Interruption of a transforming growth factor alpha autocrine loop in Caco-2 cells by antisense oligodeoxynucleotides.
OBJECTIVE
We have previously shown that Caco-2 cell proliferation is driven by basolateral membrane epidermal growth factor receptors. The aim of this study was to investigate whether autocrine production of transforming growth factor alpha (TGF-alpha) activates these receptors and stimulates proliferation using antisense oligodeoxynucleotides.


METHODS
Caco-2 cells grown on microporous membranes or Jurkat cells were exposed to conventional or 5' cholesterol-modified oligodeoxynucleotides synthesized with random, antisense, or missense base sequences. Indices of proliferation were measured, including [3H]thymidine or [3H]uridine uptake for studies of short-term stimulation and the methylthiotetrazole assay as an index of cell number increase over longer periods. Secretion of TGF-alpha by cells was detected using a soft agar bioassay.


RESULTS
Incubation with antisense oligodeoxynucleotides inhibited TGF-alpha secretion compared with controls. Random and missense oligodeoxynucleotides had no effect on proliferation. The TGF-alpha antisense oligodeoxynucleotides markedly inhibited proliferation, an effect that was abolished by adding TGF-alpha to the medium. Oligonucleotides had no effect on Jurkat cells, a lymphocytic cell line lacking epidermal growth factor receptors. Cholesterol-modified oligodeoxynucleotides were more effective and specific than unmodified oligodeoxynucleotides.


CONCLUSIONS
Caco-2 cell proliferation is driven by autocrine stimulation of epidermal growth factor receptors by TGF-alpha. This mechanism may be effectively inhibited by antisense oligodeoxynucleotides, particularly those modified by the 5' attachment of cholesterol.
D001483:Base Sequence; D018938:Caco-2 Cells / Q000378:metabolism* / Q000473:pathology; D002455:Cell Division; D002784:Cholesterol / Q000494:pharmacology; D066246:ErbB Receptors / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*; D016211:Transforming Growth Factor alpha / Q000096:biosynthesis* / Q000235:genetics
D016376:Oligonucleotides, Antisense; D016211:Transforming Growth Factor alpha; D002784:Cholesterol; D066246:ErbB Receptors