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Isolation of Mesenchymal Stem Cells of Fetal or Maternal Origin from Human Placenta

^a Department of Obstetrics,
^b Department of Immunohematology and Blood Transfusion, and
^c Department of Hematology, Leiden University Medical Center, Leiden, The Netherlands

Key Words. Mesenchymal stem cells • Placenta • Amniotic fluid • Decidua basalis • Decidua parietalis • Fetal

Correspondence: Pieternella S. in ‘t Anker, M.D., Ph.D., Department of Obstetrics, K6-32, P.O. Box 9600, 2300 RC Leiden, The Netherlands. Telephone: 31-71-5262872; Fax: 31-71-5266741; e-mail: E.in_t_Anker{at}lumc.nl

	ABSTRACT

Top Abstract Introduction Materials and Methods Results Discussion References

 
Recently we reported that second-trimester amniotic fluid (AF) is an abundant source of fetal mesenchymal stem cells (MSCs). In this study, we analyze the origin of these MSCs and the presence of MSCs in human-term AF. In addition, different parts of the human placenta were studied for the presence of either fetal or maternal MSCs. We compared the phenotype and growth characteristics of MSCs derived from AF and placenta.

Cells from human second-trimester (mean gestational age, 19⁺² [standard deviation, ± 1⁺³] weeks, n = 10) and term third-trimester (mean gestational age, 38⁺⁴ [standard deviation, ± 1] weeks, n = 10) AF, amnion, decidua basalis, and decidua parietalis were cultured in M199 medium supplemented with 10% fetal calf serum and endothelial cell growth factor. Cultured cells were immunophenotypically characterized, the adipogenic and osteogenic differentiation capacity was tested, and the growth kinetics were analyzed. The origin of fetal and maternal cells was determined by molecular human leukocyte antigen typing.

We successfully isolated MSCs from second-trimester AF, amnion, and decidua basalis as well as term amnion, decidua parietalis, and decidua basalis. In contrast, MSCs were cultured from only 2 out of 10 term AF samples. The phenotype of MSCs cultured from different fetal and maternal parts of the placenta was comparable. Maternal MSCs from second-trimester and term decidua basalis and parietalis showed a significantly higher expansion capacity than that of MSCs from adult bone marrow (p < .05).

Our results indicate that both fetal and maternal MSCs can be isolated from the human placenta. Amnion is a novel source of fetal MSCs, likely contributing to the presence of MSCs in AF. Decidua basalis and decidua parietalis are sources for maternal MSCs. The expansion potency from both fetal and maternal placenta-derived MSCs was higher compared with adult bone marrow–derived MSCs.

	INTRODUCTION

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Mesenchymal stem cells (MSCs) are capable of differentiating into different mesenchymal lineages, including adipose and connective tissue, bone, and cartilage [1–3]. MSCs were initially identified in human postnatal bone marrow (BM) and later in peripheral blood, periosteum, muscle, adipose tissue, and connective tissue of human adults [1, 4–7]. For clinical use, human adult BM is the most common source of MSCs [8, 9]; however, the frequency of MSCs in human adult BM is relatively low. Because the frequency and differentiating capacity of MSCs are decreasing with age [2, 10], different fetal tissues have been studied for the presence of MSCs. Human first-trimester fetal BM, liver, and blood [11] and second-trimester BM, liver, lung, spleen, pancreas, and kidney [12–15] have been found to be rich sources of MSCs.

Because the use of fetal tissues for stem cell therapy has ethical restrictions and is associated with a high rate of bacterial and fungal contamination, other potential sources of fetal MSCs applicable for human therapies have been sought. Umbilical cord blood (UCB) is an attractive source of fetal MSCs; however, it is shown that MSCs are present in UCB in a low frequency [16] or are even undetectable [17, 18] (in ‘t Anker et al., unpublished data). Recently, we reported that second-trimester amniotic fluid (AF) is a novel and rich source of fetal MSCs useful for clinical application [19].

