Novel Insights into Concepts and Directionality of Maternal–Fetal Cholesterol Transfer across the Human Placenta
Abstract
:1. Introduction
2. Results and Discussion
2.1. Characterization of Isolated Trophoblasts and Efficiency of In Vitro Differentiation
2.1.1. Cell Morphology
2.1.2. Protein Expression of Specific Cell Type Markers
2.1.3. Secretion of Syncytial Factors
2.2. Cholesterol Transport in CTB and STB
2.2.1. Gene Expression Studies
2.2.2. Cholesterol Uptake: Inward Transport
2.2.3. Cholesterol Efflux: Outward Transport
2.3. Directionality of the Cholesterol Transport and ABCA1 Localization
3. Materials and Methods
3.1. Human Placental Tissue Collection
3.2. Isolation of Primary Trophoblasts and Cell Culture
3.3. Cell Morphology
3.4. Flow Cytometry Analysis of Cell Purity
3.5. Quantitative RT-PCR
3.6. ELISA for hCG Secretion
3.7. Cholesterol Transport Studies
3.7.1. Cholesterol Transport in Conventional Plates
3.7.2. Cholesterol Transport in Transwell® System
3.8. Immunofluorescent Confocal Microscopy
3.9. Statistics
4. Conclusions
Author Contributions
Acknowledgments
Conflicts of Interest
References
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Gene | Accession Number | Primer Pairs (5’ end to 3’ end) | Product Length (bp) |
---|---|---|---|
ABCA1 | XM_011518342.3 | For: CCACATTTTTGCCTGGGACG Rev: AGCGATTCTCCCCAAACCTT | 88 |
ApoA-1 | NM_001318021.1 | For: AGCGGCAGAGACTATGTG Rev: CTGTCCCAGTTGTCAAGG | 83 |
Endoglin | NM_001114753.2 | For: CAAGACCAGGAAGTCCATA Rev: CGTGTGCGAGTAGATGTA | 174 |
LDL-R | XM_011528010.2 | For: GACGTGGCGTGAACATCTG Rev: CTGGCAGGCAATGCTTTGG | 111 |
SR-BI | NM_005505.4 | For: CGGCTCGGAGAGCGACTAC Rev: GGGCTTATTCTCCATGATCACC | 76 |
SREBP-2 | XM_017028922.2 | For: CCTTCCTCTGCCTCTCCTTT Rev: CACAAAGACGCTCAGGACAA | 200 |
YWHAZ | XM_024447266.1 | For: CCGTTACTTGGCTGAGGTTG Rev: AGTTAAGGGCCAGACCCAGT | 143 |
Steady State | LXR Agonist Stimulation | |||
---|---|---|---|---|
CTB | STB | CTB | STB | |
mRNA Levels | ||||
SR-B1 (×10−3) | 13.4 ± 5.90 | 15.0 ± 5.95 A | 16.4 ± 4.90 X | 21.2 ± 5.98 B,Y |
LDL-R (×10−3) | 14.8 ± 8.7 a,* | 15.8 ± 10.9 A | 67.1 ± 54.6 b,* | 44.5 ± 25.9 B |
ABCA1 (×10−3) | 13.7 ± 8.76 a | 21.0 ± 20.4 A | 132.8 ± 76.9 b | 117.8 ± 42.0 B |
SREBP-2 (×10−2) | 3.6 ± 2.2 a | 4.7 ± 4.4 A | 8.6 ± 5.5 b | 7.4 ± 3.2 B |
ApoA-1 (×10−5) | 6.9 ± 3.6 | 9.7 ± 12 | 6.6 ± 5 | 6.3± 2.7 |
Cholesterol Uptake (CU) | ||||
% CU | 13.5 ± 3.60 | 15.1 ± 3.98 A | 13.4 ± 3.75 X | 21.8 ± 3.49 B,Y |
Correlation Coefficients | ||||
CU vs. SR-B1 mRNA | r = −0.3; p = 0.302 | r = 0.5; p = 0.094 | r = 0.2; p = 0.478 | r = 0.2; p = 0.56 |
CU vs. LDL-R | r = −0.52; p = 0.078 | r = −0.16; p = 0.60 | r = 0.59; p = 0.04 | r = −0.07; p = 0.80 |
CU vs. SREBP-2 mRNA | r = 0.7; p = 0.010 | r = 0.6; p = 0.041 | r = 0.8; p = 0.004 | r = 0.2; p = 0.57 |
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Kallol, S.; Huang, X.; Müller, S.; Ontsouka, C.E.; Albrecht, C. Novel Insights into Concepts and Directionality of Maternal–Fetal Cholesterol Transfer across the Human Placenta. Int. J. Mol. Sci. 2018, 19, 2334. https://doi.org/10.3390/ijms19082334
Kallol S, Huang X, Müller S, Ontsouka CE, Albrecht C. Novel Insights into Concepts and Directionality of Maternal–Fetal Cholesterol Transfer across the Human Placenta. International Journal of Molecular Sciences. 2018; 19(8):2334. https://doi.org/10.3390/ijms19082334
Chicago/Turabian StyleKallol, Sampada, Xiao Huang, Stefan Müller, Corneille Edgar Ontsouka, and Christiane Albrecht. 2018. "Novel Insights into Concepts and Directionality of Maternal–Fetal Cholesterol Transfer across the Human Placenta" International Journal of Molecular Sciences 19, no. 8: 2334. https://doi.org/10.3390/ijms19082334