Isolation and Purification of Human Placental Plasma Membranes from Normal and Pre-eclamptic Pregnancies. A Comparative Study

Jiménez, Verónica; Henríquez, Mauricio; Llanos, Paola; Riquelme, Gloria

doi:10.1016/j.placenta.2003.10.013

Cited by 40 publications

(46 citation statements)

References 32 publications

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“…The cross-contamination of purified basal membranes with apical membranes was low, as evidenced by the lack of alkaline phosphatase enrichment in the basal fraction. The ratio of placental alkaline phosphatase activity enrichment of BM compared with that of apical membranes was 0.1 for the placentae used in this study, lower or equivalent to that from several other reports for single or paired apical and basal membrane preparations (Glazier et al, 1990;Illsley et al, 1990) as we demonstrated previously (Jimenez et al, 2004). In addition, for both MVM and BM vesicles, orientation is preferentially right-side out, as reported previously (Illsley et al, 1990).…”

Section: Introductionsupporting

confidence: 81%

“…Placentae were obtained from normal pregnancies and collected at term immediately after delivery from the San José Hospital Maternity Unit (Santiago, Chile). Human placental plasma membrane vesicles were prepared from fresh normal-term placentae by a method described previously that allows the simultaneous isolation of apical and basal membranes from the same placenta (Jimenez et al, 2004). This method involves differential centrifugation, precipitation of nonmicrovillous membranes with magnesium ions, and a sucrose gradient step.…”

Section: Introductionmentioning

confidence: 99%

“…The protein concentration was determined using a BCA protein assay kit (Thermo Fisher Scientific, Waltham, MA). The purity and enrichment of the apical and basal membrane fraction were determined routinely by assaying for classic marker protein activities as described previously (Jimenez et al, 2004). Alkaline phosphatase was used as an apical membrane marker.…”

Section: Introductionmentioning

confidence: 99%

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Expression and Distribution of Nucleoside Transporter Proteins in the Human Syncytiotrophoblast

Errasti-Murugarren¹,

Díaz²,

Godoy³

et al. 2011

Mol Pharmacol

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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.

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Section: Introductionsupporting

confidence: 81%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Expression and Distribution of Nucleoside Transporter Proteins in the Human Syncytiotrophoblast

Errasti-Murugarren¹,

Díaz²,

Godoy³

et al. 2011

Mol Pharmacol

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“…Preparation of Placental Apical Membrane Vesicles-The human placental microvillous membrane (MVM) 1 vesicles were prepared by a method we have described (22) that allows simultaneous isolation of apical and basal membranes from the same placenta. This method is a modification of the method described by Illsley et al (23).…”

Section: Methodsmentioning

confidence: 99%

Annexin 6 Modulates the Maxi-chloride Channel of the Apical Membrane of Syncytiotrophoblast Isolated from Human Placenta

Riquelme

Llanos

Tischner

et al. 2004

Journal of Biological Chemistry

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The syncytiotrophoblast separates the maternal and fetal blood and constitutes the primary barrier for maternal-fetal transport. The Maxi-chloride channel from the apical membrane of the syncytiotrophoblast plays a role in the chloride conductance. Annexins can play an important role in the regulation of membrane events. In this study we evaluate the role of annexin 6 in the Maxichloride channel properties. The results showed that annexin 6 is bound in the apical placenta membranes in a calcium-dependent phospholipid-binding manner but also in a calcium-independent fashion. The neutralization of annexin 6 decreased the total current by 39 ؎ 1.9% in the range of ؎80 mV, and the currents decrease with the time. The single-channel slope conductance was decreased from 253 ؎ 7.4 pS (control) to 105 ؎ 13 pS, and the amplitude decreased by 50%. The open probability was also affected when higher voltage steps were used, changes in either the positive or negative direction induced the channel to close, and the open probability (P o ) did not decrease. In channels with neutralized annexin 6, it was maintained at 1 at ؎40 mV and at ؎80 mV. These results suggest that endogenous annexin 6 could regulate the Maxi-chloride channel. The results obtained with normal placentae, in which annexin 6 was neutralized, are similar to those described for the Maxichloride channel isolated from pre-eclamptic placenta. Together these data suggest that annexin 6 could play an important role in ion transport of the placenta.

