脂肪干細(xì)胞的研究及應(yīng)用進(jìn)展

王賢，張培華

(廣東醫(yī)學(xué)院附屬醫(yī)院整形外科研究所，廣東 湛江 524023)

脂肪干細(xì)胞的研究及應(yīng)用進(jìn)展

王賢，張培華

(廣東醫(yī)學(xué)院附屬醫(yī)院整形外科研究所，廣東 湛江 524023)

干細(xì)胞在治療各種疾病方面擁有巨大的潛能。在細(xì)胞治療方面，脂肪干細(xì)胞是最有前景的種子細(xì)胞之一，其他的還包括胚胎干細(xì)胞和誘導(dǎo)的多能干細(xì)胞。胚胎干細(xì)胞和誘導(dǎo)的多能干細(xì)胞都具有多向分化潛能，因而起著舉足輕重的作用。但是，胚胎干細(xì)胞和誘導(dǎo)的多能干細(xì)胞分別受限于倫理學(xué)問題及細(xì)胞表型的鑒定。脂肪干細(xì)胞則不受這些限制，不僅易于獲取，而且方便擴(kuò)增。在不同的誘導(dǎo)條件下，脂肪干細(xì)胞能分化為脂肪細(xì)胞、成骨細(xì)胞、軟骨細(xì)胞、肌肉細(xì)胞、內(nèi)皮細(xì)胞和神經(jīng)細(xì)胞，這些分化潛能可應(yīng)用于再生醫(yī)學(xué)領(lǐng)域，如：皮膚重建，骨和軟骨修復(fù)等。本篇綜述著重討論目前脂肪干細(xì)胞的分離、分化和治療應(yīng)用。

脂肪干細(xì)胞；分離；分化；應(yīng)用

干細(xì)胞包括胚胎干細(xì)胞、誘導(dǎo)的多能干細(xì)胞和成體干細(xì)胞。胚胎干細(xì)胞能自我更新，并且能分化為人體的任何一種細(xì)胞。但由于涉及倫理問題，胚胎干細(xì)胞應(yīng)用于臨床研究存在很大的爭(zhēng)議，而誘導(dǎo)的多能干細(xì)胞和成體干細(xì)胞則沒有這方面的問題。自從Zuk等[1]發(fā)現(xiàn)脂肪組織中含有多分化潛能的干細(xì)胞，脂肪干細(xì)胞就逐漸成為了成體干細(xì)胞中最有前景的種子細(xì)胞之一。脂肪組織通過脂肪抽吸術(shù)或者外科手術(shù)易于獲取，對(duì)供體的創(chuàng)傷相對(duì)較小，而且不涉及倫理問題。在體外，脂肪干細(xì)胞能大量擴(kuò)增，操作簡(jiǎn)便，并能在相應(yīng)的誘導(dǎo)條件下分化為脂肪細(xì)胞、成骨細(xì)胞、軟骨細(xì)胞、肌肉細(xì)胞、內(nèi)皮細(xì)胞、神經(jīng)細(xì)胞等[1-10]。因此，同其他種子細(xì)胞相比，脂肪干細(xì)胞所受限制小，具有更加廣闊的臨床應(yīng)用前景。

1 脂肪干細(xì)胞的分離和培養(yǎng)

同其他干細(xì)胞相比，來源于脂肪組織的干細(xì)胞體內(nèi)含量更大。目前，從脂肪組中分離脂肪干細(xì)胞不僅可以人工分離，也可以采用儀器自動(dòng)離心分離。為了大量獲得脂肪干細(xì)胞，目前大多是在脂肪組織離心分離之后用膠原酶消化[3-10]。Zeng等[11]報(bào)道了另一種方法，即組織塊貼壁培養(yǎng)法，是一種更加高效而且快速的脂肪干細(xì)胞分離培養(yǎng)方法。上述兩種方法所獲得的脂肪干細(xì)胞，在倒置相差顯微鏡下觀察，外形都呈成纖維細(xì)胞樣，可融合成魚群狀或者旋渦狀。

