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Objective To investigate the influence of necroptosis pathways in microglia activation and inflammatory factor levels in cerebral ischemia/reperfusion (I/R) injury in rats and their significances.Methods A modified suture method was used to establish the models of middle cerebral artery I/R in rats.(1) SD rats according to the random number table were divided into cerebral I/R 6 h group,cerebral I/R 12 h group,cerebral I/R 24 h group,and cerebral I/R 72 h group (n=6);the expression of ionized calcium binding adaptor molecule-1 (iba-1) was detected by immunohistochemical staining,and time points enjoyed obvious iba-1 expression were selected according to the experimental results.(2) SD rats were randomly divided into sham operation group,cerebral I/R group,80 nmol necrostatin-1 (Nec-1) intervention group and 160 nmol Nec-1 intervention group (n=6),and the Nec-1 intervention was given 2 h after ischemia;Longa method was used to evaluate the neurological function scores and TTC method was used to detect the infarct volume;and the appropriate dosages of Nec-1 were selected according to these results.(3) SD rats were randomly divided into sham operation group,cerebral I/R group,solvent group and Nec-1 intervention group (n=6),and Nec-1 or DMSO intervention was given 2 h after ischemia.HE staining was used to observe the survival and proliferation ofmicroglias around the infarction tissues;immunohistochemical staining was used to detect the iba-1 expression surrounding the infarction tissues;immunohistochemistry and Western blotting were employed to observe the cytokine tumor necrosis factor (TNF)-α and interleukin (IL)-1β and glia-derived neurotrophic factor (GDNF) expressions.Results (1) The iba-1 expression at cerebral I/R 24 h group was significantly increased as compared with that in other groups (P＜0.05).(2) As compared with those in the cerebral I/R group,the neurological function scores and infarct volume were significantly decreased in the 80 nmol Nec-1 intervention group and 160 nmol Nec-1 intervention group (P＜0.05);more obvious changes were noted in the 160 nmol Nec-1 intervention group as compared with those in the 80 nmol Nec-1 intervention group,with significant difference (P＜0.05).(3) HE staining showed peri-infarct tissue inflammatory cell infiltration,reduced neuron number,cell body shrinkage and nuclear pyknosis in the cerebral I/R group,while these changes were less obvious in the Nec-1 intervention group;as compared with cerebral I/R group,the Nec-1 intervention group had significantly decreased expressions of iba-1,TNF-α,IL-1 β and GDNF (P＜0.05).Conclusion Necroptosis pathway involves in the activation ofmicroglias after I/R injury in rat brains;Nec-1 inhibits the activation of microglia,and reduces the neuronal damage by regulating inflammatory cytokines.
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Objective To investigate the expression of GDNF in the olfactory ensheathing glia cells of the adult rats and golden hamster for studying the mechanisms of the OECs in the CNS regeneration. Methods The immunohistochemical staining was performed on the slides of adult olfactory bulb from 3 rats and 3 golden hamsters.According to the primary antibody,tissue slides were divided into 3 groups:rabbit polyclonal GDNF,the rabbit polyclonal GFAP and rabbit polyclonal NGFRp75(Santa Cruz Biotechnology)respectively,and followed by the rabbit ABC immunoreactive staining system. Results It revealed that GDNF was expressed by cells in the first two lines of olfactory bulb from rat and golden hamster.The GFAP and NGFRp75 were also expressed in the same position from other two groups of slides respectively.Conclusion The olfactory ensheathing cell could secret GDNF in the progress of mediating neuronal survival and axonal elongation.