Objective To investigate the pathogenesis of acute lung injury in rats induced by intra-peritoneally injection of perforative peritonitis ascitic fluids（PPAF） and the role of L-arginine （L-Arg） in acute lung injury in this model. Methods Perforative peritonitis （PP） models were established in 60 rats and PPAF were collected. Forty-eight rats were randomly divided equally into NS group，PPAF group, and L-Arg group. Rats were randomly subjected to death at 7 h and 12 h. Peripheral blood WBC were counted，levels of NO and malondialdehyde （MDA） in serum were examined. Lung injury score and wet/dry ratio were evaluated， and level of myeloperoxidase （MPO） in lung tissues and lung cell apoptosis were tested. Results WBC count of peripheral blood, levels of NO and MDA in serum， level of MPO in lung tissue, lung injury score， wet/dry ratio， and lung cell apoptosis rate in PPAF group were significantly higher than that in NS group at each time point（P＜0.01）. Level of NO in serum in L-Arg group was higher than that in PPAF group （P＜0.01）， but lower level of MDA in serum， lower level of MPO in lung tissue and lung injury score，lower wet/dry ratio， and lung cell apoptosis rate were observed in L-Arg group（P＜0.05）. In PPAF group and L-Arg group， level of NO in serum， wet/dry ratio, and lung cell apoptosis rate were higher at 12 h than that at 7 h（P=0.000）. Serum NO level was in negative correlation with serum MDA level （r=-0.257，P=0.021）， MPO level in lung tissue（r=-0.444， P=0.011），and lung cell apoptosis（r=-0.351， P =0.010） in PPAF group and L-Arg group， but serum MDA level was in positive correlation with cell apoptosis（r=0.969， P＜0.001） in each group. Conclusions Acute lung injury rats model can be established by intra-peritoneally injection of PPAF. Enhanced oxidizing reaction and cell apoptosis take part in the occurrence of acute lung injury. L-Arg plays a protective role in acute lung injury.
Objective To study the effect of various doses of estrogen on tissue injury, blood supply and survival area of skin flap and to investigate its mechanism. Methods Thirty New Zealand white rabbits aged 3-4 months old and weighing 1.5-2.2 kg (male or female) were used. Random pattern skin flap (12 cm × 3 cm in size) taking the central l ine of the rabbit dorsum as axis and with the pedicle attached at the proximal end was prepared, and the flap pedicle division was performed 7 days after operation. The rabbits were divided randomly into three groups (n=10 rabbits per group). At 2, 4, and 6 days after operation, the proximal edge of flap in group A and B received 100 ?g/kg and 50 ?g/kg subcutaneous injection ofestradiol benzoate, respectively, while group C received no further treatment serving as control group. General condition ofthe rabbits was observed after injection, gross observation was performed 3 and 7 days after injection, survival area of the skin flap was measured 7 days after injection, contents of malondialdehyde (MDA) and nitric oxide (NO) were tested 5 days after injection, and the flaps were harvested 4 and 7 days after injection to receive histology and no significant difference was noted between group A and group B (P gt; 0.05). The NEU counts 4 days after injection were (18.20 ±6.24) cells/HP in group A, (21.27 ± 5.34) cells/HP in group B, and (28.78 ± 7.92) cells/HP in group C, and at 7 days after injection, there were (15.16 ± 7.02) cells/HP in group A, (18.12 ± 6.44) cells/HP in group B, and (29.67 ± 9.12) cells/HP in group C. The VEGF score 4 days after injection was (4.02 ± 0.48) points in group A, (4.19 ± 0.66) points in group B and (3.67 ± 0.49) points in group C, and at 7 day after injection, it was (4.96 ± 0.69) points in group A, (5.12 ± 0.77) points in group B, and (3.81 ± 0.54) points in group C. Significant difference was evident between 4 days and 7 days after injection in group A or B in terms of NEU counts and VEGF score (P lt; 0.05), and difference between 4 days and 7 days after injection in group C was not significant (P gt; 0.05), and the differences among 3 groups were significant (P lt; 0.05). Conclusion Estrogen injection can increase VEGF expression and NO content of flap, decrease MDA content and NEU infiltration of flat, and improve survival area of flap.
