The Effects Of Three Mechanical Ventilation Strategies On Inflammatory Factors And Type Ⅱ Alveolar Epithelium Cell Morphology In Newborn Piglets With Acute Lung Injury

ObjectiveMechanical ventilation is an effective therapeutic method for neonates with acute lung injury(ALI) and acute respiratory distress syndrome(ARDS). However, it may aggravate pre-existing lung injury or even cause lung injury, a phenomenon frequently referred to as ventilator-induced lung injury (VILI). Morbility of neonates with VILI is from 13% to 35%, and its mortality is as high as 31%. Because of high morbility of VILI in conventional mechanical ventilation(CMV)and to attenuate VILI, new mechanical ventilation technologies such as high frequency oscillatory ventilation(HFOV) and partial liquid ventilation(PLV) are being developed. At present, more adult animals rather than newborn animals were used as models to study VILI both at home and abroad, and it has been found that compared with CMV, both HFOV and PLV can attenuate VILI. Because the survival quantity and quality of AEC II plays a critical role in the recovery of neonates with VILI, It's being thought highly of the recovery of type II alveolar epithelium cell(AEC II) in VILI following further study in mechanisms of VILI. But there is few studies about CMV,HFOV and PLV on the VILI,especially newborn animal.This study was completely random design to investigate lung injury and the inflammatory response in the lung in newborn piglets treated 24 hours with either CMV,HFOV or PLV following induced acute lung injury. Bronchoalveolar lavage fluids(BALF) was analysed to detect the change of inflammatory factors and surfactant associated protein-A(SP-A).Fluorescence microscopy was used to quantitate AEC II damage,and transmission electron microscopy was used to qualitativly examine ultrastructural damage to AECⅡ. Methods1. Study Contents To investigate oxygenation index(OI),hemodynamics,inflammatory factors and SP-A in BALF,morphocytology of AEC II in newborn piglets treated 24 hours with either CMV,HFOV or PLV following induced acute lung injury.2.Study Methods2.1 Animal PreparationEighteen no more than 3-day-old newborn piglets were intubated after anesthesia,ventilated with a MAQUET Servo-i ventilaor,using pressure control ventilation mode.Femoral artery was cannulated for arterial blood pressure monitoring and blood sampling.After surgical procedure and a stabilization period of 30 minutes,the arterial blood gas,arterial blood pressure,heart rate(HR) were obtained,calculating oxygenation index(OI) and mean arterial blood pressure(MABP) as baseline parameters(baseline).2.2 Induction of Acute Lung InjuryAll the piglets ALI were induced using CMV mode.To induce acute lung injury,warmed normal saline(37)at a volume of 35 ml/kg was instilled continuously through a endotracheal tube,the lavage procedure was repeated after each 5 minutes inteval until the PaO2 remained below 100 mmHg under the fraction of inspired oxygen being 1.0 for one hour.2.3 Study GroupsAfter completion of lavage,piglets were randomly assigned to one of three study groups:control, CMV, HFOV, or PLV. Control group(n=3):piglets were killed after the completion of lavage series,no mechanical ventilation.CMV group(n=6):piglets were ventilated 24 hours using a MAQUET Servo-i ventilaor with pressure control ventilation mode. HFOV group(n=6):piglets were ventilated 24 hours using a SLE5000 ventilaor with HFOV mode.PLV group(n=3):for the animals randomized to PLV,warmed perfluorocarbon(37℃) was instilled into the lungs through a side port on the endotracheal tube connector in an amount 18 ml/kg,and instillation was supplemented hourly. Piglets were ventilated 24 hours using a MAQUET Servo-i ventilaor with pressure control ventilation mode 2.4 Observing Markers and Methods(1) Blood gas analysis,arterial blood pressure and heart rate were monitored at preinjury,ALI 0h,6h,12h,18h and 24h respectively.OI and MABP were calculated at each time point respectively.(2) Tumor necrosis factor-a(TNF-a),interleukin-8(IL-8),IL-1 and SP-A in BALF were analyzed using enzyme linked immunosorbent assay(ELISA).(3) Quantity,area and fluorescence concentration of AECⅡwas detected using fluorescence microscopy.