PTD-mFoxp3融合蛋白对异基因骨髓移植后移植物抗宿主病的影响

Effect of PTD-mFoxp3 Fusion Protein on Graft-versus-Host Disease after Allogeneic Bone Marrow Transplantation

本研究探讨静脉输注PTD-mFoxp3融合蛋白对同种异基因骨髓移植后移植物抗宿主病的影响。10周龄C57BL/6小鼠为受体并随机分为A、B、C 3组,每组10只,移植当天(第0天)接受直线加速器X射线6.0 Gy全身照射,4-6 h内输注供体BALB/c鼠全骨髓细胞3×107+脾细胞1.5×107个。A组于移植前2 d和移植后0、1、3、5、7、9、11、13 d间断多次输入生理盐水,B组输入mFoxp3蛋白,C组输注等量PTD-mFoxp3融合蛋白。移植后每天观察受体有无移植物抗宿主病的表现;取受体肝脏、空肠上端组织做病理学检查,判断组织损伤及淋巴细胞浸润程度;用ELISA法检测受体外周血中炎性因子IL-2及INF-γ浓度;流式细胞术测定长期存活受体外周血供体细胞的比例。结果表明,照射后60 d内A、B和C组受体平均生存期分别为(32.95±5.48)d、(38.00±5.45)d和(55.30±3.15)d;病理组织学观察示,A组和B组的小肠及肝脏发生明显的损伤,符合GVHD的表现,C组的病理损伤较轻;酶联免疫吸附测定表明,C组IL-2及IFN-γ浓度明显低于A和B组;长期存活的受体形成了稳定的嵌合体状态,移植后60 d A、B、C组活存动物供体细胞比例分别为(79.46±1.80)%、(79.13±2.23)%和(85.92±2.82)%。结论:PTD-mFoxp3融合蛋白可有效降低同种异基因骨髓移植后移植物抗宿主病的发生,并减轻其临床表现,降低死亡率。

This study was aimed to investigate the effect of PTD-mFoxp3 fusion protein on graft-versus-host disease （GVHD） after allogeneic bone marrow transplantation. The 10-weeks-old C57BL/6 mice as recipients were randomly divided into three groups（A,B and C）, 10 mice were in each group. The mice on day of transplantation as on day 0 re- ceived total body irradiation （TBI） 6.0 Gy, then the bone marrow cells （BMC） from BALB/c mice were injected through tail vein within 4 -6 hours. At 2 days before transplantation and 0 ,1,3 ,5 ,7 ,9 and 13 days after transplantation, mice in group A were injected with saline, mice in group B were injected with mFoxp3 protein and mice in group C were injected with PTD-mFoxp3 fusion protein. Symptoms of GVHD, survival time and histopathological changes were ob- served. The establishment of mixed chimerism was determined by flow cytometry in day 60, and IL-2 and IFN-γ expres- sion profiles in the recipient peripheral blood were assessed by ELISA. The results showed that the mean survival time of recipients in group A, B and C was （32.95 ±5.48 ）, （ 38.00± 5.45 ） and （ 55.30 ± 3.15 ） respectively. Graft rejection was observed in the liver and small intestine specimens of group A and group B. The serum levels of IL-2 and IFN-γ sig- nificantly decreased in the recipients of group C, as compared with the other groups. The flow cytometry analysis re- vealed that the survival recipient mice developed high chimerism levels, the percentages of donor cells in group A, B and C were （79.46 ± 1.80） %, （79.13 ± 2.23 ） % and （ 85.92± 2.82 ） % respectively. It is concluded that PTD-mFoxp3 fusion protein can reduce the incidence and mortality of GVHD after allogeneic bone marrow transplantation.