| As the largest immune organ of the body,the spleen plays an important role in the process of immune defense.The changes of its structure and function directly affect the immune function of the body.Studies have shown that obesity can not only cause changes in tissue structure such as increased spleen mass and red pulp area,but also increase the number of spleen lymphocytes and M2 macrophages.Other studies have found that the number of lymphocytes in alveolar lavage fluid of obese mice increased significantly after intranasal drip of E.coli.Whether this is related to the effect of obesity on the structure and function of the spleen remains to be studied.Peroxidation injury and inflammation are important mechanisms of pathological damage of tissues and organs,and also important factors of apoptosis.Therefore,this study investigated the effects of obesity on spleen tissue damage,antioxidant function,cytokines and cell apoptosis after intranasal drip of Escherichia coli(E.coli),which could provide reference for further elucidating the role of spleen in obesity against infection.Experiment 1 Effect on histomorphology of spleen in obese mice after E.coli nasal drops64 Kunming normal mice(80+1 day,35-38 g)were randomly divided into N-I,N-II.64 Kunming obese mice(80+1 day,46-50 g)were randomly divided into DIO-I,DIO-II.N-II and DIO-II intranasally infused with 40?L fluid containing 4×10~9 colony-forming unit(CFU)E.coli,N-I and DIO-I were given the equal volume of phosphate buffered saline.8 mice in each group were selected to collect spleen at 0 h,12 h,24 h and 72 h.The spleen weight and index were calculated,and the histological structure of spleen was also observed.The results showed that:(1)At 0 h,the spleen weight and index in DIO-I was lower than N-I(P<0.05).At 12 h,24 h and 72 h,the spleen weight and index in N-II was higher than N-I(P<0.05);at 12 h,24 h,and 72 h,the spleen weight and index in DIO-II was higher than DIO-I(P<0.05).(2)At 0 h,the demarcation of red pulp and white pulp in the spleen of N-II and DIO-II mice was clear,and there was no significant difference.At12 h and 24 h,the arrangement of white pulp lymphocytes in the spleen of N-II mice was looser,red pulp hyperemia was serious and polynuclear giant cells were increased;the arrangement of white pulp lymphocytes in DIO-II mice was looser,and red pulp hyperemia was observed.At 12 h and 24 h,compared with N-II,DIO-II exhibited more collagen fiber in spleen.The results showed that:(1)Obesity increased both body weight and spleen weight,but the increase of spleen weight was less than body weight.(2)Obese mice can alleviate the early congestion of spleen caused by intranasal drip of E.coli,and then promote the repair of damaged spleen in later period,which has protective effect on spleen.Experiment 2 Effect on oxidative stress and inflammation of spleen in obese mice after E.coli nasal dropsReferring to the grouping and treatment of experiment 1,another mice were selected for the experiment.8 mice in each group were selected to collect spleen at 0 h,12 h,24 h and 72 h.The MDA content,antioxidant enzyme activities,cytokines concentration were detected in spleen.The results showed that:(1)At 0 h,12 h,24 h and 72 h,the activities of GSH-Px,CAT,SOD in DIO-I was lower than N-I,and the content of MDA in DIO-I was higher than N-I(P<0.05).At 12 h and 24 h,the CAT,SOD activities in N-II was lower than N-I,and MDA content in N-II was higher than N-I(P<0.05);conversely,the GSH-Px,SOD activities in DIO-II was higher than DIO-I,and MDA content in DIO-II was lower than DIO-I(P<0.05).(2)At 0 h,12 h,24 h and 72 h,the Resistin,IL-6,IL-1?,TNF-?concentrations in DIO-I was higher than N-I,and Leptin concentration in DIO-I was lower than N-I.At 12 h and 24 h,the concentrations of Resistin,TNF-?,IL-6 and IL-1?in N-II was higher than N-I;DIO-II showed lower TNF-?,IL-6 and IL-1?concentrations compared to those of DIO-I at 24 h,although the concentrations of Leptin,Resistin,TNF-?,IL-6 and IL-1?were still observed to be higher than DIO-I at 12 h.The results showed that:(1)The antioxidant enzymes activity of the spleen of obese mice were lower,and the content of MDA was higher,showing relatively weak antioxidant capacity.After intranasal drip of E.coli,the spleen of obese mice showed stronger antioxidant capacity,thereby had a protective effect on oxygen free radicals caused the peroxidation damage.