MYH11 rare variant augments aortic growth and induces cardiac hypertrophy and heart failure with pressure overload

Abstract

Smooth muscle cell-specific myosin heavy chain, encoded by MYH11, is selectively expressed in smooth muscle cells (SMCs). Pathogenic variants in MYH11 predispose to a number of disorders, including heritable thoracic aortic disease associated with patent ductus arteriosus, visceral myopathy, and megacystis-microcolon-intestinal hypoperistalsis syndrome. Rare variants of uncertain significance occur throughout the gene, including MYH11 p.Glu1892Asp, and we sought to determine if this variant causes thoracic aortic disease in mice. Genomic editing was used to generate Myh11E1892D/E1892D mice. Wild-type (WT) and mutant mice underwent cardiovascular phenotyping with and without transverse aortic constriction (TAC). Myh11E1892D/E1892D and WT mice displayed similar growth, blood pressure, root and ascending aortic diameters, and cardiac function up to 13 months of age, along with similar contraction and relaxation on myographic testing. The hypertension induced by TAC was similarly in Myh11E1892D/E1892D and WT mice, but mutant mice showed augmented ascending aortic enlargement and increased elastic fiber fragmentation on histology. Unexpectedly, male Myh11E1892D/E1892D mice undergoing TAC had decreased ejection fraction, stroke volume, fractional shortening, and cardiac output compared to similarly treated male WT mice. Importantly, left ventricular mass increased significantly due to primarily posterior wall thickening, and cardiac histology confirmed cardiomyocyte hypertrophy and increased collagen deposition in the myocardium and surrounding arteries. These results further highlight the phenotypic heterogeneity associated with MYH11 rare variants. Given that MYH11 is selectively expressed in SMCs, these results implicate a role of SMCs in the arteries of the heart contributing to cardiac hypertrophy and failure with pressure overload.

Fig 1. Cardiovascular phenotyping in male mice.

(A) Generation of the Myh11^{E1892D/E1892D} mouse model using CRISPR-Cas9 genome editing. Sequencing of DNA sample from tail tissue and cDNA sample from both heart and aorta confirms single nucleotide variant in mouse Myh11 gene. (B) Tail-cuff blood pressure measurement. (C) and (D) Echocardiographic measurements of aortic root, ascending aorta, and left ventricular contractile function. N = 10 males in each group. SBP, systolic blood pressure; DBP, diastolic blood pressure; WT, wild-type; HET, heterozygous; HOMO, homozygous.

Fig 2. Assessment of myograph and smooth muscle cell contractile protein expression in ascending aortic tissues.

(A) Myographic assay of mouse ascending aortic rings at 10 months of age. N = 3 males in each group. (B) Immunoblot assay of protein lysates of the ascending aortas from wild-type (WT), heterozygous (HET), and homozygous (HOMO) mice at the age of 6 months. N = 7 (3M + 4F), 8 (5M + 3F), 5 (2M + 3F) in the WT, HET, and HOMO groups, respectively. KCl, potassium chloride; PE, phenylephrine; Ach, acetylcholine; SNP, sodium nitroprusside.

Fig 3. Assessment of ascending aortic remodeling 2 weeks after transverse aortic constriction (TAC) in male mice.

(A) Representative images of proximal aortic ultrasound measurement, H&E, VVG, and Sirius Red staining on ascending aortic tissue sections. (B) TAC induces similar levels of systolic (SBP) and diastolic (DBP) blood pressure in wild-type (WT) and Myh11^{E1892D/E1892D} mice, along with significant ascending aortic enlargement in Myh11^{E1892D/E1892D} mice. (C) Histology analysis shows significantly increased medial thickening and elastic breaks, and decreased medial cell density in the mutant aortas compare with WT TAC aortas, with no difference of adventitial collagen accumulation. N = 7 and 6 in the WT and Myh11^{E1892D/E1892D} groups, respectively. ns, non-significant; * P < 0.05, *** P < 0.001, by unpaired Mann-Whitney analysis.

Fig 4. Assessment of left ventricular (LV) remodeling 2 weeks after transverse aortic constriction (TAC) in male mice.

(A) Representative images of LV contraction measurement from short-axis parasternal view using M-mode. (B) Four functional parameters show decreased LV contractility in Myh11^{E1892D/E1892D} mice 2 weeks after TAC. (C) Structural parameters of LV show increased end diastole (d) and systole (s) thickness of posterior wall (PW), along with increased LV diameters and volumes in Myh11^{E1892D/E1892D} mice after TAC. (D) Wheat Germ Agglutinin (WGA) staining of LVPW shows significant increase of cardiomyocyte cross section area in male Myh11^{E1892D/E1892D} heart after TAC. (E) Representative images of Sirius Red staining of LVPW. Quantification of collagen deposition area shows increased peri-arterial (left circumflex artery, LCX) area and LVPW collagen density in Myh11^{E1892D/E1892D} heart after TAC. N = 7 and 6 in the WT and Myh11^{E1892D/E1892D} groups, respectively. ns, non-significant; * P < 0.05, ** P < 0.01, *** P < 0.001, by unpaired Mann-Whitney analysis.