(4) Ultrastructural damage to AECⅡwere observed using transmission electron microscopy.Results1.OI:No differences were found in OI at ALI Oh between groups(P>0.05).As the extension of ventilation time,0I was continuously decreased in three ventilated groups,however,from sixth hour to twenty-forth hour,OI in HFOV group and PLV group was significantly lower than that in CMV group(P<0.05).OI in PLV group was lower than that in HFOV group at each time point.OI in PLV group was significantly lower than that in HFOV group(P<0.05).2. HR,MABP:No differences were found in HR and MABP at each time point between groups(P>0.05).3. Inflammatory factors and SP-A in BALF:There were significant differences between each groups in TNF-α,IL-8,IL-1 and SP-A in BALF respectively(P<0.05). Multiple comparisons:IL-8,TNF-α,in HFOV were lower than those in CMV group(P <0.05),SP-A was higher than that in CMV(P<0.05). IL-8,IL-1,TNF-αwere lower than those in CMV(P<0.05),SP-A was higher than that in CMV(P<0.05). IL-8,TNF-αin PLV were lower than those in HFOV(P<0.05).8.4. AECⅡquantity:There were significant differences between each groups in the population mean of AECⅡquantity respectively(P<0.05). Multiple comparisons:AECⅡquantity in HFOV group and CMV group were significantly different from that in control group(P<0.05). AECⅡquantity in CMV group and HFOV group were significantly larger from that in PLV group(P<0.05).5.AEC II area:There were significant differences between each groups in the population mean of AECⅡarea respectively(P<0.05). Multiple comparisons:AEC II area in HFOV group and CMV group were significantly different from that in control group(P<0.05).9.6. AEC II immunofluorescent density:There were significant differences between each groups in the AECⅡimmunofluorescent density respeciively(P< 0.05).Multiple comparisons:AEC II immunofluorescent density was significantly different in control group from thant in CMV group. AEC II immunofluorescent density in HFOV gropu and PLV group were significantly different from CMV group.7. Ultrastructure of AECⅡ:Control group:The juxtaposition of AEC II to basal membrane and AEC I was tight. In AECⅡ, cellular nucleus and its boundary were close to normal, chromatin was homogeneous, electron density of some osmiophilic multilamellar body(LB) was decreased and vacuolized, but well-distributed electron density and parallel arranged lamellar structure were also found.Microvilli on AECⅡsurface could be seen clearly.CMV group:The juxtaposition of AECⅡto basal membrane and AEC I was partial dissociated, but not completely dislodged. In AECⅡ, some cellular nuclei became shrank, chromatin was inhomogeneous and disseminated around verge, nucleoli were vagued, LBs were arranged around cellular nucleus, its electron density and numbers were decreased, lamellar structure was destroyed. Microvilli on AECⅡsurface were reduced obviously.HFOV group:The juxtaposition of AECⅡto basal membrane and AEC I was close. In AECⅡ, the shape of cellular nucleus was regular, chromatin was disseminated around verge, nucleoli could be seen, LBs were arranged around cellular nucleus, more LBs could be seen in HFOV group than those in CMV group, incompletely vacuole-like deformity and lamellar structure of LBs could also be seen. Microvilli on AEC II surface were decreased.PLV group:Cellular nuclei of AECⅡs in PLV group were larger than those in CMV and HFOV group. In AECⅡ, chromatin was homogeneous; LBs were presented normal electron density, increased in number, less vacuole-like deformity and parallel arranged lamellar structure. Microvilli on AECⅡsurface could be seen clearly. Beside these, in HFOV group, dissociated and dislodged AECⅡ. ConclusionsOI:OI in PLV group at sixth hour and eighteenth hour point was improved more significanttly than that in HFOV group.OI in PLV group at each time point was better than that in CMV group.OI in HFOV group at each time point was better than than in CMV group. We can conclude that optimal oxygenation is in PLV group,and then is in HFOV group, after newborn piglet was ventilated 24 hours,Inflammatory factors in BALF:In three ventilated groups, IL-8,TNF-αin PLV gropu were lower than those in CMV group and HFOV group,IL-1 in PLV was lower than that in CMV group. IL-8,TNF-αin HFOV group were lower than those in CMV group. SP-A in PLV group and HFOV group were more than those in CMV group,which indicates inflammation were less obvious in PLV from HFOV and CMV,, after newborn piglet was ventilated 24 hours,Morphology From AECⅡquantity, AECⅡarea, AECⅡimmunofluorescent density and ultramicrostructural damage, the study show AECⅡinjury were less obvious in PLV from HFOV and CMV.ZHAO Youwei(Pediatrics)Directed by professor FU Wanhai...