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To explore the role of heme oxygenase (HO)-1 experimental system in dachengqitang (DD) ameliorating ALI induced by lipopolysaccharide (LPS) in mice. Seventy-five male Kunming mice were randomly divided into control group (normal saline was instilled intratracheally(50 microL/per mouse), LPS group (LPS was instilled intratracheally to replicate ALI model), DD + LPS group, DD + LPS + ZnPP (ZnPP, HO-1 specific inhibitor) group and the DD group. Mice were killed at 6 h after administration. Lung indexes were tested; lung histomorphological changes were observed under microscope, and neutrophils (PMN) number and protein content of bronchoalveolar lavage fluid (BALF) were measured; HO-1 mRNA and protein expression in lung tissue were detected by RT-PCR and Western blot. The results showed that intratracheal instillation of LPS in mice can cause significant morphological changes in lung tissue. Both PMN numbers and protein content in BALF were increased. meanwhile the expressions of HO-1 mRNA and protein in lung tissue were increased. Pretreated with DD and then intratracheally instillated LPS coulde ameliorat lung tissue injury, reduced PMN BALF number and protein content, but increase HO-1 mRNA and protein expression in the lung tissue when compared with LPS. HO-1 inhibitor ZnPP coulde inhibite the ameliorative effect of DD. The results suggest that the ameliorative effect of DD on ALI induced by LPS in mice were related with upregulation HO-1 mRNA and protein.
Subject(s)
Animals , Male , Mice , Acute Lung Injury , Blotting, Western , Bronchoalveolar Lavage Fluid , Chemistry , Cell Biology , Gene Expression Regulation, Enzymologic , Heme Oxygenase-1 , Genetics , Metabolism , Leukocyte Count , Lipopolysaccharides , Lung , Pathology , Neutrophils , Cell Biology , Phytotherapy , Methods , Plant Extracts , Pharmacology , Proteins , Metabolism , Protoporphyrins , Pharmacology , Random Allocation , Reverse Transcriptase Polymerase Chain Reaction , Treatment OutcomeABSTRACT
AIM: To study the signal pathway involved in up-regulation of LPS-induced HO-1 expression by CCK-8. METHODS: Forty-two SD rats were divided into 7 groups (six rats each) randomly as follows: control group, LPS group, LPS+SP600125 (JNK-specific inhibitor) group, CCK-8+LPS group, CCK-8+LPS+SP600125 group, CCK-8 group and CCK-8 +SP600125 group. Lungs from the rats in these 7 groups were excised 6 h after the agents were administered. HO-1 mRNA expression was examined by RT-PCR. The protein expression of HO-1 was detected by Western blotting and immunofluorescence flow cytometry (FCM). RESULTS: There were significant positive expression of HO-1 mRNA in LPS group compared to control group. CCK-8 enhanced LPS-induced HO-1 mRNA expression and CCK-8 alone induced HO-1 mRNA expression as well. The mRNA expressions of HO-1 in LPS group, CCK-8+LPS group and CCK-8 group were 3.01 (P<0.01), 5.88 (P<0.01) and 3.45 (P<0.01) times as many as that in control group, respectively. SP600125 inhibited the mRNA expression of HO-1 induced by CCK-8 and (or) LPS. The change of HO-1 protein expression was in accordance with that of HO-1 mRNA expression by Western blotting and immunofluorescence FCM. CONCLUSION: These results suggest that JNK/c-Jun signal pathway plays an important role in the up-regulation of LPS-induced HO-1 expression by CCK-8.
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Objective To observe the changes of urinary microalbumin(UMA),and explore the relation between early impairment of renal function and level of UMA in patients with obstructive sleep apnea hypopnea syndrome(OSAHS).Methods OSAHS patients(n=67)and healthy control subjects(n=82).Then body parameter,serum creatinine(SCr),blood urea nitrogen(BUN),glomerular filtration rate(GFR)and UMA were determined in both groups.Furthermore,linear correlation was performed between UMA and body parameter,SCr,BUN,GFR and sleep-breathing parameters of patients with OSAHS,respectively.Results UMA level was higher in OSAHS patients than that in control subjects(P＜0.01).UMA level in OSAHS patients was correlated positively with body mass index(BMI),apnea hypopnea index(AHI),percentage of sleep time below 90% oxygen saturation (SaO2＜90%)and percentage of sleep time the total duration of apnea/hypopnea,respectively,and those were correlated negatively with the lowest SaO2 and the average lowest SaO2 respectively,but those had no correlativity with age,stature,waistline,neckline,SCr,BUN and GFR(P＞0.05).Conclusions UMA level increases in patients with OSAHS.The detection of microalbumin in urine is helpful in early diagnosis in OSAHS renal damage patients.
