脂肪组织来源的干细胞的研究进展

来自中胚层的脂肪组织与骨髓组织一样含有大量能自我更新和多向系分化潜能的细胞群称为脂肪干细胞。其取材方便，来源丰富，可在体外稳定增殖传代。并与骨髓间充质干细胞有相似的多向分化表面标志CD105、STRO-1以及CD166受体。研究发现它具有多向系分化潜能，除可以分化为问充质来源的脂肪、骨，软骨、脂肪以及骨骼肌、心肌等细胞，也可诱导分化为来源于外胚层的神经细胞以及具有功能性的血管内皮细胞，用以修复骨、软骨、心肌、骨骼肌、血管以及神经等组织。其具有造血支持作用以及可被逆转录病毒、腺病毒以及慢病毒较高的转染效率等优点，可以作为基因转移良好的靶细胞。因而，脂肪来源的干细胞其有望成为组织工程、细胞治疗以及基因转染良好的种子细胞。     

脂肪来源间充质干细胞的生物学特性及临床应用前景

组织工程技术发展的关键是获得具有来源方便、广泛、具有分化能力的种子细胞。脂肪来源间充质干细胞的发现及研究,证明了它可以充当这一角色。它具有多项分化潜能,在合适的诱导条件下可以分化为骨细胞、软骨细胞、神经前体细胞、心肌细胞等。它的这些分化能力,给临床上一些疾病如组织缺损性疾病,心肌梗死,基因疾病的治疗带来了曙光。脂肪来源的干细胞,因其取材方便、安全和扩增速率高等特点必将在细胞治疗和组织工程方面有更广阔的应用前景。     

脂肪干细胞在软骨组织工程中的应用

软骨是最早应用组织工程技术成功构建的组织之一,但由于缺乏合适的软骨构建种子细胞,因此其发展相对落后。随着干细胞研究的兴起,脂肪干细胞（ASC）因其具有分布广泛、可利用细胞量大、取材容易等优点,为ASC作为种子细胞应用于组织工程研究提供了可能;但是ASC构建软骨组织的效果却不如骨髓间充质干细胞（BMSC）理想。因此ASC在软骨组织工程中的应用仍面临着诸多问题与挑战,其中最核心的问题是如何提高ASC成软骨的效率。为此从如何纯化脂肪来源细胞、尽可能保持其中干细胞的生物学特性并优化软骨诱导方案3个方面予以综述,为提高ASC成软骨的效率提供参考。     

脂肪干细胞在骨科基础研究中的新进展

背景：自脂肪来源干细胞被发现以来,培养传代及在支架材料上的生长能力就越来越受到骨科相关研究的关注。 目的：探讨脂肪来源干细胞的生物学特性及在骨科基础研究中的进展。 方法：由第一作者检索2001/2010 ISI Web of knowledge 数据库(网址http://www.isiknowledge.com)和PubMed 数据库(网址 http://www.ncbi.nlm.nih.gov/PubMed)有关脂肪来源干细胞培养特性和组织修复的相关文献并进行综合分析,检索词为“adipose derived stem cells,orthopaedic”。 结果与结论：脂肪来源干细胞具有与骨髓间充质干细胞相似的生物学特性及分化潜能,并且在骨骼、软骨、椎间盘、肌腱和韧带等组织的修复方面表现出良好的治疗效果。同时,脂肪来源干细胞在体内广泛分布,获取容易,为其在骨科相关疾病的修复治疗中奠定了良好的应用前景。     

脂肪基质干细胞的研究进展

脂肪基质干细胞是存在于脂肪组织中的一群能多向分化的细胞。它们具有与骨髓间充质干细胞相似的特性,在不同的诱导条件下能向不同组织细胞分化。脂肪基质干细胞也具有一定的免疫负调节作用,能用于造血干细胞移植后的造血支持。     

脂肪源性成熟基质细胞-干细胞研究的新热点

曾被认为是废物的脂肪组织正成为国外生物学上新的研究热点。近年来，在脂肪组织中发现一新的成体干细胞：脂肪源性成熟基质细胞（adipose-derived adult stromal cells，ADAS cells），因其具有易获取，易扩增的特性和多向分化的潜能而倍受关注。     

