慢病毒载体介导单纯疱疹病毒胸苷激酶/更昔洛韦系统抑制移植物抗宿主病的实验研究

目的研究慢病毒载体介导单纯疱疹病毒胸苷激酶／更昔洛韦（HSV-TK／GCV）系统对小鼠异基因骨髓移植（allo—BMT）后移植物抗宿主病（GVHD）的防治作用。方法将转染HSV-TK基因的慢病毒感染供鼠（C57BL／6小鼠）脾脏淋巴细胞，将感染后的淋巴细胞与供鼠骨髓细胞混合，移植给经。Co1射线照射后的受鼠（BALB／c小鼠）。分别于移植后当天、移植后7天（＋7天）和＋12天腹腔注射GCV25mg／kg×7d，观察受鼠生存期、GVHD发生率及严重程度、T细胞亚群（CD3、CD4、CD8）及异基因嵌合等指标。结果HSV-TK／GCV使用0天、＋7天、＋12天组受鼠生存时间分别为（30．10±5．21）d、（36．40±5．28）d、（28．20±4．82）d，三组生存时间均较移植对照组（15．10±0．43）d]明显延长（P〈0．05）；HSV-TK／GCV＋7天组小鼠50d存活率达60％，高于HSV-TK／GCV0天（40％）和＋12天组（30％）（P〉0．05）。对照组小鼠全部发生Ⅲ～Ⅳ级GVHD，实验组死亡小鼠有Ⅱ～Ⅲ级GVHD病理学改变，而长期生存小鼠仅出现Ⅰ～Ⅱ级GVHD。在＋5，＋10，＋15d，3个时间点实验组小鼠CD4^＋细胞明显高于对照组（P〈0．05），CD8^＋细胞均低于对照组（P〈0．05）。＋30天受鼠异基因嵌合率为100％。结论慢病毒载体介导的HSV-TK／GCV系统能有效控制小鼠allo—BMT后GVHD；＋7天外周血白细胞数开始回升时用GCV控制GVHD效果最佳。

Experimental study of the inhibiting effect of the lentiviral vector mediated herpes simplex virus-thymidine kinase/ganciclovir on GVHD

OBJECTIVE: To study the effect of lentiviral vector mediated herpes simplex virus-thymidine kinase/ganciclovir (HSV-TK/GCV) on graft- versus-host disease (GVHD) after allogeneic bone marrow transplantation (allo- BMT) in mice. METHODS: Donor splenic lymphocytes from C57BL/6 which were infected by lentiviral vectors carrying HSV-TK were transplanted into 60Co gamma ray irradiated recipient mice with donor bone marrow cells. GCV 25 mg x kg(-1) x d(-1) was administered in 3 groups on day 0, +7, +12 respectively after transplant for 7 days by intraperitoneal injection. Survival time, severity of GVHD, incidence of GVHD, T lymphocytes immune reconstruction and of allogeneic chimerism ratio were detected after allo-BMT. RESULTS: The average survival times for GCV 0 day, +7 day and +12 day group were (30. 10 +/- 5.21) d, (36.40 +/- 5.28) d and (28.20 +/- 4.82) d respectively, being significantly longer than that in the control group (15.10 +/- 0.43) d] (P < 0.05). The 50 d-survival rate for TK/GCV + 7 day group was 60%. While for 0 day and +12 day group was 40% and 30% respectively. The incidence of grade III approximately IV GVHD in the control group was 100%, and the dead mice in experimental groups showed pathological changes of II approximately III GVHD. Long-term alive recipient mice only developed grade I approximately II GVHD after allo-BMT. The number of CD4+ lymphocytes in experimental groups was higher than that in control group (P <0.05), but CD8+ lymphocytes was lower on day +5, +10, +15 day (P <0.05). Allogeneic chimerism rate of recipient mice on +30 d was 100%. CONCLUSIONS: HSV-TK/GCV induced by the lentiviral vectors has a definite effect in prevention of GVHD after allo-BMT. GCV adminstrated from 7 days post-transplantation showed the best effects.