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Acute lung injury (ALI) is a severe form of sepsis that is associated with a high rate of morbidity and death in critically ill individuals. The emergence of ALI is the result of several factors at work. Case mortality rates might range from 40% to 70%. Researchers have discovered that epigenetic alterations are important in the pathophysiology of ALI and that using epigenetic inhibitors may help reduce symptoms. In embryonic development, circadian rhythm, the cell cycle, and cancer, methylation of m6A seems to be relevant all along the way. According to recent research, posttranscriptional methylation is a key player in the development of alveolar lymphoma. In this study, we clustered ALI based on m6A-related factors, analyzed different classes of immune cell enrichment and inflammatory cytokine expression, screened clustered differential genes for ALI to construct coexpression networks, screened key ALI genes potentially regulated by m6A modifications, and then typed the disease based on key genes to compare the consistency of different clustering results. Our findings have revealed a hitherto undiscovered prognostic sign and a therapeutic target for ALI therapy in m6A and immune invading cells, respectively.

Hepatitis E virus (HEV) infection has many extrahepatic manifestations as well as liver symptoms. Multiple studies have shown that HEV infection has symptoms related to the nervous system, kidneys, cryoglobulinemia, hematological system, reproductive system, autoimmunity and pancreas. Hence, HEV infection should be considered as a systemic disease, rather than solely a liver disease. The extrahepatic manifestations induced by different genotypes of HEV vary. The severity of these diseases does not necessarily correlate with the severity of HEV infection, and even asymptomatic HEV infection may trigger and cause systemic diseases. Patients with systemic manifestations of HEV infection should have priority for antiviral therapy, which could alleviate or improve the extrahepatic manifestations related to HEV infection. However, the extrahepatic manifestations caused by different genotypes of HEV and their corresponding mechanisms have not been clearly identified. This review discusses the extrahepatic manifestations related to HEV infection and their triggering mechanisms.

To meet demand for lightweight and high-strength ceramic foams, in-situ self-reinforced Si3N4 ceramic foams, with compressive strength of 13.2–45.9 MPa, were fabricated by protein foaming method combined with sintered reaction-bonded method. For comparison, ordinary protein foamed ceramics with irregular block microstructure were fabricated via reaction-bonded method, which had compressive strength of 3.6–20.5 MPa. Physical properties of these two types of samples were systematically compared. When open porosity was about 80%, both types of Si3N4 ceramic foams had excellent thermal insulation properties (

Previously, we generated a novel bispecific antibody (BsAb) targeting c-MET and PD-1 (PDCD1), which can bridge T cells and c-MET positive tumor cells. This study investigated functional mechanisms and antitumor activities of our BsAb against c-MET/PD-L1 (CD274) positive colorectal cancer (CRC). The Cancer Genome Atlas database was used to evaluate c-MET expression in tumor tissues. The expression of plasma exosomal c-MET/PD-L1 in CRC patients was measured by enzyme-linked immunosorbent assays. Western blotting assay was performed to determine expression levels of c-MET and molecular mechanism. The scratch wound healing migration assay and transwell chamber invasion assay were conducted to determine cell migratory and invasive abilities, respectively. A humanized mouse model was used to evaluate antitumor activity of the BsAb in vivo. The BsAb inhibited c-MET/PD-L1+ CRC cell migration and invasion and mediated strong antitumor activity against HCT116 tumors in mice. The BsAb induced the degradation of c-MET protein in a dose and time-dependent manner. The BsAb suppressed the phosphorylation of c-MET downstream proteins GRB2-associated-binding protein 1 (Gab1) and focal adhesion kinase (FAK). The BsAb promoted macrophage phagocytosis. Furthermore, the level of plasma exosomal-c-MET/PD-L1 distinguished CRC patients from healthy controls. The BsAb exhibited potent anti-tumor activities by two distinguished mechanisms: inhibition of c-MET signal transduction and promotion of macrophage-mediated phagocytosis. Our BsAb may provide a novel therapeutic tool for patients with c-MET/PD-L1+ CRC, and the status of exosomal-c-MET/PD-L1 may be predictive responsiveness to treatment with our BsAb.

