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Infection and severe trauma can result in acute lung injury/acute respiratory distress syndrome (ALI/ARDS) and eventually pulmonary fibrosis. Epithelial-to-mesenchymal transition (EMT) is related to pulmonary fibrosis. Our study found that pyocyanin (PCN) could promote apoptosis and EMT in alveolar type II epithelial A549 cells. We hypothesized that there might be a common mechanism related to both apoptosis and EMT in A549 cells. The aim of this study was to determine whether reactive oxygen species (ROS) induced by PCN is the common stimulus upstream of apoptosis and EMT as well as the relevant signalling pathways. A549 cells were challenged with PCN; ROS was then detected by immunofluorescence, and apoptosis was measured by flow cytometry. Caspases, EMT markers and the TGF-β/Smad pathway were assessed by Western blot, qPCR or ELISA. The results showed that PCN promoted ROS production, and the apoptosis rate was clearly increased. E-cadherin downregulation, vimentin and α-SMA upregulation in A549 cells, cleaved caspase-9 and caspase-3, TGF-β1 and activated Smad2/3 were also detected. Interestingly, the protein expression of cleaved caspase-3 and vimentin was highly positively correlated. Inhibition of ROS could partially reverse PCN-induced EMT and apoptosis in A549 cells, and EMT could also be reversed by TGF-β1 inhibitors. In conclusion, ROS may be a common activating mechanism of apoptosis and EMT in alveolar epithelial cells, during which the degree of apoptosis is positively related to EMT. ROS may induce alveolar epithelial cell apoptosis through the mitochondrial pathway or endoplasmic reticulum pathway. ROS activates TGF-β1, followed by SMADs, eventually inducing EMT., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Purpose: More recently, literature has emerged providing findings about the novelty of microRNAs (miR)-targeted therapeutics in the treatment of retinoblastoma (RB). The prime objective of this study was to identify the potential role of miR-378a-3p and its regulation in RB cells via forkhead box G1 (FOXG1)., Methods: The expression of miR-378a-3p and FOXG1 in the clinical RB tissues was determined using RNA quantitation and Western blot assays. The interaction between miR-378a-3p and FOXG1 was identified using dual luciferase reporter gene assay. The potential effects of miR-378a-3p on the RB cell biological processes were evaluated by conducting gain- and loss-of-function studies of miR-378a-3p and FOXG1, followed by cell viability, cell cycle progression, and apoptosis measurements. Furthermore, experiments were performed in nude mice to assess its effects on tumor formation., Results: miR-378a-3p was poorly expressed, whereas FOXG1 was highly expressed in RB tissues and cells. miR-378a-3p bound to the FOXG1 3' untranslated region and negatively modulated its expression. The overexpression of miR-378a-3p was found to decrease RB cell viability and to promote cell apoptosis in vitro, whereas overexpressed FOXG1 reversed the regulatory effects of miR-378a-3p on RB cellular behaviors. In nude mice, the restoration of miR-378a-3p by miR-378a-3p agomir was shown to play a role in the reduction of tumor volume and size relative to nude mice injected with negative control-agomir., Conclusions: Our findings identified that increased miR-378a-3p exerted an inhibitory effect on RB cell proliferation by targeting FOXG1, suggesting the role of miR-378a-3p as a novel therapeutic target for RB.

Receptor-interacting protein kinase 3 (RIP3) is a critical initiator in mediating necroptosis induced by tumor necrosis factor alpha (TNFα) in L929 cells, so knockdown of RIP3 inhibits TNFα-induced L929 cell necroptosis. However, RIP3 knockdown was shown to switch TNFα-induced necroptosis to apoptosis in L929 cells in other studies. Therefore, whether RIP3 knockdown blocks the TNFα-induced death of L929 cells is controversial. In this study, TNFα activated caspase pathway and induced cell death in RIP3 knockdown L929 cells, and the RIP3-independent cell death had been blocked by Z-VAD-FMK (pan-caspase inhibitor) or caspase 8 knockdown, demonstrating that RIP3 knockdown switched TNFα-induced necroptosis to caspase-dependent apoptosis. Although both TNF receptor type 1-associated death domain protein (TRADD) and RIP1 have been reported to mediate TNFα-induced apoptosis, the knockdown of TRADD, but not RIP1, suppressed TNFα-induced activation of the caspase pathway and subsequent apoptosis in RIP3 knockdown L929 cells. In addition, TRADD bound and activated caspase 8 during the RIP3-independent apoptosis process, indicating that TRADD initiates RIP3-independent apoptosis by activating the caspase pathway. Collectively, we identified the target and mechanism underlying RIP3-independent apoptosis and elucidated the coordinated roles of RIP3 and TRADD in mediating the programmed cell death of L929 cells following TNFα stimulation.

