| The harm of heavy metals such as nickel(Ni)is the dangers of the most prominent in the development of human society,Ni exposure can cause people and livestock poultry kinds of many cardiovascular diseases,including arrhythmia,atherosclerosis,coronary heart disease,cardiac developmental delay and deformity,myocardial hypertrophy and degeneration and focal aneurysms,but the specific mechanism is unclear.Thioredoxin reductase 3(Txnrd 3)as a member of the family of selenoprotein,in cell proliferation,apoptosis,antioxidation,antitumor activity and fine mature plays a key role.Our initial findings suggest that selenium deficiency changed obviously make the heart injury,but also accompanied Txnrd 3 expression significantly reduced,so we guess Txnrd 3 and there may be some relationship between heart damage,but the specific mechanism is unclear.Melatonin(Mel)is an antioxidant,can protect a variety of tissues and organs from damage,we assume Mel may also be protective in the heart damage.One hundred and sixty 8-week-old C57 BL/6N male mice weighing 25-30 g,wild type and homozygous for Txnrd 3+/+ and Txnrd 3-/-,randomly divided into 8 groups,Wild-Control,Wild-Ni,Wild-Mel,Wild-Ni+Mel,Homozygote-Control,Homozygote-Ni,Homozygote-Mel,and Homozygote-Ni+Mel.The mice in control group were given distilled water;Ni-treated group were given freshly prepared10 mg/kg Ni Cl2 solution respectively;The Mel group was administered intragastrically at a concentration of 2 mg/kg at a dose of 0.1 m L/10 g body weight for 21 days.Histopathology and ultrastructure of heart tissue were observed,and the indexes related to oxidative stress were detected by the kit(including GSSG and MDA content,CAT and T-SOD activity),and m RNA of mouse heart was detected by RT-PCR and Western blot(including mitochondrial apoptosis-related genes Caspase3,Caspase 9,BAX,cyt.c,BCL 2,and p53,autophagy-related genes LC 3,ATG 1,ATG 7,p62,m TOR,and Beclin-1,and inflammation-related genes TNF-α,COX2,IL-1β,IL-2,IL-6,and IL-7)and proteins(including mitochondrial apoptosis-related genes Caspase3,Caspase 9,BAX,cyt.c,BCL 2 and p53,autophagy-related genes ATG 7,Beclin 1 and p62,and inflammation-related genes COX2,IL-1β,IL-2 and TNF-α).TUNEL was used to detect the occurrence of apoptosis in cardiomyocytes.The main results are as follows:1)Observe clinical manifestations in mice found that Wild-Control group,the Homozygote-Control group,the Wild-Mel group,the Homozygote-Mel group of mice were hair smooth and close to the body,in good health in mice,the change of state were not obvious.WildNi group mice dirty coat dry,angular,depressed,breathing disorders,crouching,unity state variation compared with control group.Wild-Ni+Mel group of mice were had fluffy and dull hairare,slow growth,weight decreased activity,breathing after activity,compared with the Wild-Ni group status getting better.Homozygote-Ni group hypothermia in mice,sparse coat,messy,severe wasting,spirit heavy depression,tremors,difficulty breathing,action were difficult,stimulate the reaction were small,a few late death in mice,and the Wild-state level than the Control group and Wild-Ni group mice.Homozygote-Ni+Mel group of mice were messy,loss of appetite,weight loss,heavy sleep,shortness of breath,slow,compared with the Wild-Ni+Mel group status were poor,compared with the Homozygote-Ni group status is good.2)Observed the pathological morphology and ultrastructure of mice heart tissue found that Wild-Control group mice were normal myocardial tissue form,complete structure is clear.Wild-Ni group mice myocardial fibers arranged loosely,disorder,with congestion and inflammatory cells infiltration;Homozygote-Ni group rats were further dissolve myocardial fibers,accompanied by a large number of inflammatory cells infiltration.Compared with Wild-Control group,Wild-Ni and Homozygote-Ni group rats were damage is