Adenoma detection rate (ADR) is the colonoscopy quality metric with the strongest association to interval or “missed” cancer. Accurate measurement of ADR can be laborious and costly. Our aim was to determine if administrative procedure codes for colonoscopy and text searches of pathology results for adenoma mentions could estimate ADR. We identified US Veterans with a colonoscopy using Current Procedure Terminology (CPT) codes between January 2013 and December 2016 at ten Veterans Affairs sites. We applied simple text searches using Microsoft SQL Server full-text searches to query all pathology notes for “adenoma(s)” or “adenomatous” text mentions to calculate ADRs. To validate our identification of colonoscopy procedures, endoscopists of record, and adenoma detection from the electronic health record, we manually reviewed a random sample of 2000 procedure and pathology notes from the 10 sites. Structured data fields were accurate in identification of colonoscopies being performed (PPV = 0.99; 95% CI 0.99–1.00) and identifying the endoscopist of record (PPV of 0.95; 95% CI 0.94–0.96) for ADR measurement. Simple text searches of pathology notes for adenoma mentions had excellent performance statistics as follows: sensitivity 0.99 (95% CI 0.98–1.00), specificity 0.93 (95% CI 0.92–0.95), NPV 0.99 (95% CI 0.98–1.00), and PPV 0.93 (0.91–0.94) for measurement of ADR. There was no clinically significant difference in the estimates of overall ADR vs. screening ADR (p > 0.05). Measuring ADR using administrative codes and text searches from pathology results is an efficient method to broadly survey colonoscopy quality.

Background: Lung cancer is one of most common malignancies worldwide. We have previously identified retinoic acid-induced gene G (Rig-G) as a tumor suppressor in not only acute promyelocytic leukemia, but as well in other solid tumors. However, the clinical significance of Rig-G and the underlying mechanism(s) for its biological function in lung cancer remain largely unexplored. Methods: We first compared the expression of Rig-G between lung cancer (n=138) and normal tissues (n=23), from public-available datasets and our patient cohort. We further analyzed the correlation of Rig-G expression with key clinico-pathological features and survival outcomes in a multi-site clinical cohort of 300 lung cancer patients. Functional studies for Rig-G were performed in cell lines, and an animal model to support clinical findings. Findings: We found that Rig-G was frequently downregulated in lung cancer tissues and celllines, and correlated with poor prognosis in lung cancer patients. Overexpression of Rig-G led to significantly reduced cell growth and suppressed migrationin A549 and NCI-H1944 cells, accompanied by reduced epithelial-mesenchymal transition. Likewise, restoration of Rig-G in Lewis lung carcinoma cells permitted development of fewer cancer metastases vs. controls in an animal model. Gene expression profiling results identified p53 pathway as a key downstream target of Rig-G, and p53 inhibition by pifithrin-α caused abrogation of tumor-suppressive effects of Rig-G in lung cancer. Interpretation: We for the first time have identified Rig-G as a novel and important tumor suppressor, which may serve as a potential therapeutic target for restoring p53 expression in lung cancer patients. Funding Statement: This work was supported by the National Cancer Institute, NIH (CA72851, CA184792, CA202797 and CA187956), the National Natural Science Foundation of China (81272603, 81472179 and 81873975), the Excellent Academic Leader Training Program of Shanghai Health System (2018BR31), the Clinical Research and Cultivation Project of Shanghai Tongji Hospital grant [ITJ(ZD)1803]. This work was also supported by the National Natural Science Foundation of China (81672826 and 81874179), the Municipal Human Resources Development Program for Outstanding Young Talents in Medical and Health Sciences in Shanghai (2017YQ044), the Shanghai Pujiang Talent Plan (18PJD047), the Natural Science Foundation of Shanghai (19ZR1448800), the Medical Guidance Science and Technology Support Project of Shanghai (9411964800) and the National Natural Science Foundation Training Program of Tongji Hospital (GJPY1804). Declaration of Interests: None of the authors have any potential conflicts to disclose. Ethics Approval Statement: A written informed consent was obtained from all patients and the study was approved by ethics committee of Shanghai Tongji Hospital. All animal experiments were approved by the Tongji Hospital of Tongji University Ethics Committee on the Use and Care of Animals.

