| Objective: Acute lung injury/acute respiratory distress syndrome (ALI/ARDS) refers to acute progressive respiratory failure with stubborn hypoxemia induced by different predisposing factors except cardiogenic causes. ALI and ARDS have the same manifestations in the changes of pathophysiology, ARDS is the most severe form of ALI. The patho-base of ALI/ARDS is mediated by many kinds of inflammatory cell, such as macrophage, neutrophil cell, leukomonocyte and so on, which lead to the inflammatory reaction of lung and the injury of capillary membrane induced by the loss of control of the inflammatory reaction. The high risk factors of ALI/ARDS can be divided into two cases: a direct insult on lung cells (pulmonary ARDS, ARDSp), such as severe pneumonia, aspiration of gastric contents, pulmonary contusion, and so on; indirectly (extrapulmonary ARDS, ARDSexp) such as sepsis, serious nonthoracic trauma, serious pancreatitis, and so on. As much as 30% of ALI/ARDS is associated with aspiration of gastric contents, ARDS caused by aspiration of gastric contents is the serious complication of the lung, the operation of tracheal intubation or incision of trachea can increase the incidence rate of inhalation.During the development of ARDS, the contribution of both pro- and anti- inflammatory cytokines mediates the advancement of inflammation. Some animal experiments have confirmed many drugs including PGE1, pulmonary surfactant and Bisolvon, have protect effect of ALI, which is induced by hydrochloric acid. Despite improvement in supportive care and multiple therapeutic efforts directed at modifying the course of this condition, mortality still remain about 50%. Prompt and significant solutions are required. Although considerable progress has been made in the inflammatory mechanism of ARDS, we have not more information about the similarity and difference of the inflammatory mechanism (such as the permeability and the contents of the inflammatory mediators) by different reasons. The glucocorticoids inhibit the inflammatory pathways at virtually all level, but the therapy with glucocorticoid in ARDS is ambivalent. Glucocorticoid has the effect of anti-inflammatory, adjust immune function, meanwhile, it can regulates the growth and apoptosis of cells, which is mediated by glucocorticoid receptor, glucocorticoid receptor is translate by GRmRNA in cells. Because of the complex of ALI/ARDS, ALI/ARDS caused by different reasons have the same place in pathology and pathophysiology, but they have the difference in its development, growth and prognstic. So they have the different therapeutic efficacy. Therefore, our experiment establishes two animal models of ALI/ARDS to compare their differences in the cytokines levels, the express amount of GRmRNA in lung and the effect of GCs on the prognosis of ALI/ARDS and expression of GRmRNA. We hypothesized that the effect of GCs to these two animal models may exists difference, to explore the mechanism of GCs or the change of GRmRNA and cytokine to ALI, which can provide theoretic evidences for clinical physicians to choose the of use GCs in the treatment of ALI/ARDS with aspiration of gastric contents.Methods: 48 Male SD rats of 250 to 300 g were randomly divided into six groups:①NS group,②HCl group,③LPS+HCl group,④NS+DEX group,⑤HCl+DEX group,⑥LPS+HCl+DEX group. Every group has 8 animals.④,⑤,⑥three groups were intraperitoneal injection with Dexamethasone (2mg/kg) after replicating model.