Lung Research Articles

Single Cell MALDI-MSI Analysis of Lipids and Proteins within a Replicative Senescence Fibroblast Model.

In this study, we evaluate lipids and select proteins in human lung fibroblasts (hLFs) to interrogate changes occurring due to aging and senescence. To study single cell populations, a comparison of cells adhered onto slides using poly-d-lysine versus centrifugal force deposition was first analyzed to determine whether specific alterations were observed between preparations. The poly-d-lysine approach was then utilized to interrogate the lipidome of the cell populations and further evaluate potential applications of the MALDI-immunohistochemistry (IHC) platform for single-cell-level analyses. Two protein markers of senescence, vimentin and p21, were both observed within the fibroblast populations and quantified. Lipidomic analysis of the fibroblasts found 12 lipids significantly altered because of replicative senescence, including fatty acids, such as stearic acid, and ceramide phosphoethanolamine species (CerPE). Similar to previous reports, alterations were detected in putative fatty acid building blocks, ceramides, among other lipid species. Altogether, our results reveal the ability to detect lipids implicated in senescence and show alterations to protein expression between normal and senescent fibroblast populations, including differences between young and aged cells. This report is the first time that the MALDI-IHC system has been utilized at a single-cell level to analyze both protein expression and lipid profiles in cultured cells, with a particular focus on changes associated with aging and senescence.

Acquisition of a multibasic cleavage site does not increase MERS-CoV entry into Calu-3 human lung cells.

Human-to-human transmission of the highly pathogenic Middle East respiratory syndrome coronavirus (MERS-CoV) is currently inefficient. However, there is concern that the virus might mutate and thereby increase its transmissibility and thus pandemic potential. The pandemic SARS-CoV-2 depends on a highly cleavable furin motif at the S1/S2 site of the viral spike (S) protein for efficient lung cell entry, transmission, and pathogenicity. Here, by employing pseudotyped particles, we investigated whether augmented cleavage at the S1/S2 site also increases MERS-CoV entry into Calu-3 human lung cells. We report that polymorphism T746K at the S1/S2 cleavage site or optimization of the furin motif increases S protein cleavage but not lung cell entry. These findings suggest that, unlike what has been reported for SARS-CoV-2, a highly cleavable S1/S2 site might not augment MERS-CoV infectivity for human lung cells.IMPORTANCEThe highly cleavable furin motif in the spike protein is required for robust lung cell entry, transmission, and pathogenicity of SARS-CoV-2. In contrast, it is unknown whether optimization of the furin motif in the spike protein of the pre-pandemic MERS-CoV increases lung cell entry and allows for robust human-human transmission. The present study indicates that this might not be the case. Thus, neither a naturally occurring polymorphism that increased MERS-CoV spike protein cleavage nor artificial optimization of the cleavage site allowed for increased spike-protein-driven entry into Calu-3 human lung cells.

VRAC Complex Modulates Mechano-Electrical Signal Responses in Human Airway Smooth Muscle Shortening.

Leucine-rich repeat containing 8A (LRRC8A) is an obligatory constituent of the volume-regulated anion channel (VRAC) that is fundamental to a wide range of biological processes, including regulating cell size, proliferation, and migration. Here we explored the physiological role for VRAC in excitation-contraction (E-C) coupling and shortening of human airway smooth muscle (HASM). In HASM cells, pharmacological inhibition of VRAC with DCPIB (0.1-10 μM) markedly attenuated swell-activated Cl- conductance and contractile agonist (histamine or carbachol)-induced cellular stiffening as measured by single-cell patch clamp and optical magnetic twisting cytometry, respectively. In addition, HASM cells treated with DCPIB or transfected with LRRC8A-targeting siRNA showed reduced agonist-induced phosphorylation of protein kinase B (AKT), paxillin, myosin phosphatase target subunit 1 (MYPT1), and myosin light chain (MLC). Consistent with the changes of these E-C coupling effectors, DCPIB appreciably decreased agonist-induced small airways narrowing in human precision-cut lung slices (hPCLS). Taken together, our findings shed a new light on the mechanistic link between HASM shortening and regulatory volume decrease via LRRC8A, revealing a previously unrecognized nodal point for modulation of the E-C coupling and acute airways constriction.

Branched-chain α-ketoacids aerobically activate HIF1α signalling in vascular cells.

