| Purpose: Incranial infection is one of the most common causes of epilepsy. The cytokines and chemokines may play a key role in the development of infection-related epilepsy. This study aims to observe the expression change of IL-1β, TNF-α and CX3CL1-CX3CR1, and the sensitivity to Lithium chloride – Pilocarpine(Licl-Pilo) as well as the pathological change in the brain of LPS induced intracranial infection rat model. We also aim to further explore the impact of exogenous CX3CL1 and anti- CX3CL1 antibody on the epileptogenicity of Lithium-pilocarpine and the pathological change in this intracranial infection rat model.Methods: 32 adult male SD rats were randomly divided into LPS treated group and control group with 16 rats in each. The rats in the LPS treated group were intracerebroventricularly injected with LPS of 50ug/5ul, whereas, the rats in the control group were intracerebroventricularly injected with the same volume of normal saline. 24 hours later, 10 rats in each group were selected randomly and administered intraperitoneally with 10% chloral hydrate(3.3ml/kg). After successfully anaesthetized, part of the rats were decapitated and the proteins in the hippocampal tissue were extracted. The quantity of IL-1β and TNF-α were measured with ELISA, and the expressions of CX3CL1 and CX3CR1 in the hippocampal tissue were tested by Western-blot. The other part of rats were perfused and the brains were taken out to make paraffin sections for immunohistochemical study. The remaining 6 rats in the LPS treated group and control group were used to make status epilepticus(SE) by lithium chloride-pilocarpine, and the latency to SE and the seizure severity were observed behaviorally and electrophysiologically.Another 30 adult male SD rats were selected and randomly divided into CX3CL1+LPS group, anti- CX3CR1 antibody + LPS group, and Na Cl+LPS group with 10 rats in each. All rats in the three groups were placed a catheter through the lateral ventricle and the LPS of 50μg/5μl was injected into the ventricle by way of this catheter to build intracranial infection model, followed by lateral ventricular injection with 5ul CX3CL1 or anti- CX3CR1 antibody or normal saline every 10 hours for 3 days. Then, Lithium chloride – Pilocarpine was used to induce SE, and the latency to SE and the seizure severity were observed behaviorally and electrophysiologically. In addition, the pathological change in the brains were examined by immunohistochemistry.Results:(1) Compared with the control group, the rats in LPS treated group showed a reduction in daily activity and food intake, and the inflammatory cytokines of IL-1β and TNF-α in hippocampus rose remarkably and reached to the statistically significant level(p<0.001). In addition, the rats in LPS treated group became more sensitive to Licl-Pilo. They had much shorter latency of SE and severer seizure(p<0.001) which were demonstrated by behavior and electrophysiology.(2) Western blot finding showed that the chemokine of CX3CL1 and CX3CR1 in the tissue of hippocampus in LPS group increased significantly when compared with that of control group(p<0.05).(3) Immunohistochemistry study disclosed in the LPS group that the number of neurons in the cortex decreased obviously, and the remaining neurons were unintact. Meanwhile, the arrangement of neurons in the hippocampus CA1 area became loose and disorganized, and the neurons’ shape became blur and their stain became lighter. The differences had statistical significance when compared with the control group(p<0.001). In addition, the cortex and hippocampus CA1 area emerged in large number of proliferated and activated microglia cells in the rats of LPS group when compared with control group, and the difference was significant(p<0.001)(4) Lateral intracerebroventricular injection of exogenous CX3CL1 and CX3CR1 antibody displayed by behavior and electrophysiology that CX3CR1 antibody prominently lengthened the latency of SE(p<0.01), whereas, CX3CL1 only had a tendency to shorten the latency of SE, but with no significant difference(p >0.05) when compared with simply LPS treated rats, respectively.(5) Immunohistochemical study showed that there much more neurons in cortex and adjacent hioppocampus CA1 area disappeared in the rats of CX3CL1 + LPS group, but more neurons kept in the rats of CX3CR1 antibody + LPS group, and their differences got to a significant level(p<0.01)When compared with Nacl+LPS group. In addition, the microglia cells in cortex became more active, but had no obvious change in quantity when injecting CX3CL1 together with LPS. On the other hand, the microglia cells became less active, and decreased in quantity when injecting CX3CR1 antibody together with LPS, and the differences had statistical significance when compared with Nacl+LPS group(p<0.01)Conclusions:(1) LPS can result adult rat in intracranial infection. This kind of infected rat has decreased number of neurons, but has more activated microglia cells in both cortex and hippocampus CA1 area, and becomes more sensitive to Licl-Pilo in that the rat will suffer severer seizure when compared with no LPS treated rat.(2) The LPS infected rats have increased expression of CX3CL1 and CX3CR1 in the hippocampus.(3) Lateral intracerebroventricular injection of recombinant CX3CL1 has no obvious effect on the sensitivity to the Licl-Pilo induced SE, but aggravates brain injury with increased activity of microglia cells and decreased number of neurons in the hippocampus CA1 area.(4) Lateral intracerebroventricular injection of CX3CR1 antibody can reduce sensitivity to seizure, relieve the degree of attack, and inhibit the activity of microglia cells and reduce the loss of neurons in cortex and hippocampus CA1 area. |
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