FTY720对小鼠异基因骨髓移植后急性GVHD的预防作用

目的评估FTY720对小鼠异基因骨髓移植诱发急性移植物抗宿主病(aGVHD)的预防作用.方法 36只受体BALB/c小鼠,接受直线加速器一次性全身照射后4 ～ 6 h内,经尾静脉注射供体小鼠骨髓和脾脏细胞.受体小鼠分为移植方式对照组(A组)和异基因移植治疗组(B组);同时A组分未移植组(A1)、同基因移植组(A2)和异基因移植对照组(A3),B组分CsA 5 mg/kg组(B1),FTY720 0.3 mg/kg组(B2)和FTY720 1 mg/kg组(B3).移植后受体小鼠,根据生存时间、体重变化和病理组织学检查,评估各组aGVHD严重程度.结果异基因移植各组在移植后第7 d造血开始恢复,第14 d流式细胞仪检测显示异基因骨髓细胞已植入,病理组织学检查A3组发生重度aGVHD;与B1和B2组比较,B3组aGVHD严重程度最低.结论小鼠异基因骨髓移植后给予FTY720具有预防aGVHD作用,以剂量为1 mg/kg时作用显著.     

雷帕霉素抑制小鼠骨髓移植后移植物抗宿主病作用的初步研究

目的：探讨新型免疫抑制药物雷帕霉素（RAPA）对小鼠异基因骨髓移植（allo-BMT）后急性移植物抗宿主病（aGVHD）的抑制作用，并与传统免疫抑制药物比较作用效果。方法：用主要组织相容性抗原（MHC）完全不合的纯种近交系小鼠[供鼠为雄性c57BL／6J（H-2“）鼠，受鼠为雌性BALB／c（H-2^d）鼠]建立allo-BMT／aGVHD动物模型，随机分5组，第1、2和3组分别给予RAPA、环孢菌素A（CsA）加甲氨蝶呤（MTX）、RAPA加CsA加MTX联合用药作为aGVHD抑制方案，第4组为空白对照，第5组为未移植组。观察各组小鼠骨髓移植后aGVHD的出现情况。结果：移植小鼠出现典型的aGVHD症状，未用aGVHD预防方案的移植小鼠（第4组）死亡高峰在移植后第5～7天，死亡率达100％。用不同aGVHD预防方案的第1～3组小鼠aGVHD症状明显减轻，平均生存时间（MST）较第4组显著延长（分别延长3．5、1．9和6．0d，P〈0．05）。不同aGVHD预防方案的移植小鼠之间，移植后外周血常规变化差异无显著性。经用各预防方案后，移植小鼠aGVHD病理表现减轻，且RAPA、CsA和MTX联合用药组的病理分级轻于RAPA单药组或CsA加MTX标准方案组。结论：RAPA可以显著抑制体外T淋巴细胞增殖效应，增强小鼠对骨髓移植的免疫耐受，对小鼠骨髓移植后移植物抗宿主病（GVHD）有明显抑制作用，可减轻GVHD症状和病理损害程度，延长平均生存时间。     

来氟米特预防小鼠异基因骨髓移植后aGVHD的实验研究

目的 评估来氟米特（LEF）对小鼠异基因骨髓移植（allo-BMT）术后急性移植物抗宿主病（aGVHD）的预防作用.方法 以SPF级雄性C57BL/6（H-2b）为供鼠,雌性BALB/C（H-2d）为受鼠,建立allo-BMT/aGVHD动物模型,并将大鼠随机分为：A组（模型对照组）、B组（环孢菌素A组,剂量为10 mg·kg^-1·d^-1）、C组（LEF组,剂量为10 mg·kg^-1·d^-1）,同时根据临床表现、病理组织学检查、平均生存时间评估各组aGVHD的严重程度.结果 B组和C组受鼠平均生存时间均较A组显著延长（均P〈0.01）,B组和C组间平均生存时间的差异均无统计学意义（均P〉0.05）;B组和C组受鼠的aGVHD临床表现和病理改变均较A组明显减轻.结论 LEF可有效预防小鼠allo-BMT术后出现的aGVHD,并可减轻其症状和病理损害程度,同时显著延长平均生存时间.     

