细胞移植及细胞联合移植修复脊髓损伤的研究进展

<正>脊髓损伤(spinal cord injury,SCI)是中枢神经系统的严重创伤,其发病率和致残率呈逐年增高的趋势,而且给社会和家庭带来沉重的负担[1],是近年医学研究的热点难题之一。随着神经组织工程技术的发展,利用细胞移植和细胞联合移植修复SCI成为近年研究的热点。目前可供选择的移植细胞类型有神经干细胞(neural stem cells,NSCs)、施万细胞(schwann cells,SCs)、胚胎干细胞(embryonic stem cells,ESCs)、嗅鞘细胞(olfactory ensheating cells,OECs)、骨髓间充质干细胞(bone mesenechymal stem cells,BMSCs)、少突胶质     

树突状细胞与移植免疫耐受研究进展

诱导供者特异性免疫耐受是最终克服移植排斥提高受者生存质量的有效途径之一。近年随着对树突状细胞发育成熟过程的认识步步深入，树突状细胞在移植排斥和移植耐受平衡中的双向调节作用引入注目。     

胰岛细胞移植进展

近年来,随着在供体胰腺的获取与处理、胰岛的分离与纯化、新型免疫抑制剂的应用等领域不断取得突破,胰岛细胞移植已经公认为是一种治疗1型糖尿病安全有效的治疗方法。胰岛细胞移植治疗的理想目标在于获得比目前治疗方案更符合生理的代谢控制,便患者摆脱对胰岛素的依赖,并保持移植物长期有功能存活和改善患者的生活质量。然而,大量的临床试验表明目前的胰岛移植治疗糖尿病方案仍然存在较多难题,距离临床推广应用仍需时日。本文简要介绍了胰岛细胞移植的历史,并对胰岛分离技术、移植程序、免疫抑制剂的应用、异种胰岛细胞移植、干细胞移植、基因技术在胰岛细胞移植中的应用、羊膜上皮细胞移植、胰岛细胞移植所面临的挑战及将来发展方向等方面作了简要综述。     

细胞移植的免疫生物学

细胞移植的前景十分广阔，它不仅可用于诸如糖尿病等疾病的替代治疗，也是基因治疗的途径，并且还可诱导移植时的免疫耐受。细胞移植的免疫生物学包括下述内个方面：①移植细胞的分离和纯化；②细胞的保存；③移植技术；④对细胞移植的免疫应答；⑤预防同种异体细胞移植的免疫应答；⑥移植细胞产物的释放和调节；⑦异种移植；⑧基因治疗。此文将简要讨论上述问题。     

Mdx小鼠细胞治疗的种子细胞及移植途径研究现状

Duchenne型肌营养不良(简称DMD)是最常见的X连锁隐性遗传性肌病,患病率在男性活产婴儿中约为1/3 500,DMD主要的发病机制是由于抗肌萎缩蛋白(dystrophin)基因突变,导致其基因产物缺失,引起骨骼肌坏死以及功能严重受损.依据DMD的发病机制,有效的治疗途径不外乎改变突变的基因以表达功能正常的蛋白,或者直接将正常的细胞移植发病部位替换病变的细胞.理想的细胞移植治疗就是将具有较好增殖和分化能力的细胞,高效移植入宿主病变器官并发挥治疗作用,但目前细胞移植治疗还处于动物实验研究阶段.Mdx小鼠是DMD最为常见的动物模型,目前对其研究面临的最大问题是选择怎样的种子细胞及移植途径才能达到最理想的治疗效果,国内外许多研究者就此问题进行了探索.本文就近年来的研究成果作一综述.     

树突状细胞与移植排斥研究进展

树突状细胞（ＤＣ）作为专职抗原递呈）细胞，在移植免疫应答中同样起着重要作用。近年来对ＤＣ在移植排斥中作用有了深入了解。本文介绍了移植器官内ＤＣ来尖和类别，移植一ＤＣＲ的移地及其致敏受者Ｔ细胞途径、移植前去除ＤＣ降低移植物免疫原性作用，以及ＤＣ与移植嵌合现象关系和临床移植中控制ＤＣ的策略。     

细胞移植修补心脏

细胞移植修补心脏 ,用以改善受损的心功能 ,受到越来越多的关注 ,简述供移植细胞的来源、细胞移植的方式、细胞移植改善心功能可能的机制以及目前存在的问题等。随着研究的深入 ,细胞移植将会为心衰的防治提供新的途径     

