Normal Human Lung Epithelial Cells Research Articles

Proteomic analysis reveals the molecular mechanism of Astragaloside in the treatment of non-small cell lung cancer by inducing apoptosis

BackgroundAstragaloside III (AS III), a saponin-like metabolite derived from the traditional Chinese medicine Astragali Radix, has been shown to be effective in the treatment of cancer and heart failure, and a variety of digestive disorders. However, its molecular mechanism in the treatment of non-small cell lung cancer (NSCLC) is unknown.MethodsHuman lung cancer A549 cells and NCI-H460 cells and a normal human lung epithelial cell BEAS-2B were treated with different concentrations of AS III. CCK-8 and EdU staining were used to determine the anti-proliferative effects of AS III in vitro. Quantitative proteomic analysis was performed on A549 cells treated with the indicated concentrations of AS III, and the expression levels of apoptosis-related proteins were examined by Western blotting.ResultsAS III treatment significantly inhibited proliferation and increased apoptosis in A549 and H460 cells and modulated functional signaling pathways associated with apoptosis and metabolism. At the molecular level, AS III promoted a reduction in the expression of ANXA1 (p < 0.01), with increased levels of cleaved Caspase 3 and PARP 1. In addition, AS III treatment significantly decreased the LC3-I/LC3-II ratio. The results of experiment in vitro showed that AS III promoted NSCLC apoptosis by down-regulating the phosphorylation levels of P38, JNK, and AKT (p < 0.01), inhibiting the expression of Bcl-2 (p < 0.01), and up-regulating the expression of Bax (p < 0.01).ConclusionThese findings provide a mechanism whereby AS III treatment induces apoptosis in NSCLC cells, which may be achieved in part via modulation of the P38, ERK and mTOR signaling pathways.

FOXA1 prolongs S phase and promotes cancer progression in non-small cell lung cancer through upregulation of CDC5L and activation of the ERK1/2 and JAK2 pathways.

Non-small cell lung cancer (NSCLC) causes high mortality worldwide; however, its molecular pathways have not been fully investigated. The relationship between FOXA1 and CDC5L as well as their roles in NSCLC have not been comprehensively studied. Clinical tissues were collected from 78 NSCLC patients for clinical studies. The BEAS-2B human normal lung epithelial cell line and the A549, Calu-3, H526 and H2170 human NSCLC cell lines were used for in vitro studies. sh-FOXA1 and oe-CDC5L constructs were used to generate knockdown and overexpression models, respectively. The CCK-8 assay was used to analyze cell viability. The cell cycle and apoptosis were evaluated by flow cytometry analysis. The relationship between FOXA1 and CDC5L was demonstrated using dual-luciferase and ChIP assays. Gene levels were examined via immunohistochemistry, qRT-PCR and western blot analysis. FOXA1 levels were increased in NSCLC clinical tissues and cell lines. Depletion of FOXA1 increased the apoptosis rate and increased the proportion of cells in G2/M phase. In addition, we demonstrated that FOXA1 was directly bound to the promoter of CDC5L and that depletion of FOXA1 inhibited CDC5L expression. Overexpression of CDC5L induced ERK1/2 phosphorylation, induced JAK2 phosphorylation, inhibited cell apoptosis, prolonged S phase, and significantly reversed the effects of FOXA1 knockdown on the progression of NSCLC. The present study demonstrated that FOXA1 prolongs S phase and promotes NSCLC progression through upregulation of CDC5L and activation of the ERK1/2 and JAK2 pathways.

Airborne transmission of human-isolated avian H3N8 influenza virus between ferrets

H3N8 avian influenza viruses (AIVs) in China caused two confirmed human infections in 2022, followed by a fatal case reported in 2023. H3N8 viruses are widespread in chicken flocks; however, the zoonotic features of H3N8 viruses are poorly understood. Here, we demonstrate that H3N8 viruses were able to infect and replicate efficiently in organotypic normal human bronchial epithelial (NHBE) cells and lung epithelial (Calu-3) cells. Human isolates of H3N8 virus were more virulent and caused severe pathology in mice andferrets, relative to chicken isolates. Importantly, H3N8 virus isolated from a patient with severe pneumonia was transmissible between ferrets through respiratory droplets; it had acquired human-receptor-binding preference and amino acid substitution PB2-E627K necessary for airborne transmission. Human populations, even when vaccinated against human H3N2 virus, appear immunologically naive to emerging mammalian-adapted H3N8 AIVs and could be vulnerable to infection at epidemic or pandemic proportion.

