| Objective The experiment aimed to discuss the effect of1,25dihydroxyvitaminD3towards murine immune injection in penetrating keratoplasty united skintransplantation model. Then further explore the immune regulation of murine immunesystem.Methods In the experiment, C57BL/6mice were taken as recipients, Balb/c miceas donors. The C57BL/6mice were divided into control group、intervention group andtransplanted group according to completely randomized design method. Establish theanimal model of penetrating keratoplasty united with skin transplantation, theintervention group were given a10ng dose of1,25-Dihydroxyvitamin D3dissolved inthe peanut oil every other day, the control group and transplanted group were givenequal dose of normal saline. Observe the corneal graft with slit lamp after operationand observe the skin graft with naked eye.On day13after the transplantation, the mice were executed after weighing thethree groups of mice. Then take the spleens of the mice, then calculated thespleen/weight ratio. The lymphocytes were separated with lymphocyte separationmedium. Total lymphocyte RNA was extracted with trizol, then acquire thesingle-stranded DNA with reverse transcription process. In the end, detect themessage RNA with quantitative polymerase chain reaction(Q-PCR). Observe thedifferences of T helper1associated cytokine IFN-γ mRNA and IL-2mRNA、T helper2associated cytokine IL-10mRNA and T helper17associated cytokine IL-17AmRNAof spleen lymphocyte. Then speculate the effect of1,25dihydroxyvitamin D3towardscytokines.On day13after the transplantation, get the peripheral blood of the three group,then measure the difference and variation of γδT cell and CD4+CD25+Treg, detectthe regulatory effect of1,25dihydroxyvitamin D3towards these2kinds of cells. Statistical analysis：All the experimental data was presented with mean±standarddeviation。 The SPSS19.0was used to analysis the data. There was no statisticallysignificant difference when P＞0.05，the difference was statistically significant whenP＜0.05.Results The survival period of corneal graft and skin graft in the group intervenedby1,25dihydroxyvitamin D3was longer than the transplantation group, theproportion of peripheral γδT cells of grafted group was obviously higher than controlgroup and intervened group, the percentage in intervention group was higher thancontrol group,1,25dihydroxyvitamin D3down-regulated the γδT cell.CD4+CD25+Treg in the peripheral blood of transplantation group was significantlylower than the control group and intervention group, and that in the intervention groupwas higher than control group.1,25dihydroxyvitamin D3up-regulated the thenumber of CD4+CD25+Treg. In terms of the spleen lymphocytes, the expression ofTh1related IFN-γ mRNA'IL-2mRNA in transplantation group was obviouslyhigher than the other two groups, intervention group was higher than control group,Th17related cytokine IL-17mRNA in grafted group was higher than control andintervention group. Expression of Th2related IL-10mRNA in intervention group washigher than control and transplantation group. The difference of the three groups wasstatistically significant. Therefore,1,25-dihydroxyvitamin D3might down-regulatethe Th1related IFN-γmRNA、IL-2mRNA and Th17related IL-17mRNA expression,up-regulate Th2related IL-10mRNA expression.Conclusion These results revealed that1,25dihydroxyvitamin D3might inhibitthe immune rejection of allogeneic mice corneal transplantation model viaup-regulating the quantity of CD4+CD25+Treg cells which were able to inhibit theimmune rejection through promoting immunologic tolerance and down-regulating theproportion of γδT cells which have a promoting effect to immune rejection.1,25dihydroxyvitamin D3inhibited the expression of inflammatory related cytokines, like interferon-γ、interleukin-2and interleukin-17A,1,25dihydroxyvitaminD3restrained the immune rejection by promoting the expression of interleukin-10thatshowed inhibitory effect. Therefore,1,25dihydroxyvitamin D3is likely to be aneffective measure to inhibit the immune rejection. |
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