Journal: Oncogene

Article Title: The human DEK oncogene stimulates beta catenin signaling, invasion and mammosphere formation in breast cancer

doi: 10.1038/onc.2011.2

Figure Lengend Snippet: A) Western blotting for DEK revealed high expression in ten breast cancer cell lines and the non-tumorigenic MCF10A immortalized human mammary epithelial cell line compared to two primary normal breast samples from separate donors. Actin was used as a loading control. The blot is over-exposed to emphasize low expression in normal tissue (B) A tissue microarray was stained for the DEK protein (brown) by immunohistochemistry. Normal, abnormal, and cancerous tissue are shown: (a) normal tissue, (b) hyperplasia, (c) fibrocystic changes, (d) grade I carcinoma in situ , (e) grade II-III, and (f) grade III invasive ductal carcinomas. Low power images are at 100x total magnification.

Article Snippet: Tissue sections from paraffin-embedded MDA-MB-468 NTsh xenograft tumors from the mammary gland were deparaffinized, underdwent antigen retrieval with 10 mM sodium citrate, and were blocked with 5% normal donkey serum and immunostained with antibodies for DEK (1:50, Aviva Systems Biology, San Diego, CA) and PCNA (1:50, BD Biosciences).

Techniques: Western Blot, Expressing, Microarray, Staining, Immunohistochemistry, In Situ

Journal: Oncogene

Article Title: The human DEK oncogene stimulates beta catenin signaling, invasion and mammosphere formation in breast cancer

doi: 10.1038/onc.2011.2

Figure Lengend Snippet: (A) MDA-MB-468, MCF10A, and MCF7 cells were retrovirally transduced with either empty vector (R780) or a DEK over-expression construct (R780:DEK). MDA-MB-468 and MCF7 cells were transduced with lentiviral shRNA constructs for non-targeting shRNA (NTsh) or two distinct DEK shRNAs (DEKsh2 or DEKsh5). Lysates were analyzed by Western blotting for DEK, cyclin A and Actin expression. (B) DEK over-expression significantly increases cellular growth rates in non-tumorigenic MCF10A cells and moderately increases growth rates in cancer cells. Population growth was monitored by plating equal numbers of control and DEK over-expressing cells then counting cell numbers over the course of five to seven days (p<0.05 in MCF10A cells, p=0.06 for MCF7 cells). (C) Loss of DEK expression inhibits growth in breast cancer cells. Population growth was monitored as in (B) immediately after selection in puromycin.. (D) Loss of DEK expression results in increased rates of apoptosis. MCF7 and MDA-MB-468 NTsh or DEKsh2 cells were stained for cleaved caspase 3 and analyzed by flow cytometry.

Article Snippet: Tissue sections from paraffin-embedded MDA-MB-468 NTsh xenograft tumors from the mammary gland were deparaffinized, underdwent antigen retrieval with 10 mM sodium citrate, and were blocked with 5% normal donkey serum and immunostained with antibodies for DEK (1:50, Aviva Systems Biology, San Diego, CA) and PCNA (1:50, BD Biosciences).

Techniques: Transduction, Plasmid Preparation, Over Expression, Construct, shRNA, Western Blot, Expressing, Selection, Staining, Flow Cytometry

Journal: Oncogene

Article Title: The human DEK oncogene stimulates beta catenin signaling, invasion and mammosphere formation in breast cancer

doi: 10.1038/onc.2011.2

Figure Lengend Snippet: (A) (Top) Loss of DEK expression impairs tumor growth in nude mice. MDA-MB-468 breast cancer cells transduced with lentiviral non-targeting shRNA (NTsh) or DEK shRNA (DEKsh2) were injected into the mammary fat pad of female nude mice (N=10 per cell line) and tumor growth was monitored with calipers for ten weeks. (Bottom) At necropsy, tumors were removed from the mammary gland and measured in three dimensions with calipers to calculate the final tumor volume (volume = (π/6)*(LxWxH)). (B) (Top) H&E images of representative tumors. T = tumor and N = necrosis. Arrows indicate normal mammary ducts. Immunohistochemical staining of tumors showed a correlation between DEK (middle panel) and ΔNp63 (bottom panel) staining. The H&E images are at 100x magnification. (Right) Western blotting of tumor lysates show down-regulation of ΔNp63 and upregulation of E-cadherin in DEKsh2 tumors. (C) DEKsh2 tumors had a higher percentage of cleaved caspase-3 positive cells, indicative of apoptosis, compared to NTsh-derived tumors. (Right) Representative immunohistochemistry images for caspase 3 staining. Arrows indicate representative positive cells. (D) DEKsh2 tumors had fewer BrdU-positive cells compared to NTsh-derived tumors. (Right) Representative immunohistochemistry images for BrdU staining. (E) Tumor cells exhibit co-expression of DEK with PCNA. Sections from paraffin-embedded NTsh derived tumors were immunolabelled for DEK (red) and PCNA (green).

Article Snippet: Tissue sections from paraffin-embedded MDA-MB-468 NTsh xenograft tumors from the mammary gland were deparaffinized, underdwent antigen retrieval with 10 mM sodium citrate, and were blocked with 5% normal donkey serum and immunostained with antibodies for DEK (1:50, Aviva Systems Biology, San Diego, CA) and PCNA (1:50, BD Biosciences).

Techniques: Expressing, Transduction, shRNA, Injection, Immunohistochemical staining, Staining, Western Blot, Derivative Assay, Immunohistochemistry, BrdU Staining