The plasma membrane distribution and related biological activity of nucleoside transporter proteins (NTs) were investigated in human syncytiotrophoblast from term placenta using a variety of approaches, including nucleoside uptake measurements into vesicles from selected plasma membrane domains, NT immunohistochemistry, and subcellular localization (basal, heavy, and light apical membranes as well as raft-enriched membranes from the apical domain). In contrast with other epithelia, in this epithelium, we have identified the high-affinity pyrimidine-preferring human concentrative nucleoside transporter (hCNT) 1 as the only hCNT-type protein expressed at both the basal and apical membranes. hCNT1 localization in lipid rafts is also dependent on its subcellular localization in the apical plasma membrane, suggesting a complex cellular and regional expression. Overall, this result favors the view that the placenta is a pyrimidine-preferring nucleoside sink from both maternal and fetal sides, and hCNT1 plays a major role in promoting pyrimidine salvage and placental growth. This finding may be of pharmacological relevance, because hCNT1 is known to interact with anticancer nucleoside-derived drugs and other molecules, such as nicotine and caffeine, for which a great variety of harmful effects on placental and fetal development, including intrauterine growth retardation, have been reported.

We report on the characteristics of raft domains in the apical membrane from human placental syncytiotrophoblast (hSTB), an epithelium responsible for maternal-fetal exchange. Previously, we described two isolated fractions of the hSTB apical membrane: a classical microvillous membrane (MVM) and a light microvillous membrane (LMVM). Detergent-resistant microdomains (DRMs) from MVM and LMVM were prepared with Triton X-100 followed by flotation in a sucrose gradient and tested by Western and dot blot with raft markers (placental alkaline phosphatase, lipid ganglioside, annexin 2) and transferrin receptor as a nonraft marker. DRMs from both fractions showed a consistent peak for these markers, except that the DRMs from MVM had no annexin 2 mark. Cholesterol depletion modified the segregation in both groups of DRMs. Our results show two distinguishable lipid raft subsets from MVM and LMVM. Additionally, we found significant differences between MVM and LMVM in cholesterol content and in expression of cytoskeletal proteins. MVM is enriched in ezrin and beta-actin; in contrast, cholesterol and cytokeratin-7 are more abundant in LMVM. These differences may explain the distinct properties of the lipid raft subtypes.

Intrauterine growth restriction (IUGR) and preeclampsia (PE) are leading causes of perinatal and maternal morbidity and mortality. Previously we reported the expression of lipid rafts in classical microvillous membrane (MVM) and light microvillous membrane (LMVM), two subdomains in apical membrane from the human placental syncytiotrophoblast (hSTB), which constitute the epithelium responsible for maternal-fetal transport. Here the aim was to study the raft and cytoskeletal proteins from PE and IUGR. Microdomains from MVM and LMVM were tested with raft markers (placental alkaline phosphatase, lipid ganglioside, and annexin 2) and a nonraft marker (hTf-R). No changes were detected with those markers in whole purified apical membranes in normal, PE, and IUGR pregnancies; however, their patterns of distribution in lipid rafts were different in PE and IUGR. Cholesterol depletion modified their segregation, confirming their presence in lipid rafts, although unlike normal placenta, in these pathologies there is only one type of microdomain. Additionally, the cytoskeleton proteins actin, ezrin, and cytokeratin-7 showed clear differences between normal and pathological membranes. Cytokeratin-7 expression decreased to 50% in PE, and the distribution between LMVM and MVM (~43 and 57%, respectively) changed in both PE and IUGR, in contrast with the asymmetrical enrichment obtained in normal LMVM (~62%). In conclusion, lipid rafts from IUGR and PE have different features compared to rafts from normal placentae, and this is associated with alterations in the expression and distribution of cytoskeletal proteins.

No abstract

No abstract

No abstract