TY - JOUR
T1 - Knockdown of Ift88 in fibroblasts causes extracellular matrix remodeling and decreases conduction velocity in cardiomyocyte monolayers
AU - Ernault, Auriane C.C.
AU - Kawasaki, Makiri
AU - Fabrizi, Benedetta
AU - Montañés-Agudo, Pablo
AU - Amersfoorth, Shirley C. M.
AU - Al-Shama, Rushd F. M.
AU - Coronel, Ruben
AU - de Groot, Joris R.
N1 - Funding Information: This study was supported by Netherlands Organization for Health Research and Development (ZonMW/NWO, 106.146.310) awarded to JG. Funding Information: This research was supported by grants from the National Heart Lung Blood Institute of NIH U01 HL148860, U01HL122700/HL148861. A portion of this research was performed (as part of project award 10.46936/reso.proj. 2021.60258/60007251), in the Environmental Molecular Sciences Laboratory, a DOE Office of Science User Facility sponsored by the Biological and Environmental Research program under Contract No. DE-AC05-76RL01830. Publisher Copyright: Copyright © 2022 Ernault, Kawasaki, Fabrizi, Montañés-Agudo, Amersfoorth, Al-Shama, Coronel and De Groot.
PY - 2022/11/17
Y1 - 2022/11/17
N2 - Background: Atrial fibrosis plays an important role in the development and persistence of atrial fibrillation by promoting reentry. Primary cilia have been identified as a regulator of fibroblasts (FB) activation and extracellular matrix (ECM) deposition. We hypothesized that selective reduction of primary cilia causes increased fibrosis and facilitates reentry. Aim: The aim of this study was to disrupt the formation of primary cilia in FB and examine its consequences on ECM and conduction in a co-culture system of cardiomyocytes (CM) and FB. Materials: Using short interfering RNA (siRNA), we removed primary cilia in neonatal rat ventricular FB by reducing the expression of Ift88 gene required for ciliary assembly. We co-cultured neonatal rat ventricular cardiomyocytes (CM) with FB previously transfected with Ift88 siRNA (siIft88) or negative control siRNA (siNC) for 48 h. We examined the consequences of ciliated fibroblasts reduction on conduction and tissue remodeling by performing electrical mapping, microelectrode, and gene expression measurements. Results: Transfection of FB with siIft88 resulted in a significant 60% and 30% reduction of relative Ift88 expression in FB and CM-FB co-cultures, respectively, compared to siNC. Knockdown of Ift88 significantly increased the expression of ECM genes Fn1, Col1a1 and Ctgf by 38%, 30% and 18%, respectively, in comparison to transfection with siNC. Conduction velocity (CV) was significantly decreased in the siIft88 group in comparison to siNC [11.12 ± 4.27 cm/s (n = 10) vs. 17.00 ± 6.20 (n = 10) respectively, p < 0.05]. The fraction of sites with interelectrode activation block was larger in the siIft88 group than in the siNC group (6.59 × 10−2 ± 8.01 × 10−2 vs. 1.18 × 10−2 ± 3.72 × 10−2 respectively, p < 0.05). We documented spontaneous reentrant arrhythmias in two cultures in the siIft88 group and in none of the siNC group. Action potentials were not significantly different between siNC and siIft88 groups. Conclusion: Disruption of cilia formation by siIft88 causes ECM remodeling and conduction abnormalities. Prevention of cilia loss could be a target for prevention of arrhythmias.
AB - Background: Atrial fibrosis plays an important role in the development and persistence of atrial fibrillation by promoting reentry. Primary cilia have been identified as a regulator of fibroblasts (FB) activation and extracellular matrix (ECM) deposition. We hypothesized that selective reduction of primary cilia causes increased fibrosis and facilitates reentry. Aim: The aim of this study was to disrupt the formation of primary cilia in FB and examine its consequences on ECM and conduction in a co-culture system of cardiomyocytes (CM) and FB. Materials: Using short interfering RNA (siRNA), we removed primary cilia in neonatal rat ventricular FB by reducing the expression of Ift88 gene required for ciliary assembly. We co-cultured neonatal rat ventricular cardiomyocytes (CM) with FB previously transfected with Ift88 siRNA (siIft88) or negative control siRNA (siNC) for 48 h. We examined the consequences of ciliated fibroblasts reduction on conduction and tissue remodeling by performing electrical mapping, microelectrode, and gene expression measurements. Results: Transfection of FB with siIft88 resulted in a significant 60% and 30% reduction of relative Ift88 expression in FB and CM-FB co-cultures, respectively, compared to siNC. Knockdown of Ift88 significantly increased the expression of ECM genes Fn1, Col1a1 and Ctgf by 38%, 30% and 18%, respectively, in comparison to transfection with siNC. Conduction velocity (CV) was significantly decreased in the siIft88 group in comparison to siNC [11.12 ± 4.27 cm/s (n = 10) vs. 17.00 ± 6.20 (n = 10) respectively, p < 0.05]. The fraction of sites with interelectrode activation block was larger in the siIft88 group than in the siNC group (6.59 × 10−2 ± 8.01 × 10−2 vs. 1.18 × 10−2 ± 3.72 × 10−2 respectively, p < 0.05). We documented spontaneous reentrant arrhythmias in two cultures in the siIft88 group and in none of the siNC group. Action potentials were not significantly different between siNC and siIft88 groups. Conclusion: Disruption of cilia formation by siIft88 causes ECM remodeling and conduction abnormalities. Prevention of cilia loss could be a target for prevention of arrhythmias.
KW - atrial fibrillation
KW - cardiac
KW - fibroblasts
KW - fibrosis
KW - primary cilia
UR - http://www.scopus.com/inward/record.url?scp=85143222907&partnerID=8YFLogxK
U2 - https://doi.org/10.3389/fphys.2022.1057200
DO - https://doi.org/10.3389/fphys.2022.1057200
M3 - Article
C2 - 36467697
SN - 1664-042X
VL - 13
JO - Frontiers in physiology
JF - Frontiers in physiology
M1 - 1057200
ER -