AF contains a heterogeneous population of cells from fetal origin. Potential sites contributing to the presence of cells in the AF are the fetal skin, the fetal membranes of the placenta, and the epithelial and mucosa of the fetal digestive, respiratory, and urinary tract [20, 21]. The origin of the fetal MSCs in AF is yet unknown, and therefore we studied the fetal membranes as a possible source of fetal MSCs. In addition, we analyzed the presence of MSCs in human third-trimester AF, because the presence of those cells in third-trimester AF had not been studied yet.

In their recent report, Zhang et al. [22] describe the presence of MSCs in term human placenta. They did not analyze the origin of the placenta villi-derived cells. The human placenta is an organ with both a fetal and a maternal portion (Fig. 1). The amnion and chorion are from fetal origin, and the different regions of the decidua are from maternal origin. Here we studied whether MSCs from fetal origin and maternal origin could be cultured from the fetal versus maternal side of the placenta.

View larger version (77K): [in this window] [in a new window]   Figure 1. Schematic drawing of the human placenta showing the amnion (fetal part), decidua basalis, and decidua parietalis (maternal part).

	MATERIALS AND METHODS

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Term third-trimester AF and placenta tissue were harvested from 10 deliveries at a mean GA of 38⁺⁴ (SD, ± 1) weeks. All women underwent elective cesarean section for breech presentation. AF was collected by puncture through the membranes after opening the uterine wall during the cesarean section. Decidua basalis tissue was macroscopically dissected from the central region of the maternal-facing surface of the placenta. Term amnion was obtained by removing it from the membranes, and decidua parietalis was collected by scraping it from the chorion. AF, amnion, and decidua were processed within 4 hours.

Additionally, fetal and maternal blood was collected. Second-trimester fetal UCB was collected transcervically, after dilatation, by cordocentesis using a needle (23-gauge) and syringe. Term UCB was collected after the cesarean section. Maternal blood was harvested from women undergoing the termination of pregnancy or cesarean section before the procedure. UCB and maternal blood were collected in tubes containing heparin (Belliver Industrial Estate, Plymouth, U.K.).

Human adult BM samples were obtained from harvests of healthy, young, female donors for allogeneic stem cell transplantation.

The Medical Ethical Review Board of the Leiden University Medical Center approved the protocol (P02/200).

Cell Isolation
AF samples were centrifuged for 10 minutes at 1,283 rpm. Pellets were resuspended in Iscove’s modified Dulbecco’s medium (Bio-Whittaker, Verviers, Belgium) containing 20 U/ml penicillin and 20 µg streptomycin (P/S) and 2% heat-inactivated fetal calf serum (FCS; Gibco Laboratories, Grand Island, NY)—that is, washing medium.

Tissue specimen of amnion from approximately 1 cm² and decidua basalis and decidua parietalis from approximately 1 cm³ were washed in phosphate-buffered saline (PBS). Single-cell suspensions of amnion, decidua basalis, and decidua parietalis were made by mincing and flushing the tissue parts through a 100-µm nylon filter (Falcon, Becton, Dickinson, San Jose, CA) with washing medium.

Maternal blood, UCB, and adult BM were depleted of red cells by incubation in NH₄Cl (8.4 g/l)/KHO₃ (1 g/l) buffer for 10 minutes at 4°C. Maternal and fetal blood was stored at –70°C until use.

Culture of Mesenchymal Stem Cells
Single-cell suspensions of AF, amnion, decidua basalis, and decidua parietalis were cultured in M199 (Gibco Laboratories) supplemented with 10% heat-inactivated FCS, P/S, endothelial cell growth factor (20 µg/ml, Roche Diagnostics GmbH, Mannheim, Germany), and heparin (8 U/ml)—that is, MSC culture medium. Cells were plated in six-wells plates (Greiner Bio-One GmbH, Frickenhausen, Germany). After 7 days, nonadherent cells were removed and the medium was refreshed. When grown to confluency, adherent cells were detached with trypsin/EDTA (5 minutes at 37°C) and expanded in culture flasks (T25 and T75, Greiner Bio-One GmbH). Tissue culture plates and flasks were coated with 1% gelatin (30 minutes at room temperature). Plates and flasks were kept in a humidified atmosphere at 37°C (5% vol/vol CO₂).