Background Colorectal cancer (CRC) is the leading cause of cancer-related death worldwide. Exosome shave emerged as crucial regulators of intercellular communication and that abundant Circular RNAs (circRNAs) are enriched within exosomes. CircRNAs are novel members of noncoding RNAs regulating cancer proliferation and progression. However, the function and regulatory mechanism of cancer-derived exosomal circRNAs in CRC remains unclear. Methods CRC cells-derived exosomes were characterized using transmission electron microscopy, nanoparticle tracking analysis (NTA) and western blot. CCK-8, wound healing and transwell assays, and flow cytometry assays were conducted to assess whether exosomes would affect the proliferation, metastasis, and apoptosis of CRC cells, respectively. Moreover, we performed the RNA sequencing and RT-qPCR to identify circRNAs in exosome-stimulated CRC cells. Fluorescence in situ hybridization (FISH) assay was used to detect the cellular distribution of circPACRGL. Bioinformatic analyses (StarBase 2.0) were used to pool the miRNA targets of circPACRGL. Luciferase assays were performed to verify the direct interaction. Finally, flow cytometry was used to detect the differentiation of N1-N2 neutrophils. Results Our study identified a novel CRC-derived exosomal circRNA, circPACRGL. We found circPACRGL was significantly upregulated in CRC cells after tumor-derived exosomes addition. Moreover, circPACRGL serves as a sponge for miR-142-3p/miR-506-3p to facilitate the transforming growth factor-β1 (TGF-β1) expression. As a result, circPACRGL promoted CRC cell proliferation, migration and invasion, as well as differentiation of N1 to N2 neutrophils via miR-142-3p/miR-506-3p-TGF-β1 axis. Conclusion Our study, the first to reveal that cancer-derived exosomal circPACRGL plays an oncogenic role in CRC proliferation and metastasis, providing mechanistic insights into the roles of circRNAs in CRC progression and a valuable marker for CRC treatment.

Exosomes play important roles in proliferation and microenvironment modulation of many types of cancers, including colorectal cancer (CRC). However, the inhibitory effect of CRC cells-derived exosomes in angiogenesis has not been fully discussed. In this study, the roles of microRNA-183-5p (miR-183-5p) in abundant in exosomes secreted from the CRC cells were investigated. Initially, microarray analysis was employed to determine the differentially expressed miRNAs. Exosomes isolated from CRC cells were co-cultured with HMEC-1 cells to explore the role of exosomes in angiogenesis. Further, the effects of CRC cell-derived exosomal miR-183-5p on proliferation, invasion and tube formation abilities of HMEC-1 cells were assessed. The preventative effect of exosomal miR-183-5p in vivo was measured in nude mice. Initially, it was found that FOXO1 was downregulated while miR-183-5p was upregulated in CRC. Additionally, the inhibition of miR-183-5p was suggested to suppress proliferation, invasion and tube formation abilities of HMEC-1 cells through upregulating FOXO1. Then, in vitro assays demonstrated that CRC cell-derived exosomes overexpressing miR-183-5p contributed to an enhanced proliferation, invasion and tube formation abilities of HMEC-1 cells. Furthermore, in vivo experiments confirmed the tumor-promotive effects of CRC cell-derived exosomal miR-183-5p. Collectively, our study demonstrates that the CRC cell-derived exosomes overexpressing miR-183-5p aggravates CRC through the regulation of FOXO1. Exosomes overexpressing miR-183-5p might be a potential treatment biomarker for CRC.

Dynamic changes in metabolites may affect liver disease progression, and provide new methods for predicting liver damage. We used ultra-performance liquid chromatography-mass spectroscopy to assess serum metabolites in healthy controls (HC), and patients with acute hepatitis E (AHE) or hepatitis E virus acute liver failure (HEV-ALF). The principal component analysis, partial least squares discriminant analysis, and discriminant analysis of orthogonal projections to latent structures models illustrated significant differences in the metabolite components between AHE patients and HCs, or between HEV-ALF and AHE patients. In pathway enrichment analysis, we further identified two altered pathways, including linoleic acid metabolism and phenylalanine, tyrosine, and tryptophan biosynthesis, when comparing AHE patients with HCs. Linoleic acid metabolism and porphyrin and chlorophyll metabolism pathways were significantly different in HEV-ALF when compared with AHE patients. The discriminative performances of differential metabolites showed that taurocholic acid, glycocholic acid, glycochenodeoxycholate-3-sulfate, and docosahexaenoic acid could be used to distinguish HEV-ALF from AHE patients. The serum levels of glycocholic acid, taurocholic acid, deoxycholic acid glycine conjugate, and docosahexaenoic acid were associated with the prognosis of HEV-ALF patients. Dynamic changes in serum metabolites were associated with AHE infection and severity. The identified metabolites can be used to diagnose and predict the prognosis of HEV-ALF.

