The Research On The Pathogenesis And Protective Methods For The Lung Injury Induced By Skeletal Muscle Ischemia Reperfusion

Background and Objective: The skeletal muscle ischemia-reperfusion (IR)injury is a common clinical issue. Prolonged limb ischemia followed byre-establishing reperfusion may worsen the initial ischemic injury, and paradoxicallyresult in the damage of remote vital organs. This complex damages may even lead tosystemic inflammatory response syndrome (SIRS) or multiple organ dysfunctionsyndrome, MODS). The lung is one of the most susceptible remote organs. Lunginjury is one of the most common complications of skeletal muscle IR and traumaticinjury, which can trigger acute lung injury (ALI) and acute respiratory distresssyndrome (ARDS) and cause high morbidity and mortality in critically ill patients.The underlying mechanism is complex which may involve inflammation, edema,excess oxidative stress, activation of neutrophil and inflammatory cytokines,downregulation of hydrogen sulfide (H2S)/cystathionine-γ-lyase (CSE), activation ofrennin angiotensin aldosterone system (RAAS), etc. However, the early recongnitionand treatment of lung injury resulted from limb IR and traumatic injury is notsatisfying, and the immune response mechanism and water transport abnormality isnot very clear.Our study was divided into three parts. Firstly, we determined the blood levelsof clara cell secretion protein (CC16) as a lung epithelium specific biomarker, andtumor necrosis factor (TNFα), interleukin (IL)-1β, IL-6, IL-8as imflammatorymarkers to verify lung injury during bilateral total knee arthroplasty. Secondly, weestablished the rat model of bilateral hindlimb ischemia and the model of combinedtrauma to explore the role of aquaporin1(AQP1), aquaporin5(AQP5) andTLR4-MyD88-NF-kB signal pathway in the process of lung edema and inflammatoryinjury. Thirdly, we invest the regulation roles and mechanism of exogenous H2S andaldosterone receptor blocker spiron, and provide a feasible strategy for the treatment of lung injury resulting from limb IR.Methods: In the first part,15patients who were given bilateral total kneearthroplasty were included. Blood samples were harvested before operation, at4h orat12h postoperatively. Blood CC16, TNFα, IL-1β, IL-6and IL-8level weredetermined by Elisa or Lumines. In the second part,22Wistar rats were randomlydivided into3groups:①normal control group (n=8),②IR group (n=8):4hreperfusion following4h ischemia of bilateral hind limbs,③combined trauma group(n=6): IR combined with injury of bone and soft tissues. The lung CC16level wasmeasured and the lung injury and edema were evaluated. The protein and mRNAlevels of AQP1, AQP5, TLR4, Myd88, and NF-κB were determined by Western Blotor Real-time PCR method. In the third part,40Wistar rats were randomly divided into5groups (n=8):①normal control group,②IR group,③IR+NaHS group:pretreatment of NaHS at dosage of0.78mg/kg before IR, ip.④IR+PPG group:pretreatment of DL-propargylglycine (PPG) at dosage of60mg/kg before IR, ip.⑤IR+spiron group: IR+Spiron at dosage of20mg/kg/day6days before IR, po. Theprotein and mRNA levels of AQP1, AQP5, TLR4, Myd88, and NF-κB, as well as themRNA level of CC16, were determined.Results: The blood CC16, TNFα, IL-6and IL-8levels were significantlyincreased at the time point of4h postoperatively compared with preoperative level(P<0.05). There was obvious lung injury and edema in IR group accompanied withupregulation of TLR4-Myd88-NF-κB pathway and downregulation of AQP1andAQP5. Compared with IR group, the lung injury and edema aggaravated and thechangment of AQP1, AQP5and TLR4-Myd88-NF-κB pathway were more obvious.NaHS and spiron pre-treatment reduced lung injury with increasing AQP1, AQP5expression and inhibition of TLR-Myd88-NF-κB pathway, but PPG adjusted AQP1,AQP5and TLR4pathway to the opposite side and exaggerated lung injury.Conclusions: The operation of bilateral total knee arthroplasty could lead toincrease blood levels of CC16, TNFα, IL-6and IL-8, which might be responsible for lung epithelium damage and inflammatory injury. Skeletal muscle IR and combinedtrauma could lead to lung inflammatory injury and water transport abnormality. AQP1,AQP5and TLR4-Myd88-NF-κB pathway were involved in limb IR and combinedtrauma induced lung injury. Increased endogenous H2S or the pretreatment of spironwould improve lung injury and edema, and their protective effect at least partiallydepend on the upregulation of AQP1, AQP5expression and the inhibition ofTLR4-Myd88-NFκB pathway and to improve pulmonary edema and reduceinflammatory reaction.