Hypoxia-inducible factor 1α (HIF1α) is a master regulator of biological processes in hypoxia. Yet, the mechanisms and biological consequences of aerobic HIF1α activation by intrinsic factors, particularly in normal (primary) cells, remain elusive. Here we show that HIF1α signalling is activated in several human primary vascular cells in normoxia and in vascular smooth muscle cells of normal human lungs. Mechanistically, aerobic HIF1α activation is mediated by paracrine secretion of three branched-chain α-ketoacids (BCKAs), which suppress PHD2 activity via direct inhibition and via LDHA-mediated generation of L-2-hydroxyglutarate. BCKA-mediated HIF1α signalling activation stimulated glycolytic activity and governed a phenotypic switch of pulmonary artery smooth muscle cells, which correlated with BCKA metabolic dysregulation and pathophenotypic changes in pulmonary arterial hypertension patients and male rat models. We thus identify BCKAs as previously unrecognized signalling metabolites that aerobically activate HIF1α and that the BCKA-HIF1α pathway modulates vascular smooth muscle cell function, an effect that may be relevant to pulmonary vascular pathobiology.

Recruitment of CCR5-positive T cells in pulmonary vascular lesions in idiopathic- and connective tissue disease associated- pulmonary arterial hypertension

Abstract Background C-C chemokine receptor type 5 (CCR5) is a chemokine receptor predominantly expressed on leukocytes including T cells, macrophages, and dendritic cells, and is involved in the process by which T cells are attracted to specific tissues and organs. Although CCR5 has been shown to be expressed on macrophages and pulmonary artery smooth muscle cells in lung samples from human idiopathic pulmonary arterial hypertension (PAH) and some animal models, its expression on T cells in human PAH lungs has not been investigated. Purpose To investigate the immunohistochemical expression of CCR5 on T cells infiltrating in pulmonary vascular lesions of human PAH tissues, particularly from patients with idiopathic PAH and PAH associated with connective tissue disease (CTD-PAH), where inflammation has been implicated in the pathogenesis. Methods Subjects included 25 postmortem cases of idiopathic PAH (14 cases, a mean age of 34 ± 10 years, 8 female cases) and CTD-PAH (11 cases, 48 ± 13 years, all female). We performed immunohistochemistry using antibodies against CCR5 and CD3 (for T cells) on formalin-fixed paraffin-embedded sections. To semi-quantify, we selected the three small muscular pulmonary arteries (80-150 µm in external diameter) with the strongest CCR5-positive cell infiltration in each case, counted the number of positive cells, and the average were shown as the number of cells per vessel. Results In all cases, regardless of whether IPAH or CTD-PAH, a subset of infiltrating inflammatory cells into or accumulating around vessels was CCR5-positive. Specifically, CCR5-positive inflammatory cells were found in remodelled pulmonary arteries, such as plexiform lesions and constrictive lesions with intimal thickening and/or medial hypertrophy, as well as perivascular space. The average number of CCR5-positive cells per small muscular pulmonary artery was 8.1/vessel. Where pulmonary venous occlusive lesions were seen, CCR5-positive cells were also present in and around them. The majority of CCR5-positive inflammatory cells appeared to be T cells, based on morphology and CD3 immunopositivity in the serial sections. Conclusion(s) Activated T cells showing CCR5 immunopositivity were recruited to remodelled pulmonary vascular lesions in both idiopathic PAH and CTD-PAH. This implicates CCR5-positive T cells as a common key player and that CCR5 signalling is involved in the pathogenesis of PAH in concert with other inflammatory cells, vascular endothelial cells, and pulmonary artery smooth muscle cells.

Synthesis, Characterization, and Molecular Docking of Novel Isatin-thiosemicarbazone containing 1,2,3-triazole Derivatives as Potential Anti-cancer Agents

Abstract: To improve the activity of isatin-1,2,3-triazole hybrids as anticancer agents, new derivatives of isatin- thiosemicarbazone-1,2,3-triazoles 9-16 were designed and synthesized via the condensation of isatin-1,2,3- triazole hybrids 1-8 with thiosemicarbazide. Spectral and elemental analysis confirmed the structure of the prepared derivatives 9-16. Also, as anticancer agents, the latter derivatives were screened against three human cancerous cell lines: human lung fibroblast (WI38), colorectal carcinoma colon cancer (HCT-116), and mammary gland breast cancer (MCF-7). Doxorubicin, a standard control, was used to compare viable cell percentages and IC50 values. In general, derivatives 12 and 16 revealed a higher potency against the three human cancerous cell lines. Finally, the molecular descriptors of compounds 12 and 16 were correlated with their observed cytotoxicity. result: In general, derivatives 12 and 16 revealed a higher potency against the three human cancerous cell lines.

