Damage In A549 Cells Research Articles

Forsythoside B, the active component of Frosythiae fructuse water extract, alleviates Streptococcus pneumoniae virulence by targeting pneumolysin.

To explore the therapeutic potential of Forsythoside B in treating Streptococcus pneumoniae (S. pneumoniae) infections, focusing on its ability to inhibit pneumolysin activity and protect cells from damage. Hemolysis tests were used to evaluate Forsythoside B's inhibitory effect on pneumolysin activity, while growth curve analysis assessed its impact on S. pneumoniae growth. Western blotting and oligomerization analysis were conducted to examine its influence on pneumolysin oligomerization. Cytotoxicity assays, including LDH release and live/dead cell staining, evaluated the protective effects of Forsythoside B against pneumolysin-induced damage in A549 cells. Additionally, a mouse model was employed to test the effects on survival rates, lung bacterial load, and inflammation. The results showed that Forsythoside B significantly inhibited pneumolysin activity, reduced its oligomerization, and protected A549 cells from damage without affecting bacterial growth. In the mouse model, it improved survival rates and reduced lung inflammation, indicating its potential as a therapeutic agent against S. pneumoniae infections. Forsythoside B shows potential as a therapeutic agent for treating pneumonia, particularly in infections caused by S. pneumoniae.

Aloe and coconut extracts mediated CuInS2 nanoparticles induce apoptosis in non-small lung cancer cells (A549)

The most prevalent type of cancer found in the world is lung cancer. Furthermore, chemotherapeutic intervention for lung carcinomas is frequently associated with severe off-target effects. As a result, there is a worldwide push to investigate alternative medicines with superior tolerance profiles, such as natural compounds, to substitute routinely used chemotherapeutics. The purpose of this study is to examine the anticancer potential of aloe (S1) and coconut (S2) extract-mediated CuInS2 (CIS) nanoparticles against non-small cell lung carcinoma (A549) cells. S1 and S2 both efficiently showed a dose-proportionate cytotoxic effect on A549 cells, while causing negligible toxic effects on normal healthy lung cells (WI-38), indicating their safety against healthy cells. Reduced cell viability was validated by acridine orange and ethidium bromide double staining, demonstrating that apoptosis was induced in A549 cells treated with both nanoparticles. In addition, the mitochondrial membrane potential was reduced by both nanoparticles, leading to an increase in ROS generation. Both nanoparticles increased caspase-3, caspase-9, and caspase-8 levels in A549 cells. To summarize, both nanoparticles induced apoptotic cell damage in A549 cells by targeting mitochondria, causing increased ROS production, activating the caspase cascade, and inducing apoptosis. Aloe (S1) and coconut (S2) extract-mediated CIS nanoparticles may represent viable therapeutic options for lung cancer management.

Norwogonin aids in fighting MRSA-induced pneumonia by targeting agrAC to inhibit α-hemolysin production.

The increasing prevalence of infections related to methicillin-resistant Staphylococcus aureus (MRSA) necessitates the exploration of innovative therapeutic strategies that diverge from conventional antibiotic treatments. This is imperative to effectively combat resistance and manage these infections. The adoption of antivirulence strategies has emerged as a particularly promising avenue. This approach applies a heightened selective pressure on pathogens, thereby diminishing the likelihood of bacteria evolving resistance to antibiotics. In our pursuit of novel therapeutics for treating MRSA infections, we have focused on agents that inhibit the virulence of S. aureus without impeding its growth, aiming to minimize the development of drug resistance. α-Hemolysin, a critical virulence factor encoded by the hla gene, is a cytotoxin that forms pores in host cell membranes and plays a pivotal role in the progression of disease during bacterial infections. Herein, we identified that norwogonin could effectively inhibit Hla production via targeting agrAC, a crucial protein in quorum sensing, resulting in dose-dependent inhibition of hemolytic activity without suppressing S. aureus growth. In vitro assays illustrated that norwogonin decreased the thermal stability of agrAC, providing evidence of interaction between norwogonin and agrAC. Meanwhile, norwogonin alleviated Hla-mediated A549 cell damage and reduced lactate dehydrogenase release. In vivo studies suggested that norwogonin treatment blocked the establishment of a mouse model of pneumonia caused by S. aureus USA300. Notably, norwogonin enhanced the antibacterial potency of oxacillin. In conclusion, norwogonin is a promising candidate for treating S. aureus infections, offering a novel alternative to traditional antibiotics by targeting virulence factors and enhancing the efficacy of existing treatments.

