Article Title: Sulforaphane counteracts aggressiveness of pancreatic cancer driven by dysregulated Cx43-mediated gap junctional intercellular communication
Figure Lengend Snippet: Sulforaphane induces Cx43 and inhibits CSC characteristics in primary CSCs (A) Staining of patient-derived frozen tissue from a ductal adenocarcinoma (Primary Tumor 22) and it´s derived xenograft from passage 3 (Xenograft P3) with Trichrome, c-Met or Cx43 phosphorylated at Ser 368 (Abcam), followed by microscopical evaluation under 400× magnification. The scale bar indicates 50 μm. (B) Representative picture of an anchorage-independent growing spheroidal culture established from a mouse xenograft derived from the primary patient tumor 22. The percentage of expression of the CSC markers c-Met and CD133 was determined as described in part C and is indicated. (C) One week after in vitro spheroidal culture, cells derived from pancreatic ductal adenocarcinoma 22 and 30 were left untreated or were treated with sulforaphane (10 μM). Twenty-four hours later, the cells were cytospinned to glass slides, and the expression of Cx43 phosphorylated at Ser 368 (Abcam), c-Met, CD133, E-cadherin and the cleaved, active fragment of Caspase-3 was examined by immunohistochemistry. The number of positive cells was quantified in 10 vision fields under 400× magnification and the means ± SD are shown in the diagrams. **p
Article Snippet: Primary Antibodies Rabbit polyclonal antibodies (pAbs) were used against Cx32, Cx26 (Invitrogen, Camarillo, California; USA), c-Met, total Cx43 (#3512, Cell Signaling Technology, Boston, MA, USA), Cx43 phosphorylated at Ser 368 (#ab47368; Abcam, Cambridge, UK), Cx45 phosphorylated at Ser 279/282 (Santa Cruz Biotechnology, Inc. Heidelberg, Germany), Cx45, CD44 (GeneTex, Irvine, California, USA), acetyl-Histone H3 and acetyl-Histone H4 (Merck Millipore, Darmstadt, Germany), E-cadherin (Cell Signaling), Ki67 (Thermo Scientific, Rockford, IL, USA) and the cleaved fragment of activated human caspase-3 (R & D Systems, Abingdon, UK).
Techniques: Staining, Derivative Assay, Expressing, In Vitro, Immunohistochemistry