Effects of Ligustrazine on Expression of Bone Marrow Heparan Sulfates in Syngeneic Bone Marrow Transplantation Mice

Hematopoietic reconstitution after bone marrowtransplantation (BMT ) is a complicated process,which concerns notonly with the quality and quanti-ty of hematopoietic stem cells(HSC) ,but also withthe recovery of hematopoietic inducing microenviron-ment (HIM) .The contactof HSC and HIM,whichregulates the proliferation,differentiation and self-maintenance of HSC,is critical for hematopoietic re-constitution.Heparan sulfates (HS) are a kind of gly-cosaminoglycans macromolecules composed of…     

Enhancing Effects of Ligustrazine on Expression of CD31 and Hematopoietic Reconstitution in Syngenic Bone Marrow Transplantation of Mice

Summary： The effect of ligustrazine on the expression of CD31 in syngenic bone marrow transplantation （BMT） mice was studied. Fifty-six Balb/c mice were divided into 3 groups： normal control group. BMT control group, and ligustrazine treated group. Syngenic BMT mouse models were established according to the literatures. In BMT control group and the ligustrazine treated group, the mice were given respecxively orally 0.2 mL saline and 2 mg ligustrazine twice a day. On the 7th, 14th, and 21st day after BMT, the mice were killed. The expression of CD31 on the surface of bone marrow nuclear cells （BMNC） was detected by flow cytometry. Peripheral blood leukocytes, platelets and BMNC were counted. Histological observation of bone marrow was made. The results showed thai in ligustrazine treated group the peripheral blood leukocylcs, platelets and BMNC counts, and the expression of CD31 on the day 7, 14, 21 after BMT were higher than in BMTcontrol group （P〈0.01 or P〈0.05）. In conclusion, ligustrazine could obviously enhance the CD31 expression on the surface of BMNC after syngcnic BMT in mice, which may be one of the mecha- nisms underlying the ligustrazine accelerating hematopoietic reconstitution in syngenic BMT.     

肝细胞生长因子对骨髓移植小鼠造血重建的促进作用

目的 观察肝细胞生长因子（HGF）对骨髓移植小鼠造血重建的促进作用。方法 用BALB／c小鼠建立同基因骨髓移植模型，随机将BALB／c小鼠分A、B两组，B组移植后叶7d皮下注射HGF，A组皮下注射PBS，移植后1、7、14、21和28d检测小鼠血红蛋白、白细胞、血小板、骨髓单个核细胞及骨髓造血细胞面积。结果 B组血红蛋白在移植后7、14d与同期A组无差异（P〉0．05），B组血红蛋白在移植后21d高于同期A组（P〈0．05）；B组血小板在移植后7、14及21d高于同期A组（P值分别小于0．01、0．01和0．05）：B组白细胞在移植后7、14、21及28d高于同期A组（P值分别小于0．01、0．01、0．05及0．05）；B组骨髓单个核细胞及骨髓造血细胞面积在移植后7、14、21和28d高于同期A组（P值分别小于0．05、0．01、0．01及0．05）。结论 HGF对骨髓移植小鼠造血重建有一定的促进作用。     

Effect of ligustrazine on hematopoiesis in bone marrow transplantation mice

Hematopoietic reconstitution after bone marrowtransplantation is a complicated process,whichconcerns with the quality and quantity ofhematopoietic stem cell (HSC) ,and also with therecovery of hematopoietic inducing microenvironment(HIM) .The contact of HSC and HIM is critical forhematopoietic reconstitution.1　MATERIALSAND METHODS1 .1　Materials1 .1 .1　 Animals　　 BALB/c mice of both sexes(clean class,close copulatory series (H- 2 d,MLSb) )were 8— 1 0 weeks old,and 1 7— 2 …     

骨髓干细胞移植后mdx鼠抗肌萎缩蛋白的表达

目的研究骨髓干细胞移植治疗Duchenne型肌营养不良鼠(mdx鼠)后抗肌萎缩蛋白表达的动态变化。方法取6~8周龄C57BL/6鼠骨髓干细胞,以2×107个/只干细胞尾静脉移植到经7Gyγ射线预处理后的7~9周龄mdx鼠体内,分别于移植后5、8、12、16周应用免疫荧光、逆转录-聚合酶链反应、Western blot检测腓肠肌抗肌萎缩蛋白的表达情况。结果经γ射线放疗预处理的mdx鼠,在尾静脉移植骨髓干细胞5周后可见抗肌萎缩蛋白少量表达,随移植时间延长表达逐渐增多,16周时肌纤维抗肌萎缩蛋白的表达为12%,并可见肌细胞核中移现象较同龄mdx鼠对照组减少,边居增多。结论同种异体骨髓干细胞移植治疗mdx鼠后,骨髓干细胞通过血液循环趋向病变肌组织,并分化为肌细胞表达抗肌萎缩蛋白。随移植时间延长表达增多,提示骨髓干细胞移植可长久持续参与受损骨骼肌的修复与再生。     