人髌下脂肪垫来源脂肪间充质干细胞的分离、培养及鉴定

背景：髌下脂肪垫在膝关节手术中经常要部分切除,其可以作为脂肪间充质干细胞的重要来源。 目的：探讨自髌下脂肪垫中分离、培养脂肪间充质干细胞的策略及细胞分子表面标记情况。 方法：髌下脂肪垫组织取自膝关节镜手术的患者,以Ⅰ型胶原酶消化消化脂肪组织获取干细胞,用10%低糖DMEM培养基培养,利用MTT法测定不同代细胞增殖情况并绘制生长曲线。检测第5代细胞表面CD29及CD44的表达。 结果与结论：培养24 h后可见原代细胞贴壁,1周后细胞呈纺锤型并且增殖速度加快,传代后的细胞贴壁及增殖细胞速度加快。生长曲线示第2及第5代的细胞增殖能力明显较第8代能力强。所取细胞能够分化为骨细胞和脂肪细胞。流式细胞仪检测结果显示第5代脂肪间充质干细胞重96.8%表达CD29,97.6%表达CD44。提示自髌下脂肪垫分离及提取脂肪干细胞简单易行,所得细胞的纯度及增殖能力均符合组织工程种子细胞的基本条件。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     

来源于脂肪的基质干细胞研究现状

源于中胚层的脂肪组织所含的干细胞具有多向分化的潜能，在一定的诱导备件下能分化为脂肪细胞、软骨细胞、成骨细胞，成肌细胞、心肌细胞和神经细胞，并具有自体来源、容易获取、再生能力强、低衰老性等特点，并有望成为人体最大的自体干细胞来源库，广泛应用于临床细胞治疗和组织工程。     

人脂肪来源间充质干细胞成骨及成软骨的潜能

背景：人脂肪来源间充质干细胞是种具有较强的体外增殖和多系分化能力的成体干细胞,可以从美容吸脂手术中获得,取材方便,原料来源丰富,在生物治疗应用方面蕴藏着巨大的价值。 目的：体外分离、培养人脂肪来源间充质干细胞,探讨其基本生物学特性及成骨成软骨的潜能。 方法：取美容吸脂获得的脂肪组织,采用Ⅱ型胶原酶消化法分离人脂肪来源间充质干细胞并进行体外培养;观察细胞形态、测定细胞周期、流式细胞仪鉴定细胞表面标志;取第3代细胞,分别加入成骨诱导培养基及成软骨诱导培养基行体外成骨及成软骨诱导。 结果与结论：体外培养人脂肪来源间充质干细胞呈纤维样形态,原代细胞24 h内贴壁,培养5~7 d 后开始形成细胞集落;经细胞周期检测显示G₀/G₁,S和G₂/M所占比例分别为(88±2)%,(12±2)%和0.03%。经流式细胞仪检测CD29和CD105呈阳性表达,CD34和CD45呈阴性表达。RT-PCR检测显示,人脂肪间充质干细胞经成骨诱导分化后细胞中骨桥蛋白mRNA呈阳性表达,经软骨诱导分化后细胞中Ⅱ型胶原mRNA呈阳性表达。结果证实,实验成功体外分离、培养人脂肪来源间充质干细胞,其具有向成骨细胞和软骨细胞分化的潜能。     

兔脂肪源性干细胞的成骨诱导及其与异种松质骨的生物相容性研究

探讨异种松质骨作为支架材料的前景,并观察其与成骨诱导前后脂肪源性干细胞的生物相容性。取兔脂肪源性干细胞,诱导成骨并检测。将诱导成骨和未诱导的脂肪源性干细胞接种至松质骨支架上。扫描电镜观察,并计算细胞24 h的贴壁率。将细胞-支架复合物及单纯支架分别植入家兔皮下,10天后处死取材,HE染色行组织学分析。结果表明:兔脂肪源性干细胞在体外诱导培养后,碱性磷酸酶含量升高,诱导2周后茜素红染色阳性,提示脂肪源性干细胞向成骨方向分化。扫描电镜观察显示:诱导成骨和未诱导的脂肪源性干细胞均能与异种松质骨支架材料复合良好,二者24 h的贴壁率差别无统计学意义。由此说明:脂肪源性干细胞能够在体外诱导成骨,且异种松质骨支架材料与其有良好的生物相容性,提示脂肪源性干细胞能够作为骨组织工程的种子细胞,同时也提示异种松质骨能够作为支架材料用于骨组织工程研究。     

Application of Whole Mouse Embryo Culture System on Stem Cell Research

The normal development of mouse embryo in vivo could be maintained in vitro up to 72 h in the presence of rat serum which is continuously supplied with the appropriate concentration of O₂ and CO₂. There are several applications of the whole mouse embryo culture model for study of cellular dynamics in hematopoiesis and its interaction with vasculogenesis. In this protocol, we have described details of manipulation techniques in combination with the whole embryo culture and also some advance techniques applied to the mouse embryo such as intra-cardiac inoculation of acetylated low density lipoprotein for cell-specific labeling and engraftment of donor yolk-sac from different genotype/phenotype mouse embryo onto the yolk-sac of host mouse for study of the dynamic distribution of hematopoietic cell.

