Peter van Mourik

8 Chapter 1 GENERAL INTRODUCTION Recent advances in adult stem cell biology have resulted in the development of organoid culture technologies using a variety of tissue sources such as intestine, lung, and kidney 1 . Organoids are three-dimensional, multi-cellular structures that recapitulate tissue features of the parental organ and are usually grown from donor tissue fragments 1 . As organoids are functional expressions of individual genomes, these cultures are particularly useful to understand how genetic factors contribute to individual disease. As such, they are used to study hereditary diseases like cystic fibrosis (CF), and more common diseases such as cancer where genetics can influence disease severity and drug efficacy 2,3 . Intestinal organoids have been on the forefront of these developments as culture methodology was first developed for this tissue source 4 . For CF, human intestinal organoids can be grown from intestinal crypt fragments that are isolated from rectal biopsies. Taking rectal biopsies is typically an innocuous procedure that can be performed in all age groups (including newborns) without a need for anesthesia 5,6 . Intestinal organoids can be generated with high individual success rates, which facilitates access to tissue that can be expanded, stored in liquid nitrogen and can be thawed and continuously cultured for more than 6 months while preserving individual functional phenotype 7 . While many labs worldwide have adopted organoid culturing 8 , few laboratories have expertise in using intestinal organoids to study CF. Here we focus on applications of intestinal organoids for CF and emphasize important future directions that can help to develop effective therapies for all people with CF. HOW DOES IT WORK? Genetic variability of the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) gene can result in misfolding and malfunction of the CFTR protein. This subsequently dysregulates epithelial ion and fluid transport, resulting in CF disease manifestations. CFTR plays a dominant role in rapid fluid secretory responses in the large intestine that are evoked by cyclic AMP (cAMP) inducing stimuli and its activation can lead to secretory diarrhea. Chloride is the main driver of this rapid fluid secretory response 9,10 . CFTR is expressed on the apical membrane that lines the internal lumen of rectal organoids. CFTR activation by cAMP-raising agents such as forskolin leads to chloride transport into the organoid lumen that is accompanied by luminal water secretion through osmosis (Figure 1). Two functional assays have been developed