A549 Cell Injury Research Articles

Protective Effects of Berry Volatiles on Lipopolysaccharide-Induced Acute Lung Injury in Human A549 Cells

Abstract Objectives Berry volatiles are primary or secondary compounds produced when tissue disrupts, and responsible for the flavor and aroma of berries. Berry volatiles has been recently reported to have anti-inflammatory effect. Acute lung injury is one of the severe consequences of lung inflammation increasing respiratory stress and pulmonary edema with high mortality. Thus, the objective of this study was to investigate the anti-inflammatory effect of three berry volatiles (blackberry, black raspberry, and blueberry) on lipopolysaccharide (LPS)-induced acute lung injury in lung epithelial A549 cells and its mechanism. Methods A549 cells were pretreated with three different dilutions (2, 4, and 8-fold) of blackberry, black raspberry, and blueberry volatile extract for 1 h. Cells were then stimulated with or without LPS (10 μg/mL) for 24 h. The cell viability was measured with one-step diphenyl tetrazolium bromide (MTS) assay. The level of pro-inflammatory cytokines in the culture media was examined by using enzyme-linked immunosorbent assay (ELISA) kit. Apoptosis of A549 cells were evaluated by using death detection ELISA kit. All statistical tests were analyzed by using one-way ANOVA, followed by Tukey's multiple comparisons. Significant difference was defined at P < 0.05. Results Two-fold diluted berry volatiles (blackberry, black raspberry, and blueberry) significantly decreased A549 cell viability by 24%, 52% and 61%, respectively compared to the control (P < 0.05). Concentration of interleukin-8 (IL-8) was remarkably higher when LPS stimulated A549 cells compared to the control (P < 0.05). However, 2-fold diluted berry volatiles significantly suppressed LPS-induced IL-8 level compared to LPS treated group (P < 0.01). 2-fold diluted blackberry, black raspberry, and blueberry volatiles also showed significant apoptotic effect on A549 cells compared to the control (P < 0.05). Conclusions These results showed that blackberry, black raspberry, and blueberry volatiles suppress inflammatory responses by modulating pro-inflammatory cytokines and apoptosis. This study suggests that volatiles from berries (blackberry, black raspberry, and blueberry) may have a potential impact on lung inflammation. Funding Sources Arkansas Biosciences Institute.

Dexmedetomidine alleviates pulmonary edema through the epithelial sodium channel (ENaC) via the PI3K/Akt/Nedd4-2 pathway in LPS-induced acute lung injury

Dexmedetomidine (Dex), a highly selective α2-adrenergic receptor (α2AR) agonist, has an anti-inflammatory property and can alleviate pulmonary edema in lipopolysaccharide (LPS)-induced acute lung injury (ALI), but the mechanism is still unclear. In this study, we attempted to investigate the effect of Dex on alveolar epithelial sodium channel (ENaC) in the modulation of alveolar fluid clearance (AFC) and the underlying mechanism. Lipopolysaccharide (LPS) was used to induce acute lung injury (ALI) in rats and alveolar epithelial cell injury in A549 cells. In vivo, Dex markedly reduced pulmonary edema induced by LPS through promoting AFC, prevented LPS-induced downregulation of α-, β-, and γ-ENaC expression, attenuated inflammatory cell infiltration in lung tissue, reduced the concentrations of TNF-α, IL-1β, and IL-6, and increased concentrations of IL-10 in bronchoalveolar lavage fluid (BALF). In A549 cells stimulated with LPS, Dex attenuated LPS-mediated cell injury and the downregulation of α-, β-, and γ-ENaC expression. However, all of these effects were blocked by the PI3K inhibitor LY294002, suggesting that the protective role of Dex is PI3K-dependent. Additionally, Dex increased the expression of phosphorylated Akt and reduced the expression of Nedd4-2, while LY294002 reversed the effect of Dex in vivo and in vitro. Furthermore, insulin-like growth factor (IGF)-1, a PI3K agonists, promoted the expression of phosphorylated Akt and reduced the expression of Nedd4-2 in LPS-stimulated A549 cells, indicating that Dex worked through PI3K, and Akt and Nedd4-2 are downstream of PI3K. In conclusion, Dex alleviates pulmonary edema by suppressing inflammatory response in LPS-induced ALI, and the mechanism is partly related to the upregulation of ENaC expression via the PI3K/Akt/Nedd4-2 signaling pathway.

