Messenger RNA Expression of Transporter and Ion Channel Genes in Undifferentiated and Differentiated Caco-2 Cells Compared to Human Intestines

Purpose. The purpose of this work was to study the influence of cell differentiation on the mRNA expression of transporters and channels in Caco-2 cells and to assess Caco-2 cells as a model for carrier-mediated drug transport in the intestines. Method. Gene mRNA expression was measured using a custom-designed microarray chip with 750 deoxyoligonucleotide probes (70mers). Each oligomer was printed four times on poly-lysine-coated glass slides. Expression profiles were expressed as ratio values between fluorescence intensities of Cy3 and Cy5 dye-labeled cDNA derived from poly(A) + RNA samples of Caco-2 cells and total RNA of human intestines. Results. Significant differences in the mRNA expression profile of transporters and channels were observed upon differentiation of Caco-2 cells from 5 days to 2 weeks in culture, including changes for MAT8, S-protein, and Nramp2. Comparing Caco-2 cells of different passage number revealed few changes in mRNAs except for GLUT3, which was down-regulated 2.4-fold within 13 passage numbers. Caco-2 cells had a similar expression profile when either cultured in flasks or on filters but differed more strongly from human small and large intestine, regardless of the differentiation state of Caco-2 cells. Expression of several genes highly transcribed in small or large intestines differed fourfold or more in Caco-2 cells. Conclusions. Although Caco-2 cells have proven a suitable model for studying carrier-mediated transport in human intestines, the expression of specific transporter and ion channel genes may differ substantiall

A synthetic human Agouti-related protein-(83–132)-NH2 fragment is a potent inhibitor of melanocortin receptor function

AbstractChemical synthesis of Agouti proteins – Agouti and Agouti-related proteins – is complicated by their large size and by multiple cysteine residues located in the carboxyl terminal regions. Three human Agouti-related protein (AGRP) fragments, two of which correspond to a proposed endoprotease cleavage site between amino acids 82 and 83, were synthesized and tested for anti-melanotropic activity using Xenopus laevis dermal melanophores. Amino-terminal fragments AGRP(25–51) and (54–82) were devoid of significant antagonist activity, whereas the amidated carboxyl-terminal AGRP fragment (83–132)-NH2 was potently active with an inhibitory equilibrium dissociation constant (Ki) of 0.7 nM. The ability to synthesize functionally active AGRP should help unravel its role in the central nervous system and its unusual properties with respect to interaction with the melanocortin family of G-protein coupled receptors

Genomics and Drugs: Finding the Optimal Drug for the Right Patient

ISSN:0724-8741ISSN:1573-904