Proteomic approach to identify host cell attachment proteins provides protective Pseudomonas aeruginosa vaccine antigen FtsZ

Pseudomonas aeruginosa is an opportunistic Gram-negative pathogen that causes severe nosocomial infections in susceptible individuals due to the emergence of multidrug-resistant strains. There are no approved vaccines against P. aeruginosa infections nor candidates in active clinical development, highlighting the need for novel candidates and strategies. Using a cell-blot proteomic approach, we reproducibly identified 49 proteins involved in interactions with human lung epithelial cells across four P. aeruginosa strains. Among these were cell division protein FtsZ and outer membrane protein OpmH. Escherichia coli BL21 cells overexpressing recombinant FtsZ or rOpmH showed a 66- and 15-fold increased ability to attach to 16HBE14o− cells, further supporting their involvement in host cell attachment. Both antigens led to proliferation of NK and CD8+ cytotoxic T cells, significant increases in the production of IFN-γ, IL-17A, TNF and IL-4 in immunised mice and elicited strong antigen-specific serological IgG1 and IgG2c responses. Immunisation with FtsZ significantly reduced bacterial burden in the lungs by 1.9-log CFU and dissemination to spleen by 1.8-log CFU. The protective antigen candidate, FtsZ, would not have been identified by traditional approaches relying on either virulence mechanisms or sequence-based predictions, opening new avenues in the development of an anti-P. aeruginosa vaccine.

Unveiling the anticancer potential: Exploring 4-fluoro benzoic acid and piperazine through spectroscopic, X-ray diffraction, DFT and molecular docking analysis

The crystalline substance formed from piperazine and 4-fluorobenzoic acid (PZ-PFBA) was successfully synthesized. This synthesis involved the transfer of protons between the selected acid and base, resulting in the formation of distinct crystalline salt. The SCXRD analysis conforms the salt crystallized in a monoclinic crystal system with a P 21/c space group and lattice parameters are a = 13.0688(14) Å, b = 8.2686(8) Å, and c = 8.2552(9) Å. Hirshfeld surface analysis, a technique for quantifying molecular interactions, was employed to investigate the prepared crystalline salt. Additionally, the salt was subjected to further examination using FT-IR, 1H NMR, 13C NMR, and SCXRD studies. The crystalline salt has major and established biological actions that inhibit the human lung cancer cell line A549. In vitro, and Insilico anticancer tests show that PZ-PFBA has better efficacy against the human cervical cancer cell line (HeLa) and in bioinformatics analysis. PZ-PFBA demonstrated more efficacy in reducing pathogenic strains compared to its parent reactants.

New Quadratic Discriminant Analysis Algorithms for Correlated Audiometric Data.

Paired organs like eyes, ears, and lungs in humans exhibit similarities, and data from these organs often display remarkable correlations. Accounting for these correlations could enhance classification models used in predicting disease phenotypes. To our knowledge, there is limited, if any, literature addressing this topic, and existing methods do not exploit such correlations. For example, the conventional approach treats each ear as an independent observation when predicting audiometric phenotypes and is agnostic about the correlation of data from the two ears of the same person. This approach may lead to information loss and reduce the model performance. In response to this gap, particularly in the context of audiometric phenotype prediction, this paper proposes new quadratic discriminant analysis (QDA) algorithms that appropriately deal with the dependence between ears. We propose two-stage analysis strategies: (1) conducting data transformations to reduce data dimensionality before applying QDA; and (2) developing new QDA algorithms to partially utilize the dependence between phenotypes of two ears. We conducted simulation studies to compare different transformation methods and to assess the performance of different QDA algorithms. The empirical results suggested that the transformation may only be beneficial when the sample size is relatively small. Moreover, our proposed new QDA algorithms performed better than the conventional approach in both person-level and ear-level accuracy. As an illustration, we applied them to audiometric data from the Medical University of South Carolina Longitudinal Cohort Study of Age-related Hearing Loss. In addition, we developed an R package, PairQDA, to implement the proposed algorithms.