Long non-coding RNA MALAT1 enhances the protective effect of dexmedetomidine on acute lung injury by sponging miR-135a-5p to downregulate the ratio of X-box binding proteins XBP-1S/XBP-1U

ABSTRACT Acute lung injury (ALI) is the common and clinically severe complication. Dexmedetomidine (DEX) can protect against lipopolysaccharide (LPS)-induced ALI through anti-apoptosis, anti-inflammatory and immune regulatory actions. It is well documented that major causes of LPS-induced ALI are endoplasmic reticulum stress (ERS) and abnormally elevated CHOP. Moreover, XBP-1 can enhance CHOP expression. XBP-1S can aggravate ERS and XBP-1 U can repress ERS. By querying Starbase, miR-135a-5p interacts with XBP-1 and lncRNA MALAT1 sponges miR-135a-5p. It has been reported that MALAT1 interference markedly promoted the apoptosis of pulmonary microvascular endothelial cells in ALI rats by activating TLR4/NF-κB pathway. miR-135a-5p inhibitor remarkably alleviated LPS-induced A549 cell injury through suppressing cell apoptosis. In the present work, LPS was dripped into the nasal cavity of SD rats to establish the rat model of ALI and LPS was also applied to stimulate BEAS-2B cells to imitate ALI in vitro. Then, the pathology, lung function indexes, levels of inflammatory factors, apoptosis of lung tissues in SD rats and apoptotic level of BEAS-2B cells were measured, so as to confirm whether upregulation of lncRNA MALAT1 was able to suppress ERS, thus enhancing the protective effect of DEX against ALI. Herein, overexpression of lncRNA MALAT1 strengthened the remission effects of DEX on LPS-triggered ALI, severe pulmonary edema, inflammatory response and cell apoptosis of lung tissues in SD rats and reinforced the anti-apoptosis effect of DEX on LPS-stimulated BEAS-2B cells. Mechanically, lncRNA MALAT1 enhanced the protective effect of DEX against ALI by downregulating the ratio of XBP-1S/XBP-1U to repress ERS.

Protective effect of toll-interacting protein overexpression against paraquat-induced lung injury in mice and A549 cells through inhibiting oxidative stress, inflammation, and NF-κB signaling pathway

Toll-interacting protein (Tollip) is a pivotal negative regulator of inflammatory response. In the present study, the effects of Tollip overexpression on paraquat (PQ)-induced lung injury were explored through in vivo and in vitro investigations. Upon stimulation with PQ in mice, the expression of Tollip was down-regulated. Histopathological analysis revealed that the overexpression of Tollip significantly decreased inflammatory cell infiltration. Similarly, the levels of myeloperoxidase (MPO) and interleukin-1β (IL-1β) were lowered by Tollip overexpression in PQ-administrated mice. Besides, the overexpression of Tollip reduced reactive oxygen species (ROS) generation and malondialdehyde (MDA) level but enhanced superoxide dismutase (SOD) activity in PQ-treated A549 cells. Meanwhile, Tollip overexpression lowered the level of IL-1β and decreased the protein expressions of p-p65 in the cytoplasm and nuclear p65. Importantly, inhibition of NF-κB signaling pathway probably by decreasing NF-κB p65-DNA binding activity was induced by Tollip overexpression. Taken together, Tollip overexpression attenuated PQ-initiated lung injury possibly via reduction of oxidative stress and inflammation and suppression of NF-κB signaling pathway activation, which provided some novel ideas for the treatment of lung damage mediated by PQ.

Let7f‑5p attenuates inflammatory injury in invitro pneumonia models by targeting MAPK6.