NCI 159456 PERK Inhibitor as a Targeted Therapy for Lung Cancer: An In Vitro Study.

Non-small cell lung cancer (NSCLC) represents the most common histological type of lung cancer, characterized by a five-year survival rate of 15% and poor prognosis. Accumulating evidence indicates a prominent role of endoplasmic reticulum (ER) stress and the protein kinase RNA-like ER kinase (PERK)-dependent pathway of the unfolded protein response (UPR) in the pathogenesis of NSCLC. Increased expression of downstream targets of PERK was observed in various subtypes of NSCLC, and it was associated with a more aggressive phenotype, high risk of recurrence, and poor prognosis. Therefore, the present study aimed to investigate the biological effect of the selective PERK inhibitor NCI 159456 on A549 NSCLC cells and Human Pulmonary Fibroblasts (HPF) in vitro. Treatment of both normal and ER-stressed A549 cells with NCI 159456 resulted in a significant increase in the mRNA expression level of pro-apoptotic genes like activating transcription factor 4 (ATF4), DNA damage inducible transcript 3 (DDIT3), and BCL2 Associated X, Apoptosis Regulator (BAX) as well as a decreased level of the anti-apoptotic gene B-cell lymphoma 2 (Bcl-2). Cytotoxicity and genotoxicity analyses revealed that NCI 159456 significantly decreased viability and increased DNA damage in A549 cells under normal and ER stress conditions. Caspase-3 and reactive oxygen species (ROS) detection assays demonstrated that NCI 159456 significantly induced apoptosis and increased the ROS level in normal and ER-stressed A549 cells. Importantly, treatment with the inhibitor did not affect substantially normal HPF cells at any used concentration. The results indicate that PERK inhibitors could potentially be applied as a targeted therapy for NSCLC.

FTO Deficiency Alleviates LPS-induced Acute Lung Injury by TXNIP/NLRP3-mediated Alveolar Epithelial Cell Pyroptosis.

N6-methyladenosine (m6A) plays a role in various diseases, but it has rarely been reported in acute lung injury (ALI). The FTO (fat mass and obesity-associated) protein can regulate mRNA metabolism by removing m6A residues. The aim of this study was to examine the role and mechanism of the m6A demethylase FTO in LPS-induced ALI. Lung epithelial FTO-knockout mice and FTO-knockdown/overexpression human alveolar epithelial (A549) cell lines were constructed to evaluate the effects of FTO on ALI. Bioinformatics analysis and a series of invivo and invitro assays were used to examine the mechanism of FTO regulation. Rescue assays were conducted to examine whether the impact of FTO on ALI depended on the TXNIP/NLRP3 pathway. In LPS-induced ALI, RNA m6A modification amounts were upregulated, and FTO expression was downregulated. In vivo, lung epithelial FTO knockout alleviated alveolar structure disorder, tissue edema, and pulmonary inflammation and improved the survival of ALI mice. In vitro, FTO knockdown reduced A549 cell damage and death induced by LPS, whereas FTO overexpression exacerbated cell damage and death. Mechanistically, bioinformatics analysis revealed that TXNIP was a downstream target of FTO. FTO deficiency mitigated pyroptosis in LPS-induced ALI via the TXNIP/NLRP3 pathway. Rescue assays confirmed that the impact of FTO on the TXNIP/NLRP3 pathway was significantly reversed by the TXNIP inhibitor SRI-37330. Deficiency of FTO alleviates LPS-induced ALI via TXNIP/NLRP3 pathway-mediated alveolar epithelial cell pyroptosis, which might be a novel therapeutic strategy for combating ALI.

