Hypophosphorylation of the Stiff N2B Titin Isoform Raises Cardiomyocyte Resting Tension in Failing Human Myocardium

High diastolic stiffness of failing myocardium results from interstitial fibrosis and elevated resting tension (F-passive) of cardiomyocytes. A shift in titin isoform expression from N2BA to N2B isoform, lower overall phosphorylation of titin, and a shift in titin phosphorylation from N2B to N2BA isoform can raise F-passive of cardiomyocytes. In left ventricular biopsies of heart failure (HF) patients, aortic stenosis (AS) patients, and controls (CON), we therefore related F-passive of isolated cardiomyocytes to expression of titin isoforms and to phosphorylation of titin and titin isoforms. Biopsies were procured by transvascular technique (44 HF, 3 CON), perioperatively (25 AS, 4 CON), or from explanted hearts (4 HF, 8 CON). None had coronary artery disease. Isolated, permeabilized cardiomyocytes were stretched to 2.2-mu m sarcomere length to measure Fpassive. Expression and phosphorylation of titin isoforms were analyzed using gel electrophoresis with ProQ Diamond and SYPRO Ruby stains and reported as ratio of titin (N2BA/N2B) or of phosphorylated titin (P-N2BA/P-N2B) isoforms. Fpassive was higher in HF (6.1 +/- 0.4 kN/m(2)) than in CON (2.3 +/- 0.3 kN/m2; P < 0.01) or in AS (2.2 +/- 0.2 kN/m2; P < 0.001). Titin isoform expression differed between HF (N2BA/N2B = 0.73 +/- 0.06) and CON (N2BA/N2B = 0.39 +/- 0.05; P < 0.001) and was comparable in HF and AS (N2BA/N2B = 0.59 +/- 0.06). Overall titin phosphorylation was also comparable in HF and AS, but relative phosphorylation of the stiff N2B titin isoform was significantly lower in HF (P-N2BA/P-N2B = 0.77 +/- 0.05) than in AS (P-N2BA/P-N2B = 0.54 +/- 0.05; P < 0.01). Relative hypophosphorylation of the stiff N2B titin isoform is a novel mechanism responsible for raised Fpassive of human HF cardiomyocytes. (Circ Res. 2009; 104: 780-786.)