ObjectiveTo investigate the expression of caspase-3 and Toll-like receptor 4 (TLR4) in the incised rat skin healing process and its relationship with the wound time and to provide an experimental evidence for the prediction of injury time. MethodsAfter the rat incised wound model was established, hematoxylin-eosin dyeing technology and immunohistochemical staining technique were used to observe the expression of caspase-3 and TLR4. Then Image Pro Plus Image analysis software and SPSS statistical analysis software were used to deal with the experimental results. ResultsCaspase-3- and TLR4-positive cells were detected in epidermis, hair follicle and sebaceous gland cells in the control skin. The expression of caspase-3 and TLR4 of the ante mortem groups were significantly different compared with the control group except the 0 h group (P<0.05). Caspase-3- and TLR4-positive cells were detected in neutrophils around the hair follicle half an hour later. Caspase-3- and TLR4-positive cell rate increased with the infiltration of inflammatory cells. Caspase-3- and TLR4-positive cell rate reached the maximum on the 3 rd day, and then it began to decrease, and they were mainly expressed in fibroblasts and mononuclear macrophages. Caspase-3- and TLR4-positive cells were mainly expressed in fibroblasts on the 10th day. There was no significant differences between the postmortem injury groups and the normal control groups (P>0.05). ConclusionCaspase-3- and TLR4-positive cell rate is time dependent and stable in 25℃ temperature environment which makes it possible to determine the time of injury.
ObjectiveTo observe the dynamic changes of neuroglobin (NGB) expression in hippocampus after status epilepticus(SE) in rats, and to explore the role of NGB in epileptic seizures.Methods40 healthy male Sprague Dawley rats were randomly divided into two group according to random number table method:control group (n=5) and epilepsy model group(n=35).Epilepsy model group according to observation time was divided into:0h, 1h, 3h, 12h, 24h, 10d and 30d.Intraperitoneal injection Lithium-pilocarpine (20 mg/kg~127 mg/kg, Li-PC) to establish the rat model of SE.Observe the behavioral changes in rats with epilepsy.Nissl staining was used to detect the neuronal damage in hippocampus. Streptavidin-biotin-peroxidase complex immunohistochemical method was used to detect the expression level of NGB in hippocampus;ResultsAfter SE, the neurons in hippocampus were severely damaged with the progress of epileptic seizures, the number of surviving neurons in CA1, CA3 regions showed a near linear decline.Among them, the number of surviving neurons in (12h, 24h, 10d, 30d)CA1, (0h, 12h, 24h, 10d, 30d)CA3 and(12h, 24h, 10d, 30d) DG area were significantly lower than that of the control group (P < 0.05).The expression level of NGB in CA1, CA3 and DG region of hippocampus were increased after SE, and both of CA1 and DG were reached peak in 24h after SE, but was still higher than the control group.And the CA3 area showed a continue rising trend.Among them, CA1(24h, 10d, 30d), CA3(24h, 10d, 30d) and DG(12h, 24h, 10d, 30d) were higher than that of control group significantly (P < 0.05).In addition, it was found that there was a positive correlation between the number of surviving neurons in CA3 area and the expression level of NGB (R=0.306, P=0.011).ConclusionUp-regulation of NGB expression in hippocampus after status epilepticus, and was positively correlated with the number of neurons in the CA3 area, suggesting that up regulation of NGB expression may be a compensatory protective mechanism of ischemic injury induced by seizures, and participate in the protection of epilepsy related neuronal damage.
【Abstract】ObjectiveTo investigate the effects of liver transplantation on splenic function in rats with hepatic cirrhosis. MethodsHepatic cirrhosis model was established in rats by subcutaneous injections of carbon tetrachloride. Liver transplantation model was established with twocuff technique. Spleen index, morphological changes of spleen were observed before and after liver transplantation in hepatic cirrhosis rats. Spleen T lymphocyte subgroups before and after liver transplantation were also assayed by immunofluorescence staining and flow cytometry. ResultsBefore liver transplantation, spleen index was increased from (2.42±0.11) mg/g to (3.62±0.14) mg/g， P＜0.01; pathological examination of spleen samples showed that the areas of white pulp were decreased from (23.47±2.30)% to (7.70±2.01)%， P＜0.01， and the areas of spleen trabecula were increased from (1.75±0.61)% to (4.46±0.71)%， P＜0.01. Meanwhile, the ratio of CD4/CD8 of spleen T lymphocyte subgroups was decreased from 2.67±0.15 to 1.18±0.15, P＜0.01. After liver transplantation, spleen index was decreased from (3.62±0.14) mg/g to (2.62±0.11) mg/g， P＜0.01; pathological examination of spleen showed that the areas of white pulp were increased from (7.70±2.01)% to (15.07±1.97)%， P＜0.01, and those of spleen trabecula were decreased from (4.46±0.71)% to (3.11±0.51)%， P＜0.05. Meanwhile, the ratio of CD4/CD8 of spleen T lymphocyte subgroups was increased from 1.18±0.15 to 2.32±0.11, P＜0.01. ConclusionImpaired function of spleen resulting from liver function damage can be improved in rats with hepatic cirrhosis after liver transplantation.
