Donor mesenchymal stem cells (MSCs) have been documented in fetal and maternal circulations after plain intra-amniotic injection, with diverse therapeutic effects. We sought to determine the pathway of this unique cell kinetic route. Rat fetuses (n = 226) were divided into two groups based on the content of intra-amniotic injections performed on gestational day 17 (E17): either a concentrated suspension of luciferase-labeled syngeneic amniotic fluid-derived MSCs (afMSCs; n = 111), or acellular luciferase (n = 115). Samples from placenta, chorion, amnion, amniotic fluid, stomach fluid, peripheral blood, and umbilical cord were procured at five daily time points thereafter until term (E18–22) for luminometry. In addition, 53 sets of fresh gestational membranes (chorion/amnion combined) from nonmanipulated term fetuses were secured to transwell inserts for in vitro analysis of MSC migration using luciferase-labeled afMSCs. Statistical analyses included the Mann-Whitney U-test, Wald test, nonlinear regression modeling, and Fisher's exact test. In vivo, luciferase activity was observed in the amnion, chorion, and placenta of fetuses receiving cells, but not in those receiving acellular luciferase (P < 0.001). There was a consistent nonlinear age-dependent relationship of luciferase activity between the amnion, chorion, and placenta following a parabolic bimodal pattern characterized by significantly higher early preterm (E18) and late-term (E22) activities (P < 0.001), with no differences between E21 and E22 (P = 0.12). In vitro, the presence of cells was documented by luminometry in 21/53 (39.6%) of the assays, in suspension and/or attached to the plastic substrate, and within all screened gestational membrane sets, irrespective of stimuli with collagen coating or fetal bovine serum. We conclude that, after intra-amniotic injection, donor MSCs undergo controlled cell routing, as opposed to passive clearance. Transgestational membrane transport appears to constitute the path for donor cells to reach the placenta, a known gateway to the fetal circulation, significantly expanding the potential applications of transamniotic stem cell therapy.

中文翻译：

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羊水干细胞治疗后的血供体细胞路由途径。

普通羊膜腔内注射后，在胎儿和母体循环中已记录了供体间充质干细胞（MSCs），具有多种治疗作用。我们试图确定这种独特的细胞动力学途径的途径。 根据在妊娠第17天（E17）进行羊膜内注射的内容，将大鼠胎儿（n = 226）分为两组：荧光素酶标记的同基因羊水来源的MSC的浓缩悬浮液（afMSCs；n  = 111） ）或无细胞荧光素酶（n = 115）。此后每天五个时间点从胎盘，绒毛膜，羊膜，羊水，胃液，外周血和脐带中采集样品，直到足月（E18-22）进行光度测定。另外，将53组来自未操纵足月胎儿的新鲜妊娠膜（绒毛膜/羊膜结合体）固定在transwell插入物上，以使用荧光素酶标记的afMSCs体外分析MSC的迁移。统计分析包括Mann-Whitney U检验，Wald检验，非线性回归建模和Fisher精确检验。在体内，在接受细胞的胎儿的羊膜，绒毛膜和胎盘中观察到萤光素酶活性，但在接受脱细胞萤光素酶的胎儿中则未见（P <0.001）。在抛物线双峰模式下，萤光素酶活性在羊膜，绒毛膜和胎盘之间存在非线性的年龄依赖性关系，其特征是早早（E18）和后期（E22）活动明显较高（P  <0.001），无E21和E22之间的差异（P = 0.12）。在体外，通过光度法在21/53（39.6％）的测定中，悬浮和/或附着在塑料底物上以及所有筛查的胎膜组中，不论是否有胶原蛋白涂层或胎儿刺激，都通过光度法记录了细胞的存在。牛血清。我们得出的结论是，在羊膜内注射后，供体MSC经历了受控的细胞路由，而不是被动清除。妊娠膜转运似乎构成供体细胞到达胎盘的途径，胎盘是通往胎儿循环的已知通道，大大扩大了跨羊膜干细胞疗法的潜在应用范围。