Wednesday, May 6, 2009

Acid Ceramidase

Acid ceramidase maintains the chondrogenic phenotype of expanded primary chondrocytes and improves the chondrogenic differentiation of bone marrow-derived mesenchymal stem cells.

"Acid ceramidase is required to maintain the metabolic balance of several important bioactive lipids, including ceramide, sphingosine and sphingosine-1-phosphate.  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. Short-term changes in sphingolipid metabolism may lead to longer-term effects on the chondrogenic phenotype."

"rAC was taken up by rat chondrocytes and retained biological activity by hydrolyzing ceramide and producing sphingosine and S1P. The fact that AC levels returned to baseline by 7 days was consistent with its expected intracellular half-life of 48–72 h"

rAC was tested on human osteoarthritic patient cultures and pro-chondrogenic gene expression increased although there was no difference in cell number.

"Sphingosine may have a protective effect on chondrocytes."

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