Interplay between β-carotene and lipoprotein metabolism at the maternal-fetal barrier

Abstract

Vitamin A is an essential nutrient, critical for proper embryonic development in mammals. Both embryonic vitamin A-deficiency or -excess lead to congenital malformations or lethality in mammals, including humans. This is due to the defective transcriptional action of retinoic acid, the active form of vitamin A, that regulates in a spatial- and temporal-dependent manner the expression of genes essential for organogenesis. Thus, an adequate supply of vitamin A from the maternal circulation is vital for normal mammalian fetal development. Provitamin A carotenoids circulate in the maternal bloodstream and are available to the embryo. Of all the dietary carotenoids, β-carotene is the main vitamin A precursor, contributing at least 30% of the vitamin A intake in the industrialized countries and often constituting the sole source of retinoids (vitamin A and its derivatives) in the developing world. In humans, up to 40% of the absorbed dietary β-carotene is incorporated in its intact form in chylomicrons for distribution to other organs within the body, including the developing tissues. Here, it can serve as a source of vitamin A upon conversion into apocarotenoids by its cleavage enzymes. Given that β-carotene is carried in the bloodstream by lipoproteins, and that the placenta acquires, assembles and secretes lipoproteins, it is becoming evident that the maternal-fetal transfer of β-carotene relies on lipoprotein metabolism. Here, we will explore the current knowledge about this important biological process, the cross-talk between carotenoid and lipid metabolism in the context of the maternal-fetal transfer of this provitamin A precursor, and the mechanisms whereby β-carotene is metabolized by the developing tissues. This article is part of a Special Issue entitled Carotenoids recent advances in cell and molecular biology edited by Johannes von Lintig and Loredana Quadro.

Keywords: Lipoproteins; Microsomal triglyceride transfer protein (MTP); Retinoids; β-Carotene; β-Carotene-15,15′‑oxygenase (BCO1); β-Carotene-9′,10′‑oxygenase (BCO2).

Figure 1.. Symmetric and asymmetric cleavage of β-carotene.

Central cleavage of β-carotene at the 15,15’ double bond is catalyzed by BCO1 to yield two molecules of retinal. Asymmetric cleavage at the 9’,10’ double bond is catalyzed by BCO2, and yields β-apo-10’-carotenal (depicted) and β-ionone (not shown). Retinaldehyde can be oxidized to retinoic acid or reduced to retinol, which can then be esterified to form retinyl esters.

Figure 2.. MTP transfers β-carotene.

β-carotene-containing donor vesicles were made by mixing 100 μl of β-carotene (stock 10 mg/mL in chloroform) with 70 μl of phosphatidylcholine (stock 20 mg/mL in chloroform), evaporated under nitrogen and then resuspended and sonicated in 4.5 mL of 15 mM Tris-HCl buffer, pH 7.4, 1 mM EDTA, 150 mM NaCl, 0.02% NaN₃. For the assay, 20 μL of β-carotene-containing donor vesicles (DV) and 5 μL of acceptor vesicles (AV) containing only phosphatidylcholine were used, along with 50 μL BSA (10 mg/mL).. 1 μg purified bovine liver MTP protein (A) or 200 μg mouse liver homogenate protein (B and C) were incubated for 18 h (for BC) or 1 h (for TG) in water bath at 37 C. Excitation at 355 nm and emission at 535 nm were used to measure β-carotene. TG transfer was measured as reported [53]. Data are means ± SD; n=3 replicates/condition; WT, wild-type C57Bl/6J liver homogenates; KO, MTP-deficient liver homogenates (wild type C57Bl/6J background). TG, triglycerides; BC, β-carotene. Statistical analysis within each panel by t-test, *, p<0.05.

Figure 3.. Proposed model for maternal fetal transfer of intact β-carotene (β-car) upon a single maternal administration of β-carotene.

Intact β-carotene from the maternal bloodstream is transported toward the fetal circulation in association with lipoproteins assembled within the placenta syncytiotrophoblast cells. Under condition of vitamin A sufficiency, the lipoprotein receptors LRP1 and VLDLR presumably play a role in the uptake. Placental BCO1 cleaves the provitamin A carotenoid symmetrically to yield retinaldehyde (not shown), which in turn is oxidized to retinoic acid. Through this pathway, retinoic acid levels increase in placenta early after β-carotene administration (4h), thus initially suppressing MTP transcription and activity. Asymmetric cleavage of β-carotene by BCO2 also occurs, generating one β-ionone ring and β-apo-10’carotenal (apo10AL). Apo10AL levels rise as retinoic acid declines. At 24h post maternal administration of β-carotene apo10AL then increases the transcription and activity of placental MTP, which in turn stimulates lipoprotein biosynthesis and ultimately the transfer of β-carotene toward the fetal circulation. This temporally-regulated generation of apo10AL vs. retinoic acid from β-carotene in placenta and their antagonistic transcriptional activity on MTP are proposed as the key factors that optimize placental lipoprotein biogenesis and lipoprotein-mediated transfer of β-carotene to the fetus. apoB, apolipoprotein B-100; FA, fatty acids; CE, cholesteryl esters; Syncyt I, syncytiotrophoblast layer I; Syncyt II, syncytiotrophoblast layer II; TG, triglycerides.

Figure 4.. β-carotene suppresses placental lipoprotein biosynthesis under a maternal regimen of high vitamin A intake during pregnancy.

Tissues from wild-type dams on the vitamin A excess (220 IU of vitamin A/g diet) or sufficient (14 IU of vitamin A/g diet) diet during pregnancy with or without β-carotene (BC) supplementation by intraperiotenal injection (or vehicle, Veh) are from previously published studies from our laboratories [72,84]. (A) Placental qPCR analysis of Mttp, its transcriptional activator Hnf4a, and apoB in WT dams on a vitamin A sufficient or excess diet administered vehicle (Veh) or β-carotene at mid-gestation and sacrificed after 24 h. Data are presented as mean ± S.D. of duplicate determinations and are representative of three independent determinations. n= 1–2 placentas/dam from four to six wild-type dams per group. Statistical analysis by ANOVA. *, p < 0.05. Data for the vitamin A sufficient diet group where previously published (78). (B) Placental MTP activity expressed as percent transfer of lipids/mg/h (n = 1–3 placentas/dam from 3–4 β-carotene-treated dams). Dams as in panel A. (C) Ratio of β-carotene content in each placenta and its corresponding embryo expressed as percent. β-carotene was directly injected into the placentas of wild-type dams at 14.5 dpc, as previously described [84].. Data from dams on the vitamin A sufficient diet pretreated with the MTP inhibitor lomitapide (14 IU MTPi) or not (14 IU) were previously published [84].. β-carotene levels in placenta and embryos were measured by HPLC [80] Individual values are shown. n = 1–7 placentas/dam from 4 wild-type dams per group.