| objective:To explore the effects of Recombinant Human Erythropoietin (r-HuEPO)on neuronal degeneration, cell proliferation and mossy fiber sprouting in hippocampus dentate gyrus of adult rat after status epilepticus. Methods:(1) Modeling and grouping:the 140 healthy adult male Sprague-Dawley(SD) rats were randomly divided into normal control group (20 rats) received intraperitoneal injections of an identical volume of normal saline. Each time the number of experimental animals in each group were 4 and hippocampus dentate gyrus was selected for study; model of temporal lobe epilepsy was produced in the left (120 rats) by injecting lithium chloride-pilocarpine intraperitoneally and the extent of seizure was graded according to the standard of Racine, rats that epileptic state continued for more than or equal to 1h were selected and then randomized into r-HuEPO+status epilepticus group(SE+r-HuEPO group,intervention group) and SE group (no intervention group), r-HuEPO was injected intraperitoneally once at a dose of 5000IU in each rat in the intervention group 6h after SE, The time after injection of r-HuEPO, all rats received injections of 5-bromodeoxyuridine (BrdU) (50 mg/kg, bid, i.p, interval 4h)for 6 days, the survivals were eventually used for further study. (2) On the 4th day after modeling, the Fluoro-Jade B (FJ-B) immunofluorescence technology was adopted to observe the neuronal degeneration in dentate gyrus. (3) On the 9th day after modeling, BrdU immunohistochemistry test was used to observe the proliferation of cells in the dentate gyrus; On the 14th day, Timm's staining technology was adopted to observe mossy fiber sprouting in dentate gyrus. (4) On the 28th day, rats were sacrificed for both BrdU labeled immunohistochemistry test and Timm's staining. Results:(1) The success rate of SE induced in experimental rats was 88.57% after intraperitoneally injecting lithium-pilocarpine, the mortality rate of SE+ r-HuEPO group was 68.33%, and the SE group was 71.67%; Comparing the SE+r-HuEPO group and the SE group, x2=0.690, P>0.1, the difference was not statistically significant. (2) Results of FJ-B staining: the fourth day after SE, degeneration of neurons in the dentate gyrus in SE group (no intervention group) was more serious than the SE+ r-HuEPO group (intervention group) and the control group, which mainly located in the SGZ and the dentate hilus, comparing the SE group (32.32±5.09) and the control group (6.50±1.08), P＜0.01, the difference was statistically significant. Comparing the SE group and the SE+r-HuEPO group (11.12±0.75), P＜0.01, the difference was statistically significant. Comparing the SE+r-HuEPO group and the control group, P>0.05, the difference was not statistically significant. (3) Results of BrdU immunohistochemical staining:the 9th day after SE, BrdU immunoreactive cells were significantly increased in the SE group,which mainly located and clusteredly distributed in the SGZ. But the number was not significantly increased in the SE+r-HuEPO group,which still mainly distributed scatteredly in the SGZ. In the control group, there were very small amount of immunoreactive cells scattered in the SGZ. Comparing the SE group (92.8±2.97) and the control group (12.80±2.13), P＜0.01, the difference was statistically significant. Comparing the SE group and the SE+r-HuEPO (16.95±0.54) group, P＜0.01, the difference was statistically significant. Comparing the SE+r-HuEPO group and the control group, P>0.05, the difference was not statistically significant. On the 28th day, the number of BrdU immunoreactive cells in the SE group further increased and many migrated to the hilus, part of which "distributed in the granule cell layer and inner molecular layer; although the number of BrdU immunoreactive cells in the SE+r-HuEPO group increased and also some migrated to the hilus, but the increase was not obvious; the number increased in the control group was not obvious and still mainly located in the SGZ. Comparing the SE group (146.45±5.91) and the control group (38.52±3.10), P<0.01,the difference was statistically significant. Comparing the SE group and the SE+r-HuEPO group (46.27±2.93), P＜0.01, the difference was statistically significant. Comparing the SE+r-HuEPO group and the control group, P>0.05, the difference was not statistically significant. (4) Results of Timm's staining: 14 days after SE, Timm's staining granules in each group has not increased significantly, but comparing the SE group (0.45±0.19) and the control group (0.10±0.11), P＜0.05, the difference was statistically significant. On the 28th day, mossy fiber sprouting in the SE group was significant and connected to form a dense band in the inner molecular layer; the score in the SE+r-HuEPO group has also increased, but the difference between the control group and the SE+r-HuEPO group was not significant. Comparing the SE group (3.60±0.16) and the control group (0.30±0.11), P＜0.01, the difference was statistically significant. Comparing the SE and the SE+r-HuEPO group (0.55±0.10), P＜0.01, the difference was statistically significant. Comparing the SE+r-HuEPO group and the control group, P>0.05, the difference was not statistically significant. Conclusions (1) Neuronal degeneration and cell proliferation can be induced in dentate gyrus after SE in rats, and a large number of newborn cells migrate to the hilus, part of which migrate to the inner molecular layer. SE can also result in mossy fiber sprouting of nerve cells in the dentate gyrus, which extends into the inner molecular layer. (2) r-HuEPO can't reduce statistically the mortality rate in the 7-8W old male SD rats after the onset of status epilepticus 6h later. (3) r-HuEPO can reduce the extent of neuronal degeneration and the number of cell proliferation and inhibit mossy fiber sprouting in the dentate gyrus after the onset of status epilepticus in rats.
|
PDF Full Text Request