Comparison of mRNA expression of transcriptional factors and intercalated disk constituent proteins between in vivo and cultured cardiomyocytes.

The weak contractile force exerted by engineered cardiac muscle is a big problem in cardiac muscle tissue engineering, even though the field has made great progress over the past decade. We believe that one major reason for the weak contractile force is that the expression of genes regulating cardiomyocyte differentiation and cardiac tissue syncytium may be different for in vivo and cultured cells. In the present study, we investigated the difference of mRNA expression under in vivo and culture conditions in order to seek a target for further gene transfer treatment in the process of cardiac tissue construction. To this end, mRNA expression of four major transcriptional factors (SRF, p300, Nkx2.5, and myocardin) and two intercalated disk constituent proteins (N-cadherin and connexin43) in rat cardiomyocytes was measured by means of ratiometric reverse-transcription polymerase chain reaction. Cardiomyocytes were harvested from the hearts of 18-day (about 3 days before birth) Wistar-rat embryos (embryonic cells), 12-day neonatal rat hearts (neonatal cells), or 14-day successive dish culture of the embryonic cells harvested from 18-day embryos (cultured cells). The results indicated that, except for SRF, the mRNAs had a lower expression tendency in cultured cells than in embryonic and in neonatal cells; in particular, the mRNA expression of myocardin, N-cadherin, and connexin43 of cultured cardiomyocytes was significantly lower than that of neonatal cells. Therefore, myocardin is a candidate for forced gene up-expression during the construction of engineered cardiac tissue; in addition, a plausible reason for the weak contractile force of engineered cardiac tissue is the weak constitution of intercalated disk, because it was elucidated that mRNA expression of proteins related to intercalated disk were lower in culture.