| To establish and evaluate Ischemia/reperfusion (IRI) kidney injury rat model immunosuppressed with low dose cyclosporin (CsA, 3mg/kg/d), and then the expression and its difference of metallothionein (MT) induced with exogenous zinc were investigated. The role of exogenous metallothionein together with low dose CsA in the protection of ischemia/reperfusion injury of kidney and its influence on immunological system were also observed.Methods:(1) Ischemia/reperfusion (IRI) kidney injury rat model immunosuppressed with low dose CsA was established and evaluated by examing the histological changes under microscopy and electromicroscopy as well as the dynamic changes of renal function (Cr, BUN), superoxide dismutase (SOD) and malondiadehyde (MDA).(2) On IRI kidney model immunosuppressed with low dose CsA, the induction of MT with Zinc Chloride in the kidney, spleen and thymus were measured with competitive ELISA. The dynamic expression of MT-mRNA in the kidney was measured with fluonrescent real-time quantitative RT-PCR.(3) Exogenous metallothionein and/or CsA were given on the rat IRI model. The histological changes of kidney were observed under light microscope and electromicroscope. Creatinine, BUN, SOD and MDA were all measured. The apoptosis of renal tubular cells were observed with TUNEL in situ hybridization, and BrdU(+) tubular cells were counted to observe the tubular proliferation.(4) Lymphocytes were isolated from whole blood samples. The ConA-stimulated T lymphocyte proliferation tests were performed for in vivo T cell group and in vitro group by adding MT and/or low dose CsA. PE-CD4+, FITC-CD8+ T lymphocytes were analyzied by cell flowcytometry. The production of IL-2, IL-4 and IFN-y were measured by ELISA in circulating blood serum. The apoptosis of thymus and spleen samples were also observed by TUNEL in situ hybridization.Results:(i)On successfully-established IRI kidney model immunosuppressed with low dose CsA, the damage of renal histological structure, renal function and superoxide were more severe in IRI group than that in pseudosurgery group with significance difference, Such changes were mostly apparent on the timepoint of 24h, serving for determination of the observe timepoint in other parts in this research work. (2)On IRI kidney model immunosuppressed with low dose CsA, MT induced by Zinc was abundantly expressed in proximal and distal renal tubule, but fewer in glomeruler cells. The expression of renal MT increased significantly from 6h, reached the highest level on 24h, and then decreased gradually. There was statistical significance in comparison with control group at the same postoperative timepoints. MTl-mRNA, MT2-mRNA reached the highest level on 6h.Significantly higher MT expression in either spleen or thymus was observed in Zinc group in comparison with control group on 6h, 24h and 72h. Similar to the kidney, significant increasement of MT on 6h with a peak on 24h was observed in spleen and thymus.(3)The damage of renal function and superoxide were significantly ameliorated inexogenous MT group than that in CsA or NaCl group. The apoptosis index (AI) ofrenal tubular cells in MT group was significantly lower than that in NaCl or CsAgroup. The counts of renal BrdU(+) cells in MT group were significantly higher thanthat in NaCl or CsA group.The damage of renal function and superoxide were significantly ameliorated inexogenous CsA group than that in NaCl group, but were significantly severe than thatin MT group. The apoptosis index (AI) of renal tubular cells in CsA group was significantly lower than that in NaCl group. The counts of renal BrdU(+) cells in MT group were significantly higher than that in NaCl group, but were significantly lower than that in MT group.The creatinine (Cr) and blood urea nitrogen (BUN) were lowest in the group with combination usage of MT and CsA, with significant difference comparing with either MT or CsA group. Histological deteriorations were mostly mild in MT+CsA group. SOD and MDA levels in MT...
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