Attenuation of PM2.5-induced alveolar epithelial cells and lung injury through regulation of mitochondrial fission and fusion

BackgroundExposure to particulate matter (PM) with an aerodynamic diameter less than 2.5 μm (PM2.5) is a risk factor for developing pulmonary diseases and the worsening of ongoing disease. Mitochondrial fission and fusion are essential processes underlying mitochondrial homeostasis in health and disease. We examined the role of mitochondrial fission and fusion in PM2.5-induced alveolar epithelial cell damage and lung injury. Key genes in these processes include dystrophin-related protein 1 (DRP1) and optic atrophy 1 (OPA1) respectively.MethodsAlveolar epithelial (A549) cells were treated with PM2.5 (32 µg/ml) in the presence and absence of Mdivi-1 (10µM, a DRP1 inhibitor) or BGP-15 (10µM, an OPA1 activator). Results were validated using DRP1-knockdown (KD) and OPA1-overexpression (OE). Mice were injected intraperitoneally with Mdivi-1 (20 mg/kg), BGP-15 (20 mg/kg) or distilled water (control) one hour before intranasal instillation of PM2.5 (7.8 mg/kg) or distilled water for two consecutive days.ResultsPM2.5 exposure of A549 cells caused oxidative stress, enhanced inflammation, necroptosis, mitophagy and mitochondrial dysfunction indicated by abnormal mitochondrial morphology, decreased mitochondrial membrane potential (ΔΨm), reduced mitochondrial respiration and disrupted mitochondrial fission and fusion. Regulating mitochondrial fission and fusion pharmacologically using Mdivi-1 and BGP-15 and genetically using DRP1-KD and OPA1-OE prevented PM2.5-induced celluar damage in A549 cells. Mdivi-1 and BGP-15 attenuated PM2.5-induced acute lung injury in mice.ConclusionIncreased mitochondrial fission and decreased mitochondrial fusion may underlie PM2.5-induced alveolar epithelial cell damage in vitro and lung injury in vivo.

Retracted: Mir-204 Regulates LPS-Induced A549 Cell Damage by Targeting FOXK2.

[This retracts the article DOI: 10.1155/2021/7404671.].

Natural flavone hispidulin protects mice from Staphylococcus aureus pneumonia by inhibition of α-hemolysin production via targeting AgrAC

The emergence of drug-resistant Staphylococcus aureus (S. aureus) has limited drug options for the clinical treatment of S. aureus infections. Considering recent reports, therapeutic strategies targeting bacterial virulence hold great promise, and alpha-hemolysin (encoded by hla), a critical virulence factor of S. aureus, plays a vital role during bacterial infection. Herein, we demonstrated that hispidulin effectively inhibited the hemolytic activity of S. aureus USA300 without suppressing bacterial growth, along with inhibiting hla transcription and expression in a dose-dependent manner. As heptamer formation is essential for hla-mediated invasion of cells, nevertheless, hispidulin did not affect the deoxycholate-induced oligomerization of hla, suggesting that hispidulin did not affect the protein activity of hla. In vitro assays illustrated that hispidulin bound to agrAC, a crucial protein in quorum sensing. Meanwhile, hispidulin alleviated A549 cell damage caused by S. aureus USA300 and reduced lactate dehydrogenase release. In vivo studies showed that hispidulin had a protective effect against pneumonia caused by S. aureus USA300 in mice. S. aureus did not develop resistance to hispidulin in the short term. Interestingly, our research indicated that hispidulin synergized with the antibacterial activity of cefoxitin. These results showed that hispidulin effectively inhibited α-hemolysin expression by inhibiting the agr quorum sensing of S. aureus. It has promise as an agent to treat S. aureus infection.

β-Catenin-Specific Inhibitor, iCRT14, Promotes BoHV-1 Infection-Induced DNA Damage in Human A549 Lung Adenocarcinoma Cells by Enhancing Viral Protein Expression.