Pneumonia accounts for ~1.3 million mortalities in children per year worldwide. MicroRNAs are implicated in several diseases, including cancer and pneumonia; however, the role of let7f-5p in pneumonia is not completely understood. In the present study, lipopolysaccharide (LPS) was used to establish an in vitro pneumonia model in A549 and WI-38 cells. The reverse transcription-quantitative PCR (RT-qPCR) and western blotting results demonstrated that let7f-5p expression levels were significantly decreased, whereas MAPK6 expression levels were significantly increased in the peripheral venous blood of patients with pneumonia and in LPS-induced A549 and WI-38 cells compared with healthy volunteers and control cells, respectively. Furthermore, the dual-luciferase reporter assay demonstrated that let7f-5p targeted the 3′-untranslated region of MAPK6. The ELISA and RT-qPCR results demonstrated that let7f-5p mimic ameliorated LPS-induced inflammatory injury in A549 and WI-38 cells, as demonstrated by decreased expression levels of proinflammatory cytokines, including TNF-α and IL-6. In addition, the Cell Counting Kit-8 assay results indicated that let7f-5p mimic ameliorated LPS-induced reductions in cell viability, and the western blotting results demonstrated that let7f-5p mimic reversed LPS-induced activation of the STAT3 signaling pathway. Notably, the aforementioned let7f-5p-mediated effects were reversed by MAPK6 overexpression. Collectively, the results of the present study suggested that let7f-5p inhibited inflammation by targeting MAPK6 in the in vitro pneumonia model, thus let7f-5p may serve as a potential novel therapeutic target for pneumonia.

MiR-30a-5p ameliorates LPS-induced inflammatory injury in human A549 cells and mice via targeting RUNX2.

In this study, we aim to investigate the role of miR-30a-5p in acute lung injury/acute respiratory distress syndrome (ALI/ARDS) using LPS-induced A549 cells and mice. We found cell viability was significantly declined accompanied by cell apoptosis and cell cycle arrest at G0/G1 phase in LPS-treated A549 cells. MiR-30a-5p was down-regulated by LPS treatment and miR-30a-5p significantly protected A549 cells from LPS-induced injury by increasing cell viability, reducing cell apoptosis, promoting cell cycle progression, and inhibiting inflammatory reactions. Dual-luciferase activity demonstrated that RUNX2 was a direct target for miR-30a-5p and its expression was negatively and directly regulated by miR-30a-5p. Over-expression of RUNX2 weakened the inhibitory effect of miR-30a-5p on inflammatory injury. In vivo, over-expression of miR-30a-5p alleviated LPS-induced inflammatory responses and lung injury in LPS-administrated mice. Besides, miR-30a-5p repressed LPS-elevated phosphorylation levels of the signal transducer and activator of transcription 3 (STAT3) and c-Jun N-terminal kinase (JNK), IκBα degradation, and NF-κB p65 phosphorylation. In conclusion, miR-30a-5p ameliorates LPS-induced inflammatory injury in A549 cells and mice via targeting RUNX2 and related signaling pathways, thereby influencing the progression of ARDS.

Mitophagy–lysosomal pathway is involved in silver nanoparticle-induced apoptosis in A549 cells

With the increasing use of silver nanoparticles (AgNPs) in biological materials, the cytotoxicity caused by these particles has attracted much attention. However, the molecular mechanism underlying AgNP cytotoxicity remains unclear. In this study, we aimed to systematically investigate the toxicity induced by AgNP exposure to the lung adenocarcinoma A549 cell line at the subcellular and signaling pathway levels and elucidate the related molecular mechanism. The survival rate of cells exposed to AgNPs at 0, 20, 40, 80, and 160 μg/mL for 24 or 48 h decreased in a dose- and time-dependent manner. AgNPs induced autophagy and mitophagy, determined by the transmission electron microscopy investigation and upregulation of LC3 II/I, p62, PINK1, and Parkin expression levels. AgNP treatment induced lysosomal injury, including the decline of lysosomal membrane integrity and increase in cathepsin B level. The decreased in mitochondrial membrane potential, along with upregulation of cytochrome c, caspases 9 and 3, and BAX/BCL2, further suggested that mitochondrial injury were involved in AgNP-induced apoptosis. In addition, mitochondrial injury may further lead to excessive production of reactive oxygen species and oxidative/ antioxidant imbalance. The results suggested that AgNPs could regulate autophagy via mitochondrial and lysosome injury in A549 cells. The information of the molecular mechanism will provide an experimental basis for the safe application of nanomaterials.