①,④two groups were control group, 0.9% saline was instilled into the trachea;②,⑤two groups were HCl group, HCl(pH 1.8, 2ml/kg) was instilled into the trachea;③,⑥two groups were LPS+HCl groups, LPS(4mg/kg) was injected intravenously via a tail vein, 4 hours later, HCl(pH 1.8, 0.5ml/kg) was instilled into the trachea.4 hours later, rats were given intraperitoneal injection anesthesia of 4ml/kg 10% chloral hydrate. Blood sampling was obtained from heart in a breast operation. Blood was collected and centrifugated (3000rpm, 4℃, 10min). Serum was stored in -80℃until processing. Taking the anterior lobe of right lung, which weight was 100mg, used for extract general RNA. The inferior lobe of right lung was placed in 10% neutral formalin fluid, embedded in paraffin, and stained with hematoxylin-eosin. The wet/dry was measured with the other section of the right lung (80℃, 48h). Bronchoalveloar lavage fluid was obtained by bronchoalveloar lavage with 0.9% saline from left lung. BALF was collected and centrifugated (1200rpm, 4℃, 10min). The sediment was infused with 0.9% saline to be counted the total leukocytes. Supernatant fluid was stored in -80℃until processing. The total protein in BALF was determined by using the Comas Blue assay. The pro-inflammatory cytokines TNF-alpha, IL-1beta and the anti-inflammatory cytokines IL-4, IL-10 were detected by the solid phase enzyme linked immunosorbent assay (ELISA) both in serum and BALF.Results: (1) In the groups of HCl and LPS+HCl, the total leukocytes were higher than those of the control group (P<0.01). No difference existed in HCl group and HCl+DEX group, LPS+HCl group and LPS+HCl+DEX group existed no difference too. PMN% in BALF of the two modle groups were higher compared with the control group(P<0.01)。PMN% in LPS+HCl+DEX groups was lower than LPS+HCl groups (P<0.01). No difference existed in HCl groups and HCl+DEX groups. (2) In the groups of HCl and LPS+HCl, the GRmRNA/β-actinmRNA were lower than those of the control group (P<0.01). Comparing to corresponding model groups, the GRmRNA/β-actinmRNA in LPS+HCl+DEX groups increased (P<0.05), but not in HCl+DEX groups. (3) In the groups of HCl and LPS+HCl, the protein concentration in the BALF and W/D were higher than those of the control group (P<0.05). Comparing to LPS+HCl groups, W/D in LPS+HCl+DEX groups decreased (P<0.05). The protein concentrations in HCl+DEX groups were lower than those of HCl groups (P<0.05). (4) In serum and BALF of both of HCl group and HCl+LPS group, the concentrations of TNF-αwere higher than those of the control (P<0.01). Comparing to LPS+HCl groups, TNF-αin LPS+HCl+DEX groups decreased (P<0.05). The content of IL-1βin serum of all the groups was undetected. The concentrations of IL-1βin BALF were higher than those of the control (P<0.05). (5) In serum and BALF of both of HCl group and LPS+HCl group, the concentrations of IL-4 and that of IL-10 were higher than those of the control (P<0.05). The LPS+HCl+DEX group and the LPS+HCl group had similar concentration of IL-4. In serum and BALF of LPS+HCl+DEX groups, the concentrations of IL-10 were higher than those of the control (P<0.05). Comparing to HCl groups, IL-10 in BALF in HCl+DEX groups increased (P<0.01).The lungs were edema and hemorrhage both in HCl and LPS+HCl groups. The LM result showed that two model manifested with acute and diffused alveolar damage including necrosis of the endothelia and epithelia, leukocyte accumulation in lung tissue, pulmonary edema, profound lung inflammation. The histological grades were consistent among the two model groups.Data are expressed as means±SEM. The DEX intervening differences were analyzed by group t-test. The group differences were analyzed by one-way analysis of variance (ANOVA) using SAS6.12 software. P<0.05 was considered statistically significant.Conclusion:1 The rats in the model group by instilling HCl into the trachea, which is called"one-hit"and by intravenous injection of LPS, than instilling HCl into the trachea, which is called"two-hit", both characterized with diffuse alveolar damage and manifest with severe systemic inflammatory response, which indicated we succeed in replicating ARDS models.2 The pulmonary morphology, the permeability, the inflammatory cytokines and the glucocorticoid receptor mRNA in these two models is not consistent. It can be concluded that pathogenesis of ARDS is different by different risk factors.3 Dexamethasone can alleviate the pulmonary inflammatory injury caused by"two-hit", but not useful to ALI caused by HCl. So curative effect of glucocorticoid is associated with clinical reasons.
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