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Sphingosine-kinase and sphingosine-1-phosphate regulate migration of immature dendritic cells
© Author(s); licensee BioMed Central Ltd. 2004
Received: 28 April 2004
Published: 1 July 2004
Currently, dendritic cells (DC) are tested as vectors for cancer immunotherapy. However, little is known about the mechanisms regulating DC migration. Sphingosine-kinase (SphK) and its catalytic product Sphingosine-1-phosphate (S1P) play a central role in processes such as cellular differentiation, survival and migration, which are often disregulated in cancer. Here, we examined the role of SphK and S1P in migration of DC.
DC were generated in the presence of GM-CSF and IL-4 for 5 days, then matured with PGE2, TNF-α and IL1-β for 2 days. Expression of SphK and S1P-receptors was examined by RT-PCR. In transwell assays migration of immature (i) and mature (m) DC towards SDF-1, MiP-1a, MCP, 6Ckine, MiP-3β [100 ng/ml] and S1P [10-5 M] was tested for being dependent on SphK using the SphK inhibitor Dihydrosphingosine [DHS, 10-6 M]. The role of S1P3 receptor in S1P-induced migration was tested using the S1P3-inhibitor Suramin [10-6 M]. In parallel, Ca2+-flux was assessed by FACS with Fura Red.
SphK expression was declining from iDC to mDC to antigen-loaded mDC. Expression levels of S1P receptors were S1P1>S1P2 = S1P3, unrelated to maturation stage or antigen uptake. IDC migrated on SDF-1, MIP-1α, MCP and S1P, whereby S1P combined with a chemokine acted synergistic. MDC migrated on 6Ckine and MIP-3β, but not on S1P. Pre-treatment with DHS inhibited migration of iDC but not mDC, showing that SphK is required for iDC migration. Pre-treatment with Suramin inhibited iDC migration in response to S1P, demonstrating a mediation via S1P3. Chemokine induced Ca2+-flux was inhibited by DHS, indicating that SphK-mediated migration might be Ca2+-dependent.
Our results suggest a role for SphK/S1P in accumulation of peripheral iDC at sites of antigen invasion. These findings could provide a new approach to optimise DC-based cancer immunotherapy by therapeutic modulation of SphK/S1P and have to be verified in an animal model in the next step.
This work was supported by the Friedrich-Baur-Stiftung and the Promotionsstudium Molekulare Medizin, Munich, and contains data of the doctoral thesis of Veronika Jakl at the Ludwig-Maximilians-University, Munich, Germany.