Expression of recombinant anticoagulant hirudin in the differentiated cultures of the porcine mammary epithelial cell line SI-PMEC

• Published in: Cell biology international Vol. 32; no. 7; pp. 739 - 747
• Main Authors: Sun, Y.L., Chou, Y.C., Kuan, T.C., Tu, C.F., Lin, C.S.
• Format: Journal Article
• Language: English
• Published: Oxford, UK Elsevier Ltd 01.07.2008; Blackwell Publishing Ltd
• Subjects: Amino Acid Sequence; Animals; Base Sequence; Cell Line; Collagen - pharmacology; Drug Combinations; Epithelial cell; Epithelial Cells - cytology; Epithelial Cells - metabolism; Fibrinolytic Agents - metabolism; Gene Expression; Genetic Vectors; Hirudin; Hirudins - biosynthesis; Hirudins - genetics; Laminin - pharmacology; Mammary gland; Mammary Glands, Animal; Molecular Sequence Data; Prolactin - pharmacology; Proteoglycans - pharmacology; Recombinant protein; Recombinant Proteins - biosynthesis; Recombinant Proteins - genetics; SI-PMEC; Swine; Transfection; Hi; SI-PMEC; pGB562/Hi; EGFP; pGB562/GFP/SI-PMEC; Hirudin; GB562; RT-PCR; pGB562/Hi/SI-PMEC; pGB562/GFP; Epithelial cell; Recombinant protein; Mammary gland
• ISSN: 1065-6995; 1095-8355
• DOI: 10.1016/j.cellbi.2008.02.004

To express recombinant proteins in the spontaneously immortalized porcine mammary epithelial cell line (SI-PMEC) currently established in our laboratory, a chemically synthesized DNA fragment encoding the anticoagulant hirudin was used to construct a mammalian expression vector under the control of the goat β-casein regulatory sequence. The vector, named pGB562/Hi, was transfected into the SI-PMEC cells to yield pGB562/Hi/SI-PMEC. The pGB562/Hi/SI-PMEC cells expressed recombinant hirudin only when they were differentiated into functional structures by growth on a Matrigel-coated petri dish supplemented with the lactogenic hormone prolactin. The differentiated pGB562/Hi/SI-PMEC cells produced about 0.5–0.6 μg of recombinant hirudin/mg of total cellular protein. These results suggest that the established SI-PMEC cells have pharmaceutical potential to inducibly express bioactive heterogeneous proteins.