免疫隔离在细胞移植中的应用

免疫排斥是组织移植中的最大障碍。免疫隔离可望解决这一难题，但仅适用于组织及细胞移植，且移植物必须是靠分泌某些活性物质来发挥功能。内分泌腺的移植特别适合应用免疫隔离技术。     

树突状细胞与移植排斥研究进展

树突状细胞（DC）作为专职机原递呈细胞，在移植免疫应答中同样起着重要作用。近年来对DC在移植排斥中作用有了深入了解。本文介绍了移植器官内DC来源和类别，移植后DC的移行及其致敏受者T细胞途径，移植前去除DC降低移植物免疫原性作用，以及DC与移植嵌合现象关系和临床移植中控制DC的策略。     

移植免疫耐受,移植排斥与细胞凋良

同种器官移植后植物有两种结局即移植排斥与免疫耐受。本文综述了细胞凋亡与移植排斥及免疫耐受关系方面的实验及临床研究结果。认为移植物中存在细胞凋亡现象。凋亡细胞可以是移植物实质细胞或间质浸润细胞。移植物实质细胞凋亡,产生移植排斥：间质浸润细胞凋亡,产生免疫耐受。细胞凋亡与Ｆａｓ、ＦａｓＬ关系密切。Ｆａｓ与ＦａｓＬ相互作用可以诱导Ｆａｓ阳性细胞凋亡,介导移植排斥与免疫耐受。     

miR-1231对结肠癌干细胞增殖、凋亡和侵袭的影响

BACKGROUND:Previous studies have found that miR-1231 is down-regulated in colon cancer stem cells (CCSCs), but the effect of miR-1231 on CCSCs remains unclear. OBJECTIVE:To explore the effect of miR-1231 on the proliferation, apoptosis and invasion of CCSCs (CD133⁺CD44⁺). METHODS: CD133⁺CD44⁺ cells and CD133^-CD44^- cells were separated from SW1116 cells by immunomagnetic bead separation. The expression level of miR-1231 in CD133⁺CD44⁺ and CD133^-CD44^- cells was detected by qRT-PCR. miR-1231-overexpressing CD133⁺CD44⁺ cells were transfected with miR-1231 mimics or miR-control by lipofection transfection. The effects of miR-1231 on CD133⁺CD44⁺ cell proliferation, apoptosis and invasion were investigated by MTT, flow cytometry and Transwell assays, respectively. In addition, the expression levels of Ki67, Bax, Bcl-2, MMP-2 and MMP-9 protein in miR-1231-overexpressing CD133+CD44+ cells and control cells were detected by western blot. RESULTS AND CONCLUSION:CD133⁺CD44⁺ and CD133^-CD44^- cells were obtained by the immunomagnetic bead separation. The expression level of miR-1231 in CD133⁺CD44⁺ cells was significantly lower than that in CD133^-CD44^- cells. miR-1231 suppressed CD133⁺CD44⁺ cell proliferation and invasion, but promoted the apoptosis in these cells. Western blot analysis showed that miR-1231-overexpressing CD133⁺CD44⁺ cells had obvious decreases in Ki67, Bcl-2, MMP-2 and MMP-9 protein expression and a significant increase in Bax protein expression compared with control cells. All these results further confirm that miR-1231 inhibits the proliferation and invasion but promotes the apoptosis in CD133⁺CD44⁺ cells. These findings suggest that miR-1231 can be a suppressor of CCSCs, which offers a novel potential therapeutic target for CCSCs and colon cancer.     

miR-486对胶质瘤干细胞的抑制作用

背景：既往研究发现,miR-486在胶质瘤干细胞(CD133⁺)中的表达水平显著低于其在胶质瘤非干细胞(CD133^-)中的表达水平,但是miR-486对CD133⁺细胞的影响尚不明确。 目的：探索miR-486对CD133⁺细胞的作用。 方法：利用流式细胞分选将U87胶质瘤细胞中分为CD133⁺和CD133^-细胞。通过脂质体转染构建miR-486过表达的胶质瘤干细胞。 结果与结论：流式细胞分选和纯化获得高比率的CD133⁺胶质瘤干细胞。实时反转录PCR检测发现miR-486在CD133⁺胶质瘤干细胞的表达水平比CD133^-胶质瘤细胞明显下降。脂质体转染成功构建miR-486过表达的胶质瘤干细胞,体外实验发现miR-486高表达抑制胶质瘤干细胞的增殖,将其阻滞于G₁/S期,并促进凋亡。提示miR-486对胶质瘤干细胞具有抑制作用。中国组织工程研究杂志出版内容重点：干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程     

