Antisense oligodeoxynucleotides directed against aspartyl (asparaginyl) beta-hydroxylase suppress migration of cholangiocarcinoma cells.
BACKGROUND
Aspartyl (asparaginyl) beta-hydroxylase (AAH) is an alpha-ketoglutarate-dependent dioxygenase that hydroxylates aspartate and asparagine residues in EGF-like domains of proteins. The consensus sequence for AAH beta-hydroxylation occurs in signaling molecules such as Notch and Notch homologs, which have roles in cell migration.


OBJECTIVE
This study evaluated the potential role of AAH in cell migration using cholangiocarcinoma cell lines as models due to their tendency to widely infiltrate the liver.


METHODS
Five human cholangiocarcinoma cell lines established from human tumors were examined for AAH expression and motility. The effect of antisense oligodeoxynucleotide inhibition of AAH on cholangiocarcinoma cell migration was investigated.


RESULTS
Western blot analysis detected the approximately 86 kDa AAH protein in all five cholangiocarcinoma cell lines, and higher levels of AAH in cell lines derived from moderately or poorly differentiated compared with well-differentiated tumors. Immunocytochemical staining and fluorescence activated cell sorting analysis revealed both surface and intracellular AAH immunoreactivity. Using the phagokinetic non-directional migration assay and a novel ATPLite luminescence-based directional migration assay, we correlated AAH expression with motility. Correspondingly, antisense and not sense or mutated antisense AAH oligodeoxynucleotides significantly inhibited AAH expression and motility in cholangiocarcinoma cells.


CONCLUSIONS
AAH over-expression may contribute to the infiltrative growth pattern of cholangiocarcinoma cells by promoting motility.
Maeda
Takashi
T
Departments of Medicine and Pathology, Liver Research Center, Rhode Island Hospital, Brown Medical School, 55 Claverick Street, 4th Floor, Providence, RI 02903, USA.
Sepe
Paul
P
Lahousse
Stephanie
S
Tamaki
Seishu
S
Enjoji
Munetomo
M
Wands
Jack R
JR
de la Monte
Suzanne M
SM
D001650:Bile Duct Neoplasms; D001653:Bile Ducts, Intrahepatic; D045744:Cell Line, Tumor / Q000166:cytology / Q000201:enzymology; D002465:Cell Movement / Q000502:physiology*; D018281:Cholangiocarcinoma; D015971:Gene Expression Regulation, Enzymologic; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D006899:Mixed Function Oxygenases / Q000235:genetics* / Q000378:metabolism*; D020320:Oligoribonucleotides, Antisense / Q000494:pharmacology
D020320:Oligoribonucleotides, Antisense; D006899:Mixed Function Oxygenases; C058291:aspartic acid 2-oxoglutarate-dependent dioxygenase