Human Lung A549 Cells Research Articles

Metabolomics reveals that phosphatidylethanolamine can alleviate the toxicity of silica nanoparticles in human lung A549 cells.

Silica nanoparticles (SiNPs) are widely utilized in occupational settings where they can cause lung damage through inhalation. The objective of this research was to explore the metabolic markers of SiNPs-induced toxicity on A549 cells by metabolomics and provide a foundation for studying nanoparticle-induced lung toxicity. Metabolomics analysis was employed to analyze the metabolites of SiNPs-treated A549 cells. LASSO regression was applied for selection, and protective measure experiments were conducted to validate the efficacy of selected potential toxicity mitigators. After SiNPs treatment, 23 differential metabolites were identified, including lipids, nucleotides, and organic oxidants. Pathway analysis revealed involvement in various biological processes. LASSO regression further identified six metabolites significantly associated with SiNPs toxicity. Notably, phosphatidylethanolamine (PE (14:1(9Z)/14:0)) showed enrichment in six significant metabolic pathways and with an AUC of 1 in the ROC curve. Protective measure experiments verified its protective effect on A549 cells and demonstrated its considerable inhibition of SiNPs-induced cytotoxicity. This study elucidated SiNPs-induced cytotoxicity on A549 cells and identified PE as a potential toxicity mitigator. These findings contribute to understanding the mechanisms of nanoparticle-induced lung toxicity and inform occupational health preventive strategies.

Phytochemical composition of wild pomegranate juices and their cytotoxicity

PurposeThis study aims to examine the relationship between the chemical composition of juices obtained from fruits of autochthonous wild pomegranate (Punica granatum L.) grown in Montenegro and their cytotoxic effects on cancer cells.Design/methodology/approachTo explore the potential value of wild pomegranate fruits, in vitro biological assays were carried out with juices whose composition was analyzed in detail for sugars, organic acids, vitamin C and phenolic compounds. The effect of juices on survival was determined in human lung A549, cervical HeLa and breast MCF-7 carcinoma cells by MTT assay. As a control, the cytotoxicity against normal fetal lung fibroblasts (MRC-5) was monitored.FindingsAmong cancer cell lines, considering the IC50 related to total phenolics, the lowest value – 13 µg/mL was found for the A549. The strongest effect on lung cells was assumed due to the favorable contribution of ellagitannins to total phenolics in juice as well as the given combination of anthocyanins and their synergistic action. For HeLa cells, the lowest IC50 value was obtained at 88 µg/mL, and the cytotoxicity could be matched with the effects of anthocyanins and catechin. For MCF-7 cells, the lowest IC50 was 504 µg/mL, and the elevated levels of vitamin C and ellagic acid derivatives should have a noticeable effect on these cells.Originality/valueThis study provides an important contribution to the knowledge on the effect of phytochemicals from wild pomegranate juice on lung, cervical and breast cancer cells, in vitro. The present observations suggest that the juice of wild pomegranate has the potential in the fight against cancer.

A transcriptomic biomarker predictive of cell proliferation for use in adverse outcome pathway-informed testing and assessment.

High-throughput transcriptomics (HTTr) is increasingly being used to identify molecular targets of chemicals that can be linked to adverse outcomes. Cell proliferation (CP) is an important key event in chemical carcinogenesis. Here, we describe the construction and characterization of a gene expression biomarker that is predictive of the CP status in human and rodent tissues. The biomarker was constructed from 30 genes known to be increased in expression in prostate cancers relative to surrounding tissues and in cycling human MCF-7 cells after estrogen receptor (ER) agonist exposure. Using a large compendium of gene expression profiles to test utility, the biomarker could identify increases in CP in (i) 308 out of 367 tumor vs. normal surrounding tissue comparisons from 6 human organs, (ii) MCF-7 cells after activation of ER, (iii) after partial hepatectomy in mice and rats, and (iv) the livers of mice and rats after exposure to nongenotoxic hepatocarcinogens. The biomarker identified suppression of CP (i) under conditions of p53 activation by DNA damaging agents in human cells, (ii) in human A549 lung cells exposed to therapeutic anticancer kinase inhibitors (dasatinib, nilotnib), and (iii) in the mouse liver when comparing high levels of CP at birth to the low background levels in the adult. The responses using the biomarker were similar to those observed using conventional markers of CP including PCNA, Ki67, and BrdU labeling. The CP biomarker will be a useful tool for interpretation of HTTr data streams to identify CP status after exposure to chemicals in human cells or in rodent tissues.

