Journal: iScience

Article Title: Development of human alveolar epithelial cell models to study distal lung biology and disease

doi: 10.1016/j.isci.2022.103780

Figure Lengend Snippet: Derivation of AEC lines that proliferate in culture but do not form tumors in nude mice (A) AEC line derivation scheme. Previously frozen purified human AT2 cells from Lung-FT were first cultured in Y-27632 medium. Proliferative cells (a) were then transduced with LeGO iT (tdTomato) and LeGO iG (eGFP) lentiviruses carrying hTERT (b), CDK4 R24C (c), CDK4 R24C + hTERT (d), and SV40 LgT (e). Scale bar, 100 μm. (B, C) Additional SV40 LgT-transduced cell lines were established from AT2 cells purified from (B) Lung-ON and (C) Lung-TN in Y-27632 medium (f and h), then transduced with SV40 LgT (g and i). Scale bar, 100 μm. (D) Cell proliferation assays in technical triplicates from three independent experiments (mean total number of cells ± std dev). ∗p<0.05, ∗∗p< 0.005 by independent two-sample t test relative to AEC-FT-ROCKinh. (E) Cell proliferation assays for AEC-ON and AEC-TN, under standard (1,000 cells/well) and high-density (5,000 cells/well) conditions. (F) Anchorage-independent growth assays. A549 cells, positive control. Colonies were stained with crystal violet and counted using ImageJ/Fiji after 1 month. Inset images, |×2.5×| magnified to show colonies. (G) Colony quantification of six technical replicates from at least three independent experiments (mean ± std dev). N.S., not significant; ∗p< 0.05, nonparametric Wilcoxon test. (H) Subcutaneous injection into NU/J mice to assess tumorigenicity of AEC-FT, AEC-ON, and AEC-TN cell lines over 3 months. A549 cells, positive control; AEC-hTERT line, negative control. Equal numbers of male and female mice per group. Sample labels indicate which cell line was injected into the left and right flanks (left/right). (I) Photos of excised nodules (scale bar, 5 mm) and corresponding H&E stainings. (J) Graph of nodule growth starting at 3 weeks postinjection. See also Figure S1 , and .

Article Snippet: Human A549 lung adenocarcinoma cells , ATCC , CRM-CCL-185; RRID:CVCL_0023.

Techniques: Purification, Cell Culture, Transduction, Positive Control, Staining, Injection, Negative Control

Journal: iScience

Article Title: Development of human alveolar epithelial cell models to study distal lung biology and disease

doi: 10.1016/j.isci.2022.103780

Figure Lengend Snippet:

Article Snippet: Human A549 lung adenocarcinoma cells , ATCC , CRM-CCL-185; RRID:CVCL_0023.

Techniques: Transduction, Control, Plasmid Preparation, Virus, Recombinant, Membrane, Software

Journal: Toxics

Article Title: PBDEs Found in House Dust Impact Human Lung Epithelial Cell Homeostasis

doi: 10.3390/toxics10020097

Figure Lengend Snippet: Viability of A549 cells after prolonged (24, 48 and 72 h) exposure to PBDEs at concentrations from 1.5 to 48 μg mL −1 determined with the MTS assay; 0 μg mL −1 represents untreated cells. The results are expressed as percentage of corresponding control, untreated cells and given as means ± SD ( n = 6). The dashed frame highlights a time-dependent effect. & p < 0.05; # p < 0.01; * p < 0.0001 vs. untreated control.

Article Snippet: The certified human carcinoma alveolar basal epithelial A549 cell line (ECACC 86012804) was purchased from the European Collection of Authenticated Cell Cultures (ECACC) cell bank.

Techniques: MTS Assay, Control

Journal: Toxics

Article Title: PBDEs Found in House Dust Impact Human Lung Epithelial Cell Homeostasis

doi: 10.3390/toxics10020097

Figure Lengend Snippet: Cytotoxicity curves of BDE-99 and -209 at concentrations from 1.5 to 48 μg mL −1 in A549 cells over 24, 48 and 72 h. The results of the MTS assay are given as means ± SD ( n = 6).

Article Snippet: The certified human carcinoma alveolar basal epithelial A549 cell line (ECACC 86012804) was purchased from the European Collection of Authenticated Cell Cultures (ECACC) cell bank.

Techniques: MTS Assay

Journal: Toxics

Article Title: PBDEs Found in House Dust Impact Human Lung Epithelial Cell Homeostasis

doi: 10.3390/toxics10020097

Figure Lengend Snippet: IC 50 values of BDE-99 and -209 in A549 cells over 24, 48 and 72 h.