磁珠分选U251细胞系中的CD133+细胞及其生物学特性

背景：脑肿瘤干细胞理论认为,脑肿瘤干细胞是脑肿瘤细胞中“种子”细胞,是脑肿瘤发生、浸润和复发的关键细胞。 目的：观察人多形性胶质母细胞瘤U251细胞系中CD133⁺细胞的增殖、分化及体内致瘤性等生物学特性。 方法：运用磁珠分选技术分选U251细胞系中的CD133⁺和CD133^-细胞亚群;MTT法绘制两个亚群细胞的生长曲线;单克隆形成率实验检测2个亚群细胞的增殖能力;免疫荧光检测CD133⁺细胞亚群的多向分化能力;裸鼠移植实验检测2个亚群细胞在裸鼠体内致瘤性的差异。 结果与结论：U251细胞系中只有约4.5%的CD133⁺细胞;分选后的CD133⁺细胞能增殖形成典型的脑肿瘤干细胞球,生长曲线显示CD133⁺细胞增殖能力明显强于CD133^-细胞;单克隆形成率实验显示CD133⁺细胞能形成脑肿瘤干细胞球的细胞比率达到78.5%~92.4%,而CD133^-细胞仅有0.8%~2.4%;CD133⁺细胞能分化为具有GFAP和NeuN成熟表型的肿瘤细胞;CD133⁺的致瘤率为71.42%,而CD133^-细胞无致瘤性。提示U251细胞系中存在少量具有增殖、多向分化与体内致瘤能力的CD133⁺细胞,CD133⁺细胞是符合肿瘤干细胞定义的细胞亚群。     

当归多糖对人白血病干细胞体外增殖与体内移植模型的影响

目的 探讨当归多糖(ASP)对人白血病干细胞(LSCs)增殖及体内移植的影响.方法 1.ASP 对CD34+CD38-人LSCs体外增殖的影响.免疫磁性法分选正常人和髓系白血病患者骨髓中CD34+CD38-细胞,分为正常CD34+CD38-对照组、CD34+CD38-LSCs对照组、正常CD34+CD38-ASP组和C...     

欧前胡素通过下调c-met的表达提高肺癌CD133~+细胞亚群对吉非替尼的敏感性

目的:研究PC9 CD133~+细胞亚群对吉非替尼的耐药性并探讨欧前胡素提高吉非替尼抗肺癌活性的机制。方法:MTT法检测PC9细胞在吉非替尼和欧前胡素处理下的细胞活力。Western blot实验检测吉非替尼和欧前胡素对PC9细胞c-met表达水平、caspases活化水平及表皮生长因子受体(EGFR)、PI3K、AKT磷酸化水平的影响。流式细胞术检测欧前胡素和吉非替尼对PC9细胞系的CD133~+细胞亚群种群比例的影响及PC9细胞在二者处理下的凋亡率。结果:PC9 CD133~+细胞亚群对吉非替尼的敏感性显著低于PC9 CD133~-细胞亚群。吉非替尼能显著抑制PC9 CD133~-细胞亚群EGFR/PI3K/AKT的活化,但对PC9 CD133~+细胞亚群该通路的影响不大。吉非替尼单独处理能提高PC9细胞系中CD133~+细胞亚群的比例,然而联用欧前胡素后PC9 CD133~+细胞亚群的种群比例显著下降。Western blot实验表明欧前胡素能显著降低PC9 CD133~+细胞亚群的c-met蛋白表达水平表明c-met是欧前胡素的治疗靶点。MTT、Western blot、流式细胞术实验结果表明在PC9 CD133~+细胞亚群中,欧前胡素通过抑制c-met的表达提高吉非替尼对PI3K/AKT的抑制作用,从而诱导PC9 CD133~+细胞亚群发生caspases活化和凋亡。结论:欧前胡素通过下调c-met的表达提高肺癌CD133~+细胞亚群对吉非替尼的敏感性,两者存在协同抗肿瘤效应。     

