptotic cell-induced PGE2production. Both NS-398 and COX-2-siRNA, as well as the PGE2receptor EP2 antagonist, blocked HGF expression in response to apoptotic cells. In addition, the HGF receptor antagonist suppressed increases in COX-2 and PGE2induction. Thein vivorelevance of the interaction between the COX-2/PGE2and HGF pathways through a positive feedback loop was shown in cultured alveolar macrophages followingin vivoexposure of bleomycin-stimulated lungs to apoptotic cells. Our results demonstrate that upregulation of the COX-2/PGE2and HGF in macrophages following exposure to apoptotic cells represents a mechanism for mediating the anti-inflammatory and antifibrotic consequences of apoptotic cell recognition.

Neuroinflammation plays an important role in the progression of various neurodegenerative diseases. In this study, we investigated the anti-inflammatory effects of lonchocarpine, a natural compound isolated from Abrus precatorius, under in vitro and in vivo neuroinflammatory conditions induced by challenge with lipopolysaccharide (LPS)- or polyinosinic-polycytidylic acid (poly(I:C)). Lonchocarpine suppressed the expression of iNOS and proinflammatory cytokines in LPS or poly(I:C)-stimulated BV2 microglial cells. These anti-inflammatory effects were verified in brains of mice with systemic inflammation induced by administration of LPS or poly(I:C). Lonchocarpine reduced the number of Iba-1-positive activated microglia, and suppressed the mRNA expression of various proinflammatory markers in the cortex of LPS- or poly(I:C)-injected mice. Molecular mechanistic experiments showed that lonchocarpine inhibited NF-κB activity by reducing the phosphorylation and degradation of IκBα in LPS- or poly(I:C)-stimulated BV2 cells. Analysis of further upstream signaling pathways in LPS-stimulated microglia showed that lonchocarpine inhibited the phosphorylation of IκB kinase and TGFβ-activated kinase 1 (TAK1). Moreover, lonchocarpine suppressed the interaction of myeloid differentiation factor 88 (MyD88) and intereleukin-1 receptor-associated kinase 4 (IRAK4). These data suggest that toll-like receptor 4 downstream signals such as MyD88/IRAK4-TAK1-NF-κB are at least partly involved in the anti-inflammatory mechanism of lonchocarpine in LPS-stimulated microglia. Its strong anti-inflammatory effects may make lonchocarpine an effective preventative drug for neuroinflammatory disorders that are associated with systemic inflammation.

The optimum method of donor natural killer cell infusion (DNKI) after allogeneic hematopoietic cell transplantation (HCT) remains unclear. Fifty-one patients (age range, 19 years to 67 years) with refractory acute leukemia underwent HLA-haploidentical HCT and underwent DNKI on days 6, 9, 13, and 20 of HCT. Median DNKI doses were .5, .5, 1.0, and 2.0 × 108/kg cells, respectively. During DNKI, 33 of the 45 evaluated patients (73%) developed fever (>38.3°C) along with weight gain (median, 13%; range, 2% to 31%) and/or hyperbilirubinemia (median, 6.2 mg/dL; range, 1.0 mg/dL to 35.1 mg/dL); the toxicity was reversible in 90% of patients. After transplantation, we observed cumulative incidences of neutrophil engraftment (≥500/µL), grade 2 to 4 acute graft-versus-host disease (GVHD), chronic GVHD, and nonrelapse mortality of 84%, 28%, 30%, and 16%, respectively. The leukemia complete remission rate was 57% at 1 month after HCT and 3-year cumulative incidence of leukemia progression was 75%. When analyzed together with our historical cohort of 40 patients with refractory acute leukemia who underwent haploidentical HCT and DNKI on days 14 and 21 only, higher expression of NKp30 (>90%) on donor NK cells was an independent predictor of higher complete remission (hazard ratio, 5.59) and less leukemia progression (hazard ratio, .57). Additional DNKI on days 6 and 9 was not associated with less leukemia progression (75% versus 55%).

