title: Acid ceramidase maintains the chondrogenic phenotype of expanded primary chondrocytes and improves the chondrogenic differentiation of bone marrow-derived mesenchymal stem cells.
abstract: Acid ceramidase is required to maintain the metabolic balance of several important bioactive lipids, including ceramide, sphingosine and sphingosine-1-phosphate. Here we show that addition of recombinant acid ceramidase (rAC) to primary chondrocyte culture media maintained low levels of ceramide and led to elevated sphingosine by 48 hours. Surprisingly, after three weeks of expansion the chondrogenic phenotype of these cells also was markedly improved, as assessed by a combination of histochemical staining (Alcian Blue and Safranin-O), western blotting (e.g., Sox9, aggrecan, collagen 2A1), and/or qPCR. The same effects were evident in rat, equine and human cells, and were observed in monolayer and 3-D cultures. rAC also reduced the number of apoptotic cells in some culture conditions, contributing to overall improved cell quality. In addition to these effects on primary chondrocytes, when rAC was added to freshly harvested rat, equine or feline bone marrow cultures an ~2-fold enrichment of mesenchymal stem cells (MSCs) was observed by one week. rAC also improved the chondrogenic differentiation of MSCs, as revealed by histochemical and immunostaining. These latter effects were synergistic with TGF-beta1. Based on these results we propose that rAC could be used to improve the outcome of cell-based cartilage repair by maintaining the quality of the expanded cells, and also might be useful in vivo to induce endogenous cartilage repair in combination with other techniques. The results also suggest that short-term changes in sphingolipid metabolism may lead to longer-term effects on the chondrogenic phenotype.
authors: Genetics & Genomic Sciences, Mount Sinai School of Medicine, New York, New York, United States of America.
authors: Calogera M
authors: CM
authors: Simonaro
authors: Sylvain
authors: S
authors: Sachot
authors: Yi
authors: Y
authors: Ge
authors: Xingxuan
authors: X
authors: He
authors: Victor A
authors: VA
authors: Deangelis
authors: Efrat
authors: E
authors: Eliyahu
authors: Daniel J
authors: DJ
authors: Leong
authors: Hui B
authors: HB
authors: Sun
authors: Jeffrey B
authors: JB
authors: Mason
authors: Mark E
authors: ME
authors: Haskins
authors: Dean W
authors: DW
authors: Richardson
authors: Edward H
authors: EH
authors: Schuchman
mesh_terms: D055573:Acid Ceramidase / Q000494:pharmacology*; D000368:Aged; D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D002358:Cartilage, Articular / Q000166:cytology; D002415:Cats; D002454:Cell Differentiation / Q000187:drug effects*; D049109:Cell Proliferation / Q000187:drug effects; D002469:Cell Separation; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D020219:Chondrogenesis / Q000187:drug effects*; D024043:Collagen Type II / Q000378:metabolism; D005260:Female; D006736:Horses; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D010641:Phenotype; D051381:Rats; D011994:Recombinant Proteins / Q000494:pharmacology; D013107:Sphingolipids / Q000378:metabolism
chemical_list: D024043:Collagen Type II; D011994:Recombinant Proteins; D013107:Sphingolipids; D055573:Acid Ceramidase
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