Journal: Clinical Cancer Research

Article Title: The Pancreas Cancer Microenvironment

doi: 10.1158/1078-0432.ccr-11-3114

Figure Lengend Snippet: Figure 1. PDA contains an abundant stroma. A, mouse PDA tissue was stained with picrosirius red and imaged under polarized light to visualize collagen fibers (20 magnification). B, Masson trichrome staining highlights connective tissue distribution in an example of mouse PDA. Collagen fibers are staining in blue with the cytoplasm appearing in red (40 magnification). C, immunohistochemistry for SPARC on murine PDA (a-SPARC polyclonal antibody, protein tech catalog number: 15274-1-AP, 10 magnification). D, histochemical staining of hyaluronan (biotinylated hyaluronan binding protein, Calbiochem 385911, 10 magnification) reveals pan-stromal deposition in murine PDA.

Article Snippet: C, immunohistochemistry for SPARC on murine PDA (a-SPARC polyclonal antibody, protein tech catalog number: 15274-1-AP, 10magnification).

Techniques: Staining, Immunohistochemistry, Binding Assay

Journal: Clinical Cancer Research

Article Title: The Pancreas Cancer Microenvironment

doi: 10.1158/1078-0432.ccr-11-3114

Figure Lengend Snippet: Figure 2. Murine PDA is characterized by hypovasularity. In this example of CD31 immunohistochemistry, the dotted line denotes the boundary between tumor (T) and peritumoral diseased pancreas (PT; 20 magnification).

Article Snippet: C, immunohistochemistry for SPARC on murine PDA (a-SPARC polyclonal antibody, protein tech catalog number: 15274-1-AP, 10magnification).

Techniques: Immunohistochemistry

Journal: Cancer Research Communications

Article Title: Alprazolam Reduces Inflammatory Cytokine Production in Pancreatic Cancer–Associated Fibroblasts

doi: 10.1158/2767-9764.CRC-25-0472

Figure Lengend Snippet: ALP alters the inflammatory secretome of PDAC CAFs in vitro . A, Annotated images of cytokine array membranes following incubation with conditioned media from C7-TA-PSC cells treated with 20 μmol/L ALP or DMSO for 24 hours. B, Quantification of the total pixel intensity for cytokines visualized in A . Data points reflect values of each technical duplicate from a single biological replicate ( n = 1). AU, arbitrary units. C, RT-qPCR quantification of the fold change in cytokine RNA expression from C7-TA-PSC cells after treatment with 20 μmol/L of each BZD relative to the DMSO control. Data were collected in technical duplicate and biological triplicate ( n = 3). D, Quantification of cytokine protein levels detected by individual ELISAs using the conditioned media from cells treated as in C . Data were collected in technical duplicate for three biological replicates ( n = 3). Data in B–D are represented as mean ± SD. All statistical analyses in C and D were conducted using one-way ANOVA with Dunnett correction for multiple comparisons. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

Article Snippet: KPC1245 murine PDAC cells were derived from a Kras LSL-G12D/+ ; Trp53 LSL-R172H/+ ; Pdx1-Cre tumor in a female C57BL/6 mouse in the lab of Dr. David Tuveson (Cold Spring Harbor Laboratory, Cold Spring Harbor, NY).

Techniques: In Vitro, Incubation, Quantitative RT-PCR, RNA Expression, Control

Journal: Cancer Research Communications

Article Title: Alprazolam Reduces Inflammatory Cytokine Production in Pancreatic Cancer–Associated Fibroblasts

doi: 10.1158/2767-9764.CRC-25-0472

Figure Lengend Snippet: ALP suppresses TLR4 signaling in PDAC CAFs. A, Heatmap of relative RNA expression levels for TLRs in a scRNA-seq profile of human fibroblasts from normal pancreas and PDAC tumor tissues analyzed in ref. . B, RT-qPCR validation of TLR4 RNA expression in C7-TA-PSC cells at baseline relative to GAPDH . C, Fold change in cytokine RNA expression levels determined by RT-qPCR following 24 hours of treatment with DMSO, 20 μmol/L ALP, and/or 2.5 μg/mL LPS. Experiments in B and C were conducted in technical and biological triplicate ( n = 3). Data are represented as mean ± SD. Statistical analyses in C were conducted using one-way ANOVA with a Tukey correction for multiple comparisons. ns, not significant; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

