The emergence of the circadian clock network in hiPSC-derived hepatocytes on chip.

The circadian clock has paramount implications in physiology and pathology. Although the circadian clock has been widely investigated in adults, up to now very little is known about how circadian rhythms emerge during embryonic development. Some studies about the ontology of the circadian system are focused on the development of the central pacemaker, whereas there is still no agreement about the development of the circadian clock in peripheral tissues. Our work represents the first attempt at investigating the onset of peripheral circadian clocks in the liver, which has a central role in controlling several aspects of human physiology. We profile the emergence of the circadian genes during the transition from the initial state of human pluripotency to the final state of hepatic maturation. We demonstrate that circadian rhythmicity is absent in human pluripotent stem cells, and it arises gradually during the process of hepatic commitment. The clock genes expression reaches a peak at the hepatic progenitor stage. At this point o hiPSC-derived f differentiation the gene oscillations start to be observed with a period of 13 h and approaches 24 h in a later stage when the clock primary feedback loop starts working properly. At the end of differentiation, circadian rhythmicity appears, with genes of primary and secondary feedback loops in antiphase (CLOCK, BMAL1 and REV-ERBα) a sign that the system becomes to be functional.