Radioprotective efficacy of Astilbin in mitigating radiation-induced lung injury through inhibition of p53 acetylation.

Radiation-induced lung injury (RILI) is a common side effect in thoracic tumor patients undergoing radiotherapy. At present, there is no ideal radio-protective agent which is widely used in RILI treatment. Astilbin (AST), a bioactive flavonoid, exhibits various biological effects, including anti-inflammatory, antioxidant, and anti-fibrotic activities, which partly result from reducing oxidative stress and inflammation in various pathogenic conditions. However, the protective efficacy of AST to ameliorate RILI has not been reported. In this study, we employed network pharmacology, RNA sequencing, and experimental evaluation to reveal the effects and pharmacological mechanism of AST to treat RILI in vivo and in vitro. We observed that AST reduced radiation-induced apoptosis, DNA damage, inflammatory reactions, and the reactive oxygen species (ROS) level in human normal lung epithelial cells BEAS-2B. Further study showed that AST treatment significantly ameliorated RILI by reducing the radiation-induced pathology changes and inflammatory reaction of lung tissue in C57BL/6J mice. Mechanistically, the expression of epithelial-mesenchymal transition (EMT) markers and radiation-triggered acetylation of the p53 protein were alleviated by AST treatment. Furthermore, AST alleviated the acetylation of p53 after intervention of Trichostatin A (TSA). Our data indicate that AST can alleviate RILI by inhibiting inflammatory reactions and the EMT process through decreasing the expression of p53 acetylation. In conclusion, our study suggests that AST has great potential to be a new protective and therapeutic compound for RILI.

Insight into the lncRNA-mRNA Co-Expression Profile and ceRNA Network in Lipopolysaccharide-Induced Acute Lung Injury.

Long non-coding RNAs (lncRNAs) participate in acute lung injury (ALI). However, their latent biological function and molecular mechanism have not been fully understood. In the present study, the global expression profiles of lncRNAs and mRNAs between the control and lipopolysaccharide (LPS)-stimulated groups of human normal lung epithelial cells (BEAS-2B) were determined using high-throughput sequencing. Overall, a total of 433 lncRNAs and 183 mRNAs were differentially expressed. A lncRNA-mRNA co-expression network was established, and then the top 10 lncRNAs were screened using topological methods. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analysis results showed that the key lncRNAs targeting mRNAs were mostly enriched in the inflammatory-related biological processes. Gene set variation analysis and Pearson's correlation coefficients confirmed the close correlation for the top 10 lncRNAs with inflammatory responses. A protein-protein interaction network analysis was conducted based on the key lncRNAs targeting mRNAs, where IL-1β, IL-6, and CXCL8 were regarded as the hub genes. A competing endogenous RNA (ceRNA) modulatory network was created with five lncRNAs, thirteen microRNAs, and twelve mRNAs. Finally, real-time quantitative reverse transcription-polymerase chain reaction was employed to verify the expression levels of several key lncRNAs in BEAS-2B cells and human serum samples.

MiR-183-5p promotes the progression of non-small cell lung cancer through targeted regulation of FOXO1

