13 research outputs found

    Oleic acid (OA) promotes the proliferation and migration ability of MCF10DCIS.COM cells but not SUM225 cells, whereas palmitic acid (PA) leads to cell death in both cells.

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    <p>(A and B) MTT assay of cell proliferation in MCF10DCIS.COM and SUM225 cells incubated with increasing OA or PA. OA induced significantly increased viability in MCF10DCIS.COM cells but led to cell death in SUM225 cells. PA induced the death of both MCF10DCIS.COM and SUM225 cells. (C) Trans-well assay of cell migration in MCF10DCIS.COM and SUM 225 cells. OA significantly promoted the migration of MCF10DCIS.COM cell but not SUM225 cells. (D) Wound healing assay of lateral migration of MCF10DCIS.COM cells incubated with OA. OA significantly enhanced the lateral migration of MCF10DCIS.COM cells. All the experiments were performed at least in triplicate and the values are reported as the means ± standard error. *<i>p</i><0.05, **<i>p</i><0.01.</p

    Oleic acid (OA) promotes the viability and migration through the FAK, PI3K/AKT, and MEK/ERK signaling pathway in MCF10DCIS.COM cells.

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    <p>(A and B) Representative Western blot and quantitative analysis of phosphorylated FAK, AKT and ERK1/2 in MCF10DCIS.COM cells incubated with OA. OA induced a significant increase in the phosphorylation of FAK, AKT and ERK1/2. (C) MTT assay of cell proliferation in MCF10DCIS.COM cells incubated with OA in the presence of FAK (PF573228), PI3K/AKT (LY294002) and MEK/ERK (PD98059) inhibitors. All kinase inhibitors induced cell death, and OA-promoted proliferation was reduced in the presence of all kinase inhibitors. (D) Trans-well assay of cell migration in MCF10DCIS.COM cells incubated with OA in the presence of FAK, PI3K/AKT and MEK/ERK inhibitors. OA-induced migration was suppressed by the presence of all kinase inhibitors. All the experiments were performed at least in triplicate, and the values are reported as the means ± standard error. *<i>p</i><0.05, **<i>p</i><0.01.</p

    Different Biological Action of Oleic Acid in ALDH<sup>high</sup> and ALDH<sup>low</sup> Subpopulations Separated from Ductal Carcinoma <i>In Situ</i> of Breast Cancer

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    <div><p>The mechanisms underlying breast cancer progression of ductal carcinoma in situ <b>(</b>DCIS) associated with fatty acids are largely unknown. In the present study, we compared the action of oleic acid (OA) on two human DCIS cell lines, MCF10DCIS.COM (ER/PR/HER2-negative) and SUM225 (HER2 overexpressed). OA led to a significant increase in proliferation, migration, lipid accumulation and the expression of lipogenic proteins, such as SREBP-1, FAS and ACC-1, in MCF10DCIS.COM cells but not SUM225 cells. The ALDH<sup>high</sup> subpopulation analyzed by the ALDEFLUOR assay was approximately 39.2±5.3% of MCF10DCIS.COM cells but was small (3.11±0.9%) in SUM225 cells. We further investigated the different biological action of OA in the distinct ALDH<sup>low</sup> and ALDH<sup>high</sup> subpopulations of MCF10DCIS.COM cells. OA led to an increase in the expression of ALDH1A1, ALDH1A2 and ALDH1A3 in MCF10DCIS.COM cells. SREBP-1 and ACC-1 were highly expressed in ALDH<sup>high</sup> cells relative to ALDH<sup>low</sup> cells, whereas FAS was higher in ALDH<sup>low</sup> cells. In the presence of OA, ALDH<sup>high</sup> cells were more likely to proliferate and migrate and displayed significantly high levels of SREBP-1 and FAS and strong phosphorylation of FAK and AKT relative to ALDH<sup>low</sup> cells. This study suggests that OA could be a critical risk factor to promote the proliferation and migration of ALDH<sup>high</sup> cells in DCIS, leading to breast cancer progression.</p></div

    Oleic acid (OA) further promotes the proliferation and migration abilities and upregulates lipogenic proteins in ALDH<sup>high</sup> cells.

