| Objective: A research on the effect of HeiJingPaiChiTang on the immunological rejection after High-risk Corneal transplantation in ratMethods:1.To observe the effect of immunological rejection of implant after HeiJingPaiChiTang Through Stomach-perfusion, and compared its effect with the cyclosporine A(CsA) Through Stomach-perfusion,the Combination of HeiJingPaiChiTang and CsA, appraising it effect, providing dosage regimen for clinic.Establish homogeneity variant SD-Wister rat's High-risk corneal transplantation model. randomly group A～D, Group A: negative Control; group B: CsA( l 0mg/Kg/d); group C: HeiJingPaiChiTang (4g/Kg/d);group D: HeiJingPaiChiTang (4g/Kg/d) combined with CsA( l 0mg/Kg/d). observe each group's transparency,the degree of edema, the degree of new vessels ,the index of transplantation rejection, and corneal rejection by slit lamp microscope. Adopt statistics software as a way of statistical analysis. Take the implant of cornea in 4 rats each groups, manufacture HE stain, light microscope and micro-photography analyses technique were used to observe cytological alterations and count the inflammatory cells on the 15 day in post-operation. Then use the statistics software as a way of statistical analysis. Manufacture HE stain for liver and kidney to estimate Drug toxicity on the 30 day after operation.2. To study the mechanism of action for HeiJingPaiChiTang on the immunological rejection after High-risk Corneal transplantation.Establish homogeneity variant SD-Wister rat's High-risk corneal transplantation model randomly group A,B and C. Group A: negative Control; group B: HeiJingPaiChiTang (4g/Kg/d); group C: HeiJingPaiChiTang (4g/Kg/d) combined with CsA (l 0mg/Kg/d). Collect rat's peripheral blood by cutting part of rats' tails on the7,14,21 day after operation, the percentage of CD4~+,CD8~+ T lymphocyte and the ratio of CD4~+/CD8~+ would detected by flow cytometry. The result was dealed by statistics software. Take rat's spleen in asepsis on 25 day after operation, manufacture the spleen T cells suspension. The test of T lymphocyte increment was performed to detect the ability of T lymphocyte increment. The result was dealed by statistics software.Result:1 . To observe the effect of immunological rejection of implant after HeiJingPaiChiTang through Stomach-perfusion.(1)Both C and D group can effectively control the growth of corneal new vessels. The rejection of corneal grafts could be significant inhibited by HeiJingPaiChiTang(group C), and group D is the best. From the implant's survival time we can see group A is the shortest one (10.38±1.69d), group D is the longest(22.5±3.07d) .(2)the result of histology(HE stain): on the 15 day after operation , corneas grafts in control (A group) became edema and vascularized, which were heavily infiltrated with inflammatory cells, there were few inflammatory cells and new vessels in experimental group B and C, which no inflammatory cells and new vessels in experimental groups D. There was no significant pathological change in liver and kidney.(3) The count of Neutrophils and lympholeukocyte in Medication group (B,C,D) was obviously reduced compared with the control.2.To study the mechanism of action for HeiJingPaiChiTang on the immunological rejection after High-risk Corneal transplantation.(1)The result of flow cytometry: On the 1 week, The expression of CD4~+ molecule and the ratio of CD4~+/CD8~+ in experimental C was obviously down-regulation.compared with A(P<0.05) ;The expression of CD4~+ molecule in experimental B,C was down-regulation on the 2 week, and it have a significant difference compared the control group(P<0.05). And on the 3 week, The expression of CD4~+ molecule and the ratio of CD4~+/CD8~+ in experimental B and C was down-regulation compared with the control (P<0.05) .there was no difference between control and experimental group on CD8~+ expression(P>0.05).(2)The T lymphocyte increment: group B and C had significant down-regulation compared with A (P<0.01), especially C.Conclusion:1. HeiJingPaiChiTang have obviously Anti-inflammatory Action .It can inhibit the growth of corneal new vessels.2.HeiJingPaiChiTang can prolong the mean survival time to some extent, and it have synergistic effect with CsA, which offer some reference Value for clinic.3.HeiJingPaiChiTang can effectively inhibit the expression of CD4~+ molecule and cut down the ratio of CD4~+/CD8~+, it also can control the T lymphocyte increment, which maybe some of the mechanism of action for HeiJingPaiChiTang on the immunological rejection after High-risk Corneal transplantation.. |
