Journal: Breast Cancer : Targets and Therapy

Article Title: Identification of genes involved in breast cancer and breast cancer stem cells

doi: 10.2147/BCTT.S85202

Figure Lengend Snippet: Gene expression among different cancer types. Notes: Relative gene expression analysis of nine genes that were overexpressed in breast cancer and breast CSCs. The gene expression also was studied in colon and lung cancer, colon CSCs, lung CSCs, embryonic stem cells, and in a reference sample. The experiments were performed in triplicate and a P -value of <0.05 was considered significant. Abbreviations: CSCs, cancer stem cells; ESCs, embryonic stem cells; HP , haptoglobin; SMPD1 , sphingomyelin phosphodiesterase 1, acid lysosomal; SCRIB , scribbled homolog ( Drosophila ); KCMF1 , potassium channel modulatory factor 1; FAM155B , family with sequence similarity 155, member B; PTGER3 , prostaglandin E receptor 3 (subtype EP3); GPR3 , G protein-coupled receptor 3; TMX2 , thioredoxin-related transmembrane protein 2; DDX49 , DEAD (Asp-Glu-Ala-Asp) box polypeptide 49.

Article Snippet: During the exponential phase of proliferation, commercial breast CSCs cells were plated in 24-well plates (E36102-29-24Well; Celprogen) and transfected with gene-specific small interfering RNAs (siRNAs) using Lipofectamine 2000 Reagent (11668-027; Thermo Fisher Scientific), according to the manufacturer’s instructions.

Techniques: Expressing, Sequencing

Journal: Breast Cancer : Targets and Therapy

Article Title: Identification of genes involved in breast cancer and breast cancer stem cells

doi: 10.2147/BCTT.S85202

Figure Lengend Snippet: Breast CSCs pre- and post-siRNA knockdown. Note: Representative images showing breast CSCs pre- and post-siRNA knockdown. Abbreviations: CSCs, cancer stem cells; siRNA, small interfering RNA; TMX2 , thioredoxin-related transmembrane protein 2; FAM155B , family with sequence similarity 155, member B; PTGER3 , prostaglandin E receptor 3 (subtype EP3); GPR3 , G protein-coupled receptor 3; DDX49 , DEAD (Asp-Glu-Ala-Asp) box polypeptide 49.

Article Snippet: During the exponential phase of proliferation, commercial breast CSCs cells were plated in 24-well plates (E36102-29-24Well; Celprogen) and transfected with gene-specific small interfering RNAs (siRNAs) using Lipofectamine 2000 Reagent (11668-027; Thermo Fisher Scientific), according to the manufacturer’s instructions.

Techniques: Small Interfering RNA, Sequencing

Journal: Breast Cancer : Targets and Therapy

Article Title: Identification of genes involved in breast cancer and breast cancer stem cells

doi: 10.2147/BCTT.S85202

Figure Lengend Snippet: Gene expression of stemness transcription factors in breast CSCs. Notes: Relative gene expression of transcription factors in breast CSCs following knockdown. The ∆∆Ct method was used to perform the analysis. Each bar represents the average of the Ct values. The assays were performed in triplicate and a P -value of <0.05 was considered to be significant. The assays are presented in a log2 scale. Thus, positive values indicate overexpression while negative values indicate underexpression. Abbreviations: CSCs, cancer stem cells; TMX2 , thioredoxin-related transmembrane protein 2; FAM155B , family with sequence similarity 155, member B; PTGER3 , prostaglandin E receptor 3 (subtype EP3); GPR3 , G protein-coupled receptor 3; DDX49 , DEAD (Asp-Glu-Ala-Asp) box polypeptide 49; NANOG , Homeobox protein NANOG; OCT3/4 , Octamer-binding transcription factor ¾; SOX2 , sex determining region Y-box 2; CD34 , Hematopoietic progenitor cell antigen CD34.

Article Snippet: During the exponential phase of proliferation, commercial breast CSCs cells were plated in 24-well plates (E36102-29-24Well; Celprogen) and transfected with gene-specific small interfering RNAs (siRNAs) using Lipofectamine 2000 Reagent (11668-027; Thermo Fisher Scientific), according to the manufacturer’s instructions.

Techniques: Expressing, Over Expression, Sequencing, Binding Assay

Journal: Oncology reports

Article Title: Radioresistant breast cancer cells exhibit increased resistance to chemotherapy and enhanced invasive properties due to cancer stem cells.

doi: 10.3892/or.2018.6714

Figure Lengend Snippet: Figure 2. RT‑R‑MDA‑MB‑231 cells demonstrate higher levels of CSCs marker CD44, but lower levels of CD24 compared with MDA‑MB‑231 cells. Classical CSCs markers CD44 and CD24 were detected in the low metastatic breast cancer cells MCF‑7 and RT‑R‑MCF‑7 (A) and highly metastatic breast cancer cells MDA‑MB‑231 and RT‑R‑MDA‑MB‑231 (B) using the specific antibodies by western blotting. Data represent mean values ± standard error of the mean of three independent experiments in triplicate. **P<0.01 compared with control. CD, cluster of differentiation; CSCs, cancer stem cells; RT‑R, radioresistant.

