黄芪皂甙Ⅳ联合脂肪源性干细胞对老龄大鼠脑缺血再灌注中神经营养因子表达的影响

目的 探讨黄芪皂甙Ⅳ联合脂肪源性干细胞( ADSC)移植对老龄大鼠脑缺血再灌注模型神经功能及神经营养因子表达的影响.方法 老龄大鼠随机分为模型组、ADSC组、黄芪皂甙Ⅳ组、黄芪+ADSC组,应用PKH-26标记移植的ADSC,NSS法检测神经功能恢复,免疫组化法和实时荧光定量PCR检测基质细胞衍生因子-1(SDF-1)、脑源性神经营养因子(BDNF)、胰岛素样生长因子-1(IGF-1)蛋白和mRNA的表达.结果 治疗组均能使大鼠神经功能损害评分降低,以联合组恢复最为明显(P＜0.05).黄芪皂甙Ⅳ组SDF-1蛋白和mRNA表达较模型组显著增加(P＜0.05).联合组PKH-26标记的细胞个数高于ADSC组(P＜0.05).联合组和ADSC组BDNF和IGF-1蛋白和mRNA表达高于模型组,其中联合组表达最为显著(p＜0.05).结论 联合组促进脑缺血再灌注老龄大鼠神经营养因子表达和神经功能恢复作用优于单独治疗组,其机制可能与黄芪皂甙Ⅳ上调脑缺血区SDF-1表达,促进移植的ADSC迁移和存活有关.     

二氯化钴预处理对脑梗死大鼠基质细胞衍生因子1α和趋化因子受体4表达的影响

目的探讨缺氧缺血环境中基质细胞衍生因子(SDF-1α)和趋化因子受体4(CXCR4)生物轴对脑保护作用及二氯化钴预处理的影响。方法选择成年雄性SD大鼠168只,随机分为干预组84只,模型组84只。按缺血后再灌注时间分为再灌注6、24h、3、5、7、14d6个时间点。采用改良Longa方法制备局灶性脑缺血模型,通过免疫组织化学法及Western blot法观察2组大鼠脑组织皮质区脑缺血后再灌注各个时间点SDF-1α及CXCR4表达变化。结果干预组于脑缺血再灌注6h皮质区SDF-1α、CXCR4阳性细胞表达明显增加,7d表达至最高水平,随后表达逐渐减少,14d仍有阳性表达。干预组各时间点皮质区SDF-1α、CXCR4阳性细胞表达明显高于模型组(P0.05)。模型组及干预组大鼠皮质区脑组织6h时SDF-1α、CXCR4蛋白表达即明显增加,7d达高峰,14d仍有阳性表达;干预组各时间点CXCR4蛋白表达明显高于模型组,差异有统计学意义(P0.05)。结论二氯化钴可能通过上调SDF-1α、CXCR4表达水平,诱导脑缺氧耐受,促进间充质干细胞向缺血组织迁移,发挥脑保护作用。     

电针百会、风府穴对脑I/R损伤大鼠海马区CPG15表达影响的实验研究

目的 探讨电针对局灶性脑缺血再灌注大鼠神经功能的恢复及海马区神经可塑性相关基因15(CPG15)表达影响的情况.方法 60只SD大鼠,雌雄各半,随机分为正常对照组、模型组、电针经穴组、电针非经穴组、西药对照组.采用线栓法制备局灶性脑缺血再灌注模型,电针经穴组电针百会、风府穴,电针非经穴组电针大鼠臀部非经非穴位置,电针以疏波2Hz,强度3 mA～5mA,持续电针30 min,每天1次,连续治疗2周.西药对照组以尼莫地平20 mg/(kg·d)灌胃,每日2次,连续灌胃2周.2周后Longa5分法对大鼠神经功能缺损评分,并取材,运用免疫组化法检测大鼠缺血侧海马区CPG15表达情况.结果 模型组大鼠神经功能缺损评分及缺血侧海马区CPG15表达显著高于正常对照组(P＜0.01);电针经穴组与西药治疗组大鼠神经功能评分及海马区CPG15表达差异无统计学意义(P＞0.05),而与模型组比较,电针经穴组与西药治疗组神经功能评分及海马区CPG15表达均有统计学意义(P＜0.01);电针非经穴组大鼠神经功能缺损评分及缺血侧海马区CPG15表达与模型组比较差异无统计学意义(P＞0.05).结论 电针可改善脑缺血再灌注大鼠神经功能,并提高海马区CPG15的表达,电针对脑缺血再灌注后脑细胞的神经可塑性有促进作用.     

