Impact And Mechanism Of Continuous Veno-venous Hemofiltration In Different Ultrafiltration Rate On Inflammatory Status And Immune Function Of Porcine Endotoxemic Shock

Objective1.To study the impact of continuous veno-venous hemofiltration(CVVH) in different ultrafiltmtion rate on survival time,hemodynamics,arterial oxygenation and inflammation in porcine endotoxemic shock;To investigate the mechanism, including the impact of CVVH and high volume hemofiltration(HVHF) on plasma cytokines,monocyte function,neutrophil apoptosis and nuclear factor-κB (NF-κB).2.To study the adequate dose of CVVH in sepsis.Methods1.Animal preparation and hemofiltration treatmentEighteen anesthetized mechanically ventilated pigs weighing(28.1±5.1) kg received a 15.7μg/kg endotoxin(E.coli O111:B4) infusion over 60 min.When mean arterial pressure(MAP) dropped 30%,they were randomized into three groups. Control group(n = 6) received no further intervention,CVVH group(n = 6) and HVHF group(n = 6) received pre-dilution continuous veno-venous hemofiltration for 24 hours with an ultrafiltration rate of 55 ml/kg·h and an ultrafiltration rate of 100 ml/kg·h respectively,the blood flow was 150 ml/min.A 1.2-m² polyacrylonitrile membrane(AN69) and a Baxter BM 25 hemofiltration device were used.2.Measurements1) Monitoring heart rate(HR),mean arterial pressure(MAP),body temperature,pulse, Mean Pulmonary Arterial Pressure(mPAP),Pulmonary Artery Wedge Pressure (PAWP),Central Venous Pressure(CVP),Cardial Output(CO),Cardial Index(CI), Stroke Volume(SV),Stroke Index(SI),Systemic Vascular Resistance Index(SVRI), Pulmonary Vascular Resistance Index(PVRI) and Left Ventricular Stroke Work Index (LVSWI). 2) Monitoring PaO₂,SaO₂,FiO₂,oxygenation index(OI= PaO₂/FiO₂),BE,PaCO₂ and HCO₃^-.3) Monitoring survival time and urine output.Renal function and the complete blood count were measured before endotoxin infusion(Baseline) and at 0h(T0),6h(T6), 12h(T12),24h(T24) after the establishment of endotoxic shock model.4) Blood were taken before endotoxin infusion(Baseline) and at 0h(T0),1h(T1), 6h(T6),12h(T12),24h(T24) after the establishment of endotoxic shock model.The plasma levels of TNF-α,IL-6,IL-10 and IL-18 were tested by ELISA.5) At T0,T6,T12,T24 during CVVH,peripheral monocytes were isolated and stimulated with LPS(10μg/ml )to detect the ability of production of TNF-α.MHCâ…¡expression on rnonocytes was assessed to evaluate the antigen presenting function of monocytes.Apoptotic changes in PMNs were measured by flow cytometry.6) Wedge-shaped sections from kidneys and lungs were stained with H&E.The expressions of intercellular adhesion molecule-1(ICAM-1) and p65 were assessed by immunohistochemistry.Moreover,NF-κB/DNA binding activity was analyzed by ELISA.Results1.Part OneThe natural course of endotoxemic shock in our study was characterized by increases of HR,mPAP,PAWP,SVRI and PVRI,and decreases of MAP,SVRI,SV,SI, LVSWI,OI and BE.LVSWI and OI were improved significantly in HVHF group and CVVH group at T24(P＜0.05 ).No significant differences between HVHF group and CVVH group were found.2.Part TwoThe survival time in control group was(15.4±5.2) h,CVVH group was(21.4±4.1) h,HVHF group was(22±6.7) h.The survival time in CVVH and HVHF group were significantly longer than that of control group(P＜0.05 ).The urine output in CVVH and HVHF group was obviously higher than that in control group at T18 and T24(P＜0.05).Platelet decreased progressively after the infusion of LPS. 3.Part ThreePlasma TNF-αand IL-6 peaked at T1,while IL-10 peaked at T0,then they declined gradually.While IL-18 increased and did not decline after T0.A significant decrease of plasma IL-10 levels was observed at T6,T12 and T24 in CVVH group compared with control groups(P＜0.05).HVHF group accomplished a greater decrease in plasma TNF-α(T6) and IL-10(T6,T12,T24) levels compared with control group and CVVH group(P＜0.05 ).The level of IL-6 and IL-18 showed no significant difference among three groups(P＞0.05 ).There was a significant negative correlation between IL-6 and survival time(P＜0.05).4.Part FourMHCâ…¡expression of monocyte and the level of TNF-αwere markedly decreased in porcine endotoxemic shock(P＜0.05).CVVH and HVHF had no effect on them.The infusion of endotoxin was followed by decrease ofPMN apoptosis. HVHF increased PMN apoptosis and decreased the number of peripheral PMN(P＜0.05).5.Part FiveCVVH and HVHF resulted in significantly lower scores for hyaline membranes, interstitial infiltration,edema and atelectasis in lung compared with control group(P＜0.05).CVVH and HVHF mitigated significantly severe tubular damage and glomerular damage compared with control group(P＜0.05).The expression of ICAM-1 and p65,the degree of NF-κB/DNA binding activity in lung and kidney were reduced in CVVH group and HVHF group(P＜0.05),there was no significant difference between them(P＞0.05 ).Conclusions1.HVHF and CVVH can improve blood pumping and arterial oxygenation of porcine endotoxemic shock in some degree.2.HVHF and CVVH can prolong survival time and improve urine output compared with control group.3.IL-10 can be removed effectively with CVVH and HVHF in porcine endotoxemic shock and HVHF can also remove TNF-αeffectively.IL-6 was a powerful independent predictive factor for survival time in porcine endotoxemic shock.4.HVHF can alleviate the decrease of PMN apoptosis which may result from removal of TNF-αand IL-10.CVVH and HVHF can not improve monocyte function.5.CVVH and HVHF can offer both renal and lung protection,which may be related to suppression of inflammation and NF-κB.