Article Snippet: The certified human carcinoma alveolar basal epithelial A549 cell line (ECACC 86012804) was purchased from the European Collection of Authenticated Cell Cultures (ECACC) cell bank.

Techniques:

Journal: Toxics

Article Title: PBDEs Found in House Dust Impact Human Lung Epithelial Cell Homeostasis

doi: 10.3390/toxics10020097

Figure Lengend Snippet: Prolonged exposure of A549 cells to a house dust sample extract. A549 cells were exposed to ∑PBDE in house dust at concentrations from 0.375 to 24 μg mL −1 for 24, 48 and 72 h, and cell viability was determined with the MTS assay. The results are given as means ± SD ( n = 6). The dashed frame highlights the concentration found in the house dust sample. & p < 0.05; # p < 0.01; vs. untreated control.

Article Snippet: The certified human carcinoma alveolar basal epithelial A549 cell line (ECACC 86012804) was purchased from the European Collection of Authenticated Cell Cultures (ECACC) cell bank.

Techniques: MTS Assay, Concentration Assay, Control

Journal: Toxics

Article Title: PBDEs Found in House Dust Impact Human Lung Epithelial Cell Homeostasis

doi: 10.3390/toxics10020097

Figure Lengend Snippet: LOAEL cytotoxicity of BDE-99 (7.5 μg mL −1 ) and BDE-209 (15 μg mL −1 ), alone and combined, in A549 cells treated for 24, 48 and 72 h. The results of the MTS assay are given as means ± SE ( n = 6). # p < 0.01; $ p < 0.001 vs. untreated control.

Article Snippet: The certified human carcinoma alveolar basal epithelial A549 cell line (ECACC 86012804) was purchased from the European Collection of Authenticated Cell Cultures (ECACC) cell bank.

Techniques: MTS Assay, Control

Journal: Toxics

Article Title: PBDEs Found in House Dust Impact Human Lung Epithelial Cell Homeostasis

doi: 10.3390/toxics10020097

Figure Lengend Snippet: Levels of LDH release after 4 h exposure of A549 cells to PBDE congeners and house dust sample extract. Triton (0.08%) was used as the positive control. The results are presented as percentage of LDH release and given as means ± SD ( n = 3). # p < 0.01; $ p < 0.001; * p < 0.0001 vs. untreated control.

Article Snippet: The certified human carcinoma alveolar basal epithelial A549 cell line (ECACC 86012804) was purchased from the European Collection of Authenticated Cell Cultures (ECACC) cell bank.

Techniques: Positive Control, Control

Journal: Toxics

Article Title: PBDEs Found in House Dust Impact Human Lung Epithelial Cell Homeostasis

doi: 10.3390/toxics10020097

Figure Lengend Snippet: ROS levels after the 4 h exposure of A549 cells to PBDE congeners, house dust sample extract and H 2 O 2 (100 µM) as positive control. The results of DCF-fluorescence signal measurements are presented as relative fluorescence units (RFU) and given as means ± SD ( n = 6). & p < 0.05; # p < 0.01; $ p < 0.001; * p < 0.0001 vs. untreated control.

Article Snippet: The certified human carcinoma alveolar basal epithelial A549 cell line (ECACC 86012804) was purchased from the European Collection of Authenticated Cell Cultures (ECACC) cell bank.

Techniques: Positive Control, Fluorescence, Control

Journal: Toxics

Article Title: PBDEs Found in House Dust Impact Human Lung Epithelial Cell Homeostasis

doi: 10.3390/toxics10020097

Figure Lengend Snippet: GSH levels after the 4 h exposure of A549 cells to single PBDEs and house dust sample extract at 3, 6 and 12 µg mL −1 and to tBOOH (100 µM) as positive control. The results of MCB-fluorescence measurements are presented as relative fluorescence units (RFU) and given as means ± SD ( n = 6). & p < 0.05; # p < 0.01; $ p < 0.001; * p < 0.0001 vs. untreated control.

Article Snippet: The certified human carcinoma alveolar basal epithelial A549 cell line (ECACC 86012804) was purchased from the European Collection of Authenticated Cell Cultures (ECACC) cell bank.

