脐带间充质干细胞对CD34^＋细胞在小鼠体内归巢的影响及其机制研究

Study of influence of umbilical cord mesenchymal stem cells on CD34^＋ cells in vivo homing in NOD/ SCID mice and its mechanism

目的探讨脐带来源间充质干细胞（MSC）对脐血来源CD34^＋细胞在NOD／SCID小鼠体内归巢的影响及其可能的机制。方法将CD34^＋细胞与MSC细胞共移植入经放射线照射后的NOD／SCID小鼠，采用流式细胞术和RT—PCR检测移植后20hNOD／SCID小鼠骨髓及脾脏中人CD34^＋细胞，计算其相应的骨髓和脾脏的归巢效率。将脐血CD34^＋细胞与脐带MSC体外共培养，检测MSC细胞对CD34^＋细胞趋化功能的影响；并于培养4、7d检测培养后CD34^＋细胞表面CD49e、CD31、CD62L、CD11a等归巢相关黏附分子表达情况。结果①移植后20h采用流式细胞术成功在小鼠骨髓和脾脏中检测到人CD45^＋细胞。共移植组CD34^＋细胞骨髓归巢率[（7．2±1．1）％]高于单移植组[（5．4±0．9）％]（P〈0．05）。②RT-PCR结果显示共移植组小鼠骨髓细胞和脾脏细胞，单移植组小鼠脾脏细胞扩增得到人GAPDH基因片段，而单移植组小鼠骨髓细胞未见明显扩增条带。③MSC存在时，CD34^＋细胞的体外迁移能力为（35．7±5．8）％，显著高于CD34^＋细胞自发迁移率[（3．5±0．6）％，P〈0．05]。④CD34^＋细胞与MSC体外共培养后细胞表面CD49e、CD31和CD62L黏附分子的表达水平高于CD34^＋细胞单独培养组。结论MSC细胞与CD34^＋细胞共移植有利于CD34^＋细胞向骨髓、脾脏等造血器官归巢，这可能与MSC促进CD34^＋细胞迁移以及维持CD34^＋细胞表面归巢相关黏附分子的表达相关。

Objective To investigate the effect and the potential mechanism of umbilical cord （UC） derived mesenchymal stem cells （MSCs） on umbilical cord blood（UCB） derived CD34^＋ cells in vivo homing in xenotransplanted NOD/SCID mice model. Methods CD34^＋ cells and MSCs were derived from fresh UCB and UC, respectively. CD34^＋ cells （5 × 10^5 per mice） and MSC cells（5 × 10^6 per mice） were co-transplanted into irradiated NOD/SCID mice intravenously. CD34 ^＋ cells （5 × 10^5 per mice） alone were transplanted into the mice as control group. CD34^＋ cells homed in bone marrow and spleen of recipient mice were detected 20 hours after transplant by FACS and RT-PCR, and the homing efficieneies were calculated. The effect of MSCs on CD34^＋ cells chemotactic ruction was investigated after co-cultured UCB CD34^＋ cells with UC MSCs in vitro. After 4 and 7 days coeulture, the homing related adhesion molecules （the CD49e, CD31, CD62L, CDlla） expressed on CD34^＋ cells were detected by FACS. Results ①The homing effieiencies in bone marrow in experimental and control group were （7.2 ± 1.1 ） % and （5.4 ±0.9） % , respectively（P 〈0.05）. ②Human GAPDH gene was detected in bone marrow in experimental group and in spleen in both groups. ③The migration efficiency of CD34 ^＋ cells was significantly higher in experimental group （ 35.7 ± 5.8 ） % than in control group （3.5 ± 0.6） % （ P 〈 0.05 ）. ④The expression of CD49e, CD31, CD62L on CD34 ^＋ cells kept higher level in MSCs coeuhured group than in CD34^＋ cells alone group. Conclusions MSCs can efficiently increase homing of CD34^＋ cells to bone marrow and spleen in vivo by keeping a high level of homing adhesion molecules expression and improving migration efficiency of UCB CD34^＋ cells.