The Protective Effects Of Dexmedetomidine On Liver In Polymicrobial-septic Rats

Dexmedetomidine（Dex）,one selective α2 adrenoceptor agonist,is widely used in the ICU with charateristics of sedative,analgesic,sympatholytic,while without suppression of breath.Early researches have found that:Dex could improve the survival rate of both septic rats and humans and protect organs through antiapotosis and anti-inflammation.However,still no inconclusive results implies whether Dex could protect liver and how it protect liver from sepsis.Morever,large amounts of experimental and clinical findings show that the status of liver could directly influence the survival and progonosis of sepsis,while lots of hepatocytic deaths characterised as necrosis during sepsis is the main factor of liver injury.And,one newly found regulative necrosis-necroptosis has been established to be involved in numerous inflammatory diseases,and interestingly,one study reports that specific knockout of RIP3-one essential protein in necroptosis pathway-could ameliorate infiltrations of inflammatory cells into liver so as to protect liver from sepsis.Therefore,the present study is aimed to explore whether Dex could protect liver from sepsis and be in inhibiting necroptosis.Part 1:the effects of different dosages of Dex on liver in septic ratsPurpose:Through observing the effects of different dosages of Dex on liver in septic rats,exploring the optimal dosage and mechanism of Dex.Methods:After catheterization of caudal vein,80 healthy SPF SD rats,weighing200²20g,aging 6⁸ weeks were randomized into 5 groups（n=16）:sham group（group Sh）,septic group（group S）,high dosage of Dex group（group H）,medium dosage of Dex group（group M）and low dosage of Dex group（group L）.10 rats of each group were randomly chosen to observe the 24 h survival rate.CLP were used to induce sepsis in group S,H,M and L.For rats in group Sh,the cecum was exposed without ligation or puncture.5.0、2.5 and 1.0 μg/kg/h Dex were continuously infused at 1h before CLP in group H,M and L,and normal volume of NS were infused into rats from group Sh and S.Both arterial and venous blood were taken at 6,12 and 24 h after operation aimed to test ABG and concentrations of AST and ALT in serum.All rats were sacrificed to remove liver,then HE and TUNEL staining were used to observe morphological changes and cell death in liver,besides these,concentrations of IL-6and TNF-αwere also detected.Results:Compared with group Sh,for all rats undergoing CLP,the levels of liver enzymes、lactate 、IL-6 and TNF-αwere increased（all P<0.05）,while PaO₂ were decrease（all P<0.05）with more obvious pathological changes and cell death in liver.When being compared to group S,pretreatment with 5μg/kg/h Dex could decrease levels of liver enzymes、lactate、IL-6 and TNF-α,meanwhile improved pathological changes and cell deaths in liver.Conclusion:Different dosages of Dex could implicate various effects on liver,both2.5μg/kg/h and 5.0 μg/kg/h Dex could protect liver from sepsis via anti-inflammation and anti-apoptosis,while 5μg/kg/h gets more beneficial effects.Part 2:The types of hepatocellular death during sepsisPurpose:This part of study was designed to explore whether necroptosis was involved in hepatocellular deaht during sepsis through detecting changes of proteins comprised of necroptosis signaling.Methods:After catheterization of caudal vein,24 healthy SPF SD rats,weighing200²20g,aging 6⁸ weeks were randomized into 4 groups（n=6）:sham group（group Sh）,sepsis group（group S）,dissolvant DMSO group（group D）and Nec-1 group（group N）.Rats in group Sh received same procedure as group Sh in part 1.1.0mg/kg Nec-1or DMSO in same volume were infused for rats from group N and D;same volume of NS was used in group Sh and S.All rats were sacrificed at 6h after operation,and then expressions of RIP1、RIP3 and MLKL were detected through western blot,and levels of ROS were tested in chemiluminescence.Results:Compared with group Sh,level of ROS and expressions of RIP1、RIP3 and MLKL were up-regulated in group S,N and D（all P<0.05）;compared with group S,these indexes were decreased（all P<0.05）;no differences were found between group Sh and N for all these markers（all P>0.05）.Conclusion:Necroptosis is involved in hepacellular death in septic rats.Part 3:Dexmedetomidine could protect liver from sepsis via inhibiting necroptosisPurpose:Through observing whether Dex could cause changes in expressions of proteins related to necroptosis signaling,probing whether the protective effect of Dex on liver during sepsis is related to necroptosis inhibition.Method:This part of study also contained three steps:（1）32 SD rats were randomized into 3 groups（n=8）:sham group（group Sh）,sepsis group（group S）and5μg/kg/h Dex group（group D）（2）40 SD rats were divided into 5 parts（n=8）:sham group（group Sh）,sepsis group（group S）,1.0mg/kg Nec-1 group（group N1）,0.5mg/kg Nec-1 group（group N2）,0.25mg/kg Nec-1 group（group N3）;（3）24 rats were grouped into 3 groups（n=8）:sham group（group Sh）,sepsis group（group S）and Dex combined with Nec-1 group（group ND）.Results:（1）compared with group Sh,RIP1、RIP3、MLKL、HMGB1 and ROS were all up-regulated（all P<0.05）;Dex could down-regulated expressions of all four proteins and the level of ROS（all P<0.05）;（2）4 proteins in other 4 groups were increased when being compared to group Sh,while compared to group S,there were no differences found in rats from group N3;（3）compared to group Sh,expressions of all proteins were increased in group S and ND;however,rats receiving Dex and Nec-1expressed less RIP、RIP3、MLKL and HMGB1.Conclusion:Dexmedetomidine could protect liver from sepsis via inhibiting necroptosis of hepatocytes.