RhEPO Promotes Neurological Function Recovery By Expanding Tregs In An Animal Model Of TBI

Object Inflammation contributes to secondary brain injury, resulting in neuronal apoptosis, brain edema formation and deficit of neurological function during the pathophysiological course of traumatic brain injury(TBI). It has been reported that anti-inflammatory regulatory T cells(Tregs) had the ability of immunomodulation in inflammation reaction. We hypothesize that expansion of Tregs by Recombinant human EPO (rhEPO)-induced play an important role of neuroprotective immunomodulator in TBI model. Methods:TBI was induced in adult male mice by fluid percussion injury(FPI) as a model of TBI. The TBI mice were randomly divided into sham, TBI+saline and TBI+rhEPO treatment groups. rhEPO was administered intraperitoneally1h after TBI. Animals were sacrificed24h and72h after TBI. Flow cytometry was used to detect the number of Treg cells in the spleen and immunohistochemstry to Treg cells, neutrophils, CD3positive T cells, microglias and cleaved cleave caspase-3in the injured brain tissue. ELISA was applied to measure IL-1β, TNF-α, IL-10and TGF-β in brain homogenates. The brain water content was quantified as a measure of brain edema. Brain lesions were evaluated by H.E staining at24h and72h after TBI. Morris Water Maze(MWM) was used to evaluate the cognitive function of TBI mice7to12d after FPI. Results:At72h after TBI, the number of Treg cells in the spleen was reduced in normal saline treated mice group compared to sham group. The number of Treg cells was significantly increased in mice treated with rhEPO, but not those with saline. The mice with rhEPO also markedly reduced the accumulation of neutrophils, CD3positive T cells, and activation of microglia in injured tissue, but the reduction occurred at differet time intervals after TBI. These rhEPO treated mice also had a significantly reduced number of cleaved caspase-3positive cells72h after brain injury. rhEPO treatment significantly increased the level of IL-10and TGF-β,while reduced the level of IL-1β and TNF-a as compared to normal saline treated mice and sham surgery mice. Consistent with its anti-inflammatory effects, rhEPO-treated mice also reduce brain edema24h and72h after FPI. Compared with TBI+saline group, brain lesions were also reduced in the TBI+rhEPO group72h after FPI. As a result, the cognitive function improved faster in rhEPO treated mice than normal saline treated mice. Conclusions:Animal model study have clearly indicated that Tregs, expanded by rhEPO in vivo, could effectively protect neurological function. The findings indicated that Tregs may be a potential therapy target in TBI.