Nanofibers loaded levosalbutamol sulfate fast dissolving oral films for sublingual delivery

During allergic reactions, a drug should work quickly and effectively. Levosalbutamol (LSAL) is a short-acting drug that stimulates β2 adrenergic receptors and is commonly used to treat asthma and chronic obstructive pulmonary disease (COPD). This study aimed to prepare sublingual oral patches of levosalbutamol sulfate electrospun nanofibers for ultrafast release. In this study, we fabricated nanofibers by electrospinning polyvinyl alcohol (PVA) polymer (10 %) and different concentrations (3, 5, and 10 %) of LSAL. The nanofibers are randomly aligned, with smooth surfaces and an average fiber diameter of 160, 170, and 198 nm at PVA/LSAL concentrations of 3 %, 5 %, and 10 %, respectively. The in vitro dissolution study showed that all PVA/LSAL nanofibers released the drug ultra-fast (within 5–10 min); 90 % released within 3 min. The folding endurance study revealed that the drug-loaded nanofibers had improved stability. A disintegration study showed that the nanofibers disintegrated rapidly in 1–3 min, and an in vitro cytocompatibility study showed good cell viability with A549 human lung cells. Hence, the proposed fast-dissolving sublingual oral films/patches approach is a promising alternative to existing dosage forms. They can provide an ultra-fast release of various drugs requiring a fast onset of action with increased bioavailability and patient compliance.

Air-liquid interface exposure of A549 human lung cells to characterize the hazard potential of a gaseous bio-hybrid fuel blend.

Gaseous and semi-volatile organic compounds emitted by the transport sector contribute to air pollution and have adverse effects on human health. To reduce harmful effects to the environment as well as to humans, renewable and sustainable bio-hybrid fuels are explored and investigated in the cluster of excellence "The Fuel Science Center" at RWTH Aachen University. However, data on the effects of bio-hybrid fuels on human health is scarce, leaving a data gap regarding their hazard potential. To help close this data gap, this study investigates potential toxic effects of a Ketone-Ester-Alcohol-Alkane (KEAA) fuel blend on A549 human lung cells. Experiments were performed using a commercially available air-liquid interface exposure system which was optimized beforehand. Then, cells were exposed at the air-liquid interface to 50-2000 ppm C3.7 of gaseous KEAA for 1 h. After a 24 h recovery period in the incubator, cells treated with 500 ppm C3.7 KEAA showed significant lower metabolic activity and cells treated with 50, 250, 500 and 1000 ppm C3.7 KEAA showed significant higher cytotoxicity compared to controls. Our data support the international occupational exposure limits of the single KEAA constituents. This finding applies only to the exposure scenario tested in this study and is difficult to extrapolate to the complex in vivo situation.

Salvianolic acid B inhibits the growth and metastasis of A549 lung cancer cells through the NDRG2/PTEN pathway by inducing oxidative stress.

Salvianolic acid B (Sal B) has demonstrated anticancer activity against various types of cancer. However, the underlying mechanism of Sal B-mediated anticancer effects remains incompletely understood. This study aims to investigate the impact of Sal B on the growth and metastasis of human A549 lung cells, as well as elucidate its potential mechanisms. In this study, different concentrations of Sal B were administered to A549 cells. The effects on migration and invasion abilities were assessed using MTT, wound healing, and transwell assays. Flow cytometry analysis was employed to evaluate Sal B-induced apoptosis in A549 cells. Western blotting and immunohistochemistry were conducted to measure the expression levels of cleaved caspase-3, cleaved PARP, and E-cadherin. Commercial kits were utilized for detecting intracellular reactive oxygen species (ROS) and NAD+. Additionally, a xenograft model with transplanted A549 tumors was employed to assess the anti-tumor effect of Sal B in vivo. The expression levels of NDRG2, p-PTEN, and p-AKT were determined through western blotting. Our findings demonstrate that Sal B effectively inhibits proliferation, migration, and invasion in A549 cells while inducing dose-dependent apoptosis. These apoptotic responses and inhibition of tumor cell metastasis are accompanied by alterations in intracellular ROS levels and NAD+/NADH ratio. Furthermore, our in vivo experiment reveals that Sal B significantly suppresses A549 tumor growth compared to an untreated control group while promoting increased cleavage of caspase-3 and PARP. Importantly, we observe that Sal B upregulates NDRG2 expression while downregulating p-PTEN and p-AKT expressions. Collectively, our results provide compelling evidence supporting the ability of Sal B to inhibit both growth and metastasis in A549 lung cancer cells through oxidative stress modulation as well as involvement of the NDRG2/PTEN/AKT pathway.