Objective To observe the relationship between endothelial constitutive nitric oxide synthase (ecNOS) genetic polymorphism and diabetic retinopathy（DR）of non insulindependent diabetes mellitus (NIDDM) patients of the Han nationality.Methods A total of 166 patients who clinical diagnosed with NIDDM as case group, 85 cases of patients (cataract or fracture) and healthy subjects without diabetes, hypertension and kidney disease,over 40 years old of age and without consanguinity between each other were selected as normal control group. Case group were divided into non-DR (NDR) group, nonproliferative-DR (BDR) group and proliferativeDR (PDR) group according to the result of fundus fluorescein angiography. Case group and normal control group subjects all were Han nationality. DNA was extracted from peripheral venous blood; the fourth 27 base pairs (bp) repeat polymorphism of ecNOS gene by was measured by polymerase chain reaction (PCR). Results The 27 bp repeat sequences within the ecNOS gene present in the Han nationality,allele b repeat 5 times, alleles a repeat 4 times. PCR results showed that there are 2 alleles and 3 genotypes in normal control, NDR, BDR and PDR group. The frequency of genotype bb、ab、aa were 80%, 16.5%, 3.5% in normal subjects; 77.2％, 13.9%, 8.9％ in NDR group; 80.5％, 17.1%,2.4％ in BDR group;78.3％, 13%, 8.7％ in PDR group,respectively. The allele frequency (chi;2 =1.841) and gene frequency (chi;2=3.847) were not statistically significant (P＞0.5) in normal control,NDR,BDR and PDR group. Logistic regression analysis showed that there is no relation between DR and ecNOS duplicated gene polymorphism. Conclusions There is 27 bp repeated polymorphism in 4th intron of ecNOS gene, which may not be associated with the DR of NIDDM in the Han nationality.
Objective To study the changes of endothelin (ET) and nitric oxide (NO) in the local site of vein transfer with delayed breaking pedicle and the relation with vasospasm and vein transfer in rabbits. MethodsThe ET concentration of blood was determined with the radioimmunoassay method. The plasma NO-2,NO-3 levels in the local site of vein transfer with delayed breaking pedicle, which reflected NO levels indirectly, were detected with Ultravioletvisible (UvVIS ) spectrophotometer. ResultsThe endothelin concentration of blood was increased significantly at 2, 4 hour after the operation (P＜0.01), and at 8 hour after the operation (P＜0.05). The plasma NO level was significantly decreased at 2, 4 hour after the operation (P＜0.01). But at 24 hour after the operation, the plasma NO level was increased significantly (P＜0.05). Conclusion The recovery of ET concentration of blood and the increase of plasma NO at 24 hour after the operation are the cause of the reduced incidence of vascular crisis of vein transfer with delayed breaking pedicle, and the very time point is the optimum moment for pedicle breaking.
ObjectiveTo explore the effect of endotoxin on insulin secretion from islet βcell of rat pancreas.MethodsAfter the model of endotoxemia was established in rats with intraperitoneal injection of LPS (2 mg/kg),the changes of insulin level in the serum and pancreas were dynamically determined, the expression of inducible nitric oxide synthase (iNOS) by situ hybridization and DNA damage in islet cells were also observed, the effect of sodium nitroprusside (exogenous NO) on synthesis and secretion of insulin from isolated islet βcell of normal rat pancreas under high glucose stimulation was also evaluated.ResultsThe level of glucose and insulin in plasma were significantly increased at 12th and 6th h, respectively and kept on 3 d after injection of LPS,but the insulin level in pancreas was not remarkably altered.The expression of iNOS and DNA damaged significantly enhanced at 6 d after endotoxemia. The high glucosestimulated insulin synthesis and secretion were bly inhibited by exogenous NO.ConclusionThese findings suggest that LPS be stimulate the expression of iNOS and NO product,which inhibites synthesis and secretion of insulin in islet βcells,but it stimulates insulin secretion by another mechanism,and results in dysfunction and destruction of the rat pancreas.
