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

PESCIO, LUCILA GISELE; SANTACREU, BRUNO JAIME; LOPEZ, VANINA GISELA; PAVÁN, CARLOS HUMBERTO; ROMERO, DANIELA JUDITH; FAVALE, NICOLÁS OCTAVIO; STERIN-SPEZIALE, NORMA BEATRIZ

JLR PAPERS IN PRESS

AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC

Año: 2017 vol. 58 p. 1428 - 1438

0022-2275


Cermides (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 differ- entiation 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 (cul- tured under hypertonicity). The molecular species detected were the same, but cells subjected to hypertonicity pre- sented 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 sug- gest that hypertonicity induces the endolysosomal degrada- tion of SM, generating the Cer used as substrate for the synthesis of specific molecular species of glycosphingolipids that are essential for MDCK cell differentiation.