MicroRNA-4485 ameliorates severe influenza pneumonia via inhibition of the STAT3/PI3K/AKT signaling pathway.

The present study aimed to explore the potential roles and mechanism of microRNA-4485 (miR-4485) in severe influenza pneumonia. miR-4485 expression was detected in patients with severe H1N1 pneumonia using quantitative PCR. Furthermore, the effects of aberrantly expressed miR-4485 on H1N1-infected A549 cells were investigated using Cell Counting Kit-8, terminal deoxynucleotidyl transferase dUTP nick end labeling, western blotting and (ELISA) assays. Furthermore, the regulatory relationships between miR-4485 and the STAT3-mediated PI3K/AKT/mTOR signaling pathway were explored using a luciferase reporter and rescue assay. MiR-4485 expression was downregulated following H1N1 infection and in patients with H1N1 pneumonia. In addition, miR-4485 alleviated H1N1-induced A549 cell injury by promoting cell viability and the production of cytokines, as well as reducing apoptosis in A549 cells. Furthermore, STAT3 was revealed to be a target gene of miR-4485. Additionally, STAT3 silencing reversed the protective effects of miR-4485 knockdown on H1N1-induced cell injury via inhibition of the PI3K/AKT/mTOR signaling pathway. In conclusion, miR-4485 inhibited H1N1-induced severe pneumonia in A549 cells by targeting STAT3 via the PI3K/AKT/mTOR signaling pathway.

MicroRNA-92a serves as a risk factor in sepsis-induced ARDS and regulates apoptosis and cell migration in lipopolysaccharide-induced HPMEC and A549 cell injury

AimsSepsis-induced acute respiratory distress syndrome (ARDS) is a common, high mortality complication in intensive care unit (ICU) patients. MicroRNA-92a (miR-92a) plays a role in many diseases, but its association with sepsis-induced ARDS is unclear. Materials and methodsWe enrolled 53 patients, including 17 with sepsis only, and 36 with sepsis-induced ARDS. Lipopolysaccharide (LPS) was used to stimulate pulmonary microvascular endothelial cells (HPMEC) and alveolar epithelial A549 cells, which were used to investigate the miR-92a roles in ARDS. MiR-92a expression levels in patient serum and cells were quantified using quantitative reverse transcription-polymerase chain reaction (RT-PCR), and protein expression was examined using Western blotting. The effect of miR-92a on apoptosis was examined using flow cytometry. Wound healing and transwell migration assays were used to evaluate cell migration. Key findingsSerum miR-92a expression was higher in patients with sepsis-induced ARDS, when compared to patients with sepsis only. After LPS treatment in cells, miR-92a expression was higher when compared with control group, cell apoptosis and inflammatory responses were increased and cell migration was inhibited. However, cell apoptosis and inflammatory responses were decreased and cell migration was enhanced after miR-92a downregulation, when compared with inhibitor negative control (NC) group. Moreover, phosphorylated-Akt and phosphorylated-mTOR expression were increased after miR-92a inhibition. SignificanceOur study provides evidence that circulating serum miR-92a could act as a risk factor for sepsis-induced ARDS. MiR-92a inhibition attenuated the adverse effects of LPS on ARDS through the Akt/mTOR signaling pathway.

MiR-520c-3p alleviates LPS-induced A549 cell and mice lung injury via targeting NLRC5.