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BACKGROUND:After cerebral tissue ischemia and anoxia in young rats,the cerebral edema gets serious, and the levels of nitric oxide (NO) and malondialdehyde (MDA) decrease. Radix Astagali seu Hedysari has the pharmacological effects of enhancing immunity, anti-anoxia and improving myocardial ischemic reinfusion injury.OBJECTIVE: To investigate the influences of Radix Astagali seu Hedysari (huangqi) on contents of NO and MDA in brain tissue of young rats with cerebral injury after cerebral ischemia and anoxia.DESIGN:A randomized and controlled trial.SETTING: Department of Pediatrics, Second Hospital, Hebei Medical University; Department of Internal Medicine, Institute of Traditional Chinese Medicine, Hebei Medical University; Department of Pathophysiology, Hebei Medical UniversityMATERIALS:The experiment was conducted from January to April 2004at Department of Pathophysiology, Hebei Medical University. Total 40 SD rats, 7-day old, were at random divided as normal control group, model group, humgqi low-dose group and huangqi high-dose group, with 10 rats in each group. Huangqi injection (The content in 10 mL injection is consistent with 20 g raw drug) was provided by Institute of Traditional Chinese Medicine of Hebei Medical University (produced in Chengdu Di'ou Jiuhong Pharmaceutical Factory, Batch No. 0005028).METHODS:Except rats in normal group, those in the rest groups, under conscious and local anesthesia, were all given common carotid artery ligation, establishing cerebral injury model due to ischemia and anoxia. Rats in normal group were intraperitoneally injected 0.1 mL normal saline; rats in model group were intraperitoneally injected 9 g/L normal saline, 0.1 mL each day; rats in huangqi low-dose group and huangqi high-dose group were respectively given 0.1mL, 0.5 mL huangqi injection, once a day, intraperitoneally. Cerebral blood flow was detected immediately, 2 and 4days after injection. Then the rats were decapitated for collecting the brains to measure the water content in brain, the contents of NO and MDA.MAIN OUTCOME MEASURES: [1] Water contents in brains of rats in every group. [2] Cerebral blood flow, and the contents of NO and MDA.RESULTS:Totally 40 rats were involved in the trial and all entered in the final result analysis. [1] The water content in brain of each group: Compared with normal group, the content in model group was increased immediately after model establishment [(87.316±0.275)%, (88.259±0.297)% ,P ＜ 0.05 ],and did not return to the normal level at the second day [(86.973±0.265)%,(88.173±0.445)%,P ＜ 0.05]; compared with model group, the content in huangqi high-dose group was obviously decreased at second day[(88.173±0.445)%, (86.542±0.141)% ,P ＜ 0.05]. [2] Measurement of cerebral blood flow: compared with control group, the blood flow in model group was obviously decreased immediately after model establishment[(231.88±13.33), (139.54±10.58)mV,P＜ 0.05], and did not return to normal level till the 4th day [(234.57±14.38), (145.38±13.33)mV,P ＜ 0.05];compared with model group, the blood flow in huangqi low-dose group and huangqi high-dose group, at day 4, was obviously increased [(145.38±13.33),(288.45±12.89), (313.82±21.74)mV,P ＜ 0.01]. [3] The contents of NO and MDA: The contents in model group, immediately after model establishment, were obviously higher than those in normal control group [(26.55±5.23 ), ( 19.67±7.17 )μmol/L,P ＜ 0.05; (7.88±2.55), (4.22±0.12) μmol/L, P＜ 0.01], and at day 4, were significantly higher than those in normal control group [(48.65±17.06), (18.65±2.12)μmol/L,P ＜ 0.01; (5.29±0.68),(4.06±0.39)μmol/L,P ＜ 0.05]; compared with model group, the contents in huangqi low-dose group and huangqi high-dose group were obviously decreased at day 4 [(48.65±17.06), (23.77±12.79), (24.67±11.54)μ mol/L,P＜ 0.01; (5.29±0.68), (4.51±2.30), (3.68±0.39)μmol/L,P ＜ 0.01].CONCLUSION:Huangqi could obviously reduce cerebral edema from ischemia and anoxia, increase cerebral blood flow. It could decrease the contents of NO and MDA that is metabolite of free radical injury, thus playing its role to inhibit lipid peroxidation injury.
