The Effect Of Whole Lung Lavage On The Expression Of Claudin-4 And E-cadherin In The Alveolar Epithelium Of Rabbits

Objective: To observe effect of whole lung lavage(WLL)on expression of claudin-4 and E-cadherin in alveolar epithelial cells of rabbits,and explore self-repair mechanism of lung injury caused by whole lung lavage.Methods: 15 rabbits were randomly divided into 3 groups,including control group,2 hours group after whole lung lavage(WLL 2h group),72 hours group after whole lung lavage(WLL 72 h group).Each group had 5 rabbits.The rabbits were fasted for 12 hours and given no drink for 6 hours before the operation.The rabbits were given a tracheotomy in the state of general anesthesia,and inserted into the tracheal intubation catheter.The left lung received oxygen inhalation of 160~200mL/min and the right lung was lavaged with normal saline of 10 mL(37?)in each time.The total volume of lavage was calculated according to 25~30mL/kg.Then making the rabbits,head lower,lavage fluid naturally drainaged.The rabbits in control group were given a tracheotomy in the state of general anesthesia,and inserted into the tracheal intubation catheter.The left lung received oxygen inhalation of 160~200m L/min and the right lung without lavage.After operation for two hours chest CT examination were carried out.The rabbits were sacrificed by blood-letting after chest CT examination.The rabbits in WLL 2h group and 72 h group,after chest CT examination at 2h and 72 h,respectively,were killed by blood-letting.Lung wet/dry(W/D)ratio was measured.The paraffin section of right side of the lung tissue wase used to observe pathological changes of lung tissues by hematoxylin eosin(HE)staining.The pathological changes of lung tissues were scored.Electron microscope was used to observe the ultrastructure of alveolar epithelial barrier.Immunohistochemical staining(SP method)was used to detect lung tissue expression of claudin-4 and E-cadherin and mean optical density value(OD)was used to express the relative expression quantity of claudin-4 and E-cadherin.Results:(1)Clinical manifestations: the rabbits in WLL 2h and WLL 72 h groups appeared shortness of breath,lip nasal skin cyanosis,occasional choking cough reaction when whole lung lavage,and these symptoms disappeared gradually after the operation was finished.The rabbit in control group were no the above changes.The rabbits in WLL 72 h group could eat and drink normally.(2)Rabbit lung CT scan: WLL 2h group rabbit showed no obvious abnormality in the left lung tissue,but the uneven grinding glass shadow in the right lower lung tissue.Control group and WLL 72 h group rabbit showed no obvious abnormity of bilateral lung.(3)The general morphology of lung tissue: WLL 2h group rabbits' right lung volume increased,and lower parts of the lungs became dark red.the surface of the lung can be seen the bleeding point.Section showed a small amount of foam liquid outflow,left lung showed no obvious abnormality.Both lung in WLL 72 h group and control group had smooth surface,good elasticity,and no significant bleeding points and edema.(4)The lung wet/dry weight ratio(W/D): Compared with the control group W/D ratio(4.34� 0.06),the lung W/D ratio(4.72� 0.09)in WLL 2h group was significantly higher(P<0.05).The lung W/D ratio(4.36�0.07)in WLL 72 h group decreased significantly than than in WLL 2h group(P<0.05).The lung W/D ratio in WLL 72 h group increased slightly compared with the control group(P>0.05).(5)HE staining: WLL 2h group showed capillary dilatation and congestion,and a small amount of inflammatory cell infiltration of alveolar wall,thickening of alveolar septa and exudation of alveolar liquid.But alveolar structure remained intact.WLL 72 h group rabbits showed slighter pathological change of infiltration of inflammatory cells,with basic integrated alveolar structure and no exudation of alveolar liquid.The control group showed no these pathological changes.Pathological scores(5.0�0.7)of WLL 2h group was significantly higher than that of control group(0.2�0.4)and WLL 72 h group(1.2�0.4)(P<0.05),and there was increased slightly pathological scores in WLL 72 h group than the control group(P<0.05).(6)Electron microscopy: Control group showed a integrated tight junction structural of alveolar epithelial cells.But the tight junction injury of WLL 2h group were seen,epithelial intercellular space widened,with cell breakage of type I alveolar epithelial cells.WLL 72 h group showed no these changes.(7)Immunohistochemical: Claudin-4 and E-cadherin in control group was mainly expressed in alveolar epithelial cell envelope and a little in patina.Claudin-4 and E-cadherin in alveolar epithelial cells of WLL 2h group were expressed discontinuous and significantly weaker than those in the control group.Expression of claudin-4 and E-cadherin in alveolar epithelial cells of WLL 72 h group was increased compared with that of WLL 2h group,but still slightly lower than that of control group.The OD value of claudin-4 and E-cadherin expression in WLL 2h group were(0.93� 0.12)and(1.12�0.09),respectively,there being significantly lower value compared with the OD value of claudin-4(1.44�0.05)and E-cadherin(1.74� 0.05)in the control group and WLL 72 h group.The OD value of claudin-4 and E-cadherin expression in WLL 72 h group were(1.32�0.04)and(1.62�0.07),respectively,there being a slightly lower compared with those of control group(P<0.05).Conclusion:(1)Whole lung lavage can lead to a certain degree of rabbits alveolar epithelial barrier injury.(2)The expression of claudin-4 and E-cadherin in rabbits alveolar epithelium after whole lung lavage were decreased.(3)Whole lung lavage associated lung injury can self-repair in a certain period of time.(4)Expression of claudin-4 and E-cadherin may be involved in the clearance of lung lavage fluid and the repair of lung injury.