Lung as a target for COVID-19: Mechanistic insights and probable candidate molecules for cure.

Novel coronavirus (SARS nCoV2), belonging to the family coronaviridae, remains a dreadful pathogen affecting the respiratory tract and lungs. COVID-19 declared a global pandemic by WHO, has become a serious cause of concern for clinicians and researchers, who need to understand the significant biology and pathogenicity of this virus to design better treatment modalities. Existing antiretroviral drugs remain partially ineffective in critical subjects with associated co-morbidities. This review provides an insight into the molecular mechanisms by which SARS-CoV2 targets the lungs leading to ARDS in severe cases. This also addresses the possible drug targets and certain anti-inflammatory natural compounds that can be looked upon as promising adjuvant therapeutics for COVID-19.

Alterations in mRNA Expression Levels of Tight Junction Proteins in the Blood Cells of Smokers with or without COPD.

AIM: This study aimed to assess the role of Tight junction proteins (TJPs) and claudins in smokers with and without COPD compared to healthy individuals. BACKGROUND: Chronic obstructive pulmonary disease (COPD) is a complex chronic respiratory disease, including various inflammatory mediators. The prime etiological element in the development of COPD is cigarette smoking. The lung airway epithelium comprises beneficial immunological barriers to draw in insults, such as environmental particulates, cigarette smoke, etc. Tight junctions (TJ) connected by transmembrane proteins determine epithelial permeability. Cigarette smoke is indicated to defect TJ integrity. The possible involvement of the airway epithelium in the pathogenesis of COPD has recently become apparent; however, its detailed mechanisms remain elusive. The integrity of airway epithelium is crucial for airway homeostasis; defective airway barrier activity contributes to COPD. OBJECTIVE: In the present study, the objective was to investigate mRNA expression levels of TJP's like TJP-1, TJP-2, TJP-3, Tight junction-associated proteins-1, claudin-1, claudin-3, claudin-4, claudin-7, claudin-10, claudin-15, claudin-19, and claudin-25 from blood samples of smokers with COPD and compared them with smokers without COPD and healthy individuals. METHODS: The mRNA expressions were evaluated by the quantitative PCR method. RESULTS: The gene expressions of these TJPs were significantly down-regulated, specifically in COPD patients with a history of smoking (Smokers with COPD). Besides, FEV% was also established for these patients. Similarly, smokers with COPD showed a significant increase in the expression levels of transcription factors, like ZEB-1, ZEB-2, PDGFA, and HDGF, compared to COPD patients without a history of smoking (smokers without COPD) and the healthy subjects. CONCLUSION: In conclusion, cigarette smoke disrupts TJ of the human airway epithelium, and the transcriptional factors counteract this smoke-induced COPD. Thus, TJPs may serve as protective elements for airway epithelial homeostasis during COPD.

A comprehensive network map of IL-17A signaling pathway.

Interleukin-17A (IL-17A) is one of the member of IL-17 family consisting of other five members (IL-17B to IL-17F). The Gamma delta (γδ) T cells and T helper 17 (Th17) cells are the major producers of IL-17A. Aberrant signaling by IL-17A has been implicated in the pathogenesis of several autoimmune diseases including idiopathic pulmonary fibrosis, acute lung injury, chronic airway diseases, and cancer. Activation of the IL-17A/IL-17 receptor A (IL-17RA) system regulates phosphoinositide 3-kinase/AKT serine/threonine kinase/mammalian target of rapamycin (PI3K/AKT/mTOR), mitogen-activated protein kinases (MAPKs) and activation of nuclear factor-κB (NF-κB) mediated signaling pathways. The IL-17RA activation orchestrates multiple downstream signaling cascades resulting in the release of pro-inflammatory cytokines such as interleukins (IL)-1ß, IL-6, and IL-8, chemokines (C-X-C motif) and promotes neutrophil-mediated immune response. Considering the biomedical importance of IL-17A, we developed a pathway resource of signaling events mediated by IL-17A/IL-17RA in this study. The curation of literature data pertaining to the IL-17A system was performed manually by the NetPath criteria. Using data mined from the published literature, we describe an integrated pathway reaction map of IL-17A/IL-17RA consisting of 114 proteins and 68 reactions. That includes detailed information on IL-17A/IL-17RA mediated signaling events of 9 activation/inhibition events, 17 catalysis events, 3 molecular association events, 68 gene regulation events, 109 protein expression events, and 6 protein translocation events. The IL-17A signaling pathway map data is made freely accessible through the WikiPathways Database ( https://www.wikipathways.org/index.php/Pathway : WP5242).