2 脂肪干細(xì)胞的多向分化潛能

脂肪干細(xì)胞具有多向分化潛能，在一定的誘導(dǎo)條件下，可以分化為脂肪細(xì)胞、成骨細(xì)胞、軟骨細(xì)胞、肌肉細(xì)胞、內(nèi)皮細(xì)胞和神經(jīng)細(xì)胞[1-10]。但目前對(duì)于脂肪干細(xì)胞分化的信號(hào)通路尚不十分清楚，有待進(jìn)一步研究[12-15]。分化為脂肪細(xì)胞[2-3]：需要額外添加地塞米松、胰島素、3-異丁基-1-甲基黃嘌呤(IBMX)、吲哚美辛作為誘導(dǎo)條件，成脂分化可以通過油紅染色驗(yàn)證。分化為成骨細(xì)胞[2-3]：需要額外添加地塞米松、維生素C、β-磷酸甘油作為誘導(dǎo)條件，成骨分化可以通過茜素紅染色驗(yàn)證。分化為軟骨細(xì)胞[2]：需要額外添加胰島素、轉(zhuǎn)化生長(zhǎng)因子β1(TGF β1)和維生素C作為誘導(dǎo)條件，成軟骨分化可以通過阿爾辛蘭染色驗(yàn)證。分化為肌肉細(xì)胞[3-4]：需要額外添加5-氮雜胞苷、馬血清作為誘導(dǎo)條件，成肌肉分化可通過檢測(cè)肌蛋白的生成以及體外肌小管形成實(shí)驗(yàn)來驗(yàn)證。分化為內(nèi)皮細(xì)胞[5-6]：需要在含有多種因子的內(nèi)皮細(xì)胞生長(zhǎng)培養(yǎng)基(EGM-2)中添加大劑量的血管內(nèi)皮生長(zhǎng)因子(VEGF)，成內(nèi)皮分化可以通過檢測(cè)內(nèi)皮細(xì)胞的細(xì)胞表面標(biāo)志CD31、CD105、vWF以及體外血管形成實(shí)驗(yàn)來驗(yàn)證。分化為神經(jīng)細(xì)胞[7-10]需要額外添加β-硫基乙醇、維甲酸、血小板源性生長(zhǎng)因子(PDGF)、堿性成纖維細(xì)胞生長(zhǎng)因子(bFGF)、毛喉素、神經(jīng)膠質(zhì)生長(zhǎng)因子2(GGF-2β)，成神經(jīng)分化可以通過檢測(cè)神經(jīng)細(xì)胞標(biāo)志如GFAP、S100等來驗(yàn)證。

3 脂肪干細(xì)胞的臨床應(yīng)用

脂肪干細(xì)胞儲(chǔ)量大、增殖快，而且具有自我更新和多向分化能力，因此臨床應(yīng)用前景廣闊[16]。脂肪干細(xì)胞分化形成的脂肪細(xì)胞可用于軟組織缺損、隆乳、脂肪性營(yíng)養(yǎng)不良以及軟組織填充美容。在軟組織重建方面，自體脂肪移植已廣泛應(yīng)用，但是其局限性依舊存在，其中最明顯的就是組織吸收，導(dǎo)致移植的脂肪量減少20%～90%[17]。解決這一問題的關(guān)鍵，是形成新生血管，從而給移植組織提供營(yíng)養(yǎng)支持。脂肪干細(xì)胞可以分泌多種血管生長(zhǎng)因子，包括血管內(nèi)皮生長(zhǎng)因子(VEGF)[18]，從而刺激新血管形成。

脂肪干細(xì)胞在體內(nèi)和體外都能夠分化為成骨細(xì)胞[2-3，19]，成骨細(xì)胞可做為細(xì)胞材料。脂肪干細(xì)胞能與多種生物支架結(jié)合，促進(jìn)新骨生成，用于治療骨缺損[20-22]。一定條件下，脂肪干細(xì)胞在體內(nèi)和體外都可以分化為軟骨組織[2，23]。這就給利用細(xì)胞治療來修復(fù)關(guān)節(jié)軟骨的缺陷提供了可能，如：修復(fù)骨性關(guān)節(jié)炎。脂肪干細(xì)胞能夠增強(qiáng)肌肉細(xì)胞的分化，提高肌肉的修復(fù)能力[24]。移植的脂肪干細(xì)胞，可以在受損的心肌中，促進(jìn)血管生成、神經(jīng)出芽、心肌細(xì)胞再生，并保護(hù)心功能[25-26]。脂肪干細(xì)胞在肌肉重建方面已經(jīng)取得了一些可喜的成績(jī)，進(jìn)一步的研究有待深入[27]。脂肪干細(xì)胞能夠分化為內(nèi)皮細(xì)胞，并呈現(xiàn)為小管樣結(jié)構(gòu)[5-6]，脂肪干細(xì)胞還可以促進(jìn)血管的形成[28-29]，這給缺血性疾病的治療提供了新的方向。脂肪干細(xì)胞能夠分化為神經(jīng)細(xì)胞和神經(jīng)膠質(zhì)細(xì)胞[7-10]。脂肪干細(xì)胞通過分泌一些神經(jīng)生長(zhǎng)因子，還能促進(jìn)周圍神經(jīng)損傷的修復(fù)[30-31]。脂肪干細(xì)胞還有望應(yīng)用于神經(jīng)退行性病變的治療，如：帕金森病、阿爾茨海默病、亨廷頓氏舞蹈癥等[32-35]。

4 總 結(jié)

干細(xì)胞將成為細(xì)胞治療的有力武器，應(yīng)用于各種疾病，但仍存在許多挑戰(zhàn)，尤其是干細(xì)胞臨床應(yīng)用的安全性和有效性問題，還有同種異體的移植免疫排斥問題等。脂肪干細(xì)胞易于大量獲取，損傷相對(duì)較小，并且安全有效，其臨床應(yīng)用正與日俱增。脂肪干細(xì)胞的分離培養(yǎng)和分化誘導(dǎo)方法正在不斷改進(jìn)。脂肪干細(xì)胞尚未發(fā)現(xiàn)特異性的細(xì)胞表面標(biāo)志物，通過細(xì)胞表面標(biāo)志物純化脂肪干細(xì)胞的方法有待改善。

目前關(guān)于脂肪干細(xì)胞各方面的研究正在不斷深入，各種臨床應(yīng)用問題也在不斷被克服，脂肪干細(xì)胞的臨床應(yīng)用具有廣闊的前景。

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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)

廣東省自然科學(xué)基金(編號(hào)：2014A030313535)

張培華。E-mail：zhangph1128@126.com