Development of multiplex PCR systems for expression profiling of human cardiomyocytes induced to proliferate by lentivirus transduction of upcyte genes

Severe heart diseases such as myocarditis and cardiomyopathy are often characterized by progressive damages of contractile heart tissue which ultimately can lead to terminal heart failure. There is a need for relevant in vitro cultures of human cardiomyocytes to study pathogenic processes and to perform pharmacological testing of new heart drugs. By using the upcyte/EPCC (enhanced primary cell culture) approach for direct multiplication of organ-specific cells, we established proliferating human cardiomyocyte cultures derived from atrial appendages. For qualitative cardiac expression profiling we established a comprehensive set of multiplex PCR assays, selected from a panel of 32 genes, to rapidly screen changes at the transcriptional level in human ventricular and atrial cardiomyocytes. Our multiplex PCR approach revealed some donor variability of native atrial heart tissue that need to be confirmed by further studies with more samples. Our initial studies further indicated that characteristic heart muscle cell markers such as MLC-2a, MLC-2v, CHRM2, ADRB1, DES, EDRNB, Cx40 and KCNA5 were down-regulated when isolated cardiomyocytes were taken into primary cell culture. Compared to native heart tissue, proliferating atrial cardiomyocytes lacked expression of those cardiac markers but still expressed MYCD, GATA-4, Cx43, SERCA2, BNP, Tbx5, EDNRA and ACTB. Surprisingly, atrium-derived cardiomyocytes started to express NFAc4 in passage three, and cardiomyocyte marker expressions of Cx43 and BNP were even increased over cultivation time.