Evaluation of antioxidant, anti-inflammatory and cytotoxic potential of phytochemicals from Gymnostachyum febrifugum-an in vitro study

Gymnostachyum febrifugum an ethnomedicinal plant of Acanthaceae family is used to treat fever, ulcer and metrorrhagia. The current study aimed to determine the antioxidant, anti-inflammatory and cytotoxic potential of phytochemicals from methanolic leaf and root extracts of G. febrifugum. In addition, a preliminary phytochemical analysis was carried out using thin layer chromatography (TLC) and liquid chromatography-mass spectrometry (LC-MS). The study revealed significant antioxidant and anti-inflammatory activities of methanolic extracts of G. febrifugum roots compared to that of leaves. Also, a moderate in vitro cytotoxic effect of the root methanolic extracts was observed against human lung cancer cells. LC-MS analysis of the methanolic extract of root revealed the presence of 11 phytochemicals belonging to classes of alkaloids, flavonoids, terpenoids and sugars, of which esculin, 6,7,10-trihydroxy-8-octadecenoic acid, piperlongumine, ardisiacrispin B and berbamine were detected in high abundance. Probably, these phytochemicals were responsible for the antioxidant, anti-inflammatory and cytotoxic potential of G. febrifugum. However further research on isolation and identification of active compounds is necessary to confirm these results.

Antioxidant and Anti-Proliferative Effects of Ornithogalum balansae Extracts Collected in Black Sea Region Against Lung Cancer Cells

Ornithogalum is a genus of wild herb species widely used as food and in traditional medicine. This study investigated some of the biologically active properties of Ornithogalum balansae grown in the eastern Black Sea region of Türkiye. The investigations were carried out using the methanolic extracts of the plant’s aerial and bulbs (B1 and B2) parts. The phenolic composition was examined as total phenolic content (TPC), total flavonoid content (TFC), and HPLC-PDA. The antioxidant capacity of the extracts was tested based on ferric reducing/antioxidant power (FRAP) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging methods. The anti-proliferative activity was tested against metastatic cell lines H1299 and H209, non-small-cell lung A549, and fibroblast MRC-5 cell lines using MTT and trypan blue methods. Wound-healing and invasion chamber assays were used to determine the inhibitory effects of the extracts on migration and invasion, respectively. The extract of the aerial part contained a large number of phenolic substances and high antioxidant capacity. The extract exhibited a significant anti-proliferative effect on the human lung cancer cells (A549 and H209), with IC50 values of 0.97 ± 0.04 and 1.06 ± 0.07 µg/mL, respectively. Moreover, the aerial part exhibited inhibition of migratory and invasive capacities in A549 cells at a concentration of 1.50 µg/mL. The findings associated with O. balansae suggest a promising therapeutic potential against lung cancer cells.

Dual Inhibition of Phosphodiesterase 3 and 4 Enzymes by Ensifentrine Protects against MRSA-Induced Lung Endothelial and Epithelial Dysfunction.

Acute Respiratory Distress Syndrome (ARDS) is a severe lung condition with a high mortality rate for which there are no effective therapeutics. The failure of the alveolar-capillary barrier, composed of lung endothelial (EC) and alveolar epithelial (AEC) cells, is a critical factor leading to excessive inflammation and edema characteristic of acute lung injury (ALI) pathophysiology. Phosphodiesterases (PDE) are enzymes well-recognized for their roles in regulating endothelial permeability and inflammation. Although PDE inhibitors are used as therapeutics for inflammatory diseases like COPD (chronic obstructive pulmonary disease), their efficacy in treating ARDS has not yet been established. In this study, we investigated the effects of ensifentrine, an FDA-approved novel dual PDE 3/4 inhibitor, on lung endothelial and epithelial dysfunction caused by methicillin-resistant S. aureus (MRSA), a pathogen involved in bacterial ARDS. Human primary lung endothelial cells and alveolar epithelial cell lines (A549 and immortalized AEC) were treated with heat-killed MRSA, and their responses were assessed in the presence or absence of ensifentrine. Ensifentrine given either pre- or post-exposure attenuated MRSA-induced increased lung endothelial permeability. VE-cadherin junctions, which serve to stabilize the EC barrier, were disrupted by MRSA; however, ensifentrine effectively prevented this disruption. Pre-treatment with ensifentrine protected against MRSA-induced EC pro-inflammatory signaling by inhibiting the expression of VCAM-1, ICAM-1, and by reducing the IL-6 and IL-8 release. In AEC, MRSA caused the upregulation of ICAM-1, the activation of NF-kB, and the production of IL-8, all of which were inhibited by ensifentrine. These results indicate that the dual inhibition of phosphodiesterases 3 and 4 by ensifentrine is barrier protective and attenuates MRSA-induced inflammation in both lung endothelial and epithelial cells. The PDE3/4 inhibitor ensifentrine may represent a promising novel strategy for the treatment of MRSA-induced ARDS.