Study on the effect of ligustrazine on hematopoietic reconstitution in bone marrow transplantation mice

Hematopoietic reconstitution after bone marrowtransplantation is a complicated process CXCchemokines and their receptors play important rolesin this process. One of these receptors, CXCR4 expresses on hematopoietic stem cells, and is now takenas a "homing factor" in hematopoietic reconstitu--'tionll' 2]. Ligustrazine can protect the bone marrowmicroenvironment after irradiation or immune damage[', 4]. We established allogenic bone marrow transplantation mice models and explored the effects …     

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。     

血小板第4因子对同基因骨髓移植小鼠造血重建的影响

为探讨血小板第4因子(PF4)对同基因骨髓移植(BMT)小鼠造血重建的影响及其可能机制,分别于照射前20、26 h给小鼠腹腔注射PF4 20μg/kg共2次,然后建立同基因骨髓移植小鼠模型,分别于BMT后+7、+14 d计数外周血细胞、骨髓单个核细胞(BMNC)以及脾集落形成单位(CFU-S),检测趋化因子受体CXCR4的表达.PF4处理组小鼠+7、+1 4 d BMNC以及+7 d CFU-S计数显著高于BMT组小鼠(P＜0.01或P＜0.05);BMT后+14d,PF4组小鼠CXCR4的表达显著高于BMT组(P＜0.05).表明PF4可以加速BMT后的造血重建过程,促进CXCR4的表达可能是其机制之一.     

Effects of Ligustrazine on Hematopoiesis in the Early Phase of Bone Marrow Transplantation Mice

the National NaturalScience Foundation(Serial No.3 9870 92 6)sacrificed on the1 st,3rd,5 th,7th,1 0 th,1 5 th,and 2 1 st day after BMT.The marrow from fe-murs was flushed into RPMI- 1 640 medium. BM-NCs from each femur were counted.1 .2 .7Carbon- Mold Preparation of Blood Vessels　　 A colloidal carbon solution( prepared Chineseink,0 .2 ml,2 0 mg/ml in saline) was injected1 -2 min before sacrifice on the1 st,3rd,5 th,7th,1 0 th,1 5 th,2 1 st day after BMT.One femur ofeach mouse was r…     

Effects of adenovirus mediated vascular endothelial growth factor gene transfer on reconstitution of hematopoiesis in post-bone marrow transplantation mice

Background Bone marrow transplantation (BMT) conditioning procedure is considered as the cause of damage to bone marrow microvasculature and the delay of hematopoiesis recovery. However, hematopoiesis regulation post BMT by vascular endothelial growth factor (VEGF) has not yet been studied. In this study, adenovirus were used to investigate the effects of VEGF gene transfer on preventing damages to bone marrow microenvironment and its promotion of hematopoiesis in post-BMT mice.Methods Recombinant adenovirus (Ad)-enhanced green fluorescent protein (EGFP)/hVEGF165 was injected via tail vein into BALB/c mice undergoing syngeneic BMT. During the different phases post BMT, the distribution of adenovirus and the plasma levels of hVEGF were measured as well as the numbers of white blood cells (WBC), platelet (PLT) and red blood cells (RBC) in peripheral blood. At the same time, the mice were injected with Chinese ink via tail vein, following which the tibias were separated and were used for analysis of bone marrow microvasculature surface area and cellularity.Results Significant expression of EGFP and hVEGF was observed in multiple organs at different phases post BMT, and the plasma level of hVEGF was up to (866.67±97.13) pg/ml. The recovery of WBC, PLT and RBC of the group treated with recombinant adenovirus Ad-EGFP/hVEGF165 were significantly more rapid than those of other BMT groups （P&lt;0.05, respectively）. At the 20th day post BMT, the percentage of bone marrow microvasculature surface area in group treated with VEGF ［(61.2±4.0)%］ returned to normal level ［(62.0±5.0)%, P&gt;0.05］. The restoration of hematopoiesis was retarded more than that of microvasculature. The cellularity of bone marrow in each group was still lower than that of normal control ［(62.3±4.0)%, P&lt;0.05］ at the 30th day post BMT, but the percentage in group treated with VEGF at the 20th and 30th days post BMT ［(46.5±5.0)% and (55.1±4.5)%］ exceeded those of other BMT groups (P&lt;0.05, respectively).Conclusion VEGF gene transfer mediated by adenovirus may protect the hematopoietic microenvironment to promote the restoration of hematopoiesis in post-BMT mice.     