MicroRNAs (miRNAs) play important roles in the progression of acute lung injury (ALI). So far, little is known about the role of miR-520c-3p in ALI. In this study, the mechanism of the occurrence and development of ALI was explored. We firstly found that miR-520c-3p could inhibit the expression of NLRC5. A549 cells were treated with lipopolysaccharide (LPS) in vitro to simulate ALI cells and inducing ALI model mice. Moreover, miR-520c-3p overexpression enhanced the viability of damaged cells, inhibited LPS-induced apoptosis, and decreased LPS-induced IL-1β, IL-6 and TNF-α. In addition, the NLRC5 was a direct target of miR-520c-3p. And NLRC5 partially aggravated inflammation injury. In conclusion, miR-520c-3p alleviates LPS-induced inflammatory injury via regulating NLRC5.

MiR-342 attenuates lipopolysaccharide-induced acute lung injury via inhibiting MAPK1 expression.

Micro RNA (miRNA) and mitogen-activated protein kinase (MAPK) are reported as the crucial regulators of inflammatory responses in acute lung injury (ALI). This study will explore the role of the miR-342/MAPK1 axis in regulation of lipopolysaccharide (LPS)-induced ALI. We found that miR-342 was down-regulated in LPS-induced A549 cells compared with the control group with DMSO, accompanied by elevated inflammatory cytokines and apoptosis. Over-expression of miR-342 reduced LPS-induced inflammatory responses and apoptosis in LPS-stimulated A549 cells, and had a protective role in LPS-treated mice with ALI by decreasing levels of inflammatory cytokines, improving survival of mice with ALI, and ameliorating the lung permeability. Dual-luciferase reporter gene assay demonstrated that miR-342 regulated the expression of MAPK1 by directly targeting its 3' untranslated region (3'-UTR). Mechanistically, MAPK1 silencing abrogated LPS-induced inflammatory injury in A549 cells, and partially enhanced the protective effect of miR-342. Therefore, miR-342 attenuates LPS-induced ALI by targeting MAPK1 expression, thereby protecting against A549 cell injury induced by LPS and lung injury of mice with ALI.

Bcl-2 Proteins Regulate Mitophagy in Lipopolysaccharide-Induced Acute Lung Injury via PINK1/Parkin Signaling Pathway.

Mitophagy is involved in sepsis-induced acute lung injury (ALI). Bcl-2 family proteins play an important role in mitochondrial homeostasis. However, whether targeting Bcl-2 proteins (Bcl-2 and Bad) could influence mitophagy in ALI remains unclear. In this study, lipopolysaccharide (LPS) was used to induce injury in A549 cells and ALI in mice. LPS treatment resulted in elevated cell apoptosis, enhanced mitophagy, decreased Bcl-2 expression, increased Bad expression, and activation of PINK1/Parkin signaling in cells and lung tissues. Both Bcl-2 overexpression and Bad knockdown attenuated LPS-induced injury, inhibited cell apoptosis and mitophagy, and improved survival. Atg5 knockout (KO) inhibited LPS-induced cell apoptosis. Furthermore, Bcl-2 proteins regulated mitophagy by modulating the recruitment of Parkin from the cytoplasm to mitochondria via direct protein-protein interactions. These results were further confirmed in Park2 KO cells and Park2−/− mice. This is the first study to demonstrate that Bcl-2 proteins regulated mitophagy in LPS-induced ALI via modulating the PINK1/Parkin signaling pathway, promoting new insights into the mechanisms and investigation of therapeutic strategies for a septic patient with ALI.