Glioblastoma multiforme (GBM) is the most common primary intracranial tumor in adults and has poor prognosis. Diffuse infiltration into normal brain parenchyma, rapid growth, and the presence of necrosis are remarkable hallmarks of GBM. However, the effect of necrotic cells on GBM growth and metastasis is poorly understood at present. In this study, we examined the biological significance of necrotic tissues by exploring the molecular mechanisms underlying the signaling network between necrotic tissues and GBM cells. The migration and invasion of the GBM cell line CRT-MG was significantly enhanced by treatment with necrotic cells, as shown by assays for scratch wound healing and spheroid invasion. Incubation with necrotic cells induced IL-8 secretion in CRT-MG cells in a dose-dependent manner. In human GBM tissues, IL-8 positive cells were mainly distributed in the perinecrotic region, as seen in immunohistochemistry and immunofluorescence analysis. Necrotic cells induced NF-κB and AP-1 activation and their binding to the IL-8 promoter, leading to enhanced IL-8 production and secretion in GBM cells. Our data demonstrate that when GBM cells are exposed to and stimulated by necrotic cells, the migration and invasion of GBM cells are enhanced and facilitated via NF-κB/AP-1 mediated IL-8 upregulation.

Apoptotic cell clearance results in the release of growth factors and the action of signaling molecules involved in tissue homeostasis maintenance. Here, we investigated whether and how macrophages programmed by apoptotic cells inhibit the TGF-β1-induced Epithelial-mesenchymal transition (EMT) process in lung alveolar epithelial cells. Treatment with conditioned medium derived from macrophages exposed to apoptotic cells, but not viable or necrotic cells, inhibited TGF-β1-induced EMT, including loss of E-cadherin, synthesis of N-cadherin and α-smooth muscle actin, and induction of EMT-activating transcription factors, such as Snail1/2, Zeb1/2, and Twist1. Exposure of macrophages to cyclooxygenase (COX-2) inhibitors (NS-398 and COX-2 siRNA) or RhoA/Rho kinase inhibitors (Y-27632 and RhoA siRNA) and LA-4 cells to antagonists of prostaglandin E2 (PGE2) receptor (EP4 [AH-23848]), PGD2 receptors (DP1 [BW-A868C] and DP2 [BAY-u3405]), or the hepatocyte growth factor (HGF) receptor c-Met (PHA-665752), reversed EMT inhibition by the conditioned medium. Additionally, we found that apoptotic cell instillation inhibited bleomycin-mediated EMT in primary mouse alveolar type II epithelial cells in vivo. Our data suggest a new model for epithelial cell homeostasis, by which the anti-EMT programming of macrophages by apoptotic cells may control the progressive fibrotic reaction via the production of potent paracrine EMT inhibitors.

Mer receptor tyrosine kinase (Mer) regulates macrophage activation and promotes apoptotic cell clearance. Mer activation is regulated through proteolytic cleavage of the extracellular domain. To determine if membrane-bound Mer is cleaved during bleomycin-induced lung injury, and, if so, how preventing the cleavage of Mer enhances apoptotic cell uptake and down-regulates pulmonary immune responses. During bleomycin-induced acute lung injury in mice, membrane-bound Mer expression decreased, but production of soluble Mer and activity as well as expression of disintegrin and metalloproteinase 17 (ADAM17) were enhanced . Treatment with the ADAM inhibitor TAPI-0 restored Mer expression and diminished soluble Mer production. Furthermore, TAPI-0 increased Mer activation in alveolar macrophages and lung tissue resulting in enhanced apoptotic cell clearance in vivo and ex vivo by alveolar macrophages. Suppression of bleomycin-induced pro-inflammatory mediators, but enhancement of hepatocyte growth factor induction were seen after TAPI-0 treatment. Additional bleomycin-induced inflammatory responses reduced by TAPI-0 treatment included inflammatory cell recruitment into the lungs, levels of total protein and lactate dehydrogenase activity in bronchoalveolar lavage fluid, as well as caspase-3 and caspase-9 activity and alveolar epithelial cell apoptosis in lung tissue. Importantly, the effects of TAPI-0 on bleomycin-induced inflammation and apoptosis were reversed by coadministration of specific Mer-neutralizing antibodies. These findings suggest that restored membrane-bound Mer expression by TAPI-0 treatment may help resolve lung inflammation and apoptosis after bleomycin treatment.