二氯化钴预处理对大鼠脑梗死体积及SDF-1α/CXCR4表达水平的影响

目的观察二氯化钴预处理对大鼠脑梗死体积百分比、基质细胞衍生因子1α(SDF-1α)/趋化因子受体4(CXCR4)表达的影响;探讨缺氧缺血环境中SDF-1α/CXCR4生物轴对脑的保护作用。方法将168只成年雄性SD大鼠随机分为缺氧预处理组(n=84)、模型组(n=84)。按缺血后再灌注时间不同分为再灌注6 h、24 h、3天、5天、7天、14天6个亚组。采用改良Longa方法制备局灶性脑缺血模型。观察缺氧缺血后脑组织病理学改变,通过氯化三苯基四氮唑(TTC)染色法观察脑缺血后再灌注两组大鼠脑梗死体积百分比的变化;免疫组织化学法检测大鼠脑组织皮层区各个时间点SDF-1α及CXCR4的表达变化。结果 TTC染色显示,缺氧预处理组及模型组大鼠6 h时即可见明显梗死灶,24 h脑梗死体积趋于稳定。两组大鼠24 h、3天、5天、7天、14天5个时间点脑梗死体积百分比比较,差异无统计学意义(P0.05);缺氧预处理组各时间点脑梗死体积百分比均明显小于模型组(P0.05)。免疫组织化学法结果显示,两组于脑缺血再灌注6 h皮层区SDF-1α、CXCR4表达明显增加,7天表达至高峰,随后表达逐渐减少,14天仍有表达。缺氧预处理组各时间点皮层区SDF-1α、CXCR4的阳性细胞数均显著多于模型组(P0.05)。结论二氯化钴预处理能缩小脑梗死体积,其可能通过上调SDF-1α、CXCR4的表达水平,诱导脑缺氧耐受,促进间充质干细胞向缺血组织迁移,发挥脑保护作用。     

电针对局灶脑缺血再灌注大鼠大脑皮质胎盘生长因子及其受体Flt-1 mRNA和蛋白表达的影响及其促进脑内血管再生的作用

目的 观察电针对SD大鼠局灶脑缺血再灌注后大脑皮质胎盘生长因子及其受体Flt-1 mRNA和蛋白表达的影响,探讨电针促进局灶脑缺血再灌注后脑内血管再生的可能机制.方法 采用线栓法制备SD大鼠局灶脑缺血再灌注模型.将SD大鼠随机分为正常对照组、模型组、电针组,将模型组和电针组分为局灶脑缺血2 h再灌注1、3和7天三个亚组.取双侧"合谷"穴为电针刺激穴位.采用免疫组织化学法检测胎盘生长因子及其受体Flt-1蛋白在缺血区大脑皮质的分布及其表达;逆转录聚合酶链反应检测缺血区大脑皮质胎盘生长因子mRNA和Flt-1 mRNA的表达;免疫印迹法定量检测缺血区侧大脑皮质胎盘生长因子蛋白的表达.结果 与正常对照组比较,模型组和电针组各时间点缺血区大脑皮质胎盘生长因子mRNA和蛋白及Flt-1 mRNA和蛋白的表达增加(P<0.05);电针组各时间点缺血区大脑皮质胎盘生长因子mRNA和蛋白及Flt-1 mRNA和蛋白的表达比模型组增加更为明显(P<0.05).结论 电针上调了局灶脑缺血再灌注大鼠缺血区大脑皮质的胎盘生长因子和Flt-1的表达,胎盘生长因子/Flt-1可能促进脑缺血后脑内血管再生.     