Microencapsulation of Origanum heracleoticum L. and Thymus vulgaris L. essential oils – Novel strategy to combat multi-resistant Acinetobacter baumannii

Within the global rise of antimicrobial resistance enhanced by the COVID-19 pandemic, where Acinetobacter baumannii has been distinguished as an emerging multi-resistant pathogen, essential oils, become the focus of novel therapeutic approaches. Hypothesizing that encapsulated Origanum heracleoticum L. and Thymus vulgaris L. essential oils could express multi-target approach against A. baumannii, this study aimed to develop microencapsulated systems with optimal technological qualities using 2-hydroxypropyl-β-cyclodextrin as a carrier, and to evaluate their pharmacological potential against A. baumannii, and their morphological and physicochemical characteristics, safety and stability profiles. The highest yield and encapsulation efficiency were obtained with 1:10 essential oil to carrier, and 1.5:10 carrier to water w/w ratios. The formation of inclusion complexes was confirmed by Fourier Transform Infrared Spectroscopy. Both microencapsulates achieved improved homogeneity, particle surface, and thermal stability compared with the pure carrier. Dominant bioactive compounds (carvacrol and p-cymene in O. heracleoticum essential oil, and thymol and p-cymene in T. vulgaris essential oil) remained the most abundant after encapsulation. While non-encapsulated essential oils revealed similar antimicrobial activity towards clinical A. baumannii isolates obtained from COVID-19 patients, encapsulated O. heracleoticum essential oil inhibited the bacterial growth at lower concentrations than T. vulgaris essential oil microencapsulate. All samples significantly reduced the formation of A. baumannii biofilm, for at least 53.90% towards the most infective isolate according to the Caenorhabditis elegans assay. Further, in silico molecular docking study revealed strong interaction pattern of carvacrol and thymol with the outer membrane protein A, which is the main factor for the A. baumannii biofilm formation. Cytotoxicity investigation on human lung A549 cells showed high survival rate in the presence of all tested concentrations, and the stability study revealed notable preservation of the bioactives’ content and pharmacological potential. Altogether, microencapsulated essential oils exhibited a multi-target approach towards A. baumannii, with satisfactory preserving capability during storage.

Sinomenine attenuates pulmonary fibrosis by downregulating TGF-β1/Smad3, PI3K/Akt and NF-κB signaling pathways

BackgroundSince COVID-19 became a global epidemic disease in 2019, pulmonary fibrosis (PF) has become more prevalent among persons with severe infections, with IPF being the most prevalent form. In traditional Chinese medicine, various disorders are treated using Sinomenine (SIN). The SIN’s strategy for PF defense is unclear.MethodsBleomycin (BLM) was used to induce PF, after which inflammatory factors, lung histological alterations, and the TGF-/Smad signaling pathway were assessed. By administering various dosages of SIN and the TGF- receptor inhibitor SB-431,542 to human embryonic lung fibroblasts (HFL-1) and A549 cells, we were able to examine proliferation and migration as well as the signaling molecules implicated in Epithelial-Mesenchymal Transition (EMT) and Extra-Cellular Matrix (ECM).ResultsIn vivo, SIN reduced the pathological changes in the lung tissue induced by BLM, reduced the abnormal expression of inflammatory cytokines, and improved the weight and survival rate of mice. In vitro, SIN inhibited the migration and proliferation by inhibiting TGF-β1/Smad3, PI3K/Akt, and NF-κB pathways, prevented the myofibroblasts (FMT) of HFL-1, reversed the EMT of A549 cells, restored the balance of matrix metalloenzymes, and reduced the expression of ECM proteins.ConclusionSIN attenuated PF by down-regulating TGF-β/Smad3, PI3K/Akt, and NF-κB signaling pathways, being a potential effective drug in the treatment of PF.