The unique molecular targets associated antioxidant and antifibrotic activity of curcumin in in vitro model of acute lung injury: A proteomic approach.

Bleomycin (BLM) injury is associated with the severity of acute lung injury (ALI) leading to fibrosis, a high-morbidity, and high-mortality respiratory disease of unknown etiology. BLM-induced ALI is marked by the activation of a potent fibrogenic cytokine transcription growth factor beta-1 (TGFß-1), which is considered a critical cytokine in the progression of alveolar injury. Previously, our work demonstrated that a diet-derived compound curcumin (diferuloylmethane), represents its antioxidative and antifibrotic application in TGF-ß1-mediated BLM-induced alveolar basal epithelial cells. However, curcumin-specific protein targets, as well as its mechanism using mass spectrometry-based proteomic approach, remain elusive. To elucidate the underlying mechanism, a quantitative proteomics approach and bioinformatics analysis were employed to identify the protein targets of curcumin in BLM or TGF-ß1-treated cells. With subsequent in vitro experiments, curcumin-related pathways and cellular processes were predicted and validated. The current study discusses two separate proteomics experiments using BLM and TGF-ß1-treated cells with the proteomics approach, various unique target proteins were identified, and proteomic analysis revealed that curcumin reversed the expressions of unique proteins like DNA topoisomerase 2-alpha (TOP2A), kinesin-like protein (KIF11), centromere protein F (CENPF), and so on BLM or TGF-ß1 injury. For the first time, the current study reveals that curcumin restores TGF-ß1 induced peroxisomes like PEX-13, PEX-14, PEX-19, and ACOX1. This was verified by subsequent in vitro assays. This study generated molecular evidence to deepen our understanding of the therapeutic role of curcumin at the proteomic level and may be useful to identify molecular targets for future drug discovery.

Therapeutic properties of Punica granatum L (pomegranate) and its applications in lung-based diseases: A detailed review.

Respiratory diseases are the prime cause of death and disability worldwide. The majority of lung-based diseases are resistant to treatment. Hence, research on unique drugs/compounds with a more efficient and minimum side effect for treating lung diseases is urgent. Punica granatum L (pomegranate) fruit has been used in the prevention and treatment of various respiratory disorders in recent times. In vivo and in vitro studies have demonstrated that pomegranate fruit, as well as its juice, extract, peel powder, and oil, exert anti-proliferative, anti-oxidant, anti-microbial, anti-inflammatory, anti-cancer, and anti-tumorigenic properties by attenuating various respiratory conditions such as asthma, lung fibrosis, lung cancer, chronic obstructive pulmonary disease (COPD), and alveolar inflammation via modulating various signaling pathways. The current review summarizes the potential properties and medical benefits of pomegranate against different lung-based diseases, also highlighting its possible role in the lung fibrinolytic system. The available data suggest that pomegranate is effective in controlling the disease progressions and could be a potential therapeutic target benefiting human health status. Furthermore, this review also outlines the preclinical and clinical studies highlighting the role of pomegranate in lung diseases further evoking future studies to investigate the effect of intake of this anti-oxidant fruit in larger and well-defined human clinical trials. PRACTICAL APPLICATIONS: This review outlines the putative pharmacologic benefits of P. granatum L (pomegranate) in treating various chronic lung-based diseases such as lung cancer, COPD, ARDS, asthma, lung fibrosis, and cystic fibrosis. This review also highlights the possible inhibitory role of P. granatum L (pomegranate) in the lung fibrinolytic system triggering the fibrinolytic markers. This review summarizes the preclinical and clinical studies using in vitro, in vivo, and human models highlighting the potential role of P. granatum L (pomegranate) in lung diseases. This review evokes future research to investigate the effect of intake of pomegranate fruit in well-defined human clinical trials.

CRISPR/Cas9 Genome Editing Tool: A Promising Tool for Therapeutic Applications on Respiratory Diseases.