Oncolytic bovine herpesvirus type 1 (BoHV-1) infection induces DNA damage in human lung adenocarcinoma cell line A549. However, the underlying mechanisms are not fully understood. We found that BoHV-1 infection decreased the steady-state protein levels of p53-binding protein 1 (53BP1), which plays a central role in dictating DNA damage repair and maintaining genomic stability. Furthermore, BoHV-1 impaired the formation of 53BP1 foci, suggesting that BoHV-1 inhibits 53BP1-mediated DNA damage repair. Interestingly, BoHV-1 infection redistributed intracellular β-catenin, and iCRT14 (5-[[2,5-Dimethyl-1-(3-pyridinyl)-1H-pyrrol-3-yl]methylene]-3-phenyl-2,4-thiazolidinedione), a β-catenin-specific inhibitor, enhanced certain viral protein expression, such as the envelope glycoproteins gC and gD, and enhanced virus infection-induced DNA damage. Therefore, for the first time, we provide evidence showing that BoHV-1 infection disrupts 53BP1-mediated DNA damage repair and suggest β-catenin as a potential host factor restricting both virus replication and DNA damage in A549 cells.

GSDMD-miR-223-NLRP3 axis involved in B(a)P-induced inflammatory injury of alveolar epithelial cells

Benzo(a)pyrene [B(a)P], a ubiquitous environmental pollutant, causes lung inflammatory damage. Pyroptosis，a new inflammation-dependent programmed cell death, happened when pyroptosis-related GSDMD is activated mediated by NLRP3 inflammasome. microRNA-223 (miRNA-223) is involved in inflammatory diseases by regulating NLRP3. However, whether GSDMD regulate NLRP3 inflammasome through miR-223 in B(a)P induced lung inflammatory injury remain unknown. In this study, alveolar epithelial cells (A549) were stimulated with 0, 2, 4, 8 μM B(a)P for 12 h or 24 h. The inflammatory injury and pyroptosis were determined. And the activation of NLRP3 inflammasomes and the level of miRNA-223 were detected. Then, the change of inflammatory injury and activation of NLRP3 inflammasomes in B(a)P-induced A549 cells were detected after inhibiting of GSDMD or miR-223 using siRNA-GSDMD (siGSDMD) or miR-223 inhibitor, respectively. Our results indicated that after B(a)P exposure, TNF-α and IL-6 in the supernatant were increased. Transmission electron microscope (TEM) results showed that A549 cells were obviously swollen and the cell membrane ruptured. Hoechest33342/PI staining showed that pyroptosis occurred. NLRP3, IL-1β, IL-18, GSDMD, GSDMD-N, pro caspase-1 and cleaved caspase-1 were significantly increased. Additionally, after transfecting with siGSDMD in B(a)P-induced A549 cells, the expression level of miR-223 was significantly increased. But IL-6 and TNF-α in the supernatant, the expression of NLRP3, IL-1β, IL-18 and cleaved caspase-1 protein were also decreased. And after inhibiting miR-223 in B(a)P-induced A549 cells, the expression of TNF-α and IL-6 in the supernatant, the protein expression of NLRP3, IL-1β, IL-18 and cleaved caspase-1 were increased. In conclusion, GSDMD may regulate NLRP3 inflammasome through miR-223, which is involved in B(a)P induced inflammatory damage in A549 cells.

Mir-204 Regulates LPS-Induced A549 Cell Damage by Targeting FOXK2.

Objective To assess whether miR-204 and HA affect A549 cell injury induced by lipopolysaccharide. Material and Methods. A549 cells were treated with hirsutanol A, and cell damage was induced by LPS followed by analysis of cell proliferation by CCK-8, cell apoptosis by flow cytometry, apoptosis-related protein expression by western blot, downstream target of miR-20 by dual-luciferase reporter gene, and inflammatory factors by ELISA and PCR. Results LPS can significantly inhibit the viability of A549 cells, induce cell apoptosis, and promote the release of IL-6, IL-1β, and TNF-α, while HA pretreatment can target FOXK2 by upregulating miR-204 levels, thereby alleviating apoptosis and promoting cell viability and at the same time inhibiting the release of inflammatory factors by inhibiting the activation of NF-κB. Conclusions miR-204 participates in the protection of HA acute lung injury by targeting FOXK2.