缺血后处理对缺血再灌注大鼠脑排斥导向分子A表达及轴突损伤的影响

目的探讨实施缺血后处理对大鼠中枢神经缺血损伤及修复的影响。方法 SD大鼠随机分为假手术组、缺血/再灌注组、缺血后处理组。对各组大鼠术后进行神经功能评分,脑梗死体积测量,反转录聚合酶链反应检测大鼠缺血侧皮质及海马排斥导向分子A(RGMa)mRNA表达,免疫组织化学法检测皮质及海马RGMa及轴突神经丝蛋白(NF-200)表达。结果缺血后处理组较缺血/再灌注组12、24 h、2 d神经功能缺失改善(P<0.05)。缺血后处理组梗死体积明显减少(P<0.01),其缺血侧皮质和海马RGMa mRNA及蛋白表达相比缺血/再灌注组降低(P<0.05)。缺血后处理组缺血皮质和海马NF-200光密度值比缺血/再灌注组升高(P<0.01)。结论缺血后处理能改善大鼠缺血性脑损伤导致的神经功能障碍,使轴突的损伤减少,起到神经保护作用;其作用的机制可能与减少排斥性轴突导向因子RGMa的表达相关。     

电针治疗对大鼠脑缺血再灌注中Annexin A1表达的影响

目的探讨电针治疗在大鼠脑缺血再灌注损伤中对Annexin A1表达的影响。方法 SD大鼠32只,随机分为对照组、缺血再灌注组、电针组、假手术组,每组8只。检测Annexin A1在大鼠脑组织中的表达变化及AnnexinA1转位结果。结果大鼠大脑中动脉栓塞60 min再灌注24h后,可出现明显的神经缺损性行为,在脑缺血前后以电针治疗可明显改善大脑损伤性行为异常。在海马CA1、CA2、CA3、齿状回、皮质区,缺血再灌注组大鼠Annex-in A1的表达较对照组明显升高,电针组大鼠Annexin A1的表达较缺血再灌注组明显下降(P<0.05)。在海马CA1、CA3、皮质区,缺血再灌注组大鼠Annexin A1核转位和膜转位较对照组明显上升;在海马CA1、CA3区,电针组大鼠Annexin A1核转位和膜转位较缺血再灌注组明显下降(P<0.05)。结论脑缺血再灌注后Annexin A1表达上调,Annexin A1出现核转位和膜转位现象,电针可以逆转Annexin A1表达和转位。电针有可能通过调节Annexin A1的核转位和膜转位来改变神经元的凋亡。     

白蛋白治疗对大鼠脑缺血后海马血管内皮生长因子及其受体1的影响

目的探讨人血白蛋白治疗对大鼠脑缺血早期海马血管内皮生长因子(VEGF)及fam样酪氨酸激酶受体1(flt-1)表达的影响。方法雄性SD大鼠40只,采用大脑中动脉线栓法制备大鼠局灶性脑缺血再灌注模型,随机分为假手术组10只、生理盐水组15只和白蛋白组15只。采用溴甲酚绿法测定血清白蛋白水平,RT-PCR检测脑缺血再灌注后6、24、48h大鼠海马VEGF和flt-1mRNA表达,免疫组织化学和Western blot法检测脑缺血再灌注24h海马VEGF蛋白表达。结果与生理盐水组比较,白蛋白组大鼠神经功能缺损评分于脑缺血再灌注后24、48h明显降低(P<0.05),海马VEGF mRNA和flt-1mRNA表达于脑缺血再灌注后6、24h明显降低(P<0.05,P<0.01),海马VEGF蛋白表达于脑缺血再灌注后24h明显降低(P<0.05)。结论白蛋白治疗可下调脑缺血早期海马VEGF和flt-1mRNA表达,降低海马VEGF蛋白表达水平,改善神经功能缺损。     

