203 7 General Discussion However, this study focused on removal of not only amino acids but also proteins and carbohydrates. Moreover, they studied intestinal permeability of the organoids but did not characterize metabolic (mitochondrial or peroxisomal) function. Organoids are three-dimensional cell structures that recapitulate multiple functions of the organ of origin whilst continuing to proliferate in culture3. Since their discovery4, tens of different organoid models have been described, including the pancreas5, stomach6, lungs7, and brain8. Organoids have been used for a wide range of purposes such as developmental studies3,9, modeling genetic diseases10, and drug screening11. Here, I demonstrated that hepatic and intestinal organoids make suitable models to study malnutrition in vitro. In order to mimic low protein diets (LPD), used in in vivo models to induce malnutrition12–14, organoids were grown in culture medium without any amino acids. However, CG-MS analysis of the organoid supernatant revealed that all amino acids were present, probably coming from the hydrogel or from the degradation of the growth factors in the medium, yet in much lower levels than in the control cultures. These in vitro systems present a wide range of advantages and new possibilities when compared to in vivo studies15. One main point of interest is the possibility to study one specific organ at a time16. This allowed us to study the effects of amino-acid restriction on the liver and the intestine in an isolated system without a systemic response. A key example highlighting the importance of organ-specific studies is discussed in chapter 4. Here I reported that, opposite to what was expected from in vivo studies12–14, the autophagic flux was clearly upregulated in hepatic and intestinal organoids upon amino-acid restriction. While these results might seem contradictory, it is not uncommon to see different responses between in vivo and in vitro models. Moreover, this organ-specific approach paves the way for characterizing and studying different organ-specific-pathophysiological processes which can provide great insight into the disease of interest. Another advantage of these translational in vitro models is the possibility to pre-screen large numbers of drugs prior to in vivo testing17,18. In chapter 2, to illustrate the potential of the models, I tested the effects of fenofibrate and rapamycin in the liver and the intestine, respectively, as a proof of principle. These compounds showed a similar response as to that observed in vivo14. Moreover, in chapter 4 I focused on a more extensive drug screening to prevent peroxisomal loss in the hepatic malnutrition model. I this chapter I evaluated the potential of different PPAR-α activators including synthetic and naturally occurring compounds. These studies led us to the findings that docosahexaenoic acid (DHA), an omega-3 fatty acid, could prevent the loss
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