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    <p>(A) Quantitative real-time RT-PCR of ALDH1A1, ALDH1A2 and ALDH1A3 in MCF10DCIS.COM cells. All subtypes of ALDH1 were significantly increased by OA. Notably, OA led to a remarkable increase in ALDH1A2 in MCF10DCIS.COM cells. (B) MTT assay of cell proliferation in ALDH<sup>high</sup> and ALDH<sup>low</sup> cells incubated with OA. OA-induced proliferation of ALDH<sup>high</sup> cells was greater than that of ALDH<sup>low</sup> cells. (C) Trans-well assay of cell migration in ALDH<sup>high</sup> and ALDH<sup>low</sup> cells. The OA-induced migration ability was higher in ALDH<sup>high</sup> cells than ALDH<sup>low</sup> cells. (D) Representative Western blot of SREBP-1, FAS and ACC-1 in ALDH<sup>high</sup> cells and ALDH<sup>low</sup> cells. (E, F and G) Analysis of expression levels of SREBP-1, FAS and ACC-1. Significantly higher expression of SERBP-1 and ACC-1 was observed in ALDH<sup>high</sup> cells, whereas FAS was significantly higher in ALDH<sup>low</sup> cells. OA led to the significant upregulation of SREBP-1 and FAS in ALDH<sup>high</sup> cells and the significant upregulation of SREBP-1 and downregulation of FAS in ALDH<sup>low</sup> cells. All experiments were performed at least in triplicate, and the values are reported as the means ± standard error. *<i>p</i><0.05, **<i>p</i><0.01.</p

    Oleic acid (OA) induces lipid accumulation and the upregulation of lipogenic proteins in MCF10DCIS.COM cells but not SUM225 cells.

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    <p>(A) Representative Oil Red O staining in MCF10DCIS.COM and SUM 225 cells incubated with OA. A large number of lipid droplets were observed in MCF10DCIS.COM cells but not SUM225 cells. (B) Quantitative analysis of intracellular lipid contents from Oil Red O staining. OA led to lipid accumulation in MCF10DCIS.COM cells. (C) Representative Western blot of SREBP-1, FAS and ACC-1 in MCF10DCIS.COM and SUM 225 cells incubated with OA. (D) Quantitative analysis of lipogenic protein levels in MCF10DCIS.COM and SUM225 cells. A significantly higher level of SERBP-1 was observed in MCF10DCIS.COM cells relative to SUM225 cells. The FAS level was significantly higher in SUM225 cells than MCF10DCIS.COM cells. The level of ACC-1 was similar between MCF10DCIS.COM cells and SUM225 cells (upper). OA resulted in the significant upregulation of SREBP-1, FAS and ACC-1 in MCF10DCIS.COM cells (middle) but led to the downregulation of SREBP-1 and the upregulation of ACC-1 significantly in SUM225 cells (lower). All experiments were performed at least in triplicate, and the values are reported as the means ± standard error. *<i>p</i><0.05, **<i>p</i><0.01.</p

    Distinct subpopulations of ALDH1<sup>high</sup> and ALDH1<sup>low</sup> cells were separated from MCF10DCIS.COM cells.

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    <p>(A) Representative flow cytometry for ALDEFLUOR assay showing the percentage of ALDH<sup>high</sup> cells in MCD10DCIS.COM and SUM225 cells. Graph showed that the ALDH<sup>high</sup> cell population obtained from experiments performed at least in triplicate. Cells exhibiting high ALDH activity were higher in MCF10DCIS.COM cells relative to SUM225 cells. (B) Quantitative real-time RT-PCR of ALDH1A1, ALDH1A2 and ALDH1A3 in ALDH1<sup>high</sup> and ALDH1<sup>low</sup> subpopulation cells separated from MCF10DCIS.COM cells. Significantly higher expression levels of ALDH1A2 and ALDH1A3 mRNAs were detected in ALDH<sup>high</sup> cells relative to ALDH<sup>low</sup> cells. The experiments were performed at least in triplicate, and the values are reported as the means ± standard error. *<i>p</i><0.05, **<i>p</i><0.01. (C) RT-PCR analysis of CD24 and CD44 mRNAs in ALDH<sup>high</sup> and ALDH<sup>low</sup> cells. CD44 mRNA was higher in ALDH1<sup>high</sup> cells than ALDH1<sup>low</sup> cells. (D) Flow cytometric analysis of CD44 and CD24. Of MCF10DCIS.COM cells, 70% exhibited the CD44+/CD24- phenotype. CD44+/CD24- cell populations were higher in separated ALDH1<sup>high</sup> cells than ALDH1<sup>low</sup> cells. (E) Immunofluorescence staining of CD44, CD24 and ALDH1. ALDH1<sup>high</sup> cells expressed a high level of ALDH1 and CD44, whereas ALDH1<sup>low</sup> cells displayed a high level of CD24 and a low level of ALDH1 and CD44.</p