A splicing isoform of PD-1 promotes tumor progression as a potential immune checkpoint

The immune checkpoint receptor, programmed cell death 1 (PD-1, encoded by PDCD1), mediates the immune escape of cancer, but whether PD-1 splicing isoforms contribute to this process is still unclear. Here, we identify an alternative splicing isoform of human PD-1, which carries a 28-base pairs extension retained from 5′ region of intron 2 (PD-1^28), is expressed in peripheral T cells and tumor infiltrating lymphocytes. PD-1^28 expression is induced on T cells upon activation and is regulated by an RNA binding protein, TAF15. Functionally, PD-1^28 inhibits T cell proliferation, cytokine production, and tumor cell killing in vitro. In vivo, T cell-specific exogenous expression of PD-1^28 promotes tumor growth in both a syngeneic mouse tumor model and humanized NOG mice inoculated with human lung cancer cells. Our study thus demonstrates that PD-1^28 functions as an immune checkpoint, and may contribute to resistance to immune checkpoint blockade therapy.

Ultrastructural analysis of lamellar bodies in type II alveolar epithelial cells in the human lung.

Pulmonary surfactant is produced by type II alveolar epithelial cells (AEC2) and stored in lamellar bodies (LBs) prior to secretion. Here, we characterize AEC2 and their LBs in the human lung ultrastructurally and quantitatively. Five human lungs were analyzed by transmission electron microscopy, serial section electron tomography and stereology. A human lung contained about 24 billion AEC2 with a mean size of about 650 µm³. The number of AEC2 as well as the total volume of LBs per lung, about 1.9 mL, strongly correlated with total lung volume. A single AEC2 contained an LB volume of about 74 µm³. This amount was packed in about 324 LBs with a mean size of 0.24 µm³. Three morphologically distinct subpopulations of LBs were identified: 1.) isolated LBs which make up the majority (average 300 per AEC2), 2.) LBs connected to each other via pores (average 23 per AEC2), and 3.) LBs connected to the plasma membrane via a fusion pore (average 1 per AEC2). Along this sequence of subpopulations, the mean size of LBs increased. LBs that are connected either with each other or to the plasma membrane contained about 14% of an AEC2´s LB volume. This is in line with the concept of an intermediate surfactant pool, stored in LBs either directly or indirectly connected to the plasma membrane. In summary, this study provides quantitative reference data on surfactant-storing LBs in AEC2 as well as morphological evidence for an intermediate surfactant pool in the human lung.

Synthesis of Sulfonamide-Based Schiff Bases as Potent Anticancer Agents: Spectral Analyses, Biological Activity, Molecular Docking, ADME, DFT, and Pharmacophore Modelling Studies.

The current study focuses on the synthesis and characterization of six benzenesulfonamide-based Schiff base derivatives (7-12) with various electron-withdrawing and electron-donating substituents (-F, -CI, -Br, -CH3, and -OCH3) and the assessment of their antiproliferative activities against human lung (A549) and liver (HepG2) cancer cell lines using in vitro and in silico approaches. The structures of the synthesized compounds (7-12) were elucidated by elemental analysis and FT-IR, 1D (1H, 13C, APT, and DEPT-135), and 2D (HMQC and HMBC) NMR spectroscopies. The cytotoxic activities of the targeted compounds were determined at various concentrations against these cancer cell lines for 72 h, using the MTT method. The targeted molecules (7-12) demonstrated remarkable antiproliferative activities, with IC50 values ranging from 6.032-9.533 μM against the A549 cell line and 5.244-9.629 μM against the HepG2 cell line. These compounds showed activities at lower or very similar concentrations to cisplatin against the A549 cell line and at much lower concentrations than cisplatin against the HepG2 cell line. Among them, compounds 10 and 12 were found to be more effective against A549 and HepG2 cells, respectively, than cisplatin. These compounds were analyzed by interacting with the 1BNA, 4HJO, and 4ASD crystal structures in molecular docking studies.

Cytotoxic activity, selectivity, and clonogenicity of fruits and resins of Saudi medicinal plants against human liver adenocarcinoma.