Treatment of cancer in humans requires a thorough understanding of the multiple pathways by which it develops. Recent studies suggest that nuclear receptor coactivator 4 (NCOA4) may be a predictive biomarker for renal cancer. In the present work, TCGA, GEPIA, and several bioinformatics approaches were used to analyze the NCOA4 expression patterns, prognostic relevance, and association between NCOA4 and clinicopathological features and immune cell infiltration. We investigated NCOA4 expression in malignancies. Low NCOA4 expression was associated with poor overall survival in individuals with malignancies such as cholangiocarcinoma, colon adenocarcinoma, and clear cell renal carcinoma. We also analyzed NCOA4 DNA methylation in normal and primary tumor tissues and investigated possible functional pathways underlying NCOA4-mediated oncogenesis. In conclusion, downregulation of NCOA4 is associated with poor prognosis, and NCOA4 may be a predictive biomarker for COAD.

Few studies have focused on the effect of hepatitis E virus (HEV) infection on gut microbiota. To explore the relationship between changes in gut microbiota and inflammatory factors and viral load, we conducted a comparative study of 33 patients with acute hepatitis E (AHE) patients and 25 healthy controls (HCs) using high-throughput 16S ribosomal ribonucleic acid gene sequencing. Shannon and Simpson's indices showed no significant differences in bacterial diversity between the AHE and HCs groups. Proteobacteria, Gammaproteobacteria, and Enterobacteriaceae were most abundant in the AHE group, which contributed to the difference between the gut microbiota of the AHE and HCs groups, and the same difference between the HEV-RNA-positive and HEV-RNA-negative groups. Functional prediction analysis showed that ribosome, purine metabolism, and two-component system were the top three pathways. Compared with the AHE group with normal interferon (IFN)-γ, Proteobacteria, Gammaproteobacteria, Xanthomonadaceae, and Enterobacteriaceae were more abundant in the high-IFN-γ group. The abundance of Gammaproteobacteria was positively correlated with the level of serum alanine transaminase and total bilirubin. The abundance of Gammaproteobacteria could discriminate AHE patients from HCs, and could better predict the severity of AHE patients. We believe that our findings will contribute toward a novel treatment strategy for AHE.

Rationale: Antimicrobial peptides, such as cathelicidin LL-37/hCAP-18, are important effectors of the innate immune system with direct antibacterial activity. In addition, LL-37 is involved in the regulation of tumor cell growth. However, the molecular mechanisms underlying the functions of LL-37 in promoting lung cancer are not fully understood. Methods: The expression of LL-37 in the tissues and sera of patients with non-small cell lung cancer was determined through immunohistological, immunofluorescence analysis, and enzyme-linked immunosorbent assay. The animal model of wild-type and Cramp knockout mice was employed to evaluate the tumorigenic effect of LL-37 in non-small cell lung cancer. The mechanism of LL-37 involving in the promotion of lung tumor growth was evaluated via microarray analyses, recombinant protein treatment approaches in vitro, tumor immunohistochemical assays, and intervention studies in vivo. Results: LL-37 produced by myeloid cells was frequently upregulated in primary human lung cancer tissues. Moreover, its expression level correlated with poor clinical outcome. LL-37 activated Wnt/β-catenin signaling by inducing the phosphorylation of protein kinase B and subsequent phosphorylation of glycogen synthase kinase 3β mediated by the toll-like receptor-4 expressed in lung tumor cells. LL-37 treatment of tumor cells also decreased the levels of Axin2. In contrast, it elevated those of an RNA-binding protein (tristetraprolin), which may be involved in the mechanism through which LL-37 induces activation of Wnt/β-catenin. Conclusion: LL-37 may be a critical molecular link between tumor-supportive immune cells and tumors, facilitating the progression of lung cancer.

