Rat visceral yolk sac cells: viability and expression of cell markers during maternal diabetes

Abstract

The function of the visceral yolk sac (VYS) is critical for embryo organogenesis until final fetal development in rats, and can be affected by conditions such as diabetes. In view of the importance of diabetes during pregnancy for maternal and neonatal health, the objective of this study was to assess fetal weight, VYS cell markers, and viability in female Wistar rats (200-250 g) with induced diabetes (alloxan, 37 mg/kg) on the 8th gestational day (gd 8). At gd 15, rats from control (n=5) and diabetic (n=5) groups were anesthetized and laparotomized to remove the uterine horns for weighing of fetuses and collecting the VYS. Flow cytometry was used for characterizing VYS cells, and for determining mitochondrial activity, cell proliferation, DNA ploidy, cell cycle phases, and caspase-3 activity. Fetal weight was reduced in the diabetic group. Expression of the cell markers CD34, VEGFR1, CD115, CD117, CD14, CCR2, CD90, CD44, STRO-1, OCT3/4, and Nanog was detected in VYS cells in both groups. In the diabetic group, significantly decreased expression of CD34 (P<0.05), CCR2 (P<0.001), and OCT3/4 (P<0.01), and significantly increased expression of CD90 (P<0.05), CD117 (P<0.01), and CD14 (P<0.05) were observed. VYS cells with inactive mitochondria, activated caspase-3, and low proliferation were present in the rats with diabetes. Severe hyperglycemia caused by maternal diabetes had negative effects on pregnancy, VYS cell viability, and the expression of cell markers.

Diabetes affects rat visceral yolk sac cells viability and cell markers expression

“…Experimentally-induced diabetes in pregnant animals, especially rodents, results in similar abnormalities to those observed in humans [11,12] . In Aires et al [10] study, diabetes was induced by one alloxan injection at gestational day 8 (8 gd) and at 15 gd the collection of fetuses and VYS was made. The pregnant rats showed marked hyperglycemia and reduced fetal weight, which is a common finding in severe diabetes [13] .…”

“…Aires et al [10] showed the expression of stromal (CD90, CD44) and pluripotency (STRO-1, OCT3/4, NANOG) markers on VYS cells in both the control and diabetic groups confirming the existence of MSC in the rat VYS. However in diabetes, the functional significance of the increased CD90 expression and reduced OCT3/4 expression by VYS cells is not fully understood but could influence the mesenchymal niche and the cell differentiation.…”

“…An important work showed that VYS mitochondrial function is maintained in normal pregnancy, even after the establishment of the chorioallantoic circulation (12 gd) [9] ; therefore, the reduction of VYS mitochondrial activity in diabetes may be harmful for embryo nutrition. Aires et al [10] showed the expression an increased number of VYS cells with inactive mitochondria (unviable cells) in the diabetic condition, indicating a reduction of mitochondrial transmembrane potential. An increase in sub-haploid VYS cells in the diabetic group corresponds to apoptotic cells with fragmented DNA or condensed chromatin [23] , as evidenced by the significant amount of activated caspase-3 cells in that group.…”

“…In rodents, the visceral yolk sac (VYS) acts as an active region for metabolic exchange and nutrition uptake [6,7,8] , becoming an important tissue that persists active during the days following the placentation period from gestational day 12 (12 gd) onwards [9] . Recently, Aires et al [10] expanded these studies by showing the effect of severe hyperglycemia induced by alloxan on pregnancy, VYS cell viability and cell marker expression at 15 gd in rats.…”

Diabetes consequences in the fetus liver of the non-obese diabetic mice

“…Fluorescence-activated cell sorting analysis was performed as reported previously 9 with primary antibodies: goat polyclonal anti-ALB (sc-8108, Santa Cruz, CA, USA), goat polyclonal anti-AFP (sc-46293, Santa Cruz, CA, USA) at 4 °C overnight. The corresponding isotype antibody was used as negative control and as a secondary antibody was used rabbit anti-goat IgG: Alexa 594 (Immuny, Campinas, Brazil).…”