Role of the Kir2.1 potassium channel in bone morphogenesis

Andersen's syndrome (AS) is a complex disorder characterized by a triad of symptoms: periodic paralysis, cardiac arrhythmias, and developmental disorders (Andersen 1971). It is a rare pathology, dominant hereditary, associated with genetic mutations of the KCNJ2 gene (Plaster 2001) that encodes the inward rectifier potassium channel Kir2.1. Kir2.1 is known to play a major role in stabilizing resting cell potential and in the late phase of repolarization of cardiac action potential. Functional characterizations of these mutations were performed in vitro and ex vivo and showed the dominant negative effect of mutations on this channel. Progress has been made in understanding the roles of this channel in muscle and heart tissue (Zaritsky 2001). However, few studies have addressed the role of the Kir2.1 channel in developmental manifestations, including its role in non-excitable cells. In addition, the invalidation (KO) of the KCNJ2 gene in mice is lethal within hours of birth due to cleft palate (Zaritsky 2000, Dahal 2012). This early post-natal mortality makes in vivo studies difficult, particularly those to assess its role in adult tissues. An alternative is the use of an in vitro model involving human induced pluripotent cells (iPS) (Takahashi 2007) that are capable of being differentiated in all cell types. Dr. Bendahhou's team has previously shown that the Kir2.1 functional channel is necessary for bone formation in a model of myoblast differentiation into osteoblasts (Sacco 2015). In addition, they generated iPS from muscle biopsies of healthy individuals and AS patients (Pini 2016). We have also shown that the absence of Kir2.1 in osteoblastic and chondrocytic differentiation impacts not only matrix production but also the expression of the main genes of these two lineages (Pini 2018). In osteoblastogenesis, activity of the Bone Morphogenetic Protein (BMP) signaling pathway is decreased in AS cells (Pini 2018). My project focuses on determining the role of the Kir2.1 channel in the BMP signalling pathway in in vitro osteoblasts. To do this, I have used two cell types. Taking advantage of their differentiation capacity, I used the iPS (healthy and AS) already generated by my team, but also the iPS I have generated, to study the role of Kir2.1 in bone tissue. In parallel, I used a cell line of immortalized fetal human osteoblasts (hFOB) developed by Harris in 1995. A combination of transcriptomic and proteomic analyses, as well as immunohistochemistry, were used to investigate the role of the Kir2.1 potassium channel in key stages of bone morphogenesis.