The Role Of MTOR In Lipid Metabolism Of Diaphragmatic Fibers Caused By Mechanical Ventilation

Objective:Mechanical ventilation is clinically effective for the treatment of critically ill patients with respiratory failure and respiratory support,however,there are many factors lead to diaphragmatic atrophy and contraction dysfunction in the process of mechanical ventilation,namely ventilator-induced diaphragm dysfunction(VIDD),which shows mainly weaning failure.The mechanism of VIDD is complex.The main physiopathologic changes include reduced diaphragmatic protein synthesis and increased proteolysis and oxidative stress that is associated with mitochondrial dysfunction,but early onset and regulating mechanism of VIDD is unclear.Metabolic substrate is in s state of relative overload,which and lipid accumulation are potential triggered factors of mitochondrial damage in mechanical ventilation.m TOR signaling pathways involve in the regulations of a variety of cellular activities.A study shows that activated m TOR involves in foam cell formation and development of atherosclerosis by the sterol regulatory element-binding proteins-2(SREBP-2),but its regulatory mechanism of lipid metabolism in the VIDD is not well known.Therefore,this study explores the impact of m TOR on VIDD and possible mechanism.The purposes of this study include the following:(1)to set up diaphragmatic dysfunction model of mechanical ventilation of rats;(2)to research the impact of m TOR on contraction function,structure of the fiber,lipid metabolism and mitochondria of diaphragm.Methods:Spraguee-Dawley rats were randomly divided into control group(CON group),control ventilation group(CMV group)and control ventilation and inhibitors of m TOR group(CMV+RAPA group)(n=8).In the CON and CMV groups,the equivalent volume of DMSO was injected intraperitoneal injection(i.p.)once a day for two days.In the CMV+RAPA group,20 mg/kg of rapamycin was injected i.p.once a day for two days.After CMV and CMV+RAPA groups had 12 h of control ventilation,taking diaphragm to measure diaphragmatic contractility through biological signal acquisition system.Morphological changes of diaphragm were observed by HE staining between groups.The ultrastructure of diaphragmatic fibers was observed by electron microscopy.Lipid accumulation was observed by oil red staining.The expression of m TOR,phosphorylated-m TOR and SREBP-2 was measured by western blot.Results:(1)Diaphragmatic contractility:Compared with CON group,peak twitch tension(Pt)and maximum tetanic tension(Po)of diaphragm decrease obviously in CMV group(P<0.05);CMV+RAPA group has no significant difference(P>0.05).Compared with CMV group,the Pt and Po increase obviously in CMV+RAPA group(P<0.05).Compared with CON group,the tension-frequency curves move down in CMV and CMV+RAPA groups.The tensions of 10 and 20 Hz have no significant difference(P>0.05)and the tensions of 40,60,80,100 and 120 Hz decrease obviously(P<0.05)in CMV group.CMV+RAPA group has no significant difference(P>0.05).Compared with CMV group,the curve moves up but has no significant difference in CMV+RAPA group(P>0.05).(2)Morphological detection of diaphragm:HE staining shows that the morphology of diaphragmatic fibers is almost normal in CON group.Compared with CON group,the muscle bundle of diaphragmatic fibers have small clusters of atrophic muscle fibers and interstitial edema in CMV group.Compared with CMV group,the structure of diaphragmatic fibers have slight damage in CMV+RAPA group and no significant difference compared to CON group.(3)The ultrastructure of diaphragm:The observation of electron microscope shows that the muscle bundle of diaphragm is normal in CON group.Compared with CON group,the muscle bundle is edematous and the interval widens,as well as sarcomeres are disordered and deficient in CMV group.Compared with CMV group,the damage of muscle bundle is alleviative in CMV+RAPA group.(4)Lipid metabolism of diaphragm:Oil red staining shows that compared with CON group,lipid accumulation of diaphragmatic fibers increases obviously in CMV group(P<0.05).Compared with CMV group,lipid accumulation of diaphragmatic fibers reduces significantly in CMV+RAPAP(P<0.05).(5)Activity detection of m TOR:The results of western blot show that compared with CON group,the expression of m TOR and SREBP-2 of diaphragmatic fibers increase significantly in CMV group(P<0.05).Compared with CMV group,the expression of m TOR and SREBP-2 of diaphragmatic fibers decrease significantly in CMV+RAPA group(P<0.05).Conclusion:The model of diaphragmatic dysfunction is set up successful after 12 h of control ventilation in the rats.m TOR is activated which causes increased lipid accumulation and damaged mitochondria.The structure of diaphragmatic fibers is damaged.The diaphragmatic contractility decreases and the contraction of diaphragm is impaired.Rapamycin,the inhibitors of m TOR,can inhibits activity of m TOR.Then it reduces lipid accumulation and mitochondrial damage and improves diaphragmatic dysfunction caused by mechanical ventilation.m TOR plays an important regulating role in lipid metabolism of diaphragmatic fibers caused by mechanical ventilation.The activation of m TOR leads to abnormal lipid metabolism which may be underlying mechanisms of early onset of VIDD.