亚低温预处理对大鼠局灶性脑缺血的保护作用

我们采用大鼠大脑中动脉缺血再灌注模型,在缺血开始前即应用亚低温,模拟神经外科手术中往常面临的缺血情况。研究亚低温预处理的作用效果与缺血时程的关系,为今后亚低温在实验研究和临床应用的范围提供依据。 一、材料与方法 1.实验动物及分组:72只健康雄性SD大鼠,体重250～300 g,随机分为6个实验组:1、2、3 h脑缺血再灌注常温组及亚低温组。亚低温组动物在缺血开始前     

线栓法制备大鼠局灶性脑缺血再灌注模型的研究进展

缺血性脑血管病的研究需要合适的动物模型,建立重复性好、结果可靠、稳定性强的动物模型一直是研究者追求的目标。颈内动脉线栓法制备大鼠局灶性脑缺血模型,无需开颅、重复性好、可准确控制缺血和再灌注时间,但仍有很多因素直接影响模型的成功率,给复制带来较大困难。现就模型复制过程需注意的环节和研究现状作一综述。     

单次电针预处理诱导大鼠局灶性脑缺血耐受的时程

目的 探讨单次电针预处理能否诱导局灶性脑缺血耐受及其时程。方法 48只健康雄性SD大鼠(280～320g),随机分为6组(n=8):对照组和电针1～5组(EA1～5)。对照组未行任何预处理;电针1～5组分别在电针刺激百会穴30min后0.5、1、2、3和24 h给予局灶性脑缺血。采用颈内动脉尼龙线线栓法致大脑中动脉栓塞(120 min)模型,观察再灌注后24 h时神经功能损害并取大脑行TTC染色以测量脑梗死容积。结果术后动物均存活。电针刺激百会穴30 min后2 h和24 h给予局灶性脑缺血,再灌注24 h时神经功能缺陷评分明显低于对照组(P<0.05),脑梗死容积明显小于对照组(P<0.01),其它时间点与对照组差异无显著性。结论 单次电针刺激百会穴30 min,可诱导双相大脑局灶性缺血耐受,即急性缺血耐受出现在预处理后2 h,延迟缺血耐受出现在预处理后24 h。     

脐血间充质干细胞移植对大鼠局灶性脑缺血的影响

目的　从人脐血中分离纯化间充质干细胞(MSC) ,观察其移植对大鼠大脑中动脉栓塞后神经功能恢复的影响及细胞的存活、迁移向神经细胞分化的情况。方法　雄性SD大鼠45只,用线栓法建立大鼠大脑中动脉栓塞(MCAO)模型,大鼠随机分为3组:MSC移植组、单核细胞组和生理盐水组。移植后1、7、14、2 1、2 8d采用改良神经功能损害评分(mNSS)观察大鼠神经功能恢复情况,应用免疫组织化学和免疫荧光双标记技术检测5溴 2脱氧尿核苷(BrdU )标记的MSC细胞的存活、迁移及其胶质纤维酸性蛋白(GFAP)和神经元特异性核蛋白(NeuN)的表达。结果　人脐血MSC细胞移植可显著提高大鼠局灶性脑缺血后神经功能的恢复(P <0 .0 5 )。移植的MSC细胞可在大鼠脑组织中存活,并向缺血区域迁移,11.67%MSC细胞表达GFAP ,3 .72 %MSC细胞表达NeuN。结论　人脐血中含有MSC细胞并可促进局灶性脑缺血大鼠的神经功能恢复,移植细胞可在大鼠脑缺血区域中存活、迁移并向星形胶质细胞或神经元分化     

探讨脑缺血大鼠骨髓基质细胞移植后减少神经细胞凋亡的作用

目的探讨大鼠局灶性脑缺血后骨髓基质细胞移植对神经细胞凋亡及相关蛋白表达的影响。方法取大鼠股骨髓基质细胞,分化培养为骨髓基质细胞源神经干细胞。将32只健康SD大鼠随机平均分为4组。A组不建立局灶性脑缺血模型,不做任何移植,其余步骤同其他组。B、C、D组均建立局灶性脑缺血再灌注模型。B组将1mlPBS经尾静脉注入大鼠体内;C组将1ml3×106个骨髓基质细胞经尾静脉注入大鼠体内;D组将1ml3×106个骨髓基质细胞源神经干细胞经尾静脉注入大鼠体内。分别在移植后7d和14d行脑灌注固定取材,应用免疫组织化学染色检测脑组织中Bcl2、Bax蛋白表达阳性的细胞,原位末端脱氧核糖核酸转移酶标记法(TUNEL)检测神经细胞凋亡数。结果C组和D组各时点的神经细胞凋亡数均少于B组(P<0.01),C组和D组移植14d时,神经细胞凋亡数显著少于移植7d时(P<0.01),移植14d时D组神经细胞凋亡数显著少于C组(P<0.05)。C组和D组Bcl2表达阳性的细胞数显著高于B组(P<0.01)。C组和D组Bax蛋白表达阳性的细胞数明显低于B组(P<0.01)。结论骨髓基质细胞源神经干细胞可能通过上调Bcl2蛋白,下调Bax蛋白的表达,减少神经细胞凋亡,从而对脑缺血再灌注损伤后的神经细胞起保护作用。     

L-精氨酸对大鼠局灶性脑缺血损伤的影响

目的 观察一氧化氮（NO）和一氧化氮供体L－精氨酸（L－Arginine)对大鼠局灶性脑缺血损伤的影响。方法 用线栓法建立大鼠大脑中动脉脑缺血（MCAO）模型。将42只SD大鼠随机分为4组：假手术组、对照组、L－精氨酸治疗组（500mg/kg、300mg/kg)，经腹腔给药，于规定时间处死大鼠，迅速取出大脑，观察大鼠脑缺血后脑梗死体积和脑组织中NO、丙二醛（MDA）含量、一氧化氮合酶（NOS）、超氧化物歧化酶（SOD）活性的变化。结果 L－精氨酸能明显减少脑缺血体积，改善脑损伤；大剂量比小剂量更明显；可明显增加脑组织中NO含量，减少MDA含量，增加SOD活性。结论 L－精氨酸对脑缺血早期有治疗作用，且剂量500mg/kg体重给药比300mg/kg体重给药治疗效果更为明显。     

葛根素对大鼠局灶性脑缺血再灌注后热休克蛋白70表达的影响

葛根素是从中药葛根提取的主要有效成分之一,为一种异黄酮化合物,热休克蛋白70(HSP70)是一类应激蛋白,对脑缺血损伤有保护作用,本研究拟观察局灶性脑缺血再灌注时HSP70的表达及葛根素对HSP70表达的影响。     

成年大鼠局灶性脑缺血后内源性神经前体细胞增殖迁移的研究

目的 研究局灶性脑缺血后成年大鼠内源性神经前体细胞在缺血后不同时间窗内增殖分化及前体细胞最大的增殖效应。方法 用Longa’s线栓法制作大鼠脑缺血模型，用免疫组织化学的方法检测大鼠内源性神经前体细胞最佳增殖时间及最大增殖效果。结果 脑缺血半球室管膜下区、嗅球和头端迁移流(RMS)溴化脱氧尿嘧啶(BrdU)阳性细胞数在脑缺血后1d明显增多，3～7d达高峰，14d后开始下降；BrdU阳性细胞数在脑缺血半球室管膜下区和嗅球成正相关；脑缺血侧海马齿状回未见BrdU阳性细胞数明显增多；BrdU阳性细胞最大数量在脑缺血后第3周，从第4周开始BrdU阳性细胞数下降。结论 成年大鼠局灶性脑缺血后3～7d内源性神经前体细胞增殖达高峰。在该期进行外源性细胞移植治疗可能会更行之有效。     

七氟醚对大鼠局灶性脑缺血再灌注损伤的保护作用

目的评价七氟醚对大鼠局灶性脑缺血再灌注损伤的保护作用及其机制。方法雄性SD大鼠24只，随机分为假手术组、损伤组、七氟醚组，每组8只。采用大脑中动脉线栓法阻断前脑血供3h、再灌注24h制备大鼠局灶性脑缺血再灌注损伤模型。七氟醚组于再灌注前30min经面罩吸入七氟醚（呼气末浓度维持1．0MAC，持续30min）。再灌注24h时用Zea Longa评分法进行神经功能缺陷评分，并测定体重。再灌注24h时用原位末端脱氧核苷酸转移酶标记法测定纹状体神经细胞凋亡，计算神经细胞凋亡密度，并用免疫组织化学法测定纹状体PKCγ蛋白的表达。结果与缺血前比较，再灌注24h时损伤组体重减轻（P〈0．01）；与假手术组比较，再灌注24h时损伤组和七氟醚组神经功能缺陷评分及神经细胞凋亡密度增加，七氟醚组纹状体PKCγ表达降低；与损伤组比较，七氟醚组神经功能缺陷评分、纹状体神经细胞凋亡密度降低，PKCγ表达增加（P〈0．05或0．01）。结论吸入1．0MAC七氟醚对大鼠局灶性脑缺血再灌注损伤产生保护作用，其机制与上调纹状体PKCγ蛋白表达有关。     

骨髓基质干细胞及其来源的产胰岛素细胞移植对受体胰岛和新生血管的影响

目的 观察骨髓基质干细胞(MSCs)及其来源的产胰岛素细胞(IPCs)移植对受体残余胰岛和其周围新生血管增殖的影响.方法 对大鼠MSCs进行体外诱导成为IPCs.对1型糖尿病大鼠随机分为胰岛素控制血糖组(对照组)、MSCs移植组及IPCs移植组3个治疗组(每组10只).至血糖开始下降至10mmoL/L,同期移除3组大鼠右肾及胰腺,分别进行PCNA、胰岛素和CD31抗体染色,观察3组大鼠肾脏和胰腺的胰岛素及血管内皮细胞表达情况.结果 MSCs诱导生成IPCs.移植物:移植侧的右肾均可见到较多的胰岛素和CD31阳性细胞.胰腺组织:(1)对照组:胰岛萎缩.(2)细胞移植组:残余胰岛增殖率:(20.84±3.48)%和(18.43±2.84)%(P>0.05),胰岛周围均有少量CD31阳性细胞.结论 MSCs及其来源的IPCs明显促进了受体残余胰岛周围新生血管的生成,进而使残余胰岛得到增殖,而MSCs在体内也可分化成IPCs和血管内皮细胞.     

局灶性脑缺血预处理后凋亡相关蛋白表达的动态研究

一次短暂的脑缺血能够激发脑组织对再次严重缺血产生保护性耐受,这次短暂的缺血被称为脑缺血预处理[1] 。我们通过采用开颅方法直接阻断大鼠大脑中动脉(MCA)模型,检测局灶性脑缺血预处理后凋亡相关蛋白Bcl 2、Bax及胱天蛋白酶(Caspase) 3 p2 0表达的动态改变及它们对神经细胞凋亡的影响。一、材料与方法1.动物模型制备及分组:局灶性脑缺血预处理及再次缺血动物模型采用Glazier等[2 ] 开颅阻断MCA的方法,160只雄性成年Wistar大鼠随机分为2 0min缺血预处理组、单次3、6、12、2 4h、2、5d缺血组、预处理后再次3、6、12、2 4h、2、5d缺血…     

线栓法制备大鼠局灶性脑缺血再灌注模型的研究进展

缺血性脑血管病的研究需要合适的动物模型，建立重复性好、结果可靠、稳定性强的动物模型一直是研究者追求的目标。颈内动脉线栓法制备大鼠局灶性脑缺血模型，无需开颅、重复性好、可准确控制缺血和再灌注时间，但仍有很多因素直接影响模型的成功率，给复制带来较大困难。现就模型复制过程需注意的环节和研究现状作一综述。     

异氟醚对大鼠局灶性脑缺血/再灌注损伤的保护作用

目的 探讨异氟醚对大鼠局灶性脑缺血/再灌注损伤的脑保护作用。方法 32只雄性SD大鼠,280～320 g,随机分为四组,假手术组:仅分离血管,不留置线拴;脑缺血组:缺血前吸入纯氧30min行2h大脑中动脉栓塞(MCAO);0.9％和1.5％异氟醚组:分别在MCA0前吸入0.9％和1.5％异氟醚30 min。监测缺血/再灌注期间鼓膜温度变化,测定再灌注22 h和70 h时脑梗死体积及再灌注22h时光、电镜的病理改变。结果 缺血侧鼓膜温度较非缺血侧明显降低,最大温差达0.78℃±0.35℃。异氟醚轻度缩小再灌注22 h和70 h时的脑梗死体积,且1.5％异氟醚对再灌注70 h时梗死体积百分比减小的效果优于0.9％异氟醚。光、电镜结果提示异氟醚能减轻局灶性脑缺血/再灌注损伤对神经元、线粒体和内皮细胞的损害。结论缺血前吸入0.9％和1.5％异氟醚对大鼠局灶性脑缺血,再灌注损伤,可产生一定程度的保护作用。     

骨髓基质干细胞及其来源的产胰岛素细胞移植对受体胰岛和新生血管的影响

Objective To investigate the proliferative impact of transplantation of bone marrow mesenchymal stem cells (MSCs) and insulin producing ceils (IPCs) from them on recipients' remnant islets and neovascularization around. Methods MSCs were induced to IPCs in vitro. The rats with type Ⅰ diabetic meilitus were randomly divided into control group, MSCs transplant group, and IPCs transplant group. When the blood glucose level of recipient rats was decreased to 10 mmol/L, the right kidney and pancreas were removed from each group simultaneously. All the thin tissue slices were estimated by immu-nohistochemistry staining of PCNA antibody,insulin antibody and CD31 anbibody. Results The passaged MSCs differentiated into IPCs. In the grafts, insulin-positive and CD31 -positive cells were observed in the right kidney. In control group, few proliferative cells located in the atrophic islets. In the transplant groups, a lot of proliferative islet cells were obsereved, around which few CD31 -positive ceils existed;Pancreatic proliferative islet cell rate in MSCs and IPCs transplant group was ( 20.84±3.48 ) % and ( 18.43± 2.84% ) ( P > 0.05 ) respectively. Conclusion Obviously, MSCs and IPCs promoted recipient neovascu-larization surrounding the islets, which enhanced the proliferation of remnant islet cells. MSCs differentiated to IPCs and vascular endothelial cells in the recipient transplant location.     

骨髓基质干细胞及其来源的产胰岛素细胞移植对受体胰岛和新生血管的影响

Objective To investigate the proliferative impact of transplantation of bone marrow mesenchymal stem cells (MSCs) and insulin producing ceils (IPCs) from them on recipients' remnant islets and neovascularization around. Methods MSCs were induced to IPCs in vitro. The rats with type Ⅰ diabetic meilitus were randomly divided into control group, MSCs transplant group, and IPCs transplant group. When the blood glucose level of recipient rats was decreased to 10 mmol/L, the right kidney and pancreas were removed from each group simultaneously. All the thin tissue slices were estimated by immu-nohistochemistry staining of PCNA antibody,insulin antibody and CD31 anbibody. Results The passaged MSCs differentiated into IPCs. In the grafts, insulin-positive and CD31 -positive cells were observed in the right kidney. In control group, few proliferative cells located in the atrophic islets. In the transplant groups, a lot of proliferative islet cells were obsereved, around which few CD31 -positive ceils existed;Pancreatic proliferative islet cell rate in MSCs and IPCs transplant group was ( 20.84±3.48 ) % and ( 18.43± 2.84% ) ( P > 0.05 ) respectively. Conclusion Obviously, MSCs and IPCs promoted recipient neovascu-larization surrounding the islets, which enhanced the proliferation of remnant islet cells. MSCs differentiated to IPCs and vascular endothelial cells in the recipient transplant location.     

骨髓基质干细胞及其来源的产胰岛素细胞移植对受体胰岛和新生血管的影响

Objective To investigate the proliferative impact of transplantation of bone marrow mesenchymal stem cells (MSCs) and insulin producing ceils (IPCs) from them on recipients' remnant islets and neovascularization around. Methods MSCs were induced to IPCs in vitro. The rats with type Ⅰ diabetic meilitus were randomly divided into control group, MSCs transplant group, and IPCs transplant group. When the blood glucose level of recipient rats was decreased to 10 mmol/L, the right kidney and pancreas were removed from each group simultaneously. All the thin tissue slices were estimated by immu-nohistochemistry staining of PCNA antibody,insulin antibody and CD31 anbibody. Results The passaged MSCs differentiated into IPCs. In the grafts, insulin-positive and CD31 -positive cells were observed in the right kidney. In control group, few proliferative cells located in the atrophic islets. In the transplant groups, a lot of proliferative islet cells were obsereved, around which few CD31 -positive ceils existed;Pancreatic proliferative islet cell rate in MSCs and IPCs transplant group was ( 20.84±3.48 ) % and ( 18.43± 2.84% ) ( P > 0.05 ) respectively. Conclusion Obviously, MSCs and IPCs promoted recipient neovascu-larization surrounding the islets, which enhanced the proliferation of remnant islet cells. MSCs differentiated to IPCs and vascular endothelial cells in the recipient transplant location.     

骨髓基质干细胞及其来源的产胰岛素细胞移植对受体胰岛和新生血管的影响

Objective To investigate the proliferative impact of transplantation of bone marrow mesenchymal stem cells (MSCs) and insulin producing ceils (IPCs) from them on recipients' remnant islets and neovascularization around. Methods MSCs were induced to IPCs in vitro. The rats with type Ⅰ diabetic meilitus were randomly divided into control group, MSCs transplant group, and IPCs transplant group. When the blood glucose level of recipient rats was decreased to 10 mmol/L, the right kidney and pancreas were removed from each group simultaneously. All the thin tissue slices were estimated by immu-nohistochemistry staining of PCNA antibody,insulin antibody and CD31 anbibody. Results The passaged MSCs differentiated into IPCs. In the grafts, insulin-positive and CD31 -positive cells were observed in the right kidney. In control group, few proliferative cells located in the atrophic islets. In the transplant groups, a lot of proliferative islet cells were obsereved, around which few CD31 -positive ceils existed;Pancreatic proliferative islet cell rate in MSCs and IPCs transplant group was ( 20.84±3.48 ) % and ( 18.43± 2.84% ) ( P > 0.05 ) respectively. Conclusion Obviously, MSCs and IPCs promoted recipient neovascu-larization surrounding the islets, which enhanced the proliferation of remnant islet cells. MSCs differentiated to IPCs and vascular endothelial cells in the recipient transplant location.     

骨髓基质干细胞及其来源的产胰岛素细胞移植对受体胰岛和新生血管的影响

Objective To investigate the proliferative impact of transplantation of bone marrow mesenchymal stem cells (MSCs) and insulin producing ceils (IPCs) from them on recipients' remnant islets and neovascularization around. Methods MSCs were induced to IPCs in vitro. The rats with type Ⅰ diabetic meilitus were randomly divided into control group, MSCs transplant group, and IPCs transplant group. When the blood glucose level of recipient rats was decreased to 10 mmol/L, the right kidney and pancreas were removed from each group simultaneously. All the thin tissue slices were estimated by immu-nohistochemistry staining of PCNA antibody,insulin antibody and CD31 anbibody. Results The passaged MSCs differentiated into IPCs. In the grafts, insulin-positive and CD31 -positive cells were observed in the right kidney. In control group, few proliferative cells located in the atrophic islets. In the transplant groups, a lot of proliferative islet cells were obsereved, around which few CD31 -positive ceils existed;Pancreatic proliferative islet cell rate in MSCs and IPCs transplant group was ( 20.84±3.48 ) % and ( 18.43± 2.84% ) ( P > 0.05 ) respectively. Conclusion Obviously, MSCs and IPCs promoted recipient neovascu-larization surrounding the islets, which enhanced the proliferation of remnant islet cells. MSCs differentiated to IPCs and vascular endothelial cells in the recipient transplant location.     

骨髓基质干细胞及其来源的产胰岛素细胞移植对受体胰岛和新生血管的影响

Objective To investigate the proliferative impact of transplantation of bone marrow mesenchymal stem cells (MSCs) and insulin producing ceils (IPCs) from them on recipients' remnant islets and neovascularization around. Methods MSCs were induced to IPCs in vitro. The rats with type Ⅰ diabetic meilitus were randomly divided into control group, MSCs transplant group, and IPCs transplant group. When the blood glucose level of recipient rats was decreased to 10 mmol/L, the right kidney and pancreas were removed from each group simultaneously. All the thin tissue slices were estimated by immu-nohistochemistry staining of PCNA antibody,insulin antibody and CD31 anbibody. Results The passaged MSCs differentiated into IPCs. In the grafts, insulin-positive and CD31 -positive cells were observed in the right kidney. In control group, few proliferative cells located in the atrophic islets. In the transplant groups, a lot of proliferative islet cells were obsereved, around which few CD31 -positive ceils existed;Pancreatic proliferative islet cell rate in MSCs and IPCs transplant group was ( 20.84±3.48 ) % and ( 18.43± 2.84% ) ( P > 0.05 ) respectively. Conclusion Obviously, MSCs and IPCs promoted recipient neovascu-larization surrounding the islets, which enhanced the proliferation of remnant islet cells. MSCs differentiated to IPCs and vascular endothelial cells in the recipient transplant location.     

骨髓基质干细胞及其来源的产胰岛素细胞移植对受体胰岛和新生血管的影响

Objective To investigate the proliferative impact of transplantation of bone marrow mesenchymal stem cells (MSCs) and insulin producing ceils (IPCs) from them on recipients' remnant islets and neovascularization around. Methods MSCs were induced to IPCs in vitro. The rats with type Ⅰ diabetic meilitus were randomly divided into control group, MSCs transplant group, and IPCs transplant group. When the blood glucose level of recipient rats was decreased to 10 mmol/L, the right kidney and pancreas were removed from each group simultaneously. All the thin tissue slices were estimated by immu-nohistochemistry staining of PCNA antibody,insulin antibody and CD31 anbibody. Results The passaged MSCs differentiated into IPCs. In the grafts, insulin-positive and CD31 -positive cells were observed in the right kidney. In control group, few proliferative cells located in the atrophic islets. In the transplant groups, a lot of proliferative islet cells were obsereved, around which few CD31 -positive ceils existed;Pancreatic proliferative islet cell rate in MSCs and IPCs transplant group was ( 20.84±3.48 ) % and ( 18.43± 2.84% ) ( P > 0.05 ) respectively. Conclusion Obviously, MSCs and IPCs promoted recipient neovascu-larization surrounding the islets, which enhanced the proliferation of remnant islet cells. MSCs differentiated to IPCs and vascular endothelial cells in the recipient transplant location.