肝癌干细胞分离及其细胞生物学特性

背景：正常干细胞和肿瘤干细胞在基因表达和依赖的细胞信号通路上应该存在不同,如何发现能选择性杀伤肿瘤干细胞的治疗手段是一个仍然需要大量研究的课题。 目的：分离人肝癌细胞系肿瘤干细胞MHCC97,分析其细胞生物学特性。 方法：采用流式细胞技术在人高转移肝癌细胞系MHCC97中筛选肿瘤干细胞,分离正常人肝脏干细胞CD133^-CD34^- MHCC97和人肝癌细胞系肿瘤干细胞CD133⁺CD34⁺ MHCC97,分别检测其表型、生长曲线、细胞周期和多系分化能力。 结果与结论：人肝癌细胞系CD133⁺CD34⁺ MHCC97肿瘤干细胞的表型为CD133⁺CD34⁺ ,人肝癌细胞系CD133⁺CD34⁺ MHCC97具有与CD133^-CD34^- MHCC97相似的细胞曲线和生长周期,可以向上皮和内皮细胞分化,并表达相应特异性的分子标志。提示人肝癌细胞系中CD133⁺CD34⁺MHCC97细胞具有肿瘤干细胞的特性,可以向内皮和上皮细胞分化,同时具有肿瘤干细胞的生物学特性,是肿瘤复发转移的根源,也是临床治疗的靶点。中国组织工程研究杂志出版内容重点：干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程全文链接：     

白藜芦醇通过促进Apaf-1/caspase-9复合物的形成增强5-氟尿嘧啶对骨肉瘤CD133~+细胞亚群凋亡的诱导

目的:探讨白藜芦醇联合化疗药物5-氟尿嘧啶对骨肉瘤CD133~+细胞亚群的协同杀伤效应及机制。方法:将人骨肉瘤细胞系MG-63 CD133~+细胞亚群及相应CD133~-细胞亚群用5-氟尿嘧啶及白藜芦醇进行体外处理。MG-63细胞的相对细胞活力用MTT法进行检测;细胞凋亡率用流式细胞术进行检测;用Western blot实验检测caspase-9和caspase-3的活化、Apaf-1的表达水平及细胞色素C的释放;用免疫共沉淀法检测Apaf-1与caspase-9前体的相互作用。结果:5-氟尿嘧啶对MG-63 CD133~+细胞亚群的杀伤活性和凋亡诱导活性均显著低于MG-63 CD133~-细胞亚群。但联用白藜芦醇能显著提高5-氟尿嘧啶对MG-63 CD133~+细胞亚群的细胞活力抑制率。白藜芦醇处理能显著上调MG-63 CD133~+细胞亚群中Apaf-1的表达水平,在MG-63 CD133~+细胞亚群中转染Apaf-1siRNA后,5-氟尿嘧啶联用白藜芦醇的协同效应受到显著抑制。另外,免疫共沉淀实验结果表明白藜芦醇联合5-氟尿嘧啶能显著诱导MG-63 CD133~+细胞亚群中Apaf-1/caspase-9复合物的形成,从而诱导caspase-9发生活化。结论:白藜芦醇通过促进Apaf-1/caspase-9复合物的形成,增强5-氟尿嘧啶对骨肉瘤CD133~+细胞亚群凋亡的诱导。     

