Human Fetal Pancreas Mesenchyme for Maturation of Human Embryonic Stem Cell Derived Pancreatic Orga-noids

Objective: During the earliest stage of pancreas organogenesis the pancreatic epithelium evaginates into the surrounding mes-enchyme which secrete different growth factors for pancreatic progenitor’s (PP) expansion and subsequent differentiation. Mesenchymal cells (MCs) with diverse origins differ in gene and protein expression as well as signaling pathway factors. They also show dissimilar effects in maintenance and differen-tiation capacity of their organ-matched cells. Co-culture with native pancreatic mesenchyme permits proliferation, self-re-newal or differentiation of their epithelial progenitors, demon-strating the importance of cell-cell communications and their interactions during pancreatic organogenesis. Therefore, in this study the impact of human fetal pancreatic mesenchymal cells (hFP-MCs) on further differentiation of human embryonic stem cell derived pancreatic progenitors (hESC-PPs) was examined through a three-dimensional co-culture system.Materials and Methods: The hESC-PPs differentiation was accomplished from Royan hESC lines. HFP-MCs were iso-lated from ۱۲-۱۶ week human embryos, cultured and charac-terized for VIMENTIN and DESMIN mesenchymal markers. Afterwards the cell mixture suspension containing hESC-PPs and hFP-MCs at ۱:۱ cell ratio were co-cultivated in pancreatic endoderm media in Matrigel for ۱۴ days. The hESC-PPs exist-ence and their ability to differentiate to the endocrine lineage were examined by evaluation of specific markers by immune-staining and quantitative Real-Time PCR analysis.Results: hFP-MCs showed morphological features and also VIMENTIN expression similar to bone marrow-derived mes-enchymal stem cells (BM-MSCs). However, these cells con-troversially showed higher expression of DESMIN marker. After co-culture of hESC-PPs with hFP-MCs, generated PP organoids indicated higher expression of NGN۳ and INSULIN displaying the acceleration of pancreatic mesenchyme towards beta cell maturation.Conclusion: Using native mesenchyme in co-culture provides an opportunity for studies on endocrine-niche interactions and developmental processes by mimicking the pancreatic tissue.