Growth Kinetics
For the assessment of growth characteristics of MSCs derived from AF, amnion, decidua basalis, decidua parietalis, and adult BM, 2 x 10⁵ culture-expanded MSCs (passage 3) were seeded in culture flasks (T75) with MSC culture medium. The number of cells was counted in duplicate cultures twice a week over 3 weeks. Human leukocyte antigen (HLA)–typing on cultured cells was performed to confirm that cells from AF and amnion were from fetal origin and cells from decidua basalis and decidua parietalis were from maternal origin.

Flow Cytometric Analysis
Culture-expanded MSCs derived from AF, amnion, decidua basalis, decidua parietalis, and adult BM were phenotypically characterized by flow cytometry (FACSScan, Becton, Dickinson). Fluorescein isothiocyanate (FITC)– or phycoerythrin (PE)–conjugated antibodies against CD31 (DAKO, Glostrup, Denmark), CD166 (CLB, Amsterdam, The Netherlands), SH3, SH4 (a kind gift from Dr. A. Moseley, Osiris, Baltimore), CD105 (Ancell Corporation, Bayport, MN), CD34, CD45 (Becton, Dickinson), CD90 (Pharmingen, San Diego), HLA-DR (Becton, Dickinson), CD49d, CD49e (Immunotech Coulter Company, Marseille, France), CD123 (Pharmingen), and HLA-ABC (Instruchemie, Hilversum, The Netherlands) were used. Positive cells were identified by comparison with isotypic controls (FITC- and PE-conjugated mouse immunoglobulin G1 [IgG1], IgG2a, or IgG2b). LDS-751 (4 ng/ml, Exciton, Daton, OH) was used to adjust a life gate and to exclude dead cells.

Adipogenic and Osteogenic Differentiation Testing
The adipogenic and osteogenic differentiation capacity of culture-expanded MSCs was determined as previously reported [13, 23]. Culture-expanded cells from AF, amnion, decidua basalis, decidua parietalis, and adult BM were seeded at a density of 2.5 x 10⁴ cells/cm² in 24-well plate and cultured in 500 µl -MEM supplemented with 10% heat-inactivated FCS, P/S, ascorbic acid (50 µg/ml), and dexamethasone (10^–7 M). ß-glycerophosphate (5 mM) was added to the medium from day 7 onward. For induction of adipogenesis, 50 µM indomethacin, 1.6 µM bovine insulin, and 0.5 mM 1-methyl-3-isobutylxantine were added to this medium.

Cells were cultured at 37°C (5% vol/vol CO₂), and the medium was replaced twice weekly. After 3 weeks of culture, cells were washed with phosphate-buffered 0.9% NaCl and fixed with 10% formalin in PBS for 10 minutes and analyzed for osteogenic and adipogenic differentiation.

Adipogenic differentiation was visualized after staining with 0.3% Oil red O in 60% isopropanol for 10 minutes at room temperature. Thereafter, cells were washed with 60% isopropanol and staining was stopped with AD.

To visualize osteogenic differentiation, cells were stained for alkaline phosphatase (AP) and calcium (Ca) deposition. For AP expression, cells were washed with PBS and subsequently incubated for 15 minutes with substrate solution (0.2 mg/ml -naphthyl-1-phosphate, 0.1 M Tris buffer, pH 8.9, 0.01% magnesium sulphate, and 0.6 mg/ml fast blue RR acid), resulting in the formation of a purple reaction product.

To detect calcium deposition, cells were washed with PBS and incubated with 2% Alizarin red S solution adjusted to pH 5.5 with 0.5% NH₄OH for 2–5 minutes. Mineralization was demonstrated by the presence of red depositions.

HLA Typing
To analyze the origin of culture-expanded MSCs derived from AF (collected by both transabdominal and transcervical puncture), amnion, decidua basalis, and decidua parietalis, a molecular HLA typing was performed on DNA obtained from expanded MSCs and fetal and maternal blood cells by polymerase chain reaction/sequence-specific oligonucleotide using a reverse-dot blot method with the Dynal Reli SSO (Dynal Biotech, Hamburg, Germany) reverse-line blot strip assay [24].

