Abstract
Purpose::
T cell-mediated protective autoimmunity has been shown to attenuate secondary degeneration following primary insult to the central nervous system (CNS). In animals with normal immune function, such neuroprotection can be induced via activation or passive transfer of CD4+ T cells reactive to self-antigens expressed at the injury site or through suppression of CD4+CD25+ regulatory T cells (Treg). This study was conducted to examine the effect of the bacterial nucleotide motif CpG on Treg-mediated suppression of CD4+CD25- effector T cells (Teff) and its potential for neuroprotection in an optic nerve crush injury model.
Methods::
The effect of CpG on murine Treg suppressive activity was studied in vitro by separating T cells into Teff and Treg using autoMACS. Treg were incubated with CpG, non-bacterial GpC or phosphate-buffered saline (PBS), then washed and co-cultured with Teff. Proliferation assays were performed. The ability of CpG to confer neuroprotection was tested in vivo using an optic nerve crush model in Lewis and athymic rats. The right optic nerve was crushed using calibrated forceps. Concurrently, animals were injected with 100µg CpG, 100µg GpC or PBS. Two weeks later, retinal ganglion cells (RGCs) were retrogradely labelled. Retinas were whole mounted and RGCs were counted by a masked investigator. Optic nerves were assessed for CD3 staining of T cells and neurofilament staining.
Results::
Treatment with CpG resulted in a downregulation of Treg suppressive activity and significantly (p<0.008) increased proliferation of Teff. GpC did not have any effect. In optic nerve crush of Lewis rats, CpG treatment resulted in a significantly increased RGC survival(166±27cells/mm2, mean±SEM) compared to either PBS (78±10 cells/mm2, p=0.003) or GpC (70±15 cells/mm2, p<0.01), and a significant (p<0.01) upregulation of T cell infiltration in the optic nerve (80±10 cells/mm2) compared to either PBS (43±6 cells/mm2) or GpC (40±4 cells/mm2). CpG did not confer neuroprotection in athymic rats compared to controls.
Conclusions::
Bacterial DNA suppresses Treg activity and confers T cell-dependent neuroprotection following CNS injury. These findings demonstrate that regulating the immune response to CNS insult and neurodegenerative disease improves neuronal survival. This study may lead to the development of new therapeutic modalities for neuroprotection which target the immune system to benefit the injured CNS.
Keywords: neuroprotection • immunomodulation/immunoregulation • optic nerve