Edible fruits and resins provide various benefits to mankind including potential medicinal applications. This study aimed to determine the cytotoxicity, selectivity, and clonogenicity of fruits and exudates of certain Saudi medicinal plants (Anethum graveolens (BEP-09), Opuntia ficus-indica (L.) Miller (BEP-10), Boswellia serrata Roxb. ex Colebr. (BEP-11), and Commiphora myrrha (BEP-12)) against human liver adenocarcinoma (HepG2). Initial cytotoxicity and cell line selectivity against different cell lines were screened using MTT assay. The most promising extract was subjected to gas chromatography-mass spectrometry (GC-MS) analysis to determine the main phytoconstituents. Clonogenicity was checked for the most active extract. The selected plants' fruits and resins possess a significant cytotoxic activity estimated as IC50. The fruit of BEP-10 was found to be the most active extract against liver cancer cells (IC50 = 2.82) comparable to both doxorubicin (IC50 = 1.40) and camptothecin (IC50 = 1.11). It showed a selectivity index of 4.47 compared to the normal human foetal lung fibroblast (MRC5) cells. BEP-10 showed a dose-dependent clonogenic effect against HepG2 cells comparable to the effect of doxorubicin. The GC-MS chromatogram of BEP-10 extract revealed the presence of eight small polar molecules, representing 73% of the total identified compounds and the rest three molecules (27%) were non-polar constituents. The furan derivatives represent the chief components in BEP-10 (16.3%), while the aldehyde 5-(hydroxymethyl)-2-furancarboxaldehyde was found to be the main molecule (13.2%). The fruits of BEP-10 have a potential cytotoxic effect particularly against HepG2. The identified phytoconstituents in the tested plant extract might contribute to the investigated cytotoxic activity.

Induction of Ca2+ signaling and cytotoxic responses of human lung fibroblasts upon an antihistamine drug oxatomide treatment and evaluating the protective effects of Ca2+ chelating.

Oxatomide, an antihistamine drug of the diphenylmethylpiperazine family, has anti-inflammatory effects in airway disease. Because oxatomide was shown to cause diverse physiological responses in several cell models, the impact of oxatomide on Ca2+ signaling and its related physiological effects has not been explored in IMR-90 human fetal lung fibroblasts. This study assessed the effect of oxatomide on cell viability and intracellular free Ca2+ concentrations ([Ca2+]i) and examined whether oxatomide-induced cytotoxicity through Ca2+ signaling in IMR-90 cells. Cell viability was measured by the cell proliferation reagent (WST-1). [Ca2+]i was measured by the Ca2+-sensitive fluorescent dye fura-2. Oxatomide (10-40 μM) concentration dependently reduced cell viability and induced [Ca2+]i rises in IMR-90 cells. This cytotoxic effect was reversed by chelation of cytosolic Ca2+ with BAPTA-AM. In terms of Ca2+ signaling, oxatomide-caused Ca2+ entry was inhibited by modulators of store-operated Ca2+ channels (2-APB and SKF96365) and protein kinase C (PKC) inhibitor (GF109203X). Furthermore, oxatomide-induced Ca2+ influx was confirmed by Mn2+-induced quench of fura-2 fluorescence. In a Ca2+-free medium, preincubation with the endoplasmic reticulum Ca2+ pump inhibitor thapsigargin inhibited oxatomide-evoked [Ca2+]i rises. Conversely, treatment with oxatomide abolished thapsigargin-induced [Ca2+]i rises. Inhibition of phospholipase C (PLC) with U73122 also inhibited oxatomide-caused [Ca2+]i rises. In IMR-90 cells, oxatomide-induced cytotoxicity by preceding [Ca2+]i rises involving PKC-sensitive store-operated Ca2+ entry and PLC-dependent Ca2+ release from the endoplasmic reticulum. BAPTA-AM, with its Ca2+ chelating effects, may be a potential compound for preventing oxatomide-induced cytotoxicity.

Nerolidol inhibits proliferation and triggers ROS-facilitated apoptosis in lung carcinoma cells via the suppression of MAPK/STAT3/NF-κB and P13K/AKT pathways.