Synergistic Effect of Biochemical Factors and Strain on the Smooth Muscle Cell Differentiation of Adipose-Derived Stem Cells on an Elastic Nanofibrous Scaffold

Abstract

Adipose-derived stem cells (ASCs) have been the subject of many tissue-engineering studies, mainly because of their multipotential properties. We have focused on the potential of human ASCs (hASCs) for smooth muscle cell (SMC) differentiation in cardiovascular tissue engineering. In this study, we investigated the combined effect of differentiation factors along with strain utilizing a customized device on hASCs proliferation and consequent differentiation into SMCs on an elastic nanofibrous scaffold. The cell proliferation increased in strain-stimulated culture and was more affected by media composition as compared to in static culture. Differentiation factors did not have an influence on SM α-actin (α-SMA) and myosin heavy chain (MHC) expression in static culture. However, α-SMA and MHC expression were affected by differentiation factors in strain culture, in particular, showing that treatment with retinoic acid significantly increased the expression of α-SMA (3.6-fold) and MHC (2-fold) as compared to strain alone. This study demonstrated the synergic effect of strain and biochemical factor on the SMC differentiation of hASCs and provided a useful method for the application of stem cells in mechano-active tissue engineering.     

Product Regulation and the Clinical Translation of Stem Cell Research

The recent approval by the United States Food and Drug Administration of a clinical trial involving a product derived from human embryonic stem cells, along with recent concerns about unproven stem cell therapies being offered to patients, highlight the importance of regulation at the critical stage of beginning human trials of novel therapies. The regulations governing therapeutic products (drugs and related products) are one part of the broader legal framework, but will play an increasingly prominent role as we move into clinical translation. The classification of products as drugs or biologics, on one hand, or minimally manipulated cell and tissue products for homologous use, on the other, will determine the requirements that will apply, including whether use in clinical trials requires approval. Product regulation works alongside other parts of the legal and policy framework, notably research ethics review and legal responsibilities of medical professionals, that play important though limited roles. Three key developments and challenges currently facing product regulation and related areas will affect stem cell research in this phase: regulatory reform, fragmentation, and capacity.     

红景天苷对新生大鼠海马区神经干细胞分化的研究

目的研究红景天苷药物血清诱导新生大鼠海马神经干细胞向神经元方向分化,并促进所分化神经元细胞发育的作用。方法从新生24h内的Wistar大鼠脑中分离扩增获得大量神经干细胞后,加入低、中、高剂量的红景天苷药物血清及对照血清,观察其对神经干细胞分化为神经元的影响,并通过免疫细胞化学染色检测神经干细胞分化为神经元的状况。结果各药物实验组诱导神经干细胞分化为NSE阳性细胞的个数面积与周长明显高于对照组(P<0.05),分化为GFAP阳性细胞的个数明显低于对照组,且呈量效依赖关系。结论红景天苷药物学清在体外可促进神经干细胞向神经元方向分化,在一定范围内存在量效依赖关系,对所分化的神经元细胞有促生长发育作用。     

Pressure and Distortion Regulate Human Mesenchymal Stem Cell Gene Expression

While the concept that physical forces such as tension and compression are involved in mature tissue modeling is widely accepted, the role of these specific types of mechanical loading in the differentiation and maturation of uncommitted cell types like human mesenchymal stem cells (hMSCs) is currently unknown. We observed that hMSCs have the fundamental ability to distinguish between dynamic tensile and compressive loading by regulating distinct gene expression patterns and that these differences in gene expression can be related to conformational changes in cell shape and volume. Dynamic tension was found to regulate both fibroblastic and osteogenic associated genes while dynamic compression up-regulated genes associated with chondrogenesis. Identifying genes involved in the mechanotransduction of different modes of physical loading in hMSC may greatly enhance the ability to rationally design tissue regeneration systems to restore proper tissue function.     