aggravating,which suggests that Ni exposure may induce the small rat heart damage;Compared with the Wild-Ni group,Homozygote-Ni group damage is aggravating,and compared with the Wild-Ni+Mel group,Homozygote-Ni+Mel damage is aggravating,suggesting Txnrd 3 expression reduce increased heart damage induced by Ni,show the inhibits Mel to Ni exposure of protection.Compared with the Wild-Ni group,Wild-Ni+Mel damage loss,and compared with Homozygote-Ni group,the Homozygote-Ni+Mel group damage,suggesting that Mel could reduce the heart dirty Ni induced damage.In addition,the Wild-Control group mice myocardial cell morphology was normal,not seen ultrastructure changes.Small Wild-Ni group rats were chromatin edge set,mitochondrial damage,cytoplasm vacuoles and observed a small amount of autophagosome,with the Wild-Control group than the injury worse.Homozygote-Ni group found in the ultrastructural features of typical apoptosis and autophagy,mitochondria damage is serious,also found the autophagy-lysosome,compared with the Wild-Control and Wild-Ni group injury aggravated.This suggests Txnrd 3 expression down low can promote the mice induced by Ni myocardial cell apoptosis and mitochondria autophagy(mainly)mitochondrial apoptosis.3)Observe mice heart tissue oxidative stress related indicators found that compared with Wild-Control group,the Homozygote-Control groups of mice GSSG,T-SOD activity and MDA,CAT content were no significant change(P > 0.05),and compared with wild-Mel pretreatment group,Homozygote-Mel group no significant change in the index of the mice(P > 0.05),this suggests that a simple Txnrd 3 expression of cardiac antioxidant levels is not enough to induce mice.Compared with Wild-Control group,Wild-Ni group rats were GSSG and MDA content increased significantly,the CAT and T-SOD activity significantly lower(P < 0.05),and compared with Homozygote-Control group,the Homozygote-Ni group to the above indicators show the change of mice(P < 0.05),suggesting that Ni exposure can induce mice cardiac oxidative stress.Compared with the Wild-Control group,the index of the Wild-Mel group rats were no significant change(P > 0.05),and compared with Homozygote-Control group,the Homozygote-Mel group no significant change in the index of the mice(P > 0.05),suggesting that Mel dose in this experiment,right does not cause mice oxidative damage of the heart.Compared with the Wild-Ni group,Wild-Ni+Mel group rats were GSSG and MDA content decreased significantly,the CAT and T-SOD activity was significantly increased(P < 0.05),and Homozygote-Ni group than Homozygote-Ni+Mel group had a significantly changes the index of the mice(P > 0.05),suggesting that Mel to Ni exposed rats were induced by cardiac oxidative stress effect have antagonism.Compared with the Wild-Ni,Homozygote-Ni group rats were GSSG and MDA content increased significantly,the CAT and the T-SOD activity decreased significantly(P < 0.05),and compared with the Wild-Ni+Mel group,Homozygote-Ni+Mel group had a significantly changes the index of the mice(P < 0.05),suggesting that Txnrd promote into Ni 3 expression reduce exposure induced mice cardiac oxidative stress,inhibit the Mel to Ni exposure of protection.4)Observe the TUNEL staining to detect myocardial cell apoptosis were found to Homozygote-Ni group had a significantly higher than that of Wild mice heart cells apoptosis rate-Ni group(P < 0.05),the Homozygote-Ni group and Wild-Ni myocardial cell apoptosis rate were significantly higher than in Wild-and the Control group(P < 0.05).Suggesting that Txnrd can promote the mice induced by Ni 3 expression reduce myocardial cell apoptosis.5)Observation test mice mitochondrial apoptosis related gene expression in heart tissue results show that compared with the Wild-Control group,the Homozygote-Control groups of mice Caspase3,Caspase 9,BAX,cyt.C and p53 m RNA abundance and protein