单倍相合异基因骨髓移植前后骨髓像分析

目的观察单倍相合异基因骨髓移植前后骨髓像、造血能否重建。方法对22例患者造血干细胞移植前骨髓穿刺抽吸涂片镜检;骨髓移植成功出层流室首次骨髓穿刺抽吸涂片;每月定期复查骨髓穿刺抽吸涂片镜栓。同时观察外周血涂片和骨髓涂片2张或2张以上。结果22例患者单倍相合并基因造血干细胞移植后骨髓像全部达到正常,完全缓解。移植成功随访,其中6例患者随访中骨髓细胞出现少许病态造血现象,2例出现纯红细胞性再生障碍性贫血（PRCA）,余14例骨髓正常。结论单倍相合并基因骨髓移植能够使白血病和重型再障骨髓重建。造血重建恢复期可能继发骨髓增生异常综合症（MDS）和PRCA。     

Effect of Ligustrazine on the Expression of LFA-1, ICAM-1 Following Bone Marrow Transplantation in Mice

Asablood activatingChinesedrug ,ligustrazinecanimprovethemicroenvironmentofbonemarrow ,increasetheadherentfunctionofstromalcellsandpromotethegrowthofhematopoieticcells[1] .Toin terprettherolesthatligustrazineplaysinbonemar rowhematopoieticreconstitutionan…     

小鼠脾细胞在混合骨髓移植中的作用

目的:观察同异基因小鼠脾细胞输注比例和输注量在混合半相合骨髓移植中对移植物抗宿主病(GVHD)和嵌合状态的影响.方法:受鼠全身照射后,经尾静脉回输同基因和半相合异基因供鼠骨髓及两者不同比例脾细胞.A组未加入异基因供鼠脾细胞,B组加入的同异基因供鼠脾细胞量相等,C组加入异基因供鼠脾细胞量是同基因供鼠脾细胞量的两倍.回输后观察GVHD临床表现,取肝、皮肤、小肠送病理学检查评定GVHD分级.测定 70天、 150天嵌合率.监测造血重建和受鼠生存率.结果:混合骨髓移植各组小鼠生存期均超过50天.其中A组,临床和病理无GVHD表现, 150天嵌合水平大幅度下降.B组,临床和病理无明显GVHD表现, 150天嵌合水平中等幅度下降.C组病理有GVHDⅡ级表现, 150天嵌合水平下降不明显.半相合骨髓移植对照组小鼠出现典型的GVHD临床和病理表现,仅20%小鼠生存期超过50天.结论:小鼠混合半相合骨髓移植中,同异基因小鼠脾细胞输注比例和输注量对移植排斥和GVHD有协调作用.     

血小板因子4对环磷酰胺损伤小鼠骨髓细胞增殖能力的影响

目的 探讨血小板因子4(PF4)对环磷酰胺(CTX)诱导的化学损伤小鼠骨髓细胞增殖能力的影响. 方法 36只小鼠随机分为PF4组、CTX组和对照组.小鼠腹腔注射CTX 200 mg/kg制备化学药物损伤小鼠模型.在注射CTX前26 h和20 h,PF4组和CTX组各注射PF4(40μg/kg)和等体积的PBS缓冲液,对照组只注射等体积的PBS缓冲液.通过体外双层琼脂培养法测定PF4对CTX损伤小鼠骨髓细胞粒一巨噬细胞集落(CFU-GM)形成的影响,流式细胞仪测定细胞周期的变化.结果 注射CTX后第5天CTX组骨髓细胞数(BMK)较对照组显著减少(P<0.01);第8天PF4组BMK较CTX组显著增多(P<0.01).注射CTX后第5天CTX组骨髓CFU-GM数量较对照组和PF4组显著减少(P<0.01);第8天PF4组与CTX组的骨髓CFU-GM数量均有不同程度的上升,但两组间差异仍有统计学意义(P<0.05).注射CTX后第5天PF4组骨髓有核细胞GO/G1期细胞百分率比CTX组显著提高,且细胞坏死、凋亡率显著下降(P<0.01),而与对照组相比差异无统计学意义(P>0.05).第8天各组间G0/G1期细胞百分率、细胞坏死/凋亡率差异均无统计学意义(P>0.05). 结论 PF4使处于G0/G1期的骨髓细胞数增多,降低了其坏死和凋亡率,增加骨髓细胞CFU-GM集落形成率.     

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.     