Null pointer exceptions are common software faults, but they are dangerous and can cause a program to crash if they occur. In addition, it is hard to find them by simply running test cases. When a null pointer exception occurs, the stack trace stored by the Java runtime environment can help us to locate the cause of the exception. In this paper, firstly, we propose an automatically null pointer exception localization approach guided by stack trace, from the null pointer dereference to the null value assignment point. Secondly, an empirical study is designed to evaluate the effectiveness of the tool on eight open source projects. The experimental results show that the tool is effective in locating the null pointer exception.

RationaleThe treatment options for COVID-19 patients are sparse and do not show sufficient efficacy. Alpha-1-antitrypsin (AAT) is a multi-functional host-defense protein with anti-proteolytic and anti-inflammatory activities.ObjectivesThe aim of the present study was to evaluate whether AAT is a suitable candidate for treatment of COVID-19.MethodsAAT and inflammatory markers were measured in the serum of COVID-19 patients. Human cell cultures were employed to determine the cell-based anti-protease activity of AAT and to test whether AAT inhibits the host cell entry of vesicular stomatitis virus (VSV) particles bearing the spike (S) protein of SARS-CoV-2 and the replication of authentic SARS-CoV-2. Inhaled and / or intravenous AAT was applied to nine patients with mild-to-moderate COVID-19.Measurements and Main ResultsThe serum AAT concentration in COVID-19 patients was increased as compared to control patients. The relative AAT concentrations were decreased in severe COVID-19 or in non-survivors in ratio to inflammatory blood biomarkers. AAT inhibited serine protease activity in human cell cultures, the uptake of VSV-S into airway cell lines and the replication of SARS-CoV-2 in human lung organoids. All patients, who received AAT, survived and showed decreasing respiratory distress, inflammatory markers, and viral load.ConclusionAAT has anti-SARS-CoV-2 activity in human cell models, is well tolerated in patients with COVID-19 and together with its anti-inflammatory properties might be a good candidate for treatment of COVID-19.FundingThis work was supported by grants of the Rolf M. Schwiete Stiftung, the Saarland University, the BMBF, the State of Lower Saxony, and The State of Saarland.Scientific Knowledge on the SubjectCOVID-19 is caused by “severe acute respiratory syndrome coronavirus 2” (SARS-CoV-2) and is a serious global health threat. Efficacious treatments are not available and there are no drugs that can prevent progression towards respiratory and extra-pulmonary organ failure. AAT has been studied in vitro and has activity against SARS-CoV-2. We searched PubMed and Google Scholar using the search terms “COVID-19”, “SARS-CoV-2”, “therapy”, and “α-1-antitrypsin” (AAT) for research published in 2020 and 2021.What This Study Adds to the FieldThis study shows the results of a translational program with a focus on the biology of AAT in COVID-19. The data show that there is a relative deficiency of AAT in relation to systemic inflammation. AAT inhibits serine protease activity in human airway cells and the replication of SARS-CoV-2 in human lung organoids. Inhaled and / or intravenous application of AAT in nine patients was associated with clinical stabilization. The findings of this exploratory study suggest that AAT has a mechanistic role in the pathophysiology of COVID-19 based on its anti-inflammatory and anti-viral activities. This offers the possibility to test and develop AAT application for treatment of different phenotypes or stages of COVID-19, including severe, inflammatory courses or early stages. Inhaled treatment could be an option to administer AAT non-invasively in early stages.

Background Ginkgolide B (GKB) is a major active component of the extracts of Ginkgo biloba leaves, and it has been used as an anti-cancer agent. However, it is unknown whether GKB has the therapeutic effects on lung cancer. Here, we studied the effects of GKB on lung cancer cells. Methods The effects of GKB on lung cancer cell proliferation and invasion were analyzed by cell counting kit (CCK-8) and cell invasion assays, respectively. Apoptosis was detected by flow cytometry. Western blot analysis was used to confirm the expression of autophagy-associated proteins in GKB-treated cells. Immunofluorescence analysis was used to analyze the level of light chain 3B (LC3B). Results Treatment with GKB time-dependently inhibited the proliferation and decreased the invasive capacity of A549 and H1975 cells. GKB induced apoptosis of these cells, but there was no significant effect on apoptosis compared to the control treatment. GKB-induced inhibition of cell proliferation and GKB-induced cell death were due to autophagy rather than apoptosis. GKB-induced autophagy of lung cancer cells was dependent on beclin-1, and autophagy-induced inhibition of the NLRP3 inflammasome contributed to the anti-tumor effect of GKB. Conclusions GKB-mediated autophagy of lung cancer cells is beclin-1-dependent and results in inhibition of the NLRP3 inflammasome. Therefore, GKB might be a potential therapeutic candidate for the treatment of lung cancer.

