Yuan Qilong, Zeng Xiaoyong, Chen Liang, Peng Ejun, Ye Zhangqun
Center of Assisted Reproductive Technology, Guangzhou General Hospital of Guangzhou Military Command, Guangzhou Guangdong 510010, PR China.
Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2010 Oct;24(10):1228-32.
To compare the myogenic differentiation ability in vitro of rabbit adipose-derived stem cells (ADCSs) from different sites so as to provide ideal seed cells for repair and reconstruction of urinary tract.
Adipose tissues were obtained from the nape of the neck, post peritoneum, and vicinity of epididymis of a 4-month-old male New Zealand rabbit and ADSCs were harvested through collagenase digestion. ADSCs were purified by differential attachment method. The protein marker CD44 of rabbit ADSCs was used to identify the stem cells by immunocytochemistry, then the 5th generation of ADSCs were induced to differentiate into adipogenic, osteogenic, and myogenic cells. Multi-differentiation was confirmed by Oil red O staining, von Kossa staining, and RT-PCR. Myogenic differentiation abilities of ADSCs from 3 different sites were compared between the control group (L-DMEM medium containing 10%FBS) and the experimental group (myogenic medium) by RT-PCR method.
ADSCs could be easily isolated from adipose tissues of the nape of the neck, post peritoneum, and vicinity of epididymis. ADSCs displayed a typical cobblestone morphology. Brown particles could be seen in ADSCs by CD44 immunocytochemistry staining. Oil red O staining showed red fat drops in ADSCs after 14 days of adipogenic culture. Black matrix could be seen in ADSCs by von Kossa staining after 28 days of osteogenic culture. RT-PCR detection showed moderate alpha-actin expression in the control group and strong alpha-actin expression in the experimental group after 42 days of myogenic culture. The growth rate of alpha-actin from the adipose tissue of post peritoneum (28.622% +/- 4.879%) was significantly lower (P < 0.05) than those from the adipose tissues of the nape of the neck (35.471% +/- 3.434%) and vicinity of epididymis (38.446% +/- 4.852%).
The ADSCs from different sites show different myogenic differentiation abilities in vitro. ADSCs from the adipose tissues of the nape of the neck and vicinity of epididymis can be used as ideal seed cells for tissue engineering of lower urinary tract.
比较兔不同部位脂肪来源干细胞（ADSCs）的体外成肌分化能力，为尿路修复与重建提供理想的种子细胞。
取4月龄雄性新西兰兔颈部、腹膜后及附睾附近的脂肪组织，采用胶原酶消化法获取ADSCs。通过差速贴壁法纯化ADSCs。采用免疫细胞化学法，用兔ADSCs的蛋白标志物CD44鉴定干细胞，然后将第5代ADSCs诱导分化为脂肪细胞、成骨细胞和成肌细胞。通过油红O染色、冯库萨染色和逆转录聚合酶链反应（RT-PCR）证实多向分化。采用RT-PCR法比较对照组（含10%胎牛血清的L-DMEM培养基）和实验组（成肌培养基）中3个不同部位ADSCs的成肌分化能力。
可从颈部、腹膜后及附睾附近的脂肪组织中轻松分离出ADSCs。ADSCs呈现典型的鹅卵石形态。CD44免疫细胞化学染色可见ADSCs中有棕色颗粒。脂肪生成培养14天后，油红O染色显示ADSCs中有红色脂肪滴。成骨培养28天后，冯库萨染色可见ADSCs中有黑色基质。RT-PCR检测显示，成肌培养42天后，对照组中α-肌动蛋白表达适度，实验组中α-肌动蛋白表达强烈。腹膜后脂肪组织中α-肌动蛋白的生长率（28.622%±4.879%）显著低于颈部脂肪组织（35.471%±3.434%）和附睾附近脂肪组织（38.446%±4.852%）（P<0.05）。
不同部位的ADSCs在体外表现出不同的成肌分化能力。颈部和附睾附近脂肪组织来源的ADSCs可作为下尿路组织工程的理想种子细胞。
