human cutaneous squamous cell carcinoma cell line a431  (ATCC)

Journal: Cell Death & Disease

Article Title: Gas plasma-induced bacterial PAMP release promotes skin cancer cell death

doi: 10.1038/s41419-025-08283-8

Figure Lengend Snippet: a representative image of gas plasma treatment; b metabolic activity of A375, SCC-25, and A431 cells 24 h post gas plasma treatment and calculated IC 25 values; c metabolic activity of A375, SCC-25, and A431 cells 24 h, 48 h, and 72 h after treatment with three different dilutions of bacterial pPAMP-containing supernatants, statistical analysis was performed using two-way analysis of variance (ANOVA); d metabolic activity of A375, SCC-25 and A431 cells 24 h after gas plasma or PAMP treatment alone or combination treatment, statistical analysis was performed using one-way ANOVA; e representative flow cytometry contour plots of the viable cell population; f quantification of viable cells 24 h after gas plasma or PAMP treatment alone or combination treatment, statistical analysis was performed using two-way ANOVA; g representative flow cytometry histogram of DAPI intensity for cell cycle analysis; h G2/G1 phase ratio in A375, SCC-25 and A431 cells 24 h after gas plasma or PAMP treatment alone or combination treatment, statistical analysis was performed using one-way ANOVA. cPAMPs = PAMPs derived from untreated bacteria. ctrl = control. hPAMPs = PAMPs derived from heat-inactivated bacteria. LPS = lipopolysaccharide. PL = gas plasma. pPAMPs = PAMPs derived from gas plasma-exposed bacteria.

Article Snippet: The human melanoma cell line A375 and the human squamous cell carcinoma cell lines A431, as well as SCC-25 (all ATCC, USA), were cultured in Roswell Park Memorial Institute (RPMI) 1640 medium (Pan Biotech, Germany) supplemented with 10% fetal bovine serum, 1% penicillin/ streptomycin, and 1% L-glutamine (all Corning, Germany).

Techniques: Clinical Proteomics, Activity Assay, Flow Cytometry, Cell Cycle Assay, Derivative Assay, Bacteria, Control

Journal: Cell Death & Disease

Article Title: Gas plasma-induced bacterial PAMP release promotes skin cancer cell death

doi: 10.1038/s41419-025-08283-8

Figure Lengend Snippet: a representative flow cytometry intensity histograms of CD40 levels in control and treated SCC-25 cells; b heatmap showing medians of calculated expression changes of eight different surface markers in response to gas plasma or PAMP treatment alone or a combination treatment of both ( n = 4); c quantification of intracellular γH2AX expression in A375, SCC-25 and A431 cells 24 h after gas plasma or PAMP treatment alone or combination treatment, violin plots show median (indicated as stacked line) and quartiles (indicated as dotted line), statistical analysis was performed using one-way ANOVA ( n = 4); d representative images of actin cytoskeleton and nucleus staining in control and treated tumor cells; e representative images of DiD, caspase 3/7 and DAPI staining in control and treated A375 cells; f–h fluorescence microscopy-based quantification of the cell number per well 24 h after gas plasma or PAMP treatment alone or combination treatment ( f ) ( n = 4), DAPI intensity per well ( g ) ( n = 4), and caspase 3/7 intensity per well ( h ) ( n = 4) 24 h after gas plasma or PAMP treatment alone or combination treatment, data is shown as box and whiskers (Tukey; mean is shown as ′+′), statistical analysis was performed using unpaired t test (one-tailed). cas 3/7 = caspase 3/7. cPAMPs = PAMPs derived from untreated bacteria. ctrl = control. hPAMPs = PAMPs derived from heat-inactivated bacteria. LPS = lipopolysaccharide. PL = gas plasma. pPAMPs = PAMPs derived from gas plasma-exposed bacteria.

Article Snippet: The human melanoma cell line A375 and the human squamous cell carcinoma cell lines A431, as well as SCC-25 (all ATCC, USA), were cultured in Roswell Park Memorial Institute (RPMI) 1640 medium (Pan Biotech, Germany) supplemented with 10% fetal bovine serum, 1% penicillin/ streptomycin, and 1% L-glutamine (all Corning, Germany).

