Studies on Tissue Protective Cytokines in Remyelination and Regenerative Medicine

Abstract

There is growing interest in the tissue-protective effects of some cytokines, including erythropoietin (EPO) and the IL-6 family cytokine leukaemia inhibitory factor (LIF); both have receptors, and exert their effects, on cells other than their primary targets. In the nervous system, these cytokines could aid in the treatment of demyelinating diseases, such as multiple sclerosis, by protecting myelin from damage and supporting remyelination after damage has occurred. Previous work has shown that EPO increases myelination in oligodendrocytes, the cells responsible for myelin deposition in the central nervous system. I aimed to determine if LIF shares the promyelinating effects of EPO and understand more about the mechanisms mediating tissue-protective cytokine-induced myelination. A model of rat oligodendrocyte precursor cells was used and their myelinating capacity was measured as represented by myelin oligodendrocyte glycoprotein (Mog) expression. Initially I studied EPO and LIF’s effects on these cells before defining the molecular mechanisms causing their effects using microarray gene expression analysis. EPO increased myelination by eight-fold, a level that was sustained at concentrations up to and including 400ng/ml. After treatment with LIF at 0.2ng/ml Mog expression was increased by two-fold, but concentrations above 2ng/ml caused a reduced expression of Mog. Interestingly, when LIF and EPO were added simultaneously there was a significant reduction in EPO-induced Mog expression suggesting that LIF induced an inhibitory feedback that was responsible for blocking not only its own, but also EPO’s effect. The inhibitory feedback was replicated when LIF was replaced by ciliary neurotrophic factor (CNTF) and oncostatin M (OSM), glycoprotein 130 (GP130) cytokines that use the same receptor as LIF. The signalling mechanisms that may have caused the inhibition of EPO-induced Mog were then investigated. Socs3, a known inhibitory feedback of LIF and other IL-6 cytokines, negatively correlated with Mog expression, as the higher concentration of LIF and the simultaneous EPO and LIF treatment induced the greatest Socs3 expression. Gene expression microarray analysis was performed to elucidate further mechanisms that may cause the inhibition of EPO-induced Mog. A variety of candidate genes were identified and their expression validated by qPCR. The roles of Tlr2/Myd88 and lipocalin 2 were investigated further and Tlr2 activation showed a functional effect on Mog expression. The results showed that LIF and other GP130 cytokines inhibited EPO’s positive effect on myelination and clarified some of the mechanisms that resulted in inhibition. The implications of my work could be an increase in efficacy of EPO treatment, as the work has elucidated mechanisms that could inhibit EPO’s promyelinating effect. Increased efficacy of EPO would impact new therapies and therapeutic approaches using tissue protective cytokines in regenerative medicine.

Date of Award	Jul 2016
Original language	English
Awarding Institution	University of Brighton