ObjectiveTo explore the mechanism of lung injury in Sprague-Dawley (SD) rats induced by acute organic phosphorus pesticides (AOPP) by observing the changes of the blood serum nuclear factor (NF)-κB consistence, NF-κB level of lung tissue and lung coefficient. MethodNinety-six healthy male SD rats (six weeks old) were randomly divided into group A (control, n=48) and group B (poison, n=48). The rats of group B were given omethoate by gavage (45 mg/kg), and the rats of group A accepted normal saline. Then the rats were killed at designated observing points (30 minutes; 3, 6, 12, 24, and 48 hours), and the lung coefficient, blood serum NF-κB consistence and NF-κB level of lung tissue were measured. At the same time, we observed the pathological changes of the rats' lung tissue. ResultsCompared with group A, blood serum NF-κB consistence, NF-κB level of lung tissue and the level of lung coefficient in group B were significantly higher (P<0.01). The lung tissues of group A were normal at each time point, but in group B, the lung pathological changes gradually appeared 30 minutes later with pulmonary interstitial engorging, alveolar septum widening and some alveolus being full of red blood cells, and this situation reached its peak at hour 12. Then it gradually mitigated from 24 to 48 hours. ConclusionThere are significant increases in blood serum NF-κB consistence and NF-κB level in lung tissues in rats with lung injury induced by omethoate poisoning. The NF-κB may play a role in the process of lung injury induced by organophosphorus pesticide.
Objective Using cortex convulsions threshold detector and electrical stimulation in rats cortex convulsions threshold model, compare the efficacy and aging of domestic lamotrigine (LTG) and imported LTG. Methods Electrical stimulation convulsions threshold model in rats after stability, 40 rats were randomly divided into A、B、C、D groups,AandBgroup were divided into three different dose groups: domestic LTG low dose (12.5 mg/kg/d), middle dose (25 mg/kg·d), high dose group (37.5 mg/kg·d); imported LTG low doses (12.5 mg/kg·d), middle dose (25 mg/kg·d), high dose group (37.5 mg/kg·d); Carbamazepine middle dose group (72 mg/kg·d); the control group (normal saline 2 ml/time). Recording electrical stimulation in rats cortex convulsions threshold model after administration, compare the differences before and after the administration. Results Three different dose groups of domestic LTG and imported LTG all hadahigher level of electrical stimulation cortex convulsions threshold, and showedadose-response relationship. Onset time of LTG after administration was 1 to 2 hours, peak time was 3 to 4 hours, maintaining time was 8 to 10 hours. Conclusion LTG can improve cortex convulsions threshold in the electrical stimulated rats, there was no significant difference with carbamazepine, and showedadose-response relationship; Repeat dosing for 4 days, both domestic and imported LIG can maintain effective anticonvulsive effect, the efficacy and the aging of two groups of LTG have no significant difference (P＞0.05).
Objective To study three-dimensional culturing methods of neonatal rat cardiac myocytes in simulated microgravity. Methods Neonatal rat primary cardiac myocytes were separated and seeded into polylactic acid scaffolds, stirredin spinner flasks for 24 hours, and then moved into rotary cell culture system for three-dimensional culture. The growth of cardiac myocytes was observed underinverted phase contrast microscope, scanning electron microscope and transmission electron microscope, and metabolic assay was assessed by MTT assay. Results Cardiac myocytes with sustained metabolic activity attached to the polylactic acid scaffolds, extended and confluenced. Pulsations of PLAcardiac myocytes was found in some areas. Conclusion The rotary cell culture system is suitable to develop neonatal rat cardiac myocytes culturing for three-dimensional modeling.