Article Snippet: KPC1245 murine PDAC cells were derived from a Kras LSL-G12D/+ ; Trp53 LSL-R172H/+ ; Pdx1-Cre tumor in a female C57BL/6 mouse in the lab of Dr. David Tuveson (Cold Spring Harbor Laboratory, Cold Spring Harbor, NY).

Techniques: RNA Expression, Quantitative RT-PCR, Biomarker Discovery

Journal: Cancers

Article Title: Murine Macrophages Modulate Their Inflammatory Profile in Response to Gas Plasma-Inactivated Pancreatic Cancer Cells

doi: 10.3390/cancers13112525

Figure Lengend Snippet: Cytotoxicity and migration in pancreatic cancer cells in vitro. ( a ) Macroscopic image of the atmospheric pressure argon plasma jet kINPen (left) and microscopy image of PDA6606 murine pancreatic cancer cells used in this study (right); ( b ) doubling time of PDA6606 cells in fully supplemented DMEM and RPMI culture medium; ( c ) evaporation of liquid during gas plasma treatment of cultures; ( d ) representative macroscopic image of the resazurin-based assay on cellular metabolism for control and 60 s plasma-treated cells at 24 h; ( e ) metabolic activity of cells following exposure to direct plasma treatment or plasma-treated cell culture medium (in % of values normalized to untreated controls); ( f ) H 2 O 2 generation during plasma treatment of cell culture medium; ( g ) representative brightfield and propidium iodide (PI, orange) images of PDA6606 12 h post-exposure; ( h ) kinetic of relative number of dead cells following plasma treatment; ( i ) scheme of the migration assay; ( j ) representative brightfield images of untreated and plasma-treated cells of the migration assay, the red and blue lines mark the boarders at 0 and 24 h, respectively; ( k ) quantification of absolute cell areas in untreated and plasma-treated cells. Scale bars are 20 µm ( a ), 50 µm ( g ), and 485 µm ( j ). ** = p < 0.01; *** = p < 0.001; NAC = n-acetylcysteine; gas = exposure to argon with plasma-off.

Article Snippet: Murine PDA6606 (Pancreatic Ductal Adenocarcinoma, PDA) cells were kindly provided by David Tuveson (The Cancer Center, Johns Hopkins University, Baltimore, MD, USA).

Techniques: Migration, In Vitro, Clinical Proteomics, Microscopy, Evaporation, Resazurin Assay, Control, Activity Assay, Cell Culture

Journal: Cancers

Article Title: Murine Macrophages Modulate Their Inflammatory Profile in Response to Gas Plasma-Inactivated Pancreatic Cancer Cells

doi: 10.3390/cancers13112525

Figure Lengend Snippet: Co-culture of pancreatic cancer cells with macrophages and their activity. ( a ) Live-cell imaging of DiL-stained PDA6606 (PDA) cells (red), CFSE-labeled RAW 264.7 (RAW) macrophages, and overlay; ( b ) quantification of tumor cell migration over 20 h; ( c ) representative brightfield images of untreated co-cultures of PDA and RAW cells leading to clustering of the latter (arrow) as well as plasma-treated co-cultures showing less clustering at 72 h post-exposure; ( d ) quantification of clusters and comparison to RAW cells alone (M0) and M2-skewed RAW cells per field of view; ( e ) representative brightfield images of granular release from RAW cells (arrow); ( f ) metabolic activity of untreated and plasma-treated PDA cells at the plate bottom with RAW cells co-cultured in a transwell system on top, with a pore size that did not allow for transmigration, metabolic activity was measured of PDA cells alone after removing the transwells harboring the RAW cells at 24 h; ( g ) representative flow cytometry density dot-plot showing the discrimination of RAW cells from PDA cells in co-culture assays using CD11b fluorescently labeled antibodies; ( h ) quantification and normalization of the mean fluorescent intensities (MFI) over several surface markers on RAW cells co-cultured with PDA cells for 24 and 96 h. Scale bars are 25 µm ( a ), 250 µm ( c ), and 20 µm ( e ). Gas = exposure to argon with plasma-off; * = p < 0.05, ** = p < 0.01; *** = p < 0.001.

