Expression of the mRNAs and Proteins for the Na+/H+ Exchangers and Their Regulatory Factors in Baboon and Human Placental Syncytiotrophoblast

Abstract

In polarized epithelial cells of several organ systems, e.g. the kidney, a family of Na(+)/H(+) exchangers (e.g. Na(+)/H(+) exchanger-1 and -3) and their regulatory proteins, Na(+)/H(+) exchanger regulatory factor and Na(+)/H(+) exchanger-3 kinase A regulatory protein play a major role in regulating Na(+)/H(+) exchange integral to cellular homeostasis. Because the primate placenta regulates exchange of Na(+) and H(+) between the mother and fetus critical to fetal-placental homeostasis, the current study determined whether Na(+)/H(+) exchanger-1 and -3 were compartmentalized and associated with expression of Na(+)/H(+) exchanger regulatory factor and Na(+)/H(+) exchanger-3 kinase A regulatory protein in baboon and human syncytiotrophoblast. Using RT-PCR, single 413-bp Na(+)/H(+) exchanger-1 and 190-bp Na(+)/H(+) exchanger-3 products were expressed by baboon and human syncytiotrophoblasts. The 104-kDa Na(+)/H(+) exchanger-1 protein was detected by Western blot in microvillus membranes and to a much lesser extent in the basal membranes of the baboon and human syncytiotrophoblasts. In contrast, the 85-kDa Na(+)/H(+) exchanger-3 protein was detected primarily in membranes contiguous with the basal membranes of the syncytiotrophoblast of both species. Differential localization of Na(+)/H(+) exchanger-1 and -3 was confirmed by immunocytochemistry. The Na(+)/H(+) exchanger-3 regulatory protein, Na(+)/H(+) exchanger-3 kinase A regulatory protein, resided almost exclusively in the basal membranes, whereas Na(+)/H(+) exchanger regulatory factor was localized primarily to the microvillus membranes in the baboon and human syncytiotrophoblast. Collectively, these results are the first to show that the baboon and human term placental syncytiotrophoblast expressed the mRNAs and proteins for Na(+)/H(+) exchanger-1 and -3 and their regulatory factors and that Na(+)/H(+) exchanger-1 and Na(+)/H(+) exchanger regulatory factor resided primarily in the microvillus membranes, whereas Na(+)/H(+) exchanger-3 and Na(+)/H(+) exchanger-3 kinase A regulatory protein were localized to membranes contiguous with the basal membranes and to the basal membranes, respectively. We conclude that a complete Na(+)/H(+) exchange system is present in the baboon and human term placental syncytiotrophoblast and suggest that the primate placenta exhibits polarity with respect to the capacity for regulation of Na(+)/H(+) exchange between the placenta and the maternal and fetal circulations.

Predominant Basal Directional Release of Thromboxane, but not Prostacyclin, by Placental Trophoblasts from Normal and Preeclamptic Pregnancies

“…Isolated trophoblasts were purified by Percoll (Sigma, St. Louis, MO) gradient centrifugation. Contaminated red blood cells were eliminated by incubation of isolated trophoblasts with lysis buffer (155mM NH 4 Cl, 10mM KHCO 3 and 0.14M MDTA, pH7.2) on ice for 5 minutes.…”

“…Emerging evidence has shown that trophoblasts are polarized epithelial cells with distinct functional regulations between the microvillous and the basal membranes (1)(2)(3)(4). For example, lactate produced by trophoblasts is a source of energy fuel to the fetus.…”

“…A study of trophoblast lactate transporter activities/expressions by Settle et al (3) revealed differential localizations of lactate transporter subtypes on microvillous and basal membranes, which corresponded to their roles in the lactate production, lactate release to and lactic acid removal from the fetal circulation (3). Na+/H+ exchangers also display a differential distribution across the microvillous and basal membranes (4). Differential releases of matrix metalloproteinase-2 (MMP-2) and −9 (MMP-9) by syncytiotrophoblasts in culture was reported by Sawicki et al (1).…”

Regulation of Expression and Localisation of the Na+/H+ Exchanger (NHE) 3 and the NHE Regulatory Factor 2 in Baboon Placental Syncytiotrophoblast by Oestrogen

“…After removal of the decidua basalis and chorionic plate, 6-7 randomly selected sections of villous placenta were placed in phosphate buffered formalin (Sigma Chemical Co, St. Louis, MO) for immunocytochemistry and 60-70 grams of whole villous tissue used for preparation of fractions of syncytiotrophoblast MVM, cytosol presumably containing endosomes (10,000 × g; 10K), and cytosol containing endosomes and membranes contiguous with the basal membrane (BMm) and the BM/FVE as described previously [4,9]. Although we [4,13] previously showed using these procedures that alkaline phosphatase activity was enriched in the MVM and minimal in other fractions, whereas binding of dihydroalprenolol (DHAP) was enriched in the BM/FVE fraction, activity/binding was also determined in syncytiotrophoblast membrane fractions of animals of the current study.…”

“…Moreover, the specific concentrations of NHE1 and its regulatory factor NHERF1, both localized primarily to the microvillus membrane (MVM), were similar at mid and late gestation in association with a 4-fold increase in MVM alkaline phosphatase activity and presumably microvillus surface area as shown in human placenta [5,6]. Despite the physiologic importance of the placental NHE system [3,[7][8][9], our understanding of the factors controlling NHE/NHERF expression and localization in the primate syncytiotrophoblast is incomplete.…”