大鼠异基因骨髓移植急性移植物抗宿主病模型的建立

目的建立较稳定的异基因骨髓移植急性移植物抗宿主病动物模型,为异基因骨髓移植后的急性移植物抗宿主病(aGVHD)的相关研究提供实验参照。方法以雄性SD大鼠为供鼠,雌性Wistar大鼠为受鼠,受体大鼠随机分成A、B、C、D、E 5组,移植当天所有受鼠均接受8.5 GY的全身照射(TBI),于照射后4~6 h内,A组回输等量培养液,B组经尾静脉输注供鼠骨髓细胞(2×108个/kg),C、D、E组分别回输供鼠骨髓细胞(2×108个/kg) 不同比例的脾细胞。观察各组大鼠生存期、外周白细胞计数、及有无aGVHD的临床及病理表现。结果A组大鼠于15d内全部死亡,外周血白细胞计数明显减低,骨髓病理示造血组织减少,提示死于造血衰竭。B、C、D、E组大鼠外周血白细胞计数均有明显恢复,B组大鼠8只存活超过50 d,C、D、E组大鼠均于50 d观察期内死亡,并有aGVHD的临床表现及病理表现,但C组大鼠aGVHD的程度较轻且时间不集中,其中D、E组大鼠可于相对集中的时间内观察到典型aGVHD临床及病理。结论TBI预处理的方式是可行的,单纯输入异基因骨髓细胞不能引起明显的aGVHD,骨髓细胞与脾细胞1∶1及1∶1.5混合组均可作为异基因骨髓移植后理想的aGVHD动物模型。     

核转录因子-kB在同种异基因小鼠角膜移植排斥反应中的表达

目的：检测小鼠角膜移植后核转录因子KB（NF-kB）活性变化。方法：建立以C57BL／6小鼠为供体，以BALB／c小鼠为受体角膜移植实验模型，随机分为A、B、C三组，A组为正常小鼠组，B组为自体角膜移植作为对照组，C组同种异基因角膜移植作为实验组，ELASA方法检测小鼠角膜移植后3d、5d、7d角膜NF-kB的活性变化。结果：组织学检查证实，同种异基因小鼠角膜移植后14d见明显淋巴细胞浸润。3d、5d角膜NF-kB的活性升高，7d达到高峰，与原位移植组有显著差异（P〈0．01）。结论：小鼠同种异基因角膜移植后，角膜NF-kB的活性升高，可能参与移植排斥。     

雷帕霉素抑制小鼠骨髓移植后移植物抗宿主病作用的初步研究

目的:探讨新型免疫抑制药物雷帕霉素(RAPA)对小鼠异基因骨髓移植(allo-BMT)后急性移植物抗宿主病(aGVHD)的抑制作用,并与传统免疫抑制药物比较作用效果。方法:用主要组织相容性抗原(MHC)完全不合的纯种近交系小鼠[供鼠为雄性C57BL/6 J(H-2b)鼠,受鼠为雌性BALB/c(H-2d)鼠]建立allo-BMT/aGVHD动物模型,随机分5组,第1、2和3组分别给予RAPA、环孢菌素A(CsA)加甲氨蝶呤(MTX)、RAPA加CsA加MTX联合用药作为aGVHD抑制方案,第4组为空白对照,第5组为未移植组。观察各组小鼠骨髓移植后aGVHD的出现情况。结果:移植小鼠出现典型的aGVHD症状,未用aGVHD预防方案的移植小鼠(第4组)死亡高峰在移植后第5~7天,死亡率达100%。用不同aGVHD预防方案的第1~3组小鼠aGVHD症状明显减轻,平均生存时间(MST)较第4组显著延长(分别延长3.5、1.9和6.0 d,P<0.05)。不同aGVHD预防方案的移植小鼠之间,移植后外周血常规变化差异无显著性。经用各预防方案后,移植小鼠aGVHD病理表现减轻,且RAPA、CsA和MTX联合用药组的病理分级轻于RAPA单药组或CsA加MTX标准方案组。结论:RAPA可以显著抑制体外T淋巴细胞增殖效应,增强小鼠对骨髓移植的免疫耐受,对小鼠骨髓移植后移植物抗宿主病(GVHD)有明显抑制作用,可减轻GVHD症状和病理损害程度,延长平均生存时间。     