Statistical Analysis
Logistic growth curves were fitted to the data of the growth curves using the function nls in the free software system R (r-project). Student’s t-test was used to calculate differences in growth. A p value of < .05 was considered significantly significant.

	RESULTS

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Identification of Mesenchymal Stem Cells
The mean volume of AF collected from second-trimester pregnancies by transabdominal puncture was 8.7 ml (SD, ± 1.7) and by transcervical punctures was 32.3 ml (SD, ± 13.9). MSCs were cultured from all 10 consecutive samples of both transabdominally and transcervically collected second-trimester AF. In 8 of 10 samples, MSCs were isolated from second-trimester amnion, and in 8 of 10 samples, MSCs were isolated from second-trimester decidua (Table 1).

Culture-expanded cells derived from AF, amnion, decidua basalis, and decidua parietalis were immunopheno-typically analyzed. The phenotype of the cultured cells was similar to that of MSC derived from adult BM and fetal second-trimester tissues [1, 13], i.e., CD90, CD105, CD166, CD49e, SH3, SH4, and HLA-ABC positive and CD31, CD34, CD45, CD49d, CD123, and HLA-DR negative. No difference was found among the expression of one of these markers on MSCs from the different sources.

Culture-expanded cells from AF, amnion, decidua basalis, and decidua parietalis and adult BM were all able to differentiate into both osteoblasts and adipocytes.

HLA Typing
HLA analysis of the culture-expanded cells from transabdominally collected second-trimester (n = 10) and term (n = 2) AF showed that all these samples were of fetal origin, i.e., only fetal-specific and no maternal alleles were present (Table 1). On the basis of HLA typing, all second-trimester (n = 8) and term (n = 7) amnion-derived culture-expanded cells were of fetal origin (Fig. 2A). In contrast, only 4 of the 10 transcervically collected AF samples were of fetal origin. Six of the 10 transcervically collected AF samples expressed both fetal- and maternal-specific alleles.

View larger version (37K): [in this window] [in a new window]   Figure 2. HLA typing of culture-expanded MSCs. The Dynal Reli SSO (Dynal Biotech, Hamburg, Germany) reverse-line blot strip assay was used for molecular typing of the HLA-A and HLA-B locus alleles of maternal cells, fetal cells, and culture-expanded MSCs from the same sample. The HLA-A and HLA-B type of the culture-expanded amnion-derived MSCs (A) is identical to the fetal HLA-A and HLA-B type and mismatched with the maternal HLA-A and HLA-B type. The HLA-A and HLA-B type of the culture-expanded deciduas basalis–derived MSCs (B) is identical to the maternal HLA-A and HLA-B type and mismatched with the fetal HLA-A and HLA-B type. Upward arrows indicate maternal-specific HLA antigens, and downward arrows indicate fetal-specific HLA antigens. Abbreviations: C, cultured amnion-derived MSCs; F, fetal cells; M, maternal cells; MSCs, mesenchymal stem cells.

Growth Characteristics
The growth characteristics of MSCs derived from different fetal (second-trimester transabdominal-collected AF [n = 7] and amnion [n = 3]), maternal (decidua from second-trimester placenta [n = 5] and decidua basalis [n = 2] and decidua parietalis [n = 5] from term placenta), and adult (BM [n = 2]) sources were compared during 3 weeks. The growth of second-trimester fetal AF (collected transabdominally) and amnion-derived MSCs was similar (Fig. 3). There was no significant difference in cell numbers and growth kinetics during this culture period. After 11 days, a plateau in the growth was reached. Maternal cells derived from second-trimester decidua tissue and term decidua parietalis tissue had a similar growth pattern (Fig. 4). The number of cells derived from these two sources at the days of counting was not significantly different. From day 11, the number of adherent cells derived from term decidua basalis was significantly higher (p < .05) compared with the amount of cells derived from second-trimester decidua tissue and term decidua parietalis. The growth velocity and cell number of MSCs derived from adult BM was from 11 days significantly lower (p < .05) than that from MSCs derived from the three maternal sources we tested, i.e., second-trimester decidua and term decidua parietalis and decidua basalis.

