RESUMO
The ovine developmental model represents the standard in vivo model for studies involving maternofetal physiology, amniotic fluid (AF) research, and fetal cell therapy prior to human clinical use. Although being close to the human fetal anatomy, 2 separate extraembryonic fluid compartments remain during gestation, known as the amnion and the allantois. A clear distinction between AF versus allantoic fluid (AL) is therefore indispensable for correct scientific conclusions with regard to human translation. In the presented study, the biochemical composition of AF and AL was evaluated in ovine gravid uteri postmortem (n = 31) over the entire gestation. Four parameters, consisting of Na+, Cl-, Mg2+, and total protein, have been found to allow for specific discrimination of the 2 fetal fluids at all gestational phases and therefore as potential surrogate parameters for gestational age. In addition, volumetric changes of the developing fetus and the 2 fetal fluid cavities were analyzed by contrast-enhanced computed tomography (n = 12). AF showed a significant, linear volumetric increase over gestation, whereas AL volume maintained relatively static independent of gestational age. These results serve as a basis for future studies by providing surrogate markers enabling a reliable distinction of isolated fetal fluids and contained cells in the ovine developmental model over the entire gestation.
RESUMO
Amniotic fluid represents an abundant source of multipotent stem cells, referred as broadly multipotent given their differentiation potential and expression of pluripotency-related genes. However, the origin of this broadly multipotent cellular fraction is not fully understood. Several sources have been proposed so far, including embryonic and extraembryonic tissues. In this regard, the ovine developmental model uniquely allows for direct comparison of fetal fluid-derived cells from two separate fetal fluid cavities, the allantois and the amnion, over the entire duration of gestation. As allantoic fluid mainly collects fetal urine, cells originating from the efferent urinary tract can directly be compared with cells deriving from the extraembryonic amniotic tissues and the fetus. This study shows isolation of cells from the amniotic [ovine amniotic fluid cells (oAFCs)] and allantoic fluid [ovine allantoic fluid cells (oALCs)] in a strictly paired fashion with oAFCs and oALCs derived from the same fetus. Both cell types showed cellular phenotypes comparable to standard mesenchymal stem cells (MSCs), with trilineage differentiation potential, and expression of common ovine MSC markers. However, the expression of MSC markers per single cell was higher in oAFCs as measured by flow cytometry. oAFCs exhibited higher proliferative capacities and showed significantly higher expression of pluripotency-related genes OCT4, STAT3, NANOG, and REX1 by quantitative real-time polymerase chain reaction compared with paired oALCs. No significant decrease of pluripotency-related gene expression was noted over gestation, implying that cells with high differentiation potential may be isolated at the end of pregnancy. In conclusion, this study suggests that cells with highest stem cell characteristics may originate from the fetus itself or the amniotic fetal adnexa rather than from the efferent urinary tract or the allantoic fetal adnexa.