The Effect of 1'-acetoxychavicol Acetate on A549 Human Non-small Cell Lung Cancer

Lung cancer, a global leading cause of cancer mortality, requires safer and more effective treatments. 1'-acetoxychavicol acetate (ACA) is notable for its wide-ranging therapeutic properties, spanning anticancer, anti-inflammatory, and anti-obesity effects. While its efficacy has been explored in various cell lines, its potential against lung cancer remains few reported. This research aimed to investigate the impact of ACA on cytotoxicity, anti-proliferation, and antimigration potential in A549 human non-small cell lung cell lines. The investigation utilized MTT assays to assess cell viability and determined the IC50 values of ACA at 24, 48, and 72 hours to be 50.42 µM, 33.22 µM, and 21.66 µM, respectively. These results reveal a concentration- and time-dependent reduction in cell viability following ACA treatment. In addition, anti-proliferation was performed using colony-forming assays, and ACA showed notable efficacy in significantly decreasing colony formation, highlighting its strong impact on cell proliferation and viability. In the final part, ACA had promising inhibitory effects on migration, which varied depending on the dose used in the scratch assay. In conclusion, ACA highlighted cytotoxic and anti-proliferation properties of A549 cell line, supporting its potential role in lung cancer therapy pending further investigation and development.

Antibacterial activity, cytotoxicity and chemotaxonomic significance of chemical constituents from Alsophila manniana Hook R.M.Tryon (Cyatheaceae) rhizomes

Phytochemical investigation of Alsophila manniana rhizomes led to the isolation of twelve known compounds, including two triterpenes (1 and 2), four steroids (3–6), one glycerol (7), three phenols (8–10), and two flavonoids (11 and 12). Their structures were elucidated by analyses of the NMR and MS data and by comparisons with those in the literature. Their antibacterial and cytotoxic activities were evaluated against Escherichia coli, Klebsiella pneumoniae, Staphylococcus aureus, and Bacillus subtilis; and on human lung A549, cervical HeLa, and breast MCF-7 cancer cell lines, respectively. Fractions FD and FF had significant activities against E. coli (MIC = 75 μg/mL) and K. pneumoniae (MICs = 75 and 37.5 μg/mL). Compounds 3 and 7–12 had moderate antibacterial activities with MIC values ranging from 75 to 150 μg/mL against the bacteria. All samples were inactive against all tested human cancer cell lines, even at 100 μg/mL. This is the first investigation of the chemical constituents of A. manniana, as well as the first report of compounds 3, 7, and 10–12 from the genus Alsophila. The chemotaxonomic significance of these compounds is discussed.

Genotoxicand biomarker response of mussel gill cells, haemocyte and A549 human lung cells to radiation from X-rays

A comparative study on the genotoxic effect of radiation from X-rays of different levels on Mussel gill cells, haemocyte and human lung cell A549 using the comet assay was carried out. The results revealed an increasing DNA damage with increasing doses of X-ray, single strand breaks and alkali labile sites in the haemocyte of the control had a mean value of 5.92 ± 1.27 while the highest treatment group 15Gray (Gy) had a mean value of 78.65 ± 4.36,withsignificant differences (P <0.05), A mean value of 21.10 ± 13.10 was recorded for Mussel gill cells in the control group while the highest treatment group 15 Gray (Gy) had 85.55 ± 1.43. Statistical analysis revealed significant differences (P <0.05) for univariate tests of the control group and the treatment groups. The A549 human lung cells had a minimal damage in the control group 2.30 ± 0.30 while the highest exposed group had a mean value from the median % tail intensity of 43.10 ± 2.81. Pair wise comparison of gills cells, haemocytes from mussels and A549 human lung cells exposed to 15 Gy of X-rays shows that the highest damaging effect was observed on the gill cells followed by the haemocytes with the least damaging effect on the A549 human lung cells exposed to the same dose rate of X-rays with significant differences (P <0.05) between them.This study revealed that A549 human lung cells were more resistant to DNA damage induced by the X-rays when compared to mussel gill and haemocyte cells and also indicates that mussel gill cells and haemocytes can serve as biomarkers of genotoxicity of radiation exposure using the comet assay.

