RhEPO Up-regulates The Level Of Circulating EPCs And Promotes Neuro-vascular Repair In Injured Brain Tissue

Objective: To explore the inducing effect of the recombinant human erythropoietin (rhEPO) on the level of circulating endothelial progenitor cell (EPCs) in the Wistar rat models of traumatic brain injury(TBI), on the neuro-vascular repairing effect and on the potential cellular mechanisms.Methods: 174 health male Wistar rats (8-week-old, weight: 300g-350g) were divided randomly into group I (n=144) and group II (n=30). Group I was further divided into three subgroups: Sham injury, TBI~+Saline and TBI~+rhEPO respectively (n=48 for every group). To assay the level of the circulating EPCs, 6 investigating time points were set as 3h, 6h, 24h, 3d, 7d, 14d post-trauma and each point contains 8 rats. Gorup II was also divided into tree subgroups: Sham injury, TBI~+Saline and TBI~+rhEPO respectively (n=10 for every group). The TBI rat models were made by attacking the opened cranial of the rats with a fluid-percussion power (1.5-2.0atm) of the fluid-percussion craniocerebral injury instrument. Blood samples of the rats were collected from their posterior orbits and the brain tissues were collected at each corresponding time point after trauma. Marked with both positive CD34 and CD133 antibodies, the level of the circulating EPCs was assayed in the flow cytometry instrument. The immunofluorescence staining was also performed to investigate the expression changes of CD31~+ on hippocampus and cortex of the injured brain tissue, which is the marker of the endothelial cells of the matured vascular indicating a degree of the vascular repairing. The spatial learning and memory ability of the rats from the subgroups of group II was assessed to assay the rhEPO therapeutic effect on the neuro-function recovery of the TBI rats.Results: 1. Compared with sham group, in general, the level of circulating EPCs in TBI~+Saline and TBI~+rhEPO Wistar rats post-TBI decreased immediately, then increased later. The level of EPCs of TBI~+rhEPO rats were significantly higher than the one of the TBI~+Saline 6h post-TBI, the former retained high until 7d and then decreased to normal level several days later. However, the level of the EPCs of TBI~+Saline rats peaked at 6h after trauma and decreased to a normal level thereafter. 2. The immunofluorescence staining of CD31~+ in cortex and hippocampus tissues was stronger in TBI~+Saline and TBI~+rhEPO rats than in sham rats 1day post-TBI, however, no significant difference was detected between the TBI+Saline and TBI+rhEPO rats. Another 6 days later(7 d post-trauma), the CD31 expression in brain tissue from both TBI+Saline and TBI+rhEPO rats was more significant than the sham rats, with the highest one in the TBI+rhEPO rats when compared with the TBI+Saline rats (P<0.05). 3. In general, the percent time spending in target quadrant increasesd following the days practiced in the Morris water maze test. Compared with the TBI+Saline rats, the percent time spending in the target quadrant of the TBI+rhEPO rats increased apparently from the fourth days on (P<0.05), which indicates that the rats treating with rhEPO improved their ability of spatial learning and memory. In addition, the total neurological severity scores of the TBI+rhEPO rats were lower than TBI+Saline rats, even much lower on 21d post-trauma, which also indicates that the rats treating with rhEPO improved their severity of TBI rats.Conclusion: rhEPO could increase the number of circulating EPCs, promote the neurovascular repair in injured brain tissue and improve spatial learning ability and memory functionh of the TBI rats. It may be a potential therapeutic drug to cure TBI.