expression level had no significant change(P > 0.05),and compared with the Wild-Mel group,Homozygote-Mel group no significant change in the index of the mice(P > 0.05),suggesting that pure Txnrd 3 lower expression not promote mice heart dirty tissue mitochondrial apoptosis.Compared with the Wild-Control group,the Wild-Ni group significantly increased the index of the mice(P < 0.05),and compared with Homozygote-Control group,the Homozygote-Ni group significantly increased the index of the mice(P < 0.05),suggesting that Ni can promote mice heart mitochondrial apoptosis.Compared with the Wild-Control group,Wild-Mel group no significant change in the index of the mice(P > 0.05),and Homozygote-compared to the Control group,the index of the small rats were Homozygote-Mel group had no significant change(P > 0.05),suggesting that Mel dose in this test.With Wild-Ni group than the Wild-Ni+Mel group of the index of the rats were significantly lower(P < 0.05),and compared with Homozygote-Ni group,the Homozygote-the index of the Ni+Mel group rats were significantly reduced(P > 0.05),suggesting that Mel to Ni exposed rats were induced by cardiac mitochondrial apoptosis role has antagonism.Compared with the Wild-Ni group,Homozygote-the index of the Ni group rats were significantly increased(P <0.05),and compared with the Wild-Ni+Mel group,Homozygote-Ni+Mel group rats were above refers to the elevated significantly(P < 0.05),suggesting that Txnrd promoted the Ni 3 expression reduce exposure induced mice heart mitochondria conditioning wu,suppresses the Mel to Ni exposure of protection.6)Observation test mice autophagy in the heart tissue and inflammation,the expression of related genes found that,compared with the Wild-Control,Homozygote-Control groups of mice LC 3,ATG1,ATG 7,p62,ATG m TOR,Beclin 1,TNF-α,COX2,IL-1,IL-2,IL-6 and IL-7 no significant change(P > 0.05),and compared with the Wild-Mel group,Homozygote-Mel group no significant change in the index of the mice(P > 0.05),suggesting that pure Txnrd 3 expression of lower won’t promote mice heart tissue autophagy and inflammation.Compared with the Wild-Control group,the Wild-Ni group rats were LC 3,ATG 1,ATG 7,Beclin 1,TNF-α,COX2,IL-1,IL-2,IL-6 and IL-7 significantly increased,the expression of p62 and m TOR lower(P <0.05),and Homozygote-compared to the Control group,the index of the small rats were Homozygote-Ni group changed significantly(P < 0.05),suggesting that Ni exposure can promote the mice heart autophagy and inflammation.Compared with Wild-Control group,Wild-Mel group no significant change in the index of the mice(P > 0.05),and compared with Homozygote-Control group,the Homozygote-Mel group no significant change in the index of the mice(P > 0.05),suggesting that Mel dose in this test.Compared with the Wild-Ni group,Wild-Ni+Mel mice set of LC 3,ATG1,ATG 7,Beclin 1,TNF-α,COX2,IL-1,IL-2,the expression of IL-6 and IL-7decreased significantly,the expression of p62 and m TOR elevated(P < 0.05),and compared with Homozygote-Ni group,the Homozygote-Ni+Mel group changes the index of the mice significantly(P < 0.05),suggesting that Mel to Ni exposed rats were induced by cardiac function autophagy and inflammation have antagonism.Compared with Wild-Ni group,the Homozygote-Ni group of LC 3,ATG 1,ATG 7,Beclin 1,TNF-α,COX2,IL-1,2,IL-6 and IL-7 significantly increased,the expression of p62 and m TOR lower(P < 0.05),and compared with the Wild-Ni+Mel group,Homozygote-Ni+Mel group changes the index of the mice significantly(P < 0.05).In conclusion,the reduction of Txnrd 3 expression promotes Ni induced cardiac mitochondrial apoptosis,autophagy and inflammation in mice through oxidative stress.Mel can protect the heart of mice by reducing ROS,but this protective effect becomes worse after the reduction of Txnrd 3 expression. |
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