View larger version (5K): [in this window] [in a new window]   Figure 3. Growth curves of amnion and amniotic fluid. At t = 0, 200,000 mesenchymal stem cells were seeded in culture flasks. Duplicate cultures were harvested twice weekly for 3 weeks, and adherent cells were counted. Results are expressed as mean ± standard error of the mean. Growth curves of second-trimester amnion (n = 3, gray line) and second-trimester amniotic fluid (n = 7, black line).

View larger version (8K): [in this window] [in a new window]   Figure 4. Growth curves of decidua basalis, decidua parietalis, and adult BM. At t = 0, 200,000 mesenchymal stem cells were seeded in culture flasks. Duplicate cultures were harvested twice weekly for 3 weeks, and adherent cells were counted. Results are expressed as mean ± standard error of the mean. Growth curves of adult BM (n = 2, ), second-trimester decidua (n = 5, ), term decidua basalis (n = 2,), and term decidua parietalis (n = 5,x).Abbreviation: BM, bone marrow.

	DISCUSSION

Top Abstract Introduction Materials and Methods Results Discussion References

Recently we described that second-trimester AF is a rich source of MSCs [19]. HLA typing confirmed that these isolated MSC were exclusively of fetal origin. It has been reported that AF contains different fetal cells that originate from several fetal tissues [20]. Potential sites contributing to MSCs in AF are the fetal membranes. In this study we show that amnion is a rich source for MSCs. From 8 of the 10 second-trimester and 7 of 10 term amnion samples, MSCs could be isolated and expanded. Moreover, the growth characteristics of MSCs from amnion and AF, analyzed in growth curves, were similar. From these data we hypothesize that MSCs in AF are likely, at least in part, derived from amnion.

In contrast to second-trimester AF, we could only culture MSCs from a minority of term AF samples. Adherent cells were identified in only 2 of 10 AF samples. The total number of cells in AF increased steadily, but the proportion of viable cells decreased with GA [20]. The latter may explain our failure to culture MSCs from term AF. Moreover, the nonviable cells possibly interfere with the culture of MSCs. In our experiments we tried to deplete nonviable cells of term AF by filtering the cells and by density-gradient centrifugation. However, we were also unable to culture MSCs from the low-density fraction, suggesting that the frequency of MSCs in term AF is significantly lower than in second-trimester AF.

The human placenta is an organ consisting of a fetal and a maternal part (Fig. 1). In their report, Zhang et al. [22] described the presence of MSCs in placenta; however, they did not study the maternal or fetal origin of the cultured placenta villi-derived cells. After transcervical collection of AF, we unexpectedly found that the MSCs from some of the samples were not of pure fetal origin but also contained maternal cells. A reasonable explanation for the presence of maternal cells in this population is contamination of the AF with cells derived from the decidua.

We analyzed the origin of MSCs present in maternal parts of the placenta. For this, HLA typing was performed on culture-expanded cells from decidua and compared with the HLA of the mother and fetus. We confirmed the pure maternal origin of MSCs derived from decidua basalis and decidua parietalis from both second-trimester and term pregnancies. However, MSCs from two of eight samples of second-trimester decidua tissue were not from pure maternal origin but showed fetal alleles as well. This may be related to the small size of the placenta at the second-trimester of pregnancy; it is almost impossible to obtain placenta tissue that contains only maternal cells. MSCs could be derived from 8 of 10 second-trimester decidua samples, 6 of 10 third-trimester decidua parietalis samples, and 4 of 10 third-trimester deciduas basalis samples. A possible explanation for the lower success rate of third-trimester deciduas samples is a decreasing frequency of MSCs with GA.