MiR-297 alleviates LPS-induced A549 cell and mice lung injury via targeting cyclin dependent kinase 8

In recent decades, microRNAs (miRNAs) have been reported to play an important role in the pathogenesis of acute lung injury/acute respiratory distress syndrome (ALI/ARDS). To explore the underlying mechanisms of miR-297 in ALI/ARDS, we investigated its role in lipopolysaccharide (LPS)-induced A549 cell and mice lung injury model. We found that the expression of miR-297 decreased in LPS-induced A549 cells or serum of ALI/ARDS mice. Moreover, LPS suppressed A549 cell viability, promoted apoptosis and increased the expressions of inflammatory and autophagy-related factors. Conversely, overexpression of miR-297 increased cell proliferation and reduced cell apoptosis, and alleviated inflammatory cytokines secretion and autophagy in the presence of LPS. Importantly, miR-297 directly inhibited transcription of cyclin dependent kinase 8 (CDK8) by binding to the 3′-untranslated region (3′-UTR) of CDK8 mRNA, and the expression of CDK8 was negatively regulated by miR-297. Silencing of CDK8 promoted the anti-inflammatory and anti-apoptotic effects of miR-297 in LPS-mediated A549 cells. The expressions of phosphorylated p65 (p-p65)/p65 and phosphorylated inhibitor kappa B-α (p-IκBα)/IκBα were dramatically decreased by miR-297 mimics and silencing of CDK8. In vivo, miR-297 alleviated LPS-induced inflammatory responses and lung injury in ALI/ARDS mice. In conclusion, our findings suggested that miR-297 affects nuclear factor-kappa B (NF-κB) pathway to alleviate LPS-induced A549 cell and mice lung injury via targeting CDK8 expression.

Ability of plasma-activated acetated Ringer's solution to induce A549 cell injury is enhanced by a pre-treatment with histone deacetylase inhibitors.

Non-thermal plasma (NTP) is applicable to living cells and has emerged as a novel technology for cancer therapy. NTP affect cells not only by direct irradiation, but also by an indirect treatment with previously prepared plasma-activated liquid. Histone deacetylase (HDAC) inhibitors have the potential to enhance susceptibility to anticancer drugs and radiation because these reagents decondense the compact chromatin structure by neutralizing the positive charge of the histone tail. The aim of the present study was to demonstrate the advantage of the combined application of plasma-activated acetated Ringer’s solution (PAA) and HDAC inhibitors on A549 cancer cells. PAA maintained its ability for at least 1 week stored at any temperature tested. Cell death was enhanced more by combined regimens of PAA and HDAC inhibitors, such as trichostatin A (TSA) and valproic acid (VPA), than by a single PAA treatment and was accompanied by ROS production, DNA breaks, and mitochondria dysfunction through a caspase-independent pathway. These phenomena induced the depletion of ATP and elevations in intracellular calcium concentrations. The sensitivities of HaCaT cells as normal cells to PAA were less than that of A549 cells. These results suggest that HDAC inhibitors synergistically induce the sensitivity of cancer cells to PAA.

Hydrogen Sulfide Attenuates Hydrogen Peroxide-Induced Injury in Human Lung Epithelial A549 Cells.

Lung tissues are frequently exposed to a hyperoxia environment, which leads to oxidative stress injuries. Hydrogen sulfide (H2S) is widely implicated in physiological and pathological processes and its antioxidant effect has attracted much attention. Therefore, in this study, we used hydrogen peroxide (H2O2) as an oxidative damage model to investigate the protective mechanism of H2S in lung injury. Cell death induced by H2O2 treatment could be significantly attenuated by the pre-treatment of H2S, resulting in a decrease in the Bax/Bcl-2 ratio and the inhibition of caspase-3 activity in human lung epithelial cell line A549 cells. Additionally, the results showed that H2S decreased reactive oxygen species (ROS), as well as neutralized the damaging effects of H2O2 in mitochondria energy-producing and cell metabolism. Pre-treatment of H2S also decreased H2O2-induced suppression of endogenous H2S production enzymes, cystathionine-beta-synthase (CBS), cystathionine-gamma-lyase (CSE), and 3-mercapto-pyruvate sulfurtransferase (MPST). Furthermore, the administration of H2S attenuated [Ca2+] overload and endoplasmic reticulum (ER) stress through the mitogen-activated protein kinase (MAPK) signaling pathway. Therefore, H2S might be a potential therapeutic agent for reducing ROS and ER stress-associated apoptosis against H2O2-induced lung injury.

