Malgorzata Burek, Aiden Haghikia, Ralf Gold, Norbert Roewer, Andrew Chan and Carola Y Förster
Objective: Multiple sclerosis (MS) is a neurodegenerative disorder of the central nervous system (CNS). Damage of the blood-brain barrier integrity is a key pathogenic event leading to the migration of lymphocytes into the CNS and subsequent demyelination. This process is tightly regulated by chemokines and cytokines which are target of therapeutic strategies in MS, such as anti-inflammatory glucocorticosteroid treatment. Here, we examine the effects of dexamethasone-treatment and MS patient sera on the expression of cytokines and chemokines in brain microvascular cell line, cEND in vitro.
Methods: We conducted 96-well Mouse Cytokines and Receptors qPCR arrays to quantitatively compare the cytokine and chemokine expression profiles after treatment with dexamethasone. For selected cytokines, we studied the effects of pre-treatment with MS patient sera from active phase of disease (exacerbation) or in relapse (remission) in combination with dexamethasone.
Results: After dexamethasone treatment, colony stimulating factor 3 (Csf3) and interleukin 17F (IL17f) were significantly up-regulated, whereas the chemokine (C-C motif) ligand 12 (CCL12/MCP-5), chemokine (C-X-C motif) receptor 3 (CXCR3) and kit oncogene (Kit) were significantly down-regulated. These results were confirmed in qRT-PCR using gene-specific primers. For Csf3 and CCL12 we analyzed the dexamethasone-mediated changes in protein levels secreted into the cell culture medium. Dexamethasone treatment increased the release of Csf3 into the culture medium and decreased the release of CCL12 by cEND. Additionally, we examined the effects of MS-patient sera on dexamethasone-induced cytokine secretion. Pretreatment with MS-patient serum from exacerbation phase augmented dexamethasone effects on Csf3 and CCL12 release in cEND cells. The expression of Csf3- and CCL12-receptors was demonstrated on protein and mRNA level in cEND cells.
Conclusion: We identified Csf3 (G-Csf) and CCL12 as cytokines differentially regulated by dexamethasone on mRNA and protein level. This effect was even more pronounced after pretreatment with MS patient serum, especially from patients with acute relapses.