Remodelling of adult cardiac muscle cells in culture: dynamic process of disorganization and reorganization of myofibrils

Abstract

The myofibrils of adult rat cardiac muscle cells in long-term culture initially break down and later reassemble into mature myofibrils. The objective of this study is to examine the disorganization process of myofibrils and to determine how disorganized myofibrillar proteins, myosin, titin, actin, and alpha-actinin are reorganized into mature myofibrils in adult cells. After dismantlement of myofibrils during initial culture period (24-72 h), myofibrillar proteins became disorganized into amorphous form. These proteins later were observed in vesicular, amorphous, and nonstriated fibrillar forms. Some vesicular structures, containing mainly myosin, titin, alpha-actin, and alpha-actinin were observed on the outer surfaces of the cell and outside the cell body. Such vesicles containing F-actin were rare. Punctate structures of alpha-actinin emerged from the pre-existing amorphous alpha-actinin along with the appearance of mostly titin periodicities. The periodicities of alpha-actinin later became prevalent, followed by the appearance of periodicities of actin. alpha-actinin provided an initiation point on which titin and actin became associated, forming titin-associated I-Z-I structures. Titin traversed the I-bands on either side of the Z-line. The phalloidin-stained I-Z-I structures bound to antibodies to muscle specific sarcomeric proteins (titin, alpha-actin, alpha-actinin). The differentiation of myosin periodicities lagged behind those of titin, alpha-actinin, and actin although presarcomeric structures of immunolabelled titin and myosin were very closely linked in their distributions in the formative myofibrils. Variations in the temporal sequence of emergence of periodicities of alpha-actinin and myosin were observed among certain myocytes. Also observed was the variation of the temporal sequence of emergence of titin and actin periodicities among different myocytes and within a single myocyte. Even in the late stage of culture (30 days), when the cell body was packed with myofibrils, the myocytes contained remnants of amorphous myofibrillar proteins.