Changes in Gene Expression During the Formation of Bioengineered Heart Muscle

Abstract

A three-dimensional bioengineered heart muscle (BEHM) construct model had been previously developed, exhibiting contractile forces up to 800 microN. The interest of this study was to determine gene expression levels of biologic markers involved in calcium-handling between BEHM, cell monolayer, and neonatal heart. Cardiac cells were isolated from one litter of F344 rats and organized into groups (n = 5): 4-, 7-, 10-day BEHM and cell monolayer; BEHM was evaluated for cell viability and contractility. Groups were then analyzed for mRNA expression of calcium-handling proteins: myosin heavy chain (MHC) alpha and beta, Sarcoplasmic reticulum Ca++ ATPase (SERCA) 2, phospholamban (PBL), and ryanodine receptor. BEHM exhibited electrically stimulated active force (208 +/- 12 microN day 4, 361 +/- 22 microN day 7, and 344 +/- 29 microN day 10) and no decrease in cell number. Real-time polymerase chain reaction (PCR) showed an increase in gene expression of all calcium-handling proteins in BEHM at 7 and 10 days compared with monolayers, for example, comparing BEHM to monolayer (7 and 10 days, respectively), MHC-alpha: 2600-fold increase and a 100-fold increase; MHC-beta: 70-fold increase at 10 days; ryanodine receptor: 74-fold increase at 10 days; SERCA: 19-fold increase and sixfold increase; PBL: 158-fold increase and 24-fold increase. It was concluded that a three-dimensional environment is a better culturing condition of cardiac cells than a monolayer. Also, BEHM constructs demonstrated a high similarity to a native myocardium, and is, thus, a good starting foundation for engineered heart muscle.

Knockdown of embryonic myosin heavy chain reveals an essential role in the morphology and function of the developing heart

“…In this study, we have shown the differing effects following the manipulation of e-, α- or βMYH in both the atria and ventricle at early stages of heart development, which might result in alterations between the generation of the electrical activity of the cells and their Ca 2+ transients. These Ca 2+ transient changes are fundamental to proper activation of the machinery of contraction, in which MYH plays a key role (Chandra et al, 2007; Dillmann, 2009; Khait and Birla, 2009), but also to the regulation of gene expression (Webb and Miller, 2003). Evidence is presented suggesting that eMYH has a major impact on the AP properties of the atrial and ventricular cells, with AP characteristics that are normally found during very early heart development (Arguello et al, 1986).…”

Heavy and light roles: myosin in the morphogenesis of the heart

“…Further, as seen with αMHC, the atrial and ventricular cardiomyocytes had normal action potentials, although irregular Ca 2+ transients were seen in atrial and ventricular cells [42]. Defects in calcium signaling may lead to defects in contraction of the heart, with MHC known to be important for induction of contraction [67–69]. Two fish models have been used to analyze the presumptive functional orthologue to MYH7 .…”

Artificial Organs 2009: A Year in Review

“…Luda Khait and Ravi K. Birla (110) of the University of Michigan, Ann Arbor, MI, USA evaluated changes in gene expression during the formation of bioengineered heart muscle (BEHM). Cardiac cells were isolated from one litter of F344 rats and organized into the groups 4‐, 7‐, 10‐day BEHM and cell monolayer.…”