Tanshinone IIA derivatives induced S-phase arrest through stabilizing c-myc G-quadruplex DNA to regulate ROS-mediated PI3K/Akt/mTOR pathway

Herein, a derivate from tanshinone IIA, 1,6,6-trimethyl-11-phenyl-7,8,9,10-tetrahydro-6H-furo[2′,3′:1,2]phenanthro[3,4-d]imidazole (TA25), has been synthesized and investigated as potential inhibitor against the proliferation, migration and invasion of lung cancer cells. MTT assay and cell colony formation assay results showed that TA25 exhibits acceptable inhibitory effect against the proliferation of lung cancer A549 cells, and the value of IC50 was about 17.9 μM. This result was further confirmed by the inhibition of TA25 against the growth of xenograft lung cancer cells on zebrafish bearing tumor (A549 lung cancer cells). The results of wound-healing assay and FITC-gelatin invasion assay displayed that TA25 could inhibit the migration and invasion of lung cancer A549 cells. Moreover, the studies on the binding properties of TA25 interact with c-myc G-quadruplex DNA suggested that TA25 can bind in the G-quarter plane formed from G7, G11, G16 and G20 with c-myc G-quadruplex DNA through π-π stacking. Further study of the potential anti-cancer mechanism indicated that TA25 can induce S-phase arrest in lung cancer A549 cells, and this phenomenon resulted from the promotion of the production of reactive oxygen species and DNA damage in A549 cells under the action of TA25. Further research revealed that TA25 could inhibit the PI3K/Akt/mTOR signal pathway and increase the expression of p53 protein. Overall, TA25 can be developed into a promising inhibitor against the proliferation, migration and invasion of lung cancer cells and has potential clinical application in the near future.

Astragalin flavonoid inhibits proliferation in human lung carcinoma cells mediated via induction of caspase-dependent intrinsic pathway, ROS production, cell migration and invasion inhibition and targeting JAK/STAT signalling pathway.

The aim of the current study was to investigate the anti-lung cancer effects of astragalin. Studies were also undertaken to evaluate its effects on apoptosis induction, ROS production, cellular migration and invasion and JAK/STAT3 signalling pathway. MTT assay was used to evaluate cell viability in NSCLC A549 cells after exposure to astragalin molecule. Apoptosis was investigated using AO/EB staining, comet assay and western blotting assay. Fluorescence microscopy was implemented to estimate ROS production. Cell migration and invasion were measured using transwell chambers assay. Effects of astragalin on JAK/STAT pathway were investigated using western blotting assay. Results showed astragalin molecule induced inhibition of proliferation in A549 cells in a dose-dependent fashion. Further, the antiproliferative effects were found to mediate via apoptosis as suggested by AO/EB staining and western blotting assay. Astragalin modulated the expressions of caspase-3, caspase-9, Bax, Bak, Cyt-c Bcl-2, XIAP and Bcl-xL. Astragalin induced DNA damage in A549 cells which too indicated apoptotic cell death. Astragalin molecule enhanced the production of ROS by A549 cells. It inhibited both cell migration and invasion of A549 cells in a concentration-dependent manner. Finally, astragalin drug was observed with remarkable potential of targeting JAK/STAT pathway in A549 NSCLC cells. These results indicated that astragalin drug could prove helpful in lung cancer treatment and research provided more in-vivo studies are performed.