整合素传递力学信号影响间充质干细胞分化的研究进展

简述硬度的可控性及其控制方法、不同硬度调控间充质干细胞(mesenchymal stem cells,MSCs)分化的相应方向和整合素在硬度调控MSCs分化的信号通路中的作用。其中,重点说明整合素在硬度调控MSCs分化的信号通路中的作用。硬度调控MSCs分化的信号通路包括:Rho/ROCK信号通路、整合素/FAK信号通路、ERK信号通路、JNK信号通路、Wnt-β-catenin信号通路和PI3K/Akt信号通路等。而整合素作为跨膜异二聚体糖蛋白,参与部分信号通路传递力学信号给MSCs。不同的整合素家族参与不同的信号通路来调控MSCs向不同方向分化,且这些信号通路间存在相互影响。研究结论为组织修复、器官再造和再生医学等方面的应用提供理论依据。     

Early Cytomegalovirus Reactivation Leaves a Specific and Dynamic Imprint on the Reconstituting T Cell Compartment Long-Term after Hematopoietic Stem Cell Transplantation

Human cytomegalovirus (CMV) reactivation frequently occurs during the early phase of immune recovery after allogeneic hematopoietic stem cell transplantation (HSCT). Whereas the recovery of virus-specific immunity in the early phase after HSCT is extensively studied, the impact of CMV on the reconstitution and composition of the T cell compartment long-term after HSCT is unknown. We analyzed T cell reconstitution 1 to 2 years after HSCT in 131 pediatric patients. One year after HSCT, patients with early CMV reactivation (n = 46) had 3-fold higher CD8⁺ T cell numbers (median, 1323 versus 424 cells/μL; P < .0001) compared with patients without CMV reactivation (n = 85). This effect, caused by a major expansion of CD8⁺ effector memory (EM) and end-stage effector (EMRA) T cells, was independent of pretransplantation donor and recipient CMV serostatus and not seen after Epstein-Barr virus or adenovirus reactivations. At 1 and 2 years after HSCT, the absolute numbers of CD8⁺ naive and central memory T cells, as well as CD4⁺ naive, CM, EM, and EMRA T cells, did not differ between patients with or without CMV reactivation. In the second year after HSCT, a significant contraction of the initially expanded CD8⁺ EM and EMRA T cell compartments was observed in patients with early CMV reactivation. In conclusion, CMV reactivation early after pediatric HSCT leaves a specific and dynamic imprint on the size and composition of the CD8⁺ T cell compartment without compromising the reconstitution of CD8⁺ and CD4⁺ naive and central memory T cells pivotal in the response to neo and recall antigens.     

骨髓间充质干细胞定向诱导成类软骨细胞的实验研究

目的　体外诱导骨髓间充质干细胞 (MSCs)向软骨细胞表型分化 ,探讨其分化的条件 ,为其作为软骨组织工程种子细胞提供实验基础。方法　取第二代成年大鼠骨髓间充质干细胞 ,实验组用无血清培养液诱导 ;对照组用含 10 %胎牛血清的培养液自然分化。Ⅱ型胶原免疫组化、甲苯胺蓝染色、透射电镜检测其分化情况。统计学分析不同时间、不同接种密度的软骨分化率。结果　诱导后的MSCs具有软骨细胞特点 ,不同诱导时间、不同接种密度的Ⅱ型胶原免疫组化阳性率有显著差异。诱导 14d为最佳 ,诱导率与细胞接种密度成正相关。结论　成年大鼠骨髓间充质干细胞在无血清培养基能向软骨细胞方向转化。     

Participation in Clinical Research: Perspectives of Adult Patients and Parents of Pediatric Patients Undergoing Hematopoietic Stem Cell Transplantation

Despite major improvements over the past several decades, many patients undergoing hematopoietic stem cell transplantations (HSCT) continue to suffer from significant treatment-related morbidity and mortality. Clinical research studies (trials) have been integral to advancing the standard of care in HSCT. However, 1 of the biggest challenges with clinical trials is the low participation rate. Although barriers to participation in cancer clinical trials have been previously explored, studies specific to HSCT are lacking. The current study was undertaken to examine the knowledge, attitudes, and perceptions of HSCT patients regarding clinical trials. As members of focus groups, participants responded to open-ended questions that assessed factors influencing decision-making about HSCT clinical trials. Suggestions for improvements in the recruitment process were also solicited among participants. Seventeen adult HSCT patients and 6 parents of pediatric HSCT patients participated in the study. The median age was 56 years (range, 18 to 70) and 44 years (range, 28 to 54) for adult patients and parents, respectively. Participants universally indicated that too much information was provided within the informed consents and they were intimidated by the medical and legal language. Despite the large amount of information provided to them at the time of study enrollment, the participants had limited knowledge retention and recall of study details. Nevertheless, participants reported overall positive experiences with clinical trial participation and many would readily choose to participate again. A common concern among participants was the uncertainty of study outcome and general lack of feedback about results at the end of the study. Participants suggested that investigators provide more condensed and easier to understand informed consents and follow-up of study findings. These findings could be used to help guide the development of improved consent documents and enhanced participation in research studies, thereby affecting the future design of HSCT research protocols.