Respiratory diseases are one of the prime topics of concern in the current era due to improper diagnostics tools. Gene-editing therapy, like Clustered regularly interspaced palindromic repeats- associated nuclease 9 (CRISPR/Cas9), is gaining popularity in pulmonary research, opening up doors to invaluable insights on underlying mechanisms. CRISPR/Cas9 can be considered as a potential gene-editing tool with a scientific community that is helping in the advancement of knowledge in respiratory health and therapy. As an appealing therapeutic tool, we hereby explore the advanced research on the application of CRISPR/Cas9 tools in chronic respiratory diseases such as lung cancer, Acute respiratory distress syndrome (ARDS) and cystic fibrosis (CF). We also address the urgent need to establish this gene-editing tool in various other lung diseases such as asthma, Chronic obstructive pulmonary disease (COPD) and Idiopathic pulmonary fibrosis (IPF). The present review introduces CRISPR/Cas9 as a worthy application in targeting epithelial-mesenchymal transition and fibrinolytic system via editing specific genes. Thereby, based on the efficiency of CRISPR/Cas9, it can be considered as a promising therapeutic tool in respiratory health research.

Proteomics Analysis Revealed the Importance of Inflammation-Mediated Downstream Pathways and the Protective Role of Curcumin in Bleomycin-Induced Pulmonary Fibrosis in C57BL/6 Mice.

Bleomycin (BLM)-induced pulmonary fibrosis is characterized by inflammation in the alveoli, subsequent deposition of extracellular matrix (ECM) and myofibroblasts, and an impaired fibrinolytic system. Here, we describe major hematological changes, the IL-17A-mediated p53-fibrinolytic pathway, and the high throughput hits of liquid chromatography-mass spectrometry (LC-MS) analysis during the progression of pulmonary fibrosis and the therapeutic potential of curcumin against disease progression. C57BL/6 mice were exposed to BLM, followed by curcumin intervention after 24 and 48 h. Mice were sacrificed after 7 days to validate the hematological parameters, molecular pathways, and proteomics. Various techniques such as western blotting, immunofluorescence, reverse transcriptase polymerase chain reaction (RT-PCR), hematoxylin and eosin staining, Masson's trichrome staining, and immunohistochemistry were used to validate the proposed theory. LC-MS analysis was performed using a Q-Orbitrap mass spectrometer. The Schrödinger approach was used to perform the in silico molecular docking studies. BLM-exposed mice exhibited gradual weight loss and altered lung morphology; however, these were reversed by curcumin treatment. Significant changes in the hematological parameters confirmed the severity of BLM exposure in the mice, and expression of IL-17A-mediated p53-fibrinolytic system components and alveolar epithelial cell (AEC) apoptosis further confirmed the pathophysiology of pulmonary fibrosis. Differentially expressed proteins were characterized and mapped using the proteomics approach. A strong interaction of curcumin is observed with p53, uPA, and PAI-I proteins. The key role of IL-17A-mediated inflammation in the impairment of the p53-fibrinolytic system and AEC apoptosis was confirmed during BLM-induced pulmonary fibrosis. Therapeutic efficacy of curcumin exhibited a protective role against the progression of pulmonary fibrosis, which promises potent therapeutic modality to target the IL-17A-mediated p53-fibrinolytic system during pulmonary fibrosis.

Curcumin attenuates IL-17A mediated pulmonary SMAD dependent and non-dependent mechanism during acute lung injury in vivo.

