Analysis Of Differentially Expressed Genes And Study Of The Expression Of EPHX2 In Rat Liver Following Hemorrhagic Shock

Background And ObjectiveDifferent rats have distinct ability of anti-shock. Even with the same race,age,sex and weight, few rats can survived from severe trauma hemorrhagic shock. The gene background of the survived need to be uncovered and the function of potential key gene need to be investigated in the pathology of hemorrhagic shock.MethodsProtocol A: In a controlled 25ml/Kg total body weight(TBW) fixed-volume hemorrhage and left lobular hepatectomy rat model , differential genes were screened from the liver of rats which survived over 24h( SH group) and died within 1h (control group) by illuminaTM chips for rat cDNA microarray(27,342 genes, >700,000 probes). Quantitative real-time PCR were used to confirm the differential expression genes identified by microarrays. Regulation network of differential genes were further queried using KEGG.Protocol B: In a fixed-pressure hemorrhage rat model, rats were randomly divided into control group,HS group ( hemorrhagic shock no resuscitation),HSR1( 1h after hemorrhagic shock and resuscitation),HSR3( 3h after hemorrhagic shock and resuscitation),HSR6 ( 6h after hemorrhagic shock and resuscitation) group. We made the MAP down to 30±5mmHg sustaining 1h, and the HSR group infusion the 1/2 all blood and 3/2 lactated ringer solution within 20 min. The Expression of EPHX2 and YY1 mRNA and protein was detected in the liver of rats.ResultsProtocol A: Rats with 2.5ml/100g total body weight(TBW) fixed-volume hemorrhage at the rate of 2.0ml.Kg-1.min-1 with left lateral lobular hepatectomy have about 50% survival rate. 100 different genes of 21,793 assessed genes were identified by microarrays in the liver of rats between the survived and the dead(fold>2), among which 47 genes were up-regulated and 53 genes were down-regulated. Of these genes ,the signaling pathways linked to circadian rhythm,beta-Alanine metabolism,Histidine metabolism,Biosynthesis of unsaturated fatty acids,Glycine, serine and threonine metabolism,Vitamin B6 metabolism,Arginine and proline metabolism were identified. Real-time polymerase chain reaction confirmed the differential expression for Aldh1a7,Aoc3,Cyp26al,HDC and EPHX2 genes. Of these genes, EPHX2 is lower expression in the survival rats livers contrast to the dead, and we chose it for the potential key gene to the deep research. Protocol B: EPHX2 was notably increased after hemorrhage with MAP 30±5mmHg sustaining 1h contrast to the control group(P<0.05). After resuscitation, EPHX2 became lower by degrees in turns of HSR1,HSR3 and HSR6 group wherever on mRNA level or on western level(P<0.05). The nuclear factor YY1 have an opposite change tendency contrast to EPHX2 in the liver of the rats, which suggest that YY1 may have a negative regulation to EPHX2.ConclusionOur study provided a unique insight into the gene changes contributed for the survival of rats suffered severe fixed volume hemorrhagic shock and left lobular hepatectomy. We considered that EPHX2 may be one of the key genes and over-expression of EPHX2 screened from the rats survived severe trauma hemorrhagic shock may contributor to the apoptosis of liver cells in the rats. The nuclear factor YY1 have a negatively control to EPHX2, and to increase YY1 may contributor to reduce the apoptosis of liver cells of the rats.