Sphingomyelin synthase1 activity is essential for the renal cell differentiation

FAVALE, NO; SANTACREU B; PESCIO, LG; MARQUEZ, MG; STERIN SPEZIALE, NB

Banff,

Conferencia; 53th International Conference on the Bioscience of Lipids (ICBL); 2012

ICBL - ASBMB

We have previously demonstrated that sphingomyelin (SM) biosynthesis is essential for hypertonicity-induced Madin-Darby canine kidney (MDCK) cell differentiation. Under inhibition of SM synthesis, instead MDCK cells switch to a mesenchymal phenotype to differentiate, thus performing an epithelial to mesenchymal transition. Our aim was to study the sphingolipid metabolic pathway as well as the sphingomyelin synthase isoform involved in such a process. Confluent MDCK cells were subjected to hypertonicity and concomitantly treated or not (control) with 15 μM D609 or siRNA-SMS1. Cell viability was determined by trypan blue, cell phenotype was visualized by confocal immunofluorescence of the cytoskeleton using phalloidin, and adherens junctions were determined by visualization of E-cadherin distribution. In these condition total SM level decreased, and ceramide (Cer) level increased. To evaluate whether the effect of SMSinhibition was due to SM decrease or Cer augmentation, sphingolipid metabolism was determined by using [14C]palmitic acid precursor in the presence or absence of cycloserine (CS) or fumonisin B1 (FB1). By using D609 as well as siRNA-SMS1 the characteristic polarized phenotype of the cells was lost, and no retrieval was observed by a concomitant treatment with CS or FB1, which denotes no involvement of Cer accumulation. Acquiring the mesenchymal phenotype was accompanied by loss of the epithelial marker (cad16) and the rise of the mesenchymal marker (vimentin). These results demonstrate implication of SM synthesis in the epithelial-mesenchymal transition (EMT). It is important to note that EMT has been implicated in the development ofcancer and renal fibrosis, consequently SMS activity emerges as a possible target molecule for the study of such important human pathologies.