| Introduction:Acute lung injury, a pulmonary complication induced by infection, shock and wound, may develop to the acute respiratory distress syndrome (ARDS) or multisystem organs failure (MOF). Its clinical features are serious hypoxemia, diffuse pulmonary soakage and the descent of pulmonary compliance. Due to its high incidence and mortality, ALI/ARDS has become one of the main concerns. In United States, approximately 100,000 people are infected with ARDS every year, with the survival rate of 50-60%. With the discovery and identification of toll-like receptors (TLRs), the new drugs aiming at the TLRs become a new hotspot in the field of medical research. Intervention in the TLR -NF-κB (nuclear factor-κB) signal transduction pathway may be a new therapy strategy and may bring hope for the septic shock and other inflammatory diseases.Eucalyptus globulus oil is extracted from eucalyptus globules Labill., with maincomponents of 1,8-cineole and a-pinene. It has been proved that the Eucalyptus globulus oil could alleviate the chronic airway inflammation, alleviate the injury of airway epithelium, and reduce the infiltration of inflammatory cells. It needs further research to prove, however, whether the Eucalyptus globulus oil has the effect on the acute lung injury. Therefore, our research aims at the toll-like receptor 4 (TLR4) to study the effect of Eucalyptus globulus oil on the acute lung inflammation. Objective:Setting up the rat's acute lung injury models induced by lipopolysaccharide inhalation and mice's acute lung injury models induced by lipoposaccharide via intranasal instillation, to measure the leucocyte in BALF, to observe lung pathology, as well as to measure the expression of TLR4. After treatment of prednisone and different doses of Eucalyptus globulus oil, effect of glococorticosteriod on acute lung inflation and TLR4 expression will be observed. The nuclear factor-KB will be measured using Western-blot analysis. Methods:The Sprague-Dawley rats were intratracheally instilled with lipopolysaccharide (LPS, 2mg-kg"' per day) for two days to induce acute lung injury, then treated with prednisone and three different doses of Eucalyptus globulus oil for three days. The mice were intranasally instilled with lipopolysaccharide to induce acute lung inflammation.(1) The lung was lavaged with saline, and the total number of leukocyte in BALF was counted.(2) Pathological sections were made of trachea, bronchia and lung, which fixed by 10% formalin. The change of inflammation in airway tissue was investigated after being stained with HE. Semi-quantitative analysis had been done according to the standard made by our laboratory,(3) Immunohistochemistry dye was made with the Envision two-step method.The result was measured by the image-analysis system.(4) NF-KB/p65 in rat's lung tissue was extracted and measured with Western-blot analysis.(5) After LPS inhalation, rats were treated with prednisone and short-term treatment with different doses of Eucalyptus globulus oil. The leucocyte number in B ALF, the lung pathology, TLR4 and the nuclear factor-KB were detected.(6) Measurement data was expressed as means! SD and analyzed with t-test and q -test. Ranked data was analyzed with u-test.Results:1. The model of rats' acute lung injury induced by LPSThe Sprague-Dawley rats were intratracheally instilled with lipopolysaccharide (LPS, 2mg-kg"' per day) for two days to induce acute lung injury. The leucocyte number in BALF increased apparently and the lung inflammation was more serious than that of the contrast.2. The expression of TLR4 in rats' respiratory tractThere was little expression of TLR4 in normal rats' respiratory tract, and no difference of distribution was shown in different sections of the tract. After being stimulated with LPS, the expression of TLR4 in bronchia and bronchiole was significantly higher than that in the contrast, and the expression in the bronchiole was higher than that in the trachea and the bronchia.3. The model of mice' acute lung injury induced by LPSIntranasal instillation with LPS (6mg-mll) could induce acute lung inflammation in mice model. The leucocyte number in BALF increased more significantly than that in the contrast and the lung inflammation was more serious than the contrast.4. The expression of TLR4 in respiratory tract of miceThere was little expression of TLR4 in respiratory tract of normal mice and thedistribution was asymmetric. The stimulation of LPS could increase the TLR4 expression in respiratory tract, and the TLR4 expression in trachea increased most apparently.5. The effect of Eucalyptus globulus oilThe number of leucocyte in the group of Eucalyptus globulus oil treatment, especially the 300 mg-kg"1 group, had significantly decreased than that in the models. The inflammation in the group of Eucalyptus globulus oil treatment had apparently been alleviated compared with the models. The expression of TLR4 decreased more significantly after Eucalyptus globulus oil treatment than that in the models. The treatment of Eucalyptus globulus oil could not inhibit the expression of NF-KB/p65.6. The effect of glococorticosteriod in therapy dosageThe prednisone in therapy dosage could alleviate the rat's lung inflammation. The leucocyte in BALF decreased apparently, and the lung inflammation was less serious than that in the model. The prednisone could alleviate the expression of TLR4, especially in the bronchia and the bronchiole. The expression of NF-icB/p65 in pulmonary histocyte was inhibited. Conclusions:1. The model of rat's acute lung injury could be replicated by intratracheal instillation with lipopolysaccharide (LPS, 2mg-kg~' per day) for two days. The model of mice's acute lung injury could be replicated via intranasal instillation with LPS.2. TLR4 was expressed in normal rats' and mice's respiratory tract. No significant difference of the distribution of TLR4 was found in rats' different sections of respiratory tract, but the distribution in mice' respiratory tract was asymmetric. The expression of TLR4 was reinforced in main bronchus and bronchioles when being stimulated by LPS, which showed that the LPS might activate the TLR4 signal transduction pathway.3. The acute lung injury could be induced by LPS intratracheal instillation.Eucalyptus globulus oil treatment could inhibit the infiltration of leucocytes in BALF as well as the inflammation in the lung tissue, and could inhibit the increase of TLR4 expression stimulated by LPS.4. The NF-icB/p65 in lung tissue could increase along with the TLR4 increase, which showed LPS could activate NF-icB/p65 through the TLR4 signal transduction pathway. Eucalyptus globulus oil treatment in a short term (3 days) could not inhibit the increase of NF-KB/p65 in lung tissue. However, in previous research, Eucalyptus globulus oil treatment (21 days) could inhibit the increase of NF- k B/p65 induced by LPS.
|
PDF Full Text Request