Acute lung injury (ALI) is a pathologic condition responsible for incurable human chronic respiratory diseases. Recent studies have shown the involvement of the glycoprotein, IL17A secreted by IL-17 producing cells in chronic inflammation. The current investigation was carried out to study the IL-17A mediated activation of SMAD and non- SMAD signaling in alveolar epithelial cells and to assess the putative modulatory role of curcumin. C57BL/6 mice were exposed to IL-17A and curcumin was administered as an intervention to modulate the IL-17A-induced alveolar damage. Techniques like Immunofluorescence and real-time PCR were used. We found elevated expression of IL-17A and IL-17A-associated signaling pathways to be activated in mice lung tissues. Curcumin intervention in vivo promoted the resolution of IL-17A-induced ALI and attenuated pulmonary damage. Increase phosphorylation of non- SMAD proteins like P-EGFR, P-STAT-1, STAT-3, P-JAK-1/2, P-JNK, and also SMAD proteins like P- SMAD 2/3 and TGF-ß1 was encountered upon IL-17A exposure, while curcumin intervention reversed the protein expression levels. Curcumin was found to block mRNA expressions of non- SMAD genes EGFR, JNK-1, JAK1, JAK2, STAT-1, STAT-3, MAPK14, also of TGF-ß1 and SMAD genes like SMAD 2, SMAD 3. However, mRNA expressions of SMAD 6 and SMAD 7 were increased upon curcumin intervention. Our study indicates that IL-17A participates in the development of ALI in both SMAD dependent and independent manner and the IL-17A signaling components were effectively controlled by curcumin, suggesting probable anti-inflammatory use of curcumin during ALI.

Curcumin Suppresses Epithelial Growth Factor Receptor (EGFR) and Proliferative Protein (Ki 67) in Acute Lung Injury and Lung Fibrosis In vitro and In vivo.

BACKGROUND: Acute lung injury is one of the common conditions caused due to bleomycin therapy which leads to pulmonary fibrosis, which is one of the severe interstitial lung diseases most commonly affecting the elderly individuals. EGFR and Ki67 can be marked as beneficial markers for detecting pulmonary fibrosis based on which clinicians can guide the therapy. OBJECTIVE: The aim of the study was to evaluate the effect of curcumin as an intervention on two prognostic markers EGFR and Ki67 in bleomycin-induced basal alveolar epithelial cells and C57BL/6 mice. Protein expressions and pathological expressions of EGFR and Ki67 were evaluated to analyze the effect of curcumin via both in vitro and in vivo approaches. METHODS: The effect of curcumin was investigated both on cell lines (A549) and animal model (both normal and bleomycin-induced mice, n=6) via techniques like western blotting for protein expression. Techniques like immunofluorescence and immunohistochemistry were carried out and examined under confocal microscopy and phase contrast microscopy to analyze the expressions of the said biomarkers. Bleomycin was used as a causative agent to induce inflammation. RESULTS: The natural polyphenol curcumin could downregulate the expressions levels of Ki67 and EGFR both in vitro and in vivo. Immunofluorescence analysis of proliferative marker Ki67 showed a reduced expression on curcumin treatment in vitro. The pathological sections from treated lungs showed a significant decrease in EGFR and Ki67 levels when exposed to curcumin. CONCLUSION: We conclude that curcumin, a well-known natural bioactive compound holds strong antiproliferative effects on Ki67 and EGFR expressions.We observed that a clinical outcome in the diagnosis of pulmonary fibrosis remains to be unconvincing so far. Curcumin can be considered as a potential therapeutic.

Caveolin-1-derived peptide limits development of pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a fatal fibrotic lung disease with a median 5-year survival of ~20%. Current U.S. Food and Drug Administration-approved pharmacotherapies slow progression of IPF, providing hope that even more effective treatments can be developed. Alveolar epithelial progenitor type II cell (AEC) apoptosis and proliferation, and accumulation of activated myofibroblasts or fibrotic lung fibroblasts (fLfs) contribute to the progression of IPF. Full-length caveolin-1 scaffolding domain peptide (CSP; amino acids 82 to 101 of Cav1: DGIWKASFTTFTVTKYWFYR) inhibits AEC apoptosis and fLf activation and expansion and attenuates PF in bleomycin (BLM)-induced lung injury in mice. Like full-length CSP, a seven-amino acid deletion fragment of CSP, CSP7 (FTTFTVT), demonstrated antifibrotic effects in murine models of lung fibrosis. When CSP7 was administered during the fibrotic phase in three preclinical models [single-dose BLM, repeated-dose BLM, and adenovirus expressing constitutively active transforming growth factor-ß1 (Ad-TGF-ß1)-induced established PF], CSP7 reduced extracellular matrix (ECM) markers characteristic of PF, increased AEC survival, and improved lung function. CSP7 is amenable to both systemic (intraperitoneal) or direct lung delivery in a nebulized or dry powder form. Furthermore, CSP7 treatment of end-stage human IPF lung tissue explants attenuated ECM production and promoted AEC survival. Ames testing for mutagenicity and in vitro human peripheral blood lymphocyte and in vivo mouse micronucleus transformation assays indicated that CSP7 is not carcinogenic. Together, these findings support the further development of CSP7 as an antifibrotic treatment for patients with IPF or other interstitial lung diseases.

