Journal: Breast cancer research : BCR

Article Title: PTHrP intracrine actions divergently influence breast cancer growth through p27 and LIFR.

doi: 10.1186/s13058-024-01791-z

Figure Lengend Snippet: Fig. 4 PTHrP differentially regulates p27 through LIFR in breast cancer cells. (A) Immunofluorescence staining and quantification of LIFR in primary tumors from mice inoculated with MSCV, FLSEC, DNLS, or DNLS + CTERM cells. All panels = 40X and scale bars = 50 μm. (B) Western blot analysis of p27, pERK, ERK, p-p38, p38 and tubulin (loading control) protein levels in MSCV, FLSEC, DNLS, or DNLS + CTERM cells treated with vehicle (DMSO) or LIFR inhibitor (EC359, 50nM or 100nM) for 24 h. Densitometry for western blot analysis of (C) p27, (D) pERK/ERK and (E) p-p38/p38 described in (B). (A) **p < 0.01 vs. DNLS by unpaired t-test. (C) *p < 0.05 vs. DNLS by unpaired t-test or *p < 0.05 vs. MSCV by one-way ANOVA with multiple comparisons. (D & E) *p < 0.05 vs. MSCV by one-way ANOVA with multiple comparisons or *p < 0.05, **p < 0.01, ***p < 0.001 versus vehicle by two-way ANOVA. Graphs represent mean ± SEM

Article Snippet: The following day, cells were treated with EC359, a leukemia inhibitory factor receptor (LIFR) inhibitor (50nM or 100nM; MedChemExpress; Catalog No. HY-1,201,420) or vehicle (0.1% dimethyl sulfoxide, DMSO) for 1, 6, or 24 h in full-serum media.

Techniques: Immunofluorescence, Staining, Western Blot, Control

Journal: Breast cancer research : BCR

Article Title: PTHrP intracrine actions divergently influence breast cancer growth through p27 and LIFR.

doi: 10.1186/s13058-024-01791-z

Figure Lengend Snippet: Fig. 6 Model of PTHrP domain-specific actions in breast cancer progression and bone colonization. In the primary breast site (top left panel, left of ar rows), PTHrP lacking the NLS and C-terminal domain decreases tumor cell proliferation through p27 induction driven by the tumor suppressor leukemia inhibitory factor receptor (LIFR). PTHrP lacking the NLS and C-terminal domain also preferentially induces p38 phosphorylation and signaling to inhibit cell cycling downstream of LIFR activation. In the breast, truncated PTHrP lacking the NLS alone (top left panel, right of arrows) downregulates LIFR expression (denoted by transparent coloring) and prevents induction of p27 expression and activation of p38 signaling (denoted by dashed arrows, dot ted outlines and transparent coloring) to drive cell proliferation and tumor growth. In bone disseminated tumor cells (bottom panel), LIFR expression is downregulated and the induction of p27 by PTHrP lacking the NLS and C-terminal domain persists, but is not sufficient to repress metastatic outgrowth (denoted by dashed inhibitor line), in contrast to the primary tumor. In the bone, tumor cells expressing PTHrP peptides lacking the NLS or NLS and C- terminal domain readily proliferate into metastatic tumors. Image created with Biorender.com

Article Snippet: The following day, cells were treated with EC359, a leukemia inhibitory factor receptor (LIFR) inhibitor (50nM or 100nM; MedChemExpress; Catalog No. HY-1,201,420) or vehicle (0.1% dimethyl sulfoxide, DMSO) for 1, 6, or 24 h in full-serum media.

Techniques: Phospho-proteomics, Activation Assay, Expressing

Journal: Cancers

Article Title: Gp130-Mediated STAT3 Activation Contributes to the Aggressiveness of Pancreatic Cancer through H19 Long Non-Coding RNA Expression

doi: 10.3390/cancers14092055

Figure Lengend Snippet: Active gp130/STAT3 pathway in PANC-1 sphere cells. ( A ) Real-time qPCR analysis of IL-6 , LIF , IL-6R , LIFR , and gp130 in PANC-1 cells cultured in 2D (adherent) or 3D (sphere) conditions. The results presented are normalized to values obtained for adherent cells (value = 1). Results are presented as means ± SD from three independent experiments. ( B ) FACS analysis of IL-6R, LIFR, and gp130 expression in PANC-1 cells cultured in 2D (adherent) or 3D (sphere) conditions. Controls are indicated by thin lines with gray color. ( C ) Cell surface levels of IL-6R, LIFR, and gp130 expression in PANC-1 cells cultured in 2D (adherent) or 3D (sphere) conditions. Mean fluorescence intensities (MFIs) relative to those of adherent cells are presented. Results are presented as means ± SD from three independent experiments. ( D ) Western blot analysis of p-gp130, gp130, p-STAT3, and STAT3 was performed in PANC-1 cells cultured in 2D (adherent) or 3D (sphere) conditions. Relative band intensity is provided. ** p < 0.01.

