title: Inhibition of phosphodiesterase-4 (PDE4) activity triggers luminal apoptosis and AKT dephosphorylation in a 3-D colonic-crypt model.
abstract: BACKGROUND
  We previously established a three-dimensional (3-D) colonic crypt model using HKe3 cells which are human colorectal cancer (CRC) HCT116 cells with a disruption in oncogenic KRAS, and revealed the crucial roles of oncogenic KRAS both in inhibition of apoptosis and in disruption of cell polarity; however, the molecular mechanism of KRAS-induced these 3-D specific biological changes remains to be elucidated.
  
  
  RESULTS
  Among the genes that were upregulated by oncogenic KRAS in this model, we focused on the phosphodiesterase 4B (PDE4B) of which expression levels were found to be higher in clinical tumor samples from CRC patients in comparison to those from healthy control in the public datasets of gene expression analysis. PDE4B2 was specifically overexpressed among other PDE4 isoforms, and re-expression of oncogenic KRAS in HKe3 cells resulted in PDE4B overexpression. Furthermore, the inhibition of PDE4 catalytic activity using rolipram reverted the disorganization of HCT116 cells into the normal physiologic state of the epithelial cell polarity by inducing the apical assembly of ZO-1 (a tight junction marker) and E-cadherin (an adherens junction marker) and by increasing the activity of caspase-3 (an apoptosis marker) in luminal cavities. Notably, rolipram reduced the AKT phosphorylation, which is known to be associated with the disruption of luminal cavity formation and CRC development. Similar results were also obtained using PDE4B2-shRNAs. In addition, increased expression of PDE4B mRNA was found to be correlated with relapsed CRC in a public datasets of gene expression analysis.
  
  
  CONCLUSIONS
  These results collectively suggested that PDE4B is upregulated by oncogenic KRAS, and also that the inhibition of PDE4 catalytic activity can induce both epithelial cell polarity and luminal apoptosis in CRC, thus highlighting the utility of our 3-D culture (3 DC) model for the KRAS-induced development of CRC in 3-D microenvironment. Indeed, using this model, we found that PDE4B is a promising candidate for a therapeutic target as well as prognostic molecular marker in CRC. Further elucidation of the signaling network of PDE4B2 in 3 DC would provide a better understanding of CRC in vivo.
authors: Department of Cell Biology, Faculty of Medicine, Fukuoka University, Fukuoka 814-0180, Japan.
authors: Toshiyuki
authors: T
authors: Tsunoda
authors: Takeharu
authors: T
authors: Ota
authors: Takahiro
authors: T
authors: Fujimoto
authors: Keiko
authors: K
authors: Doi
authors: Yoko
authors: Y
authors: Tanaka
authors: Yasuhiro
authors: Y
authors: Yoshida
authors: Masahiro
authors: M
authors: Ogawa
authors: Hiroshi
authors: H
authors: Matsuzaki
authors: Masato
authors: M
authors: Hamabashiri
authors: Darren R
authors: DR
authors: Tyson
authors: Masahide
authors: M
authors: Kuroki
authors: Shingo
authors: S
authors: Miyamoto
authors: Senji
authors: S
authors: Shirasawa
mesh_terms: D017209:Apoptosis / Q000187:drug effects / Q000235:genetics*; D045744:Cell Line, Tumor; D016000:Cluster Analysis; D015179:Colorectal Neoplasms / Q000235:genetics* / Q000378:metabolism*; D054703:Cyclic Nucleotide Phosphodiesterases, Type 4 / Q000235:genetics* / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D011905:Genes, ras; D045325:HCT116 Cells; D006801:Humans; D058988:Phosphodiesterase 4 Inhibitors / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D034622:RNA Interference; D012008:Recurrence; D020889:Rolipram / Q000494:pharmacology; D018874:Spheroids, Cellular; D019108:Tight Junctions / Q000378:metabolism; D014407:Tumor Cells, Cultured
chemical_list: D058988:Phosphodiesterase 4 Inhibitors; D051057:Proto-Oncogene Proteins c-akt; D054703:Cyclic Nucleotide Phosphodiesterases, Type 4; D020889:Rolipram