骨化三醇及复方丹参对大鼠异基因骨髓移植后急性移植物抗宿主病的影响

目的 观察骨化三醇及丹参对异基因骨髓移植(Allo-BMT)后急性移植物抗宿主病(aGVHD)的影响.方法 以雄性SD大鼠为供鼠,雌性Wistar大鼠为受鼠,受体大鼠随机分成A、B、C、D组,移植当天所有受鼠均接受8.5Gy的全身照射(TBI),于照射后4～6h,A组,为骨髓细胞与脾细胞1:1.5联合输注组:输注供鼠的骨髓细胞2×108个/kg+脾细胞3×108个/kg,即GVHD模型组.B、C、D组除建立GVHD模型外,B组为骨化三醇组;C组为中药制剂复方丹参组;D组为骨化三醇和中药制剂复方丹参联合组.观察各组大鼠生存期及有无aGVHD的临床及病理表现.结果 A、B、C、D组大鼠均于50d观察期内死亡,并有aGVHD的临床及病理表现,但是B、C、D组大鼠aGVHD的发病时间较A纽延迟,临床aGVHD评分亦较A组大鼠低(P<0.05),平均生存时间较A组大鼠明显延长(P<0.05),病理表现较A组大鼠轻.结论 骨髓细胞与脾细胞混合榆注可以引起明显的aGVHD,骨化三醇及丹参能够减轻Allo-BMT后aGVHD的发生.     

延迟连续移植预防小鼠移植物抗宿主病的实验研究

目的研究延迟连续移植对小鼠异基因移植后急性移植物抗宿主病（aGVHD）的影响。方法近交系BALB/c（H-2d）受鼠接受8.0 Gy全身放疗（TBI）后,分4组（每组n=20）输注供鼠C57BL/6（H-2b）骨髓和脾细胞的混合液：①经典移植组（TBI后4 h输注）;②连续移植组（TBI后4 h、1-3天连续输注）;③延迟移植组（TBI后4天输注）;④延迟连续移植组（TBI后4-7天连续输注）。流式细胞仪检测移植后各组T细胞亚群、NK细胞、H-2b的表达,比较各组受鼠生存率、aGVHD情况及造血重建。结果经典移植组均于3周内死于aGVHD,连续移植组、延迟移植组60天生存率分别是30%、50%。延迟连续移植组60天生存率为70%,高于其他3组（P＜0.05）。延迟连续移植组20天时外周血白细胞计数恢复正常,30天时T细胞亚群和NK细胞的表达恢复正常,60天时H-2b细胞的百分率为98.13%±1.11%。结论延迟连续骨髓移植可减轻小鼠异基因移植后的aGVHD,提高生存率,同时不影响骨髓植入、造血和免疫重建,是一种简单有效的预防aGVHD的新方法。     

异基因造血干细胞移植后小鼠共刺激分子表达与急性移植物抗宿主病的关系

目的 探究异基因造血干细胞移植后小鼠共刺激分子表达及其与急性移植物抗宿主病（aGVHD）发病的关系。方法 供体为5只C57BL/6J（H2KD^-H2KB⁺）雄性小鼠,受体为30只CB6F1（H2KD⁺H2KB⁺）雌性小鼠。将受体小鼠随机分为单纯照射组、骨髓移植组和aGVHD组,每组10只。单纯照射组：受体小鼠接受γ射线照射后不注射任何细胞;骨髓移植组：受体小鼠接受γ射线照射后注射5×10⁶个供体来源去T淋巴细胞的骨髓有核细胞;aGVHD组：受体小鼠接受γ射线照射后,同时注射5×10⁶个供体来源去T淋巴细胞的骨髓有核细胞和3×10⁷个供体来源脾细胞。采用log-rank生存曲线分析各组小鼠移植后生存情况。分别在移植后7、14、21、28 d采用流式细胞术检测骨髓移植组和aGVHD组小鼠外周血CD4⁺T淋巴细胞和CD8⁺T淋巴细胞表面共刺激分子细胞毒性T淋巴细胞相关抗原4（CTLA-4）、程序性死亡蛋白1（PD-1）、可诱导共刺激分子（ICOS）和CD28的表达情况,以及移植后21 d两组小鼠脾组织CD4⁺T淋巴细胞内白细胞介素（IL）-17、γ干扰素、IL-4的表达情况,并在移植后21 d采用3,3''-二氨基联苯胺（DAB）染色检测两组小鼠结肠组织共刺激分子配体的表达情况。结果 单纯照射组受体小鼠均在γ射线照射后19 d内死亡;骨髓移植组小鼠在γ射线照射后均未出现aGVHD症状,30 d内均存活;aGVHD组小鼠在γ射线照射后出现aGVHD症状的时间为14（11~18）d,生存时间为22（13~30）d。随着时间的推移,aGVHD组小鼠外周血CD4⁺和CD8⁺T淋巴细胞表面CTLA-4、PD-1表达在移植后均呈下降趋势,ICOS、CD28表达均呈上升趋势。在移植后28 d时aGVHD组小鼠外周血CD4⁺和CD8⁺T淋巴细胞表面CTLA-4、PD-1表达均低于骨髓移植组,ICOS、CD28表达均高于骨髓移植组,差异均有统计学意义（P均<0.05）。DAB染色结果示,骨髓移植组结肠组织CD80、ICOS配体（ICOSL）、PD-1配体（PD-L1）表达均为阴性,aGVHD组结肠组织CD80、ICOSL、PD-L1阳性表达率分别为40%、80%、80%。与骨髓移植组比较,aGVHD组小鼠脾组织CD4⁺T淋巴细胞内IL-17、γ干扰素表达均增加,IL-4表达减少,差异均有统计学意义（P均<0.05）。结论 异基因造血干细胞移植小鼠发生aGVHD后,共刺激分子CTLA-4表达随时间推移逐渐降低。CTLA-4、ICOS、PD-1均可能通过调节辅助性T细胞（Th）1、Th2、Th17等T淋巴细胞亚群分布参与aGVHD的发生、发展。     