Article Snippet: Isolation of CD24low/CD44high breast cancer stem cells was performed using a MagCollect CD24- CD44+ Breast Cancer Stem Cell Isolation kit (R&D Systems, Inc., Minneapolis, MN, USA) following the manufacturer's protocol.

Techniques: Marker, Western Blot, Control

Journal: Oncology reports

Article Title: Radioresistant breast cancer cells exhibit increased resistance to chemotherapy and enhanced invasive properties due to cancer stem cells.

doi: 10.3892/or.2018.6714

Figure Lengend Snippet: Figure 3. RT‑R‑MDA‑MB‑231 cells increase the populations of CD24low/CD44high cells, and expression levels of other CSCs markers Notch‑4, Oct‑3/4 and ALDH1. (A) CD24low/CD44high cells were isolated from MDA‑MB‑231 and RT‑R‑MDA‑MB cells, and then the number of CD24low/CD44high breast cancer stem cells was quantified by flow cytometry. (B) MDA‑MB‑231 and RT‑R‑MDA‑MB‑231 were labeled with anti‑CD24 and anti‑CD44 antibodies, and the percentages of CD24 or CD44‑expressed subpopulation in cells were determined by flow cytometry. (C) Other cancer stem cell markers, including Notch‑4, Oct3/4 and ALDH1 were detected in the MDA‑MB‑231 and RT‑R‑MDA‑MB‑231 cells using the specific antibodies by western blotting. Data represent mean values ± standard error of the mean of three independent experiments in triplicate. *P<0.05 and **P<0.01 compared with MDA‑MB‑231 cells. ALDH1, aldehyde dehydrogenase 1; CD, cluster of differentiation; CSCs, cancer stem cells; Oct, octamer‑binding transcription factor; RT‑R, radioresistant.

Article Snippet: Isolation of CD24low/CD44high breast cancer stem cells was performed using a MagCollect CD24- CD44+ Breast Cancer Stem Cell Isolation kit (R&D Systems, Inc., Minneapolis, MN, USA) following the manufacturer's protocol.

Techniques: Expressing, Isolation, Flow Cytometry, Labeling, Western Blot

Journal: PLoS ONE

Article Title: Induction of AMPK activation by N , N’- diarylurea FND-4b decreases growth and increases apoptosis in triple negative and estrogen-receptor positive breast cancers

doi: 10.1371/journal.pone.0209392

Figure Lengend Snippet: Equal numbers of MCF-7, T-47D, MDA-MB-231, HCC-1806 or breast cancer stem cells were treated with fresh medium that contained different FND-4b concentrations (0, 1, 2.5, 5, 10, and 20 μM) for 24 h. Western blot analysis was then performed for phosphorylated and total forms of AMPKα, ACC, and S6 as well as cyclin D1 and cleaved PARP. Beta-actin was used as the loading control. Treatment of the HCC-1806 cell line was repeated to confirm the spike in AMPKα activation at the 5 μM dosage with subsequent decreases at higher concentrations.

Article Snippet: Human breast cancer stem cell complete growth medium was from Celprogen (Torrance, CA).

Techniques: Western Blot, Activation Assay

Journal: PLoS ONE

Article Title: Induction of AMPK activation by N , N’- diarylurea FND-4b decreases growth and increases apoptosis in triple negative and estrogen-receptor positive breast cancers

doi: 10.1371/journal.pone.0209392

Figure Lengend Snippet: (A) Equal numbers of MCF-7, T-47D, MDA-MB-231, HCC-1806, or breast cancer stem cells were grown in medium containing 0 or 5 μM FND-4b for 72 h followed by cell counting. (B) The same procedure described in (A) was followed with the exception of growing cells in medium containing 0 or 1 mM AICAR instead. (C) MCF-7, T-47D, MDA-MB-231, and HCC-1806 cells were grown in medium containing different concentrations of FND-4b (0, 2.5, 5, 10, and 20 μM) for 72 h before SRB growth assays were performed. Data are presented as mean ± SD from experiments performed in triplicate (cell counting) or sextuplicate (SRB assays) and are representative of three independent experiments. * indicates p-value < 0.01.

Article Snippet: Human breast cancer stem cell complete growth medium was from Celprogen (Torrance, CA).

