Changes in ceramide metabolism are essential in Madin-Darby canine kidney cell differentiation.

Ceramides (Cers) and complex sphingolipids with defined acyl chain lengths play important roles in numerous cell processes. Six Cer synthase (CerS) isoenzymes (CerS1-6) are the key enzymes responsible for the production of the diversity of molecular species. In this study, we investigated the changes in sphingolipid metabolism during the differentiation of Madin-Darby canine kidney (MDCK) cells. By MALDI TOF TOF MS, we analyzed the molecular species of Cer, glucosylceramide (GlcCer), lactosylceramide (LacCer), and SM in nondifferentiated and differentiated cells (cultured under hypertonicity). The molecular species detected were the same, but cells subjected to hypertonicity presented higher levels of C24:1 Cer, C24:1 GlcCer, C24:1 SM, and C16:0 LacCer. Consistently with the molecular species, MDCK cells expressed CerS2, CerS4, and CerS6, but with no differences during cell differentiation. We next evaluated the different synthesis pathways with sphingolipid inhibitors and found that cells subjected to hypertonicity in the presence of amitriptyline, an inhibitor of acid sphingomyelinase, showed decreased radiolabeled incorporation in LacCer and cells did not develop a mature apical membrane. These results suggest that hypertonicity induces the endolysosomal degradation of SM, generating the Cer used as substrate for the synthesis of specific molecular species of glycosphingolipids that are essential for MDCK cell differentiation.

Aptamers: current challenges and future prospects.

INTRODUCTION: Aptamers are oligonucleotide molecules raised in vitro from large combinatorial libraries of nucleic acids and developed to bind to targets with high affinity and specificity. Whereas novel target molecules are proposed for therapeutic intervention and diagnostic, aptamer technology has a great potential to become a source of lead compounds. AREAS COVERED: In this review, the authors address the current status of the technology and highlight the recent progress in aptamer-based technologies. They also discuss the current major technical limitations of aptamer technology and propose original solutions based on existing technologies that could result in a solid aptamer-discovery platform. EXPERT OPINION: Whereas aptamers have shown to bind to targets with similar affinities and specificities to those of antibodies, aptamers have several advantages that could outweigh antibody technology and open new opportunities for better medical and diagnostic solutions. However, the current status of the aptamer technology suffers from several technical limitations that slowdown the progression of novel aptamers into the clinic and makes the business around aptamers challenging.