Techniques: Flow Cytometry, Control, Expressing, Clinical Proteomics, Staining, Fluorescence, Microscopy, One-tailed Test, Derivative Assay, Bacteria

Journal: Journal of Clinical Medicine

Article Title: An Evaluation of Prognostic Factors in Cutaneous Squamous Cell Carcinoma: A Single-Center Study of 237 Japanese Cases

doi: 10.3390/jcm14041243

Figure Lengend Snippet: List of top 10 differentially expressed genes (DEG) in A431 stimulated with LL-37.

Article Snippet: Human epidermoid squamous carcinoma A431 cell lines were purchased from the American Type Culture Collection (Manassas, VA, USA).

Techniques: Expressing

Journal: bioRxiv

Article Title: An extended and apical supracellular actin network interconnects squamous carcinoma cells

doi: 10.1101/2025.01.10.632450

Figure Lengend Snippet: (A) Representative bright-field images of the gaps between both A431 cell sheets at 0, 24, 48 and 72h of migration after removing the insert. The yellow lines represent the contour of the gaps not covered by the cells (wound area). Scale bar = 200 µm. (B) Representative bright-field images of the outside periphery of both A431 cell sheets at 0, 24, 48 and 72h of migration after removing the insert. The yellow arrowheads point towards multicellular structures called fingers. Scale bar = 200 µm. (C) Wound area after 0, 24, 48 and 72h of A431 cell migration. For each condition, 8 pictures per experiment obtained in 3 independent experiments were analysed. (D-F) Mean number of fingers/mm (D) , elongation of fingers (E) and area of fingers (F) formed after 24, 48 and 72h of A431 cell migration. The elongation was calculated by the ratio length/width of fingers and the area by the entire surface over the base of the fingers. For each condition, 24 pictures per experiment obtained in 3 independent experiments were analysed. All the data are presented as mean ± standard deviation (s.d.) and tested by Kruskal-Wallis (ns: not significant; *: p<0,05; **: p<0,01; ****: p<0,0001).

Article Snippet: Human skin squamous carcinoma A431 cell line (#CRL-1555), human tongue squamous carcinoma CAL27 cell line (#CRL-2095) and immortalized human embryonic kidney HEK-293T cell line (#CRL-3216) were purchased from ATCC.

Techniques: Migration, Standard Deviation

Journal: bioRxiv

Article Title: An extended and apical supracellular actin network interconnects squamous carcinoma cells

doi: 10.1101/2025.01.10.632450

Figure Lengend Snippet: (A) Time-lapse DIC imaging of a finger formed by A431 cells after 56h of migration (top). Three images took at 15 min intervals show the protrusion and retraction dynamics of cells at the front of migration. Scale bar = 100 µm. Two kymographs, generated by drawing lines in the middle of a leader cell (pink line, left rectangle) and a side cell (blue line, right rectangle), show the protrusion and retraction dynamics of both cell types from 56 to 60h of migration (bottom). (B) Mean velocity of leader cells and side cells at different timepoint intervals during A431 cell migration. For each time interval, 12 leader cells and 12 side cells from one experiment were analysed. (C) Z-projections of representative confocal images of A431 cell sheets stained for vimentin and fibronectin after 0, 24, 48 and 72h of migration. Orange and yellow lines delimit respectively the front and rear of cell sheets. Scale bar = 100 µm (D) Vimentin mean fluorescence intensity at the front and at the rear of A431 cell sheets after 0, 24, 48 and 72h of migration. For each condition, 7 to 10 pictures per experiment of 3 independent experiments were analysed. (E) Difference in vimentin mean fluorescence intensity between the front and the rear in A431 cell sheets after 0, 24, 48 and 72h of migration. 7 to 10 pictures per experiment of 3 independent experiments were analysed. (F) Fibronectin mean fluorescence intensity at the front and at the rear of A431 cell sheets after 0, 24, 48 and 72h of migration. For each condition, 7 to 8 pictures per experiment of 3 independent experiments were analysed. (G) Difference in fibronectin mean fluorescence intensity between the front and the rear in A431 cell sheets after 0, 24, 48 and 72h of migration. 7 to 8 pictures per experiment of 3 independent experiments were analysed. All the data are presented as mean ± standard deviation (s.d.) and Two-way ANOVA (B), Wilcoxon (D, F) or Kruskall-Wallis (E, G) (ns: not significant; *: p<0,05; **: p<0,01; ***: p<0,001; ****: p<0,0001).

