Acute respiratory distress syndrome enhances tumor metastasis into lungs: Role of BRD4 in the tumor microenvironment.

Acute respiratory distress syndrome (ARDS) is associated with severe lung inflammation, edema, hypoxia, and high vascular permeability. The COVID-19-associated pandemic ARDS caused by SARS-CoV-2 has created dire global conditions and has been highly contagious. Chronic inflammatory disease enhances cancer cell proliferation, progression, and invasion. We investigated how acute lung inflammation activates the tumor microenvironment and enhances lung metastasis in LPS induced in vitro and in vivo models. Respiratory illness is mainly caused by cytokine storm, which further influences oxidative and nitrosative stress. The LPS-induced inflammatory cytokines made the conditions suitable for the tumor microenvironment in the lungs. In the present study, we observed that LPS induced the cytokine storm and promoted lung inflammation via BRD4, which further caused the nuclear translocation of p65 NF-κB and STAT3. The transcriptional activation additionally triggers the tumor microenvironment and lung metastasis. Thus, BRD4-regulated p65 and STAT3 transcriptional activity in ARDS enhances lung tumor metastasis. Moreover, LPS-induced ARDS might promote the tumor microenvironment and increase cancer metastasis into the lungs. Collectively, BRD4 plays a vital role in inflammation-mediated tumor metastasis and is found to be a diagnostic and molecular target in inflammation-associated cancers.

Amelioration of Endotoxin-Induced Acute Lung Injury and Alveolar Epithelial Cells Apoptosis by Simvastatin Is Associated with Up-Regulation of Survivin/NF-kB/p65 Pathway.

Disruption of the alveolar-endothelial barrier caused by inflammation leads to the progression of septic acute lung injury (ALI). In the present study, we investigated the beneficial effects of simvastatin on the endotoxin lipopolysaccharide (LPS)-induced ALI and its related mechanisms. A model of ALI was induced within experimental sepsis developed by intraperitoneal injection of a single non-lethal LPS dose after short-term simvastatin pretreatment (10-40 mg/kg orally). The severity of the lung tissue inflammatory injury was expressed as pulmonary damage scores (PDS). Alveolar epithelial cell apoptosis was confirmed by TUNEL assay (DNA fragmentation) and expressed as an apoptotic index (AI), and immunohistochemically for cleaved caspase-3, cytochrome C, and anti-apoptotic Bcl-xL, an inhibitor of apoptosis, survivin, and transcriptional factor, NF-kB/p65. Severe inflammatory injury of pulmonary parenchyma (PDS 3.33 ± 0.48) was developed after the LPS challenge, whereas simvastatin significantly and dose-dependently protected lung histology after LPS (p < 0.01). Simvastatin in a dose of 40 mg/kg showed the most significant effects in amelioration alveolar epithelial cells apoptosis, demonstrating this as a marked decrease of AI (p < 0.01 vs. LPS), cytochrome C, and cleaved caspase-3 expression. Furthermore, simvastatin significantly enhanced the expression of Bcl-xL and survivin. Finally, the expression of survivin and its regulator NF-kB/p65 in the alveolar epithelium was in strong positive correlation across the groups. Simvastatin could play a protective role against LPS-induced ALI and apoptosis of the alveolar-endothelial barrier. Taken together, these effects were seemingly mediated by inhibition of caspase 3 and cytochrome C, a finding that might be associated with the up-regulation of cell-survival survivin/NF-kB/p65 pathway and Bcl-xL.

NSP9 of SARS-CoV-2 attenuates nuclear transport by hampering nucleoporin 62 dynamics and functions in host cells.

Nuclear pore complexes (NPC) regulate molecular traffics on nuclear envelope, which plays crucial roles during cell fate specification and diseases. The viral accessory protein NSP9 of SARS-CoV-2 is reported to interact with nucleoporin 62 (NUP62), a structural component of the NPC, but its biological impact on the host cell remain obscure. Here, we established new cell line models with ectopic NSP9 expression and determined the subcellular destination and biological functions of NSP9. Confocal imaging identified NSP9 to be largely localized in close proximity to the endoplasmic reticulum. In agreement with the subcellular distribution of NSP9, association of NSP9 with NUP62 was observed in cytoplasm. Furthermore, the overexpression of NSP9 correlated with a reduction of NUP62 expression on the nuclear envelope, suggesting that attenuating NUP62 expression might have contributed to defective NPC formation. Importantly, the loss of NUP62 impaired translocation of p65, a subunit of NF-κB, upon TNF-α stimulation. Concordantly, NSP9 over-expression blocked p65 nuclear transport. Taken together, these data shed light on the molecular mechanisms underlying the modulation of host cells during SARS-CoV-2 infection.

