To explore the role of cell apoptosis in denervated skeletal muscle atrophy in rats and the effect of losartan on it. Methods Forty-two Sprague Dawley rats were randomly divided into 3 groups: group I (n =14, normal control group), group II (n =14, denervated group) and group III (n =14, losartan group). The rats were not treated in group I, and were made denervated gastrocnemius models in groups II and III. In group III, the rats were treated with losartan 10 mg /kg• d by gavage and with normal sal ine in groups I and II. After 4 weeks, gastrocnemius mass to body mass ratio (GAS/BM) served as the degree of muscle atrophy. Apoptotic cells in gastrocnemius were stained in situ by using TUNEL. Gastrocnemius Bcl-2 and Bax protein were quantified by immunohistochemistry and Western blot. Bax /Bcl-2 served as the degree of apoptosis. Results The ratio of apoptosis was higher in group II than that in group I (11.32% ± 4.51% vs 0.56% ± 0.21%, P lt; 0.05). The ratio of apoptosis was lower in group III than that in group II (7.21% ± 2.05% vs 11.32% ± 4.51%, P lt; 0.05). The atrophy of skeletal muscle（GAS/BM） in group II was more serious than that in group I (11.68 ± 1.98 vs 12.86 ±0.74, P lt; 0.05), there was no significant difference between group III and group II (12.11 ± 0.65 vs 11.68 ± 1.98, P gt; 0.05). The expression of Bcl-2 in group II (18.3% ± 4.9%) was significantly lower than that in group I (27.5% ± 2.8%) and group III (25.5% ± 3.5%); there was no significant difference between group III and group I (P gt; 0.05). The expression of Bax in group II (24.1% ± 3.1%) was significantly higher than that in group I (22.1% ± 3.6%) and group III (21.7% ± 2.3%); there was no significant difference between group III and group I (P gt; 0.05). Western blot results showed that: the expressions of Bcl-2 were 122.5 ± 14.6 in group II, 135.3 ± 6.2 in group I and 139.2 ± 16.2 in group III; showing significant diffeerences between group II and group I, between group III and group II (P lt;0.05). The expressions of Bax were 107.1 ± 15.8 in group II, 89.3 ± 8.4 in group I, and 94.2 ± 9.5 in group III; showing significant diffeerences between group II and group I, between group III and group II (P lt; 0.05). There was no significant difference in the expression of Bcl-2 and Bax between group Ⅲ and group I (P gt; 0.05). Conclusion Cell apoptosis plays an important role in denervated skeletal muscle atrophy in rats and may be one of the factors causing skeletal muscle atrophy. Losarton can decrease skeletal muscle cell apoptosis through regulating the ratio of Bax / Bcl-2.
Objective To explore the effects of several immunosuppressants on the cell numbers of cultured rat macrophages and Schwann’s cells. Methods The macrophages and Schwann’s cells were cultured from the newborn Wistar rats. Different concentrations of methylprednisolone(10-3, 10－4,10－6 and 10－8 mol/L), CsA(10－5, 10－6, 10－7 and 10－8 mol/L) and FK506(10-6, 10－7, 10－8 and 10－9 mol/L) were administrated to the cells, while control group was given no drugs. Twentyfour, 48 and 72 hours after administration, the cells from different concentrations were measured with MTT methods respectively. Theresults were compared and analyzed statistically. Results Only high concentration methylprednisolone (10-4 mol/L) and a certain range of concentrations of CsA (10-6，10-7 and 10-8 mol/L) and FK506 (10-7,10-8 and 10-9 mol/L) can provide protection to culturedrat macrophages. Under most concentrations, CsA and FK506 had no effects onthe cell number of cultured rat Schwann’s cell. Only with high concentration CsA (10-5 mol/L) and methylprednisolone (10-3 mol/L) could significantly decreased the cell number of Schwann’s cell. Long time (72 hours) and low dosage (10-8 mol/L) administration of methylprednisolone could significantlyprotect Schwann’s cell. Conclusion High concentration methylprednisolone and some certain concentration CsA and FK506 can protect cultured rat macrophages. But high concentration CsA and methylprednisolone prohibit the proliferation of Schwann’s cells. Only long time and low dosage methylprednisolonecan protect cultured rat Schwann’s cells.
ObjectiveTo investigate the changes of inflammatory cytokines in bronchoalveolar lavage fluid (BALF) in rats exposed to paraquat (PQ). MethodsAdult healthy SD rats were randomly divided into a control group (n=8) and three experimental groups (PQ in low dosage of 15 mg/kg,medium dosage of 30 mg/kg,and high dosage of 60 mg/kg,n=24 in each group). The rats in three experimental groups were intragastrically administered with PQ,and the rats in the control group were treated with saline by gavage. Two rats in the control group and six rats in three experimental groups were sacrificed on 1st,7th,14th,and 21st day after exposure respectively. BALF was collected for measurement of interleukin-1(IL-1),IL-6,macrophage inflammatory protein-2(MIP-2),monocyte chemoattractant protein-1(MCP-1),and biopterin by ELISA. ResultsThe levels of cytokines in all experimental groups were higher than those in the control group at any time point. In the exposure day 1 to day 14, IL-1 and biopterin levels in BALF increased significantly with the increase in PQ dose. On 14th and 21st day,IL-6 level in BALF increased significantly with the increase in PQ dosage. The levels of IL-1,IL-6,and biopterin in the experimental groups reached the peak on 14th day. On 14th day,the MIP-2 level in BALF of high-dosage group was significantly higher than that of low-dosage and medium-dosage groups (all P<0.05). The level of MCP-1 in the low-dosage group was lower than that in the medium-dosage and high-dosage groups at any time point (P<0.05). ConclusionIL-1,IL-6,MIP-2,MCP-1,and biopterin may play important roles in the development and progression of PQ-induce lung inflammation.
