The Effects Of Oxidative Stress On The Apoptosis Of Lymphocytes In Peyer's Patches After Burn And Delayed Fluid Resuscitation In Rats At High Altitude

Objective To evaluate the influence of oxidative stress on the apoptosis of lymphocytes in the Peyer's patches after severe burn with delayed fluid resuscitation in rats at high altitude,and the relation between the expression of apoptosis-related gene and oxidative stress in Peyer's patches.Methods A total 132 Wistar rats were applyed as objects of this study.They were randomly divided into 3 groups at different altitudes(1517m and 3848m) and employed as the experimental models(TBSA30%,Ⅲdegree),and then they were randomly divided into 3 subgroups at different altitudes(1517m and 3848m):delayed fluid resuscitation group(DFR),immediate fluid resuscitation group(IFR) and sham group(SG).The Peyer's patches were harvested from the ileum of rats at 6h,12h, 24h,48h and 72h after burn respectively.Large slice technique,TUNEL,TBA method, colorimetric method,immunohistochemistry and image analysis techniques were used to detect apoptosis and the expression of Bax,HIF-1αand CD3 in Peyer's patches, the content of MDA and TSH of Peyer's patches were also observed.Results 1.With the altitude increased,the follicular germinal center of Peyer's patches expanded and the inflammatory cell infiltration could be seen,clustered distribution of apoptotic cells in the follicular center,nuclear pyknosissed,chromatin mass assembled in the cells membrane;apoptotic bodies of various sizes were observed and cytoplasmic condensation,and were round or oval.2.The positive expression of Bax appeared in nucleus of lymphocytes in Peyer's patches.The expression level of Bax were much higher in DFR and IFR than those in SG,as with the increase in altitude Bax expression enhanced at 3848m altitude than those at 1517m,and in DFR than those in IFR,the experimental groups in 3848m higher respectively than those in 1517m at 6h and 72h after injured(P<0.05).3.The positive expression of HIF-1αappeared in nucleus of lymphocytes in Peyer's patches.The expression level of HIF-1αwere much higher in DFR and IFR than those in SG,as with the increase of altitude HIF-1αexpression enhanced at 3848m altitude than those at 1517m,and in DFR than those in IFR,the experimental groups in 3848m higher respectively than those in 1517m at 6h and 24h after injured(P<0.05). 4.The positive expression of TUNEL appeared in nucleus of lymphocytes in Peyer's patches.The expression level of TUNEL were much higher in DFR and IFR than those in SG,as with the increase in altitude TUNEL expression enhanced at 3848m altitude than those at 1517m,and in DFR than those in IFR,the experimental groups in 3848m higher respectively than those in 1517m at some time points(P<0.05).5.The positive expression of CD3 appeared in cytoplasmic and cell membrane of T lymphocytes in Peyer's patches.The CD3~+ expression level of T lymphocytes in the Peyer's patches were much lower in DFR and IFR than those in SG,as with the increase in altitude,CD3~+expression weakened at 3848m altitude than those at 1517m,and in DFR than those in IFR,the experimental groups in 3848m lower respectively than those in 1517m at 6h and 24h after injured(P<0.05).6.Content of MDA in the Peyer's patches were much higher in DFR and IFR than those in SG,as with the increase in altitude content of MDA enhanced at 3848m altitude than those at 1517m,and in DFR than those in IFR,the experimental groups in 3848m higher respectively than those in 1517m at 12h～72h after injured(P<0.05).7.Content of TSH in the Peyer's patches were much lower in DFR and IFR than those in SG,as with the increase in altitude content of TSH enhanced at 3848m altitude than those at 1517m,and in DFR than those in IFR,the experimental groups in 3848m lower respectively than those in 1517m at 12h～72h after injured(P<0.05).8.There were significant positive correlation between cell apoptosis and expression of HIF-1α, MDA(r=0.991,0.965,P<0.01) in DFR at 3848m.Conclusion1.With the altitude increased,there were obviously differences of pathological changes of Peyer's patches after severe scald with delayed fluid resuscitation,nuclear pyknosissed,chromatin mass assembled in the cells membrane;there are seen that visible apoptotic bodies of various sizes and cytoplasmic condensation,and were round or oval.2.The apoptotic rate of Peyer's patches increased after severe scald with delayed fluid resuscitation at high altitude.3.Bax,HIF-1αmay regulate the apoptosis of lymphocytes in the Peyer's patches after severe scald with delayed fluid resuscitation in rats at different altitude.4.There was a severe oxidative stress in Peyer's patches after severe scald with delayed fluid resuscitation in rats at high altitude,and may be responsible for the dramatic apoptosis in Peyer's patches.5.The decrease of CD3~+T lymphocytes may induce the intestinal cell immune depression after severe scald with delayed fluid resuscitation at high altitude.