Connexin 43 influences lineage commitment of human neural progenitor cells

Gap junctions (GJs) are intercellular channels connecting the cytoplasm of adjacent cells. This type of connection is an efficient way of cellular communications in many tissues including the central nervous system. Connexins are the proteins that constitute mammalian GJs, and Connexin43 (Cx43) is the most abundant isoform expressed in body cells. Cx43 has been detected within immature neural populations, but only in astrocytes in the adult brain and investigations have shown that Cx43 channel and adhesive properties largely influence neuronal differentiation of mouse neural progenitor (NP) cells. To date the role of Cx43 in neuronal differentiation remains unexplored in human systems, hence our study aimed to investigate the Cx43 participation in human NP differentiation. We largely detected Cx43 protein within the immature neural populations showing that protein expression occurred by fibroblast growth factor (FGF _2) stimulation through the ERKlj2 pathway; FGF _2 withdrawal induced NP differentiation and a progressive loss in Cx43 expression. Cx43's role in neuronal differentiation was explored by cloning lentiviral vectors (LV) coding for Cx43 or anti-Cx43 shRNA constructs and the protein knockdown resulted in an increase in neurons and a decrease in astrocytes, suggesting a role for Cx43 in human stem cell differentiation and neuronal fate. GJs mediate intercellular communications of several mouse and human embryonic stem (ES) cell lines; in our investigation we also showed the presence of functional GJ channels in human ES cells, as well as Cx43 protein expression. Manipulation of ES cells was attempted using LVs and results indicated that the CMV promoter in ES cells is largely inactive. In summary I demonstrated the active role of Cx43 in mechanisms that govern neurogenesis and differentiation of neural progenitor cells, furthermore we highlighted the importance of the internal promoter in LV constructs for genetic manipulation of embryonic ES cells.EThOS - Electronic Theses Online ServiceGBUnited Kingdo