ABSTRACT:As the fastest self-regenerating organ, the intestinal epithelium serves primarily to absorb nutrients and minerals, while its other distinct functional characteristics include the capacity to proliferate and replicate many lineage precursors. Currently, in vitro studies using small intestine cells are hindered by the lack of a standard cell culture model. Here, we characterize primary human prenatal small intestine epithelial cells (HPIEC). The data demonstrate that HPIEC express markers indicative of stem cells (NOTCH1, SOX9, OLFM4, LGR5), epithelial cells (CK18, CDH1), and immunological and entero-endocrinal cells (DEFA5, GPR-120, GLP-1). More importantly, HPIEC are shown to secrete GLP-1 and lysozyme, and express sucrase-isomaltase, maltase-glucoamylase, and drug transport function that is cyclosporin A repressible, which demonstrates functionality. These data indicate that HPIEC are a heterogeneous cell population that contains stem cells and epithelial cells that may be valuable for various functional, developmental, and pharmacologic studies.