    Ferritin-based MRI of dendritic cell

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    Figure 1. Analysis of dendritic cell (DC) transduced with myc-tagged human ferritin heavy chain (FTH) and green fluorescence prtein (GFP) using lentivirus. Figure 2. Analyses of proliferation and migration activities and co-stimulatory molecules expressions in dendritic cell (DC) and human ferritin heavy chain-transduced DC (FTH-DC). Figure. 3. Cellular iron staining, iron amount measurement and in vitro MRI analysis of dendritic cell (DC) and human ferritin heavy chain-transduced DC (FTH-DC). Figure 4. in vivo and ex vivo MRI of popliteal lymph nodes (LNs) of mouse injected with dendritic cell (DC) and ferritin heavy chain-transduced DC (FTH-DC). Figure 5. Histological analysis of cryosectioned popliteal lymph nodes (LNs) with dendritic cell (DC) and ferritin heavy chain-transduced DC (FTH-DC)

    Analyses of proliferation and migration activities and co-stimulatory molecules expressions in dendritic cell (DC) and human ferritin heavy chain-transduced DC (FTH-DC).

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    <p>(A) A standard 3-,5-diphenyltetrazolium bromide (MTT) assay for proliferation activity of DCs and FTH-DCs cultured for 24 h, 48 h and 72 h. (B and C) Trans-well assay for migration abilities of DCs and FTH-DCs incubated with TNF-α (20 ng/mL) and IFN-γ (20 ng/mL) in the presence or absence of CCL19 and CCL21 for 24 h. Representative fluorescent image of nuclear stained with diamidino-2-phenylindole (DAPI) in DCs and FTH-DCs that migrated to the lower chamber. (D) RT-PCR analysis of C-C chemokine receptor type-7 (CCR-7) in DCs and FTH-DCs. Both DCs and FTH-DCs, which were incubated in the medium supplemented with TNF-α (20 ng/mL) and IFN-γ (20 ng/mL) for 24 h, highly expressed the CCR-7 as compared to untreated cells. (E and F) Representative flow cytometric analysis of co-stimulatory molecules such as CD40, CD80 and CD86 in DCs and FTH-DCs treated with or without LPS (100 ng/mL) for 24 h. Flow cytometric results obtained from 3 independent experiments. All data are presented as the mean ± standard deviations of at least three independent experiments. *, <i>p</i> ≤0.05.</p

    Histological analysis of cryosectioned popliteal lymph nodes (LNs) with dendritic cell (DC) and ferritin heavy chain-transduced DC (FTH-DC).

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    <p>(A) Representative GFP fluorescence images of popliteal LNs isolated from mouse at 48 h after injection of DCs and FTH-DCs. (B) Representative hematoxylin and eosin staining of cryosectioned popliteal LNs with DCs and FTH-DCs. (C) Immunofluorescence images of GFP (green) and FTH (red) detected with anti-GFP and anti-myc antibodies in LNs with DCs and FTH-DCs. (D) Merge images (yellow) of co-immunofluorescence staining of activation marker, CD25 (red) and GFP (green) detected with anti-CD25 and anti-GFP antibodies in FTH-DCs of cryosectioned LNs. Nuclear was stained with diamidino-2-phenylindole (DAPI, blue).</p

    Analysis of dendritic cell (DC) transduced with myc-tagged human ferritin heavy chain (FTH) and green fluorescence prtein (GFP) using lentivirus.

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    <p>(A) Representative GFP fluorescence image (green) and FTH immunofluorescence image (red) detected with anti-myc antibody in DCs and FTH-DCs. (B) Representative Western blots for FTH from whole cell lysate of DCs and FTH-DCs detected by using anti-myc antibody.</p
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