Heme oxygenase-1 is expressed in embryonic stem cells and increases during differentiation

TORO A; COSENTINO S; GUBERMAN A; PETRONE MV; VAZQUEZ ECHEGARAY C; VAZQUEZ ELBA; SOLARI C; FRANCIA M

Ciudad de Buenos

Congreso; Reunión Conjunta de Biociencias 2017; 2017

Embryonic stem cells (ESCs) are pluripotent, which means that they have the ability to differentiate into cells of all three germ layers, and they possess several mechanisms that ensure the genome stability avoiding the propagation of genetic damage. Heme oxygenase (HO) is the limiting enzyme in the oxidative catabolism of the heme group, and particularly, the inducible isoform HO-1 is known by its antioxidant and antiapoptotic activities. Some reports have studied HO-1 action in multipotent stem cells. However, little is known about HO-1 role in pluripotent stem cells. Based on these evidences, we aimed to study if HO-1 is involved either in ESCs survival or in pluripotency maintenance. In order to achieve this, we first differentiated W4 ESCs with a non-directed differentiation protocol, culturing them in absence of the cytokine LIF for 4 days. RNA and proteins were extracted from undifferentiated cells, as a control, and from distinct time points along the differentiation protocol (days 1, 2, 3 and 4). HO-1 expression profile was analyzed by RT-qPCR and Western blot (WB). We found that HO-1 mRNA and protein levels increased during differentiation. Furthermore, we analyzed HO-1 protein by immunofluorescence in order to study HO-1 subcellular localization throughout the differentiation process. We found that this protein was ubiquitously expressed, both in cytoplasm and in the nucleus, in ESCs in undifferentiated state, and that its levels were upregulated during differentiation, along with the decreased expression of Oct4 and Nanog, transcriptional factors that are crucial to pluripotency. RT-qPCR and WB results were analyzed using randomized block design ANOVA and Tukey test. These findings show that HO-1 expression is modulated along the differentiation suggesting that this enzyme could be relevant to leave behind the undifferentiated state or to the differentiation process and could be associated with Oct4 and Nanog regulation.