MiR-150 attenuates LPS-induced acute lung injury via targeting AKT3

Acute lung injury (ALI) and its severe manifestation of acute respiratory distress syndrome (ARDS) in human lung are induced by inflammatory cytokines and endogenous factors such as miRNAs. However, the role of miR-150 in lipopolysaccharide (LPS)-induced ALI is not clear. Here, we found miR-150 expression was significantly reduced in the serum of patients with ARDS, and negatively associated with the disease severity and 28-day survival of ARDS. In vivo, miR-150 decreased total cell and neutrophil counts, and production of inflammatory cytokines interleukin-1β (IL-1β), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) as well as levels of total protein, albumin and IgM in the bronchoalveolar lavage (BAL) fluid in LPS-induced ALI mice. Meanwhile, miR-150 improved the 72 h survival rate of LPS-induced ALI mice. In-vitro assays demonstrated that miR-150 alleviated LPS-induced A549 cell apoptosis, autophagy, and release of inflammatory cytokines. Further, AKT3 was a direct target of miR-150. Silencing of AKT3 partially reversed LPS-induced A549 cell injury, and enhanced the protective effects of miR-150. In addition, miR-150 or si-AKT3 effectively inhibited the phosphorylation levels of c-Jun N-terminal kinase (JNK) and nuclear factor-κB (NF-κB) (p65 and IκBα). In conclusion, miR-150 alleviated LPS-induced acute lung injury via directly targeting AKT3 expression or regulating JNK and NF-κB pathways, which may be a promising therapeutic strategy to treat ALI/ARDS.

Knockdown of the lncRNA MALAT1 alleviates lipopolysaccharide‑induced A549 cell injury by targeting the miR‑17‑5p/FOXA1 axis.

Long‑noncoding RNAs (lncRNAs) are crucial for the pathophysiology of acute lung injury (ALI). Metastasis‑associated lung adenocarcinoma transcript 1 (MALAT1) suppresses inflammatory responses via microRNA (miR)‑146a in lipopolysaccharide (LPS)‑induced ALI. However, the molecular mechanisms underlying the MALAT1‑mediated regulation of cell proliferation and apoptosis in LPS‑induced ALI remain unclear. In the present study, it was found that LPS treatment upregulated MALAT1 expression and suppressed the proliferation of A549 cells. MALAT1 knockdown significantly promoted the proliferation and G1/S phase transition and inhibited apoptosis in LPS‑treated A549 cells. In addition, miR‑17‑5p was a direct target of MALAT1. miR‑17‑5p expression was downregulated and FOXA1 expression was upregulated in LPS‑treated A549 cells. Further, MALAT1 knockdown promoted miR‑17‑5p expression and inhibited FOXA1 expression, whereas the combined suppression of MALAT1 and miR‑17‑5p induced FOXA1 expression. Moreover, miR‑17‑5p knockdown reversed the effects of MALAT1 suppression on LPS‑induced A549 cell proliferation. These results indicated that MALAT1 serves as a competing endogenous lncRNA that, by sequestering miR‑17‑5p, stimulates FOXA1 expression and mediates LPS‑induced A549 cell injury. In conclusion, the present study demonstrated that MALAT1 knockdown alleviates LPS‑induced A549 cell injury by targeting the miR‑17‑5p/FOXA1 axis.

The mechanism of miR-16-5p protection on LPS-induced A549 cell injury by targeting CXCR3