骨髓间充质干细胞调节心脏移植大鼠的免疫功能

BACKGROUND:Heart transplantation is an effective method for treatment of end-stage heart failure, but immune rejection that seriously impact therapeutic effacicy is easy to occur after transplantation. OBJECTIVE:To investigate the regulatory effect of bone marrow mesenchymal stem cells on the immune function of rats undergoiong heart transplantation. METHODS:Twenty Lewis rats were enrolled as donors, and 20 Wistar rats as recipients. Heart transplantation models were established in the Wistar rats. These 20 model rats were randomized into cell transplantation and control group with 10 rats in each group. Forty-eight hours after heart transplantation, rats in the cell transplantation group were given bone marrow mesenchymal stem cell suspension (1 mL, 2×10⁸ cells/L) via the tail vein, while rats in the control group were given normal saline in the same dose. Then, the expression levels of serum interleukin-2, interleukin-10 and percentage of CD4⁺, CD8⁺, CD4⁺/CD8⁺, CD4⁺CD25^high, CD4⁺CD25^high Foxp3⁺ T cells in the venous blood were detected in the two groups at 7 days after cell transplantation. Additionally, rat myocardial tissues were taken and observed pathologically. RESULTS AND CONCLUSION:The survival time of the cell transplantation group was significantly longer than that of the control group (P < 0.05). The expression level of interleukin-2 showed no significant difference between the two groups (P > 0.05), but the level of interleukin-10 in the cell transplantation group was significantly higher than that in the control group (P < 0.05). Compared with the control group, the percentage of CD4⁺/CD8⁺, CD4⁺CD25^high, CD4⁺CD25^high Foxp3⁺ and CD4⁺ T cells was significantly higher, and the percentage of CD8⁺ T cells was significantly lower in the cell transplantation group (P < 0.05). Histopathological findings showed that there were a small amount of infiltrated lymphocytes in the cell transplantation group with the presence of slight bleeding and edema, and these inflammatory reactions were milder than those in the control group. These findings indicate that bone marrow mesenchymal stem cell transplantation can effectively reduce the rejection in rats undergoing heart transplantation.     

“Nk-Like” T Cytotoxicity Against B Lymphocytes in a Hypogammaglobulinemic Patient

Physiologically, cells with NK activity appear to exert a negative control on immunoglobulin production. The clinical association of large granular lymphocyte (LGL) proliferation with hypogammaglobulinemia suggests that these functional NK cells could also be involved in pathological situations.

We studied in vitro lymphocyte functions in a patient presenting LGL proliferation associated with hypogammaglobulinemia. The CD3⁺ CD8⁺ CD57⁺ CD16^- phenotype lymphocytes expressed a high NK type cytotoxicity towards K562 targets, suggesting that they may be considered as “NK-like” T cells. We cultured the patient peripheral blood mononuclear cells (PBMC) with control subject PBMC and with PBMC from two other subjects with B chronic lymphocytic leukemia (B- CLL) of the CD20⁺ CD21^- CD10^- phenotype. Patient PBMC exhibited a lytic activity on control PBMC and on the B lymphocytes of one of the two B- CLL but only in the presence of PWM. This activity was not exerted by the culture supernatant and required a cell - to - cell contact. We suggest that the hypogammaglobulinemia observed in this patient may be related to a cytotoxic effect exerted on B lymphocytes by a CD3⁺ CD8⁺ CD57⁺ CD16^- LGL proliferation.     

恶性肿瘤干细胞标记物CD34在鼻咽癌细胞系的表达

BACKGROUND:Previous research have confirmed that CD34 is closely related to oncogenesis, progress, recurrence, metastasis and drug-resistance of various cancers, but its role in nasopharyngeal carcinoma remains unclear. OBJECTIVE:To sort cells positive and negative for CD34 in nasopharyngeal carcinoma cell lines and to detect cell proliferation and migration. METHODS:Expressions of CD34 in nasopharyngeal carcinoma cell lines 5-8F, 6-10B, CNE1 and CNE2 were detected by flow cytometry. And CD34+ and CD34^- cells were sorted based on cell surface markers for purity identification. Afterwards, proliferation and migration of CD34⁺ and CD34^- cells were detected by MTT assay, colony-formation assay and scratch assay. RESULTS AND CONCLUSION:All four nasopharyngeal carcinoma cell lines expressed CD34 in 0.1%-0.2%, and the level of CD34 was closely related to the cell growth density. The purity of CD34⁺ cell was more than 98% in the sorted CD34⁺ cell populations, but no CD34⁺ cells were found in the sorted CD34^- cell populations. At 1, 3, 5 and 7 days the proliferation rate of CD34⁺ cell, populations was significantly higher than that of CD34^- cells (P < 0.05). Consistently, the colony-formation efficiency of CD34⁺ cell was significantly higher than that of CD34^- cells (P < 0.05). Moreover, CD34⁺ cells migrated significantly faster than CD34^- cells by scratch assay (P < 0.05). In conclusion, CD34⁺ cells cultured in vitro display higher proliferation and migration capacities, indicating that CD34⁺ cells have the potential of nasopharyngeal carcinoma stem cells.