189 6 iPSC-derived liver organoids as a tool to study Medium Chain Acyl-CoA Dehydrogenase deficienc Human Precision-Cut Liver Slices (hPCLS) PCLS with approximately 250–300 µM thickness were prepared using a Krumdieck Tissue Slicer (Alabama Research and Development), as described elsewhere. Individual slices were kept in culture for 24h in Williams E medium (WE), containing 25 mM glucose, 0.5% BSA, 1 mM l-carnitine, and gentamycin (Invitrogen). The plates were kept under a continuous supply of 80% O2/5% CO2, shaking at 70 rpm. Human liver tissue was collected from a transplantation donor at the University Medical Center Groningen (UMCG, Netherlands). The liver was used for research purposes after being rejected for transplantation with the approval of the Research Ethics Committee of UMCG. Statistical Analyses All results are expressed as mean ± standard error of the mean (SEM). Analyses were performed using GraphPad Prism Software Version 9.02 (Graphpad Software). Statistical significance was determined as *P value < 0.5, **P value < 0.01 and ***P value < 0.001; no indication means no significant changes (ns). For iPSC work, biological replicates are considered individual donors. For organoid work, due to low number of patients, biological replicates are considered organoids coming from the two same patients or healthy controls that were at least one passage apart. Informed consent The use of historical patient fibroblasts was approved by the Medical Ethical Committee of the University Medical Center Groningen and confirmed according to the Dutch law. FUNDING This work was supported by the European Union’s Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie Actions Grant Agreement PerICo No 812968 and PoLiMeR, No 812616; the Stichting De CockHadders and the Stichting Beatrix Kinderziekenhuis.
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