Abstract Objective To investigate miR-183-5p targeting to forkhead box protein O1 (FOXO1) and its corresponding effect on the proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) of non-small cell lung cancer (NSCLC) cells. Methods NSCLC tissues and adjacent normal tissues from 60 patients with NSCLC adenocarcinoma were obtained via pathological biopsy or intraoperative resection. Several cell lines were cultured in vitro, including the human normal lung epithelial cell line BEAS-2B and human NSCLC cell lines A549, SPCA-1, PC-9, and 95-D. miR-183-5p and FOXO1 mRNA expression in tissues and cells were detected by qRT-PCR; the corresponding correlations in NSCLC tissues were analyzed using the Pearson test, and the relationship between miR-183-5p expression and clinicopathological parameters was analyzed. The miR-183-5p-mediated regulation of FOXO1 was verified by bioinformatics prediction alongside double luciferase, RNA-binding protein immunoprecipitation (RIP) assay, and pull-down experiments. A549 cells were divided into control, anti-miR-NC, anti-miR-183-5p, miR-NC, miR-183-5p, miR-183-5p+pcDNA3.1, and miR-183-5p+pcDNA3.1-FOXO1 groups. Cell proliferation, invasion, migration, apoptosis, and cell cycle distribution were detected using an MTT assay, clone formation assay, Transwell assay, scratch test, and flow cytometry, respectively. The expression of EMT-related proteins in the cells was analyzed by western blotting. The effect of miR-185-3p silencing on the development of transplanted tumors was detected by analyzing tumor formation in nude mice. Results miR-183-5p expression was significantly higher in NSCLC tissues and cells than in adjacent normal tissues, whereas FOXO1 mRNA expression was significantly down-regulated. There was a significant negative correlation between miR-183-5p and FOXO1 mRNA in NSCLC tissues (P &lt; 0.05). Additionally, the expression of miR-183-5p was significantly correlated with tumor size, tumor differentiation, and tumor-node-metastasis stage in patients with NSCLC (P &lt; 0.05). miR-183-5p targeted and inhibited FOXO1 expression. Compared to the anti-miR-NC group, the cell proliferation, scratch healing rate, N-cadherin and vimentin protein expression, and the proportion of S phase cells were significantly lower in the anti-miR-183-5p group, whereas the protein expression of E-cadherin and α-catenin and the proportion of G0/G1 phase cells were significantly higher; additionally, the frequency of colony formation and invasion were significantly lower in the anti-miR-183-5p group (P &lt; 0.05). Compared to the miR-NC group, the cell proliferation, scratch healing rate, N-cadherin and vimentin protein expression, and the proportion of S phase cells in the miR-183-5p group were significantly higher, whereas the E-cadherin and α-catenin protein expression and the proportion of G0/G1 phase cells were significantly lower; furthermore, the frequency of colony formation and invasion were significantly higher in the miR-183-5p group (P &lt; 0.05). Compared with the miR-183-5p+pcDNA3.1 group, the OD value, scratch healing rate, N-cadherin and vimentin protein expression, and the proportion of S phase cells were significantly lower in the miR-183-5p+pcDNA3.1-FOXO1 group, whereas E-cadherin and α-catenin protein expression and the proportion of G0/G1 phase cells were significantly higher; additionally, the frequency of colony formation and invasion was significantly lower in the miR-183-5p+pcDNA3.1-FOXO1 group (P &lt; 0.05). Overall, silencing miR-185-3p inhibited the growth of transplanted tumors and promoted FOXO1 expression. Conclusion Overexpression of miR-183-5p can inhibit apoptosis and promote the proliferation, migration, invasion, and EMT, of NSCLC cells by down-regulating FOXO1 expression.

LncRNA SH3BP5-AS1 Regulates the Proliferation and Cell Cycle of Non-Small Cell Lung Cancer Cells

Background: Non-small cell lung cancer (NSCLC) consists of a class of heterogeneous diseases. Objective: LncRNAs are exceedingly implicated in the pathogenesis of NSCLC. Herein, the current study set out to illustrate the molecular mechanism of SH3BP5-AS1 in NSCLC cells. Methods: SH3BP5-AS1 expression in clinical NSCLC tissues and its impact on prognosis were analyzed by bioinformatics database. SH3BP5-AS1 expression patterns in NSCLC cell lines (A549/H1299/H1975/H460) and human normal lung epithelial cell lines (BEAS-2B) were examined by RT-qPCR. SH3BP5-AS1 was overexpressed in A549 or silenced in H1975 cells through transfection to assess its effect on proliferation, cell cycle distribution, and apoptosis, apoptosisrelated protein (Cleaved Caspase-3, Bax, Bcl-2) levels, invasive, migratory, and healing capacity through CCK-8, colony formation assay, flow cytometry, Western blot, Transwell, and cell scratch test. Results: SH3BP5-AS1 was under-expressed in NSCLC clinical tissues, and NSCLC patients with low SH3BP5-AS1 expression showed poor prognosis. A549/H1299/H1975/H460 cells had reduced levels of SH3BP5-AS1, with the relative level lowest/highest expression in A549/H1975 cells, respectively. SH3BP5-AS1 overexpression repressed A549 cell proliferation, slowed down cell cycle progression, enhanced apoptosis, elevated Cleared Caspase-3, Bax, suppressed Bcl-2 protein levels, and inhibited migratory, invasive, and scratch healing capacities, while SH3BP5-AS1 silencing brought about the opposite results in H1975 cells. Conclusion: SH3BP5-AS1 could suppress NSCLC cell proliferation, slow down cell cycle progression, stimulate apoptosis, and limit invasion and migration.