Cystic fibrosis (CF) and chronic obstructive pulmonary disease (COPD) are associated with acute and chronic bacterial infections of the lung. Excessive differentiation of basal cells to mucus-producing goblet cells can result in mucus hyperproduction and loss of mucociliary clearance in the airways of CF and COPD patients. Here, we aimed to investigate the effect of pathogen-associated molecular patterns (PAMPs) on the differentiation of human 3D bronchospheres. Primary human bronchial epithelial cells (HBECs) were differentiated to bronchospheres in the presence of bacterial flagellin and LPS and the synthetic Toll-like receptor (TLR) ligands Pam3CSK4 (TLR-2) and polyinosinic:polycytidylic acid (pIC, TLR-3). Electron and fluorescence microscopy showed that the differentiation of bronchospheres associated with the formation of lumina and appearance of cilia within 30 days after seeding. Incubation with flagellin resulted in a decreased formation of lumina and loss of cilia formation. Incubation with Pam3CSK, pIC, and LPS did not significantly affect formation of lumina and ciliation. Mucus production was strongly increased in response to flagellin and, to a lesser degree, in response to Pam3CSK4. Our results indicate that bacterial factors, such as flagellin, drive the differentiation of the respiratory epithelium towards mucus hyperproduction.

The [2 + 2] condensation between 1,8-dihydroxyanthraquinone diester and 1,3-diamino-2-propanol in ethanol affords pseudocalixarene macrocyclic L2 in high yield. Heating L2 to 200 °C in DMSO can generate much smaller [1 + 1] macrocycle L1. Crystal structure of L1 shows that all non-hydrogen atoms are coplanar except one carbon from 1,3-diamino-2-propanol. L1 is quite inert, but L2 can interact with alkaline, alkaline-earth, as well as Zn2+, Cu2+, Fe3+ and Al3+ cation (chloride) evidenced by their CV and 1H NMR. With a L2:M (1:2), there are discernable UV–vis spectrum/1H NMR changes in the presence Cu2+, Fe3+, Al3+. Anion plays a key role in the cation binding process as the chromogenic response to Li+, Na+, K+, Ba2+ and TBA+ hydroxide. L2 forms 1:1 complex with TBAOH and 2:3 complex with NaOH, KOH, Ba(OH)2 in DMSO. L2 reacts with LiOH in 1:4 stoichiometric and is one of the highest lithium loading ligand. The reaction is second-order with t1/2 ~ 30 min for 0.25 mM L2 in the presence of 4.0 eq. LiOH. Besides hydroxide, L2 can interact with TBAF to form 1:1 host–guest complexes evidenced by their color and NMR change. With bulky cation, TBAOH shifts Ep(II) more negative, while other cations shift Ep(II) to positive. L2 is an ion pair acceptor that binds hydroxide and fluoride salt chromogenic.

Background Globus pharyngeus is common and has a low cure rate. Its etiology is complex and reported to be associated with laryngopharyngeal reflux (LPR). However, some patients with globus do not exhibit any reflux symptoms or respond to proton pump inhibitors (PPIs) treatments. The purpose of this study was to clarify the related risk factors of these patients with a final objective of improving the curative effect. Methods Forty two patients afflicted with globus pharyngeus (G group) and 38 patients without globus pharyngeus (NG group) were included in this study. According to the laryngopharyngeal Reflux Symptom Index and the response to PPIs treatments, the patients were further divided into reflux groups (G-R, NG-R) and non-reflux groups (G-NR, NG-NR). High Resolution Manometry (HRM) was performed to assess esophageal motility. Questionnaires, including categories such as life exposure factors, were conducted. Results a) The average resting and residual pressures of the upper esophageal sphincter (UES) in the G-NR group was higher than in the NG-NR and NG-R groups (P 0.05). c) The esophageal distal contractile integral score of the G-NR group was not different from the NG-NR group (P > 0.05). d) Compared to the NG-NR group, the G-NR group showed higher incidence of stress, smoking, drinking, high salt and anxiety (P