B7反义肽预处理的受体树突状细胞抑制移植动脉的内膜增生

目的建立同种异基因小鼠颈总动脉原位移植模型,探讨B7反义肽预处理的受体树突状细胞（DC）抑制移植动脉内膜增生的有效性。方法利用B7反义肽封闭负载供体抗原的受体DC,对受体C3H小鼠进行预处理后,建立异基因小鼠（Cm7BL／6→C3H）颈总动脉原位移植模型,2个月后对移植动脉作病理分析。结果DC预处理组移植动脉内膜增生减弱（P〈0．05）．免疫组化分析示TGFβ1在动脉壁内的表达减少（P〈0．05）。结论B7反义肽预处理的负载供体抗原的受体DC能够抑制移植动脉内膜的增生。     

昆明山海棠提取物预防急性移植物抗宿主病的实验研究

目的观察昆明山海棠（THH）提取物在急性移植物抗宿主病（aGVHD）中的作用及其可能的作用机制。方法建立aGVHD模型,BALB/c受鼠清髓性预处理后分别用生理盐水、环孢素A（CsA）、THH及CsA＋THH混合物灌胃。观察移植后受鼠aGVHD、生存时间、CD4^＋/CD8^＋比率、CD4^＋CD25^＋T细胞比率及脾细胞Foxp3 mRNA水平。结果生理盐水组出现典型aGVHD,20天内全部死亡。其余3组aGVHD临床、病理表现及生存时间均改善,组间无差异;CD4^＋/CD8^＋比率、受鼠CD4^＋CD25^＋T细胞比率及脾Foxp3 mRNA表达较生理盐水组升高,以联用组明显。结论THH可减轻小鼠aGVHD,延长其生存时间。机制可能与增加CD4^＋CD25^＋T细胞比率,上调Foxp3 mRNA表达,调节CD4^＋/CD8^＋比率有关。THH和小剂量CsA联合有协同作用。     

小鼠骨髓移植修复卵巢功能损伤

目的研究骨髓移植对化疗引起的小鼠卵巢功能损伤的修复作用。方法 48只小鼠随机分为3组:正常对照组(A组)、化疗并骨髓移植组(B组)和化疗组(C组)。B、C组小鼠每天给予环磷酰胺150 mg/kg腹腔注射,连续3 d。1周后,对B组小鼠进行骨髓移植。给药起第21、50天分两批处死小鼠,称量卵巢、子宫湿重,观察卵巢组织学变化。用Hoechest33342荧光染料标记骨髓细胞,移植入预先注射环磷酰胺的小鼠体内,2 d后取小鼠卵巢做冰冻病理切片,荧光显微镜下观察。结果给药第21、50天,B组小鼠的卵巢系数、各级卵泡数目明显高于C组而低于A组,均具有统计学意义(P<0.05)。受体鼠的卵巢内发现显示荧光的供体鼠骨髓细胞。结论骨髓移植对化疗药物引起的卵巢功能损伤具有修复作用。     

Graft-versus-leukemia effects from donor lymphocyte infusion after nonmyeloablative allogeneic bone marrow transplantation in mice

