3-DIMENSIONAL RECONSTITUTION OF EMBRYONIC CARDIOMYOCYTES IN A COLLAGEN MATRIX - A NEW HEART-MUSCLE MODEL SYSTEM

A method has been developed for culturing cardiac myocytes in a collag en matrix to produce a coherently contracting 3-dimensional model hear t tissue that allows direct measurement of isometric contractile force . Embryonic chick cardiomyocytes were mixed with collagen solution and allowed to gel between two Velcro-coated glass tubes. During culture, the cardiomyocytes formed spontaneously beating cardiac myocyte-popul ated matrices (CMPMs) anchored at opposite ends to the Velcro-covered tubes through which they could be attached to a force measuring system . Immunohistochemistry and electron microscopy revealed a highly organ ized tissue-like structure of alpha-actin and alpha-tropomyosin-positi ve cardiac myocytes exhibiting typical cross-striation, sarcomeric myo filaments, intercalated discs, desmosomes, and tight junctions. Force measurements off paced or unpaced CMPMs were performed in organ baths after 6-11 days of cultivation and were stable for up to 24 h. Force i ncreased with frequency between 0.8 and 2.0 Hz (positive ''staircase'' ), increasing rest length (Starling mechanism), and increasing extrace llular calcium. The utility of this system as a test bed for genetic m anipulation was demonstrated by infecting the CMPMs with a recombinant beta-galactosidase-carrying adenovirus. Transduction efficiency incre ased from about 5% (MOI 0.1) to about 50% (MOI 100). CMPMs display mor e physiological characteristics of intact heart tissue than monolayer cultures. This approach, simpler and faster than generation of transge nic animals, should allow functional consequences of genetic or pharma cological manipulation of cardiomyocytes in vitro to be studied under highly controlled conditions.