Background: Colorectal cancer (CRC) remains one of the leading causes of cancer-related death. The current study aimed to elucidate the mechanism by which exosomes carrying KRAS mutant contribute to neutrophil recruitment as well as the formation of the neutrophil extracellular traps (NETs) in CRC. Methods: APC-WT and APC-KRASG12D mouse models were initially established. The peripheral blood, spleen, bone marrow (BM) and mesenteric lymph nodes (mLN) were isolated to detect neutrophil content. Next, exosomes isolated from APC-WT and APC-KRASG12D mice were injected into APC-WT and APC-KRASG12D mice. The neutrophil ratio, NETs formation and IL-8 protein content in colon tissue were subsequently quantified. DKs-8 (wild type) and DKO-1 (KRAS mutant) cells were employed for in vitro experimentation. Then, DKs-8 cells were cultured with exosome-treated PMA stimulated neutrophil-forming NETs culture medium, with cell viability, invasion, migration, and adhesion evaluated. Results: Compared with APC-WT mice, the number of polyps and neutrophils in the peripheral blood, spleen and mLNs increased in APC-KRASG12D mice, with increased NETs formation, IL-8 expression, and exosomes. IL-8 upregulation, neutrophil recruitment and NETs formation were observed in mice injected with exosomes derived from APC-KRASG12D. The in vitro investigation results revealed that more NETs were formed by DKO-1-Exosomes, which were inhibited by DNAse. In addition, DKs-8 and DKO-1 cells-derived exosomes can adhere to NETs under static conditions in vitro. Exosomal KRAS mutants were noted to exert a stimulatory effect on the production of IL-8 while promoting NETs formation as well as CRC promotion. Conclusion: The results obtained provide evidence suggesting that exosomes may transfer mutant KRAS to recipient cells and trigger an increase in IL-8 production, promote neutrophil recruitment and form NETs, ultimately leading to the promotion of CRC. Funding Statement: This work was supported by the National Natural Science Foundation of China (81873975, 81802084, 81974314, 81902984), the Excellent Academic Leader Training Program of Shanghai Health System (2018BR31), the Medical Guidace Science and Technology Support Project of Shanghai (19411964800), the Natural Science Foundation of Shanghai (19ZR1448800), the Clinical Research and Cultivation Project of Shanghai Tongji Hospital [ITJ(ZD)1803, ITJ(ZD)1905, ITJ(QN)1905]. Declaration of Interests: The authors declare that they have no competing interests. Ethics Approval Statement: The study protocol was approved by the Ethics Committee of Tongji Hospital of Tongji University. All subjects have written informed consent. All animal use and experiments were performed in strict accordance with the procedures approved by the National Cancer Institute Animal Care and Use Committee (ACUC).

Background and Purpose PGE2 inhibits cytokine generation from human lung macrophages. However, the EP receptor that mediates this beneficial anti-inflammatory effect of PGE2 has not been defined. The aim of this study was to identify the EP receptor by which PGE2 inhibits cytokine generation from human lung macrophages. This was determined by using recently developed EP receptor ligands. Experimental Approach The effects of PGE2 and EP-selective agonists on LPS-induced generation of TNF-α and IL-6 from macrophages were evaluated. The effects of EP2-selective (PF-04852946, PF-04418948) and EP4-selective (L-161,982, CJ-042794) receptor antagonists on PGE2 responses were studied. The expression of EP receptor subtypes by human lung macrophages was determined by RT-PCR. Key Results PGE2 inhibited LPS-induced and Streptococcus pneumoniae-induced cytokine generation from human lung macrophages. Analysis of mRNA levels indicated that macrophages expressed EP2 and EP4 receptors. L-902,688 (EP4 receptor-selective agonist) was considerably more potent than butaprost (EP2 receptor-selective agonist) as an inhibitor of TNF-α generation from macrophages. EP2 receptor-selective antagonists had marginal effects on the PGE2 inhibition of TNF-α generation, whereas EP4 receptor-selective antagonists caused rightward shifts in the PGE2 concentration–response curves. Conclusions and Implications These studies demonstrate that the EP4 receptor is the principal receptor that mediates the anti-inflammatory effects of PGE2 on human lung macrophages. This suggests that EP4 receptor agonists could be effective anti-inflammatory agents in human lung disease.