Background Nonmyeloablative allogeneic bone marrow transplantation has been used since the 1990s as a new hematological stem cell transplantation strategy for treating hematological diseases. The purpose of this study was to explore the graft-versus-leukemia (GVL) effects of donor lymphocyte infusions (DLIs) after nonmyeloablative allogeneic bone marrow transplantations, while assessing the declines in treatment-associated morbidity, mortality, and graft-versus-host disease (GVHD).Methods A total of 615 (H-2k) mice were injected with L615 tumor cells and received 500 cGy (60Coγ-ray) irradiation three days later, followed by an allogeneic bone marrow transplantation (allo-BMT). The allo-grafts consisted of 3×10（7） bone marrow cells and 1×10（7） spleen cells from BALB/C (H-2d) donor mice. Two days after the allo-BMT, the recipient mice were given 200 mg/kg of cyclophosphamide. Subsequently, recipient mice were infused with either donor spleen cells ［2×10（7）］ on day 14 or 21, or donor spleen cells ［5×10（7）］ pretreated with hydrocortisone and cyclosporin A (CsA) in vitro on day 14 post-BMT.Results The median survival time of mice that received DLI on day 21 and pretreated DLI on day 14 post-BMT was longer than that of controls and the day 14 DLI group (P&lt;0.01). No evidence of severe GVHD was observed in the day 21 DLI group nor in the day 14 treated DLI group. Mixed chimerism was confirmed in the day 14 DLI group, the day 14 treated DLI group, and the day 21 DLI group on the thirteenth day post-transplantation; full donor chimerism was observed two weeks after DLI.Conclusion Donor lymphocyte infusion after nonmyeloablative bone marrow transplantation may reduce transplantation-associated morbidity and mortality while strengthening graft-versus-leukemia effects.     

小鼠同种异基因骨髓移植aGVHD动物模型的建立

目的：建立一个德定可靠的异基因骨髓移植（allogeneic bone marrow transplantation,Allo-BMT）急性移植物抗宿主病（acute graft-vetsus-host disease,aGVHD）动物模型,为Allo-BMT中aGVHD研究提供理想的实验平台。方法：以雄性C57BL／6（H-2b）为供鼠,雌性BALB／C（H-2d）为受鼠,受鼠致死性全身照射（total body irradiation,TBI）（8.0Gy）预处理后,在移植物中混合不同比例的供鼠骨髓细胞和脾细胞以引起不同程度的aGVHD。根据存活期、外周血白细胞计数、临床表现及病理检查等判断aGVHD程度。结果：单纯异基因骨髓组只有部分小鼠出现aGVHD,75％的小鼠长期存活（〉30d）。混合不同比例的供鼠骨髓细胞和脾细胞的Allo-BMT小鼠出现aGVHD的时间和程度均有差异,其中骨髓与脾细胞1.5：1或1：1混合组小鼠,均可在相对集中的时间内（8-15d）观察到典型的aGVHD临床和病理改变。结论：建立Allo-BMT aGVHD模型需要加入一定比例的脾细胞,供鼠骨髓与脾细胞1.5：1或1：1混合进行的Allo-BMT,可作为理想的aGVHD动物模型。     

重组人白细胞介素18对小鼠骨髓细胞移植后急性移植物抗宿主病的影响

目的：探讨重组人白细胞介素18（rhIL-18）对小鼠异基因骨髓移植后急性移植物抗宿主病（aGVHD）的影响，为rhIL-18用于临床移植免疫提供理论基础。方法：受体鼠（BALB／c）接受9．0Gy剂量的X-射线照射之后，通过其尾静脉注射供体鼠（C57BL／6）的骨髓细胞和脾细胞，并于移植的-2～＋2d腹腔注射rhIL-18。移植后观察受体鼠的临床症状和存活状况，同时检测其肝脏、结肠和耳朵的病理变化。结果：移植后aGVHD期内，用rhIL-18治疗的受体鼠GVHD症状积分较未接受rhIL-18治疗的受体鼠低（P ＜ 0．05），肝脏和结肠的病理变化轻（P ＜ 0．05），短期存活时间延长（P ＜ 0．05），但对长期生存率无明显影响。结论：受体鼠早期接受rhIL-18治疗，可在移植后aGVHD期内降低aGVHD的病变程度，为rhIL-18在临床上预防和治疗aGVHD提供了实验依据。     

