| BACKGROUND:Mesenchymal stem cells (MSCs) are a kind of adult stem cells that can be self-renewal and have the ability of multi-lineage differentiation. MSCs were first found in bone marrow. Subsequently, MSCs are also found in fat, periosteum, synovium, muscle, and dental pulp. The intriguing biological characteristic of MSCs makes them become one of the major seed cells in tissue engineering and regeneration medicine. Currently, MSCs are mainly derived from bone marrow. However, aspirating bone marrow is an invasive procedure; the number and differentiating potential of bone marrow also decrease with ages. All the reasons constrain us from applying MSCs in biological engineering and the clinic. However, many researches have proved that MSCs also appear in the human umbilical cord and placenta. Furthermore, comparing to the MSCs derived from bone marrow, MSCs derived from human umbilical cord and placenta have characteristics of easier vitro expansions and low immunogenicity. Nevertheless, until now there are few studies articulating the biological characteristics of MSCs derived from umbilical cord and placenta.OBJECTION:This study was designed to isolate, identify, and cultivate human mesenchymal stem cells derived from umbilical cord, amnion and chorion, and to compare the biological characteristics of MSCs from the three sources. Furthermore, the role of the three sources of MSCs in immune regulation was also stuided.METHODS:In this study, MSCs derived from umbilical cord, amnion, and chorion were isolated separately by using density gradient centrifugation or enzyme (collagenase/trypsin) digestion. MSCs from the 3 sources were further cultured and purified by adherent culture. Short tandem repetitive sequence (STR) analysis was used to detect whether the MSCs obtained from placenta is fetal chorionic villous original. Cell growth pattern was detected by MTT assay. Flow cytometry was applied to analyze the three cells' phenotype and stem cell markers. The different differential medium was used to detect their multi-directional differentiation ability. The immunomodulatory properties of these umbilical cord, amnion, and chorion-derived cells were evaluated by coculturing the cells with peripheral blood mononuclear cells stimulated by phytohemagglutinin (PHA), and the level of interferon-y (IFN-y) was detected by enzyme-labeled immunosorbent assay (ELISA).RESULTS:The short tandem prepeat analysis showed that these chorionic-derived cells are derived from fetal chorionic. The three types of cells show as typical fibroblast morphology and are adherent to the tissue culture substrate. MSCs derived from chorionic are slightly larger than the MSCs derived from umbilical cord and amniotic membrane. The speed of doubling time of the UC-MSCs is faster than that of AM-MSC and CM-MSC (the lowest). Flow cytometric analysis reveals that these cells are consistently positive for markers of MSCs including CD90, CD73, CD 105, CD44, and negative for the hematopoietic markers CD31, CD34 and CD45. Meanwhile, all the three types of cells expressed pluripotent transcription factors like SSEA-4, Nestin and Sox-2. The three types of cells can differentiate into osteoblasts and adipogenesis under different conditioned medium cultured conditions. When cocultured with human peripheral blood mononuclear cells, the three types of MSCs can inhibit y-interferon secretion by peripheral blood mononuclear cells after stimulation of PHA. The effects of inhibition were (from strong to weak): UC-MSCs, CM-MSCs and AM-MSCs.CONCLUSION:We have successfully isolated and identified mesenchymal stem cells from human umbilical cord, amnion and chorion. The results showed that there are differences among these three types of cells, both in terms of biological characteristics and in terms of immune regulations. |
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