ObjectiveTo study the effect of Schwann cells (SCs) promoting the function of nitric oxide (NO) secretion of bone marrow mesenchymal stem cells (BMSCs) derived endothelial cells so as to lay the experimental foundation for research of the effect of nerves on vessels during the process of tissue engineering bone formation.
MethodsSCs were collected from 1-day-old Sprague Dawley (SD) rats,and identified through S100 immunohistochemistry (IHC).BMSCs were collected from 2-week-old SD rats and induced into endothelial cells (IECs),which were identified through von Willebrand factor (vWF) and CD31 immunofluorescence (IF).Transwell system was used for co-culture of SCs and IECs without contact as the experimental group,and simple culture of IECs served as the control group.The NO concentration in the medium was measured at 1,3,5,and 7 days after culture; the mRNA expressions of nitric oxide synthetase 2 (NOS2) and NOS3 were detected by real-time fluorescence quantitative PCR (RT-qPCR) at 1,3,7,and 10 days.
ResultsSCs and IECs were identified through morphology and immunology indexes of S100 IHC,vWF and CD31 IF.Significant differences were found in the NO concentration among different time points in 2 groups (P<0.05); the NO concentration of the experimental group was significantly higher than that of the control group at the other time points (P<0.05) except at 3 days.NOS2 mRNA expression of the experimental group was significantly higher than that of the control group (P<0.05); difference was significant in the NOS2 mRNA expression among different time points in 2 groups (P<0.05).NOS3 mRNA expression of the experimental group was significantly higher than that of the control group at the other time points (P<0.05) except at 10 days.No significant difference was found in NOS3 mRNA expression among different time points in the experimental group (F=6.673,P=0.062),but it showed significant differences in the control group (F=36.581,P=0.000).
ConclusionSCs can promote NO secretion of BMSCs derived endothelial cells,which is due to promoting the activity of NOS.
Objective To study the effects of L-arginine （L-Arg） on cell proliferation, inducible nitric oxide synthase (iNOS) expression and cell cycle in human colon carcinoma cell line LS174 through nitric oxide (NO) pathway. Methods LS174 cells were cultured in medium with L-Arg at different concentrations for different times. MTT method was employed to evaluate the level of the cell proliferation. The production of NO in culture supernatants of LS174 cell was detected with enzyme reduction of nitrate. The distribution of the cell cycle was detected with the flow cytometry (FCM). The expression level of iNOS in the cells was determined by Western blot and SP immunocytochemical staining method. Results The growth of LS174 was promoted by the L-Arg at low concentration (0.125 mmol/L) and inhibited at high concentrations (0.5， 2， 8 and 32 mmol/L). The level of NO was increased with the increasing concentration of L-Arg in culture medium. To compare with the control group, the ratio of cells at S phase was increased after 48 hours’ treatments with high concentrations (0.5, 2, 8 and 32 mmol/L) of L-Arg (P＜0.05, P＜0.01); while there was no obvious difference after treatments with low concentration (0.125 mmol/L) of L-Arg (Pgt;0.05). With the increase of the concentration of L-Arg, the expression of iNOS was increased as compared with control group. The higher the concentration of L-Arg was, the better the effect. Conclusion L-Arg can induce the expression of iNOS resulting in increase the production of nitric oxide (NO). Low concentration of L-Arg can promote the growth of LS174 cells, while high concentration ones can inhibit growth and proliferation. The high concentration of L-Arg could induce S phase arrestion in the cell cycle.
