Artikel
Neuroprotective effect of Sphingosine-1-phosphate receptor modulation in organotypic slice cultures, independent of the peripheral immune system
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Veröffentlicht: | 2. Juni 2015 |
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Gliederung
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Objective: The secondary cell death of neurons after lesions is an inflammatory mediated process involving intrinsic microglia and infiltrating immune cells. The sphingosinkinase1/Sphingosin1-phosphate (S1P)-receptor signal pathway is involved in the modulation of the immune answer of most cell populations. Here we investigated Fingolimod, the most popular member of the S1P receptor modulator without the influence of the peripheral immune system.
Method: Hippocampal organotypic slice cultures from postnatal C57Bl/6 P2-P4 pups were prepared and cultivated using the static membrane protocol in the presence or absence of Fingolimod. After 10d a pro-inflammatory stimulus using TNFalpha, Interferon gamma and LPS (Lipopolysaccarid from gram-negative bacteria) was applied. The amount of neurodegeneration in both pyramidal-cell-layers (CA1 and CA3) and the granule cell layer (dentate gyrus) was quantified by intensity measurement of propidium iodide uptake within the respective regions. After fixation, vibratom-reslicing and immunfluorescence staining for microglia and glia-cells was applied. The amount of microglia activation and astroglioses were evaluated via volumetric measurements in confocal captured stacks. Supernatant was scanned for pro- and anti-inflammatory cytokines using ELISA.
Results: Using organotypic slice culture, we were able to detect differences of the modulative effect of Fingolimod on the respective neuronal population of the hippocampus and the intrinsic cytokine response in an inflammatory milieu suggesting a direct impact on survival of neurons which is more dependent on their structure than on their environment. Volumetric measurement after treatment with Fingolimod showed a reduced glial scar in comparison to the controls.
Conclusions: Pretreatment of Fingolimod – a FDA approved medication in multiple sclerosis – seems to have a protective effect on neurons confronted with inflammation. Here we show for the first time data of this effect without the adaptive immune system, suggesting a possible role for immune modulatory therapy, for instance in treating tumour or injury related inflammation. A better understanding of the S1P receptor signal pathway could lead to a therapeutic approach reducing morbidity.