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24 C. Huill el a/. / Biology oflhe Ce1l94 (2002) 15-27
Total RNA
1.5 PPAR a PPARy
1.2
1.0
0.8
0.6
0.4
Poly (A+) RNA
PPAR a
PPAR y
0.2
o.\--l--'-----L.,...-J-__---L:.:;....L-".L.:...e..L,
Days of culture
5 10 15
5 10 15
PPARa
PPAR Y
G3PDH
5 10
Total RNA
15 5 10
PolyA
15
Fig. 6. PPARa and y transcript levels analyzed by Northern blotting. Total and poly(A+)RNA from Caco-2 cells cultured for 5, 10, 15 days were analyzed
by Northern blotting. Membranes were hybridized with PPARa DNA probe, dehybridized and reprobed \Vith PPARy and G3PDH probes, successively. The
bands \Vere scanned and PPAR values \Vere standardized against G3PDH values then plotted. Results are expressed in arbitrary units.
polyp and tumor formation in the Min/+mouse model (Lefebvre
et al., 1998; Saez et al., 1998). It is evident from our
resu1ts that the spontaneous differentiation of Caco-2 cells is
accompanied by an increase in PPARy expression. Our data
are in good agreement with those of Wachtershauser et al.
(2000) showing that PPARy mRNA and protein increase
significant1y during butyrate-induced Caco-2 ceH differentiation.
PPARy also plays a major ro1e in the differentiation
of other ceHs such as adipocytes (Tontonoz et al., 1994; Wu
et al., 1999), monocytes/macrophages (Tontonoz et al.,
1998), preputia1 sebocytes (Rosenfie1d et al., 1999), epidermal
cells (Rivier et al., 1998) as well as colon (Brockman et
al., 1998; Sarraf et al., 1998; Kitamura et al., 1998), prostate
(Kubota et al., 1998) and breast (MueHer et al., 1998) cancer
epithelial cells. Furthermore, activation of PPARy results in
differentiation in patients with 1iposarcoma (Demetri et al.,
1999). PPARy does not only control genes responsible for
the differentiated cell phenotype, but a1so participates to the
regulation of cell cycle withdrawa1 (Altiok et al., 1997). In
fact, PPARy activation inhibits the DNA-binding and transcriptiona1
activity of E2F/DP factors, which are invo1ved in
cell growth. At the present time, it is difficult to speculate
about the precise ro1e played by PPARy in the colon cell's
life.
In summary, the long-term culture of the human adenocarcinoma
Caco-2 ceHs leads to their differentiation. The
latter is accompanied by the development of the peroxisomal
compartment marked by an increase in both number
and enzyme activity of peroxisomes. Concomitantly to the
differentiation of Caco-2 celIs, the expression of PPARa,
PPARYI and PPARY2 is increased. A possible involvement
of PPARa in the maturation process of peroxisomes is
suggested because this transcription factor controis genes
encoding peroxisomal proteins. On the other hand, the
consequences of the inCl'ease in PPARy expression in cell
differentiation remain unclear.
Acknowledgements
We wish to thank Dr. M. Rousset (Inserm U505, Paris,
France) and Prof. D. Louvard (Institut Curie, Paris, France)
for providing us with the Caco-2 cell line and the anti-villin
antibody, respectively; Dr. L. Domenjoud for reading the
manuscript. We express our gratitude to Prof. D. Desor for
the statistical analysis of our results, to A. Stoekel and
1. Chanel for the expert technical assistance. This study was
supported by grants from the Association de la recherche
contre le cancer (contrat ARC No.o9233), the Ligue contre
le cclllcer (comité de Meurthe et Moselle), the Fondation de
la recherche médicale (comité de Lorraine) and ARERS.