【Abstract】ObjectiveTo investigate the protective effect of melatonin on renal injury induced by bile duct ligation in rats. MethodsSixtyfour rats were randomly divided into four experimental groups (n=16 rats per group): the control group (CN), sham operative group (SO), bile duct ligation group (BDL) and bile duct ligation melatonin treatment group (BDL+Mel). Obstructive jaundice was induced by common bile duct ligation. Plasma level of nitric oxide (NO), total bilirubin (TB), direct bilirubin (DB), alanine aminotransferase (ALT), aspartate aminotransferase (AST), urea nitrogen (BUN) and creatinine (Cr) were measured 4 d and 8 d after operation. NO and inducible nitric oxide synthase (iNOS) in renal tissue were detected at the same time point, too. Histopathological changes of kidneys were examined by HE staining. ResultsIn BDL group, the plasma levels of NO, TB, DB, ALT, AST, BUN and Cr were higher than those of SO group (P＜0.01）, and the level of NO and activities of iNOS in renal tissue were significantly increased (P＜0.01）. However, in BDL+Mel group, the plasma levels of NO, ALT, AST, BUN and Cr were lower than those of the BDL group (P＜0.01）, and the level of NO and activities of iNOS in renal tissue were significantly suppressed (P＜0.01）; histopathological changes of kidneys were improved.ConclusionAugmentation of iNOS activities and increasing of NO production in local tissue contributed to renal injury induced by bile duct ligation, and the mode of melatonin’s protective actions, at least in part, relates to interference with no pathways.
Objective To investigate the effects of nitric oxide precursor L-arginine on traumatic pulmonary contusion. Methods Sixty Sprague-Dawley rats were randomly divided into three groups, ie. a normal group, a model group, and a L-arginine group. The model of traumatic pulmonary contusion was established with self-made chest-impacter. Then the rats in the L-arginine group was injected intravenously with L-arginine in a dose of 250 mg/kg. All rats were sacrificed at 24 hours after these models established.Levels of TNF-α and nitric oxide ( NO2 - /NO3- ) in serum were measured by ELISA and diazo-reaction method. Lung wet/dry weight ratio, NF-κB, endothelin-1, apoptotic cell, and ICAM-1 ( intercellular adhesion molecule-1) mRNA expressions in the lung tissue were measured. Results Compared with the model group,TNF-αand lung wet/dry weight ratio decreased significantly in the L-arginine group( P lt; 0. 05) . After the L-arginine treatment, the concentration of nitric oxide, apoptotic index were significantly higher than the model group ( P lt; 0. 05) . The expressions of NF-κB, endothelin-1, and ICAM-1 mRNA in the L-arginine group were lower than those in the model group ( P lt;0. 05) . Conclusion L-arginine treatment can downregulate the expressions of NF-κB, ET-1, ICAM-1 mRNA and apoptosis obviously, and ameliorate the microcirculation of rats lung with traumatic pulmonary contusion.
ObjectiveTo analyze the expression of VEGF, IL-33 and NO concentration after laser photocoagulation and subthreshold micropulse laser photocoagulation conventional in proliferative diabetic retinopathy (PDR) patients.MethodsA case control study. The clinical data of 39 patients of PDR and 11 patients of idiopathic macular pucker (IMP) from Department of Ophthalmology, Central Theater General Hospital during November 2015 were collected in this study. PDR patients were assigned randomly into three groups. Fifteen PDR patients with 15 eyes were treated with conventional laser as group A. Thirteen PDR patients with 13 eyes were treated with subthreshold micropulse laser as group B. Eleven PDR patients with 11 eyes without any laser therapy were grouped as C. Eleven IMP patients were grouped as D. There was no difference of age (F=0.53, P=0.23), gender ratio (χ2=0.55, P=0.91), body mass index (F=2.62, P=0.07), duration diabetes (F=0.29, P=0.75), glycoslated hemglobin (F=1.72, P=0.19) in four groups. All PDR patients were examined with FFA. Total protein was quantified by a bicinchoninic acid assay kit. Levels of VEGF, IL-33, NO were determined using enzyme-linked immunosorbent assay kits.ResultsThere was no difference of total protein in four groups (F=1.78, P=0.17). Group C had a higher VEGF level than group A and B (F=7.84, P=0.002). Group A had a higher IL-33 level than group C (t=4.15, P=0.02). There was no difference of IL-33 level in group B and C (t=1.34, P=0.20). Group D had a lower NO level than group A, B, C (F=38.42, P＜0.001). There was no difference of NO level in group A, B and C (F=3.29, P=0.06).ConclusionsBoth conventional laser photocoagulation and subthreshold micropulse laser photocoagulation can decrease vitreous VEGF level and subthreshold micropulse laser photocoagulation can induce less IL-33 level.
