王贤，张培华

(广东医学院附属医院整形外科研究所，广东 湛江 524023)

脂肪干细胞的研究及应用进展

王贤，张培华

(广东医学院附属医院整形外科研究所，广东 湛江 524023)

干细胞在治疗各种疾病方面拥有巨大的潜能。在细胞治疗方面，脂肪干细胞是最有前景的种子细胞之一，其他的还包括胚胎干细胞和诱导的多能干细胞。胚胎干细胞和诱导的多能干细胞都具有多向分化潜能，因而起着举足轻重的作用。但是，胚胎干细胞和诱导的多能干细胞分别受限于伦理学问题及细胞表型的鉴定。脂肪干细胞则不受这些限制，不仅易于获取，而且方便扩增。在不同的诱导条件下，脂肪干细胞能分化为脂肪细胞、成骨细胞、软骨细胞、肌肉细胞、内皮细胞和神经细胞，这些分化潜能可应用于再生医学领域，如：皮肤重建，骨和软骨修复等。本篇综述着重讨论目前脂肪干细胞的分离、分化和治疗应用。

脂肪干细胞；分离；分化；应用

干细胞包括胚胎干细胞、诱导的多能干细胞和成体干细胞。胚胎干细胞能自我更新，并且能分化为人体的任何一种细胞。但由于涉及伦理问题，胚胎干细胞应用于临床研究存在很大的争议，而诱导的多能干细胞和成体干细胞则没有这方面的问题。自从Zuk等[1]发现脂肪组织中含有多分化潜能的干细胞，脂肪干细胞就逐渐成为了成体干细胞中最有前景的种子细胞之一。脂肪组织通过脂肪抽吸术或者外科手术易于获取，对供体的创伤相对较小，而且不涉及伦理问题。在体外，脂肪干细胞能大量扩增，操作简便，并能在相应的诱导条件下分化为脂肪细胞、成骨细胞、软骨细胞、肌肉细胞、内皮细胞、神经细胞等[1-10]。因此，同其他种子细胞相比，脂肪干细胞所受限制小，具有更加广阔的临床应用前景。

1 脂肪干细胞的分离和培养

同其他干细胞相比，来源于脂肪组织的干细胞体内含量更大。目前，从脂肪组中分离脂肪干细胞不仅可以人工分离，也可以采用仪器自动离心分离。为了大量获得脂肪干细胞，目前大多是在脂肪组织离心分离之后用胶原酶消化[3-10]。Zeng等[11]报道了另一种方法，即组织块贴壁培养法，是一种更加高效而且快速的脂肪干细胞分离培养方法。上述两种方法所获得的脂肪干细胞，在倒置相差显微镜下观察，外形都呈成纤维细胞样，可融合成鱼群状或者旋涡状。

2 脂肪干细胞的多向分化潜能

脂肪干细胞具有多向分化潜能，在一定的诱导条件下，可以分化为脂肪细胞、成骨细胞、软骨细胞、肌肉细胞、内皮细胞和神经细胞[1-10]。但目前对于脂肪干细胞分化的信号通路尚不十分清楚，有待进一步研究[12-15]。分化为脂肪细胞[2-3]：需要额外添加地塞米松、胰岛素、3-异丁基-1-甲基黄嘌呤(IBMX)、吲哚美辛作为诱导条件，成脂分化可以通过油红染色验证。分化为成骨细胞[2-3]：需要额外添加地塞米松、维生素C、β-磷酸甘油作为诱导条件，成骨分化可以通过茜素红染色验证。分化为软骨细胞[2]：需要额外添加胰岛素、转化生长因子β1(TGF β1)和维生素C作为诱导条件，成软骨分化可以通过阿尔辛兰染色验证。分化为肌肉细胞[3-4]：需要额外添加5-氮杂胞苷、马血清作为诱导条件，成肌肉分化可通过检测肌蛋白的生成以及体外肌小管形成实验来验证。分化为内皮细胞[5-6]：需要在含有多种因子的内皮细胞生长培养基(EGM-2)中添加大剂量的血管内皮生长因子(VEGF)，成内皮分化可以通过检测内皮细胞的细胞表面标志CD31、CD105、vWF以及体外血管形成实验来验证。分化为神经细胞[7-10]需要额外添加β-硫基乙醇、维甲酸、血小板源性生长因子(PDGF)、碱性成纤维细胞生长因子(bFGF)、毛喉素、神经胶质生长因子2(GGF-2β)，成神经分化可以通过检测神经细胞标志如GFAP、S100等来验证。

