| Background and purposeThere are approximate 15 million premature born every year, and premature infants have become the leading cause of newborn deaths. In recent years, with the development of perinatal medicine science and the improvement of technical of intensive care, premature mortality was significantly lower than before, the incidence of neurological complications in preterm infants were on the rise. Infection is one of the important causes of brain damage. Research has shown that there were obvious correlation between infection / inflammation and the development of neonatal brain damage. At present effective treatment is still lacking.It has become a worldwide problem of public health,so we should find intervention ways of brain damage in preterm infants, and reduceing the cognitive and motor dysfunction is particularly important.Caffeine citrate has been widely used in clinical apnea treatment in premature infants as a non-selective adenosine receptor antagonist. Studies have shown that caffeine can not only treat apnea in premature infants,but also improve the chances of withdrawal and reduce the incidence of complications such as BPD, and the incidence of cerebral palsy and abnormal behavior is reducing according to long-term follow-up results. Animal experiments showed that caffeine citrate can reduce the apoptosis of neuron in the hippocampus with hypoxic-ischemic brain damage and improve cognitive impairment and the long-term function of learning and memory. Currently the research of the effect of caffeine is less in brain damage caused by infection in the perinatal period. In this study, perinatal brain damage due to infectious was induced by intraperitoneal injection of LPS 0.6 mg/kg to 2-day-old newborn S-D rats, and treated with caffeine citrate. The purpose was to investigate the protective effect of caffeine citrate in perinatal brain damage caused by infection.Materials and methods 2-day-old neonatal SD rats were randomly divided into control group(group A), LPS infection group(group B), caffeine citrate treatment group(group C),each group of 24. Group B and C were continuously intraperitoneally injected of LPS 0.6 mg/kg for 5 days from 2 days old, and group A was continuously intraperitoneally injected of equal volume of saline. Group C was continuously intraperitoneally injected of caffeine citrate 10 mg/kg for 7 days from 4 days old and the other of equal volume of saline.After 24 hours of last injection, 8 rats of 11 days old were randomly selected in each group, the level of Iba-1 m RNA was measured by RT-PCR. At 12 days old, 8 rats were randomly picked out to observe the pathological changes in brain tissue of rats by HE staining and evaluate the expression of myelin basic protein(MBP) subcortical white matter、neuron specific enolase(NSE) subcortical gray matter by immu-nohistochemical. The rest brains were divided into left and right hemispheres marked by sagittal suture and the hippocampal tissue was taken out in ice surface rapidly. The left was to detect the expression of MBP、Caspase-3 by RT-PCR method, and the right was to evaluate the MBP、Caspase-3 protein level by western blot method.Result 1 Pathological changes of newborn rats brain tissue in three groupsWhen adjusted the optical microscope to 400 times, we discovered changes in different groups. In the control group, we find cells arranged neatly, clear layer, intact structure, normal morphology. The number of cells reduced, arranged in disorder, disorganized and ill-defined margins in LPS infection group. It was better in caffeine citrate intervention group than in LPS infection group.2 The expression of MBP in neonatal rats brain in each groupsImmunohistochemistry detection: the mean optical density values(IOD) of cortex MBP positive expression in the control group, LPS infection group,and caffeine citrate treatment group were: 132.64 ± 1.94、102.43 ± 2.12、114.25 ± 2.04; the difference was statistically significant in 3 groups(P <0.05). RT-PCR detected: the relative expression levels of MBP m RNA of 3 groups in hippocampus were0.79 ± 0.013、0.39 ± 0.032、0.55 ± 0.019, the difference was statistically significant in 3 groups(P <0.05). The relative expression of MBP protein of 3 groups in hippocampus detected by Western blot were 0.64±0.029、0.31±0.028、0.51±0.049, the difference was statistically significant in 3 groups(P <0.05).3 The expression of Caspase-3 in neonatal rats brain in each groupsRT-PCR detected: the relative expression levels of Caspase-3 m RNA of 3 groups in hippocampus were: 0.34±0.021、0.74±0.029、0.57±0.041, the difference was statistically significant in 3 groups(P <0.05). The relative expression of Caspase-3 protein of 3 groups in hippocampus detected by Western blot were 0.11±0.032、0.36±0.021、0.23±0.028, the difference was statistically significant in 3 groups(P <0.05).4 The expression of NSE in neonatal rats brain in each groupsImmunohistochemical staining showed that the mean optical density values(IOD) of NSE positive expression in grey matter in the control group、 LPS infection group and caffeine citrate treatment group and were123.75±3.12、92.46±3.03、108.24±2.97,the difference was statistically significant in 3 groups(P <0.05).5. The expression of Iba-1 in neonatal rats brain in each groupsThe expression of Iba-1 m RNA was measured by RT-PCR technique showed that the relative expression of Iba-1 in 3 group were0.60±0.026、0.82±0.023、0.69±0.037;the difference was statistically significant in 3 groups(P <0.05).Conclusion Caffeine citrate have showed protective effect on white and grey matter damage in neonatal rats of two days old. The mechanism may be related to inhibition of the inflammatory response and reducing apoptosis. |
PDF Full Text Request