LINC00638 promotes the progression of non-small cell lung cancer by regulating the miR-541-3p/IRS1/PI3K/Akt axis

BackgroundPreceding works reveal the function of long non-coding RNAs (abbreviated to lncRNAs) during non-small cell lung cancer (NSCLC) evolvement. We explored the profile and biological functions of the lncRNA LINC00638 in NSCLC. MethodsReverse transcription-quantitative PCR examined LINC00638 level in NSCLC and corresponding non-tumor tissues, human normal lung epithelial cells BEAS-2B, and NSCLC cells (NCI–H460, HCC-827, A549, H1299, H1975, H460). The gain- and loss-of-function assay of LINC00638 ascertained its function in modulating the proliferation, apoptosis, and invasion of NSCLC cells (HCC-827 and H460). Bioinformatics analysis investigated the underlying mechanisms. Dual luciferase reporter gene and RNA immunoprecipitation (RIP) checked the interactions between LINC00638 and microRNA (miR)-541-3p, miR-541-3p and insulin receptor substrate 1 (IRS1). ResultsLINC00638 was upregulated in NSCLC tissues by contrast to the profiles found in the corresponding non-tumor normal tissues, as well as in NSCLC cells vis-à-vis BEAS-2B cells. LINC00638 upregulation pertained to the poorer survival rates of NSCLC patients. Overexpressing LINC00638 augmented NSCLC cells’ proliferation, growth, migration, and invasion but inhibited their apoptosis, while down-regulating LINC00638 led to the opposite. miR-541-3p might be an underlying target of LINC00638, which targeted IRS1, inhibited NSCLC progression, and reversed the carcinogenic effects of LINC00638. Mechanistically, LINC00638/miR-541-3p regulated the IRS1/phosphoinositide 3-kinase (PI3K)/Akt signaling pathway. Repressing IRS1/2 using its inhibitor NT157 repressed LINC00638-mediated oncogenic effects. ConclusionLINC00638 may function as an oncogene in NSCLC by modulating the miR-541-3p/IRS1/PI3K/Akt axis.

Linc01515 regulates PM2.5-induced oxidative stress via targeting NRF2 in airway epithelial cells

Dysregulation of long non-coding RNAs (lncRNAs) is involved in the adverse effects caused by fine particulate matter (PM2.5). However, the molecular mechanism is not fully clarified. In this study, we performed lncRNA sequencing on PM2.5-treated human bronchial epithelial (HBE) cells to identify vital lncRNAs, and verified the differential expression of the lncRNAs by RT-qPCR in HBE and human normal lung epithelial (BEAS-2B) cells. A total of 657 and 652 lncRNAs were dysregulated after exposure to 125 and 250 μg/mL of PM2.5, respectively. Of these, lncRNA linc01515 was upregulated in HBE and BEAS-2B cells with PM2.5 treatment. Subcellular localization experiments showed that linc01515 was mostly localized in the nucleus. Functionally, we downregulated the expression of linc01515 in HBE and BEAS-2B cells before PM2.5 treatment, which can decrease malonydialdehyde (MDA) and reactive oxygen species (ROS) levels, and improve superoxide dismutase (SOD) activity. Correspondingly, linc01515 overexpression enhanced PM2.5-induced oxidative injury in airway epithelial cells. Mechanistically, N6-methyladenosine RNA binding protein immunoprecipitation (MeRIP) assay showed that the enrichment level of m6A on linc01515 was increased after PM2.5 treatment, and the m6A modification level and expression of linc01515 was decreased in the HBE cells with 3-deazaadenosine (DAA) treatment or knockdown of METTL3 to inhibit the RNA methylation level. Western blot found that NRF2, a vital transcription factor, was enhanced remarkably in linc01515-silenced cells and decreased in linc01515-overexpressed cells. Furthermore, inhibition of NRF2 activity significantly rescued effect of downregulated linc01515 expression on PM2.5-induced cytotoxicity. In addition, we observed the similar effect when downregulating linc01515 and NRF2 expression in HBE and BEAS-2B cells before PM2.5 treatment. Taken together, our findings demonstrated that PM2.5 treatment may upregulate the expression of linc01515 by enhancing its m6A modification, and then regulate NRF2 to induce oxidative damage of airway epithelial cells.