大小鼠之间造血干细胞嵌合体抗异种移植物抗宿主病

OBJECTIVE: To find a method for preventing graft-versus-host disease (GVHD) in BALB/c mice receiving rat bone marrow transplantation. METHODS: Firstly 12 SD rats were conditioned with 5.0 Gy sublethal total body irradiation(TBI), followed by infusion of 8 x 10(7) bone marrow cells from 28 BALB/c mice within 4 h and intraperitoneal administration of 100 mg/kg cyclophosphamide (Cy) 2 d later, for the purpose of inducing chimeric SD rats with specific immunological tolerance. Secondly, 24 BALB/c mice were exposed to 9.0 Gy lethal TBI and divided randomly into 3 equal groups designated respectively as groups A, B and C. Mice in group A were injected with 4 x 10(7) bone marrow cells from 4 normal rats, and mice in group B were injected with 4 x 10(7) bone marrow cells and 2 x 10(7) spleen cells from 4 normal rats, while those in group C were given 4 x 10(7) bone marrow cells and 2 x 10(7) spleen cells from 6 chimeric rats. The mice were then observed for 150 d for GVHD. RESULTS: In the second procedure, 2 mice in group A failed to survive due to infection and radiation injury respectively, and none of the rest mice presented signs of GVHD. The mice in group B all developed GVHD of varied degrees with symptoms as wasting, diarrhea, fur loss, arched body posture, and even bloody stool, and all died within 5 to 15 d with an average survival of 10.0 d (95% confidence interval 8,12). Pathological examination of the liver and intestinal tissues revealed inflammatory lymphocyte infiltration and necrosis. The majority of the mice in group C, in contrast, survived for more than 150 d with only two died on the postoperative days 18 and 31 respectively. The survival time of group B was significantly shorter than that of the other two groups. CONCLUSION: Donor/recipient chimerism may help prevent GVHD in xenogeneic bone marrow transplantation.     

TH17细胞在小鼠皮肤急性移植物抗宿主病中的作用

目的 观察TH17细胞在小鼠皮肤急性移植物抗宿主病(aGVHD)中的作用.方法 将小鼠分为3组:单纯照射组(照射组,60Co照射1次)、异基因骨髓移植组(移植组,60Co照射后进行异基因骨髓移植)和常山酮组(60Co照射后进行异基因骨髓移植,移植前1 d予TH17抑制剂常山酮腹腔注射),每组10只.移植后观察皮肤aGVHD发生情况并进行临床和病理评分.移植后6 d流式细胞仪检测血中TH1/TH17、γ干扰素(IFN-γ)及白细胞介素17(IL-17)变化.结果 小鼠在移植后6 d出现典型的系统性aGVHD表现,常山酮组小鼠系统性aGVHD较移植组重.常山酮组小鼠皮肤aGVHD临床表现和病理评分较移植组轻;常山酮组TH1/TH17中位数为17.57,移植组TH1/TH17中位数为5.31,两组比较差异有统计学意义(P<0.05).照射组IL-17低于移植组[(1.47±0.18)pg/ml比(2.81±0.19)pg/ml,P<0.05],常山酮组未测到.对照组、移植组、常山酮组IFN-γ分别为(3.86±0.32)、(42.97 ±0.42)、(9.89±0.51)pg/ml,各组间比较差异均有统计学意义(均P<0.05).结论 阻断TH17能够减轻小鼠皮肤aGVHD但加重系统性aGVHD.