Article Snippet: Cells were harvested, and dissociated single cells were incubated on ice for 30 min with PE-conjugated anti-human IL-6 receptor α (IL-6Rα) antibody (BioLegend, San Diego, CA), PE-conjugated anti-human LIF receptor (LIFR) antibody (Becton Dickinson, Franklin Lakes, NJ, USA), PE-conjugated anti-human gp130 antibody (BioLegend), or PE-conjugated isotype control (Becton Dickinson) diluted in FACS buffer (0.5% ( w / v ) BSA and 0.1% ( w / v ) sodium azide in PBS).

Techniques: Cell Culture, Expressing, Fluorescence, Western Blot

Journal: Cancers

Article Title: Gp130-Mediated STAT3 Activation Contributes to the Aggressiveness of Pancreatic Cancer through H19 Long Non-Coding RNA Expression

doi: 10.3390/cancers14092055

Figure Lengend Snippet: The correlation of gp130/STAT3 pathway-related factor expression with overall survival and H19 expression correlated with gp130/STAT3 pathway-related factors in patients with PDAC. Each survival curve was presented according to the online database GEPIA. ( A ) IL-6 , ( B ) LIF , ( C ) IL-6R , ( D ) LIFR , ( E ) gp130 (IL-6ST) , ( F ) JAK1 , ( G ) STAT3 , ( H ) TGF β-RII (TGFBR2) , ( I ) MT1-MMP (MMP14) , and ( J ) H19 . ( K ) The correlation of H19 and gp130/STAT3 pathway-related factor expression in patients with PDAC was assessed using Spearman rank correlation analysis according to the online database GEPIA.

Article Snippet: Cells were harvested, and dissociated single cells were incubated on ice for 30 min with PE-conjugated anti-human IL-6 receptor α (IL-6Rα) antibody (BioLegend, San Diego, CA), PE-conjugated anti-human LIF receptor (LIFR) antibody (Becton Dickinson, Franklin Lakes, NJ, USA), PE-conjugated anti-human gp130 antibody (BioLegend), or PE-conjugated isotype control (Becton Dickinson) diluted in FACS buffer (0.5% ( w / v ) BSA and 0.1% ( w / v ) sodium azide in PBS).

Techniques: Expressing

Journal: Cancers

Article Title: Gp130-Mediated STAT3 Activation Contributes to the Aggressiveness of Pancreatic Cancer through H19 Long Non-Coding RNA Expression

doi: 10.3390/cancers14092055

Figure Lengend Snippet: Schematic representation of autocrine/paracrine IL-6 or the LIF/gp130/STAT3 pathway in PDAC sphere cells. In PDAC sphere cells in which CSCs are enriched (CSC-like cells), binding of autocrine/paracrine IL-6 or LIF to each receptor (IL-6R or LIFR, respectively) induces gp130 homodimerization or LIFR/gp130 complex formation to thereby activating JAKs followed by phosphorylation of gp130, ultimately leading to STAT3 activation. This pathway contributes to the maintenance of stemness features and expression of MT1-MMP and TGFβ-RII. Expression of TGFβ-RII via the gp130/STAT3 pathway affects TGF-β1/Smad signaling to promote EMT induction. Additionally, p-STAT3 can access the active promoter region of H19 and contribute to its transcription. Therefore, autocrine/paracrine IL-6 or the LIF/gp130/STAT3 pathway in PDAC CSC-like cells is believed to eventually lead to invasion and metastasis, both of which are hallmarks of malignancy.

Article Snippet: Cells were harvested, and dissociated single cells were incubated on ice for 30 min with PE-conjugated anti-human IL-6 receptor α (IL-6Rα) antibody (BioLegend, San Diego, CA), PE-conjugated anti-human LIF receptor (LIFR) antibody (Becton Dickinson, Franklin Lakes, NJ, USA), PE-conjugated anti-human gp130 antibody (BioLegend), or PE-conjugated isotype control (Becton Dickinson) diluted in FACS buffer (0.5% ( w / v ) BSA and 0.1% ( w / v ) sodium azide in PBS).

Techniques: Binding Assay, Phospho-proteomics, Activation Assay, Expressing