Balakrishnan Chakrapani Narmadaa, Yeek Teck Goha, Huan Lib, Sanjay Sinhac,d, Hanry Yube,f,g and Christine Cheunga,h
a Institute of Molecular and Cell Biology, Proteos, Singapore
b Institute of Bioengineering and Nanotechnology, Nanos, Singapore
c The Anne McLaren Laboratory of Regenerative Medicine, Wellcome Trust-Medical Research Council Cambridge Stem Cell Institute, University of Cambridge, Cambridge, United Kingdom
d Division of Cardiovascular Medicine, Addenbrooke's Hospital, University of Cambridge, Cambridge, United Kingdom
e Department of Physiology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore
f Mechanobiology Institute, Singapore
g Singapore-MIT Alliance for Research and Technology, BioSyM, Singapore
h Lee Kong Chian School of Medicine, Nanyang Technological University, Singapore
Correspondence: Christine Cheung (email@example.com)
Published in Stem Cells Translational Medicine 7th October 2016
Atherosclerosis underlies many cardiovascular and cerebrovascular diseases. Nutraceuticals are emerging as a therapeutic moiety for restoring vascular health. Unlike small molecule drugs, the complexity of ingredients in nutraceuticals often confounds evaluation of their efficacy in preclinical evaluation. It is recognized that liver is a vital organ in processing complex compounds into bioactive metabolites. In this work, we developed a co-culture system of human pluripotent stem cell-derived endothelial cells (hPSC-ECs) and hepatocytes (hPSC-HEPs) for predicting vascular-protective effects of nutraceuticals. To validate our model, two compounds (quercetin and genistein) known to have anti-inflammatory effects on vasculatures, were selected. We found that both quercetin and genistein were ineffective at suppressing inflammatory activation by interleukin-1β due to limited metabolic activity of hPSC-ECs. On the other hand, hPSC-HEPs demonstrated metabolic capacity to break down both nutraceuticals into primary and secondary metabolites. When hPSC-HEPs were co-cultured with hPSC-ECs to permit paracrine interactions, the continuous turnover of metabolites mitigated interleukin-1β stimulation on hPSC-ECs. We observed significant reductions in inflammatory gene expressions, nuclear translocation of nuclear factor kappa B, and interleukin-8 production. Thus, integration of hPSC-HEPs could accurately reproduce systemic effects involved in drug metabolism in vivo to unravel beneficial constituents in nutraceuticals. This physiologically-relevant endothelial-hepatic platform would be a great resource in predicting the efficacy of complex nutraceuticals and mechanistic interrogation of vascular-targeting candidate compounds.
Figure Legend: Figure 5. Coculture with H9-embryonic stemcell-derived hepatocytes (H9-ESC-HEPs) abrogates inflammatory activation in IL-1b-stimulated H9-embryonic stem cell-derived endothelial cells (H9-ESC-ECs). (A): Schematics showing two experimental setups to study endothelial-hepatic paracrine
interactions. (Bi): Immunostaining for NFkB in unstimulated and IL-1b-stimulated hPSC-ECs, treated with or without quercetin and genistein.
NFkB, red; nucleus, blue. Scale bars = 100 mm. (Bii): Quantification of NFkB nuclear translocation shows that quercetin or genistein significantly decreased levels of NFkB nuclear colocalization in coculture of hPSC-ECs with hPSC-HEPs. (C): IL-8 protein levels were significantly reduced in endothelial-hepatic coculture but not in conditionedmedia setup. Statistical differenceswere compared with their respective stimulated groups withoutnutraceutical treatment.ppp, p # .001 (n = 3 independent biological replicates).Abbreviations:hPSC-ECs, humanpluripotent stemcell-derived endothelial cells; hPSC-HEPs, human pluripotent stem cell-derived hepatocytes; IL, interleukin; NFkB, nuclear factor kB.
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