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Endocrinology, Vol 125, 850-856, Copyright © 1989 by Endocrine Society

ARTICLES

Establishment of a cholecystokinin-producing rat medullary thyroid carcinoma cell line

RS Haun, MC Beinfeld, BA Roos and JE Dixon
Department of Biochemistry, Purdue University, West Lafayette, Indiana 47907.

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.


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