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1、1胎儿心脏黏附细胞具有类似间充质祖细 胞特征作者：江小霞，苏永锋，李秀森，张毅，吴英，毛宁【摘要】 为了观察人心脏是否含具有间充质祖细胞特性的细胞，从胎儿心脏分离、培养单个核细胞并从形态、表型和功能 3 个方面与骨髓间充质祖细胞进行比较和鉴定。结果表明，从心脏分离培养的细胞为成纤维样，表面抗原为 CD73, CD105, CD29, CD44, HLA-ABC, CD166 阳性，而 CD45, CD34, CD86, HLA-DR 阴性。在不同的分化体系中，细胞能分化为脂肪细胞、成骨细胞和软骨细胞。细胞扩增迅速，具有低免疫原性特性。结论：从心脏分离培养的细胞具有间充质祖细胞特性。 【关键词】
2、 胎儿心脏； 心脏黏附细胞； 间充质祖细胞Human Fetal Heart-derived Adherent Cells with Characteristics Similar to Mesenchymal Progenitor CellsAbstractThis study was aimed to investigate if human heart harbored a population of primitive undifferentiated cells with the characteristics of MPC. Cells were isolated from hum
3、an fetal heart and were cultured under conditions appropriate for bone marrow-derived MPCs. Their morphology, phenotypes and functions were tested by methods developed for MPC from other sources. The results showed that morphologically, cells were spindle shaped and resembled fibroblasts. In their u
4、ndifferentiated state, cells were CD73, CD105, CD29, CD44, HLA-ABC, CD166 positive and CD45, CD34, CD86, HLA-DR negative. When cultured in adipogenic, osteogenic or chondrogenic media, cells differentiated into adipocytes, osteocytes and chondrocytes respectively. They could be extensively expanded
5、in vitro and exhibited very low immunogenicity as evaluated by T cell proliferation assays. It is concluded that cells isolated from fetal heart possess simi-larity to their adult and fetal bone marrow counterparts in morphologic, immunophenotypic, and functional characteristics.Key wordsfetal heart
6、; heart derived adherent cell; mesenchymal progenitor cell2Bone marrow-derived mesenchymal progenitor cells (MPC) have attracted great attention because of their capability for renewal and differentiation into various lineages of mesenchymal tissues1 and their potential platforms for the systemic de
7、livery of therapeutic proteins in vivo following gene transfer using oncogenic retroviruses2. Studies involving a variety of animal models have shown that adult bone marrow-derived MPC can migrate and engraft in numerous organs and differentiate along tissue-specific lineages under the stimulation o
8、f local factors, and may be useful in the repair or regeneration of damaged or mutated bone, cartilage, or myocardial tissues3-5.Recent work has shown that MPC are present in many tissues, including umbilical cord6, umbilical cord blood7, bone marrow, fetal blood, and fetal li-ver8. Though the use o
9、f MPC in the treatment of acute myocardial infarction has become a novel therapeutic option, the knowledge of their presence in heart is limited 9. Our aim was to investigate whether there were cells with the characteristics of MPC in human fetal heart.Materials and MethodsIsolation and culture of a
10、dult and fetal bone mar-row, fetal heart mesenchymal progenitor cells The Research Ethics Committees of Xuanwu Hospital approved human tissue for research purposes.Human fetal heart samples were obtained from accidental abortus of 4.0-5.0 months under consent. Single-cell suspensions of fetal heart
11、mesenchymal tissue were prepared by carefully rinsing out of the blood and mincing myocardial tissue, away from the blood vessel, through a 70-m-nylon filter. Then cells from adult and fetal bone marrow samples and fetal heart samples were diluted by using 10% fetal bovine serum (FBS; Hyclone, Logan
12、, UT, USA) in Dulbeccos Modified Eagles Medium-Low Glucose (DMEM-LG; Gibco BRL, Life Technologies, Paisley, United Kongdom) with 50 U/ml penicillin, 50 g/ml streptomycin, and 2 mmol/L L-glutamine. Cells were plated into 6-well plate at a density of 100 000 cells/cm2 and incubated at 37 in 5% CO2. Af
13、ter 36 hours, nonadherent cells were removed, and the medium was replaced. At 80% confluence, cells were harvested with 0.25% trypsin and 2 mmol/L EDTA for 5 minutes at 37 and were replated in 75-cm2 flasks. To expand the cells through successive passages, they were plated at 104 cells/cm2, grown to
14、 near confluence, and harvested with the same protocol.Fluorescence-activated cell sorting (FACS) analysis of cultured cellsMonolayer adherent cells from adult bone marrow (n = 4), fetal bone marrow (n = 4), and fetal heart (n = 4) were trypsinized and labbled with anti-CD73-phycoerythrin (PE; PharM
15、ingen, USA), CD105-fluorescein isothiocyanate (FITC; Serotec, Oxford, United Kingdom), HLA-ABC-FITC, CD44-PE, CD29-PE, CD166-PE, CD45-FITC, CD34-PE, HLA-DR-FITC, CD86-PE (Becton Dickinson, USA) and were analyzed by flow cytometry (Beckman Coulter, USA).3Adipogenic, osteogenic, and chondrogenic diffe
16、- rentiationAdipogenic differentiation was assessed by incubation with DMEM with 10% FBS supplemented with 1 mol/L dexamethasone, 10 g/ml insulin, 0.5 mmol/L isobutyl methylxanthine, and 200 mol/L indomethacin (Sigma, St. Louis, MO, USA) for 2 weeks. The presence of adipocytes was assessed by the cellular accumulation of neutral lipid vacuoles that stained with Oil red O (Sigma). Osteogenic differentiation was assessed
