Effect Of Parenteral Glutamine Supplement On Lymphoid Organs’s Apoptosis In Sepsis Rats

Objective:Sepsis is still one of the important causes of death in critically ill patients, the apoptosis of lymphocyte cell is one of the important pathophysiological mechanisms of sepsis. we investigated the effects of the apoptosis of lymphoid organs in sepsis rats which was received different doses of glutamine by parenteral administration.Methods:1 、 SD rats were randomly divided into five groups:The sham operation group(SHAM), Model group(CLP), Low doseglutamine group(L-GLN), Medium dose glutamine group(M-GLN), High dose glutamine group(H-GLN),established the rat sepsis model by cecal ligation and puncture in the group of CLP、L-GLN、M-GLN、H-GLN,rats of every group were respectively received the tail intravenous injection of saline、0.35g/kg glutamine、0.50g/kg glutamine、0.75g/kg glutamine after surgey immediately.At the same time, in order to eliminate the influence of the operation itself, rat tail vein injection of saline was served as sham operation group.2 、 Observe and record the performance of rats in every group after surgery,measured respectively rectal temperature, heart rate, breath of rats in every group in abdominal anesthesia after established models for 24 hours, plucked blood by common carotid artery and centrifuged,taked the serum for detection,at same time,plucked blood by common carotid artery from three rats of every group randomly for blood culture,and taked their spleen and thymus into 10% formalin fixed, paraffin embedded for immunohistochemistry.3 、 The cell apoptosis of spleen and thymus tissue was deteced by TUNEL,Immunohistochemistry was employed for detecting the expression of caspase3 、HSP90 protein in spleen and thymus tissue,the serum level of IgA、IgG、IgM and TNF-α、IL-10 in rats was detected by ELISA.Results:1、Comfirmed the rat sepsis model was successfully established:monitored therectal temperature, heart rate, breath of rats in every group in abdominal anesthesia,compared with SHAM,the rectal temperature of rats was reduced more than 1 ℃,the heart rate was increased more than 50%, respiratory rate was increased more than 2 times in the group of CLP、L-GLN 、M-GLN、H-GLN,corresponding with the diagnostic standard of SIRS animal model. By dissected abdominal cavity,we can see the performance of abdominal infection of primary lesion,such as a large number of bloody peritoneal exudate with stench 、 cecal ligation endsurface presented black and yellow pus.meanwhile,blood culture which were randomly selected in the CLP group and the different doses of glutamine intervention groups were all cultured in mixed bacteria,that could proved the infection,so we can think the rat sepsis model by cecal ligation and puncture was sucessfully establised.2、The result of cell apoptosis index in every group :the cell apoptosis was low in the spleen and thymus tissue of rat in SHAM group;compared with SHAM group,the cell apoptosis was obvious in the spleen and thymus tissue of rat in CLP group,and have a significance difference(p<0.01).the cell apoptosis was reduced gradually in the spleen and thymus tissue of rat in the group of L-GLN、M-GLN、H-GLN(p<0.05 or p<0.01),the anti- apoptosis effect of the group of high dose glutamine was better than the group of low dose glutamine.(p<0.05).3、The result of the expression of caspase3 protein in every group: the expression of caspase3 protein in the spleen and thymus tissue of rat in the group of SHAM was low,the point of expression of caspase3 protein in the group of CLP was significantly increased compared with SHAM group(p<0.01), the point of expression of caspase3 protein in the group of L-GLN 、 M-GLN 、 H-GLN was significantly decreased compared with CLP group(p<0.05 or p<0.01),the point of expression of caspase3 protein in the group of high dose glutamine was lower than the group of low dose glutamine(p<0.05).4、The result of the expression of Hsp90 protein in every group: the expression of Hsp90 protein in the spleen and thymus tissue of rat in the group of SHAM was low,the point of expression of Hsp90 protein in the group of CLP was significantly increased compared with SHAM group(p<0.01),the point of expression of Hsp90 protein in the group of L-GLN 、 M-GLN 、 H-GLN was significantly increased compared with CLP group(p<0.05 or p<0.01),the point of expression of Hsp90 protein in the group of high dose glutamine was higher than the group of low dose glutamine(p<0.05).5、The serum level of IgA、IgG、IgM in every group:compared with SHAM group, the serum level of IgA 、 IgG 、 IgM in the group of CLP were decreased significantly(p<0.01), the serum level of IgA、IgG、IgM in the group of L-GLN、M-GLN、H-GLN were higher than the group of CLP(p<0.05 or p<0.01), the serum level of IgA、IgG、IgM in the group of high dose glutamine was lower than the group of low dose glutamine(p<0.05).6、The serum level of TNF-a、IL-10 in every group:compared with SHAM group, the serum level of TNF-a 、 IL-10 in the group of CLP were increased significantly(p<0.01), the serum level of TNF-a、IL-10 in the group of L-GLN、M-GLN、H-GLN were lower than the group of CLP(p<0.05 or p<0.01), the serum level of TNF-a、IL-10 in the group of high dose glutamine was lower than the group of low dose glutamine(p<0.05).Conclusions:Parenteral glutamine supplement can inhibit lymphoid organs’ s apoptosis in sepsis rats,and the anti-apoptotic effects was positively correlated with glutamine dose. Glutamine probably achieve the anti-apoptotic effects by inducing the expression of Hsp90,inhibiting the activation of caspase-3.