(2)The spleen of obese mice has more cytokines.After intranasal drip of E.coli,the splenic cytokines in obese mice increased temporarily and then decreased significantly,which was beneficial to alleviate the inflammatory immune damage of the spleen and promote the tissue repair;while the splenic cytokines in normal mice increased significantly and maintained for a certain period of time,which would aggravate the inflammatory immune damage of the spleen and then aggravate the inflammatory reaction.Experiment 3 Effect on apoptosis of spleen cells(SC)in obese mice after E.coli nasal drops128 Kunming normal mice(80+1 day,35-38 g)were randomly divided into N-I,N-II group.128 Kunming obese mice(80+1 day,46-50 g)were randomly divided into DIO-I,DIO-II.Referring to the treatment of experiment 1.8 mice in each group were selected to collect spleen at 0,12,24,and 72 h to detecte the SC apoptosis rate.In addition,8 mice in each group were selected to collect spleen to detecte the expressions of Bcl-2,Bax,Caspase-3 and Caspase-9 genes and proteins by qRT-PCR and Western blot.The results showed that:(1)At 0,12,24,and 72 h,the rate of SC apoptosis in DIO-I was higher than N-I(P<0.05);at 12 h and 24 h,its in N-II was higher than N-I;at 12 h,24 h,the rates of SC apoptosis in DIO-II were lower than DIO-I and N-I(P<0.05).(2)At 0,12,24 and 72 h,the expressions of Bax,Caspase-3,Caspase-9 in spleen of DIO-I were significantly higher than N-I(P<0.05).and Bcl-2/Bax ratio were significantly lower than N-I(P<0.05).At 12 h,the spleen Bax,Caspase-3,Caspase-9 gene and protein expression,Bcl-2/Bax ratio of N-II were significantly higher than N-I(P<0.05),while the spleen Bax,Caspase-3,Caspase-9gene and protein expression,Bcl-2/Bax ratio of DIO-II were significantly lower than DIO-I,and the expression of Bcl-2 was significantly higher than DIO-I(P<0.05).The results showed that:(1)SC apoptotic rate in obese mice was higher than normal mice,however,the SC apoptotic rate in normal mice was elevated,while SC apoptotic rate in obese mice was declined after intranasal drip of E.coli.(2)After intranasal administration of E.coli,the Bcl-2 in obese mice was significantly up-regulated,while the expression of Bax,Caspase-3 and Caspase-9 was significantly down-regulated,suggesting that obesity has a stronger ability to regulate apoptotic molecules in the spleen,which is more conducive to protecting the spleen and improving the immune status of the body.Conclusions:1.Obesity not only increases the body weight,but also increases the spleen weight,but the increase of spleen quality is less than that of body weight.Obesity can alleviate the spleen congestion in the early period after E.coli infection,and promote the repair of the damaged spleen in the later period,which has a protective effect on the spleen.2.The antioxidant enzymes activities in spleen of obese mice were lower,and the MDA content was higher,which showed that the antioxidant capacity was relatively weak.After intranasal drip of E.coli,the spleen of obese mice exhibited stronger antioxidant capacity,thus protecting the spleen from oxidative damage caused by oxygen free radicals.3.The spleen of obese mice has more cytokines.After intranasal drip of E.coli,only the inflammatory cytokines in the spleen of obese mice increased temporarily and then decreased significantly,which was beneficial to alleviate the inflammatory immune damage of the spleen caused by infection.4.The SC apoptotic rate in obese mice was higher.After intranasal drip of E.coli,the SC apoptotic rate in obese mice decreased.Obesity has stronger regulation ability on anti-apoptotic Bcl-2 and pro-apoptotic Bax,Caspase-3 and Caspase-9,which is more conducive to protecting the spleen and improving the immune defense ability of the body. |
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