【Abstract】 Objective To investigate the effectiveness of all-trans-retinoic acid (ATRA) at different concentrationson prol iferation and differentiation of the rat embryonic neural stem cells (NSCs), and to find the optimal concentration of ATRA that promoting the differentiation of NSCs into neurons. Methods NSCs were isolated from cerebral cortex of rat embryos (embryonic day 12-16, average 15 days), and were cultured in serum-free medium (DMEM/F12 medium containing 20 ng/mL bFGF and 20 ng/mL EGF) at the concentration of 1×106 cells/mL. Subcultures were performed 7 days after the primary culture. The cell clusters of the 3rd passage were centrifuged and divided into 5 groups. In the experimental groups (groups A, B, C, D), the ATRA concentration was 0.5, 1.0, 5.0, 10.0 μmol/L in DMEM/F12 complete medium respectively, while in control group (group E), the ATRA concentration was 0 in DMEM/F12 complete medium. The prol iferation rate of each group was analyzedby cell counting day by day till 7th day, and BrdU positive cell counting 1, 3, 5, 7, 9 days after culture. In addition, collecting the 3rd passage NSCs and divided into 5 groups. In the experimental groups (groups A, B, C, D), the ATRA concentration was 0.5, 1.0, 5.0, 10.0 μmol/L in DMEM/F12 medium containing 5% FBS respectively, while in control group (group E), the ATRA concentration was 0 in DMEM/F12 medium containing 5% FBS. The capacity of NSCs differentiation toward neurons was determined by immunofluorescence double-labell ing and flow cytometry. Results Cell counting 1-7 days after culture in each experimental group (groups A, B, C, D) showed no significant differences (P gt; 0.05). Cell counting at each time point of all the experimental groups were less than those of control group (P lt; 0.05). BrdU positive cells were increased 1, 3, 5, 7, 9 days after culture in each experimental group (groups A, B, C, D), but there was no significant difference between each experimental group(P gt; 0.05). BrdU positive cells at each time point of control groups were more than those of all the experimental groups (P lt;0.05). The differentiation ratio of neurons was enhanced in experimental groups and the optimal ATRA treatment concentration was 1.0 μmol/ L (experimental group B). The differentiation ratio of neurons induced by ATRA in group B was 29.46% ± 0.47%, 47.25% ± 0.46% and 66.81% ± 0.57% respectively after cultured 3, 5 and 7 days, whereas the differentiation ratio of neurons was 11.11% ± 0.59%, 14.10% ± 0.32% and 15.92% ± 0.70% respectively in control group. The majority of NSCs differentiated into astrogl ial phenotypes in control group. By flow cytometry detection, the differentiation ratio of neurons after cultured 3 days and 7 days in experimental groups were more than those in control group (P lt; 0.05). Conclusion ATRA treatment remarkably promoted the differentiation of NSCs into neurons and the optimal concentration was 1.0 μmol/L.
ObjectiveTo investigate the effects of hippocampal long-term potentiation (LTP) on cognitive dysfunction in immature epileptic rats. MethodsImmature epileptic rats were established by intraperitoneal injection of lithium chloride-pilocarpine (li-pilo). Racine classification standard modified by Becker was used to evaluate behavior of epileptic seizure, and the survival rats within RacineⅣmagnitude were selected in the experiment. The function of learning and memory of epileptic rats when they were adult was assessed using Morris water maze experiment, and their independent exploratory behavior was evaluated by the open-field test. Field potential was recorded by electrophysiological technology to detecte whether hippocampal LTP was essential of cognitive dysfunction. ResultsThe function of learning and memory was significantly impaired when compared with controls(n=8, t=10.86, P < 0.05;n=8, t=9.98, P < 0.05). In addition, independent exploratory behavior was significantly reduced when compared with controls(n=8, t=12.89, P < 0.05). Besides, CA1 hippocampal LTP induced by high-frequency stimulation presented the significant inhibition in epileptic rats with cognitive dysfunction when compared with controls(Slope:n=8, t=13.32, P < 0.05;Amplitude:n=8, t=20.02, P < 0.05). ConclusionInhibition of CA1 hippocampal LTP may be implicated in cognitive dysfunction of epileptic rats.
Objective To investigate the effects of granulocyto-colony stimulating factor (G-CSF) on the mobil ization of endothel ial progenitor cells (EPCs) in the rats with myocardial infarction (MI), to observe the density of neovascularization and the mRNA expressions of vascular endothel ial growth factor (VEGF) and its receptor (Flk-1) in the border area of MI. Methods Thirty-six adult male rats (weighing 250-280 g) were divided randomly into control group, MI group, and G-CSF group. In MI group and G-CSF group, the models of MI were establ ished by left anterior descenting coronary artery l igation and were treated with intraperitoneal injection of sal ine (0.3 mL/d) or G-CSF [30 μg/(kg•d)] for 5 days. In control group, after open chest operation, chest was closed without treatment. The level of EPCs was surveyed and the plasma concentrations of VEGF and C-reaction protein (CRP) were measured at 7 days. The mRNA expressions of VEGFand its receptor Flk-1 in the border area of infarct myocardium were determined through RT-PCR. Results Compared withcontrol group, the number of circulating white blood cell (WBC) and EPCs levels, and the serum concentrations of VEGF and CRP were all significantly increased in MI group and G-CSF group (P lt; 0.05); when compared with MI group, the number of circulating WBC and EPCs levels, and the serum concentrations of VEGF were increased and the concentration of CRP was decreased in G-CSF group (P lt; 0.05). Compared with control group, the mRNA expressions of VEGF and Flk-1, and the density of neovascularization in the border area of infarct myocardium were increased in MI group and G-CSF group, whereas those in G-CSF group were significantly augmented compared with MI group (P lt; 0.05). Conclusion In the rats with MI, G-CSF could promote EPCs mobil ization, increase the mRNA expressions of VEGF and Flk-1, and augment the density of neovascularization in the border area of infarct myocardium.