MicroRNAs based regulation of cytokine regulating immune expressed genes and their transcription factors in COVID-19.

BACKGROUND: Coronavirus disease 2019 is characterized by the elevation of a broad spectrum of inflammatory mediators associated with poor disease outcomes. We aimed at an in-silico analysis of regulatory microRNA and their transcription factors (TF) for these inflammatory genes that may help to devise potential therapeutic strategies in the future. METHODS: The cytokine regulating immune-expressed genes (CRIEG) were sorted from literature and the GEO microarray dataset. Their co-differentially expressed miRNA and transcription factors were predicted from publicly available databases. Enrichment analysis was done through mienturnet, MiEAA, Gene Ontology, and pathways predicted by KEGG and Reactome pathways. Finally, the functional and regulatory features were analyzed and visualized through Cytoscape. RESULTS: Sixteen CRIEG were observed to have a significant protein-protein interaction network. The ontological analysis revealed significantly enriched pathways for biological processes, molecular functions, and cellular components. The search performed in the miRNA database yielded ten miRNAs that are significantly involved in regulating these genes and their transcription factors. CONCLUSION: An in-silico representation of a network involving miRNAs, CRIEGs, and TF, which take part in the inflammatory response in COVID-19, has been elucidated. Thus, these regulatory factors may have potentially critical roles in the inflammatory response in COVID-19 and may be explored further to develop targeted therapeutic strategies and mechanistic validation.

BRD4 targeting nanotherapy prevents lipopolysaccharide induced acute respiratory distress syndrome.

Acute respiratory distress syndrome (ARDS) is a life threatening respiratory disease associated with pulmonary edema, alveolar dysfunction, hypoxia, and inflammatory cell accumulation. The most contagious form of COVID-19 associated with ARDS caused by SARS-CoV-2. SARS-CoV-2 majorly produces the cytokine storm and severe lung inflammation and ultimately leads to respiratory failure. ARDS is a complex disease and there is no proper therapeutics for effective therapy. Still, there is a huge scope to identify novel targets to combat respiratory illness. In the current study, we have identified the epigenetic regulating protein BRD4 and developed siRNA based nanomedicine to treat the ARDS. The liposomes were prepared by thin-film hydration method, where BRD4 siRNA complexed with cationic lipid and exhibited 96.24 ± 18.01 nm size and stable even in the presence of RNase. BRD4 siRNA lipoplexes (BRD4-siRNA-LP) inhibited inflammatory cells in lungs and suppressed the lipopolysaccharide (LPS) induced the neutrophil infiltration and mast cell accumulation. Also, BRD4 siRNA based nanomedicine significantly reduced the LPS induced cytokine storm followed by inflammatory signaling pathways. Interestingly, BRD4-siRNA-LP suppressed the LPS-induced p65 and STAT3 nuclear translocation and ameliorated the lung inflammation. Thus, BRD4-siRNA-LP could be a plausible therapeutic option for treating ARDS and might be useful for combating the COVID-19 associated respiratory illness.

MDA5 Governs the Innate Immune Response to SARS-CoV-2 in Lung Epithelial Cells.

Recent studies have profiled the innate immune signatures in patients infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and suggest that cellular responses to viral challenge may affect disease severity. Yet the molecular events that underlie cellular recognition and response to SARS-CoV-2 infection remain to be elucidated. Here, we find that SARS-CoV-2 replication induces a delayed interferon (IFN) response in lung epithelial cells. By screening 16 putative sensors involved in sensing of RNA virus infection, we found that MDA5 and LGP2 primarily regulate IFN induction in response to SARS-CoV-2 infection. Further analyses revealed that viral intermediates specifically activate the IFN response through MDA5-mediated sensing. Additionally, we find that IRF3, IRF5, and NF-κB/p65 are the key transcription factors regulating the IFN response during SARS-CoV-2 infection. In summary, these findings provide critical insights into the molecular basis of the innate immune recognition and signaling response to SARS-CoV-2.