We found that the expansion potential of maternal decidua-derived MSCs from both second and third trimester was significantly higher than that of adult BM-derived MSCs. BM from young (mean age, 30 years), healthy female donors was used for these experiments. This difference in growth possibly is related to pregnancy. To evaluate this, BM from healthy pregnant women should be analyzed in a growth curve and compared with the growth of BM MSCs from nonpregnant women. Another explanation is that MSCs in the placenta exhibit higher expansion potency because of their role in the growth and differentiation of this developing organ. Molecular studies and cell-cycle analysis on MSCs from different sources possibly may give insight in the reasons for the difference in growth.

Adult BM is the common source of MSCs used in clinical settings [8, 25]. However, the use of adult BM has some limitations. First, the frequency of MSCs in adult BM is low. Moreover, harvesting BM from a patient is an invasive procedure. Therefore, the search for alternative sources of MSCs useful for clinical application is important. We previously found that second-trimester AF is an abundant source of MSCs [19]. However, the risk of amniocentesis prohibits the use of AF for routine clinical use. In this study, we show that placenta is an additional source for MSCs. One of the most important benefits of using placenta-derived MSCs for clinical use is the availability. Moreover, it is possible to obtain placenta and UCB from the same donor. Therefore, amnion is an attractive source of MSCs for cotransplantation in conjunction with UCB-derived hemaopoietic stem cells [12, 19]. Furthermore, the finding of MSCs in fetal membranes might help to understand wound healing in the fetal membranes and represent a first step in the development of tissue engineering strategies in cases of fetal membrane rupture. The presence of MSCs at the fetal–maternal interface suggests a role of these cells in the induction of tolerance. Experiments on the immunosuppressive properties of decidua-derived MSCs have now been initiated. Furthermore, MSCs of fetal and maternal origin might play a role in the treatment of disorders of mesenchymal origin.

In conclusion, both second-trimester and term third-trimester amnion represent a rich source for fetal MSCs and likely contribute to the presence of MSCs in second-trimester AF. Maternal MSCs can be cultured from both second-trimester and term decidua. The phenotype of these cells is comparable with adult BM-derived MSCs, and the expansion potency is significantly higher than that of MSCs from adult BM.

	ACKNOWLEDGMENTS

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We thank the gynecologists from the department of Obstetrics of the Leiden University Medical Center and W. Beekhuizen of the Center of Human Reproduction in Leiden for collecting the amniotic fluid and P.H.C. Eilers, Ph.D., from the Leiden University Medical Center for help with statistics.

	REFERENCES

Top Abstract Introduction Materials and Methods Results Discussion References

 

1. Pittenger MF, Mackay AM, Beck SC et al. Multilineage potential of adult human mesenchymal stem cells. Science 1999;284:143–147.[Abstract/Free Full Text]

2. Caplan AI. The mesengenic process. Clin Plast Surg 1994;21:429–435.[Medline]

3. Deans RJ, Moseley AB. Mesenchymal stem cells: biology and potential clinical uses. Exp Hematol 2000;28:875–884.[CrossRef][Medline]

4. Zuk PA, Zhu M, Ashjian P et al. Human adipose tissue is a source of multipotent stem cells. Mol Biol Cell 2002;13: 4279–4295.[Abstract/Free Full Text]

5. Zvaifler NJ, Marinova-Mutafchieva L, Adams G et al. Mesenchymal precursor cells in the blood of normal individuals. Arthritis Res 2000;2:477–488.[CrossRef][Medline]

6. Nakahara H, Dennis JE, Bruder SP et al. In vitro differentiation of bone and hyertrophic cartilage from periosteal-derived cells. Exp Cell Res 1991;195:492–503.[CrossRef][Medline]

7. Nathanson MA. Bone matrix-directed chondrogenesis of muscle in vitro. Clin Orthop 1985;200:142–158.

8. Frassoni F, Labopin M, Bacigalupo A. Expanded mesenchymal stem cells (MSC), co-infused with HLA identical hemopoietic stem cell transplants, reduce acute and chronic graft versus host disease: a matched pair analysis. Bone Marrow Transplant 2002;29(suppl):S2.