N6-methyladenosine modification contributes to respiratory syncytial virus infection

BackgroundRespiratory syncytial virus (RSV) is the second leading cause of death due to lower respiratory tract infections. Effective prevention and treatment measures are lacking, posing a huge socioeconomic burden to the world. N6-methyladenosine (m6A) is the most common internal modification in messenger RNA and noncoding RNA. Numerous recent studies have shown that the dysregulation of m6A modification is associated with diseases caused by pathogenic viruses. MethodsThe changes in m6A modification were evaluated using m6A RNA methylation assay. The differences in gene expression levels of various m6A-modifying enzymes were observed using Quantitative Real-time PCR (qRT-PCR) during RSV infection. The autophagosomes were observed using transmission electron microscopy, and the expression of autophagy-associated protein Microtubule Associated Protein 1 Light Chain 3 Beta Ⅱ/Ⅰ (LC3B Ⅱ/Ⅰ) and Beclin1 in Human Normal Lung Epithelial Cells (BEAS-2B) cells using Western blot during RSV infection. The significantly differentially expressed genes were screened guided by bioinformatics. Their relationship with m6A-modifying enzymes was analyzed through protein–protein interaction network and expression correlation analysis. ResultsThe m6A abundance decreased and demethylase Fat Mass and Obesity- Associated Protein (FTO) significantly increased during RSV infection after 24 h. We also found that the DNA Damage-Inducible Transcript 3 Protein (DDIT3) level significantly increased during RSV infection after 24 h and observed autophagosomes in BEAS-2B cells. In addition, RSV infection could cause the upregulation of LC3B Ⅱ/Ⅰ and Beclin1. The expression correlation analysis showed that DDIT3 levels were positively correlated with the FTO level, and Methyltransferase Like 3 (METTL3), RNA Binding Motif Protein 15B (RBM15B), YTH Domain-Containing Family Protein 1 (YTHDF1), and levels were negatively correlated with the DDIT3 level. ConclusionsWe uncovered a significant role for m6A modification during RSV infection. Also, a correlation was found between m6A and autophagy, providing new ideas for therapeutic advancements in RSV treatment.

Long term exposure of cigarette smoke condensate (CSC) mediates transcriptomic changes in normal human lung epithelial Beas-2b cells and protection by garlic compounds.

Chronic cigarette smoke condensate (CSC) exposure is one of the preventable risk factors in the CS-induced lung cancer. However, understanding the mechanism of cellular transformation induced by CS in the lung remains limited. We investigated the effect of long term exposure of CSC in human normal lung epithelial Beas-2b cells, and chemopreventive mechanism of organosulphur garlic compounds, diallyl sulphide (DAS) and diallyl disulphide (DADS) using Next Generation Sequencing (NGS) transcriptomic analysis. CSC regulated 1077 genes and of these 36 genes are modulated by DAS while 101 genes by DADS. DAS modulated genes like IL1RL1 (interleukin-1 receptor like-1), HSPA-6 (heat shock protein family A, member 6) while DADS demonstrating ADTRP (Androgen-Dependent TFPI Regulating Protein), ANGPT4 (Angiopoietin 4), GFI1 (Growth Factor-Independent 1 Transcriptional Repressor), TBX2 (T-Box Transcription Factor 2), with some common genes like NEURL-1 (Neuralized E3-Ubiquitin Protein Ligase 1), suggesting differential effects between these two garlic compounds. They regulate genes by influencing pathways including HIF-1alpha, STAT-3 and matrix metalloproteases, contributing to the chemoprotective ability of organosulfur garlic compounds against CSC-induced cellular transformation. Taken together, we demonstrated CSC induced global gene expression changes pertaining to cellular transformation which potentially can be delayed with dietary chemopreventive phytochemicals like DS and DADS influencing alterations at the transcriptomic level.

MiR-508-5p serves as an anti-oncogene by targeting S100A16 to regulate AKT signaling and epithelial-mesenchymal transition process in lung adenocarcinoma cells

