ABSTRACT
OBJECTIVE@#To study the effect of different melatonin treatment regimens on long-term behavior and white matter damage in neonatal rats with hypoxic-ischemic brain damage (HIBD), and to seek an optimal melatonin treatment regimen.@*METHODS@#Healthy Sprague-Dawley rats, aged 7 days, were randomly divided into four groups: sham-operation, HIBD, single-dose immediate treatment (SDIT), and 7-day continuous treatment (7DCT), with 8 rats in each group. A neonatal rat model of HIBD was prepared according to the classical Rice-Vannucci method. On day 21 after HIBD, the Morris water maze test was used to evaluate spatial learning and memory abilities. On day 70 after HIBD, immunofluorescence assay was used to measure the expression of neuronal nuclear antigen (NeuN) in the cerebral cortex and the hippocampal CA1 region of neonatal rats, and double-label immunofluorescence was used to measure the expression of myelin basic protein (MBP) and neurofilament 200 (NF200) in the corpus striatum and the corpus callosum.@*RESULTS@#The results of the Morris water maze test showed that the SDIT and 7DCT groups had a significantly shorter mean escape latency than the HIBD group, and the 7DCT group had a significantly shorter mean escape latency than the SDIT group (@*CONCLUSIONS@#Both SDIT and 7DCT can improve long-term behavior and reduce white matter damage in neonatal rats with HIBD, and 7DCT is more effective than SDIT.
Subject(s)
Animals , Rats , Animals, Newborn , Hypoxia-Ischemia, Brain/drug therapy , Melatonin/pharmacology , Rats, Sprague-Dawley , White MatterABSTRACT
<p><b>BACKGROUND</b>Cigarette smoking has an influence on both arterial-type and venous-type thrombosis. However, little is known about the direct effect of cigarette smoke extract (CSE) on fibrinolytic activity of human umbilical vein endothelial cells (HUVECs). Most recently, simvastatin has been marked in its effect on endothelial cells protection and anticoagulation. In this study, the effect of CSE on the expression of tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor-1 (PAI-1) in HUVECs was addressed. The role of simvastatin in CSE-induced fibrinolytic activity changes was investigated as well.</p><p><b>METHODS</b>The fourth to fifth generation of HUVECs were incubated respectively with 0, 5%, 10% and 20% CSE for 6 hours or exposed to 5% CSE for 0, 4, 6, 8, 12, 24 hours to determine the expression changes of t-PA and PAI-1 protein. Meanwhile, cells were also accordingly exposed either to 5% CSE alone or simvastatin pre-treated and 5% CSE for 24 hours to assess the role of simvastatin in CSE-induced t-PA and PAI-1 protein and mRNA expression in HUVECs. RT-PCR and ELISA techniques were used for detecting the t-PA or PAI-1 mRNA and protein.</p><p><b>RESULTS</b>After 6-hour exposure to CSE, the expression levels of t-PA protein in 10% and 20% CSE-treated groups reduced significantly ((0.0365 +/- 0.0083) ng/ml, (0.0255 +/- 0.0087) ng/ml) when compared with that of control group ((0.0660 +/- 0.0120) ng/ml) (P < 0.05). In contrast, the levels of PAI-1 protein in 5%, 10% and 20% CSE-treated groups increased remarkably ((13.3225 +/- 0.5680) ng/ml, (14.2675 +/- 1.5380) ng/ml, (14.4292 +/- 1.6230) ng/ml) when compared with that of control group ((8.5193 +/- 0.7537) ng/ml) (P < 0.05). After stimulation with 5% CSE for 0, 4, 6, 8, 12, 24 hours, the levels of PAI-1 protein increased over time and reached the peak at 24 hours ((14.6400 +/- 1.0651) ng/ml), which was significantly higher than that of control group ((12.0656 +/- 0.6148) ng/ml) (P < 0.05). Additionally, CSE could up-regulate PAI-1 expression at both the mRNA and the protein levels. The levels of PAI-1 mRNA and protein increased significantly in 5% CSE-treated group ((8.8030 +/- 0.4745) ng/ml, (1.8155 +/- 0.0412) ng/ml) compared with those of control groups ((5.0588 +/- 0.2315) ng/ml, (1.3030 +/- 0.0647) ng/ml) (P < 0.01), and decreased after 2-hour simvastatin pre-treatment ((5.4875 +/- 0.3166) ng/ml, (1.3975 +/- 0.0297) ng/ml) (P < 0.01). No significant difference was found at the levels of t-PA protein and mRNA (P > 0.05).</p><p><b>CONCLUSIONS</b>CSE inhibits the fibrinolytic activity of HUVECs in vitro. Simvastatin plays a protective role in CSE-induced fibrinolytic malfunction.</p>