| Objective:1.To study the role of lncRNA MALAT1 in lung ischemia reperfusion in jury(LIRI);2.To explore the intrinsic relationship between lncRNA MALAT1 expressi on and IL-8 high expression in lung ischemia-reperfusion injury,and to provid e a new therapeutic target for lung transplantation ischemia-reperfusion injury.Method:1.(1)Thirty male rats were randomly divided into three groups with 10 rats in each group,namely Blank group,Sham group and LIRI group.After thoracotomy,LIRI group dissociated the hilum and severed the lower lung ligament,and used non-invasive vascular clamp to clamp the left lung hilum for 90min.After reperfusion for 180min,the rats were euthanized and pulmonary vein blood and left lung tissue were collected for detection.After thoracotomy,Sham group only dissociated the hilum and severed the lower lung ligament without clamping the hilum,closed the chest 90min later,closed the chest 180min later,euthanized the rats and collected pulmonary vein blood and left lung tissue for detection.Blank does not do anything.(2)The partial pressure of pulmonary vein oxygen was measured to verify the successful establishment of LIRI rat model.(3)HE staining of lung tissue was observed under optical microscope to evaluate pathological changes of lung injury.(4)Calculate the wet/dry(W/D)weight ratio of lung.(5)Immunohistochemistry(IHC)was used to detect the positive expression of macrophages in lung tissue.(6)The expression level of inflammatory factors in serum was detected by enzyme linked immunosorbent assay(ELISA).(8)qRT-PCR was used to detect the expression level of MALAT1 in lung tissue.2.(1)Forty male rats were randomly divided into four groups with 10 rats in eachgroup,namely Blank group,Sham group,sh-NC group and sh-MALAT1 group.The construction of Blank group and Sham group is the same as the previous part,sh-NC group and sh-MALAT1 group were injected with adenovirus of sh-NC or sh-MALAT1 via tail vein respectively on the basis of LIRI rat model.After one week,rats were euthanized and pulmonary vein blood and left lung tissue were collected for detection.(2)The expression of MALAT1 was detected by qRT-PCR and the pulmonary vein oxygen partial pressure after pulmonary ischemia reperfusion was measured to verify the successful establishment of sh-MALAT1 rat model.(3)HE staining of lung tissue was observed under optical microscope to evaluate pathological changes of lung injury.(5)Calculate the W/D weight ratio of lung.(6)IHC was used to detect the positive expression of macrophages in lung tissue.(7)The expression of interleukin-8(IL-8)in serum was measured by ELISA.Result:1.(1)Pulmonary vein oxygen partial pressure(PO2)in LIRI group was significantly lower than that in Blank group and Sham group,the difference was statistically significant(P<0.05).(2)HE staining showed that compared with Blank group and Sham group,LIRI group observed more severe alveolar hemorrhage,edema and interstitial inflammation.(3)The positive expression of macrophages in LIRI group was higher than that in Blank group and Sham group,the difference was statistically significant(P<0.05).(4)The number of apoptotic cells in LIRI group was higher than that in Blank group and Sham group,the difference was statistically significant(P<0.05).(5)Compared with the Blank group and the Sham group,the wet/dry(W/D)ratio of LIRI group increased,and the difference was statistically significant(P<0.05).(6)IL-6,IL-8,tumor necrosis factor-?(TNF-?),IL-4 and IL-10 all increased in LIRI group.Among all inflammatory factors tested,the expression of IL-8 was highest in the peripheral vein.The expression of IL-8 in LIRI group was significantly higher than that in Blank group and Sham group,the difference was statistically significant(P<0.05).(7)The expression of MALAT1 was detected by qRT-PCR.the results showed that the expression of MALAT1 in LIRI group was significantly higher than that in Blank group and Sham group,and the difference was statistically significant(P<0.05).2.(1)qRT-PCR results showed that MALAT1 in sh-MALAT1 group was significantly lower than that in sh-NC group,and the difference was statistically significant(P<0.05).(2)Pulmonary vein blood gas analysis showed that pulmonary vein PO2 in sh-MALAT1 group was higher than that in sh-NC group and Sham group,the difference was statistically significant(P<0.05).(3)HE staining showed that compared with sh-NC group,sh-MALAT1 group significantly reduced lung injury.(4)The number of positive macrophages in sh-MALAT1 group was significantly lower than that in sh-NC group,the difference was statistically significant(P<0.05).(5)Compared with sh-NC group,the W/D ratio of sh-MALAT1 group(4.56±0.77)was decreased significantly,and the difference was statistically significant(P<0.05).(6)ELISA showed that the expression of IL-8 in sh-MALAT1 group was significantly lower than that in sh-NC group,the difference was statistically significant(P<0.05).Conclusion:LIRI causes obvious lung tissue injury,and silencing MALAT1 can reduce the inflammatory injury of LIRI rats by reducing the expression level of IL-8. |
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