Memory cells specific for myelin oligodendrocyte glycoprotein (MOG) govern the transfer of experimental autoimmune encephalomyelitis

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Abstract

Multiple sclerosis (MS) is an inflammatory disease of the CNS mediated by CD4+ T cells directed against myelin antigens. Experimental autoimmune encephalomyelitis (EAE) is induced by immunization with myelin antigens like myelin oligodendrocyte glycoprotein (MOG). We have explored the transfer of EAE using MOG35-55-specific TCR transgenic (2D2) T cells. Unsorted 2D2 Th1 cells reliably transferred EAE. Further, we found that CD44hiCD62Llo effector/memory CD4+ T cells are likely responsible for the disease transfer due to the up-regulation of CD44. Given the importance of MOG in MS pathogenesis, mechanistic insights into adoptively transferred EAE by MOG-specific Th1 cells could prove valuable in MS research.

Introduction

Myelin oligodendrocyte glycoprotein (MOG), located on the outermost lamellae of the myelin sheath and on the surface of oligodendrocytes, is highly conserved among species (Johns and Bernard, 1999). Although MOG is a quantitatively minor component of CNS myelin, there is strong evidence that MOG serves as an autoantigen and plays an important role in the pathogenesis of multiple sclerosis (MS) (Sun et al., 1991, Kerlero de Rosbo et al., 1997, Egg et al., 2001). T cells from MS patients readily proliferate and secrete IFN-γ in response to MOG peptides relative to other myelin antigens such as myelin basic protein (Kerlero de Rosbo et al., 1997). In addition, anti-MOG IgG-secreting B cells are more abundant in both the peripheral blood and cerebrospinal fluid of MS patients compared to controls (Sun et al., 1991), suggesting a role for MOG in the pathogenesis of MS.

Experimental autoimmune encephalomyelitis (EAE) is the most frequently studied animal model of multiple sclerosis (MS) and can be credited for the development of several currently approved therapies (Zamvil and Steinman, 2003). EAE shares many clinical and histopathological features with MS and is mediated by myelin-reactive CD4+ T cells. EAE is induced in genetically susceptible mouse strains by immunization with myelin antigens including myelin basic protein (Zamvil et al., 1986), proteolipid protein (Tuohy et al., 1989), and MOG (Mendel et al., 1995). MOG35-55 is the major immunodominant epitope of MOG and elicits neurological impairment following immunization with MOG35-55 and adjuvants (Mendel et al., 1995). Although the active induction of EAE is an important tool in MS research, it introduces many confounding factors such as adjuvants, which are not present in the human disease. To circumvent the drawbacks of active immunization, models of passive immunization have been developed and used for decades (Pettinelli and McFarlin, 1981). In adoptively transferred EAE, lymphoid cells are harvested from the myelin antigen immunized mice, stimulated ex vivo and transferred, without the use of adjuvants, to the recipient mice.

In 2003, a MOG35-55-specific TCR transgenic (2D2) mouse was reported in which transgenic T cells were not deleted or tolerized, and were found to be fully competent. The unique feature of this mouse strain was that more than 30% of mice spontaneously developed optic neuritis without clinical or histological evidence of EAE. Only 4% of 2D2 mice were reported to develop spontaneous EAE (Bettelli et al., 2003). Since 2003, substantial difficulties have been encountered in achieving successful transfer of EAE using 2D2 donor cells. In 2009, Jäger et al. reported that naïve (CD4+CD62L+) 2D2 T cells, following primary differentiation in Th1 or Th17 culture conditions, were unable to transfer disease reliably. However, when those cells were restimulated, both Th1 and Th17 cells were able to transfer EAE indicating that secondary T cell stimulation is a critical event in establishing encephalitogenicity (Jäger et al., 2009). Given the importance of MOG in EAE and MS, using 2D2 T cells to induce EAE could provide insight into the development and pathogenesis of MS.

The progression of MS and EAE has been attributed to several properties of T cells. Both IFN-γ-producing Th1 and IL-17-secreting Th17 cells are thought to contribute to the advancement of CNS pathology in EAE and MS (Stromnes et al., 2008). More recently, the Th1 transcription factor, T-bet has been shown to be essential for the encephalitogenicity of T cells (Yang et al., 2009). T-bet expression is restricted to Th1 cells and controls the induction of IFN-γ, repressing IL-4, creating a Th1-biased environment for the perpetuation of an inflammatory response (Szabo et al., 2000). Myelin-reactive T cells from MS patients more readily proliferate in response to antigen in the absence of costimulation compared to healthy controls, suggesting that T cells from MS patients have previously been activated and have a memory phenotype (Lovett-Racke et al., 1998). In addition to accelerated activation, memory cells are able to more readily extravasate across endothelial barriers as they express high amounts of surface CD44 (DeGrendele et al., 1996). This suggests that memory cells are well poised to access the CNS parenchyma and initiate inflammation.

We examined the adoptive transfer of EAE using Th1- and Th17-differentiated 2D2 cells. We demonstrate that IFN-γ-producing, T-bet+ Th1 2D2 cells are capable of inducing EAE. Further, we show that memory T cells are necessary, and likely responsible, for the transfer of disease following Th1 differentiation. While our results are consistent with those of Jäger et al. (2009), we provide here a more direct method of transferring EAE using 2D2 Th1 cells. Additionally, we observed that Th1-differentiated 2D2 cells from male mice transfer more severe disease into sex-matched WT recipients than their female counterparts, corresponding to a difference in the number of memory cells between the sexes.

Section snippets

Mice

C57Bl/6 mice were purchased from The Jackson Laboratory. 2D2 mice were a kind gift from Dr. Vijay Kuchroo (Center for Neurologic Disease, Brigham and Women's Hospital, Harvard Medical School, Boston, MA). Mice were housed in a specific pathogen-free animal facility at The Ohio State University on a 12-h light/dark cycle and given food and water ad libitum. All 2D2 mice were screened using flow cytometric analysis of PBMCs using specific antibodies to Vα3.2 and Vβ11 (BD Biosciences). Mice used

2D2 T cells are able to infiltrate the CNS and induce severe EAE

2D2 transgenic mice, with an over-representation of T cells specific for MOG35-55, were first described by Bettelli et al. (2003). A high proportion of these mice demonstrated optic neuritis, but only 4% of 2D2 mice were reported to develop spontaneous EAE (Bettelli et al., 2003). To examine the ability of 2D2 cells to initiate EAE, WT and 2D2 mice were immunized with MOG35-55 and adjuvants. Following immunization, 2D2 mice developed more severe clinical signs of EAE and earlier onset compared

Discussion

Multiple sclerosis is a complex neuroimmune disorder perpetuated by the targeting of myelin antigens by self-reactive CD4+ T cells. Although myelin reactive T cells have been found in healthy individuals and MS patients, those found in MS patients have an activated, memory phenotype (Lovett-Racke et al., 1998). This suggests that effector/memory T cells may be a fundamental component in MS pathogenesis. MOG is a prominent neuroantigen in MS pathogenesis and by using a MOG35-55-specific TCR

Disclosure

The authors have no financial conflicts of interest.

Acknowledgements

We thank Dr. Vijay K. Kuchroo for his generous gift of 2D2 mice. In addition, we would like to thank Drs. Amy E. Lovett-Racke and Michael Racke for their helpful scientific discussion and Ingrid Gienapp for her outstanding technical support.

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