Article Snippet: Human skin squamous carcinoma A431 cell line (#CRL-1555), human tongue squamous carcinoma CAL27 cell line (#CRL-2095) and immortalized human embryonic kidney HEK-293T cell line (#CRL-3216) were purchased from ATCC.

Techniques: Imaging, Migration, Generated, Staining, Fluorescence, Standard Deviation

Journal: bioRxiv

Article Title: An extended and apical supracellular actin network interconnects squamous carcinoma cells

doi: 10.1101/2025.01.10.632450

Figure Lengend Snippet: (A-B) Representative maximum intensity projections of the basal (A) and apical (B) Z-planes of a finger formed after 72h of A431 migration. The actin and nuclei were stained. The red, yellow and blue rectangles show respectively the actin arcs, hubs and cables. Scale bar = 50 µm. (C-E) Magnifications of the red, yellow and blue rectangles from the previous images showing areas with different actin structures: arcs (C), hubs (D) and cables (E). Each panel displays the actin signal (top) and a schematic representation (bottom) of the structure. Scale bar = 20 µm. (F) Representative xy (top) and xz (bottom) images at enhanced resolution acquired with an Airyscan detector, of a confocal Z-stack (left) showing an area with hubs (black arrowheads) and cables (black arrows) in A431 cells after 72h of migration. A white arrow was drawn from the substrate to the top on the xz merge image and used to generate a plot profile (right) representing the normalized fluorescence intensity of each signal along the arrow. Scale bar = 5 µm. (G) Representative maximum intensity projections of the apical Z-planes showing an area with cables in A431 cells after 72h of migration (left). A mosaic of cells stably expressing the filamentous actin probe LifeAct fused to either GFP or iRFP670 was generated and stained for β-catenin. Full images (top) and magnifications (bottom) are shown. Magnifications are high resolution images captured using an Airyscan detector. A white arrow was drawn from a LifeAct-GFP expressing cell to a LifeAct-iRFP670 expressing cell and used to generate a plot profile (right) representing the normalized fluorescence intensity of each signal along the arrow. Scale bar = 20 µm (top) and 5 µm (bottom). (H) Representative maximum intensity projections of the apical Z-planes showing a cable (arrows, left) and a hub linked to a cable (arrowhead and arrows, right) in A431 cells after 72h of migration. α-actinin 1, actin and β-catenin were stained. These are high resolution images captured using an Airyscan detector. Scale bar = 5 µm

Article Snippet: Human skin squamous carcinoma A431 cell line (#CRL-1555), human tongue squamous carcinoma CAL27 cell line (#CRL-2095) and immortalized human embryonic kidney HEK-293T cell line (#CRL-3216) were purchased from ATCC.

Techniques: Migration, Staining, Fluorescence, Stable Transfection, Expressing, Generated

Journal: bioRxiv

Article Title: An extended and apical supracellular actin network interconnects squamous carcinoma cells

doi: 10.1101/2025.01.10.632450

Figure Lengend Snippet: (A) Representative maximum intensity projections of the apical Z-planes of the A431 cell sheet periphery after 0, 24, 48 and 72h of migration and actin staining. Scale bar = 50 µm. Red and blue arrowheads point towards hubs and cables. (B-D) Number of hubs per mm 2 (B), area of hubs (C) and total length of cables per mm 2 (D) after 0, 24, 48 and 72h of A431 cell migration. 7 pictures per experiment of 3 independent experiments were analysed. (E) Mean orientation of the apical actin structures after 0, 24, 48 and 72h of A431 cell migration. The orientation goes from −90° to 90° with 0° corresponding to the direction of migration. 12 pictures per experiment of 3 independent experiments were analysed. (F) Percentage of apical actin structures after 0, 24, 48 and 72h of A431 cell migration with angles from −30° to 30°, angles from −60° to −30° and from 30° to 60° and angles from −90° to −60° and from 60° to 90°. 12 pictures per experiment of 3 independent experiments were analysed. All the data are presented as mean ± s.d. and tested by Kruskal-Wallis (B-D) or Two-way ANOVA (F) (ns: not significant; **: p<0,01; ***: p<0,001; ****: p<0,0001)

Article Snippet: Human skin squamous carcinoma A431 cell line (#CRL-1555), human tongue squamous carcinoma CAL27 cell line (#CRL-2095) and immortalized human embryonic kidney HEK-293T cell line (#CRL-3216) were purchased from ATCC.

