[The effects of vitamin E and dexamethasone on inflammation of acute lung injury and expression of myosin light chain kinase].

OBJECTIVE: To observe the effect of vitamin E (VitE) combined with dexamethasone (DXM) on inflammation of acute lung injury and expression of myosin light chain kinase. METHODS: Forty female Balb/c mice were randomly divided into 4 groups, a saline control group (1.5 ml/kg), a LPS group (1 mg/kg), a VitE and DXM group (VitE 50 mg/kg, DXM 1 mg/kg), and a VitE group (50 mg/kg). Lung tissue histopathological changes were observed. Immunohistochemistry assays (SABC) were used to determine the myosin light chain kinase (MLCK) immunoreactive cells in the lung tissues, and the MLCK mRNA and the MLCK protein was assayed by RT-PCR and by Western blot, respectively. Means were compared with analysis of variance and Student-Newman-Keuls were used to compare 2 means. RESULTS: Histological examination showed that extensive lung inflammation were seen in the LPS group, which manifested by accumulation of significant numbers of neutrophils, accompanied by marked pulmonary edema and hemorrhage. The inflammation and hemorrhage in the 2 treatment groups were significantly improved. Immunoreactive cells of MLCK numbers in BALF in the control group, the LPS group, the VitE and DXM group, and the VitE group was (1.1 +/- 0.4), (5.6 +/- 2.1), (4.0 +/- 1.0), (4.2 +/- 1.3) x 10(9)/L respectively. Compared with other groups, the difference of LPS group was significant (F = 14. 53, all P < 0.05). Immunoreactive cells of MLCK located airway epithelial and endothelia in the LPS group were more than which in the control group, decreased immunoreactive cells of MLCK in two treatment groups. Compared with LPS group, the difference of MLCK mRNA expression (A) of lung tissue in two treatment groups was no significant (F = 2.76, all P > 0.05). Compared with LPS group, the difference of A values of MLCK protein of lung tissue in two treatment groups was statistical significance (F = 12.06, all P < 0.01). CONCLUSIONS: Vitamin E combined with low dose of DXM could effectively inhibit inflammation and expression of MLCK protein in acute lung injury induced by LPS lung. It is suggested that, inhibition of MLCK activation leads to stabilize vascular barrier function and attenuation of pulmonary edema and inflammation, which also suggests a possible role of MLCK in the pathogenesis of acute lung injury.

[Protective effect of myosin light-chain kinase inhibitor on acute lung injury].

OBJECTIVE: To investigate the influence of inhibitor of myosin light-chain kinase (MLCK) on the human pulmonary arterial endothelial cell (HPAEC) challenged with lipopolysaccharide (LPS) and LPS induced of acute lung injury (ALI) in mice. METHODS: HPAECs were cultured in ECM medium and its passages 4-6 were used. After treatment with inhibitor of MLCK (ML-7) for 60 minutes, the HPAECs were incubated in LPS for another 60 minutes, and then cell viability was measured by the methyl thiazolyl tetrazolium (MTT) assay. Immunofluorescence microscope was used to detect phosphorylated-MLCK (p-MLCK) immunoreactive cells. Twenty female BALB/c mice were randomly divided into two groups. The mice of LPS group were exposed to LPS (1 microg/g) through nasal instillation, and the mice of ML-7 group were pretreated with ML-7 before intranasal instillation of LPS. Wet/dry weight (W/D) ratio of lung, bronchoalveolar lavage fluid (BALF) protein content, myeloperoxidase (MPO) activity and histopathological changes of lung tissue were observed. Immunohistochemistry assays were used to determine the status of MLCK and CD11b immunoreactive cells in lung tissue, and expression of MLCK mRNA in lung tissue was assessed by reverse transcription-polymerase chain reaction (RT-PCR). Expression of MLCK protein in lungs was assayed by Western blotting. RESULTS: Compared with LPS group, increased absorbance (A) value of HPAEC was found in ML-7 group (P<0.01). Immunoreactive cells of p-MLCK were more reduced in the ML-7 group (P<0.05), and W/D ratio of lung, MPO activity and BALF protein content of lung tissue were decreased in ML-7 group (P<0.05 or P<0.01). Histological examination showed that an extensive lung inflammation was seen in mice of LPS group, with an accumulation of a large number of neutrophils, marked pulmonary edema and hemorrhage, but the inflammation and parenchymal hemorrhage was significantly alleviated in ML-7 group. Both MLCK immunoreactive cells located in endothelium and CD11b in infiltrated inflammatory cells were decreased in ML-7 group compared with those in LPS group. Compared with LPS group, MLCK mRNA and protein expressions (A) in ML-7 group were significantly decreased (both P<0.05). CONCLUSION: ML-7, an MLCK inhibitor, enhances activity of HPAEC induced by LPS and reduces expression of p-MLCK. It also reduces the LPS-induced infiltration of neutrophils in lung tissues, pulmonary edema and expression of MLCK and CD11b protein and MLCK mRNA in lung tissues, demonstrating that inhibition of activation of MLCK, leading to an abatement of phosphorylation of myosin light chain or MLCK, resulting in stabilization of vascular barrier function. The results suggest that MLCK has a crucial role in the pathogenesis of ALI.

