Protective Effect Of Apelin-13 On Ventilator-induced Acute Lung Injury

Objective:Mechanical ventilation(MV)is an essential clinical life support modality;while serving a therapeutic purpose,it may also lead to mechanical ventilation-associated lung injury(VILI).It has been found that the apelin/APJ axis is an endogenous countermeasure that is activated during ARDS to counteract the injury response and prevent uncontrolled lung injury.Therefore,in the present study,we investigated whether apelin-13 further activates the apelin/APJ axis during VILI by constructing a rat model of VILI,which results in a protective effect.Methods:SD rats were divided into 4 groups: control group(group C),high tidal volume group(HV group),high tidal volume + NS group(NS group),and high tidal volume +apelin-13 group(apelin-13 group).The control rats were tracheotomized and retained to breathe on their own,and the remaining rats in each group were tracheotomized and connected to a small animal ventilator for 4 hours after ventilation in order to establish a VILI rat model.The expression of apelin m RNA was detected by quantitative real-time polymerase chain reaction(q RT-PCR),and the expression levels of APJ and apoptotic proteins Bax and Bcl-2 protein were detected by immunoblotting(WB)and immunofluorescence.The degree of lung injury was assessed by lung histopathological staining and wet-to-dry ratio of lung tissue,total protein concentration of bronchoalveolar lavage fluid(BALF),the degree of inflammatory response of lung tissue was assessed by enzyme-linked immunosorbent assay(ELISA)to detect the expression of inflammatory factors in BALF,and the activity of myeloperoxidase(MPO),an antioxidant stress indicator,was measured in lung tissue.Results:(1)A VILI rat model was successfully established after 4 hours of mechanical ventilation.The expression of apelin m RNA was measured by q RT-PCR,and the expression level of APJ protein was measured by WB and immunofluorescence.The results showed that during VILI,the expression of apelin m RNA and APJ protein in HV and NS groups increased significantly compared with group C.After apelin-13 agonist intervention,the expression of apelin m RNA and APJ protein expression were further significantly increased in the apelin-13 group after apelin-13 agonist intervention.This suggests that there is a compensatory increase in apelin m RNA and receptor APJ during VILI,and that the signaling of the apelin/APJ system is further enhanced after the administration of activated apelin-13.(2)Lung injury and edema by detecting the total protein concentration of bronchoalveolar lavage fluid,lung tissue wet/dry ratio,lung histopathological staining and Smith score showed that the total protein content of BALF and lung wet/dry ratio were significantly increased in HV and NS groups compared with C group;HE staining showed severe lung injury in HV and NS groups compared with C group,and lung histology was characterized by perivascular edema,interstitial and intra-alveolar leukocyte infiltration,and significant heterogeneity in alveolar swelling;while the lung injury in the apelin-13 group was significantly less than that in the HV and NS groups;the Smith score assessment of lung injury in each group was consistent with the pathological staining results.(3)The levels of IL-1β,IL-6 and TNF-α in BALF,an inflammatory factor in BALF,were measured by ELISA,and the results showed that the protein expression of IL-1β,IL-6 and TNF-α in BALF in the HV and NS groups was significantly increased compared with group C;after apelin-13 intervention,the expression of IL-1β,IL-6 and TNF-α significantly After apelin-13 intervention,the expression of IL-1β,IL-6 and TNF-α decreased significantly,indicating that apelin-13 could effectively reduce the lung inflammatory response in VILI rats.(4)The expression of anti-apoptotic protein Bcl-2 and pro-apoptotic protein Bax was detected by WB technique,and the results showed that the Bcl-2/Bax ratio in the lung tissue of apelin-13 group was significantly higher after intraperitoneal injection of apelin-13 compared with HV and NS groups,indicating that apelin-13 was involved in the process of inhibiting apoptosis.(5)The viability of MPO,an antioxidant stress indicator,was measured by microenzymatic assay,and the results showed that a significant increase in MPO viability was found in the HV group as well as the NS group compared to the C group.After the administration of Apelin-13 intervention,MPO activity was significantly lower in the apelin-13 group compared to the HV and NS groups,indicating that apelin-13 was able to slow down the degree of oxidative stress.Conclusions:The apelin/APJ system is activated in VILI.Overexpression of apelin further enhanced the signal transduction of apelin/APJ system,which resulted in protective effects,including reducing the inflammatory response in lung tissue,inhibiting apoptosis and anti-oxidative stress,and delaying the development of VILI.