Abstract Body (Do not enter title and authors here): Introduction
Cardiac troponin I (cTnI, TNNI3 gene) is a highly conserved subunit of the sarcomere that inhibits contraction by preventing actin-myosin interaction. Pathogenic TNNI3 variants can cause both hypertrophic and restrictive cardiomyopathies but lack targeted therapies. We identified a family with restrictive cardiomyopathy carrying a cTnI variant at amino acid 157 (A157V). Using CRISPR-Cas9, we generated a knock-in mouse model reflecting this mutation (A158V in mouse). Our previous studies showed that homozygous A158V mice had impaired cardiac relaxation on invasive hemodynamics but normal lifespan, no echocardiographic changes, and no fibrosis.
Hypothesis
Myofibril mechanics and transcriptomic studies will better discriminate TNNI3 A158V mice from WT than previous phenotyping studies.
Methods
Heart tissue was obtained from wild-type (WT) and homozygous A158V mice at 9-10 months. Myofibrils were isolated from heart tissues and mechanical measurements were completed using the fast solution switching method. RNA was extracted for bulk RNA sequencing.
Results
Isolated myofibril studies showed significant prolongation of the linear relaxation phase in A158V mice versus controls (p=0.02) but no significant differences in active tension. Unsupervised clustering of bulk RNA samples separated A158V samples from controls, with 44% transcriptional variation derived from genotype. Cardiac changes in the A158V model were confirmed by significant depletion of pathways involved in structural components of the contractile apparatus, including contractile fiber, sarcomere, and I-Band. On an individual gene level, A158V mutated mice displayed higher expression of cardiac remodeling genes such as Timp1, Postn, and Tnc.
Conclusion
The TNNI3 A158V variant consistently produces impaired relaxation with myofibril studies showing prolonged linear relaxation phase. Transcriptomic studies highlight distinct clustering of TNNI3 A158V mice from WT, with prominent depletion in contractile apparatus pathways. Our findings suggest traditional methods to characterize mouse models may fail to capture disease phenotype in cardiomyopathies; however, myofibril studies and transcriptomic profiling may reveal earlier phenotypic changes.