[The expression levels of endothelin-1 and nuclear factor-kappaB in the lung tissue of acute pulmonary embolism and the effects of thrombolysis and dexamethasone].

OBJECTIVE: To study the expression levels of endothelin-1(ET-1) and nuclear factor-kappaB (NF-kappaB) in lung vascular endothelium, bronchial and alveolar epithelia in acute pulmonary thromboembolism (APTE), and to explore the effects of thrombolytic (urokinase, UK) or ant-inflammatory therapy (dexamethasone, Dex) on their expressions. METHODS: Forty rabbits were randomly divided into a control (C) group, a pulmonary thromboembolism (PTE) model group, an UK therapy group, a Dex therapy group, and an UK + Dex therapy group, with 8 rabbits each. The PTE model was established by intravenous injection of autologous blood clots. The pathological changes of the lung were examined with light microscope. By using immunohistochemistry, the expression levels of ET-1 and NF-kappaB P65 in lung vascular endothelium, bronchial and alveolar epithelia were examined. RESULTS: Histopathological study showed that lung injury was evident in the PTE, UK and Dex groups, but was mild in the UK + Dex group. In the PTE group, the expression levels of ET-1 in lung vascular endothelium, bronchial and alveolar epithelia were 0.331 +/- 0.036, 0.229 +/- 0.014 and 0.191 +/- 0.046, respectively, while the expression levels of NF-kappaB P65 were 0.245 +/- 0.036, 0.190 +/- 0.040 and 0.204 +/- 0.054, respectively. Their levels were significantly higher as compared to those of the control group and the UK + Dex group (all P < 0. 01). In the UK group, the ET-1 expression in lung vascular endothelium, bronchial and alveolar epithelia were 0.204 +/- 0.020, 0.163 +/- 0.017 and 0.137 +/- 0.013 respectively, significantly lower as compared to the PTE group (all P < 0. 05). In the Dex group, the NF-kappaB P65 expression was 0.175 +/- 0.024, 0.104 +/- 0.022 and 0.144 +/- 0.022 respectively, the difference being significant as compared with the PTE group (all P < 0. 05). In the UK + Dex group, the ET-1 expression was 0.186 +/- 0.033, 0.107 +/- 0.012 and 0.098 +/- 0.026 respectively, significantly different from the PTE group (all P < 0.05); the NF-kappaB P65 expression was 0.109 +/- 0.018, 0.062 +/- 0.023 and 0.093 +/- 0.019 respectively, significantly lower as compared with the PTE and the UK groups (all P < 0. 01). CONCLUSIONS: After APTE, thrombolytic and anti-inflammatory treatment could decrease acute lung injury induced by ET-1 and NF-kappaB activation. Thus, anti-inflammatory therapy and the use of ET-1 inhibitor or receptor antagonist should be considered in APTE.