9. Lazarus HM, Haynesworth SE, Gerson SL et al. Ex vivo expansion and subsequent infusion of human bone marrow-derived stromal progenitor cells (mesenchymal progenitor cells): implications for therapeutic use. Bone Marrow Transplant 1995;16:557–564.[Medline]

10. D’Ippolito G, Schiller PC, Ricordi C et al. Age-related osteogenic potential of mesenchymal stromal stem cells from human vertebral bone marrow. J Bone Miner Res 1999;14:1115–1122.[CrossRef][Medline]

11. Campagnoli C, Roberts IA, Kumar S et al. Identification of mesenchymal stem/progenitor cells in human first-trimester fetal blood, liver, and bone marrow. Blood 2001; 98:2396–2402.[Abstract/Free Full Text]

12. Noort WA, Kruisselbrink AB, in ‘t Anker PS et al. Mesenchymal stem cells promote engraftment of human umbilical cord blood-derived CD34⁺ cells in NOD/SCID mice. Exp Hematol 2002;30:870–878.[CrossRef][Medline]

13. in ‘t Anker PS, Noort WA, Scherjon SA et al. Mesenchymal stem cells in human second-trimester bone marrow, liver, lung, and spleen exhibit a similar immunophenotype but a heterogeneous multilineage differentiation potential. Haematologica 2003;88:845–852.[Abstract/Free Full Text]

14. Almeida-Porada G, El Shabrawy D, Porada C et al. Differentiative potential of human metanephric mesenchymal cells. Exp Hematol 2002;30:1454–1462.[CrossRef][Medline]

15. Hu Y, Liao L, Wang Q et al. Isolation and identification of mesenchymal stem cells from human fetal pancreas. J Lab Clin Med 2003;141:342–349.[CrossRef][Medline]

16. Erices A, Conget P, Minguell JJ. Mesenchymal progenitor cells in human umbilical cord blood. Br J Haematol 2000; 109:235–242.[CrossRef][Medline]

17. Mareschi K, Biasin E, Piacibello W et al. Isolation of human mesenchymal stem cells: bone marrow versus umbilical cord blood. Haematologica 2001;86:1099–1100.[Free Full Text]

18. Wexler SA, Donaldson C, Denning-Kendall P et al. Adult bone marrow is a rich source of human mesenchymal "stem" cells but umbilical cord and mobilized adult blood are not. Br J Haematol 2003;121:368–374.[CrossRef][Medline]

19. in ‘t Anker PS, Scherjon SA, Kleijburg-van der Keur C et al. Amniotic fluid as a novel source of mesenchymal stem cells for therapeutic transplantation. Blood 2003;102:1548–1549.[Free Full Text]

20. Gosden CM. Amniotic fluid cell types and culture. Br Med Bull 1983;39:348–354.[Free Full Text]

21. Priest RE, Marimuthu KM, Priest JH. Origin of cells in human amniotic fluid cultures: ultrastructural features. Lab Invest 1978;39:106–109.[Medline]

22. Zhang X, Nakaoka T, Nishishita T et al. Efficient adeno-associated virus-mediated gene expression in human placenta-derived mesenchymal cells. Microbiol Immunol 2003;47:109–116.[Medline]

23. Dang ZC, van Bezooijen RL, Karperien M et al. Exposure of KS483 cells to estrogen enhances osteogenesis and inhibits adipogenesis. J Bone Miner Res 2002;17:394–405.[CrossRef][Medline]

24. Erlich H, Bugawan T, Begovich AB et al. HLA-DR, DQ and DP typing using PCR amplification and immobilized probes. Eur J Immunogenet 1991;18:33–55.[Medline]

25. Koc ON, Gerson SL, Cooper BW et al. Rapid hematopoietic recovery after coinfusion of autologous-blood stem cells and culture-expanded marrow mesenchymal stem cells in advanced breast cancer patients receiving high-dose chemotherapy. J Clin Oncol 2000;18:307–316.[Abstract/Free Full Text]

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				K. Sudo, M. Kanno, K. Miharada, S. Ogawa, T. Hiroyama, K. Saijo, and Y. Nakamura Mesenchymal Progenitors Able to Differentiate into Osteogenic, Chondrogenic, and/or Adipogenic Cells In Vitro Are Present in Most Primary Fibroblast-Like Cell Populations Stem Cells, July 1, 2007; 25(7): 1610 - 1617. [Abstract] [Full Text] [PDF]

