Impaired in vitro growth response of plasma-treated cardiomyocytes predicts poor outcome in patients with transthyretin amyloidosis

Abstract

Objectives Direct toxic effects of transthyretin amyloid in patient plasma upon cardiomyocytes are discussed. However, no data regarding the relevance of this putative effect for clinical outcome are available. In this monocentric prospective study, we analyzed cellular hypertrophy after phenylephrine stimulation in vitro in the presence of patient plasma and correlated the cellular growth response with phenotype and prognosis. Methods and results Progress in automated microscopy and image analysis allows high-throughput analysis of cell morphology. Using the InCell microscopy system, changes in cardiomyocyte’s size after treatment with patient plasma from 89 patients suffering from transthyretin amyloidosis and 16 controls were quantified. For this purpose, we propose a novel metric that we named Hypertrophic Index, defined as difference in cell size after phenylephrine stimulation normalized to the unstimulated cell size. Its prognostic value was assessed for multiple endpoints (HTX: death/heart transplantation; DMP: cardiac decompensation; MACE: combined) using Cox proportional hazard models. Cells treated with plasma from healthy controls and hereditary transthyretin amyloidosis with polyneuropathy showed an increase in Hypertrophic Index after phenylephrine stimulation, whereas stimulation after treatment with hereditary cardiac amyloidosis or wild-type transthyretin patient plasma showed a significantly attenuated response. Hypertrophic Index was associated in univariate analyses with HTX (hazard ratio (HR) high vs low: 0.12 [0.02–0.58], p = 0.004), DMP: (HR 0.26 [0.11–0.62], p = 0.003) and MACE (HR 0.24 [0.11–0.55], p < 0.001). Its prognostic value was independent of established risk factors, cardiac TroponinT or N-terminal prohormone brain natriuretic peptide (NTproBNP). Conclusions Attenuated cardiomyocyte growth response after stimulation with patient plasma in vitro is an independent risk factor for adverse cardiac events in ATTR patients

Molecular Mechanisms of Cardiac Amyloidosis

“…Furthermore, Hein et al demonstrated that the plasma of healthy individuals or patients with hereditary transthyretin amyloidosis with polyneuropathy promotes hypertrophy of neonatal rat cardiomyocytes induced by phenylephrine, while the plasma of patients with hereditary transthyretin cardiac amyloidosis or wild-type ATTR cardiac amyloidosis attenuated the hypertrophic response in vitro. They also reported that an attenuated cardiomyocyte hypertrophic response after stimulation with patients’ plasma in vitro is an independent risk factor for adverse cardiac events (death, transplantation and/or cardiac decompensation) in ATTR patients [ 69 ].…”

Transthyretin deposition alters cardiomyocyte sarcomeric architecture, calcium transients, and contractile force

“…In vitro, it has been demonstrated that isolated cardiomyocytes, when cultured with various TTR species in the culture media, exhibit cytotoxicity with increased ROS production and altered calcium transient and action potential repolarization profiles (Sartiani et al, 2016). Additionally, human‐induced pluripotent stem cell‐derived cardiomyocytes (iPSC‐CM) cultured with plasma from wtATTR exhibit impaired growth responses compared to iPSC‐CM cultured with plasma from healthy patients (Hein, Furkel, et al, 2021). Here, the focus is to understand how underlying TTR fibrils affect cardiomyocytes cultured on different engineered substrata in order to best recapitulate cardiomyocyte biology in vitro.…”

C-MORE: A high-content single-cell morphology recognition methodology for liquid biopsies toward personalized cardiovascular medicine

“…7,8 Morphological readout in cardiovascular research is still limited to cell size and a few selected features (e.g., perimeter, elongation, and form factor). 9,10 Alternative methods focus on sarcomere organization, 3D volume/ shape, proliferation, or reporter readouts. [11][12][13][14] However, these established methods address only specific parts of morphology, and integrated approaches are not established for use in cardiovascular research.…”