Lung cancer (LC) is the leading cause of malignancy-related mortalities globally, and the existing treatment interventions are associated with harmful side effects. In the current study, we evaluated the anti-tumor efficiency of nerolidol (NRD) on human non-small cell lung cancer (NSCLC) cells. Nerolidol is a sesquiterpene alcohol extracted from the essential oils of aromatic flora with known anti-cancer activities. The latent action of NRD on antiproliferative and apoptotic effects in A549 cells is uncertain. Thus, our work is designed to explore the antiproliferative and apoptotic actions of NRD (20 and 25 μM/mL) against A549 cells. The activity of NRD on A549 cell cytotoxicity, intracellular reactive oxygen species (ROS), mitochondrial membrane potential (MMP), apoptosis, anti-apoptotic proteins, and MAPK/TAT3/NF-κB and P13K/AKT signaling pathways were assessed using MTT tests, dichlorodihydrofluorescein diacetate (DCFH-DA), dual acridine orange/ethidium bromide (AO/EB), DAPI, Rh-123, reverse transciption polymerase chain reaction (RT-PCR), and western blot analyses. We found that NRD could inhibit NSCLC cell viability through elevated intracellular ROS and MMP loss and elicited apoptosis in a quantity-dependent manner. Similarly, NRD can reduce inflammatory cytokines and anti-apoptotic elements, as well as trigger apoptotic signaling pathways. Our data established that NRD decreases A549 cell proliferation through ROS-mediated apoptosis, triggering the MAPK/STAT3/NF-κB and P13K/AKT pathways, suggesting that NRD is a possible protective remedy for NSCLC.

Bee venom prompts the inhibition of gefitinib on proliferation, migration, and invasion of non-small cell lung cancer cells via EGFR-mediated autophagy

It has been confirmed that bee venom (BV) can inhibit tumor metastasis of lung cancer cells induced by epidermal growth factor, suggesting the inhibitory role of BV on the regulation of epidermal growth factor receptor (EGFR), and may synergistically promote the anti-lung cancer effect of EGFR tyrosine kinase inhibitor gefitinib. This paper aims to ascertain the therapeutic potentials of BV combined with gefitinib against non-small cell lung cancer (NSCLC) in vitro. As results, the content of the main component melittin in air-dried BV was determined by HPLC. Subsequently, it was found that BV significantly inhibited the proliferation of NSCLC PC-9 and NCI-H1299 cells, but not generated apparent toxicity to human normal lung epithelial BEAS-2B cells. Meanwhile, the combination of BV and gefitinib also significantly inhibited the proliferation of these two cells, and suppressed the migration and invasion of PC-9 cells. By bioinformatics analysis and molecular docking, it was predicted that the main component melittin in BV could act on the cell membrane and transmembrane protein EGFR. Ultimately, Western blot assays showed BV alone or combined with gefitinib significantly decreased the protein expression of phosphorylated EGFR (p-EGFR) and the protein expression ratio of p-EGFR to EGFR, and increased the protein expression ratio of LC3-II to LC3-I in PC-9 cells or epidermal growth factor-activated PC-9 cells. The results demonstrated that BV could prompt the inhibition of gefitinib on proliferation, migration, and invasion of NSCLC cells via EGFR-mediated autophagy, showing the synergistic anti-NSCLC potential when combined with gefitinib.

Assessment of toxicity changes induced by exposure of human cells to lunar dust simulant

The toxicity of lunar dust (LD) to astronauts' health has been confirmed in the Apollo missions and subsequent biological experiments. Therefore, it is crucial to understand the biological toxicity of lunar dust for future human missions to the Moon. In this study, we exposed human lung epithelial cells (BEAS-2B) and peripheral blood B lymphocytes (AHH-1) to varying concentrations (0, 500, 1000, and 1500 μg/ml) of a lunar dust simulant (LDS) called CLDS-i for 24 and 48 h. The results provided the following key findings: (1) LDS induction of cell damage occurred through oxidative stress, with the levels of reactive oxygen species (ROS) in BEAS-2B cells being dependent on the duration of exposure. (2) Necrosis and early apoptosis were observed in BEAS-2B cells and AHH-1 cells, respectively. In addition, both cells showed lysosomal damage. (3) Genes CXCL1, SPP1, CSF2, MMP1, and POSTN are implicated in immune response and cytoskeletal arrangement regulation in BEAS-2B cells. Considering the similarities in composition and properties between CLDS-i and real lunar dust, our findings not only enhance the understanding of LDS toxicity, but also contribute to a better comprehension of the genomic alterations and molecular mechanisms underlying cellular toxicity induced by LD. These insights will contribute to the development of a biotoxicology framework aimed at safeguarding the health of astronauts and, consequently, facilitating future human missions to the Moon.