3 脂肪干细胞的临床应用

脂肪干细胞储量大、增殖快，而且具有自我更新和多向分化能力，因此临床应用前景广阔[16]。脂肪干细胞分化形成的脂肪细胞可用于软组织缺损、隆乳、脂肪性营养不良以及软组织填充美容。在软组织重建方面，自体脂肪移植已广泛应用，但是其局限性依旧存在，其中最明显的就是组织吸收，导致移植的脂肪量减少20%～90%[17]。解决这一问题的关键，是形成新生血管，从而给移植组织提供营养支持。脂肪干细胞可以分泌多种血管生长因子，包括血管内皮生长因子(VEGF)[18]，从而刺激新血管形成。

脂肪干细胞在体内和体外都能够分化为成骨细胞[2-3，19]，成骨细胞可做为细胞材料。脂肪干细胞能与多种生物支架结合，促进新骨生成，用于治疗骨缺损[20-22]。一定条件下，脂肪干细胞在体内和体外都可以分化为软骨组织[2，23]。这就给利用细胞治疗来修复关节软骨的缺陷提供了可能，如：修复骨性关节炎。脂肪干细胞能够增强肌肉细胞的分化，提高肌肉的修复能力[24]。移植的脂肪干细胞，可以在受损的心肌中，促进血管生成、神经出芽、心肌细胞再生，并保护心功能[25-26]。脂肪干细胞在肌肉重建方面已经取得了一些可喜的成绩，进一步的研究有待深入[27]。脂肪干细胞能够分化为内皮细胞，并呈现为小管样结构[5-6]，脂肪干细胞还可以促进血管的形成[28-29]，这给缺血性疾病的治疗提供了新的方向。脂肪干细胞能够分化为神经细胞和神经胶质细胞[7-10]。脂肪干细胞通过分泌一些神经生长因子，还能促进周围神经损伤的修复[30-31]。脂肪干细胞还有望应用于神经退行性病变的治疗，如：帕金森病、阿尔茨海默病、亨廷顿氏舞蹈症等[32-35]。

4 总 结

干细胞将成为细胞治疗的有力武器，应用于各种疾病，但仍存在许多挑战，尤其是干细胞临床应用的安全性和有效性问题，还有同种异体的移植免疫排斥问题等。脂肪干细胞易于大量获取，损伤相对较小，并且安全有效，其临床应用正与日俱增。脂肪干细胞的分离培养和分化诱导方法正在不断改进。脂肪干细胞尚未发现特异性的细胞表面标志物，通过细胞表面标志物纯化脂肪干细胞的方法有待改善。

目前关于脂肪干细胞各方面的研究正在不断深入，各种临床应用问题也在不断被克服，脂肪干细胞的临床应用具有广阔的前景。

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Research and application progress of adipose-derived stem cells.

WANG Xian,ZHANG Pei-hua.Plastic Surgery Institute,the Affiliated Hospital of Guangdong Medical College,Zhanjiang 524023,Guangdong,CHINA

Stem cells have a great potential to treat various kinds of diseases.For these cell-based therapies,adipose-derived stem cells(ADSCs)are one of the most promising seed cells,with others including embryonic stem cells (ESCs)and induced pluripotent stem cells(iPSCs).ESCs and iPSCs play important roles due to their pluripotency.However,ESCs and iPSCs have limitations in ethical issues and in identification of cell phenotypes,respectively.Unlike ESCs and iPSCs,ADSCs do not have such limitations,which are not only easily obtained but also uniquely expandable. ADSCs can differentiate into adipocytes,osteoblasts,chondrocytes,myocytes,endotheliocytes and neurons under different differentiation conditions,and these kinds of differentiation potential ofADSCs could be applied in regenerative medicine e.g.,skin reconstruction,bone and cartilage repair.In this review,the current status of ADSC isolation,differentiation and their therapeutic applications are discussed.

Adipose-derived stem cells;Isolation;Differentiation;Application

R329.2+1

A

1003—6350（2016）06—0965—03

10.3969/j.issn.1003-6350.2016.06.037

2015-10-13)

广东省自然科学基金(编号：2014A030313535)

张培华。E-mail：zhangph1128@126.com