Fragment-Based Design, Synthesis, and Characterization of Aminoisoindole-Derived Furin Inhibitors.

A 1H-isoindol-3-amine was identified as suitable P1 group for the proprotein convertase furin using a crystallographic screening with a set of 20 fragments known to occupy the S1 pocket of trypsin-like serine proteases. Its binding mode is very similar to that observed for the P1 group of benzamidine-derived peptidic furin inhibitors suggesting an aminomethyl substitution of this fragment to obtain a couplable P1 residue for the synthesis of substrate-analogue furin inhibitors. The obtained inhibitors possess a slightly improved picomolar inhibitory potency compared to their benzamidine-derived analogues. The crystal structures of two inhibitors in complex with furin revealed that the new P1 group is perfectly suited for incorporation in peptidic furin inhibitors. Selected inhibitors were tested for antiviral activity against respiratory syncytial virus (RSV) and a furin-dependent influenza A virus (SC35M/H7N7) in A549 human lung cells and demonstrated an efficient inhibition of virus activation and replication at low micromolar or even submicromolar concentrations. First results suggest that the Mas-related G-protein coupled receptor GPCR-X2 could be a potential off-target for certain benzamidine-derived furin inhibitors.

Preparation of water-based dextran-coated Fe3O4 magnetic fluid for magnetic hyperthermia

Abstract Fe3O4 nanoparticles were prepared by chemical co-precipitation, modified with dextran, and dispersed in water to form a magnetic fluid (MF) for use in biomedical areas. The analyses of stability and magnetic property demonstrate that the prepared functional MF possesses outstanding stability (stability index within 60 days, high dilution stability, and autoclaved stability) and high magnetization (the values χ and M s of MF are 5.87 × 10−4 and 20.57 emu/g, respectively). Due to the coating of dextran, the toxicity of MF is minimal (in vitro survival rate of MCF-7 cells, blood compatibility, and in vivo toxicity). In addition, although the outer layer is coated with dextran, the M s intensity remains high, so the Fe3O4 MF owns a fast magnetic temperature response (when the MF concentration is 55 mg/mL, it can rapidly rise to 55°C within 800 s), which plays an extremely vital role in MF hyperthermia. So, the MF can effectively cause necrosis of human lung A549 cells, which shows a certain application potential.

Investigating PM2.5 toxicity in highly polluted urban and industrial areas in the Middle East: human health risk assessment and spatial distribution

Exposure to particulate matter (PM) can be considered as a factor affecting human health. The aim of this study was to investigate the concentration of PM2.5 and heavy metals and their influence on survival of A549 human lung cells in exposure to PM2.5 breathing air of Ahvaz city. In order to assess the levels of PM2.5 and heavy metals, air samples were collected from 14 sampling stations positioned across Ahvaz city during both winter and summer seasons. The concentration of heavy metals was determined using ICP OES. Next, the MTT assay [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] was employed to ascertain the survival rate of A549 cells. The findings from this research demonstrated that average PM2.5 of the study period was (149.5 μg/m3). Also, the average concentration of PM2.5 in the urban area in winter and summer was (153.3- and 106.9 μg/m3) and in the industrial area this parameter was (191.6 and 158.3 μg/m3). The average concentration of metals (ng/m3) of urban areas against industrial, Al (493 vs. 485), Fe (536 vs. 612), Cu (198 vs. 212), Ni (128 vs. 129), Cr (48.5 vs. 54), Cd (118 vs. 124), Mn (120 vs. 119), As (51 vs. 67), Hg (37 vs. 50), Zn (302 vs. 332) and Pb (266 vs. 351) were obtained. The results of the MTT assay showed that the highest percentage of cell survival according to the exposure concentration was 25 > 50 > 100 > 200. Also, the lowest percentage of survival (58.8%) was observed in the winter season and in industrial areas with a concentration of 200 μg/ml. The carcinogenic risk assessment of heavy metals indicated that except for Cr, whose carcinogenicity was 1.32E−03, other metals were in the safe range (10–4–10–6) for human health. The high concentration of PM2.5 and heavy metals can increase respiratory and cardiovascular diseases and reduce the public health level of Ahvaz citizens.

