Shox2 influences mesenchymal stem cell fate in a co-culture model in vitro

Sinoatrial node (SAN) dysfunction is a common cardiovascular problem, and the development of a cell sourced biological pacemaker has been the focus of cardiac electro- physiology research. The aim of biological pacemaker therapy is to produce SAN-like cells, which exhibit spontaneous activity characteristic of the SAN. Short stature homeobox 2 (Shox2) is an early cardiac transcription factor and is crucial in the formation and differentiation of the sinoatrial node (SAN). The present study aimed to improve pacemaker function by overexpression of Shox2 in canine mesenchymal stem cells (cMSCs) to induce a phenotype similar to native pacemaker cells. To achieve this objective, the cMSCs were transfected with lentiviral pLentis-mShox2-red fluorescent protein, and then co-cultured with rat neonatal cardiomyocytes (RNCMs) in vitro for 5-7 days. The feasibility of regulating the differentiation of cMSCs into pacemaker-like cells by Shox2 overexpression was investigated. Reverse transcription- quantitative polymerase chain reaction and western blotting showed that Shox2-transfected cMSCs expressed high levels of T box 3, hyperpolarization-activated cyclic nucleotide-gated cation channel and Connexin 45 genes, which participate in SAN development, and low levels of working myocardium genes, Nkx2.5 and Connexin 43. In addition, Shox2-transfected cMSCs were able to pace RNCMs with a rate faster than the control cells. In conclusion, these data indicate that overexpres- sion of Shox2 in cMSCs can greatly enhance the pacemaker phenotype in a co-culture model in vitro.