Electric pulse current stimulation increases electrophysiological properties of If current reconstructed in mHCN4-transfected canine mesenchymal stem cells
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- Published online on: February 11, 2016 https://doi.org/10.3892/etm.2016.3072
- Pages: 1323-1329
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Copyright: © Feng et al. This is an open access article distributed under the terms of Creative Commons Attribution License.
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Abstract
The 'funny' current, also known as the If current, play a crucial role in the spontaneous diastolic depolarization of sinoatrial node cells. The If current is primarily induced by the protein encoded by the hyperpolarization-activated cyclic nucleotide-gated channel 4 (HCN4) gene. The functional If channel can be reconstructed in canine mesenchymal stem cells (cMSCs) transfected with mouse HCN4 (mHCN4). Biomimetic studies have shown that electric pulse current stimulation (EPCS) can promote cardiogenesis in cMSCs. However, whether EPCS is able to influence the properties of the If current reconstructed in mHCN4‑transfected cMSCs remains unclear. The present study aimed to investigate the effects of EPCS on the If current reconstructed in mHCN4‑transfected cMSCs. The cMSCs were transfected with the lentiviral vector pLentis‑mHCN4‑GFP. Following transfection, these cells were divided into two groups: mHCN4‑transfected cMSCs (group A), and mHCN4‑transfected cMSCs induced by EPCS (group B). Using a whole cell patch‑clamp technique, the If current was recorded, and group A cMSCs showed significant time and voltage dependencies and sensitivity to extracellular Cs+. The half‑maximal activation (V1/2) value was ‑101.2±4.6 mV and the time constant of activation was 324±41 msec under ‑160 mV. In the group B cells the If current increased obviously and activation curve moved to right. The absolute value of V1/2 increased significantly to ‑92.4±4.8 mV (P<0.05), and the time constant of activation diminished under the same command voltage (251±44 vs. 324±41, P<0.05). In addition, the mRNA and protein expression levels of HCN4, connexin 43 (Cx43) and Cx45 were upregulated in group B compared with group A, as determined by reverse transcription-quantitative polymerase chain reaction and western blot analyses. Transmission electron micrographs also confirmed the increased gap junctions in group B. Collectively, these results indicated that reconstructed If channels may have a positive regulatory role in EPCS induction. The cMSCs transfected with mHCN4 induced by EPCS may be a source of effective biological pacemaker cells.