CXC趋化因子受体3和其配体在大鼠肝脏缺血再灌注损伤中的表达

目的:探讨CXC趋化因子受体3(CXCR3)及其配体IP-10和Mig在大鼠肝脏缺血/再灌注(I/R)损伤中的表达及作用.方法:32只Wistar大鼠随机分成4组,每组8只.即假手术组,部分肝脏缺血再灌注6,12和24 h组.应用酶联免疫吸附实验(ELISA)法检测肝组织肿瘤坏死因子(TNF)-α水平.应用半定量聚合酶链式反应(RT-PCR)法测定肝组织CXCR3及其配体IP-10,Mig mRNA的表达.同时检测血清丙氨酸转氨酶(ALT)及天冬氨酸转氨酶(AST)的含量.结果:假手术组肝组织中CXCR3,IP-10,Mig mRNA低表达.缺血再灌注各组肝组织中CXCR3和IP-10 mRNA表达水平均明显高于假手术组(CXCR3:0.925±0.109,0.786±0.074,0.606±0.082 vs 0.125±0.028,均P<0.01;IP-10:0.863±0.091,0.680±0.075,0.543±0.284 vs 0.128±0.027,均P<0.01),6 h组高于12 h组(P<0.01),12h组与24h组无明显差别(P>0.05).Mig mRNA水平较假手术组相比,无显著差异(P>0.05).缺血再灌注各组TNF-α水平较假手术组明显升高(154.88±14-35 ng/L,258.88±13.73 ng/L,182.87±10.95 ng/L vs 23.63±4.00 ng/L,均P<0.01),再灌注12 h达高峰.结论:CXCR3及其配体IP-10在肝脏缺血再灌注早期表达上调,在缺血再灌注损伤中起重要的作用.     

黄芪皂甙Ⅳ联合BMSCs移植对大鼠脑缺血再灌注海马神经细胞凋亡及Livin、Caspase-9蛋白的影响

目的 探讨黄芪皂甙Ⅳ联合骨髓间充质干细胞(BMSCs)移植对缺血再灌注大鼠海马Livin、Caspase-9蛋白及神经细胞凋亡表达的影响.方法 实验动物随机分为假手术组、模型组、BMSCs组、联合组.采用线栓法建立大鼠大脑中动脉缺血(MCAO)模型,假手术组不予处理.BMSCs组于造模后3 h移植BMSCs.联合组既移植BMSCs,又给予黄芪皂甙Ⅳ治疗.观察缺血72 h后,各组神经功能评分,PKH-26标记的移植BMSCs分布,采用Western blot和免疫组化方法检测大鼠海马Livin、Caspase-9蛋白的表达,TUNEL方法检测细胞凋亡指数.结果 PKH-26标记的BMSCs在海马有表达.各组中联合组神经功能恢复最为明显(P<0.05).与模型组比较,联合组和BMSCs组均可上调Livin蛋白表达(P<0.05),下调Caspase-9蛋白表达(P<0.05),降低神经元细胞凋亡指教(P<0.05),以联合组作用更为显著(P<0.05).结论 黄芪皂甙Ⅳ联合BMsCs移植对脑缺血再灌注神经元细胞凋亡有协同抑制作用,其作用机制可能与黄芪皂甙Ⅳ促进BMSCs上调Livin蛋白表达,下调Caspase-9蛋白表达,抑制细胞凋亡有关.     