Allergic asthma is characterized by infiltration of eosinophils, elevated Th2 cytokine levels, airway hyperresponsiveness, and IgE. In addition to eosinophils, mast cells, and basophils, a variety of cytokines are also involved in the development of allergic asthma. The pivotal role of eosinophils in the progression of the disease has been a subject of controversy. To determine the role of eosinophils in the progression of airway inflammation, we sensitized and challenged BALB/c wild-type (WT) mice and eosinophil-deficient ΔdblGATA mice with ovalbumin (OVA) and analyzed different aspects of inflammation. We observed increased eosinophil levels and a Th2-dominant response in OVA-challenged WT mice. In contrast, eosinophil-deficient ΔdblGATA mice displayed an increased proportion of mast cells and a Th17-biased response following OVA inhalation. Notably, the levels of IL-33, an important cytokine responsible for Th2 immune deviation, were not different between WT and eosinophil-deficient mice. We also demonstrated that mast cells induced Th17-differentiation via IL-33/ST2 stimulation in vitro. These results indicate that eosinophils are not essential for the development of allergic asthma and that mast cells can skew the immune reaction predominantly toward Th17 responses via IL-33 stimulation.

Bone morphogenetic proteins (BMPs) and their antagonists have roles in scar formation and regeneration after central nervous system injuries. However, temporal changes in their expression during astroglial scar formation in the ischemic brain are unknown. Here, we examined protein levels of BMP2, BMP7, and their antagonist noggin in the ischemic brain up to 4 weeks after experimental stroke in mice. BMP2 and BMP7 levels were increased from 1 to 4 weeks in the ischemic brain, and their expression was associated with astrogliosis. BMP7 expression was more intense and co-localized in reactive astrocytes in the ischemic subcortex at 1 week. Noggin expression began to increase after 2 weeks and was further increased at 4 weeks only in the ischemic subcortex, but the intensity was weak compared to the intensity of BMPs. Noggin was co-localized mainly in activated microglia. These findings show that expression of BMPs and noggin differed over time, in intensity and in types of cell, and suggest that BMPs and noggin have different roles in the processes of glial scar formation and neurorestoration in the ischemic brain.

The receptor protein tyrosine kinases Tyro3, Axl, and Mer play important roles in macrophage function. Study of the induction of HGF during the interaction of macrophages with apoptotic cells shows that only Mer is responsible for mediating transcriptional HGF production through a RhoA-dependent pathway., The TAM receptor protein tyrosine kinases Tyro3, Axl, and Mer play important roles in macrophage function. We investigated the roles of the TAM receptors in mediating the induction of hepatocyte growth factor (HGF) during the interaction of macrophages with apoptotic cells. Mer-specific neutralizing antibody, small interfering RNA (siRNA), and a recombinant Mer protein (Mer/Fc) inhibited HGF mRNA and protein expression, as well as activation of RhoA, Akt, and specific mitogen-activated protein (MAP) kinases in response to apoptotic cells. Inhibition of Axl or Tyro3 with specific antibodies, siRNA, or Fc-fusion proteins did not prevent apoptotic cell–induced HGF mRNA and protein expression and did not inhibit activation of the postreceptor signaling molecules RhoA and certain MAP kinases, including extracellular signal-regulated protein kinase and c-Jun NH2-terminal kinase. However, Axl- and Tyro3-specific blockers did inhibit the activation of Akt and p38 MAP kinase in response to apoptotic cells. In addition, none of the TAM receptors mediated the effects of apoptotic cells on transforming growth factor-β or epidermal growth factor mRNA expression. However, they were involved in the induction of vascular endothelial growth factor mRNA expression. Our data provide evidence that when macrophages interact with apoptotic cells, only Mer of the TAM-family receptors is responsible for mediating transcriptional HGF production through a RhoA-dependent pathway.