Objective: To study the effect of miR-16-5p on lung cancer cell injury and apoptosis, and its mechanism. Methods: LPS induced lung cancer cell A549 injury; qRT-PCR method was applied to detect the expression of miR-16-5p and CXCR3 in A549 cells. Con (without LPS treatment), LPS + miR-NC group (transfected negative control samples), LPS + miR-16-5p group (transfected miR-16-5p mimics); LPS + si-NC group (transfected negative control samples), LPS + si-CXCR3 group (transfected si-CXCR3); LPS + miR-16-5p + pcDNA3.1 group (co-transfected miR-16-5p mimics and pcDNA3.1), LPS + miR-16-5p + pcDNA3.1-CXCR3 group (co-transfected miR-16-5p mimics and pcDNA3.1-CXCR3) were transfected into A549 cells by liposome method. Western blot was used to detect protein expression of CXCR3, IL-6 and TNF-α in A549 cells; apoptosis of A549 cells was detected by flow cytometry. Results: Compared with the control group, the expression of miR-16-5p mRNA was significantly decreased in A549 cells in LPS group, and the mRNA and protein expression of CXCR3 were significantly increased (p < .05). Overexpression of miR-16-5p and knockdown of CXCR3 both can down-regulated protein expression of IL-6 and TNF-α, and up-regulated apoptosis in LPS-induced A549 cell; CXCR3 is a target of miR-16-5p. Overexpression of CXCR3 rescued the protective effect of miR-16-5p on LPS-induced A549 cell injury. Conclusion: miR-16-5p can protect LPS-induced A549 cell injury, and its mechanism may be related to the targeted regulation of CXCR3, which could provide a new target for targeted therapy of lung cancer.

Baicalin Weakens Staphylococcus aureus Pathogenicity by Targeting Sortase B.

Staphylococcus aureus (S. aureus) is a human and other animal pathogen that contributes to the primary etiology of nosocomial pneumonia, a disease with high mortality rates and costs. Treatment of multidrug-resistant S. aureus infection is extremely challenging, and new therapeutic strategies beyond antibiotic treatment are needed. Anti-virulence agents that specifically target the molecular determinants of virulence may be a novel method for treating drug-resistant nosocomial infections. Sortase B (SrtB) is a crucial virulence factor in S. aureus and plays an important role during infection. In this study, we find that baicalin suppresses the activity of SrtB. Minimum inhibitory concentration and growth curve assays confirmed that baicalin has no anti-S. aureus properties. We performed live/dead, lactate dehydrogenase (LDH), adherence, and enzyme-linked immunosorbent assays to confirm that baicalin reduced human alveolar epithelial A549 cell injury caused by S. aureus, reduced the adherence of S. aureus to A549 cells, and significantly attenuated the inflammatory response of mouse macrophage J774 cells to S. aureus. Additionally, we were able to elucidate the binding mechanics and identify the interacting sites of baicalin and SrtB via a molecular dynamics simulation, site-directed mutagenesis, and fluorescence spectroscopy quenching. Finally, we confirmed that baicalin directly binds to the active center of SrtB, and the residues Asn92 and Tyr128 perform an important function in the interaction of SrtB and baicalin. Taken together, these data indicate that baicalin is a promising candidate to combat S. aureus infections.

Enhanced ability of plasma-activated lactated Ringer's solution to induce A549 cell injury

Non-thermal plasma (NTP) is applicable to living cells and has emerged as a novel technology for cancer therapy. Plasma-activated medium (PAM), which is prepared by the irradiation of culture medium with NTP, induces cell death in cancer cells. However, difficulties are associated with applying PAM to the clinical phase because culture media cannot be used for medical treatments. The objectives of the present study were to demonstrate the inhibitory effects of plasma-activated lactated Ringer's solution (PAL) on the viability of the A549 cancer cell line and elucidate the underlying mechanisms. The anti-tumor activity of PAL was significantly stronger than that of PAM, whereas their concentrations of H2O2 and nitrite were similar. Lactated Ringer's solution (Lac-R) consists of lactate and three types of inorganic salts. The results showing that NTP irradiation of the lactate solution rather than the inorganic salt solution induced the inactivation of catalase were dependent on the presence or absence of nitrite in these solutions. We detected nitrotyrosine in A549 cells treated with PAM or PAL, and the addition of catalase to PAM rather than to PAL reduced its production. The PAL treatment of A549 cells led to mitochondrial dysfunction with the down-regulation of NF-κB-Bcl2 signaling.