The Research Of Relationship Between Liver And Acute Lung Injury In The Early Intestinal Endotoxemia

Objective: To reveal the relationship between liver and lung injury in the early stage of intestinal endotoxemia by investigating the different levels of acute lung injury in the method of injecting LPS at the same dose via portal vein and vena cave. Method: 72 healthy and clean SD Rats weight 220±20g, then randomly assigned them to one of two groups: control group(FITC, F Group=36), and LPS group(FITC-LPS, L Group=36) by using the random number table. The two groups divided into the portal vein subgroup(M Group=18) and vena cava subgroup(Q Group=18); each subgroup subdivided into 3 sections in accordance with 15 min, 30 min and 60 min, there were 12 sections totally and each section contained 6 rats. Rats of control group and LPS group were injected by 1 ml FITC-0.9% saline solution and 1ml FITC-LPS-0.9% saline solution via vein and vena cava respectively. Each group abstracted blood of the rats to measure blood serum LPS, CC16, and plasma TNF-α level before executing them according to different time points. Lung tissue was taken to measure lung water content(the ratio of left lung wet / dry weight) after the rats were executed; Measure distribution of fluorescence labelled endotoxin in lung tissue; Damages of lung tissue were measured by HE staining; Examined the expression of CC16 in lung tissue by Immunohistochemical methods; and observed the ultra-microstructure of Clara cell by transmission electron microscope.Results: 1. The content of plasma lipopolysaccharide: LPS group was significantly higher than the control group, this was statistically significant(P <0.05). Comparing LM group and LQ group, Plasma lipopolysaccharide levels in the LM group was significantly reduced compared with LQ group, Both differences were statistically significant at all time points(P <0.05). 2. The content of plasma Clara cell protein(CC16): L group was higher than F group, this was statistically significant(P <0.05). And plasma CC16 content was higher in the LM group than in the LQ group, and there were statistically significants in three time periods(P <0.05). In the LM group and LQ group,the growth rate of CC16 content at 30min-60 min were lower than it at 15min-30 min. 3. The content of rat plasma TNF-α: L group was higher than F group, which was statistically significant(P <0.05). The content of rat plasma TNF-α in the LM group was higher than in the LQ group, differences at 15 min,30min,60 min were statistically significants(P <0.05). 4. Distribution and intensity of fluorescein in lung tissue: expression of fluorescein of the L group was more than F group,which was statistically significant with time increasing. The LQ group was significantly higher than the LM group. 5. Detection of rat lung tissue water content(the ratio of left lung wet / dry weight) : L group was higher than the control group, compare was statistically significant(P < 0.05). lung tissue water content in the LM group was higher than in the LQ group, differences at 15 min,30min,60 min were statistically significant(P <0.05). 6. Observe damages of lung tissue by light microscopy: F group: Alveolar structural integrity,alveolar sacs clear,alveolar septum normal.At 15 min,LM group: obvious infiltration of neutrophils and macrophages; LQ group: Alveolar epithelial cells moderate edema, and widened the alveolar septum. At 30 min, LM group: A large number of inflammatory cell infiltrates in alveoli and alveolar cell edema or bleeding, and alveolar septum broadening, forming lymphoid follicles; LQ group:Alveolar epithelial height edema, alveolar expansion, a small amount of inflammatory cell infiltration. At 60 min,LM group and LQ group:alveolar structure disappeared, alveolar consolidation and expansion, infiltration of a large number of neutrophils and macrophage, cell significant edema. The lung injure of LM group was more serious than LQ group, there was significant difference between the two groups in lung injure score(P <0.05). 7. Lung tissue Clara cell protein(CC16) Immunohistochemistry: Positive staining: cytoplasm showing brown is rat lung Clara cell protein(CC16). Compare L group, the score of F Group was significantly higher, expression was moderately positive,the difference was statistically significant(P <0.05). LM group and LQ group showed a weak positive at each time point, LQ group scores was higher than theLM group, the difference was statistically significant(P <0.05). 8. observe the ultra-microstructure of Clara cell by transmission electron microscope: FM group and FQ group: Cell body large, round or oval, There are a small number of short microvilli in the cell surface, top cells hat-shaped were broke into the lumen. There were slip surface endoplasmic reticulum, ribosomes and mitochondria in the clara cell cytoplasm. The cell cytoplasm have dense granules,which membrane wrapped and uniform electron density. These dense particles were Clara particles. These particles shifted side, rounding the top of the cell membrane. LM group and LQ group at 60 min: Clara cells were denaturation obviously, nuclear shrinkage, nuclear membrane retraction, perinuclear space passed off. Mitochondria, smooth endoplasmic reticulum, secretory granules were reduced in the cytoplasm. Clara particle density was uneven, which was degranulation state. Conclusion: 1. Kuffer cells were activated and large amount of pro-inflammatory cytokine were released into the blood circulation in the early stage of intestinal endotoxemia. It may explain why severe lung injury occurred. 2. Direct stimulation of endotoxin to lung may be relative to cytokine pre-booting lung protection system,etc.