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J Biol Chem, Vol. 273, Issue 34, 21569-21577, August 21, 1998
From the Eicosanoid Biochemistry Section, Laboratory of Molecular
Carcinogenesis, NIEHS, National Institutes of Health, Research Triangle
Park, North Carolina 27709
We studied arachidonic acid metabolism and the
expression of cyclooxygenase (Cox) and 15-lipoxygenase (15-LO) in the
human colorectal carcinoma cell line, Caco-2, which undergo apoptosis and cell differentiation in the presence of sodium butyrate (NaBT). Caco-2 cells expressed very low levels of Cox-1 but highly expressed Cox-2. NaBT treatment shifted the arachidonic acid metabolites by cell
lysates from prostaglandins to 15-hydroxyeicosatetraenoic acid,
indicating the presence of a 15-LO. Linoleic acid, an excellent substrate for 15-LO, was metabolized poorly by the Caco-2 cells, but
NaBT treatment shifted metabolism to 15-LO metabolite,
13(S)-hydroxyoctadecadienoic acid. Caco-2 cells expressed a
15-LO but only after treatment with NaBT, as determined by Northern
blotting. Immunoblotting with anti-human 15-LO antibody detected a
72-kDa band in NaBT-treated Caco-2 cells. Expression of 15-LO mRNA
was dependent on the duration of NaBT treatment, with the highest
expression observed between 10 and 24 h. Results from expression
and metabolism studies with arachidonic and linoleic acid cells
indicated Cox-2 was responsible for the lipid metabolism in control
cells, whereas 15-LO was the major enzyme responsible after NaBT
induction of apoptosis and cell differentiation. The 15-LO in Caco-2
cells was characterized as human reticulocyte 15-LO by reverse
transcription-polymerase chain reaction and restriction enzyme
analysis. The expression of 15-LO and 15-hydroxyeicosatetraenoic acid
or 13(S)-hydroxyoctadecadienoic acid formation correlates
with cell differentiation or apoptosis in Caco-2 cells induced by NaBT.
The addition of nordihydroguaiaretic acid, a lipoxygenase inhibitor,
significantly increased NaBT-induced apoptosis, whereas the addition of
indomethacin did not alter NaBT-induced apoptosis in the Caco-2 cells.
However, indomethacin treatment decreased the expression of Cox-2 in
NaBT-treated cells and significantly increased the expression of 15-LO
during NaBT treatment. These studies suggest a role for 15-LO, in
addition to Cox-2, in modulating NaBT-induced apoptosis and cell
differentiation in human colorectal carcinoma cells.
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