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Cellular growth and tube formation of HTR8/SVneo trophoblast: effects of exogenously added fatty acid-binding protein-4 and its inhibitor

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Abstract

Adequate placental angiogenesis is critical for the establishment of the placental circulation and thus for normal feto-placental growth and development. Fatty acid-binding protein-4 (FABP4) plays a pro-angiogenic role in endothelial cells; however, very little information is available in placental first trimester trophoblast cells. Here we report that exogenously added FABP4 (exo-FABP4) stimulated tube formation (as a measure of in vitro angiogenesis) in HTR8/SVneo trophoblastic cells. HTR-8/SVneo cells were incubated in the presence of exogenously added FABP4 at different concentrations and time points. Cellular growth, proliferation, in vitro tube formation, expression of growth stimulatory-, fatty acid transporters, and angiogenic genes were investigated. Internalization of exo-FABP4 was carried out using immunocytochemistry. Radioactive fatty acid uptake was determined in the presence and absence of FABP4 metabolic inhibitor. Exo-FABP4 (10–100 ng/ml) stimulated proliferation of HTR8/SVneo cells as compared to control. Exo-FABP4 dose dependently increased growth and viability of the cells to the similar extent as done by 50 µM of arachidonic acid. Exo-FABP4-induced tube formation and proliferation were significantly inhibited by FABP4 (BMS309403) inhibitor. Exo-FABP4 stimulated the expression of growth stimulatory genes such as tissue inhibitor of matrix metalloproteinases-1 (TIMP1), insulin-like growth factor 1 (IGF1), and also prokineticin 2 (PROK2), the pro-angiogenic mediators in these cells. In addition, expressions of genes associated with proliferation and differentiation such as sonic hedgehog (SHH) and WNT1 inducible signalling pathway protein 1 (WISP1) were significantly expressed when cells were exposed to exo-FABP4. Our findings reveal a pro-angiogenic role of FABP4 in first trimester placental trophoblast cells and its regulation may have impact in placental physiology.

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Acknowledgements

We are grateful to Aud Jørgensen for her technical assistance. We are grateful to Camilla Solberg for her help in thymidine assay. The authors convey their thanks to Srinivas Vilasagaram for his assistance in siRNA experiments. This study was supported by the grant from the Thune Holst Foundation and HRD fellowship, Department of Health Research (Dr. Sanjay Basak), Government of India.

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Authors and Affiliations

  1. Department of Nutrition, Faculty of Medicine, Institute of Basic Medical Sciences, University of Oslo, Oslo, Norway

    Sanjay Basak, Arnab Sarkar, Santosh Mathapati & Asim K. Duttaroy

  2. Molecular Biology Division, National Institute Nutrition, Indian Council of Medical Research, Hyderabad, India

    Sanjay Basak

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  1. Sanjay Basak
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  2. Arnab Sarkar
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Correspondence to Asim K. Duttaroy.

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Supplementary material 1 (PPTX 108 kb)

Supplementary Fig. 1. Silencing of FABP4 mRNA in the first trimester trophoblast cell line, HTR8/SVneo and endothelial cell line, EA.hy926 Cells (104 cells/well/96 well) were transfected with scramble siRNA (control) and target siRNAs (FABP4 or Cyclophilin B) at final concentration of 25 nM for 48 h. Knockdown efficiency was measured by mRNA expression levels of FABP4 or Cyclophilin after normalized with TBP (endogenous control) in both HTR8/SVneo and EA.hy926 cells . A, B Fold mRNA expression of cyclophilin B (positive control), and FABP4 are presented for HTR8/SVneo cells and C FABP4 level for EA.hy926 cells after silencing of the respective genes for 48 h

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Basak, S., Sarkar, A., Mathapati, S. et al. Cellular growth and tube formation of HTR8/SVneo trophoblast: effects of exogenously added fatty acid-binding protein-4 and its inhibitor. Mol Cell Biochem 437, 55–64 (2018). https://doi.org/10.1007/s11010-017-3095-9

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