Techniques: Migration, Staining

Journal: bioRxiv

Article Title: An extended and apical supracellular actin network interconnects squamous carcinoma cells

doi: 10.1101/2025.01.10.632450

Figure Lengend Snippet: (A) Representative maximum intensity projections of the apical Z-planes of a part of an A431 cell sheet that migrated during 72h before being treated with the vehicle (control, left) or 4 mM of the calcium chelator EGTA (right). 0 min corresponds to the beginning of the treatment. Cells are stably expressing the actin probe LifeAct-iRFP670 and the adherens junction protein E-cadherin-mCitrine. Green, red and blue arrowheads point towards cables and hubs that are maintained in the control condition but disappear after EGTA treatment. Scale bar = 50 µm. (B) Representative maximum intensity projections of the apical Z-planes of the front (left) and rear (right) of migration of A431 cell sheets. Using non-target and specific shRNAs, control (top) and p120-catenin (bottom) knocked down cells were generated, allowed to migrate during 72h and stained for actin. Red and blue arrowheads point towards hubs and cables. Scale bar = 50 µm. (C, D) Number of hubs per mm 2 (C) and total length of cables per mm 2 (D) at the front and at the rear of migration of control and p120-catenin knocked down A431 cells that migrated during 72h. For each condition, 4 to 5 pictures per experiment of 3 independent experiments were analysed. (E) Representative bright-field images of the gaps between control and p120-catenin knocked down A431 cells sheets after 0, 24, 48 and 72h of migration. The yellow lines represent the contour of the gaps not covered by the cells (wound area). Scale bar = 200 µm. (F) Wound area after 0, 24, 48 and 72h of migration of control and p120-catenin knocked down A431 cells. For each condition, 4 pictures per experiment obtained in 3 independent experiments were analysed. All the data are presented as mean ± s.d. and tested by Two-way ANOVA (ns: not significant; **: p<0,01; ****: p<0,0001)

Article Snippet: Human skin squamous carcinoma A431 cell line (#CRL-1555), human tongue squamous carcinoma CAL27 cell line (#CRL-2095) and immortalized human embryonic kidney HEK-293T cell line (#CRL-3216) were purchased from ATCC.

Techniques: Control, Stable Transfection, Expressing, Migration, Generated, Staining

Journal: bioRxiv

Article Title: An extended and apical supracellular actin network interconnects squamous carcinoma cells

doi: 10.1101/2025.01.10.632450

Figure Lengend Snippet: (A) Representative maximum intensity projections of the apical Z-planes of a finger formed after 72h of A431 migration. The phospho-myosin light chain 2 (pMLC2) and actin were stained. Black and white arrowheads point towards supracellular actin structures that colocalize with the pMLC2 staining. Scale bar = 50 µm. (B) Representative maximum intensity projections of the apical Z-planes of fingers formed after 72h of migration of A431 treated with DMSO (control) or 1 µM of the ROCK inhibitor Y-27632. Cells were stained for actin. Red and blue arrowheads point towards hubs and cables. Scale bar = 50 µm. (C-D) Number of hubs per mm 2 (C) and total length of cables per mm 2 (D) after 72h of treatment of A431 cells with DMSO (control) or 1 µM of Y-27632. For each condition, 7 to 8 pictures per experiment of 3 independent experiments were analysed. (E) Representative maximum intensity projections of the apical Z-planes of a part of an A431 cell sheet that migrated during 72h before being treated with DMSO (control, top) or 5 µM of Y-27632 (bottom). 0 min corresponds to the beginning of the treatment. Cells are stably expressing the actin probe LifeAct-iRFP670. Green, red and blue arrowheads point towards cables and hubs that are maintained in the control condition but disappear after Y-27632 treatment. Scale bar = 50 µm. (F) Representative bright-field images of the gaps between sheets of A431 cells treated with DMSO (control) or different concentrations of Y-27632 after 0, 24, 48 and 72h of migration. The yellow lines represent the contour of the gaps not covered by the cells (wound area). Scale bar = 200 µm. (G) Wound area after 0, 24, 48 and 72h of migration of A431 cells treated with DMSO (control) or different concentrations of Y-27632. For each condition, 4 pictures per experiment obtained in 3 independent experiments were analysed. All the data are presented as mean ± s.d. and tested by Mann-Whitney (C, D) or Two-way ANOVA (G) (ns: not significant; *: p<0,05; **: p<0,01; ****: p<0,0001)

Article Snippet: Human skin squamous carcinoma A431 cell line (#CRL-1555), human tongue squamous carcinoma CAL27 cell line (#CRL-2095) and immortalized human embryonic kidney HEK-293T cell line (#CRL-3216) were purchased from ATCC.