Article Snippet: Murine PDA6606 (Pancreatic Ductal Adenocarcinoma, PDA) cells were kindly provided by David Tuveson (The Cancer Center, Johns Hopkins University, Baltimore, MD, USA).

Techniques: Co-Culture Assay, Activity Assay, Live Cell Imaging, Staining, Labeling, Migration, Clinical Proteomics, Comparison, Cell Culture, Pore Size, Transmigration Assay, Flow Cytometry

Journal: Cancers

Article Title: Murine Macrophages Modulate Their Inflammatory Profile in Response to Gas Plasma-Inactivated Pancreatic Cancer Cells

doi: 10.3390/cancers13112525

Figure Lengend Snippet: Chemokine and cytokine profiles of co-cultures. ( a ) Mean absolute concentrations of 14 different chemokines and cytokines analyzed in supernatants of untreated and plasma-treated PDA6606 (PDA) monocultures as well as RAW 264.7 (RAW) co-cultures (CC) collected at 24 and 96 h, where arrows indicate significant tendencies and values in boxes represent absolute concentrations in pg/mL; ( b ) principal component analysis of all chemokines and cytokines and reducing to untreated and plasma-treated samples, showing three distinct clusters of similarity for 96 h co-cultures (green), 24 h untreated conditions for the monoculture and co-culture (orange), and all other samples (violet); ( c ) supernatant chemokine and cytokine release normalized to untreated controls (0 s) of PDA and RAW cells co-cultured in transwell systems over 24 h. Arrows indicate non-significant ( ) or significant trend for linearity (↑ or ↓). CCL = CC-chemokine ligand; CXCL = (C-X-C motif) ligand; IL = interleukin; IFN = interferon; MCP = macrophage-chemoattractant protein; TGF = transforming growth factor; TNF = tumor necrosis factor; VEGF = vascular endothelial growth factor.

Article Snippet: Murine PDA6606 (Pancreatic Ductal Adenocarcinoma, PDA) cells were kindly provided by David Tuveson (The Cancer Center, Johns Hopkins University, Baltimore, MD, USA).

Techniques: Clinical Proteomics, Co-Culture Assay, Cell Culture

Journal: Pancreas

Article Title: Ocoxin Oral Solution Exerts an Antitumoral Effect in Pancreatic Cancer and Reduces the Stromal-Mediated Chemoresistance

doi: 10.1097/MPA.0000000000001277

Figure Lengend Snippet: Ocoxin oral solution effect on the viability of the murine pancreatic adenocarcinoma 266-6 cell line. The viability of 266-6 cells was analyzed by means of the Presto Blue assay after 48 hours with different treatment combinations. A, Cell viability after OOS treatment according to 1:1000 to 1:50 (V/Vf) concentrations of (B) paclitaxel from 1 to 10 μM and gemcitabine from 200 to 1000 nM (C) combinations of all 3 of them: paclitaxel 1 μM + OOS 1:50 (V/Vf), gemcitabine 1 μM + OOS 1:50 (V/Vf), and paclitaxel 1 μM + gemcitabine 1 μM + OOS 1:50 (V/Vf). Data are expressed as the mean value (SD) of at least 3 independent experiments. Differences were considered significant for * P < 0.05.

Article Snippet: The murine pancreatic adenocarcinoma 266-6 cell line (ATCC, LGC Standards S.L.U., Barcelona, Spain) was used for in vitro and in vivo experiments.

Techniques: