注射用七叶皂苷钠联合脐带间充质干细胞移植改善脑梗死大鼠神经功能

背景：单纯的脐带间充质干细胞移植修复受损脑组织的作用并不十分理想。 目的：观察脐带间充质干细胞移植联合注射用七叶皂苷钠治疗大鼠脑梗死的效果。 方法：应用线栓法建立大鼠大脑中动脉阻塞模型,随机分为对照组、细胞移植组、七叶皂苷钠+ 细胞移植组,分别尾静脉注射细胞培养液、1×1010 L-1脐带间充质干细胞悬液、尾静脉1×1010 L-1脐带间充质干细胞悬液同时经腹腔注射七叶皂苷钠 5 mg/(kg•d),连续5 d。 结果与结论：移植后1周,七叶皂苷钠+细胞移植组大鼠神经功能障碍评分低于细胞移植组及对照组(P < 0.05);七叶皂苷钠+细胞移植组大鼠脑梗死周围组织AQP9 及AQP4 mRNA的表达低于细胞移植组,却高于对照组(P < 0.05);七叶皂苷钠+细胞移植组CM-Dil阳性细胞和神经元数量多于细胞移植组及对照组(P < 0.05)。提示脐带间充质干细胞移植联合注射用七叶皂苷钠治疗大鼠脑梗死可明显改善大鼠的神经功能。     

原子力显微镜观察人脐带间充质干细胞：生物学特性与超微结构的关系*

背景：细胞形态结构和功能密不可分,但目前对人脐带间充质干细胞超微结构的报道还很少。 目的：探讨人脐带间充质干细胞的生物学特性与原子力显微镜下超微结构的关系。 方法：采用酶消化法体外分离培养并扩增人脐带间充质干细胞,取第3代细胞,用原子力显微镜在接触模式下观察成像,通过流式细胞仪检测细胞免疫表型和细胞周期,油红O染色及碱性磷酸酶染色鉴定其成脂、成骨分化潜能。 结果与结论：第3代人脐带间充质干细胞强表达CD44,CD29,低表达CD106,不表达CD34;有80%以上的细胞处在G0/G1期,增殖指数为19.9%;成脂诱导后,油红O染色可见胞浆中有大量桔红色的小脂滴;成骨诱导后,碱性磷酸酶染色可见立方形、多边形细胞的胞质染成棕褐色。原子力显微镜下人脐带间充质干细胞以长梭形为主,细胞骨架丝明显,并形成网状连接,这与其强大的增殖、迁移、分化功能相适应。     

人脐血干细胞移植修复大鼠脊髓损伤的功能评价

背景：在脊髓损伤模型的研究中，BBB评分系统和斜板试验均与脊髓损伤程度高度相关。 目的：建立改进大鼠脊髓全横断模型，对比观察BBB评分和斜板试验在人脐血干细胞移植术后功能评价中的作用。 设计、时间及地点：观察对比实验，于2007-04/2008-07在广东省组织构建与检测重点实验室完成。 材料：脐血来自健康足月产妇。实验动物为SPF级健康成年雌性SD大鼠50只，随机分为假手术对照组（n＝10）、磷酸盐注射组（n=20）、脐血干细胞移植组（n=20）。 方法：分离和体外培养人脐血干细胞。在显微镜下纵行剖开SD大鼠硬脊膜囊后，于硬脊膜内、蛛网膜外将超薄刀片刀尖直抵椎体骨质，将蛛网膜、脊髓、两侧壁和腹侧的硬脊膜作为一个整体完全划断，制作大鼠脊髓全横断模型，假手术对照组仅打开椎板，脐血干细胞移植组大鼠于脊髓两断端分别显微注射人脐血干细胞悬液6×109 ~7×109L-1，磷酸盐注射组注射等量磷酸盐缓冲液。 主要观察指标：术后12周内每2周分别应用BBB评分和斜板试验进行1次后肢运动功能评价。 结果：假手术对照组后肢运动功能于手术前后无显著变化。从术后第2周开始，磷酸盐注射组和脐血干细胞移植组逐渐恢复部分后肢运动功能，两组的BBB评分和斜板试验检测结果均表现出一致的增长趋势，呈线性正相关关系。从术后4周开始， BBB评分<13分的磷酸盐注射组和BBB评分≥13分的脐血干细胞移植组大鼠，在斜板试验中差异非常显著（P < 0.01）。到术后第12周时，磷酸盐注射组和脐血干细胞移植组斜板试验角度和恢复程度分别为34.25°和52.94％、53.85°和83.23％，并且脐血干细胞移植组高于磷酸盐注射组（P < 0.01），而同期磷酸盐注射组和脐血干细胞移植组对应的BBB评分和恢复程度仅分别为8.15和38.81％、13.90和66.19％，可见两组斜板试验角度恢复程度亦明显高于其自身对应的BBB评分结果。 结论：与BBB评分系统相比，斜板试验能更敏感地反映大鼠爪的功能恢复，但不能特异性地反映爪的精细运动。而作为主观评分体系的BBB评分系统，虽然特异性很高，但容易受主观因素的影响，敏感性较低。因此，联合应用BBB评分和斜板试验能更好地反映脊髓神经功能恢复情况。     

人脐带间充质干细胞冻存复苏后的生物学特征*◆

背景：冻存脐带间充质干细胞,并有效地保持其生物学特性,是储存脐带间充质干细胞以供临床使用的重要工艺之一。 目的：观察冻存后脐带间充质干细胞的生物学特性,验证其是否仍具有间充质干细胞的基本特征。 方法：从脐带分离得到间充质干细胞后,将其冻存于液氮之中。比较经冻存复苏后的脐带间充质干细胞和新鲜制备的脐带间充质干细胞活率、抑制人外周血单个核细胞分泌γ-干扰素等方面的异同。检验经冻存复苏后的脐带间充质干细胞是否具有多向分化潜能,其表型是否满足间充质干细胞的基本特征。 结果与结论：在细胞活率和抑制人外周血单个核细胞分泌γ-干扰素方面,冻存和新鲜的脐带间充质干细胞差异无显著性意义。经冻存复苏后的脐带间充质干细胞保持了间充质干细胞的基本形态,其表型满足间充质干细胞的基本要求,具有多向分化的潜能。     

人脐带间充质干细胞生物学特性及其研究进展

背景：人脐带间充质干细胞是一类具有自我更新和多向分化潜能的成体干细胞,具有来源丰富,对供者无影响,易于采集和运输,无异体排斥反应,避免伦理争议等诸多优点。 目的：综述人脐带间充质干细胞的生物学特性及其应用进展。 方法：以“人脐带间充质干细胞,生物学特征,基因分析,诱导分化, human umbilical cord mesenchymal stem cells(hUMSCs),biocharacteristics, gene analysis,induce differentiation”为关键词应用计算机检索CNKI数据库、万方数据库、PubMed数据库文章。 结果与结论：人脐带间充质干细胞呈典型的成纤维状,其表达的表面标志抗原具有非单一性,高表达间质细胞标志、整合素受体,不表达造血系标志、协同刺激分子CD80、CD86 和CD40、人白细胞抗原HLA-DR,HLA-G,HLA-DP,HLA-DQ、内皮标志CD31或CD33、CD14、CD56等。人脐带间充质干细胞与造血干细胞和胚胎干细胞类似,在体外可以分化为骨细胞、软骨细胞、肝细胞、心肌细胞等;在体内可以分化为多巴胺能神经元、骨骼肌细胞、内皮细胞、胰岛细胞等。但人脐带间充质干细胞的研究仍存在亟需解决的问题,如分离方法、培养条件的规范化,如何控制其生长和分化等。     

Neural cell injury microenvironment induces neural differentiation of human umbilical cord mesenchymal stem cells

This study aimed to investigate the neural differentiation of human umbilical cord mesenchymal stem cells (hUCMSCs) under the induction of injured neural cells. After in vitro isolation and culture, passage 5 hUCMSCs were used for experimentation. hUCMSCs were co-cultured with normal or Aβ_1-40-injured PC12 cells, PC12 cell supernatant or PC12 cell lysate in a Transwell co-culture system. Western blot analysis and flow cytometry results showed that choline acetyltransferase and microtubule-associated protein 2, a specific marker for neural cells, were expressed in hUCMSCs under various culture conditions, and highest expression was observed in the hUCMSCs co-cultured with injured PC12 cells. Choline acetyltransferase and microtubule-associated protein 2 were not expressed in hUCMSCs cultured alone (no treatment). Cell Counting Kit-8 assay results showed that hUCMSCs under co-culture conditions promoted the proliferation of injured PC12 cells. These findings suggest that the microenvironment during neural tissue injury can effectively induce neural cell differentiation of hUCMSCs. These differentiated hUCMSCs likely accelerate the repair of injured neural cells.     

自体分选造血干细胞和异体脐带间充质干细胞移植治疗神经系统变性疾病

背景：将自体分选造血干细胞和异体脐带间充质干细胞技术应用于临床,试图寻找一种新的治疗神经系统变性疾病的可行方法。 目的：探讨自体分选CD34+造血干细胞和异体脐带间充质干细胞治疗神经系统变性病的可行性。 方法：入选神经系统变性病患者21例,其中运动神经元病15例,脊髓小脑共济失调6例。取4 mL的脐带间充质干细胞或自体分选干细胞液经腰穿注射到患者蛛网膜下腔,每次注射的细胞数1.0×107。于干细胞治疗后3个月进行评分。 结果与结论：15例运动神经元病患者,治疗总有效率为80%,治疗前后的脊髓侧索硬化功能分级量表和自我评估问卷评分差异有显著性意义(P < 0.05);6例脊髓小脑共济失调患者,按ICARS评分,4例病情等级下降,治疗总有效率为77%。21例中4例有轻度低颅压性头痛(腰穿后),14例采用脐带间充质干细胞治疗的患者中2例在治疗后2 h出现短暂发热,其余患者治疗后未见明显不良反应。结果初步表明自体分选造血干细胞和异体脐带间充质干细胞治疗神经系统变性病临床疗效肯定,安全可行。     

Stem cell transplantation for treatment of cerebral ischemia in rats Effects of human umbilical cord blood stem cells and human neural stem cells

BACKGROUND:Exogenous neural stem cell transplantation promotes neural regeneration. However, various types of stem cells transplantation outcomes remain controversial. OBJECTIVE:To explore distribution, proliferation and differentiation of human neural stem cells (hNSCs) and human umbilical cord blood stem cells (hUCBSCs) following transplantation in ischemic brain tissue of rats, and to compare therapeutic outcomes between hNSCs and hUCBSCs. DESIGN, TIME AND SETTING:Randomized controlled animal studies were performed at the Experimental Animal Center of Nanjing Medical University and Central Laboratory of Second Affiliated Hospital of Nanjing Medical University of China from September 2008 to April 2009. MATERIALS:hNSCs were harvested from brain tissue of 10-13 week old fetuses following spontaneous abortion, and hUCBSCs were collected from umbilical cord blood of full-term newborns at the Second Affiliated Hospital of Nanjing Medical University of China. hNSCs and hUCBSCs were labeled by 5-bromodeoxyuridine (BrdU) prior to transplantation. METHODS:Rat models of cerebral ischemia were established by the suture method. A total of 60 healthy male Sprague Dawley rats aged 7-9 weeks were randomly assigned to hNSC transplantation, hUCBSC transplantation and control groups. The rat models in the hNSC transplantation, hUCBSC transplantation and control groups were infused with hNSC suspension, hUCBSC suspension and saline via the caudal vein, respectively. MAIN OUTCOME MEASURES:The distribution, proliferation and differentiation of hNSCs and hUCBSCs in ischemic brain tissue were observed using immunohistochemical methods. Neurological function in rats was assessed using the neurological severity score. RESULTS:The number of BrdU-positive cells was significantly greater in the hNSC transplantation group compared with hUCBSC transplantation group at 14 days following transplantation (P < 0.05). The number of BrdU-positive cells reached a peak at 28 days following transplantation. Nestin-positive, glial fibrillary acidic protein-positive, cyclic nucleotide 3' phosphohydrolase-positive and neuron specific enolase-positive cells were visible following transplantation. No significant difference was determined in the constituent ratio of various cells between hNSC and hUCBSC transplantation groups (P > 0.05). The neurological severity score was significantly decreased in rats at 21 days following transplantation (P < 0.05). No significant difference was detected in neurological severity score between hNSC and hUCBSC transplantation groups at various time points (P > 0.05). CONCLUSION:The transplanted hNSCs and hUCBSCs can migrate into ischemic brain tissue, proliferate and differentiate into neuron-like, astrocyte-like and oligodendrocyte-like cells, and improve neurological function in rats with cerebral ischemia.     

Transplantation of mesenchymal stem cells from human umbilical cord versus human umbilical cord blood for peripheral nerve regeneration

BACKGROUND: Mesenchymal stem cells (MSCs) appear to be a good alternative to Schwann cells in the treatment of peripheral nerve injury. Fetal stem cells, like umbilical cord blood (UCB) and umbilical cord (UC) stem cells, have several advantages over adult stem cells.OBJECTIVE: To assess the effects of UC-derived MSCs (UCMSCs) and UCB-derived MSCs (UCBMSCs) in repair of sciatic nerve defects. DESIGN, TIME AND SETTING: A randomized controlled animal experiment was performed at the laboratory of Department of Oral and Maxillofacial Surgery, Seoul National University Dental Hospital, from July to December 2009. MATERIALS: UCMSCs were provided by the Research Institute of Biotechnology, Dongguk University. UCBMSCs were provided by the Laboratory of Stem Cells and Tumor Biology, College of Veterinary Medicine, Seoul National University. Dulbecco's modified Eagle's medium (DMEM) was purchased from Gibco-BRL, USA. METHODS: Seven-week-old Sprague-Dawley rats were randomly and evenly divided into three groups: DMEM, UCBMSCs, and UCMSCs. A 10-mm defect in the left sciatic nerve was constructed in all rats. DMEM (15 μL) containing 1 × 106 UCBMSCs or UCMSCs was injected into the gap between nerve stumps, with the surrounding epineurium as a natural conduit. For the DMEM group, simple DMEM was injected. MAIN OUTCOME MEASURES: At 7 weeks after sciatic nerve dissection, dorsal root ganglia neurons were labeled by fluorogold retrograde labeling. At 8 weeks, electrophysiology and histomorphometry were performed. At 2, 4, 6, and 8 weeks after surgery, sciatic nerve function was evaluated using gait analysis.RESULTS: The UCBMSCs group and the UCMSCs group exhibited similar sciatic nerve function and electrophysiological indices, which were better than the DMEM group, as measured by gait analysis (P < 0.05). Fluorogold retrograde labeling of sciatic nerve revealed that the UCBMSCs group demonstrated a higher number of labeled neurons; however, the differences were not significant. Histomorphometric indices were similar in the UCBMSCs and UCMSCs groups, and total axon counts, particularly axon density (P < 0.05), were significantly greater in the UCBMSCs and UCMSCs groups than in the DMEM group. CONCLUSION: Transplanting either UCBMSCs or UCMSCs into axotomized sciatic nerves could accelerate and promote sciatic nerve regeneration over 8 weeks. Both treatments had similar effects on nerve regeneration.     

Therapeutic effect of human umbilical cord mesenchymal stem cells on neonatal rat hypoxic–ischemic encephalopathy

The therapeutic potential of umbilical cord blood mesenchymal stem cells has been studied in several diseases. However, the possibility that human umbilical cord Wharton's jelly‐derived mesenchymal stem cells (hUCMSCs) can be used to treat neonatal hypoxic–ischemic encephalopathy (HIE) has not yet been investigated. This study focuses on the potential therapeutic effect of hUCMSC transplantation in a rat model of HIE. Dermal fibroblasts served as cell controls. HIE was induced in neonatal rats aged 7 days. hUCMSCs labeled with Dil were then transplanted into the models 24 hr or 72 hr post‐HIE through the peritoneal cavity or the jugular vein. Behavioral testing revealed that hUCMSC transplantation but not the dermal fibroblast improved significantly the locomotor function vs. vehicle controls. Animals receiving cell grafts 24 hr after surgery showed a more significant improvement than at 72 hr. More hUCMSCs homed to the ischemic frontal cortex following intravenous administration than after intraperitoneal injection. Differentiation of engrafted cells into neurons was observed in and around the infarct region. Gliosis in ischemic regions was significantly reduced after hUCMSC transplantation. Administration of ganglioside (GM1) enhanced the behavioral recovery on the base of hUCMSC treatment. These results demonstrate that intravenous transplantation of hUCMSCs at an early stage after HIE can improve the behavior of hypoxic–ischemic rats and decrease gliosis. Ganglioside treatment further enhanced the recovery of neurological function following hUCMSC transplantation. © 2013 Wiley Periodicals, Inc.     

脐带间充质干细胞向成骨细胞分化的潜能

背景：人脐带间充质干细胞含量丰富,较为原始,分化能力强,免疫原性低,是细胞治疗的理想靶细胞。 目的：体外分离培养脐带间充质干细胞并将其定向诱导为成骨细胞。 方法：无菌条件下培养脐带间充质干细胞,分为诱导组和对照组,诱导组用成骨诱导液处理、对照组为干细胞培养液处理。倒置光显微镜观察细胞形态,MTT法测细胞增殖,荧光双染法检测细胞活力,流式细胞仪检测细胞周期与细胞表面标记。诱导后：检测碱性磷酸酶,Von Kossa染色分析钙盐沉积,RT-PCR检测骨桥蛋白基因、碱性磷酸酶、骨唾液蛋白mRNA的表达。 结果与结论：传代细胞形态稳定、活力好,高标达CD44。诱导后von Kossa染色表现为阳性。碱性磷酸酶活性诱导组比对照组高(P < 0.05),不同时间点比较21 d最高(P < 0.05)。RT-PCR显示：诱导组21,28d 碱性磷酸酶mRNA表达均较与对照组增强(P < 0.05)。诱导组有骨唾液蛋白和骨桥蛋白基因mRNA表达。提示,人脐带间充质干细胞能够定向分化为成骨细胞。     

体外诱导人脐带间充质干细胞向肝样细胞的分化

背景：多项研究表明间充质干细胞具有向肝细胞分化的潜能,这将为肝细胞移植、生物人工肝、肝组织工程提供大量种子细胞,也有望在急性肝衰竭、终末期肝病和遗传代谢性肝脏疾病治疗方面带来较大突破。 目的：拟在体外诱导人脐带间充质干细胞向肝样细胞分化,观察其形态及功能变化。 方法：取体外培养第3代处于对数生长期的人脐带间充质干细胞,以1×109 L-1的细胞浓度种植在培养瓶中,加入含体积分数为10%胎牛血清的DMEM/F12培养基,设立3组：实验1组加入20 μg/L肝细胞生长因子+10 μg/L碱性成纤维细胞生长因子,实验2组在其基础上另加入20 μg/L制瘤素,空白对照组不加入任何生长因子。根据细胞生长情况,每周换液两三次。分别于培养7,14,18 d在倒置显微镜下观察细胞形态,采用免疫细胞化学检测甲胎蛋白、白蛋白及细胞角蛋白18的表达,运用PAS法检测糖原的表达。 结果与结论：人脐带间充质干细胞可在含20 μg/L肝细胞生长因子、10 μg/L碱性成纤维细胞生长因子、20 μg/L制瘤素、体积分数为10%胎牛血清的DMEM/F12培养体系中诱导分化为具有肝细胞表型和功能的细胞,且肝细胞生长因子、碱性成纤维细胞生长因子、制瘤素联合应用的诱导效果优于单纯应用前两者。     

人脐带来源间充质干细胞分离培养方法的优化

背景：关于人脐带间充质干细胞的分离培养方法不一,如何提高人脐带来源间充质干细胞获得效率的问题尚未解决。 目的：优选人脐带来源间充质干细胞制备的消化酶组分。 方法：无菌条件下取正常足月剖腹产的脐带近胎儿段,按不同混合酶浓度比值划分为3组,混合酶Ⅰ组成分为0.4%Ⅱ型胶原酶、0.1%胰酶、0.02%EDTA和0.1%透明质酸酶;混合酶Ⅱ组成分为0.1%Ⅱ型胶原酶、0.4%胰酶、0.02%EDTA和0.1%透明质酸酶;混合酶Ⅲ组成分为0.3%Ⅱ型胶原酶、0.1%胰酶、0.02%EDTA、0.1%透明质酸酶和0.1%DNA酶。每组再按消化时间分为1,2,3 h 3个亚组。最后重悬细胞终体积均定位4 mL并计数,采用含血清的DMEM培养液体外培养细胞。 结果与结论：采用3种不同混合酶浓度进行消化,发现相同消化时间内,混合酶Ⅲ组消化细胞最多(P < 0.05)。在作用3 h时,混合酶Ⅲ组获取的细胞中活细胞比值显著低于混合酶Ⅰ组和混合酶Ⅱ组(P < 0.01)。提示0.3%Ⅱ型胶原酶、0.1%胰酶、0.02%EDTA、0.1%透明质酸酶和0.1%DNA酶混合液消化脐带组织块2 h为获取脐带间充质干细胞的最佳条件。     

人脐血间充质干细胞与β-磷酸三钙的生物相容性

背景：体内实验显示,β-磷酸三钙多孔陶瓷是较为理想的骨组织工程支架材料,但由于体内植入实验受多种因素的影响,不能很好反映细胞的生长、增殖和表型变化。 目的：观察体外人脐血间充质干细胞与β-磷酸三钙多孔陶瓷的生物相容性。 方法：将培养的第6代人脐血间充质干细胞悬液滴注入β-磷酸三钙内部进行复合,然后将干细胞-支架材料复合物置入含体积分数为10%胎牛血清的α-MEM培养体系中培养,于培养第4,8,12天电镜下观察人脐血间充质干细胞在材料表面及内部生长情况,采用MTT测试法绘制细胞生长曲线,并进行DNA含量、蛋白质含量测定。 结果与结论：人脐血间充质干细胞与β-磷酸三钙体外复合后能够在β-磷酸三钙支架材料表面及内部的孔隙内贴附,且生长良好,其DNA复制和蛋白合成功能不受β-磷酸三钙的影响。说明人脐血间充质干细胞和β-磷酸三钙支架材料生物相容性良好,二者可作为种子细胞和支架材料用于组织工程化骨与软骨的构建。     

人脐带间充质干细胞治疗糖尿病大鼠周围神经病变的研究

目的 探讨人脐带间充质干细胞移植对糖尿病周围神经病的治疗作用.方法 贴壁法培养人脐带间充质干细胞,流式鉴定免疫表型.STZ诱导糖尿病周围神经病大鼠模型,左后肢肌肉注射移植干细胞,观察糖尿病周围神经病在神经电生理和病理病变以及干细胞整合情况.结果 移植干细胞后坐骨神经的电生理及病理显著改善,但移植的干细胞并未整合到神经纤维中.结论 人脐带间充质干细胞移植可改善糖尿病周围神经病大鼠的坐骨神经电生理和病理,但干细胞并未直接参与神经重建,提示可能通过其他间接途径起作用.     

CM-Dil体外标记人脐带间充质干细胞传代示踪的可行性

背景：掌握人脐带间充质干细胞的移植示踪方法是研究其生物学特性的关键。 目的：观察用CM-Dil标记人脐带间充质干细胞及在体外传代示踪的可行性。 方法：采用酶消化法体外分离培养人脐带间充质干细胞,通过流式细胞仪检测细胞免疫表型和细胞周期、体外成脂成骨诱导鉴定该细胞。将第5代细胞用CM-Dil标记,并将细胞传代,荧光显微镜观察体外标记情况。 结果与结论：第3代人脐带间充质干细胞强表达CD44,CD29,低表达CD106,不表达CD34、CD40;有80%以上的细胞处在G0/G1期,成脂成骨诱导后,油红O染色和碱性磷酸酶染色分别阳性。CM-Dil标记人脐带间充质干细胞细胞标记率达90%以上,体外传代后荧光强度逐渐减退,传8代后,荧光基本消失。说明人脐带间充质干细胞增殖、分化能力强,CM-Dil标记细胞示踪方法简单易行。     

移植人脐带间充质干细胞修复大鼠脊髓损伤

背景：已知人脐带间充质干细胞对脊髓损伤存在着潜在的治疗价值,然而,当前对移植人脐带间充质干细胞治疗脊髓损伤及机制方面研究很少。 目的：观察人脐带间充质干细胞对脊髓损伤大鼠的治疗效果。 方法：40只Wistar大鼠建立脊髓损伤模型,38只造模成功后随机摸球法分为3组：空白对照组：只接受单纯损伤,不做任何移植;DMEM移植组：损伤后1周予以5 μL DMEM局部移植;细胞移植组：损伤后1周予以5 μL准备好的人脐带间充质干细胞局部移植(细胞数1×106)。移植后对实验动物通过BBB评分、体感诱发电位与运动诱发电位观察后肢功能恢复情况。分别于损伤后2,4,6,8,10周随机于细胞移植组抽取大鼠2只,免疫组织化学染色观察人脐带间充质干细胞存活、迁移、分化,通过胶质纤维酸性蛋白阳性细胞染色比较各组损伤局部胶质瘢痕形成面积。 结果与结论：BBB评分损伤后4周细胞移植组高于其他两组(P < 0.05),损伤后12周细胞移植组与其他两组相比SEP、MEP潜伏期缩短、波幅值增高(P < 0.05)。免疫组织化学染色示人脐带间充质干细胞可向神经元、星形胶质细胞和少突胶质细胞分化,分化的少突胶质细胞并包绕轴突形成髓鞘。细胞移植组损伤局部胶质瘢痕面积均小于其他两组(P < 0.05),空白对照组、DMEM移植组间差异无显著性(P > 0.05)。提示未经体外诱导的人脐带间充质干细胞可于损伤大鼠脊髓体内向神经元、星形胶质细胞、少突胶质细胞分化,减小胶质瘢痕,并促进脊髓损伤大鼠神经功能的恢复。     