Citrus macroptera induces apoptosis via death receptor and mitochondrial mediated pathway as prooxidant in human non-small cell lung cancer cells

The role of plant-based prooxidants as the selective anticancer agents is of a great promise. Citrus macroptera commonly known as “Satkara”, is an important herbal and medicinal plant native to Southeast Asia. Its fruit is used in cooking different kinds of meat and pickle preparation for extra flavour and as folk medicine. In this study, the antiproliferative effect of ethyl acetate extract of C. macroptera peel (CPEA) was demonstrated on human non-small cell lung cancer cells through prooxidant induced apoptosis. Fatty acids, eight coumarins (three furanocoumarins) and one triterpenoid were identified from the CPEA through rigorous spectroscopic (UV, 1H and 13C NMR) and spectrometric (GC-MS, LC-ESI-HRMS) studies. Cytotoxicity, apoptotic rates, expression of apoptosis related genes, caspases activity and oxidative stress induced by reactive oxygen species (ROS) overproduction and mitochondrial membrane potential (MMP) damage in A549 cells were evaluated. MTT and flow cytometry analysis confirmed inhibition of A549 cell proliferation in a concentration-dependent manner with early apoptosis. qRT-PCR studies showed CPEA significantly upregulated p53, Bax, FasL, FADD and Bid mRNA expression levels and lowered Cdk1 and Bcl-2 compared to control. Caspase −9, −8 and −3 were also activated in cells treated by CPEA in both real-time and colorimetric/flurometric analysis. These results indicate that C. macroptera extract acts as prooxidant and induces cell death targeting both mitochondrial-and death receptor pathways of apoptosis in A549 cells.

Cytotoxicity, mitochondrial impairment, DNA damage and associated mechanisms induced by tris(1,3-dichloro-2-propyl) phosphate and tris(2-butoxyethyl) phosphate in A549 cells

Tris(1,3-dichloro-2-propyl) phosphate (TDCPP) and tris(2-butoxyethyl) phosphate (TBOEP), two of the most widely applied phosphorus-containing flame retardants (PFRs), have been shown to have adverse effects in different organisms, but their underlying toxicity mechanism remains to be elucidated. In the present study, the toxicological effects of TDCPP and TBOEP on human lung carcinoma (A549) cells and their associated mechanisms were investigated at the cellular, organelle and transcriptional levels. Cell Counting Kit-8 assay results showed that TDCPP and TBOEP reduced cell viability in a concentration-dependent manner. High-content screening assays showed that both PFRs caused mitochondrial impairment at the organelle level, decreasing the mitochondrial membrane potential and increasing the mitochondrial mass, and leading to increased cell cycle arrest-associated DNA damage that decreased the ratio of cells in S phase and increased the DNA content. In addition, oxidative stress was identified as a possible initial mechanistic process underlying PFR-induced cytotoxicity and mitochondrial impairment, as evidenced by intracellular reactive oxygen species and mitochondrial superoxide overproduction from environmentally relevant concentrations. Furthermore, p53 activation was shown to be involved in regulating DNA damage after A549 cell exposure to TDCPP and TBOEP. Quantitative PCR analyses showed that TDCPP and TBOEP triggered p53/p21-mediated cell cycle arrest pathways, indicating that these PFRs can activate p53, and promote associated functional repair. However, the p53/bax-mediated apoptosis pathway was not activated following exposure to PFRs at specific concentrations. Our results showed that TDCPP and TBOEP can cause cytotoxicity, mitochondrial impairment and cell cycle arrest-associated DNA damage in A549 cells, suggesting that oxidative stress is the initial molecular event by which these adverse outcomes are induced.

Aspirin eugenol ester ameliorates paraquat-induced oxidative damage through ROS/p38-MAPK-mediated mitochondrial apoptosis pathway