异体骨髓干细胞移植后假肥大型肌营养不良症模型鼠膈肌dystrophin表达及病理改变

目的探讨骨髓干细胞移植对假肥大型肌营养不良症(DMD)模型鼠-mdx鼠膈肌的治疗效果。方法取雄性SD大鼠骨髓干细胞经尾静脉植入放疗处理后的8周龄雌性mdx鼠(n=18)。于移植后4、8、12周各取6只mdx鼠的膈肌行 HE染色、抗肌萎缩蛋白(dystrophin)免疫荧光检测以及dystrophin mRNA的RT-PCR分析,同时用正常C57鼠及放疗而未移植的mdx鼠作为对照,另进行PCR反应检测实验鼠膈肌内Sry(Y染色体的性别决定区)基因。结果移植后mdx 鼠膈肌间质内炎性细胞浸润较放疗而未移植mdx鼠有所减少;移植后核中心移位纤维比例[移植后4、8、12周分别为 (15．58±0．91)％、(12．50±1．87)％、(10．17％±1．17)％]较未移植mdx鼠(19．5±1．87)％显著减少。移植后dystrophin免疫荧光阳性细胞比例[移植后4、8、12周分别为(1．00±0．32)％、(6．00±1．05)％、(11．92±1．11％)]较未移植mdx鼠(O．17±0．41)％显著增加。RT-PCR结果显示C57鼠膈肌的dystrophin mRNA相对含量(0．63±0．04)最高;而未移植mdx鼠的膈肌中未检测到dystrophin mRNA;移植后的mdx鼠膈肌中mRNA的表达水平(移植后4、8、12周分别为0．19±0．05、0．26±0.06、0．36± 0．04)随时间推移逐渐增高;移植后各时间点mdx鼠膈肌Sry基因均为阳性。结论骨髓干细胞系统移植mdx鼠可以恢复部分膈肌的dystrophin表达,改善膈肌的病理,骨髓干细胞移植有希望成为全身治疗DMD的有效方法。     

小鼠不同细胞数骨髓移植后回巢与生存率观察

目的:探讨移植细胞数对小鼠骨髓移植回巢及生存率的影响。方法:采用不同细胞数移植、不同细胞数加不同时问间隔程序移植及流式细胞仪检测等方法,观察不同细胞数骨髓移植后小鼠骨髓内回巢效果和生存情况。结果:不同细胞数骨髓移植中5×105组的回巢细胞数与高细胞数组无差异,回巢率远远高于后者(P<0.01);生存率1×105组为95％,与高细胞数组无差异。不同细胞数加不同时间间隔程序移植以细胞总数在1×105、间隔24 h移植组存活率高,组间差异非常显著(P<0.01)。结论:在一定细胞数量范围内,分次移植可能提高移植干细胞的回巢效率。     

非清髓性骨髓移植对移植物抗宿主病的影响

目的 探讨非清髓性预处理方案对大鼠异基因骨髓移值模型移植物抗宿主病(GVHD)的影响。方法 Wistar大鼠为供鼠,SD大鼠为受鼠,受鼠分两组,分别予以不同的预处理方案,A组受鼠自移植前4-1d,腹腔注射氟达拉宾(1mg/kg),移植前4h接受2Gy的全身照射;B组受鼠仅在移植当天分别接受6Gy的全身照射。各实验组受鼠均在移植当天经尾静脉输注供鼠骨髓细胞3.8×107;对照组大鼠输注等量的生理盐水,观察各组大鼠GVHD反应。结果 移植前接受氟达拉宾和2Gy全身照射预处理的大鼠生存期较长,且GVHD临床和病理评分也较低。结论 非清髓性骨髓移植预处理方案可有效预防GVHD,拓宽临床进行异基因骨髓移植的适用范围。     

肠坏死是异种骨髓移植后SCID鼠发生急性排斥反应死亡的主要原因

目的 通过骨髓移植后观察严重联合免疫缺陷（SCID）鼠的生存状况，探讨免疫排斥反应中导致动物死亡的主要原因。方法 培养去原基胸腺细胞的引产儿胸腺小叶，将其与分离的银屑病患者骨髓单一核细胞共移植于经60Coγ射线照射的SCID鼠，移植后第2天开始每鼠皮下注射重组人粒细胞集落刺激因子（rhG-CSF），观察动物生存状况。实验共使用了10只SCID鼠。结果 7例SCID鼠于移植后第7天死亡，进行尸解发现全部为肠坏死。2例于移植后经短暂反应后逐渐恢复正常，一直存活至今（移植后第35天）。1例对照鼠无任何异常反应，存活至今。结论 肠坏死是异种骨髓移植后SCID鼠发生急性排斥反应死亡的主要原因。     

骨髓移植患者的HLA分型研究

目的 探讨ＨＬＡ位点与移植物存活的关系。方法 应用微量淋巴细胞试验方法对３例骨髓移植患者及其家庭成员共１２例血液进行了ＨＬＡ分型研究，结果 １例患者两条ＨＬＡ单倍型与其家族成员供者相同，并移植成功；另一例患者的ＨＬＡ型仅其供者Ａ、Ｂ两位点相同，ＤＲ位点可疑最终移植失败，第３例患者ＨＬＡ－Ａ、Ｂ、ＤＲ位民其供者无任何相同，未行移植。结论 在骨髓移植中，供、受者双方ＨＬＡ单倍型完全相同，特别是ＤＲ位点