Techniques: Migration, Staining, Control, Stable Transfection, Expressing, MANN-WHITNEY

Journal: bioRxiv

Article Title: An extended and apical supracellular actin network interconnects squamous carcinoma cells

doi: 10.1101/2025.01.10.632450

Figure Lengend Snippet: (A) Representative maximum intensity projections of the apical Z-planes of fingers formed after 72h of migration of A431 treated with DMSO (control) or 0,5 µM of the MAP4K4 inhibitor GNE-495. Cells were stained for actin. Red and blue arrowheads point towards hubs and cables. ––Scale bar = 50 µm. (B-D) Number of hubs per mm 2 (B), area of hubs (C) and total length of cables per mm 2 (D) after 72h of treatment of A431 cells with DMSO (control) or 0,5 µM of GNE-495. For each condition, 8 pictures per experiment obtained in 3 independent experiments were analysed. (E) Representative bright-field images of the gaps between sheets of A431 cells treated with DMSO (control) or different concentrations of GNE-495 after 0, 24, 48 and 72h of migration. The yellow lines represent the contour of the gaps not covered by the cells (wound area). Scale bar = 200 µm. (F) Wound area after 0, 24, 48 and 72h of migration of A431 cells treated with DMSO (control) or different concentrations of GNE-495. For each condition, 4 pictures per experiment obtained in 5 independent experiments were analysed. (G) Representative bright-field images of the periphery of sheets of A431 cells after 72h of migration and treatment with DMSO or different concentrations of GNE-495. The yellow arrowheads point towards fingers. Scale bar = 200 µm. (H-J) Mean number of fingers/mm (H), elongation of fingers (I) and area of fingers (J) formed after 72h of migration of A431 cells treated with DMSO or different concentrations of GNE-495. The elongation was calculated by the ratio length/width of fingers and the area by the entire surface over the base of the fingers. For each condition, 12 pictures per experiment obtained in 5 independent experiments were analysed. All the data are presented as mean ± s.d. and tested by Mann-Whitney (B-D), Two-way ANOVA (F) or Kruskal-Wallis (H-J) (ns: not significant; *: p<0,05; ***: p<0,001; ****: p<0,0001)

Article Snippet: Human skin squamous carcinoma A431 cell line (#CRL-1555), human tongue squamous carcinoma CAL27 cell line (#CRL-2095) and immortalized human embryonic kidney HEK-293T cell line (#CRL-3216) were purchased from ATCC.

Techniques: Migration, Control, Staining, MANN-WHITNEY

Journal: bioRxiv

Article Title: An extended and apical supracellular actin network interconnects squamous carcinoma cells

doi: 10.1101/2025.01.10.632450

Figure Lengend Snippet: (A) Confocal images of a cable that is cut by laser ablation in A431 cells that migrated during 96 h. The nuclei, actin and plasma membranes were stained. 0 sec is just before the ablation of the cable. The red line shows where the structure has been severed. The black dotted lines delimit the maximum recoil of the cable. Cells are stably expressing the actin probe LifeAct-iRFP670. Scale bar = 10 µm. (B) Representative maximum intensity projections of the basal (top) and apical (bottom) Z-planes showing a finger formed after 72h of A431 migration. The actin, vinculin and p120-catenin were stained. In each panel, full images and magnifications are shown. Scale bar = 50 µm (full image) or 20 µm (magnification).

Article Snippet: Human skin squamous carcinoma A431 cell line (#CRL-1555), human tongue squamous carcinoma CAL27 cell line (#CRL-2095) and immortalized human embryonic kidney HEK-293T cell line (#CRL-3216) were purchased from ATCC.

Techniques: Clinical Proteomics, Staining, Stable Transfection, Expressing, Migration