Paraquat (PQ) is an effective and commercially important herbicide that is widely used worldwide. However, PQ is highly toxic and can cause various complications and acute organ damage. Aspirin eugenol ester (AEE) is a potential new compound with anti-inflammatory and antioxidant stress pharmacological activity. The present study was to reveal the therapeutic effects and the protective effect of AEE against PQ-induced acute lung injury (ALI) with the help of PQ-induced oxidative damage in A549 cells and PQ-induced lung injury in rats. AEE might have no significant therapeutic effect on PQ-induced lung injury in rats. However, AEE had a significant protective effect on PQ-induced lung injury in rats. AEE pretreatment significantly reduced the stimulatory effect of PQ on malondialdehyde (MDA), the inhibitory effect of PQ on catalase (CAT) activity, superoxide dismutase (SOD) activity, glutathione peroxidase (GPx) activity, the ratio of GSH/GSSH, the activity of caspase-3 and the overexpression of p38 mitogen-activated protein kinase (MAPK) phosphorylation in vivo. In vitro, A549 cells were treated with 250 μM PQ for 24 h. Incubation of A549 cells with PQ led to apoptosis, and increased the level of superoxide anions, reactive oxygen species (ROS), malondialdehyde and the activity of caspase-3 and up-regulation of phosphorylated p38-MAPK, reduced mitochondrial membrane potential (ΔΨm) and the activity of SOD. However, after 24 h on AEE pretreatment of A549 cells, the above-mentioned adverse reactions caused by PQ were significantly alleviated. In addition, AEE pretreatment reduced p38-MAPK phosphorylation in PQ-treated A549 cells. SB203580, the specific p38-MAPK inhibitor, and p38-MAPK shRNA attenuated the activation of the p38-MAPK signaling pathway. N-acetylcysteine (NAC) reduced the level of phosphorylated p38-MAPK and the production of intracellular ROS and inhibited apoptosis. The results showed that AEE may inhibit PQ-induced cell damage through ROS/p38-MAPK-mediated mitochondrial apoptosis pathway.

Cyclophilin A inhibits A549 cell oxidative stress and apoptosis by modulating the PI3K/Akt/mTOR signaling pathway.

The excessive and inappropriate production of reactive oxygen species (ROS) can cause oxidative stress and is implicated in the pathogenesis of lung cancer. Cyclophilin A (CypA), a member of the immunophilin family, is secreted in response to ROS. To determine the role of CypA in oxidative stress injury, we investigated the role that CypA plays in human lung carcinoma (A549) cells. Here, we showed the protective effect of human recombinant CypA (hCypA) on hydrogen peroxide (H2O2)-induced oxidative damage in A549 cells, which play crucial roles in lung cancer. Our results demonstrated that hCypA substantially promoted cell viability, superoxide dismutase (SOD), glutathione (GSH), and GSH peroxidase (GSH-Px) activities, and attenuated ROS and malondialdehyde (MDA) production in H2O2-induced A549 cells. Compared with H2O2-induced A549 cells, Caspase-3 activity in hCypA-treated cells was significantly reduced. Using Western blotting, we showed that hCypA facilitated Bcl-2 expression and inhibited Bax, Caspase-3, Caspase-7, and PARP-1 expression. Furthermore, hCypA activates the PI3K/Akt/mTOR pathway in A549 cells in response to H2O2 stimulation. Additionally, peptidyl-prolyl isomerase activity was required for PI3K/Akt activation by CypA. The present study showed that CypA protected A549 cells from H2O2-induced oxidative injury and apoptosis by activating the PI3K/Akt/mTOR pathway. Thus, CypA might be a potential target for lung cancer therapy.

In Vitro Photodynamic Therapy of Mononuclear and Dinuclear Iridium(III) Bis(terpyridine) Complexes

Three mononuclear or dinuclear bis(terpyridine) (tpy) iridium(III) complexes bearing pyren-1-yl (pyr) group(s) were synthesized. Their photophysical properties in water and in vitro photodynamic therapy (PDT) effects toward the human lung epithelial cancer cell line A549 and the human epidermal skin cancer cell line A431 were investigated to evaluate the effects of dinuclear versus mononuclear complexes and the impact of the oligoether substituent at the ligand. All complexes possessed pyr-tpy ligand-associated charge transfer (1CT)/1π,π* absorption bands at 350-550 nm, with the dinuclear complex Ir3 showing the much enhanced absorptivity of this band. These complexes exhibited dual emission upon excitation at >430 nm in most cases, with the emitting states being ascribed to 1ILCT (intraligand charge transfer) and 3π,π*/3CT states, respectively. All complexes exhibited relatively weak to moderate cytotoxicity in the dark but high photocytotoxicity upon broadband visible light irradiation. Among them, the dinuclear complex Ir3 showed the highest intracellular reactive oxygen species (ROS) generation and PDT efficiency compared to its mononuclear counterpart Ir1. Introducing an oligoether substituent on one of the tpy ligands in Ir2 also improved its intracellular ROS generation and PDT efficacy compared to those induced by Ir1. Ir3 induced both mitochondrial dysfunction and lysosomal damage upon light activation toward both cell lines, whereas Ir1 and Ir2 caused both mitochondrial dysfunction and lysosomal damage in A431 cells but only lysosomal damage in A549 cells. The dominant cell death pathway induced by Ir1-Ir3 PDT is apoptosis.