BackgroundOur aim was to expose the effect of miR-508-5p on the developmental and biological behaviour of lung adenocarcinoma (LUAC). MethodsThe KM plotter was used to analyze the survival significance of miR-508-5p and S100A16 expression in LUAC patients. qRT-PCR was performed to detect the expression of miR-508-5p and S100A16 in LUAC tissue and LUAC cell lines. CCK8, colony formation and Transwell were performed to evaluate the effects of miR-508-5p and S100A16 on cell proliferation and metastasis. Dual luciferase reporter assay was used to verify that S100A16 were targets of miR-508-5p. Western blot analysis was performed to analyze protein expression. ResultsResults showed that low miR-508-5p expression in LUAC tissues indicated poorer overall survival of LUAC patients and miR-508-5p was downregulated in LUAC cell lines compared to the normal human lung epithelial cell line. miR-508-5p mimics could inhibit A549 cell proliferation and metastasis abilities, while miR-508-5p Antagomir showed the opposite effect. We identified S100A16 as one direct target of miR-508-5p, and rescuing S100A16 expression could reverse the effect of miR-508-5p mimics on A549 cell proliferation and metastasis. miR-508-5p could involve the coordination of AKT signaling and epithelial-mesenchymal transition (EMT) progress using western-blot assays and rescuing S100A16 expression could reverse the inhibited AKT signaling and EMT progress induced by miR-508-5p mimics. ConclusionsWe found that miR-508-5p targeted S100A16 to regulate AKT signaling and EMT progress in A549 cells, resulting in impaired cell proliferation and metastasis activity, suggesting that miR-508-5p might be a promising therapeutic target and an important diagnostic and prognostic marker for improved LUAC therapeutic schedule.

Downregulation of angulin-1/LSR induces malignancy via upregulation of EGF-dependent claudin-2 and TGF-β-dependent cell metabolism in human lung adenocarcinoma A549 cells.

Abnormal expression of bicellular tight junction claudins, including claudin-2 are observed during carcinogenesis in human lung adenocarcinoma. However, little is known about the role of tricellular tight junction molecule angulin-1/lipolysis-stimulated lipoprotein receptor (LSR). In the lung adenocarcinoma tissues examined in the present study, expression of claudin-2 was higher than in normal lung tissues, while angulin-1/LSR was poorly or faintly expressed. We investigated how loss of angulin-1/LSR affects the malignancy of lung adenocarcinoma cell line A549 and normal human lung epithelial (HLE) cells. The EGF receptor tyrosine kinase inhibitor AG1478 prevented the increase of claudin-2 expression induced by EGF in A549 cells. Knockdown of LSR induced expression of claudin-2 at the protein and mRNA levels and AG1478 prevented the upregulation of claudin-2 in A549 cells. Knockdown of LSR induced cell proliferation, cell migration and cell metabolism in A549 cells. Knockdown of claudin-2 inhibited the cell proliferation but did not affect the cell migration or cell metabolism of A549 cells. The TGF-β type I receptor inhibitor EW-7197 prevented the decrease of LSR and claudin-2 induced by TGF-β1 in A549 cells and 2D culture of normal HLE cells. EW-7197 prevented the increase of cell migration and cell metabolism induced by TGF-β1 in A549 cells. EW-7197 prevented the increase of epithelial permeability of FITC-4kD dextran induced by TGF-β1 in 2.5D culture of normal HLE cells. In conclusion, downregulation of angulin-1/LSR induces malignancy via EGF-dependent claudin-2 and TGF-β-dependent cell metabolism in human lung adenocarcinoma.

Immune infiltration patterns and identification of new diagnostic biomarkers GDF10, NCKAP5, and RTKN2 in non-small cell lung cancer

This study aimed to identify potential biomarkers for non-small cell lung cancer (NSCLC) and analyze the role of immune cell infiltration in NSCLC. R software was used to screen differentially expressed genes (DEGs) from NSCLC datasets obtained from the Gene Expression Omnibus (GEO) database, and functional correlation analysis was performed. The machine learning algorithms were used to screen the potential biomarkers of NSCLC. The diagnostic values were assessed through receiver operating characteristic (ROC) curves. The protein and mRNA expression levels of potential biomarkers were verified based on the Human Protein Atlas (HPA) database and qRT-PCR. CIBERSORT was used to evaluate the infiltration of immune cells in NSCLC tissues, and the correlation between potential biomarkers and infiltrated immune cell was analyzed. Finally, specific siRNAs were utilized to reduce the GDF10, NCKAP5, and RTKN2 expression in A549 and H1975 cells. The proliferation ability of A549 and H1975 cells was detected by MTT assay. A total of 848 upregulated DEGs and 1308 downregulated DEGs were identified. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses showed that the DEGs were mainly related to cell division. Disease ontology (DO) enrichment analysis showed that the diseases with these DEGs were mainly lung diseases, including NSCLC. In addition,three potential biomarkers were identified: GDF10, NCKAP5, and RTKN2. Immune cell infiltration analysis showed that resting NK cells, activated dendritic cells, and Tregs may be involved in the pathogenesis of NSCLC. Meanwhile, GDF10, NCKAP5, and RTKN2 were negatively correlated with Tregs and naïve B cells but were positively correlated with activated dendritic cells and resting NK cells. Immunohistochemical staining showed that the expression of GDF10, NCKAP5, and RTKN2 in the lung tissue of patients with NSCLC was lower than that of normal lung tissue. qRT-PCR also confirmed that the mRNA expression of three biomarkers in NSCLC cell lines A549 and H1975 were significantly lower than those in human normal lung epithelial cells BEAS-2B. An MTT assay showed that GDF10, NCKAP5, and RTKN2 knockdown significantly promoted the proliferation of A549 and H1975 cells. The in vitro experiments showed that GDF10, NCKAP5, and RTKN2 played the inhibitory effects on NSCLC cell lines proliferation. Hence, GDF10, NCKAP5, and RTKN2 can be used as diagnostic biomarkers for NSCLC.