Acute Lung Injury: IL-17A-Mediated Inflammatory Pathway and Its Regulation by Curcumin.

Acute lung injury (ALI) is characterized by acute inflammation and tissue injury results in dysfunction of the alveolar epithelial membrane. If the epithelial injury is severe, a fibroproliferative phase of ALI can develop. During this phase, the activated fibroblast and myofibroblasts synthesize excessive collagenous extracellular matrix that leads to a condition called pulmonary fibrosis. Lung injury can be caused by several ways; however, the present review focus on bleomycin (BLM)-mediated changes in the pathology of lungs. BLM is a chemotherapeutic agent and has toxic effects on lungs, which leads to oxidative damage and elaboration of inflammatory cytokines. In response to the injury, the inflammatory cytokines will be activated to defend the system from injury. These cytokines along with growth factors stimulate the proliferation of myofibroblasts and secretion of pathologic extracellular matrix. During BLM injury, the pro-inflammatory cytokine such as IL-17A will be up-regulated and mediates the inflammation in the alveolar epithelial cell and also brings about recruitment of certain inflammatory cells in the alveolar surface. These cytokines probably help in up-regulating the expression of p53 and fibrinolytic system molecules during the alveolar epithelial cells apoptosis. Here, our key concern is to provide the adequate knowledge about IL-17A-mediated p53 fibrinolytic system and their pathogenic progression to pulmonary fibrosis. The present review focuses mainly on IL-17A-mediated p53-fibrinolytic aspects and how curcumin is involved in the regulation of pathogenic progression of ALI and pulmonary fibrosis.

Interleukin-17A: A Potential Therapeutic Target in Chronic Lung Diseases.

BACKGROUND: Interleukin-17A (IL-17A) is a pro-inflammatory cytokine that has gained a lot of attention because of its involvement in respiratory diseases. Interleukin-17 cytokine family includes six members, out of which, IL-17A participates towards the immune responses in allergy and inflammation. It also modulates the progression of respiratory disorders. OBJECTIVE: The present review is an insight into the involvement and contributions of the proinflammatory cytokine IL-17A in chronic respiratory diseases like Idiopathic Pulmonary Fibrosis (IPF), Chronic Obstructive Pulmonary Distress (COPD), asthma, pneumonia, obliterative bronchiolitis, lung cancer and many others. CONCLUSION: IL-17A is a major regulator of inflammatory responses. In all the mentioned diseases, IL- 17A plays a prime role in inducing the diseases, whereas the lack of this pro-inflammatory cytokine reduces the severity of respective respiratory diseases. Thereby, this review suggests IL-17A as an instrumental target in chronic respiratory diseases.

Targeting anti-aging protein sirtuin (Sirt) in the diagnosis of idiopathic pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a severe, incurable, age-associated respiratory disorder that has gained significance because of its unknown etiology and lack of therapeutic approaches. IPF causes maximum damage to the alveolar epithelial cells, thereby leading to lung remodeling and initiating epithelial to mesenchymal transition (EMT). The actual molecular mechanisms underlying IPF still remain unclear, and knowledge about these mechanisms would be helpful in its diagnosis. Sirtuins (Sirt) are class of NAD+-dependent proteins, widely known to exert positive and protective effects on age-related diseases such as diabetes, cancer, and so on, and are also involved in regulating IPF. The sirtuin family comprises of seven members (Sirt1 to Sirt7), out of which Sirt1, Sirt3, Sirt6, and Sirt7 exert positive effects on IPF. Sirt1 is associated with aging and inhibits cellular senescence and fibrosis. Sirt1 is well recognized in controlling pulmonary fibrosis and is also considered as a prime positive mediator of EMT. The expressions of Sirt3 protein tend to decline in IPF patients; hence it is known as an anti-fibrotic protein. Sirt6 indeed has been proven to reduce EMT during IPF. Decreased levels of Sirt7 during IPF regulate lung fibroblasts. Hence, active levels of Sirt1, Sirt3, Sirt6, and Sirt7 can be attractive target models to elucidate a novel potential therapeutic approach for IPF. In this prospect, we have discussed the role of Sirtuins in pulmonary fibrosis by exploring the recent research evidence that highlight the role of sirtuins and also describes their protective effects.