ObjectiveTo confirm that conduction of the epileptiform discharge activity out of the cranium can by reduce the seizure and consequence neuron injury, and introduce the novel micro invasive neurosurgical approach to epileptic therapy. MethodsTo build penicillin-induced neocortical epilepsy rat (n=60, 4 groups, 15 per group, including control group, conducting group, pseudo-conduction group and model control group). The specialized self-made conducting electrode was used to building conducting epileptic rat model. The epileptiform discharges were recorded by EEG with deep needle electrode for 2 hours under anesthesia, and seizures were monitored by video for 48 hours in waking. At last, the apoptosis ratio of neocortex was tested with flow cytometer. ResultsWe built 41 (91.1%) penicillin-induced neocortical epilepsy rats successfully. The mean frequency of total epileptiform discharges and frequency of diffused epileptiform discharges in EEG in conducting group were significantly less than the numbers in model control group and pseudo-conduction group(P < 0.01). However, significant difference was not found in times of focal epileptiform discharges among 3 test groups. During video monitoring, significant difference presented in the frequency of clinical seizure between conducting group and model control group or pseudo-conducting group. Furthermore, apoptosis ratios of neuron in conducting group was significantly less than the other two groups (P < 0.001). ConclusionsConducting the epileptiform discharge activity out of the cranium can prevent the seizure and reduce epileptiform discharge and apoptosis ratio of neocortex in neocortical epilepsy rats.
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 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.
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.
ObjectiveIn order to evaluate that whether the P-glycoprotein-inhibitor verapamil (VPM) could effect the distribution of antiepileptic drug phenytoin (PHT) in a rat model of mesial temporal lobe epilepsy (MTLE).MethodsThe rat models of MTLE were induced by li-pilocarpine and were randomly divided into two groups (PHT group and VPM+PHT treatment group) to compare the PHT distribution in brain, liver and kidney. Brain dialysate samples were collected by microdialysis technology. And the analysis of samples for PHT concentration was performed by high performance liquid chromatography (HPLC). The comparisons were carried out by t test (or Wilcoxon test).ResultsIn VPM+PHT treatment group, 4 out of 9 rats were dead within 30 minutes after drug administration. The significantly decreased area under the curve (AUC) ratio of brain/plasma in VPM+PHT group was 0.11±0.06 when compared with PHT group 0.21±0.02 (t=3.237, P=0.025), while there were no significant differences in ratios of liver/plasma ［PHT (1.12±0.37) vs. VPM+PHT (0.99±0.27), Z=−0.490, P=0.624］ and kidney/plasma ［PHT (0.74±0.16) vs. VPM+PHT (0.49±0.26), t=1.872, P=0.103］ between two groups.ConclusionsThe P-glycoprotein-inhibitor VPM significantly decreased PHT level in brain of rat with MTLE.
【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.
Objective To study the relationship between the expression ratio of heat shock protein (HSP) 70 to C-fos in organs outside the brain after brain concussion and the time of injury in rats, in order to provide a new visual angle for determining injury time of brain concussion. Methods The model of brain concussion was established through free falling method. Then the rats were executed at 30 minutes, 1 hour, and 3, 6, 12, 24, 48, 96, 168, 240, 336 hours after injury. Immunohistochemistry staining of C-fos and HSP70 were used in the materials from the main organs including heart, liver, spleen, lung and kidney. All related experiment results were studied by using a microscope with image analytical system and homologous statistics. Results From 30 minutes to 6 hours after injury, the proportion of HSP70 immuno-positive cells increased slowly, while the proportion of C-fos immuno-positive cells increased rapidly, and the ratio of HSP70/C-fos positive cells was on the decline. From 6 to 12 hours after injury, the proportion of HSP70 immuno-positive cells rose continuously, while the proportion of C-fos immuno-positive cells started to decrease, and the HSP70/C-fos ratio showed a rising tendency. From 12 to 336 hours after injury, the proportion of HSP70 immuno-positive cells decreased slowly, while the proportion of C-fos immuno-positive cells decreased rapidly, and the HSP70/C-fos ratio was still on the rise. Conclusions The proportion of positive cells and ratio of the two markers in the main organs including heart, liver, spleen, lung and kidney are similar to those in the brain of rats after brain concussion. Observing the proportion of positive cells of the two markers together with their ratio in the main organs outside the brain may provide a reference for the determination of injury time after brain concussion.