Establishment of a cholecystokinin-producing rat medullary thyroid carcinoma cell line.
A rat medullary thyroid carcinoma, which was previously shown to produce high levels of immunoreactive cholecystokinin (CCK), was used to establish a stable cell line. Transplantable tumors were subjected to four series of alternate in vitro and in vivo passages. Cells were prepared from the fourth series of tumors under serum-free medium conditions that prevent fibroblast growth. Subcloning of these cells yielded several propagatable clonal cell lines. One cell line with immunoreactive CCK-8 production was selected for further studies. This high CCK cell line, WE4/2, produces and secretes a CCK-immunoreactive product that coelutes with synthetic CCK-8 sulfate during Sephadex chromatography and HPLC. Northern analysis with a rat CCK cDNA revealed that the cultured cells produce a CCK RNA the same size and with the same 5' end as that previously reported for brain and intestines. In addition, a recombinant plasmid containing about 800 basepairs of 5' flanking sequence of the rat CCK gene linked to the coding sequence of the bacterial chloramphenicol acetyltransferase gene elicited a high level of chloramphenicol acetyltransferase activity when transfected into the WE4/2 cell line. Therefore, the WE4/2 cell line provides a model system for studying CCK gene expression and biosynthesis.
Haun
R S
RS
Department of Biochemistry, Purdue University, West Lafayette, Indiana 47907.
Beinfeld
M C
MC
Roos
B A
BA
Dixon
J E
JE
D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002471:Cell Transformation, Neoplastic / Q000473:pathology; D002766:Cholecystokinin / Q000235:genetics / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D004247:DNA / Q000032:analysis; D005786:Gene Expression Regulation; D009368:Neoplasm Transplantation; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D051381:Rats; D013964:Thyroid Neoplasms / Q000378:metabolism* / Q000473:pathology; D014407:Tumor Cells, Cultured / Q000378:metabolism / Q000473:pathology
D012333:RNA, Messenger; D004247:DNA; D002766:Cholecystokinin