Inflammatory and Fibrinolytic System in Acute Respiratory Distress Syndrome.

Acute respiratory distress syndrome (ARDS) is the most advanced form of acute lung injury (ALI). This is characterized by bilateral pulmonary infiltrates and severe hypoxemia. According to Berlin definition of ARDS, this is defined based on the timings, radiographic changes, edema formation, and severity on the PaO2/FiO2 ratio. During ARDS, the loss of integrity of the epithelium causes the septic shock. The degree of epithelial injury is the major prognostic marker of ARDS. In addition to this, inflammatory cell migration, fibro-proliferation, and activation of apoptosis also play an important role in the pathophysiology of ARDS. The alveolar epithelial cell is the prime target during injury where this cell either undergo apoptosis or epithelial-mesenchymal transition (EMT). Injury to the AECs triggers the changes in the DNA fragmentation and activation of certain apoptotic markers such as caspases at the same time some cells undergo biochemical changes and loses its epithelial morphology as well epithelial biomarkers and gain mesenchymal biomarkers and morphology. In both the cases, the fibrinolytic system plays an important role in maintaining the integrity of the disease process efficiently. This review highlights the research evidence of apoptosis and EMT in lung development, injury and its prognosis in ARDS thereby to develop an effective strategy for the treatment of ARDS.

IL-17A suppresses and curcumin up-regulates Akt expression upon bleomycin exposure.

Pro-inflammatory cytokine IL-17A modulates the expression of Akt in bleomycin (BLM) administered alveolar basal epithelial cells, the mechanism behind which remains unclear. This investigation was carried out to assess IL-17A mediated down-regulation of Akt expression and the pivotal role of curcumin as a regulatory molecule. Alveolar basal epithelial cells were treated with BLM and IL-17A and curcumin was administered as an intervention to regulate the BLM-induced oxidative damage. Cell proliferation was evaluated by clonogenic assay. Akt phosphorylation and total Akt expressions were studied using western blot analysis. Cell proliferation reduced upon treatment with BLM and this phenomenon was reversed in cells upon administration with curcumin. Administrations of BLM and IL-17A to the alveolar basal epithelial cells showed significant down-regulation of Akt expression which was reversed by treatment with curcumin. BLM and IL-17A mediated inflammation was intervened effectively with curcumin. Results of this study suggest the probable use of curcumin as an anti-inflammatory therapeutic for lung injury.

Role of the urokinase-fibrinolytic system in epithelial-mesenchymal transition during lung injury.

Alveolar type II epithelial (ATII) cell injury precedes development of pulmonary fibrosis. Mice lacking urokinase-type plasminogen activator (uPA) are highly susceptible, whereas those deficient in plasminogen activator inhibitor (PAI-1) are resistant to lung injury and pulmonary fibrosis. Epithelial-mesenchymal transition (EMT) has been considered, at least in part, as a source of myofibroblast formation during fibrogenesis. However, the contribution of altered expression of major components of the uPA system on ATII cell EMT during lung injury is not well understood. To investigate whether changes in uPA and PAI-1 by ATII cells contribute to EMT, ATII cells from patients with idiopathic pulmonary fibrosis and chronic obstructive pulmonary disease, and mice with bleomycin-, transforming growth factor ß-, or passive cigarette smoke-induced lung injury were analyzed for uPA, PAI-1, and EMT markers. We found reduced expression of E-cadherin and zona occludens-1, whereas collagen-I and α-smooth muscle actin were increased in ATII cells isolated from injured lungs. These changes were associated with a parallel increase in PAI-1 and reduced uPA expression. Further, inhibition of Src kinase activity using caveolin-1 scaffolding domain peptide suppressed bleomycin-, transforming growth factor ß-, or passive cigarette smoke-induced EMT and restored uPA expression while suppressing PAI-1. These studies show that induction of PAI-1 and inhibition of uPA during fibrosing lung injury lead to EMT in ATII cells.