Molecular autopsy and family screening in a young case of sudden cardiac death reveals an unusually severe case of FHL1 related hypertrophic cardiomyopathy

Abstract

Background: Hypertrophic cardiomyopathy (HCM) is a genetic cardiomyopathy with a prevalence of about 1:200. It is characterized by left ventricular hypertrophy, diastolic dysfunction and interstitial fibrosis; HCM might lead to sudden cardiac death (SCD) especially in the young. Due to low autopsy frequencies of sudden unexplained deaths (SUD) the true prevalence of SCD and especially of HCM among SUD remains unclear. Even in cases of proven SCD genetic testing is not a routine procedure precluding appropriate risk stratification and counseling of relatives.

Methods: Here we report a case of SCD in a 19-year-old investigated by combined forensic and molecular autopsy.

Results: During autopsy of the index-patient HCM was detected. As no other possible cause of death could be uncovered by forensic autopsy the event was classified as SCD. Molecular autopsy identified two (probably) pathogenic genetic variants in FHL1 and MYBPC3. The MYBPC3 variant had an incomplete penetrance. The FHL1 variant was a de novo mutation. We detected reduced FHL1 mRNA levels and no FHL1 protein in muscle samples suggesting nonsense-mediated mRNA decay and/or degradation of the truncated protein in the SCD victim revealing a plausible disease mechanism.

Conclusion: The identification of the genetic cause of the SCD contributed to the rational counseling of the relatives and risk assessment within the family. Furthermore our study revealed evidences for the pathomechanism of FHL1 mutations.

Keywords: cardiomyopathy; hypertrophic cardiomyopathy; molecular autopsy; nonsense-mediated decay; sudden cardiac death.

Figure 1

(a) Pedigree of the index patient, who died at the age of 19 years by sudden cardiac death. Relatives of the patient were examined by a cardiologist. Patient III‐9 is the only patient with a documented cardiac disease (filled symbol). (b) Explanted heart from patient III‐9 revealing a hypertrophic cardiomyopathy phenotype. The left ventricle was opened during autopsy (for data on the explanted heart s. text)

Figure 2

Histology from the right (RV) and left ventricle (LV) revealed hypertrophic myocytes with focal myofiber disarray and a severe diffuse interstitial fibrosis (Trichrome staining) especially in the RV. There was no significant inflammation in immunohistochemical stainings for T cells and macrophages and no evidence for an infection with cardiotropic viruses as determined using RT‐PCR (data not shown). Bars represent 100 µm

Figure 3

Quantitative RT‐PCR of FHL1 mRNA expression in the index patient (III‐9) and control tissues. Samples of the skeletal muscle of the index patient (SKM_I; 0.45 ± 0.1) and from a control individual (SKM_c; N = 1; 2.51 ± 0.63) were measured as duplicates and given as means ± standard error of the mean (SEM). Left ventricular myocardial samples of controls without FHL1 mutations (LV_c; N = 5; box and whiskers plot: mean and median given as “+” or line, respectively; whiskers extend from 1–99 percentile) and the index patient (LV_I; 0.07 ± 0.01). RQ, relative quantity

Figure 4

(A) Immunoblotting using an anti‐FHL1 antibody for labeling of muscle tissue extracts. The major isoforms are marked by black arrows and the apparent molecular masses are given. In preparations of the skeletal muscle (SKM) or the left ventricular myocardium (LV) of the index patient (I) FHL1 was not detectable. The molecular mass of the putative truncated peptide, as predicted from sequencing results (9.7 kDa), is marked by a gray arrow. Protein extracts of controls (C) for SKM or LV were used as a reference. (B) Immunohistochemistry of FHL1 in SKM (b) or LV (a, c, d), respectively, using diaminobenzidine as a substrate for HRP staining. Of note, FHL1‐staining was not detectable in the index patient (III‐9; b + d) but in the control tissue (a + c). Bar = 20 µm

Figure 5

Echocardiography of family members (II‐5, II‐6, III‐6 and III‐7) reveals no further cases of hypertrophic cardiomyopathy. (a) Parasternal long axis and (b) four‐chamber view. LA = left atrium, LV = left ventricle, RA = right atrium, RV = right ventricle, Ao = aorta. For echocardiographic details s. Table 2