Microglia-related increase in NTPDase1 expression during EAE

Abstract

Ectonucleoside triphosphate diphosphohydrolase 1 (NTPDase1/CD39) is the main ATP- and ADPdegrading enzyme in extracellular fluid of the central nervous system. In the hydrolysis cascade NTPDase1 removes ATP and ADP and produces AMP, which is hydrolysed by ecto-5'-nucleotidase to adenosine. During neuroinflammation, increased extracellular ATP levels exert proinflammatory effects at microglia as resident immune cells, while adenosine effects are antiinflammatory. Literature data indicate involvement of purinergic signaling in experimental autoimmune encephalomyelitis (EAE), while decreased number of NTPDase1/CD39+ regulatory T-cells was evidenced in multiple sclerosis. Downregulation of NTPDase1 expression was observed in proinflammatory activation phenotype of macrophages. However, data on the role of NTPDase1 on glial cells in neuroinflammation are still scarce. We have shown increase in ATP-, ADP- and AMP-hydrolysis, together with upregulated mRNA and protein expression of NTPDase1 in lumbar spinal cord, correlated to the disease course during EAE. In this study we aimed to explore contribution of particular cell subsets to the observed changes in NTPDase1 expression. Acute monophasic EAE was induced in female rats of Dark Agouti strain by active immunization with a mixture of spinal cord homogenate in complete Freund’s adjuvant. Immunized animals were sacrificed at the onset, peak and end of symptoms, while naïve animals were used as control. Significant increase of NTPDase1 immunofluorescence in lumbar spinal cord cross-sections was related to prominent infiltrates at the peak of EAE and increased expression of NTPDase1 among isolated mononuclear cells. Analysis of triple-labeled Arginase1/NTPDase1/Iba1 and iNOS/NTPDase1/Iba1 immunofluorescent micrographs showed prevalent contribution of Arginase1+ microglia in comparison to iNOS+ microglia in NTPDase1 immunofluorescence, at the peak of EAE. Further studies are needed to reveal possible association of NTPDase1 with antiinflammatory phenotype in microglia.