Journal: JCI Insight

Article Title: Loss of TRPV4 reduces pancreatic cancer growth and metastasis

doi: 10.1172/jci.insight.196280

Figure Lengend Snippet: ( A ) Graphical illustration of experimental design. KPC cells (100,000 cells) were injected into the pancreas, followed by injection of KPCY cells (500,000 cells) into the spleen 10 days later. ( B ) Representative images of immunostaining of mouse liver for Ki67 and GFP (marking KCPY cells) at low (upper panels) and high magnification (lower panels). Scale bars: 100 μm (top) and 50 μm (bottom). ( C and D ) Quantitative analysis of Ki67 positively stained cells and liver lesions of WT and TRPV4-KO liver. The analysis was performed on large sections of pancreatic tissues using tiling and stitching of images obtained with a Zeiss 880 Airyscan confocal microscope. Lesions were defined as an aggregate of at least 10 KPCY cells. ( E and F ) Representative images of immunostaining of liver sections with CD68 and quantitative analysis of CD68 + cells in liver sections from WT or TRPV4-KO mice. Scale bars: 100 μm. Statistical analyses were performed using Student’s t test. Animal number, n = 4–5. Results were expressed as mean ± SEM. * P ≤ 0.05; ** P ≤ 0.01.

Article Snippet: KPC cells were purchased from Cancertools (catalog 153474).

Techniques: Injection, Immunostaining, Staining, Microscopy

Journal: Cancer Science

Article Title: Downregulation of 15‐hydroxyprostaglandin dehydrogenase by interleukin‐1β from activated macrophages leads to poor prognosis in pancreatic cancer

doi: 10.1111/cas.13467

Figure Lengend Snippet: Low 15‐hydroxyprostaglandin dehydrogenase (15‐ PGDH ) expression is implicated in poor pancreatic ductal adenocarcinoma ( PDAC ) prognosis. A, Workflow diagram of patients who underwent pancreatic resection and contributed samples for immunohistochemical ( IHC ) analysis. B, Representative IHC staining of 15‐ PGDH expression in 107 PDAC tissues. Scale bar = 100 μm. C,D, Relationship between 15‐ PGDH expression and relapse‐free survival (C) or overall survival (D) using the Kaplan‐Meier method

Article Snippet: The human PDAC cell lines PK‐8 and S2‐013 were obtained from the Japanese Collection of Research Bioresource Cell Bank (Ibaraki, Japan) and RIKEN Bioresource Center Cell Bank (Tsukuba, Japan).

Techniques: Expressing, Immunohistochemical staining, Immunohistochemistry

Journal: Cancer Science

Article Title: Downregulation of 15‐hydroxyprostaglandin dehydrogenase by interleukin‐1β from activated macrophages leads to poor prognosis in pancreatic cancer

doi: 10.1111/cas.13467

Figure Lengend Snippet: 15‐Hydroxyprostaglandin dehydrogenase (15‐ PGDH ) downregulation by interleukin‐1β ( IL ‐1β) enhances pancreatic ductal adenocarcinoma cell growth. A,B, Expression of HPGD (the gene coding 15‐ PGDH protein, upper panel) or 15‐ PGDH (lower panel) in PK ‐8 cells (A) or S2‐013 cells (B) after treatment with si RNA targeting 15‐ PGDH or with control si RNA , evaluated by quantitative RT ‐ PCR (upper panel) or Western blot analysis (lower panel). Data are presented as the treated/control cell ratio. C,D, PK ‐8 cells (C) or S2‐013 cells (D) transfected with si RNA s targeting 15‐ PGDH or with control si RNA were incubated for up to 96 hours and assayed for cell number; data are presented as the treated/control (time = 0) cell ratio. E,F, Expression of 15‐ PGDH in PK ‐8 cells or S2‐013 cells after IL ‐1β (E) or tumor necrosis factor‐α ( TNF ‐α) (F) treatment for 24 and 48 hours and distilled water treatment for 48 hours as a control was evaluated by Western blotting. G, Column graph showing relative 15‐ PGDH levels in PK ‐8 cells or S2‐013 cells after IL ‐1β and TNF ‐α treatment for 24 and 48 hours, and distilled water treatment for 48 hours as a control, were evaluated using ImageJ software. H, Expression of HPGD and IL 1B in six PDAC patients determined by quantitative RT ‐ PCR . Data were normalized to the ACTB mRNA level and are shown as the mean ± SD of three independent experiments. **P < .01

Article Snippet: The human PDAC cell lines PK‐8 and S2‐013 were obtained from the Japanese Collection of Research Bioresource Cell Bank (Ibaraki, Japan) and RIKEN Bioresource Center Cell Bank (Tsukuba, Japan).

