Low Molecular Weight Heparin Prevents CLP-induced Acute Lung Injury Of Septic Rats By Anti-inflammatory Coagulation Effect

[Objective]1.To find the possible mechanism of low molecular weight heparin protective effect on ALI of septic rats,this study investigated the effect of low molecular weight heparin on mRNA and protein expression of high mobility group box1(HMGB1) in lung tissue of septic rats which induced by cecal ligation puncture (CLP).2. Observe the low molecular weight heparin effect on plasma TNF-a and IL-6 of septic rats,discussing its influence on releasing of early and late mediators of inflammation during the sepsis.[Methods]144 sepcific-pathogen free(SPF) healthy male Sprague-Dawley(SD) rats were randomly divided into three groups as follow:Sham operation group(A),nomal treatment group (B),the LMWH treatment group(C),n=48.Group A received a sham operation and the other groups were underwent CLP operation.Groups A and B accepted intraperitoneal injection(i.p.)of normal saline(NS) at a dose of 2.0ml/kg and ceftriaxone(30mg/kg),Group C were intraperitoneal injection additional LMWH(150U/kg). Observe points were made at3,6,12,18,24,48h,the rats were anesthesized and killed, mortality,lungs wet/dry ratio and pathologic change were determined.HMGB-1 mRNA and protein of lung tissues were measured by RT-PCR and Western blot.TNF-a and IL-6 of blood plasma calculated by ELSIA.MPO activity in lung homogenate was detected according to the kit instruction.The right upper lobe were removed when they were killed,then weighted(wet weight)and dried in a 80℃oven until weight was constant(dry weight).The ratio of wet weight/dry weight of the lung was calculated.The effect of LMWH on the septic severity score,survival time,expression of HMGB1 mRNA.HMGB1,MPO activity.WW/DW ratio,TNF-αand IL-6 were observed.[Result]1.The septic severity score of Group A at defferent time had no statistics significant difference,the septic severity score of Group were obviousy higher than Group A,the different between each other had ststistics significance(P<0.01);the septic severity score reached to the peak during 12h-24h,and it was still higher than the 6h level.The septic severity score of Group C was remarkably lower than Group B.The half survival time of Group A,B,C were 120h,28h,20.4h respectively,and there were remarkably ststistics significance between Group A,B,C.2.Group A had very weakly expression of HMGB1 mRNA and HMGB1 on lung tissue at all time points,and no statistics significance were observed.The accrescence of HMGB1 mRNA and HMGB1 of Group B,C were detected since 6h,and reached the peak at time 12-24h,and the decrease were detected since 48h,but still higher than Group A even at time point of 48h.Group C was remarkably lower than Group B (P<0.05).3.Group A had very weakly expression TNF-a in plasma of at all time points,and no statistics significance were observed.The accrescence of TNF-a content in plasma of Group B,C reached the peak at time 6h, and the decrease were detected since 12h,and get the same level as 3h at time 24h.Group B,C level higher than Group A even at time point of 24h.Group C was remarkably lower than Group B (P<0.05). Group A had very weakly expression IL-6 in plasma of at all time points,and no statistics significance were observed. The accrescence of IL-6 content in plasma of Group B,C reached the peak at time 3h, and the decrease were detected since 18h,and get the same level as 3h at time 24h.Group B,C level higher than Group A even at time point of 24h.Group C was remarkably lower than Group B (P<0.05).4.The MPO activity of Group A had no statistics significance between each other,the activity of Group B,C begin to increase since 3h,and also reach the peak at point 24h,and the decrease could be detected since 48h.Group C are remarkably lower than Group B(P<0.05),there were dtill higher than the basic of Group A (P<0.05).5.The WW/DW ratio of Group A was remarkably lower than Group B,. C(P<0.05),and GroupC were remarkably lower than Group B (P<0.05).6.Through light microscope,there were obviously manifest of ALI on lung tissue of B Group,including:hyperemia,edma,congestion and inflammation.Group C had lightet lung injury and the lung injury score of them lower compared to Group B(P<0.05).[Conclusion]1.The LMWH can inhibit the release of early and late mediators of inflammation in the course of sepsis which induced by CLP.2.The LMWH can significantly inhibit the mRNA,protein expression of HMGB-1 on lung tissue;attenuates the degree of acute lung injury induced by CLP by down regulation of MPO activity and WW/DW ratio. LMWH may through inhibition of HMGB1 of lung tissue to lessen the PMN infiltration and improve the survive rate of septic rats by CLP.3.The protective effection of LMWH in acute lung injury induced by CLP may through anti-inflammatory influence on the axis of coagulation and inflammation.