Toxicity assessment of CeO₂ and CuO nanoparticles at the air-liquid interface using bioinspired condensational particle growth

CeO2 and CuO nanoparticles (NPs) are used as additives in petrodiesel to enhance engine performance leading to reduced diesel combustion emissions. Despite their benefits, the additive application poses human health concerns by releasing inhalable NPs into the ambient air. In this study, a bioinspired lung cell exposure system, Dosimetric Aerosol in Vitro Inhalation Device (DAVID), was employed for evaluating the toxicity of aerosolized CeO2 and CuO NPs with a short duration of exposure (≤10 min vs. hours in other systems) and without exerting toxicity from non-NP factors. Human epithelial A549 lung cells were cultured and maintained within DAVID at the air-liquid interface (ALI), onto which aerosolized NPs were deposited, and experiments in submerged cells were used for comparison. Exposure of the cells to the CeO2 NPs did not result in detectable IL-8 release, nor did it produce a significant reduction in cell viability based on lactate dehydrogenase (LDH) assay, with a marginal decrease (10%) at the dose of 388 μg/cm2 (273 cm2/cm2). In contrast, exposure to CuO NPs resulted in a concentration dependent reduction in LDH release based on LDH leakage, with 38% reduction in viability at the highest dose of 52 μg/cm2 (28.3 cm2/cm2). Cells exposed to CuO NPs resulted in a dose dependent cellular membrane toxicity and expressed IL-8 secretion at a global dose five times lower than cells exposed under submerged conditions. However, when comparing the ALI results at the local cellular dose of CuO NPs to the submerged results, the IL-8 secretion was similar. In this study, we demonstrated DAVID as a new exposure tool that helps evaluate aerosol toxicity in simulated lung environment. Our results also highlight the necessity in choosing the right assay endpoints for the given exposure scenario, e.g., LDH for ALI and Deep Blue for submerged conditions for cell viability.

The ribosome-inactivating proteins MAP30 and Momordin inhibit SARS-CoV-2.

The continuing emergence of SARS-CoV-2 variants has highlighted the need to identify additional points for viral inhibition. Ribosome inactivating proteins (RIPs), such as MAP30 and Momordin which are derived from bitter melon (Momordica charantia), have been found to inhibit a broad range of viruses. MAP30 has been shown to potently inhibit HIV-1 with minimal cytotoxicity. Here we show that MAP30 and Momordin potently inhibit SARS-CoV-2 replication in A549 human lung cells (IC50 ~ 0.2 μM) with little concomitant cytotoxicity (CC50 ~ 2 μM). Both viral inhibition and cytotoxicity remain unaltered by appending a C-terminal Tat cell-penetration peptide to either protein. Mutation of tyrosine 70, a key residue in the active site of MAP30, to alanine completely abrogates both viral inhibition and cytotoxicity, indicating the involvement of its RNA N-glycosylase activity. Mutation of lysine 171 and lysine 215, residues corresponding to those in Ricin which when mutated prevented ribosome binding and inactivation, to alanine in MAP30 decreased cytotoxicity (CC50 ~ 10 μM) but also the viral inhibition (IC50 ~ 1 μM). Unlike with HIV-1, neither Dexamethasone nor Indomethacin exhibited synergy with MAP30 in the inhibition of SARS-CoV-2. From a structural comparison of the two proteins, one can explain their similar activities despite differences in both their active-sites and ribosome-binding regions. We also note points on the viral genome for potential inhibition by these proteins.

Variations in Intracellular Organometallic Reaction Frequency Captured by Single-Molecule Fluorescence Microscopy.

Studies of organometallic reactions in living cells commonly rely on ensemble-averaged measurements, which can obscure the detection of reaction dynamics or location-specific behavior. This information is necessary to guide the design of bioorthogonal catalysts with improved biocompatibility, activity, and selectivity. By leveraging the high spatial and temporal resolution of single-molecule fluorescence microscopy, we have successfully captured single-molecule events promoted by Ru complexes inside live A549 human lung cells. By observing individual allylcarbamate cleavage reactions in real-time, our results revealed that they occur with greater frequency inside the mitochondria than in the non-mitochondria regions. The estimated turnover frequency of the Ru complexes was at least 3-fold higher in the former than the latter. These results suggest that organelle specificity is a critical factor to consider in intracellular catalyst design, such as in developing metallodrugs for therapeutic applications.