Over-expression of CXCR4 on mesenchymal stem cells augments myoangiogenesis in the infarcted myocardium

Bone marrow mesenchymal stem cells (MSCs) participate in myocardial repair following myocardial infarction. However, their in vivo reparative capability is limited due to lack of their survival in the infarcted myocardium. To overcome this limitation, we genetically engineered male rat MSCs overexpressing CXCR4 in order to maximize the effect of stromal cell-derived factor-1α (SDF-1α) for cell migration and regeneration. MSCs were isolated from adult male rats and cultured. Adenoviral transduction was carried out to over-express either CXCR4/green fluorescent protein (Ad-CXCR4/GFP) or Ad-null/GFP alone (control). Flow cytometry was used to identify and isolate GFP/CXCR4 over-expressing MSCs for transplantation. Female rats were assigned to one of four groups (n = 8 each) to receive GFP-transduced male MSCs (2 × 10⁶) via tail vein injection 3 days after ligation of the left anterior descending (LAD) coronary artery: GFP-transduced MSCs (Ad-null/GFP-MSCs, group 1) or MSCs over-expressing CXCR4/GFP (Ad-CXCR4/GFP-MSCs, group 2), or Ad-CXCR4/GFP-MSCs plus SDF-1α (50 ng/μl) (Ad-CXCR4/GFP-MSCs/SDF-1α, group 3), or Ad-miRNA targeting CXCR4 plus SDF-1α (Ad-miRNA/GFP-MSCs + SDF-1α treatment, group 4). Cardiodynamic data were obtained 4 weeks after induction of regional myocardial infarction (MI) using echocardiography after which hearts were harvested for immunohistochemical studies. The migration of GFP and Y-chromosome positive cells increased significantly in the peri- and infarct areas of groups 2 and 3 compared to control group (p < 0.05), or miRNA-CXCR4 group (p < 0.01). The number of CXCR4 positive cells in groups 2, 3 was intimately associated with angiogenesis and myogenesis. MSCs engraftment was blocked by pretreatment with miRNA (group 4). Cardiac function was significantly improved in rats receiving MSCs over-expressing CXCR4 alone or with SDF-1α. The up-regulation of matrix metalloproteinases (MMPs) by CXCR4 overexpressing MSCs perhaps facilitated their engraftment in the collagenous tissue of the infarcted area. CXCR4 over-expression led to enhance in vivo mobilization and engraftment of MSCs into ischemic area where these cells promoted neomyoangiogenesis and alleviated early signs of left ventricular remodeling.     

早期强化运动训练与电针治疗对脑缺血再灌注后大鼠神经功能恢复的作早期强化运动训练与电针治疗对脑缺血再灌注后大鼠神经功能恢复的作用

目的:观察早期强化运动训练与电针治疗对局部脑缺血大鼠再灌注后神经功能恢复的疗效并探讨其机制。方法:将65只健康成年SD雄性大鼠随机分为假手术组（5只）、大脑中动脉闭塞再灌注（MCAO）模型组(20只)、电针组(20只)、强化运动训练组（20只）。用线栓法建立右侧MCAO模型，电针组选取人中、百会穴给予电针刺激。强化运动训练组采用跑笼运动试验,观察4组大鼠的运动功能。选取24h、3d、7d、14d4个时间点，应用免疫组化方法检测脑缺血区皮层GAP-43及Neurocan阳性细胞的表达情况。采用神经症状评分评价造模情况及神经功能的恢复情况，TTC(2,3,5-三苯基氯化四氮唑)染色观察大鼠局部脑缺血再灌注后梗死体积的大小。结果: 2周后,强化运动训练组及电针组的神经症状评分明显低于模型组(P＜0.05),并高于假手术组(P<0.05)；强化运动训练组及电针组的脑梗死体积明显小于模型组(P<0.05)，电针组与运动训练组比较差异无统计学意义。结论:脑缺血再灌注后早期强化运动训练或电针刺激能够减小脑梗死体积,促进脑缺血大鼠神经功能的恢复。早期康复治疗可上调脑缺血区GAP-43表达与下调Neurocan表达，可能是其促进脑损伤区中枢神经修复的重要机制之一。     