Protective effects of human umbilical cord mesenchymal stem cell vein transplantation against spinal cord ischemia/reperfusion injury in rats

BACKGROUND: The majority of studies addressing spinal cord ischemia/reperfusion injury (SCIRI) have focused on drugs, proteins, cytokines, and various surgical techniques. A recent study reports that human umbilical cord mesenchymal stem cell (hUCMSC) transplantation achieves good therapeutic effects, but the mechanisms underlying nerve protection remain poorly understood.OBJECTIVE: To observe survival of transplanted hUCMSCs in SCIRI rat models and the influence on motor function in the hind limbs, to determine interleukin-8 expression and cellular apoptosis in spinal cord tissues, and to verify the hypothesis that hUCMSC transplantation exhibits protective effects on SCIRI.DESIGN, TIME AND SETTING: A randomized, controlled, animal experiment was performed at the Laboratory of the Department of Orthopedics in the First Affiliated Hospital of Soochow University,China between January 2007 and December 2008.MATERIALS: hUCMSCs were harvested from umbilical cord blood of healthy pregnant women after parturition in the Obstetrical Department of the First Affiliated Hospital of Soochow University, China. Rabbit anti-human BrdU monoclonal antibody was provided by DAKO, USA. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) Kit and enzyme-linked immunosorbent assay (ELISA) Kit were purchased by Wuhan Boster, China. METHODS: A total of 72 healthy, Wistar, adult rats were randomly assigned to three groups: sham-surgery, model, and transplantation, with 24 rats in each group. SCIRI was induced in the model and transplantation groups via the abdominal aorta block method. The inf rarenal abdominal aorta was not blocked in the sham-surgery group. Prior to abdominal aorta occlusion, 0.2-0.3 mL bromodeoxyuridine (BrdU)-labeled hUCMSCs suspension (cell concentration 5 × 10~3/μL) was injected through the great saphenous vein of the hind limb, and an equal volume of physiological saline was administered to the model and sham-surgery groups.MAIN OUTCOME MEASURES: Pathological observation of rat spinal cord tissues was performed by hematoxylin-eosin staining at 6, 24, and 48 hours post-surgery. Immunohistochemistry was applied to determine hUCMSCs survival in the spinal cord. The amount of cellular apoptosis and interleukin-8 expression in spinal cord tissues was assayed utilizing the TUNEL and ELISA methods, respectively. Motor function in the hind limbs was evaluated according to Jacob's score. RESULTS: Numerous BrdU-positive cells were observed in spinal cord tissues from the transplantation group. The number of apoptotic cells and interleukin-8 levels significantly decreased in the transplantation group (P < 0.05), pathological injury was significantly ameliorated, and motor function scores significantly increased (P < 0.05) compared with the model group. CONCLUSION: Via vein transplantation, hUCMSCs were shown to reach and survive in the injury area. Results suggested that the transplanted hUCMSCs contributed to significantly improved pathological changes in the injured spinal cord, as well as motor function, following SCIRI. The protective mechanism correlated with inhibition of cellular apoptosis and reduced production of inflammatory mediators.     

BDNF expression is up-regulated by progesterone in human umbilical cord mesenchymal stem cells

Objective: To investigate whether promotion of neuronal differentiation of human umbilical cord mesenchymal stem cells (HUMSCs) by progesterone (PROG) involves changes in brain-derived neurotrophic factor (BDNF) levels.

Methods: We used rat brain tissue extracts to mimic the brain microenvironment. Quantitative sandwich enzyme-linked immunosorbent assay was performed to measure levels of BDNF in cultured medium with or without PROG.

Results: Progesterone increased levels of BDNF in HUMSCs.

Conclusion: Progesterone enhancement of brain-derived neurotrophic factor levels may be involved in PROG activated-pathways to promote neuronal differentiation of HUMSCs.