As an important pathway maintaining the balance of intracellular calcium (Ca(2+)), store-operated Ca(2+) entry (SOCE) is critical for cellular functions. Stromal interaction molecule 1 (STIM1), a key component of SOCE, plays a dual role as an endoplasmic reticulum Ca(2+) receptor and an SOCE exciter. Aberrant expression of STIM1 could be discovered in several human cancer cells. However, the role of STIM1 in regulating human hypopharyngeal carcinoma still remains unclear. Real-time polymerase chain reaction (PCR) was used to detect expression of STIM1 in human hypopharyngeal carcinoma cell line FaDu. STIM1 on FaDu cells was knocked down by lentiviral transduction method. The biological impacts after knocking down of STIM1 on FaDu cells were investigated in vitro and in vivo. The result of real-time PCR showed that STIM1 was expressed in FaDu cells. Lentiviral transduction efficiently downregulated the expression of STIM1 in FaDu cells at both mRNA and protein levels. Significant downregulation of STIM1 on FaDu cells inhibited cell proliferation, induced cell cycle arrest in G0/G1 phase, promoted cell apoptosis, and restrained cell growth rate. The antigrowth effect of STIM1 silencing was also discovered in FaDu hypopharyngeal tumor model. Our findings indicate that STIM1 is likely to become a new therapeutic target for hypopharyngeal carcinoma treatment.

Background: GBM astrocytes may adopt fetal astrocyte transcriptomic signatures involved in brain development and migration programs to facilitate diffuse tumor infiltration. Our previous data show that ETS variant 6 (ETV6) is highly expressed in human GBM and fetal astrocytes compared to normal mature astrocytes. We hypothesized that ETV6 played a role in GBM tumor progression. Methods: Expression of ETV6 was first examined in two American and three Chinese tissue microarrays. The correlation between ETV6 staining intensity and patient survival was calculated, followed by validation using public databases—TCGA and REMBRANDT. The effect of ETV6 knockdown on glioma cell proliferation (EdU), viability (AnnexinV labeling), clonogenic growth (colony formation), and migration/invasion (transwell assays) in GBM cells was tested. RNA sequencing and Western blot were performed to elucidate the underlying molecular mechanisms. Results: ETV6 was highly expressed in GBM and associated with an unfavorable prognosis. ETV6 silencing in glioma cells led to increased apoptosis or decreased proliferation, clonogenicity, migration, and invasion. RNA-Seq-based gene expression and pathway analyses revealed that ETV6 knockdown in U251 cells led to the upregulation of genes involved in extracellular matrix organization, NF-κB signaling, TNF-mediated signaling, and the downregulation of genes in the regulation of cell motility, cell proliferation, PI3K-AKT signaling, and the Ras pathway. The downregulation of the PI3K-AKT and Ras-MAPK pathways were further validated by immunoblotting. Conclusion: Our findings suggested that ETV6 was highly expressed in GBM and its high expression correlated with poor survival. ETV6 silencing decreased an aggressive in vitro phenotype probably via the PI3K-AKT and Ras-MAPK pathways. The study encourages further investigation of ETV6 as a potential therapeutic target of GBM. [ABSTRACT FROM AUTHOR]

Retinoblastoma (RB) represents an aggressive malignancy in the eye during the period of infancy and childhood. We delineated the ability of microRNA‐186 (miR‐186) to influence viability, invasion, migration, angiogenesis, and apoptosis of RB via the Hedgehog signaling pathway by targeting AAA domain‐containing protein 2 (ATAD2). The microarray‐based analysis was adopted to identify differentially expressed genes (DEGs) related to RB. Subsequently, RB cells were treated with miR‐186 mimic, miR‐186 inhibitor, or si‐ATAD2. The expression of miR‐186, ATAD2, Hedgehog signaling pathway‐related genes were evaluated, and the target relationship between miR‐186 and ATAD2 was verified. Finally, cell proliferation, invasion, migration, apoptosis, and angiogenesis were assessed. ATAD2 was identified as a DEG and modulated by miR‐186. Moreover, we revealed that ATAD2 was highly expressed, whereas miR‐186 was lowly expressed, and the Hedgehog signaling pathway was activated in RB. Then, ATAD2 as a putative target of miR‐186 was validated using a luciferase assay. miR‐186 mimic or siRNA‐ATAD2 in RB cells reduced cell viability, invasion, and migration coordinating with elevated apoptosis via impairing the Hedgehog signaling pathway, where repressed angiogenesis was observed. Overexpression of miR‐186 attenuates RB via the inactivation of the Hedgehog signaling pathway by downregulating ATAD2. [ABSTRACT FROM AUTHOR]