Techniques: Positive Control, Fluorescence, Control

Journal: Toxics

Article Title: PBDEs Found in House Dust Impact Human Lung Epithelial Cell Homeostasis

doi: 10.3390/toxics10020097

Figure Lengend Snippet: Mitochondrial membrane potential after 4 h exposure of A549 cells to single BDE congeners and house dust sample extract at 3, 6 and 12 µg mL −1 and CCCP (50 µM) as positive control. The results are presented as relative fluorescence units (RFU) and given as means ± SD ( n = 6). & p < 0.5, # p < 0.01, * p < 0.001 vs. untreated control.

Article Snippet: The certified human carcinoma alveolar basal epithelial A549 cell line (ECACC 86012804) was purchased from the European Collection of Authenticated Cell Cultures (ECACC) cell bank.

Techniques: Membrane, Positive Control, Fluorescence, Control

Journal: Toxics

Article Title: PBDEs Found in House Dust Impact Human Lung Epithelial Cell Homeostasis

doi: 10.3390/toxics10020097

Figure Lengend Snippet: Percentage of different stages of apoptosis of A549 cells after 4 h exposure to single PBDE congener and house dust sample extract at 12 µg mL −1 and PFA (0.08%, v / v ) as a positive control. The results are presented as percentage of total apoptotic cells compared to vehicle control (2.4% methanol) and given as means ± SD ( n = 6). * p < 0.001 vs. control.

Article Snippet: The certified human carcinoma alveolar basal epithelial A549 cell line (ECACC 86012804) was purchased from the European Collection of Authenticated Cell Cultures (ECACC) cell bank.

Techniques: Positive Control, Control

Journal: Cell Death & Disease

Article Title: Reciprocal positive regulation between Cx26 and PI3K/Akt pathway confers acquired gefitinib resistance in NSCLC cells via GJIC-independent induction of EMT

doi: 10.1038/cddis.2015.197

Figure Lengend Snippet: Increased Cx26 is positively correlated with gefitinib resistance in NSCLC cells. ( a ) Differential expression of Cx26, Cx31.1, Cx32, and Cx43 in different gefitinib-sensitive NSCLC cell lines was determined by RT-PCR. ( b and c ) High level of Cx26 in gefitinib-insensitive A549 and H1299 cells than that in gefitinib-sensitive HCC827 and PC9 cells was detected by RT-PCR and western blotting. GAPDH or β -actin was used as internal loading control

Article Snippet: Human NSCLC cell lines (HCC827, PC9, A549, and H1299) were originally obtained from the ATCC (Manassas, VA, USA).

Techniques: Quantitative Proteomics, Reverse Transcription Polymerase Chain Reaction, Western Blot, Control

Journal: Cell Death & Disease

Article Title: Reciprocal positive regulation between Cx26 and PI3K/Akt pathway confers acquired gefitinib resistance in NSCLC cells via GJIC-independent induction of EMT

doi: 10.1038/cddis.2015.197

Figure Lengend Snippet: Cx26 induces acquired gefitinib resistance in NSCLC cells via GJIC-independent manner. ( a ) Functional GJIC was detected by parachute assay and no detectable GJIC was found in HCC827 GR, PC9 GR, and their parental cells. Top: fluorescence images. Bottom: overlaid the corresponding phase-contrast images. Original magnification, × 200. ( b ) No enhancement of GJIC in these cells incubated with 10, 20, and 40 μ M of RA (a well-defined GJIC enhancer) for 4, 8, 12, 24, and 48 h, respectively. Top: fluorescence images. Bottom: overlaid the corresponding phase-contrast images. Original magnification, × 200. ( c and d ) Immunofluorescence staining of the cellular localization of Cx26 with or without RA treatment. All scare bars represent 50 μ m

Article Snippet: Human NSCLC cell lines (HCC827, PC9, A549, and H1299) were originally obtained from the ATCC (Manassas, VA, USA).

Techniques: Functional Assay, Fluorescence, Incubation, Immunofluorescence, Staining

Journal: Cell Death & Disease

Article Title: Reciprocal positive regulation between Cx26 and PI3K/Akt pathway confers acquired gefitinib resistance in NSCLC cells via GJIC-independent induction of EMT

doi: 10.1038/cddis.2015.197

Figure Lengend Snippet: Cx26 and PI3K/Akt pathway functionally interplay to promote EMT and gefitinib resistance in NSCLC cells. ( a and b ) Effect of LY294002 or Akt overexpression on Cx26 expression in HCC827, PC9, and their GR cells was determined by western blotting. ( c ) Effects of Akt overexpression alone or combined with Cx26 overexpression or Cx26 depletion on cell morphology changes in HCC827 and PC9 cells. Original magnification, × 400. ( d – f ) Effects of Akt overexpression alone or combined with Cx26 overexpression or Cx26 depletion on the expression of EMT markers (E-cadherin, vimentin, and slug), cell migration, and invasion, as well as cell sensitivity to gefitinib in HCC827 and PC9 cells, respectively. Error bars are mean±S.D. from four independent experiments, ** P <0.01 versus vector group. # P <0.05 and ## P <0.01 versus Akt-overexpressing group

Article Snippet: Human NSCLC cell lines (HCC827, PC9, A549, and H1299) were originally obtained from the ATCC (Manassas, VA, USA).