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AIM:To observe the changes in heme oxygenase-1(HO-1) expression in the lung after ischmia-reperfusion of hind limbs in rats.METHODS:Hind limbs ischemia was made by clamping infrarenal aorta with a microvascular clip and lung injury was made by following reperfusion. Lung tissue was obtained from the animals subjected to sham operation, 4 h ischemia without reperfusion and 4 h, 8 h, 16 h, 24 h, 48 h reperfusion following 4 h ischemia. The levels of HO-1 mRNA and protein were measured at different times by Northern blot and Western blot. Immunohistochemistry technique was used to determine the cell types responsible for limb ischemic reperfusion induced HO-1 expression. RESULTS:After ischemia-reperfusion of limbs, HO-1 mRNA increased by 4 h, reached a peak at 16 h, and returned toward baseline at 24-48 h. This time course correlated with increased HO-1 protein. Immunohistochemical studies showed HO-1expressed in a variety of cell types, including the airway epithelium, alveolar macrophages and vascular smooth muscular cells. There were no positive signals in sham group and ischemia group both in mRNA levels and protein levels. CONCLUSION:The expression of HO-1 in the lung is not induced by limb ischemia or sham operation, but induced by limb reperfusion after ischemia in rats.
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AIM: To study the regulatory effect of curcumin on expression of heme oxygenase-1 (HO-1) in the lung of rat treated with LPS. METHODS: Eighteen rats were divided into three groups injected with different agents via lingua vein: control group (animals received equivalent saline) , LPS group (animals received a bolus dose of LPS 5 mg?0.5 mL-1?kg-1) and LPS+ curcumin group (animals received AP- 1 inhibitor curcumin 20 mg?0.5 mL-1?kg-120 min before the injection of LPS 5 mg ?0.5 mL-1?kg-1) . The expression of HO-1 mRNA and protein in the lung were examined 7 h after LPS administration by reverse transcribed polymerase chain reaction (RT- PCR) and Western blotting, respectively. Carboxyhemoglobin (HbCO) formation within pulmonary tissue was measured to represent CO content. RESULTS: The results showed that HO- 1 mRNA and protein expression as well as CO content in the lung of rats in LPS group were significantly higher than those in control group (P
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Objective:To investigate the vascular protective mechanism of different levels of herbs through detecting activity of HO-1/CO system in arterial tissues of rats with stagnancy of collateral-Qi in deficiency condition.Methods:Healthy male Wistar rats of cleanness level were randomly divided into control group,stagnancy of collateral-Qi in a deficiency condition group(SCQDC group),Renshen group,Shuangshen group,and Tongxinluo group.The ultrastructure of endothelium cells in the artery was observed.RT-PCR,Western blot,and immunohistochemical approaches were used to detect JNK,c-Jun mRNA and albumen expression in the arterial tissues.Meanwhile,the content of CO in the arterial tissue was also detected.Results:In SCQDC group,the counts of pinocytotic cells and microvili on the free surface of vascular endothelial cells were significantly decreased;most crests of mitochondria fused with membrane,some even disappeared;serious degranulation was observed in the rough surfaced endoplasmic reticulum.Immunohistochemistry showed that the HO-1 positive signals in the arterial histiocyte weakened significantly compared with the control group.The HO-1 expression was increased in the 3 treatment groups.Compared with control group,SCQDC group had decreased HO-1 mRNA and albumen expression(P
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AIM: To observe the changes in nitric oxide(NO) and peroxynitrite anion (ONOO - ) in the injuried lung following the ischemia-reperfusion of hind limbs and evaluate the contribution of NO and ONOO - to tissue injury. METHODS: A model of hind limbs ischemia was made by clamping infrarenal aorta with a microvascular clip and lung injury occurring after reperfusion. Lung tissue was obtained from the animals received sham operation(group 1),4 hours ischemia without reperfusion(group2), 1 hour reperfusion following 4 hours ischemia (group3) and 4 hours reperfusion following 4 hours ischemia (group4) . The contents of MDA, NO - 2/NO - 3 and the activities of SOD in the lung were examined. Immunohistochemical technique was used to determine the immunoreactivity to iNOS and nitrotyrosine(NT)-a specific "footprint" of peroxynitrite. RESULTS: Compared with group1 and group2,the contents of MDA and NO - 2/NO - 3 increased significantly (P