| Objectives:Ischemia-reperfusion can induce a variety of body organs injure including the spleen, liver and kidney. A large number of inflammatory cytokines are expressed in these organs, finally resulting in apoptosis. There were three sections in this study:(1) The feeding and breeding process of metallothionein Ⅰ/Ⅱ (Metallothionein Ⅰ/Ⅱ, MT Ⅰ/Ⅱ) knockout (Knockout, KO) mice were observed;(2) The effects of N-acetylcysteine (NAC) and diethyldithiocarbamic acid (DETC, the inhibitor of SOD) on the antioxidant activity of the spleen cells were investigated in the MT-Ⅰ/Ⅱ KO cerebral ischemia-reperfusion injury model established by the left middle cerebral artery occlusion (Middle Cerebral Artery Occlusion, MCAO).Methods:1. The processes of feeding and breeding MT Ⅰ/Ⅱ KO mice were observed2. The MT-Ⅰ/Ⅱ KO cerebral ischemia-reperfusion injury model was established by MCAO. Compaerd with the sham group, the following indexes were tested at the24h time point after ischemia-reperfusion injury:(1) neurological deficit by five-point method.(2) spleen cell viability with and without the treatment of NAC (2h) or DETC (2h) by MTT colorimetric method.(3) the dilation of mPTP in MT-Ⅰ/Ⅱ KO mice cerebral ischemia and reperfusion at the24h time point in spleen cells and spleen cells given NAC or DETC treatment after2hours were detected.(4) Ca2+uptake release was detected by Ca2+transport assay.(5) lactate dehydrogenate (LDH) leakage rate in culture supernatant with and without the treatment of NAC (2h) or DETC (2h) were assayed by microplate reader.Results:1. Compared with wild mice, MT-I/II KO mice had the following characteristics including lower reproductive capacity, like quiet, easy mass, and low tolerance of dry.2. Effect of NAC on spleen cells in KO MT mice after cerebral ischemia reperfusion injury:(1) at the24h time point after ischemia-reperfusion injury, neural functional defect was observed in KO MT mice.(2) For both WT and MT-Ⅰ/Ⅱ KO mice, compared with that in the sham control group, spleen cell viability in ischemia reperfusion injury group was reduced (P <0.01); after NAC treatment, spleen cell viability in mice of the sham control group and ischemia reperfusion injury group were significantly increased (P<0.01). Compared with WT, MT-Ⅰ/Ⅱ KO mice in ischemia reperfusion injury group had lower spleen cell viability (P<0.01). The increase of spleen cell viability of MT-Ⅰ/Ⅱ KO mice in ischemia reperfusion injury after NAC treatment was lower than that of WT mice (P<0.05). Compared with the sham control group, DETC treatment inhibited the spleen cell viability in both WT and MT-Ⅰ/Ⅱ KO mice (P<0.01). Compared to the ischemia reperfusion injury control group, DETC treatment further inhibited the spleen cell viability in both WT (P<0.05) and MT-Ⅰ/Ⅱ KO mice (P<0.01). The inhibition of spleen cell viability of MT-Ⅰ/Ⅱ KO mice in ischemia reperfusion injury after DETC treatment was enhanced than that of WT mice.(3) For both WT and MT-Ⅰ/Ⅱ KO mice, compared with that in the sham control group, the dilation of mPTP in ischemia reperfusion injury group was increased (P<0.01); after NAC treatment, the dilation of mPTP in mice of the sham control group and ischemia reperfusion injury group were significantly reduced (P<0.01). Compared with WT, MT-Ⅰ/Ⅱ KO mice in ischemia reperfusion injury group had increased the dilation of mPTP (P<0.01). The decrease of dilation of mPTP of MT-Ⅰ/Ⅱ KO mice in ischemia reperfusion injury after NAC treatment was lower than that of WT mice (P<0.05). No significant difference in the dilation of mPTP was observed after DETC treatment (P>0.05). (4) For both WT and MT-Ⅰ/Ⅱ KO mice, compared with that in the sham control group, the Ca2+overload in ischemia reperfusion injury group was enhanced (P <0.01); after NAC treatment, the Ca2+overload in mice of the sham control group and ischemia reperfusion injury group were significantly declined (P<0.01). Compared with WT, MT-Ⅰ/Ⅱ KO mice in ischemia reperfusion injury group had enhanced Ca2+overload (P<0.01). The reduction of Ca2+overload of MT-Ⅰ/Ⅱ KO mice in ischemia reperfusion injury after NAC treatment was lower than that of WT mice (P<0.01). No significant difference in the Ca2+overload was observed after DETC treatment (P>0.05).(5) For both WT and MT-Ⅰ/Ⅱ KO mice, compared with that in the sham control group, the LDH leakage rate in ischemia reperfusion injury group was significantly increased (P<0.01); after NAC treatment, the LDH leakage rate in mice of the sham control group and ischemia reperfusion injury group were significantly declined (P<0.01). Compared with WT, MT-Ⅰ/Ⅱ KO mice in ischemia reperfusion injury group had increased LDH leakage rate (P<0.01). The decrease of LDH leakage rate of MT-Ⅰ/Ⅱ KO mice in ischemia reperfusion injury after NAC treatment was lower than that of WT mice (P<0.01). Compared with the sham control group, DETC treatment induced the LDH leakage rate in both WT and MT-Ⅰ/Ⅱ KO mice (P<0.01). Compared to the ischemia reperfusion injury control group, DETC treatment further increased the LDH leakage rate in both WT (P<0.01) and MT-Ⅰ/Ⅱ KO mice (P<0.01). The induction of LDH leakage rate of MT-Ⅰ/Ⅱ KO mice in ischemia reperfusion injury after DETC treatment was enhanced than that of WT mice.Conclusions:1. MT-Ⅰ/Ⅱ KO mice feeding should pay attention to the environment clean to prevent water shortages and environmental clean-induced generation of tumor.2. Cerebral ischemia and reperfusion have an negative impact on spleen cells of the peripheral immune organs, while antioxidant NAC has a protective effect on peripheral immune organs spleen cells after cerebral ischemia and reperfusion. DETC, the inhibotor of SOD, showed enhanced ischemia reperfusion injury by inhibiting the spleen cell viability and increasing LDH leakage rate. |
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