Techniques: Over Expression, Expressing, Western Blot, Migration, Plasmid Preparation

Journal: PLoS ONE

Article Title: Norepinephrine and Epinephrine Enhanced the Infectivity of Enterovirus 71

doi: 10.1371/journal.pone.0135154

Figure Lengend Snippet: Cells were stained with (A) anti-α 1A ADR or (B) β 2 -ADR antibodies. After incubation, washing and centrifugation, alexa 488-conjugated gaot anit-rabbit IgG antibody was added and determined by flow cytometry (red filled region). PBS stained cells (blue lines) and secondary antibody stained cells (green lines) were showed as negative controls in each experiment. A549, RD, SK-N-SH, HL-60, THP-1, Jurkat and hPBMCs exhibited (A) α 1A - and (B) β 2 -ADRs. (C) The expression of β 2 -ADR on PMA-induced THP-1 cells were observed by immunofluorescent microscopy. The green fluorescence represented β 2 -adrenergic receptor. DAPI (blue) indicated cell nucleus. hPBMC, human peripheral blood mononuclear cells. The data were represented from three independent experiments.

Article Snippet: Cells of A549 (human lung carcinoma) (BCRC No. 60074; October 2008), RD (human rhabdomyosarcoma) (BCRC No. 60113; October 2008), SK-N-SH (human neuroblastoma) (ATCC No. HTB-11; December 2008), HL-60 (human promyelocytic leukemia) (BCRC No. 60027; March 2009), THP-1 (human monocytic leukemia) (BCRC No. 60430; October 2008), and Jurkat (human T cell leukemia) (BCRC No. 60424; January 2009) were maintained in Dulbecco's Modified Eagle Medium (DMEM) or RPMI 1640 medium (Life Technologies Gibco, USA) with heat-inactivated 10% fetal bovine serum (FBS) (Biological Industries, Israel) according to the instructions of the manufacturer [ , , ].

Techniques: Staining, Incubation, Centrifugation, Flow Cytometry, Expressing, Microscopy, Fluorescence

Journal: PLoS ONE

Article Title: Norepinephrine and Epinephrine Enhanced the Infectivity of Enterovirus 71

doi: 10.1371/journal.pone.0135154

Figure Lengend Snippet: (A) A549 cells were infected with EV71 (MOI = 5) for 24 hrs and then treated with NE for 12 hrs. (B) SK-N-SH cells were infected with EV71 (MOI = 1) for 18 hrs and then treated with NE for 6 hours. (C, D) hPBMCs were infected with EV71 (MOI = 10) for 24 hrs and then treated with NE and EP for 24 hrs. The virus titers were significantly higher in (A) EV71-infected A549 cells and (B) SK-N-SH with various concentrations of NE than in EV71-infected cells without NE treatment. Both (C) NE and (D) EP elevated virus titers in EV71-infected hPBMCs at concentrations of 10 2 and 10 6 pg/mL, respectively. The data are shown as mean ± SEM of three independent experiments. White bar: virus only; black bar: virus plus NE or EP. *, P < 0.05; and **, P < 0.01 compared to virus only group and analyzed by one-way ANOVA and Post-Hoc Tukey test.

Article Snippet: Cells of A549 (human lung carcinoma) (BCRC No. 60074; October 2008), RD (human rhabdomyosarcoma) (BCRC No. 60113; October 2008), SK-N-SH (human neuroblastoma) (ATCC No. HTB-11; December 2008), HL-60 (human promyelocytic leukemia) (BCRC No. 60027; March 2009), THP-1 (human monocytic leukemia) (BCRC No. 60430; October 2008), and Jurkat (human T cell leukemia) (BCRC No. 60424; January 2009) were maintained in Dulbecco's Modified Eagle Medium (DMEM) or RPMI 1640 medium (Life Technologies Gibco, USA) with heat-inactivated 10% fetal bovine serum (FBS) (Biological Industries, Israel) according to the instructions of the manufacturer [ , , ].

Techniques: Infection, Virus