电针对抑郁症大鼠海马CREB-BDNF受体后信号转导通路的作用

目的 探讨电针对抑郁症大鼠的治疗作用及海马cAMP反应元件结合蛋白(CREB)、脑源性神经营养因子(BDNF)的影响.方法 SD成年大鼠32只随机分为空白组、模型组、针刺组(合谷+太冲)和药物组(盐酸氟西汀),每组8只.空白组进行正常饲养,不给予其他处理;模型组大鼠采用长期不可预见性温和刺激造成抑郁症模型;电针组大鼠造模后电针合谷穴和太冲穴,每日1次,每次15 min,左右两侧穴位交替进行,持续21 d.药物组大鼠造模后进行氟西汀灌胃,每日1次,持续3 w.应激后第7、14、22天观察大鼠Open field行为学的变化,应激后22 d取材观察海马神经元形态结构、BDNF和CREB的表达.结果 (1)应激后第7、14、22天模型组大鼠水平运动及垂直运动较应激前有不同程度的减弱,针刺组于应激后第14天可有效地改善抑郁型大鼠模型的行为学表现,其效果优于药物组(P<0.05).(2)模型组海马齿状回神经元排列散乱,可见明显肿胀、固缩、变性、脱落等病理改变;针刺组和药物组大鼠海马神经元的病变程度较模型组有不同程度的减轻.(3)模型组大鼠BDNF阳性细胞数量较空白组明显减少(P<0.05),针刺组和药物组海马BDNF和CREB的表达较模型组上调(P<0.05).结论 针刺能改善抑郁症大鼠的行为学及海马神经元的病理变化,CREB-BDNF受体后信号转导通路是针刺抗抑郁的重要途径和作用靶点之一.     

外周血SDF-1/CXCR4轴与急性心肌梗死患者冠状动脉侧支循环形成的关系

目的 检测急性心肌梗死(acute myocardial infarction,AMI)患者外周血基质细胞衍生因子-1(stromalcell derived factor 1,SDF-1)、CXC趋化因子受体-4(C-X-C chemokine receptor 4,CXCR4)的浓度,探讨两者在AMI患者冠状动脉侧...     

Preconditioning enhances cell survival and differentiation of stem cells during transplantation in infarcted myocardium

AIMS: We hypothesized that preconditioning (PC) with stromal-derived factor 1 alpha (SDF-1) significantly enhances cell survival, proliferation, and engraftment of bone marrow-derived mesenchymal stem cells (MSCs) via SDF-1/CXCR4 signaling. METHODS AND RESULTS: MSCs were cultured and then incubated in medium for 60 min without SDF-1 (control group) or with SDF-1 0.05 microg/mL (SDF-1 group) or CXCR4-selective antagonist, AMD 3100 (AMD) (5 microg/mL, AMD group) or SDF-1 and AMD (0.05 microg/mL, 5 microg/mL, respectively, SDF-1+AMD group). MSCs were treated for 60 min, washed in normal medium, and then exposed to H2O2 (100 micromol/L) for 60 min to determine the effects of various treatments on cell injury, viability, and proliferation. For in vivo studies, rats were grouped (n = 6) after left anterior descending coronary artery ligation to receive 20 microL Dulbecco's modified Eagle's medium without cells or with 5 x 10(5) non-preconditioned MSCs (control group), SDF-1 preconditioned MSCs (SDF-1 group), AMD (AMD group), or MSCs treated with SDF-1 plus AMD (SDF-1+AMD group). Heart function, infarct size, fibrosis, and MSC proliferation and differentiation in infarcted myocardium were determined after 4 weeks. In vitro data showed a marked increase in cell viability and proliferation following SDF-1 PC. In vivo data in preconditioned group showed a robust cell proliferation, reduction in infarct size and fibrosis, and significant improvement in cardiac function. Effects of SDF-1 PC were abrogated by CXCR4 antagonist. CONCLUSION: We conclude that PC with the chemokine SDF-1 suppresses MSCs apoptosis, enhances their survival, engraftment, and vascular density, and improves myocardial function via SDF/CXCR4 signaling. Chemokine PC is a novel approach for enhancing stem cell survival and regeneration of infarcted myocardium.     

