0.5MAC Isoflurane In60%Oxygen Protected Against Sepsis And The Role Of MiRNA

Sepsis is the generalized inflammatory response elicited by an infectious process withthe confirmed presence of bacteria or highly suspicious foci, is believed the main causesof death in Surgical Intensive Care Unit (ICU). It’s reported that approximately40%ofICU patients have sepsis, and septic shock has prohibitive mortality rate (36%). Up to now,the incidence of sepsis is rising, hospital costs are very expensive and effective treatmentmeans are very little. Therefore, to find a safety and effective treatment measure is urgent.It has been reported that hyperoxia and anesthetic dose of isoflurane are beneficial toanimal models of sepsis. Oxygen and isoflurane have been used widely in clinical practice.However, hyperoxia may aggravate tissue injury, resulting from overproduction ofreactive oxygen species and nitrogen species, and anesthetic dose of isoflurane is limitedto be applied to critically ill patients who may not tolerate the effects of anesthesia. Wehave demonstrated that combined inhalation isoflurane at subanesthetic dose (0.5MAC)and60%oxygen is beneficial to zymosan-induced generalized inflammation. Here, weproved the beneficial effects of0.5MAC isoflurane in60%oxygen in CLP-induced invivo murine model of sepsis and LPS-induced in vitro cell model of sepsis, respectively. We also demonstrated that the immuno-inflammatory mechanisms contributed to thisprotective action. We further illustrated that some miRNAs participated in theimmuno-inflammatory mechanisms underlying this protective effects using miRNAmicroarray studies.Materials and methodsWe used male Imprinting Control Region/Kunming mice (ICR/Km) weighing25to30grams for the study. Animals were randomly assigned to the following groups:CLP+Air, CLP+100%Oxy, CLP+0.5MAC ISO+60%Oxy, Sham+Air, Sham+100%Oxyand Sham+0.5MAC ISO+60%Oxy groups. In vivo, sepsis model was induced by cecalligation and puncture (CLP), and animals in sham groups were given an operation withoutCLP procedure. Animals were treated by exposure to100%oxygen or by0.5MACisoflurane in60%oxygen for1hour starting at1and6hours after CLP/Sham,respectively. Survival was recorded for7days after CLP/Sham in mice. Blood sample,organs and bronchoalveolar lavage fluid (BALF) were harvested for determining thehistopathologic observation, serum levels of biochemical parameters, lactate, C reactiveprotein (CRP), Wet-to-dry weight ratio of lung, total protein of BALF and seruminflammatory cytokines. For miRNAs assays, blood samples were harvested fordetermining inflammatory cytokines and miRNA expression profiling simultaneously.We used RAW264.7cell lines for in vitro sepsis study. Cells were randomly assignedto the following groups: LPS, LPS+100%Oxy, LPS+0.5MAC ISO+60%Oxy, Vehicle,Vehicle+100%Oxy and Vehicle+0.5MAC ISO+60%Oxy groups. Sepsis was induced byLPS stimulation, and DMEM as the control. Cells were exposured to100%oxygen or0.5MAC isoflurane in60%oxygen at the start of LPS stimulation. Then, cell culturesupernatants, nuclear proteins and inoculated cell slides were harvested for determininginflammatory cytokines and activation of NF-κB.ResultsWe observed that CLP induced abnormal changes in organ histopathology,biochemical parameters, lactate, CRP and inflammatory cytokines in serum, as well as the lower survival rate in CLP-challenged mice, which were reversed by100%oxygen or byinhaling0.5MAC isoflurane in60%oxygen. We observed that inhalation of100%oxygenor0.5MAC isoflurane in60%oxygen had significant anti-inflammatory effects at9and24hours after CLP/Sham, and at these two timepoints, changes in the expression profilingof miRNAs were detected using miRNAs assay. We also showed that LPS induced theincrease of inflammatory cytokines in cell supernatants (ELISA) and the activation ofNF-κB (western blot and confocal microscopy), which were prevented by100%oxygen orby0.5MAC isoflurane in60%oxygen.ConclusionInhalation of0.5MAC isoflurane in60%oxygen may protect against sepsis inmurine sepsis model induced by CLP and in cell sepsis model induced by LPS. AndmiRNAs modulated immuno-inflammatory pathways may play an important role in theprotective effects of inhalation of0.5MAC isoflurane in60%oxygen on sepsis. Theseresults demonstrated the potential beneficial effects of subanesthetic dose of isoflurane in60%oxygen in the patients with sepsis.