Functional analysis of human cardiac troponin by the in vitro motility assay: comparison of adult, foetal and failing hearts

Objective: Human cardiac development and heart failure are associated with altered troponin isoform expression and phosphorylation. As the functional effects of these changes in troponin are unknown, we isolated troponin from human foetal, normal adult and failing adult hearts and investigated their regulatory function. Methods: Human cardiac troponin was assayed for regul atory function by in vitro motility assay and for protein content by SDS PA GE and immunoblotting. Results: Human cardiac troponin regulated movement o f actin-tropomyosin filaments over a bed of immobilised heavy meromyosin. A t pCa 9, troponin from foetal and adult hearts reduced the fraction of fila ments moving from 90% to less than 15% with a modest (25-30%) decrease in v elocity. At pCa 5, troponin from normal adult hearts increased filament vel ocity by up to 47+/-3% with no change in the fraction of filaments moving. Foetal troponin increased velocity by only 4+/-6% and the effect of troponi n from failing hearts was between these values at 31+/-5%. Foetal hearts sh owed different troponin I and T isoform expression compared with adult hear ts. No differences in troponin isoform expression were demonstrated between normal and failing adult hearts. Conclusions: Functioning troponin and tro pomyosin may be isolated from human heart and their properties investigated by in vitro motility assay. Both functional and isoform expression differe nces exist between foetal and adult cardiac troponin. The regulatory functi on of troponin from adults with end stage heart failure is different from n ormal adult troponin. These data suggest a role for altered troponin functi on in human cardiac development and heart failure. (C) 1999 Elsevier Scienc e B.V. All rights reserved.