Biocompatible polymers with tunable mechanical properties and conductive functionality on two-photon 3D printing.

Two-photon polymerization (TPP)-based 3D printing technology utilizes the two-photon absorption process of near-infrared radiation, enabling the fabrication of micro- and nano-scale three-dimensional structures with extremely high resolution. It has been widely applied in scientific fields closely related to living organisms, such as tissue engineering, drug delivery, and biosensors. Nevertheless, the existing photoresist materials have poor mechanical tunability and are hardly able to be doped with functional materials, resulting in constraints on the preparation of functional devices with micro-nano structures. In this paper, TPP printable polymer formulas with good mechanical tunability, high resolution, strong functional scalability, and excellent biocompatibility are proposed, by using the synergistic effects of a hydroxyl group-containing photocurable resin prepolymer, UV acrylate monomer, long-chain hydrophilic crosslinking monomer and photo-initiator. This can ensure the printability and help to improve the flexibility of the printed polymer, thereby solving the problem the photosensitive materials suitable for two-photon 3D printing in previous research had in balancing the formability and flexibility. The results of nanoindenter analysis showed that the Young's modulus of the printed structure can be adjusted between 0.3 GPa and 1.43 GPa, realizing mechanical tunability. Also, complex structures, such as micro-scaffold structures and high aspect ratio hollow microneedles were printed to explore the structural stability as well as the feasibility of biodevice application. Meanwhile, the proposed polymer formula can be functionalized to be conductive by doping with functional nanomaterial MXene. Finally, the biocompatibility of the proposed polymer formula was studied by culturing with human normal lung epithelial cells. The results indicated a good potential for biodevice applications.

Adsorption of porcine pulmonin can reduce the toxicity of gold nanorods and reduce lung injury

It is well known that porcine lung phospholipid injection (Curosurf) can effectively inhibit lung inflammation and reduce inflammatory cell damage to lung tissue, but whether it can reduce nanoparticles damage to lung remains to be seen. Poractant was co-incubated with gold nanorods as the model particle in this study to adsorb poractant on the surface of gold nanorods. Elisa kit was used to determine the adsorption degree of human lung surfactant associated protein poractant on the surface of gold nanorods. Gold nanorods adsorbed human lung surfactants associated proteins were evaluated further in vitro and in vivo. The toxicity of gold nanorods after absorption of porcine lung phospholipid was studied using human normal lung epithelial cells BEAS-2B; TNF- and IL-1 in serum were studied after absorption of porcine lung phospholipid using acute lung injury as a model. The results of the experiments show that adsorption of porcine lung phospholipids can reduce the toxicity of gold nanorods and the lung damage caused by gold nanorods.

Decursin alleviates LPS-induced lung epithelial cell injury by inhibiting NF-κB pathway activation.