Techniques: Cell Counting

Journal: Cells

Article Title: Chronic High-Salt Diet Activates Tumor-Initiating Stem Cells Leading to Breast Cancer Proliferation

doi: 10.3390/cells13110912

Figure Lengend Snippet: Enumeration of TISC frequency in various murine passages following dietary salt modification. The frequency of CD44+CD24−TISCs cells in the isolated tumor cells among RS (grey) and HS (red) diet cohorts was enumerated by flow cytometry. ( A , B ) Representative flow cytometry plot of TISCs from passage 1 and passage 4 of Py230-C57Bl/6J HS diet cohort. ( C , D ) Changes in TISC frequency with each passage in Py230-C57Bl/6J ( C ) and 4T1-BALB/cJ ( D ) murine tumor models. ( E – N ) The mRNA expression of TISC markers Cadherin 1 ( E , J ), Snail2 ( F , K ), Aldh1A1 ( G , L ), Sox2 ( H , M ), and ITGA6 ( I , N ) in four passages of Py230-C57Bl/6J and 4T1-BALB/cJ tumor models (respectively). Data analyzed by one-way ANOVA for multiple comparisons and presented as mean ± SEM; n = 8 (biological replicates) per cohort; (*) p -value < 0.05.

Article Snippet: The TISCs were isolated from murine tumors by CD24 − CD44 + breast tumor stem cells using immunomagnetic beads as per the manufacturer’s specifications and reagents (catalog#MAGH111, R&D Systems, Minneapolis, MN, USA).

Techniques: Modification, Isolation, Flow Cytometry, Expressing

Journal: Cells

Article Title: Chronic High-Salt Diet Activates Tumor-Initiating Stem Cells Leading to Breast Cancer Proliferation

doi: 10.3390/cells13110912

Figure Lengend Snippet: TGFβ signaling induced the expression of immune-inhibitory CD80 on TISCs. ( A ) Representative flow cytometry histogram of TGFβR2 expression in passages 1 (blue), 2 (pink), 3 (olive green), and 4 (purple) from the HS diet cohort of the Py230-C57Bl/6J tumor model. ( B ) Comparison of the relative expression of TGFβR2 in each of the four passages on CD4+CD24-TISCs isolated from RS (grey) and HS (red) diet cohorts of the Py230-C57Bl/6J tumor model. ( C ) Representative flow cytometry histogram of pSMAD2/3 intracellular expression in each of the four passages from the HS diet cohort of the Py230-C57Bl/6J tumor model. ( D ) Comparison of the relative expression of pSMAD2/3 in each of the four passages on CD4+CD24-TISCs isolated from the RS and HS diet cohorts of the Py230-C57Bl/6J tumor model. ( E ) Representative flow cytometry histogram of CD80 surface expression in each of the four passages from the HS diet cohort of the Py230-C57Bl/6J tumor model. ( F ) Comparison of the relative expression of CD80 in each of the four passages on CD4+CD24-TISCs isolated from the RS and HS diet cohorts of the Py230-C57Bl/6J tumor model. ( G ) Representative flow cytometry plot to determine the TGFβR2/CD80 double-positive TISCs in passage 4 of the HS diet cohort from the Py230-C57Bl/6J tumor model. ( H ) Comparison of the relative expression of TGFβR2/CD80 double-positive TISCs in each of the four passages on CD4+CD24-TISCs isolated from the RS and HS diet cohorts of the Py230-C57Bl/6J tumor model. ( I ) Comparison of the relative expression of CD80 following TGFβ (80 ng/mL) stimulation for 5 days on CD4+CD24-TISCs isolated from each of the four passages of the RS and HS diet cohorts in the Py230-C57Bl/6J tumor model. ( J – M ) Comparison of the relative expression of TGFβR2 ( J ), pSMAd2/3 ( K ), CD80 ( L ), and TGFβR2/CD80 double-positive cells ( M ) in each of the four passages on CD4+CD24-TISCs isolated from the RS (grey) and HS (red) diet cohorts of the 4T1-BALB/cJ tumor model. ( N ) Comparison of the relative expression of CD80 following TGFβ (80 ng/mL) treatment for 5 days on CD4+CD24-TISCs isolated from each of the four passages of the RS and HS diet cohorts of the 4T1-BALB/cJ tumor model. Data analyzed by one-way ANOVA for multiple comparisons and presented as mean ± SEM; n = 8 (biological replicates) per cohort; (*) p -value < 0.05.

Article Snippet: The TISCs were isolated from murine tumors by CD24 − CD44 + breast tumor stem cells using immunomagnetic beads as per the manufacturer’s specifications and reagents (catalog#MAGH111, R&D Systems, Minneapolis, MN, USA).

Techniques: Expressing, Flow Cytometry, Comparison, Isolation