Journal: Breast Cancer Research : BCR

Article Title: OSM potentiates preintravasation events, increases CTC counts, and promotes breast cancer metastasis to the lung

doi: 10.1186/s13058-018-0971-5

Figure Lengend Snippet: Oncostatin M (OSM) is highly expressed in ductal carcinoma in situ (DCIS) and invasive ductal carcinoma (IDC). a To detect the presence of OSM in breast cancer tissue, histological microarrays from 72 patients with breast cancer were stained with human OSM antibody by IHC. Twelve patients had in situ DCIS, 54 patients had nonmetastatic IDC, and 16 patients had IDC with metastasis to lymph nodes (see Additional file ). The results showed that normal adjacent tissue expresses little OSM but that OSM is highly expressed in DCIS and IDC. Secondary antibody alone did not produce any background signals. b Intensity quantification of OSM stained tissues. Mean staining intensity for DCIS (2.00) and IDC (1.66) tissues was significantly higher than that of normal adjacent tissue (1.33) and metastatic tissue (1.24). There was no statistically significant difference between normal and metastatic tissue. Multiple cores from the same patients were averaged. Data are expressed as mean ± SD. * p < 0.05 by one-way analysis of variance with Tukey’s multiple comparisons test

Article Snippet: The serum was then diluted 1:3 in PBS and used in an hOSM ELISA (catalogue number DY295; R&D Systems, Minneapolis MN, USA), which was performed in accordance with the manufacturer’s instructions.

Techniques: In Situ, Staining

Journal: Breast Cancer Research : BCR

Article Title: OSM potentiates preintravasation events, increases CTC counts, and promotes breast cancer metastasis to the lung

doi: 10.1186/s13058-018-0971-5

Figure Lengend Snippet: Oncostatin M (OSM) produced by tetracycline (TET)-inducible MDA-MB-231 (MDA TO/OSM ) cell MDA TO/OSM ) tumors increase metastasis and decrease survival. a MDA TO/OSM human breast cancer cells were treated with (+TET) or without TET (−TET), and the resultant conditioned media (CM) from the treated cells were applied to parental MDA-MB-231, MDA-MB-231-LUC, and T47D cells. Left : The activity of OSM accumulated in the CM was compared with commercially obtained recombinant human OSM (rhOSM) (25 ng/ml). There was no significant difference between OSM produced by MDA TO/OSM versus rhOSM versus its ability to induce pSTAT3. Middle : Western blot analysis depicting that CM produced by MDA TO/OSM cells stimulated with TET contain OSM. Right : Enzyme-linked immunosorbent assay (ELISA) analysis showed that CM from TET-treated MDA TO/OSM cells contain 10.1 ng/ml of hOSM. b Left : Animals with MDA TO/OSM tumors were given drinking water with or without TET, and whole blood was collected at the experimental endpoint. After allowing the blood to clot and serum was separated by centrifugation, the resultant serum OSM levels were measured by ELISA. Animals with MDA TO/OSM tumors with drinking water containing TET had 67-fold higher serum OSM levels. Center : Platelet counts were higher in +TET MDA TO/OSM tumor-bearing mice than in −TET mice. Right : +TET MDA TO/OSM tumor-bearing mice had lower body weight than −TET mice. c Animals with MDA TO/OSM tumors were given drinking water containing TET for 1 week, and their lung metastasis levels were assessed by ex vivo bioluminescence imaging. Left : Representative ex vivo bioluminescence image. Right : Average radiance analysis of the ex vivo bioluminescence imaging in photons per second per square centimeter per square radian (p/s/cm 2 /sr). Animals with MDA TO/OSM tumors +TET had a fivefold higher bioluminescent radiance than −TET mice (−TET, n = 3; +TET, n = 6). Data are expressed as mean ± SEM. d Kaplan-Meier survival curve for mice with MDA TO/OSM tumors ± TET. Mice that did not receive TET survived, on average, 11 days longer (−TET, n = 9; +TET, n = 10). *** p < 0.001 by log-rank test. Data are expressed as mean ± SEM. * p < 0.05, ** p < 0.01, and *** p < 0.001 by two-tailed t test or one-way analysis of variance with Tukey’s posttest where appropriate

Article Snippet: The serum was then diluted 1:3 in PBS and used in an hOSM ELISA (catalogue number DY295; R&D Systems, Minneapolis MN, USA), which was performed in accordance with the manufacturer’s instructions.

Techniques: Produced, Activity Assay, Recombinant, Western Blot, Enzyme-linked Immunosorbent Assay, Centrifugation, Ex Vivo, Imaging, Two Tailed Test

Journal: Cell Death & Disease

Article Title: Blocking STAT3 signaling augments MEK/ERK inhibitor efficacy in esophageal squamous cell carcinoma

doi: 10.1038/s41419-022-04941-3

Figure Lengend Snippet: A Western blotting shows the expression of pSTAT3 and pERK in KYSE30 WT, ERK1/2-DKO-1, and ERK1/2-DKO-2 after EGF (50 ng/ml), OSM (10 ng/ml), or IL-6 (10 ng/ml) treatment. B KYSE150 WT and ERK1/2-DKO cells were treated with EGF (50 ng/ml), OSM (10 ng/ml), or IL-6 (10 ng/ml) for 1 h. Expressions of pSTAT3 and pERK were determined using western blotting. C RNA-seq heat map analyses of differentially expressed genes (DEGs) involved in the JAK-STAT3 signaling pathway after trametinib or DMSO treatment. A fold change cutoff of log2 < −0.58 or >0.58 and a p value cutoff of P < 0.05 were selected and overlapped for JAK-STAT3 signaling pathway genes. D qRT-PCR results show the transcription level of SOCS3 mRNA after trametinib treatment at various time points. Error bars represent the mean ± SD. ** P < 0.01, *** P < 0.001. E KYSE30 and KYSE150 cells were incubated with trametinib (1 μM) for 0, 1, 2, 4 h, immunoblotted for SOCS3 expression. F , G Western blotting results show the expression of SOCS3 after EGF (50 ng/ml) treatment in WT, ERK-DKO KYSE30 ( F ) and KYSE150 ( G ) cells. H KYSE30 ERK1/2-DKO cells stably expressing EV, ERK2-HA, and ERK2-K54R-HA were used for western blotting to determine SOCS3 expression. I Western blotting results show the expression of pSTAT3 and Flag-SOCS3 in KYSE30 and KYSE150 cells transduced with EV or Flag-SOCS3. Trametinib (1 μM) treatment was applied at various time points. J Western blotting results show the expression of pSTAT3 and pERK1/2 in KYSE30 and KYSE150 cells infected with shNC or shSOCS3 lentiviral with two target sites.

Article Snippet: OSM (10452-HNAH) and human recombinant IL-6 (10395-HNAE) were purchased from Sino Biological Inc. (Beijing, China).

Techniques: Western Blot, Expressing, RNA Sequencing Assay, Quantitative RT-PCR, Incubation, Stable Transfection, Transduction, Infection