骨髓基质细胞移植对局灶性脑缺血大鼠内源性神经前体细胞增殖的动态影响

目的探讨骨髓基质细胞(bone marrow stromal cells,BMSCs)移植对局灶性脑缺血大鼠内源性神经前体细胞增殖的动态影响。方法建立大鼠大脑中动脉缺血再灌注模型。24只健康SD大鼠分为假手术组、缺血对照组、缺血移植组。大鼠局灶性脑缺血再灌流24 h后各组腹腔注射5-溴脱氧尿核苷(bromodeoxyuridine,BrdU)标记处于增殖状态的神经前体细胞,最后一次注射后2 h处死。通过神经缺损评分观察移植后神经功能改善情况。应用免疫组织化学染色及原位细胞凋亡检测脑组织BrdU阳性及凋亡细胞。结果脑梗死后侧脑室室管膜下区、海马齿状回区及梗死灶边缘BrdU阳性细胞迅速增多(P<0.05)。移植BMSCs后BrdU阳性细胞明显多于对照组(P<0.05),随移植时间的延长,增加更趋明显。同时,BMSCs移植后大鼠神经功能得到明显康复,缺血边缘区凋亡细胞明显减少伴梗死体积缩小。结论BMSCs可促进局灶性脑缺血大鼠内源性神经前体细胞的增殖、迁移,对脑缺血再灌注损伤起保护作用。     

基质细胞衍生因子-1及其受体对大鼠主动脉血管平滑肌细胞增殖与凋亡的影响

目的研究基质细胞衍生因子-1(SDF-1)对氧化低密度脂蛋白(ox-LDL)诱导的血管平滑肌细胞(VSMC)增殖与凋亡的影响。方法选用体外培养的大鼠主动脉VSMC,并分为正常对照组、ox-LDL组[动脉粥样硬化(AS)模型]、SDF-1组(AS模型+SDF-1)、SDF-1+12G5组(AS模型+SDF-1+CXCR4单克隆抗体)和12G5组(AS模型+CXCR4单克隆抗体),应用MTT法测VSMC细胞增殖,TUNEL法测细胞凋亡率,RT-PCR法测凋亡基因bax、bcl-2mRNA的表达。结果ox-LDL组的增殖率0.38±0.01,明显高于正常对照组0.28±0.02(P<0.01),明显低于SDF-1组0.44±0.02(P<0.01),与SDF-1+12G5组和12G5组比较差异无显著性(P>0.05);ox-LDL组的凋亡率24.2±2.4,高于正常对照组19.8±2.7和SDF-1组20.7±2.8(P<0.05),而与SDF-1+12G5组和12G5组比较差异无显著性(P>0.05);ox-LDL组bcl-2和bax的比值明显低于正常对照组和SDF-1组(P<0.01),而与SDF-1+12G5组和12G5组差异无显著性(P>0.05)。结论SDF-1可明显促进ox-LDL诱导的VSMC增殖,并抑制细胞凋亡;SDF-1及其受体CXCR4构成的生物学轴可能通过bcl-2/bax途径影响VSMC凋亡。     