Techniques: Expressing, Control, Quantitative RT-PCR, Western Blot, Transfection, Incubation, Software

Journal: Cancer Science

Article Title: Downregulation of 15‐hydroxyprostaglandin dehydrogenase by interleukin‐1β from activated macrophages leads to poor prognosis in pancreatic cancer

doi: 10.1111/cas.13467

Figure Lengend Snippet: Tumor‐associated macrophages are inversely correlated with pancreatic ductal adenocarcinoma ( PDAC ) cells harboring high 15‐hydroxyprostaglandin dehydrogenase (15‐ PGDH ) expression. A,B, Representative immunohistochemical ( IHC ) staining of 15‐ PGDH (upper panel) and CD 163 (lower panel) expression in high 15‐ PGDH (A) and low 15‐ PGDH (B) serial PDAC specimens. Scale bar = 200 μm. C, Graph showing Pearson's correlation between the expression of 15‐ PGDH and the number of CD 163‐positive cells in 107 PDAC patients. D, Schematic representation of the findings of this study. IL ‐1βR, interleukin‐1β receptor

Article Snippet: The human PDAC cell lines PK‐8 and S2‐013 were obtained from the Japanese Collection of Research Bioresource Cell Bank (Ibaraki, Japan) and RIKEN Bioresource Center Cell Bank (Tsukuba, Japan).

Techniques: Expressing, Immunohistochemical staining, Immunohistochemistry

Journal: OncoTargets and therapy

Article Title: Targeting Endoglin Expressing Cells in the Tumor Microenvironment Does Not Inhibit Tumor Growth in a Pancreatic Cancer Mouse Model

doi: 10.2147/OTT.S322276

Figure Lengend Snippet: Endoglin is highly expressed on CAFs in human pancreatic tumors. ( A ) Representative images of human pancreatic cancer (representative from n = 25 PDAC patients) and normal pancreas stained for α-SMA, endoglin, cytokeratin, and vimentin. Endothelial cells (black arrow) and endoglin expressing CAFs (white arrow). ( B ) Immunofluorescent double staining for α-SMA and endoglin in human PDAC tumors. ( C ) Endoglin mRNA expression by human cells; ECRF endothelial cells, MIA PaCa-2, PANC-1 and BxPC-3 PDAC cells and 8 patient derived primary pancreatic CAFs. ( D ) Endoglin protein expression on human pancreatic fibroblasts. Basal and BMP9-induced downstream signaling (pSMAD1) was inhibited with TRC105 (full-length blot shown in Supplementary figure 4A – C ).

Article Snippet: The mouse PDAC cell line KPC-3 (Kras G12D/+ LSL-Trp53 R172H/+ Pdx-1-Cre), (kindly supplied by the department of Immunology, LUMC) with a targeted insertion of codon-optimized Luc-2 (pGL4.10) [luc2] (Promega Leiden, the Netherlands), mouse MC38 cells (kindly supplied by the department of Immunology, LUMC) and primary fibroblasts were all cultured in DMEM/F12 glutamax medium (Invitrogen, Landsmeer, the Netherlands), with 10% fetal bovine serum (FBS) (Gibco, Bleiswijk, the Netherlands), 0.01 M HEPES, 0.1 μg/mL Gentamycin, 40U/mL Penicillin and 40 μg/mL Streptomycin (all Invitrogen Landsmeer, the Netherlands) at 37°C and 5% CO2.

Techniques: Staining, Expressing, Double Staining, Derivative Assay

Journal: PLoS ONE

Article Title: IL-1α Expression in Pancreatic Ductal Adenocarcinoma Affects the Tumor Cell Migration and Is Regulated by the p38MAPK Signaling Pathway

doi: 10.1371/journal.pone.0070874

Figure Lengend Snippet: IL-1α positive PC013 and IL-1α negative PC077 cells were starved in 1%FBS overnight and seeded on Transwell filters pre coated with 10 µg/ml fibronectin. PC013 cells were incubated for 40 h in PC013/vehicle/CAF and PC013/SB203580/CAF supernatants (A), and 100 ng/ml IL-1RA (B). IL-1α negative PC077 cells were cultured for 72 h in 1% FBS medium containing mock, 500 pg/ml IL-1α, 100 ng/ml IL-1RA alone, 100 ng/ml IL-1RA together with either 500 pg/ml IL-1α, CAF supernatant (7 days), or IL-1α (500 pg/ml) activated CAF supernatant (7 days) (C). The adherent cells were fixed in 4% formaldehyde, visualized using crystal violet and quantified manually using 10× magnification in an inverted microscope. The result is presented as cells/cm 2 . * = P<0.05, ** = P<0.005, and *** = P<0.001.

Article Snippet: The primary PDAC cell lines PC013 and PC077, and primary CAFs were cultured in RPMI 1640 (Fisher Scientific, Pittsburgh, PA), supplemented with 1% FCS (Invitrogen), 2 mM HEPES (Invitrogen), 30 μg/ml Gentamycin (Invitrogen) (R10).

Techniques: Incubation, Cell Culture, Inverted Microscopy

Journal: Oncotarget

Article Title: Perspectives of TGF-β inhibition in pancreatic and hepatocellular carcinomas

doi:

Figure Lengend Snippet: At the cellular level, TGF-β induces proliferation and survival of PDAC cells in the late phase of PDAC carcinogenesis (after SMAD4 inactivation), and promotes epithelial-to-mesenchymal transition (EMT), invasion, and metastasis. At the microenvironment level, TGF-β is a key mediator of the dialogue between cancer and stellate cells (fibrotic cells), involved in the production of a dense fibrotic stroma and the resulting low vascularization of PDAC. TGF-β also deregulates the immune microenvironment toward immunosuppression and inappropriate inflammation.

Article Snippet: Consistent with this, Bachem et al.[ ] showed that PDAC cell lines stimulated PSC proliferation.

Techniques: