Modulation of Cardiomyocyte Electrical Properties Using Regulated Bone Morphogenetic Protein-2 Expression

Abstract

Because cardiomyocytes lose their ability to divide after birth, any subsequent cell loss or dysfunction results in pathologic cardiac rhythm initiation or impulse conduction. Strategies to restore and control the electrophysiological activity of the heart may, therefore, greatly affect the regeneration of cardiac tissue functionality. Using lentivirus-derived particles to regulate the bone morphogenetic protein-2 (BMP-2) gene expression in a pristinamycin- or gaseous acetaldehyde-inducible manner, we demonstrated the adjustment of cardiomyocyte electrophysiological characteristics. Complementary metal oxide semiconductor-based high-density microelectrode arrays (HD-MEAs) were used to monitor the electrophysiological activity of neonatal rat cardiomyocytes (NRCs) cultured as monolayers (NRCml) or as microtissues (NRCmt). NRCmt more closely resembled heart tissue physiology than did NRCml and could be conveniently monitored using HD-MEAs because of their ability to detect low-signal events and to sub-select the region of interest, namely, areas where the microtissues were placed. Cardiomyocyte-forming microtissues, transduced using lentiviral vectors encoding BMP-2, were capable of restoring myocardial microtissue electrical activity. We also engineered NRCmt to functionally couple within a cardiomyocyte monolayer, thus showing pacemaker-like activity upon local regulation of transgenic BMP-2 expression. The controlled expression of therapeutic transgenes represents a crucial advance for clinical interventions and gene-function analysis.

Microelectronic system for high-resolution mapping of extracellular electric fields applied to brain slices

“…Recordings from a variety of biological preparations, including dissociated cardiomyocytes and neurons, have been performed (Sanchez-Bustamante et al, 2008), however, the focus is here on acute sagittal cerebellar slices with the aim to demonstrate the performance of the developed microelectrode system. The chip is immersed in the culture solution of the biological preparation, and the cells are placed or grown directly on the chip surface so that a good passivation and corrosion protection is necessary.…”

Adipose Tissue Engineering in Three-Dimensional Levitation Tissue Culture System Based on Magnetic Nanoparticles

“…While simpler approaches to spheroid culture in an agitated medium and/or low-adherence have been reported for many cell types, including ASC, 52,54,55 the rationale behind using magnetic levitation for WAT-derived cells was to enforce retention of cell types other than ASC composing the organ, which tend to be dominated by ASC and progressively lost in conventional culture settings. Furthermore, the magnetic levitation method assures the cell-cell interaction between different cell types at the onset of levitation by magnetically guiding cells together, in contrast to relying on random cell interactions when using an agitated medium and/or low-adherence methods.…”

A synthetic mammalian electro-genetic transcription circuit

“…Lentivectors pCD20 and pCD22, enabling acetaldehyde-responsive expression of human bone morphogenetic protein 2 (BMP-2), have been described previously (32). In brief, pCD20 (5′LTR-ori SV40 -cPPT-RRE-P AIR - bmp-2 -3′LTR ΔU3 ) encodes the bmp-2 gene under control of the acetaldehyde-responsive promoter P AIR and pCD22 (5′LTR-ori SV40 -cPPT-RRE-P hEF1α - alcR -3′LTR ΔU3 ) encodes constitutive expression of the acetaldehyde-dependent transactivator AlcR (26).…”

“…In brief, pCD20 (5′LTR-ori SV40 -cPPT-RRE-P AIR - bmp-2 -3′LTR ΔU3 ) encodes the bmp-2 gene under control of the acetaldehyde-responsive promoter P AIR and pCD22 (5′LTR-ori SV40 -cPPT-RRE-P hEF1α - alcR -3′LTR ΔU3 ) encodes constitutive expression of the acetaldehyde-dependent transactivator AlcR (26). pBP253 (5′LTR-ori SV40 -cPPT-RRE-P hEF1α - bmp-2 -3′LTR ΔU3 ) is the control lentivector for constitutive bmp-2 expression (32). Vesicular stomatitis virus-pseudotyped lentiviral particles were produced and titrated as described before (33).…”

“…The HD-MEAs were manufactured as described previously (32,35). The arrays were coated with 20 µg/ml laminin for 3 h prior to seeding 105 neonatal rat cardiomyocytes (NRCs) in 1 ml medium.…”