SDF—1α／CXCR4对大鼠颈总动脉球囊损伤后血管内膜修复的影响

目的观察基质细胞衍生因子1α（stromal derived factor-1,SDF-1α）与其受体CXCR4（CXC chemokiner receptor4）间的相互作用对血管损伤后内膜修复的影响。方法雄性SD大鼠分成对照组（C组,12只）、损伤组（S组）和AMD3100（octahydrochloride hydrate）干预组（A组,AMD3100200ng／kg,腹腔注射）,S组和A组大鼠均进行左侧颈总动脉球囊损伤术。S组和A组又分成术后即刻（So/Ao组,各12只）、术后1dS1d／A1d组,各12只）、术后4d（S4d／A4d组,各12只）、术后7d（S7d／A7d组,各12只）和术后1月（S1m／A1m组,各12只）,S组和A组大鼠均进行左侧颈总动脉球囊损伤术。所有大鼠均采血,应用流式细胞仪检测外周血CD34^＋CXCR4^＋细胞,应用ELISA法检测血浆SDF—1α水平,同时取左侧颈总动脉进行SDF-1α和CXCR4的mRNA和蛋白表达的检测,并应用免疫组化法检测损伤后血管内膜中CXCR4的表达。结果（1）在S组和A组中均发现损伤后外周血循环中CD34^＋CXCR4^＋细胞显著增加（P〈0．01）,以手术后即刻上升最明显,随后逐渐下降,在术后第7天恢复到基础水平,A0和A1d组的细胞数量多于So和S1d组。（2）ELISA法结果显示：S组和A组大鼠在颈总动脉球囊损伤术后,血浆内SDF—1α水平急剧上升,在术后1d达峰值（P〈0．01）,随后迅速下降,至术后7d已回复至术前水平（P〉0．05）。（3）在S组和A组中均可见到SDF—1α和CXCR4mRNA的表达,其中SDF-1α在损伤后即刻就有表达,在术后第7天表达减弱,CXCR4mRNA则在术后第4天开始出现,在术后1m时仍有表达,但是S组中CXCR4mRNA的表达强于A组;C组中无SDF—1α和CXCR4mRNA的表达。（4）在S组和A组中均可见到CXCR4蛋白的表达（67kD）,但是S组的蛋白表达是A组的1．05～1．36倍,C组中未见CXCR4蛋白的表达。（5）应用CXCR4抗体与新生内膜中的CXCR4受体结合,发现S组中颈总动脉损伤后第1天,损伤内膜处已有CXCR4抗体与相应受体结合的阳性反应,随后反应逐渐增强,而A组中直至第4天起才发现损伤内膜处有CXCR4抗体与相应受体结合的阳性反应。结论血管损伤后,损伤部位的SDF—1α表达增加,并吸引CD34^＋CXCR4^＋细胞定植于损伤处,参与损伤血管内膜的修复,应用CXCR4的受体拮抗剂AMD3100后可减少这一效应,说明SDF—1α／CXCR4之间的相互作用在血管损伤后修复、新生内膜的形成中具有重要的作用。     

Altered SDF-1/CXCR4 axis in patients with primary myelofibrosis and in the Gata1 low mouse model of the disease

OBJECTIVE: To assess whether alterations in the stromal cell-derived factor-1 (SDF-1)/CXCR4 occur in patients with primary myelofibrosis (PMF) and in Gata1 low mice, an animal model for myelofibrosis, and whether these abnormalities might account for increased stem/progenitor cell trafficking. MATERIALS AND METHODS: In the mouse, SDF-1 mRNA levels were assayed in liver, spleen, and marrow. SDF-1 protein levels were quantified in plasma and marrow and CXCR4 mRNA and protein levels were evaluated on stem/progenitor cells and megakaryocytes purified from the marrow. SDF-1 protein levels were also evaluated in plasma and in marrow biopsy specimens obtained from normal donors and PMF patients. RESULTS: In Gata1 low mice, the plasma SDF-1 protein was five times higher than normal in younger animals. Furthermore, SDF-1 immunostaining of marrow sections progressively increased with age. Similar abnormalities were observed in PMF patients. In fact, plasma SDF-1 levels in PMF patients were significantly higher (by twofold) than normal (p < 0.01) and SDF-1 immunostaining of marrow biopsy specimens demonstrated increased SDF-1 deposition in specific areas. In two of the patients, SDF-1 deposition was normalized by curative therapy with allogenic stem cell transplantation. Similar to what already has been reported for PMF patients, the marrow from Gata1 low mice contained fewer CXCR4 pos CD117 pos cells and these cells expressed low levels of CXCR4 mRNA and protein. CONCLUSION: Similar abnormalities in the SDF-1/CXCR4 axis are observed in PMF patients and in the Gata1 low mice model of myelofibrosis. We suggest that these abnormalities contribute to the increased stem/progenitor cell trafficking observed in this mouse model as well as patients with PMF.