To reveal the possible effects of decursin on viability, oxidative stress, and inflammatory response in lipopolysaccharide (LPS)-treated human bronchial epithelial cells-2B (BEAS-2B) and human pulmonary artery endothelial cells (HPAEC) cells, and revealed the potential mechanisms. LPS was used to induce acute lung injury (ALI) in normal human lung epithelial cells, including BEAS-2B and HPAEC cells. Cell viability and apoptosis in response to LPS and decursin in BEAS-2B and HPAEC cells were, respectively, evaluated by MTT colorimetric and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assays. The oxidative stress and inflammatory response in LPS-treated BEAS-2B and HPAEC cells were detected by enzyme-linked-immunosorbent serologic assay. In addition, the role of decursin in nuclear -factor-kappa B (NF-κB) activation was analyzed by immunoblot and immunofluorescence assays. Our data revealed that decursin could alleviate the viability of LPS-induced BEAS-2B and HPAEC cells. Decursin could also reduce LPS-induced oxidative stress in BEAS-2B and HPAEC cells. In addition, it could reduce LPS-induced inflammation in BEAS-2B and HPAEC cells. Mechanically, decursin suppressed the activation of NF-κB pathway. Decursin suppressed NF-κB pathway, and therefore alleviated ALI.

Artemisinin-isatin hybrids tethered via ethylene linker and their anti-lung cancer activity.

The synthesized 11 artemisinin-isatin hybrids 5a-c and 6a-h tethered via ethylene linker were assessed for their in vitro antiproliferative activity against A549 and H1299 nonsmall-cell lung cancer cell lines as well as their cytotoxicity towards BEAS-2B human normal lung epithelial cells. The preliminary results showed that hybrids 5a-c and 6a-h did not show any cytotoxicity (IC50 : >100 µM) on BEAS-2B cells, and also possessed potential activity (IC50 : 6.99-76.49 µM) against A549 and H1299 lung cancer cell lines. The representative hybrid 6c (IC50 : 6.99 and 7.57 µM) was far more potent than artemisinin (IC50 : >100 µM) and dihydroartemisinin (IC50 : >100 µM), and was slightly less active than doxorubicin (IC50 : 4.14 and 2.77 µM). Moreover, hybrid 6c also exhibited an excellent safety profile and good selectivity with SI values of >13.21. Therefore, hybrid 6c could serve as a promising candidate for further in vivo evaluations.

Circ-PGAM1 Enhances Matrine Resistance of Non-Small Cell Lung Cancer via the miR-326/CXCR5 Axis.

Background: Circular RNAs (circ-RNAs) have been demonstrated to influence initiation, drug resistance, and metastasis of tumors. However, the effects of circular-phosphoglycerate mutase 1 (circ-PGAM1) on matrine resistance in nonsmall cell lung cancer (NSCLC) remain unknown. Materials and Methods: The reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was used to determine gene expression. The terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and cell colony formation assays were used to evaluate NSCLC apoptosis and cell proliferation after indicated treatments, respectively. Results: circ-PGAM1 was upregulated in human NSCLC cell lines (H1299 and A549) compared with the human normal lung epithelial (BEAS-2B) cells. circ-PGAM1 overexpression reversed the matrine treatment-induced inhibition on proliferation of NSCLC cells (A549 and H1299) and rescued the matrine treatment-stimulated apoptosis of these cells. miR-326 was demonstrated to interact with circ-PGAM1. circ-PGAM1 knockdown enhanced the antitumor effect of matrine on NSCLC cell proliferation and apoptosis, which was reversed by miR-326 inhibition. The authors also identified CXCR5 as a key downstream target of miR-326 in A549 cells. Conclusions: circ-PGAM1 enhances matrine resistance of NSCLC cells through the miR-326/CXCR5 axis. The authors' findings provide new insights into NSCLC-targeted therapy.

Synthesis, biological evaluation and preliminary mechanisms of 6-amino substituted harmine derivatives as potential antitumor agents

To explore the effect of the introduction of the amino and substituted amino groups on the antitumor activity of harmine, twenty-five novel 6-amino substituted harmine derivatives (3a–3j and 5a–5o) were synthesized and evaluated for anti-proliferative activity on a panel of cancer cell lines. Compounds 3i and 5n exhibited the most potent antiproliferative activity with IC50 values lower than 2.2 μM. Especially, compound 5n possessed extremely potent antitumor activity with IC50 values of 0.34 μM and 0.65 μM against HL-60 and A549 cell lines, respectively. Further, the preliminary studies of mechanisms showed that compound 5n could significantly induce cell apoptosis in a dose-dependent manner, cause cell cycle arrest at the G2/M phase and intercalate into ct-DNA via the competition with EB, while displaying very weak topoisomerase I (Topo I) inhibition activity. More importantly, 5n showed mild cytotoxicity against human normal lung epithelial cells BEAS-2B. Based on these considerations, 5n may be a good antitumor candidate compound for further exploration.