Sun Ning, Huang Yizhou, Zhang Yi, Pi Jinkui, Jaznie Law, Deng Li, Wu Chengguang
Laboratory of Stem Cell and Tissue Engineering, State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu Sichuan, 610041, P. R. China.
Core Facility, West China Hospital, Sichuan University, Chengdu Sichuan, 610041, P. R. China.
Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2016 May 8;30(5):619-625. doi: 10.7507/1002-1892.20160125.
To investigate the effectiveness of human placental decidua basalis derived mesenchymal stem cells (PDB-MSCs) in repairing full-thickness skin defect of nude mice.
Human placenta samples were obtained from healthy donor mothers with written informed consent. PDB-MSCs were isolated through enzymic digestion and density gradient centrifugation; the 4th passage cells were identified by cellular morphology, cell adipogenic and osteogenic differentiation, and phenotype evaluation. Forty-two 4-5-week-old BALB/c female nude mice were randomly divided into experimental group (=21) and control group (=21). The 4th passage PDB-MSCs solution (200 μL, 5×10/mL) was injected into the mice of experimental group via caudal vein; the mice of control group were given equal volume of PBS. The full-thickness skin defect model of 1.5 cm×1.5 cm in size was made after 3 days. The wound healing was observed generally at 1, 2, 4, 7, 14, 18, 21, 25, and 30 days after operation, and the wound healing rate was calculated after wound decrustation. HE staining was used to observe the wound repair at 1, 7, 14, 21, and 31 days; immunofluorescent staining was used for cellular localization at 7, 14, and 31 days after operation.
Cells isolated from human placenta were MSCs which had multipotential differentiation ability and expressed MSCs phenotype. Animals survived to the end of the experiment. The general observation showed that the experimental group had a faster skin repairing speed than the control group; the time for decrustation was 12-14 days in experimental group and was 14-17 days after operation in the control group. The wound healing rate of experimental group was significantly higher than that of control group at 14, 18, and 21 days (=4.001, =0.016; =3.380, =0.028; =3.888, =0.018), but no significance was found at 25 and 30 days (=1.565, =0.193; =1.000, =0.423). HE staining showed lower inflammatory reaction, and better regeneration of the whole skin and glands with time in the experimental group. The immunofluorescent staining was positive in skin defect area of experimental group at different time points which displayed that human PDB-MSCs existed.
Through enzymic digestion and density gradient centrifugation, PDB-MSCs can be obtained. Pre-stored PDB-MSCs can mobilize to the defect area and participate in repair of nude mice skin.
探讨人胎盘基蜕膜间充质干细胞（PDB-MSCs）修复裸鼠全层皮肤缺损的有效性。
经健康供体母亲书面知情同意后获取人胎盘样本。通过酶消化和密度梯度离心法分离PDB-MSCs；对第4代细胞进行细胞形态、细胞成脂及成骨分化和表型评估鉴定。将42只4-5周龄雌性BALB/c裸鼠随机分为实验组（n=21）和对照组（n=21）。将第4代PDB-MSCs溶液（200μL，5×10⁶/mL）经尾静脉注入实验组小鼠；对照组小鼠给予等量PBS。3天后制作大小为1.5 cm×1.5 cm的全层皮肤缺损模型。术后1、2、4、7、14、18、21、25和30天对伤口愈合情况进行大体观察，伤口结痂脱落后计算伤口愈合率。术后1、7、14、21和31天采用HE染色观察伤口修复情况；术后7、14和31天采用免疫荧光染色进行细胞定位。
从人胎盘中分离出的细胞为具有多向分化能力并表达MSCs表型的间充质干细胞。动物存活至实验结束。大体观察显示，实验组皮肤修复速度较对照组快；实验组脱痂时间为术后12-14天，对照组为术后14-17天。实验组在术后14、18和21天的伤口愈合率显著高于对照组（P=4.001，P=0.016；P=3.380，P=0.028；P=3.888，P=0.018），但在术后25和30天无显著性差异（P=1.565，P=0.193；P=1.000，P=0.423）。HE染色显示，实验组炎症反应较轻，随着时间推移全层皮肤及腺体再生较好。实验组不同时间点皮肤缺损区免疫荧光染色呈阳性，表明人PDB-MSCs存在。
通过酶消化和密度梯度离心法可获取PDB-MSCs。预先储存的PDB-MSCs可动员至缺损区域并参与裸鼠皮肤修复。