Abstract：

Objective To explore the functions of TH17 cell in cutaneous graft-versus-host disease (GVHD). Methods A model of acute GVHD (aGVHD) was established with a major histocompatibility complex class Ⅰ /Ⅱ -disparate allogeneic bone marrow transplantation( BMT). Bone marrow monocytes and splenic T cells from donor C57/BL6 were enriched. The recipient BABL mice were irradiated ( Co source)with 7. 5 Gy total body irradiation (TBI) and injected with 5 × 106 marrow monocytes and 5 × 105 T cells.The experimental mice were divided into 3 groups: lethal total body irradiation (TBI); allogeneic bone marrow transplantation (BMT) and recipients of holafugine ( HF). The symptoms of aGVHD were observed daily and detailed histopathologic analyses of recipient skin were performed at Day 6 post-transplantation.And Tri-color flow cytometry ( FCM) was performed at Day 6 post-transplantation to measure the levels of interleukin (IL)-17, interferon (IFN)-γ and TH1/TH17. Results Clinical GVHD symptoms were observed in recipient mice. The administration of HF to lethally irradiated recipients led to very modest GVHD-induced cutaneous changes manifested predominantly by fur loss. However, the experimental animals receiving only allogeneic BMT showed significant fur loss and pathologic skin conditions. Consistent with the clinical evaluations, the histopathologic results demonstrated significantly increased pathologic cutaneous lesions in recipients undergoing only BMT. The median ratios of TH1/TH17 cells were 17. 57 and 5. 31 in the HF and BMT groups respectively. The difference had statistical significance (P < 0. 05). The serum levels of IL-17 were(1.47 ±0. 18)and(2. 81 ±0. 19)pg/ml in the TBI and BMT groups respectively(P <0. 05). But IL-17 could not be detected in the HF group. The serum levels of IFN-γ were (3. 86 ±0. 32) ,(42. 97 ± 0. 42) and (9. 89 ± 0. 51) pg/ml in the TBI, BMT and HF groups respectively. The inter-group differences had statistical significance (P <0. 05). Conclusion An absence of TH17 cell may alleviate the cutaneous GVHD but exacerbate the systemic GVHD.     

Effects of long-term administration of low-dose FTY720 on survival of murine cardiac allograft

This study examined the effect of long-term administration of low-dose FTY720 on survival of murine cardiac allograft and the possible mechanism. Murine models of abdominal heterotopic heart transplantation were established. Low-dose FTY720 (0.3 mg/kg) was administrated to the animals 4 days before the transplantation of cardiac allografts until the occurrence of rejection or the observation terminals. The animals without FTY720 treatment and those with syngeneic cardiac grafts transplanted served as controls. The mean survival time (MST) of grafts, and T lymphocyte subsets in grafts, periph-eral blood and lymphoid organs were measured by histopathological examination or flow cytometry, and compared among groups. The results showed that the MST of allografts in FTY720-treated mice was more than 40 days, significantly longer than that in the untreated group (MST=8 days, P<0.01). After the long-term administration of FTY720, the proportion of CD4+ and CD8+ lymphocytes in pe-ripheral blood was diminished significantly, but the proportion of CD4+ lymphocytes was increased in mesenteric lymph nodes (MLNs) and spleen. Immunofluorescence staining revealed that the infiltration of CD4+ and CD8+ lymphocytes in allografts was significantly inhibited after long-term administration of low-dose FTY720. It was concluded that low-dose long-term administration of FTY720 could pro-mote T lymphocytes in lymphatic organs and decrease their infiltration in allografts, resulting in the in-hibition of rejection and the long-term survival of allografts.     

FTY720对百草枯中毒小鼠肺损伤的保护作用

目的:探讨新型免疫抑制剂FTY720对急性百草枯(PQ)中毒小鼠肺损伤的保护机制。方法:8周龄C57BL/6j小鼠随机分为百草枯40mg/kg染毒对照组(PQ组)、百草枯40mg/kg染毒+FTY720 0.5mg/kg/d干预组(PQ+FTY720组)、生理盐水对照组(Control组)。PQ组和PQ+FTY720组分别经腹腔注射PQ40mg/kg,PQ+FTY720组于2小时后腹腔注射FTY720 0.5mg/kg,每日一次,连续14天;Control组腹腔注射等量生理盐水。各组分别于第3、第28天处死,观察28天生存率,留取支气管肺泡灌洗液(BALF)及肺组织。HE染色观察各组肺组织病理变化。BCA法测定BALF中总蛋白含量。ELISA法检测BALF中IL-6水平。Real-time PCR法测定肺组织中alpha-smooth muscle actin(α-SMA)、type I col1agen、type Ⅲcollagen mRNA的表达水平,免疫组化法观察肺组织中α-SMA的表达。结果:28天生存率PQ组与PQ+FTY720组分别为50%和70%。 HE染色显示FTY720干预后小鼠肺损伤程度较PQ组减轻。PQ+FTY720组在急性期BALF总蛋白含量、IL-6水平明显低于PQ组,慢性期肺组织中α-SMA、type I col1agen、type Ⅲcollagen在mRNA水平上表达较PQ组下调,且均具统计学意义。肺组织α-SMA阳性表达颗粒较PQ组减少。结论: FTY720可抑制部分肺损伤/纤维化相关因子的表达,对百草枯中毒肺损伤有治疗保护作用。