// Dong Li 1, * , Junjun Sun 1, * , Wenfang Liu 2, * , Xuan Wang 3 , Robert Bals 4 , Junlu Wu 1 , Wenqiang Quan 1 , Yiwen Yao 1 , Yu Zhang 1 , Hong Zhou 1 , Kaiyin Wu 5 1 Department of Clinical Laboratory, Shanghai Tongji Hospital, Tongji University School of Medicine, 200065 Shanghai, China 2 Department of General Surgery, Shanghai Tongji Hospital, Tongji University School of Medicine, 200065 Shanghai, China 3 Department of Pharmacy, Putuo People’s Hospital, 200060 Shanghai, China 4 Department of Internal Medicine V – Pulmonology, Allergology, Respiratory Intensive Care Medicine, Saarland University Hospital, 66424 Homburg, Germany 5 Institute of Pathology, Charite Medical University, 10117 Berlin, Germany * These authors contributed equally as first authors Correspondence to: Dong Li, email: 186ld@163.com Keywords: Rig-G, lung cancer, growth inhibition, NF-κB, STAT3 Received: February 25, 2016 Accepted: August 24, 2016 Published: September 01, 2016 ABSTRACT The expression of the retinoic acid-induced G (Rig-G) gene, an all trans retinoic acid (ATRA)-inducible gene, was observed in multiple cancer cells, including lung cancer cells. However, whether Rig-G is a tumor suppressor in lung cancer is unknown. Here, we found that ectopic expression of Rig-G can lead to a significant decrease in proliferation of lung cancer cells, resulting in an inhibition of tumor growth. Rig-G knockdown results in a modest increase in cell proliferation, as well as confers an increase in colony formation. Furthermore, transcriptome and pathway analyses of cancer cells revealed a fundamental impact of Rig-G on various growth signaling pathways, including the NF-κB pathway. Rig-G inhibits NF-κB activity by suppressing STAT3 in lung cancer cells. The downregulation of miR21 and miR181b-1 and subsequent activation of PTEN/Akt and CYLD/IκB signaling axis leading to decreased NF-κB activity required to maintain the tumor-inhibiting effect of Rig-G.. Our findings contribute to a better understanding of the antitumor effect mechanism of Rig-G, as well as offer a novel strategy for lung cancer therapy.

Placenta-specific 8 (PLAC8) is closely associated with the proliferation, apoptosis and autophagy of several tumor cells. However, the expression and function of PLAC8 in oral squamous cell carcinoma (OSCC) remain unknown. Therefore, the present study investigated the function and mechanism of PLAC8 in OSCC. Reverse transcription-quantitative PCR and western blot analyses were performed to quantify the expression of PLAC8 in OSCC cell lines. The function of PLAC8 in OSCC was investigated via transfection, the Transwell and Cell Counting Kit-8 assays, immunofluorescence staining and western blotting. The results demonstrated that PLAC8 exspression was downregulated in OSCC cell lines. PLAC8 inhibited the cell proliferation in OSCC. In addition, PLAC8 restrained invasion and epithelial-mesenchymal transition of OSCC cells. Furthermore, β-catenin helped to repress PLAC8 expression by regulating the Wnt/β-catenin and PI3K/Akt/GSK3β signaling pathways in OSCC cells. Collectively, the results of the present study suggest that PLAC8 acts as a tumor suppressor in OSCC by downregulating β-catenin.

Cathelicidin is an antimicrobial peptide that plays an essential role in cell proliferation, angiogenesis, and also has been indicated in tumor promotion. However, it is unclear how cathelicidin causes tumor growth, and the pathogenic mechanisms based on gain or loss of function have not been proposed. Here, a cathelicidin related antimicrobial peptide (CRAMP) knockout mouse was generated using an A/J background (A/J-CRAMP-/- mice), and lung carcinoma growth was induced using 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and Non-typeable Haemophilus influenzae (NTHi). Compared with A/J mice, A/J-CRAMP-/- mice were found to have a lower tumor burden and longer survival times, with a significant reduction in both PCNA and Ki-67 positive cells. However, there was no difference between the number of apoptotic lung-cancer cells between the A/J and A/J-CRAMP-/- mice. This indicated cathelicidin might be a tumor growth factor for lung cancer, which was associated for proliferation of tumor cells. In the future, this animal model will be useful to study the distinct role of cathelicidin in induced-lung cancer development.

The reflux symptom idex. (DOCX 68 kb)