In vitro investigation into the potential regulating role of sphingolipids in lymphocyte proliferation.

Sphingosine-1-phosphate (S1P) is a bioactive lipid involved in a wide range of
cellular processes, such as cell migration, differentiation and protection from
apoptosis. S1P has both extracellular effects through five G-coupled receptors
(S1PR1-5) as well as intracellular effects by interactions with target
proteins. Lymphocytes have important functions in specificity, diversity,
self-tolerance and immunological memory. The egress and recirculation of
lymphocytes from thymus and secondary lymphoid organs depends on the S1PR1
receptor. Release of T-cells from the thymus and the interaction between T- and
B-lymphocytes in secondary lymphoid organs is essential for the functioning of
the adaptive immune system. Activation of S1PR1 on the lymphocyte membrane
leads to lymphopenia, probably due to sequestration of lymphocytes in secondary
lymphoid organs, while entering into lymphoid organs depends on loss of S1PR1
due to internalization. Since activation of S1PR1 leads to lymphopenia,
development of S1P agonists seems promising for immunosuppressive therapy in
transplantation medicine and autoimmune disease.

More sustained knowledge of the intracellular processes regarding S1P in
lymphocytes could be beneficial in modulating specific immune responses in
transplantation medicine, autoimmune diseases and sepsis. Therefore, in this
study, we will investigate S1P induced lymphocyte proliferation and its
underlying cellular mechanisms.

In vitro met vers geïsoleerde lymfocyten . in vitro eindpunten zijn de
vaststelling van een merkbare verandering van de proliferatiesnelheid van de
lymfocyten gedurende 72 uur in aanwezigheid of afwezigheid van specifieke
agonisten of antagonisten voor de receptoren en sfingosine - substraten of
remmers van de sfingosine - kinase en - transporter (de ABC C1 - transporter )
. Derhalve zal het aantal cellen schatting door het meten van fluorescentie
signaalintensiteit via de NF celproliferatie Assay Kit ( Invitrogen Molecular
Probes , Oregon , USA ) . De gemeten fluorescentiesignaal is een continue
variabele die wordt gebruikt als een surrogaat voor het aantal cellen in het
experiment . Statistisch verschil wordt berekend met behulp van T - toets van
de student voor onafhankelijke monsters ( in het geval waarin twee groepen
vergeleken worden ) of een One - Way ANOVA met post - hoc Games - Howell ( in
het geval waarin drie of meer groepen zijn vergeleken ) . Gegevens zullen
worden geanalyseerd met behulp van de meest recente beschikbare versie van SPSS
voor Windows -werkstation

The study poses a negligible risk for the volunteer, as the only intervention
will be drawing of blood at one single occasion. Increasing our fundamental
knowledge about the regulating role of sphingolipids on lymphocyte
proliferation might lead to the discovery of novel targets for pharmacological
intervention that will allow modulation of lymphocyte function. The results
could contribute to a better understanding of cellular mechanisms in adaptive
immunity which potentially allow targeted immunomodulation in transplantation
medicine, autoimmune diseases and sepsis.