Journal: Breast Cancer Research

Article Title: WISP3 and RhoC guanosine triphosphatase cooperate in the development of inflammatory breast cancer

doi: 10.1186/bcr755

Figure Lengend Snippet: Inhibition of WISP3 in human mammary epithelial (HME) cells results in an increase in RhoC transcript levels. Reverse transcriptase polymerase chain reaction was conducted on vector and HME cells that have inhibition of WISP3 expression using full-length WISP3 antisense mRNA. HME/AS WISP3 cells showed increased levels of RhoC transcript in comparison with controls.

Article Snippet: MCF10A cells were stably tranfected with full-length RhoC cDNA (MCF10A/RhoC). pFlag control vectors were used as controls (FuGene TM 6 transfection reagent; Roch–Boehringer-Mannheim, Germany).

Techniques: Inhibition, Reverse Transcription, Polymerase Chain Reaction, Plasmid Preparation, Expressing, Comparison

Journal: Breast Cancer Research

Article Title: WISP3 and RhoC guanosine triphosphatase cooperate in the development of inflammatory breast cancer

doi: 10.1186/bcr755

Figure Lengend Snippet: Restoration of WISP3 expression in SUM149 inflammatory breast cancer cells decreases RhoC protein expression. Western immunoblot of cell culture of SUM 149 cells, empty vector control (SUM149/Flag), and two WISP3-expressing clones with antibodies against RhoC and actin. Gels were scanned and pixel intensity values were obtained. Values for RhoC were corrected for loading by dividing the RhoC pixel intensity by the actin pixel intensity.

Article Snippet: MCF10A cells were stably tranfected with full-length RhoC cDNA (MCF10A/RhoC). pFlag control vectors were used as controls (FuGene TM 6 transfection reagent; Roch–Boehringer-Mannheim, Germany).

Techniques: Expressing, Western Blot, Cell Culture, Plasmid Preparation, Control, Clone Assay

Journal: Breast Cancer Research

Article Title: WISP3 and RhoC guanosine triphosphatase cooperate in the development of inflammatory breast cancer

doi: 10.1186/bcr755

Figure Lengend Snippet: RhoC overexpression in MCF10A cells results in a decrease in WISP3 mRNA level. Reverse transcriptase polymerase chain reaction was conducted on vector and MCF10A cells stably overexpressing RhoC. RhoC-overexpressing cells had decreased levels of WISP3 mRNA.

Article Snippet: MCF10A cells were stably tranfected with full-length RhoC cDNA (MCF10A/RhoC). pFlag control vectors were used as controls (FuGene TM 6 transfection reagent; Roch–Boehringer-Mannheim, Germany).

Techniques: Over Expression, Reverse Transcription, Polymerase Chain Reaction, Plasmid Preparation, Stable Transfection

Journal:

Article Title: TAp63 Is a Transcriptional Target of NF-?B

doi: 10.1002/jcb.22449

Figure Lengend Snippet: NF-κB regulates TAp63 mRNA expression. MCF10A cells were transiently transfected with p50- and RelA-expression plasmids for 24 h (A) or treated with a specific NF-κB inhibitor Bay 11-7082 (10 μM) for the indicated times (B). Total RNA was extracted and subjected to quantitative-PCR (Q-PCR) analysis. The amount of TAp63 mRNA was normalized to GAPDH in the same samples and presented as mean ± SE. Three independent experiments were performed in triplicate. ** P < 0.01. C: Primary cortical neuron cells isolated form E15.5 embryonic mice were treated with Bay 11-7082 (10 μM) for the indicated times followed by Q-PCR. Two independent experiments were performed in triplicate. * P < 0.05.

Article Snippet: Since these cells have detectable levels of NF-κB, we next treated MCF10A cells with Bay 11-7082, a specific inhibitor of NF-κB [ Pierce et al., 1997 ].

Techniques: Expressing, Transfection, Real-time Polymerase Chain Reaction, Isolation

Journal: PLOS ONE

Article Title: New oncogenic functions of LINE1 retroelement as a ceRNA for tumor suppressive microRNA miR-126 on ENPP5

doi: 10.1371/journal.pone.0286814

Figure Lengend Snippet: (A) Hsa-miR-126 levels in BRCA public database. (B) Hsa-miR-126-5p expression in HBC cell lines compared to normal MCF-10A cells. (C) Kaplan-Meier plot showed that highly expressed hsa-miR-126 associated with better overall survival in HBC patients. (D) cfa-miR-126 expression was down-regulated in CMT cell lines when compared with normal mammary gland. (E) ENPP5 expression was significantly up regulated in BRCA public database. (F) ENPP5 expression was also significantly up-regulated in HBC related cell lines of SKBR-3, MCF-7 and MDA-MB-231. Log2-transformed expression was used (G) miR-126-5p (mimic) transfection significantly reduced the ENPP5 mRNA levels. (H) miR-126-5p transfection (mimic) also significantly reduced the luciferase activity conjugated with the 3’UTR of ENPP5. (I) The luciferase activity conjugated with the 3’UTR of ENPP5 was significantly increased by the overexpression of LINE in MDA-MB-231 cells. (J) ENPP5 mRNA was significantly increased by LINE overexpression in MCF-10A cells. *, **, *** and **** indicate significance p<0.05, 0.01, 0.001, and 0.0001, respectively.

Article Snippet: ENPP5 3’UTR sequence was amplified with MCF-10A genomic DNA, digested with AsiSi and XhoI enzyme (NEB, MA, USA), and inserted into pEZX-GA02 vector (GeneCopoeia, MD, USA).

Techniques: Expressing, Transformation Assay, Transfection, Luciferase, Activity Assay, Over Expression