Human amniotic fluid as a potential new source of organ specific precursor cells for future regenerative medicine applications.

Division of Urology, Laboratory for Organ Regenerative Research and Cell Therapeutics, Saban Research Institute, Children's Hospital Los Angeles and Developmental Biology Program, Keck School of Medicine, University of Southern California, Los Angeles, California 90027, USA.
The Journal of urology (Impact Factor: 3.75). 03/2010; 183(3):1193-200. DOI: 10.1016/j.juro.2009.11.006
Source: PubMed

ABSTRACT Human amniotic fluid contains multiple cell types, including pluripotent and committed progenitor cells, and fully differentiated cells. We characterized various cell populations in amniotic fluid.
Optimum culture techniques for multiple cell line passages with minimal morphological change were established. Cell line analysis and characterization were done with reverse transcriptase and real-time polymerase chain reaction. Immunoseparation was done to distinguish native progenitor cell lines and their various subpopulations.
Endodermal and mesodermal marker expression was greatest in samples of early gestational age while ectodermal markers showed a constant rate across all samples. Pluripotent and mesenchymal cells were always present but hematopoietic cell markers were expressed only in older samples. Specific markers for lung, kidney, liver and heart progenitor cells were increasingly expressed after 18 weeks of gestation. We specifically focused on a CD24+OB-cadherin+ population that could identify uninduced metanephric mesenchyma-like cells, which in vivo are nephron precursors. The CD24+OB-cadherin+ cell line was isolated and subjected to further immunoseparation to select 5 distinct amniotic fluid kidney progenitor cell subpopulations based on E-cadherin, podocalyxin, nephrin, TRKA and PDGFRA expression, respectively.
These subpopulations may represent different precursor cell lineages committed to specific renal cell fates. Committed progenitor cells in amniotic fluid may provide an important and novel resource of useful cells for regenerative medicine purposes.

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    ABSTRACT: Context: Amniotic Fluid Derived Stem Cells (AFSC) has mesenchymal origin and is multipotent. Having played their role in the detection of genetic abnormalities in the unborn children, they are gaining attention in the regenerative medicine because of their pluripotency. Evidence Acquisition: AFSCs possess great proliferating ability and have no ethical and religious issues in their use. AFSCs may also be studied for the stem cells differentiation such as production of multiple lineages of different cells like heart, liver, pancreas, etc. The potential of their use in regenerative medicine as well as their differentiation into multiple cells is possible. Results: AFSCs have the potential to be used in tissue repair and regeneration of bladder and kidney injuries, for the treatment of congenital anomalies like tracheal anomalies and spina bifida therapy etc. However, like every therapeutic potential, AFSCs also have some limitations such as low rate of differentiation of transplanted AFSCs and immune rejection. Conclusions: AFSCs have great therapeutic potential, but extensive research is warranted to overcome the limitations to use AFSC as therapy.
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    ABSTRACT: Amniotic fluid (AF) is a source of multipotent mesenchymal stem cells (MSCs), very promising cells for tissue engineering in clinical application. The aim of this work was to isolate and characterize cells isolated from bovine AF as alternative sources of primitive multipotent stem cells in a species that could be a large-animal model for biomedical and biotechnology researches. Samples were recovered, at slaughterhouse, from 39 pregnant cows at different trimesters of pregnancy and cells were cultured in vitro. At passages (P) 3 and 7 differentiation towards chondrogenic, osteogenic and adipogenic lineages was induced. Flow cytometry analysis for CD90, CD105, CD73, CD44, CD34, CD45 and CD14 was performed, immunocytochemistry (ICC) for Oct4, SSEA4, α-SMA, Vimentin, N- and E- Cadherin and CK and qPCR analysis for OCT4, NANOG and SOX2 were carried out. The cell yield was significantly higher in the first trimester compared to the second and the third one (P < 0.05). Cells were isolated from 25/39 samples and cell population appeared heterogeneous. Two main cell types were identified in samples from all trimesters: round- (RS) and spindle-shaped (SS) cells. 17/25 samples showed both populations (mixed, MX). Both cell types showed MSC-markers and differentiation capability with some variability related to the passages. The SS-population also expressed low levels of stemness markers such as NANOG and SSEA4 but not OCT4. Bovine AF shows a heterogeneous cell population containing also MSCs, multipotent cells that represent an intermediate stage between embryonic stem cells and adult ones.
    Stem Cell Reviews and Reports 06/2014; 10(5). DOI:10.1007/s12015-014-9525-0 · 3.21 Impact Factor
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    ABSTRACT: Congenital malformations are the leading cause of deaths during the neonatal period. Infants with prenatally diagnosed soft tissue defects may benefit from readily available autologous tissue for surgical implantation perinatally. In this study we investigated the cell content of amniotic fluid (AF) and its suitability for isolation and expansion of autologous cells for clinical use. Second trimester AF was obtained at routine amniocentesis at mean gestational week+day 16+2 (n= 54). To investigate the cell content, freshly harvested AF cells (AFC) were analyzed for different cell lineages by flow cytometry. To evaluate the isolation and expansion potential of AFC, isolation by plastic adherence was evaluated with three cell culture media and positive selection of CD117, CD133, CD271 and fibroblast cells were evaluated with Minimum Essential medium alpha (MEMα) culture media. Both the positive (+) and the negative (-) fractions were analyzed. Surface expression, senescence, immunogenicity, colony forming, proliferation and differentiation capacity was examined. In the non-hematopoietic, non-endothelial fraction of freshly harvested AF (n=17), 0.13 % cells were CD73+/CD117+ and 0.12 % CD73+/CD271+. AF displayed large donor variability with varying isolation and expansion success (n=30). The proliferative, differentiative, colony forming capacity and time before senescence also showed high variance. AFC had a preference for osteogenic rather than adipogenic lineages. Most culture-expanded AFC expressed mesenchymal but not hematopoietic surface epitopes. AFC expanded in DMEM showed higher immunogenic characteristics. This study shows that AF is a heterogeneous cell source, with high donor variation. Therefore it may not be the best source for autologous cell therapy..
    Stem Cells and Development 02/2015; DOI:10.1089/scd.2014.0426 · 4.20 Impact Factor

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