Casticin Induces DNA Damage and Affects DNA Repair Associated Protein Expression in Human Lung Cancer A549 Cells (Running Title: Casticin Induces DNA Damage in Lung Cancer Cells).

Casticin was obtained from natural plants, and it has been shown to exert biological functions; however, no report concerns the induction of DNA damage and repair in human lung cancer cells. The objective of this study was to investigate the effects and molecular mechanism of casticin on DNA damage and repair in human lung cancer A549 cells. Cell viability was determined by flow cytometric assay. The DNA damage was evaluated by 4’,6-diamidino-2-phenylindole (DAPI) staining and electrophoresis which included comet assay and DNA gel electrophoresis. The protein levels associated with DNA damage and repair were analyzed by western blotting. The expression and translocation of p-H2A.X were observed by confocal laser microscopy. Casticin reduced total viable cell number and induced DNA condensation, fragmentation, and damage in A549 cells. Furthermore, casticin increased p-ATM at 6 h and increased p-ATR and BRCA1 at 6–24 h treatment but decreased p-ATM at 24–48 h, as well as decreased p-ATR and BRCA1 at 48 h. Furthermore, casticin decreased p-p53 at 6–24 h but increased at 48 h. Casticin increased p-H2A.X and MDC1 at 6–48 h treatment. In addition, casticin increased PARP (cleavage) at 6, 24, and 48 h treatment, DNA-PKcs and MGMT at 48 h in A549 cells. Casticin induced the expressions and nuclear translocation of p-H2AX in A549 cells by confocal laser microscopy. Casticin reduced cell number through DNA damage and condensation in human lung cancer A549 cells.

Synthesis of caffeic acid sulfonamide derivatives and their protective effect against H2O2 induced oxidative damage in A549 cells.

Exogenous antioxidants are considered as important therapeutic tools for oxidative stress associated disorders as they can regulate the redox state, which is associated with cell and organ function. Inspired by natural polyphenols, six new caffeic acid sulfonamide derivatives were synthesized by coupling sulfonamides to the backbone of caffeic acid with good yields. Their structure and lipophilicity were characterized by 1H nuclear magnetic resonance (NMR), 13C{1H} NMR, infrared spectroscopy (IR) and oil–water partition coefficient assay. Their free radical scavenging activity and antioxidant activity were assessed by DPPH assay and hydrogen peroxide (H2O2) induced oxidative stress in human lung carcinoma A549 cells. The oil–water partition coefficient results indicate that the conjugation of sulfonamides increases the lipophilicity of caffeic acid. The CASMD, CASDZ and CASN results show higher free radical scavenging effects compared with vitamin C. The derivatives do not show any inhibitory effect on the proliferation of A549 cells up to a concentration of 200 μM, except CASDZ which significantly inhibits the growth of A549 cells at a concentration of 200 μM. In addition, the obtained derivatives markedly attenuate H2O2 induced decrease of cell viability, inhibit the production of ROS and MDA, and promote the activities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px). Besides, treatment of H2O2 stimulated A549 cells with caffeic acid sulfonamide derivatives further increases mRNA expression of NF-E2-related factor 2 (Nrf2) and its target genes, including heme oxygenase-1 (HO-1), NAD(P)H quinone dehydrogenase 1 (NQO1) and thioredoxin reductase 1 (TXNRD1). These results suggest that these new caffeic acid sulfonamide derivatives have higher lipophilicity and better antioxidant activities than the parent caffeic acid, and they might be able to control the antioxidant response in cells via the Nrf2 pathway.