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“…The placental plasma membrane homogenates were prepared following a described protocol (Jimenez et al, 2004) with some modifications. Firstly, fetal placental pieces of the fetal face were thawed, and the amount needed was taken (~10 g was used for each assay).…”

Section: Placental Plasma Membrane Homogenates Preparationmentioning

confidence: 99%

Influence of prenatal exposure to environmental pollutants on human cord blood levels of glutamate

et al. 2014

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Alteration of the glutamatergic system may be one mechanistic pathway. We aimed to determine whether mercury and seven OCs, including PCBs 138, 153, and 180, DDT and DDE, hexachlorobenzene (HCB), and beta-hexachlorocyclohexane (β-HCH) influence the cord levels of two excitatory amino acids, glutamate and aspartate. Secondly, we evaluated if this association was mediated by glutamate uptake measured in human placental membranes. The study sample included 40 newborns from a Spanish cohort selected according to cord mercury levels. We determined the content of both amino acids in cord blood samples by means of HPLC and assessed their associations with the contaminants using linear regression analyses, and the effect of the contaminants on glutamate uptake by means of [ 3 H]-aspartate binding in human placenta samples. PCB138, β-HCH, and the sum of the 3 PCBs and seven OCs showed a significant negative association with glutamate levels (decrease of 51, 24, 56 and 54 %, respectively, in glutamate levels for each 10-fold increase in the contaminant concentration). Mercury did not show a significant correlation neither with glutamate nor aspartate levels in cord blood, however a compensatory effect between THg and both PCB138, and 4,4'-DDE was observed. The organo-metallic derivative methylmercury completely inhibited glutamate uptake in placenta while PCB138 and β-HCH partially inhibited it (IC50 values: 4.9 ± 0.8 μM, 14.2± 1.2nM and 6.9± 2.9nM, respectively).We conclude that some environmental toxicants may alter the glutamate content in the umbilical cord blood, which might underlie alterations in human development.Key words: methylmercury, polychlorobiphenyl (PCB), organochlorine pesticides, placenta, umbilical cord blood, glutamate, transport, pollutants. 3 IntroductionSocial and economic development have been mediated by strong industrial growth involving the synthesis, use and release of large numbers of chemicals many of which are potentially harmful to human health and the environment (Li and Macdonald, 2005;Porta et al., 2012).Several of these chemicals, e.g. lead, mercury, polychlorinated biphenyls (PCBs), arsenic and toluene, have been associated to subclinical brain dysfunction and neurodevelopmental disorders such as poorer cognitive and motor development, attention deficit, hyperactivity, and sensory deficits but the underlying processes leading to these effects are still unknown (Grandjean and Landrigan, 2006). Pollutants come from several different sources, move on the environment and eventually accumulate in the food chain. Daily exposure to environmental persistent pollutants results in detectable levels of metals, PCBs and organochlorine pesticides in woman bodies (Woodruff et al., 2011). During pregnancy, the pollutants accumulated by the women can reach the fetus crossing the placenta. Although the placenta regulates the exchange of nutrients and oxygen between mother and fetus and acts as a barrier of many toxicants, it does not provide protection against certain environmental contaminants (Proui...

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Isolation and Purification of Human Placental Plasma Membranes from Normal and Pre-eclamptic Pregnancies. A Comparative Study

Cited by 40 publications

References 32 publications

Expression and Distribution of Nucleoside Transporter Proteins in the Human Syncytiotrophoblast

Expression and Distribution of Nucleoside Transporter Proteins in the Human Syncytiotrophoblast

Annexin 6 Modulates the Maxi-chloride Channel of the Apical Membrane of Syncytiotrophoblast Isolated from Human Placenta

Influence of prenatal exposure to environmental pollutants on human cord blood levels of glutamate

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