Contributions
Abstract: 59
Type: Oral
Abstract Category: Pathology and pathogenesis of MS - Genetics /Epigenetics and Pharmacogenetics
Background: Genetic analysis has identified more than 100 single nucleotide polymorphisms (SNPs) that are associated with increased susceptibility to multiple sclerosis (MS) including rs74796499 (p=2.4 x 10-20) which maps to chromosome 14q3 in a region containing the genes for the enzyme galactosylceramidase (GALC) and the G-protein coupled receptor 65 (GPR65). GALC is found in lysosomes, where it hydrolyses glycolipid including psychosine. GPR65 is a pH sensitive receptor for the glycosphingolipid psychosine, and is believed to plays an important role as an immune response inhibitor in an acidic microenvironment. Given that pH is known to be acidic within MS lesions we hypothesized that the rs74796499 genotype might influence GPR65 dependent inhibition of an inflammatory reaction.
Methods: To explore the role of GPR65 in the regulation of an immune reaction we collected peripheral blood mononuclear cells (PBMCs) from 100 healthy controls (50 homozygous for the common allele and 50 carrying the MS protective allele) and measured activation markers (CD25 and CD69) in T cells and NK cells using flow cytometry. We assessed activation after culturing PBMCs for 18 hours in stimulated (using anti-CD3 and anti-CD28 antibodies) and unstimulated samples, both in the presence and the absence of the GPR65 specific ligand (BTB09089) and at different pHs. We also explored the effects of genotype on the ex-vivo surface expression of the glycosphingolipid lactosylceraminde (CD17) on PBMCs.
Results and discussion: Our results to date show that human T cell activation is pH dependent with the greatest activation occurring at pH7.0, which is 2.2 times higher than activation at pH6.2 (p=0.0007). The GPR65 ligand BTB09089 is an efficient inhibitor of such activation, especially at acidic pH. The effect of genotype on the function of GPR65 and GALC is now being analysed. Our study will expand our understanding of the functional consequences of GPR65 and GALC in MS.
Disclosure:
W. Liao is funded by The Multiple Sclerosis Society
M. Ban: nothing to disclose
B. Fiddes: nothing to disclose
L. Azzopardi: nothing to disclose
D. He: nothing to disclose
S. Sawcer: nothing to disclose
Abstract: 59
Type: Oral
Abstract Category: Pathology and pathogenesis of MS - Genetics /Epigenetics and Pharmacogenetics
Background: Genetic analysis has identified more than 100 single nucleotide polymorphisms (SNPs) that are associated with increased susceptibility to multiple sclerosis (MS) including rs74796499 (p=2.4 x 10-20) which maps to chromosome 14q3 in a region containing the genes for the enzyme galactosylceramidase (GALC) and the G-protein coupled receptor 65 (GPR65). GALC is found in lysosomes, where it hydrolyses glycolipid including psychosine. GPR65 is a pH sensitive receptor for the glycosphingolipid psychosine, and is believed to plays an important role as an immune response inhibitor in an acidic microenvironment. Given that pH is known to be acidic within MS lesions we hypothesized that the rs74796499 genotype might influence GPR65 dependent inhibition of an inflammatory reaction.
Methods: To explore the role of GPR65 in the regulation of an immune reaction we collected peripheral blood mononuclear cells (PBMCs) from 100 healthy controls (50 homozygous for the common allele and 50 carrying the MS protective allele) and measured activation markers (CD25 and CD69) in T cells and NK cells using flow cytometry. We assessed activation after culturing PBMCs for 18 hours in stimulated (using anti-CD3 and anti-CD28 antibodies) and unstimulated samples, both in the presence and the absence of the GPR65 specific ligand (BTB09089) and at different pHs. We also explored the effects of genotype on the ex-vivo surface expression of the glycosphingolipid lactosylceraminde (CD17) on PBMCs.
Results and discussion: Our results to date show that human T cell activation is pH dependent with the greatest activation occurring at pH7.0, which is 2.2 times higher than activation at pH6.2 (p=0.0007). The GPR65 ligand BTB09089 is an efficient inhibitor of such activation, especially at acidic pH. The effect of genotype on the function of GPR65 and GALC is now being analysed. Our study will expand our understanding of the functional consequences of GPR65 and GALC in MS.
Disclosure:
W. Liao is funded by The Multiple Sclerosis Society
M. Ban: nothing to disclose
B. Fiddes: nothing to disclose
L. Azzopardi: nothing to disclose
D. He: nothing to disclose
S. Sawcer: nothing to disclose