Highlights: [•] ZFX expression was significantly upregulated in gastric cancer (GC) cell lines and tissues. [•] Knockdown of ZFX induced significant apoptosis and cell cycle arrest in SGC7901 and MGC803 cells. [•] Knockdown of ZFX inhibited gastric cancer cell growth in vitro and in vivo. [Copyright &y& Elsevier]

Cancer Targeting Gene-Viro-Therapy (CTGVT) is a promising cancer therapeutical strategy that strengthens the anti-tumour effect of oncolytic virus by expressing inserted foreign anti-tumour genes. In this work, we constructed a novel adenoviral vector controlled by the tumour-specific survivin promoter on the basis of the ZD55 vector, which is an E1B55KD gene deleted vector we previously constructed. Compared with the original ZD55 vector, this new adenoviral vector (ZD55SP/E1A) showed much better ability of replication and reporter gene expression. We then combined anti-tumour gene interleukine-24 (IL-24) with an RNA polymerase III-dependent U6 promoter driving short hairpin RNA (shRNA) that targets M-phase phosphoprotein 1 (MPHOSPH1, a newly identified oncogene) by inserting the IL-24 and the shRNA of MPHOSPH1 (shMPP1) expression cassettes into the new ZD55SP/E1A vector. Our results demonstrated excellent anti-tumour effect of ZD55SP/E1A-IL-24-shMPP1 in vitro on multiple cancer cell lines such as lung cancer, liver cancer and ovarian caner. At high multiplicity-of-infection (MOI), ZD55SP/E1A-IL-24-shMPP1 triggered post-mitotic apoptosis in cancer cells by inducing prolonged mitotic arrest; while at low MOI, senescence was induced. More importantly, ZD55SP/E1A-IL-24-shMPP1 also showed excellent anti-tumour effects in vivo on SW620 xenograft nude mice. In conclusion, our strategy of constructing an IL-24 and shMPP1 dual gene expressing oncolytic adenoviral vector, which is regulated by the survivin promoter and E1B55KD deletion, could be a promising method of cancer gene therapy. [ABSTRACT FROM AUTHOR]

Uncoupling protein-2 (UCP2) is a mitochondrial inner membrane anion carrier and is emerging as a negative regulator of ROS production. Overexpression of UCP2 has been detected in various tumors, but its role in glioblastoma remains unclear. Using tissue microarrays and interrogations of public databases, we explored that the expression of UCP2 is upregulated in glioma, especially in GBM, and overexpression of UCP2 correlates with poor prognosis in glioma patients. To further reveal the role of UCP2 in glioma, UCP2-slienced cell lines (U251, U87MG and A172) by lentivirus were constructed to study how silenced UCP2 expression affects cellular functions in vitro, and tumorigenicity in vivo. RNA-Seq based genome and pathway analysis were performed to elucidate the underlying mechanisms of action of UCP2. Our results revealed that UCP2 silenced glioma cells show inhibited migration, invasiveness, clonogenicity, proliferation, promoted cell apoptosis in vitro, and weaker tumorigenicity in nude mice. Transcriptome analysis suggested a UCP2-dependent regulation of p38 MAPK (Mitogen-activated protein kinase) signaling networks, which was further validated by qRT-PCR and Western blot. Our research provides a new insight into the biological significance of UCP2 in glioma and its potential application in treatment and diagnosis. • UCP2 is overexpressed in GBM and associated with poorer prognosis. • UCP2 gene silencing inhibits migration and invasion and enhances apoptosis of glioma cells, while attenuates growth of glioblastomas in vivo. • There is an alteration of p38 MAPK cascade to a inhibited pattern in UCP2 shRNA transfected cells. [ABSTRACT FROM AUTHOR]

Avian pathogenic Escherichia coli (APEC) induce septicemia in chickens by invading type II pneumocytes to breach the blood–air barrier. The virulence of APEC can be regulated by quorum sensing (QS). Andrographolide is a QS inhibitor of Pseudomonas aeruginosa ( P. aeruginosa ). Therefore, we investigate whether andrographolide inhibits the injury of chicken type II pneumocytes by avian pathogenic E. coli O78 (APEC-O78) by disrupting the bacterial QS system. The results showed that sub-MIC of andrographolide significantly reduced the release of lactate dehydrogenase (LDH), F-actin cytoskeleton polymerization, and the degree of the adherence to chicken type II pneumocytes induced by APEC-O78. Further, we found that andrographolide significantly decreased the autoinducer-2 (AI-2) activity and the expression of virulence factors of APEC-O78. These results suggest that andrographolide reduce the pathogenicity of APEC-O78 in chicken type II pneumocytes by interfering QS and decreasing virulence. These results provide new evidence for colibacillosis prevention methods in chickens. [ABSTRACT FROM AUTHOR]