Variations in Intracellular Organometallic Reaction Frequency Captured by Single‐Molecule Fluorescence Microscopy

AbstractStudies of organometallic reactions in living cells commonly rely on ensemble‐averaged measurements, which can obscure the detection of reaction dynamics or location‐specific behavior. This information is necessary to guide the design of bioorthogonal catalysts with improved biocompatibility, activity, and selectivity. By leveraging the high spatial and temporal resolution of single‐molecule fluorescence microscopy, we have successfully captured single‐molecule events promoted by Ru complexes inside live A549 human lung cells. By observing individual allylcarbamate cleavage reactions in real‐time, our results revealed that they occur with greater frequency inside the mitochondria than in the non‐mitochondria regions. The estimated turnover frequency of the Ru complexes was at least 3‐fold higher in the former than the latter. These results suggest that organelle specificity is a critical factor to consider in intracellular catalyst design, such as in developing metallodrugs for therapeutic applications.

Alginate-Based 3D A549 Cell Culture Model to Study Paracoccidioides Infection

A three-dimensional (3D) lung aggregate model based on sodium alginate scaffolds was developed to study the interactions between Paracoccidioides brasiliensis (Pb) and lung epithelial cells. The suitability of the 3D aggregate as an infection model was examined using cell viability (cytotoxicity), metabolic activity, and proliferation assays. Several studies exemplify the similarity between 3D cell cultures and living organisms, which can generate complementary data due to the greater complexity observed in these designed models, compared to 2D cell cultures. A 3D cell culture system of human A549 lung cell line plus sodium alginate was used to create the scaffolds that were infected with Pb18. Our results showed low cytotoxicity, evidence of increased cell density (indicative of cell proliferation), and the maintenance of cell viability for seven days. The confocal analysis revealed viable yeast within the 3D scaffold, as demonstrated in the solid BHI Agar medium cultivation. Moreover, when ECM proteins were added to the alginate scaffolds, the number of retrieved fungi was significantly higher. Our results highlight that this 3D model may be promising for in vitro studies of host–pathogen interactions.

Prenol, but Not Vitamin C, of Fruit Binds to SARS-CoV-2 Spike S1 to Inhibit Viral Entry: Implications for COVID-19.

Fruit consumption may be beneficial for fighting infection. Although vitamin C is the celebrity component of fruit, its role in COVID-19 is unclear. Because spike S1 of SARS-CoV-2 binds to angiotensin-converting enzyme 2 (ACE2) on host cells to enter the cell and initiate COVID-19, using an α-screen-based assay, we screened vitamin C and other components of fruit for inhibiting the interaction between spike S1 and ACE2. We found that prenol, but neither vitamin C nor other major components of fruit (e.g., cyanidin and rutin), reduced the interaction between spike S1 and ACE2. Thermal shift assays indicated that prenol associated with spike S1, but not ACE2, and that vitamin C remained unable to do so. Although prenol inhibited the entry of pseudotyped SARS-CoV-2, but not vesicular stomatitis virus, into human ACE2-expressing HEK293 cells, vitamin C blocked the entry of pseudotyped vesicular stomatitis virus, not SARS-CoV-2, indicating the specificity of the effect. Prenol, but not vitamin C, decreased SARS-CoV-2 spike S1-induced activation of NF-κB and the expression of proinflammatory cytokines in human A549 lung cells. Moreover, prenol also decreased the expression of proinflammatory cytokines induced by spike S1 of N501Y, E484K, Omicron, and Delta variants of SARS-CoV-2. Finally, oral treatment with prenol reduced fever, decreased lung inflammation, enhanced heart function, and improved locomotor activities in SARS-CoV-2 spike S1-intoxicated mice. These results suggest that prenol and prenol-containing fruits, but not vitamin C, may be more beneficial for fighting against COVID-19.