				C. Ventura, S. Cantoni, F. Bianchi, V. Lionetti, C. Cavallini, I. Scarlata, L. Foroni, M. Maioli, L. Bonsi, F. Alviano, et al. Hyaluronan Mixed Esters of Butyric and Retinoic Acid Drive Cardiac and Endothelial Fate in Term Placenta Human Mesenchymal Stem Cells and Enhance Cardiac Repair in Infarcted Rat Hearts J. Biol. Chem., May 11, 2007; 282(19): 14243 - 14252. [Abstract] [Full Text] [PDF]

				L. Duplomb, M. Dagouassat, P. Jourdon, and D. Heymann Concise Review: Embryonic Stem Cells: A New Tool to Study Osteoblast and Osteoclast Differentiation Stem Cells, March 1, 2007; 25(3): 544 - 552. [Abstract] [Full Text] [PDF]

				P. V. Guillot, C. Gotherstrom, J. Chan, H. Kurata, and N. M. Fisk Human First-Trimester Fetal MSC Express Pluripotency Markers and Grow Faster and Have Longer Telomeres Than Adult MSC Stem Cells, March 1, 2007; 25(3): 646 - 654. [Abstract] [Full Text] [PDF]

				C.-J. Chang, M.-L. Yen, Y.-C. Chen, C.-C. Chien, H.-I. Huang, C.-H. Bai, and B. L. Yen Placenta-Derived Multipotent Cells Exhibit Immunosuppressive Properties That Are Enhanced in the Presence of Interferon-{gamma} Stem Cells, November 1, 2006; 24(11): 2466 - 2477. [Abstract] [Full Text] [PDF]

				C.-C. Chien, B. L. Yen, F.-K. Lee, T.-H. Lai, Y.-C. Chen, S.-H. Chan, and H.-I Huang In Vitro Differentiation of Human Placenta-Derived Multipotent Cells into Hepatocyte-Like Cells Stem Cells, July 1, 2006; 24(7): 1759 - 1768. [Abstract] [Full Text] [PDF]

				M. L. Weiss, S. Medicetty, A. R. Bledsoe, R. S. Rachakatla, M. Choi, S. Merchav, Y. Luo, M. S. Rao, G. Velagaleti, and D. Troyer Human Umbilical Cord Matrix Stem Cells: Preliminary Characterization and Effect of Transplantation in a Rodent Model of Parkinson's Disease Stem Cells, March 1, 2006; 24(3): 781 - 792. [Abstract] [Full Text] [PDF]

				M.-S. Tsai, S.-M. Hwang, Y.-L. Tsai, F.-C. Cheng, J.-L. Lee, and Y.-J. Chang Clonal Amniotic Fluid-Derived Stem Cells Express Characteristics of Both Mesenchymal and Neural Stem Cells Biol Reprod, March 1, 2006; 74(3): 545 - 551. [Abstract] [Full Text] [PDF]

				S. Sartore, M. Lenzi, A. Angelini, A. Chiavegato, L. Gasparotto, P. D. Coppi, R. Bianco, and G. Gerosa Amniotic mesenchymal cells autotransplanted in a porcine model of cardiac ischemia do not differentiate to cardiogenic phenotypes Eur. J. Cardiothorac. Surg., November 1, 2005; 28(5): 677 - 684. [Abstract] [Full Text] [PDF]

				A. Leri, J. Kajstura, and P. Anversa Cardiac Stem Cells and Mechanisms of Myocardial Regeneration Physiol Rev, October 1, 2005; 85(4): 1373 - 1416. [Abstract] [Full Text] [PDF]

				T. Miki, T. Lehmann, H. Cai, D. B. Stolz, and S. C. Strom Stem Cell Characteristics of Amniotic Epithelial Cells Stem Cells, October 1, 2005; 23(10): 1549 - 1559. [Abstract] [Full Text] [PDF]

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