Related Experiment Video
Updated: Sep 26, 2026
Simultaneous Isolation of High Quality Cardiomyocytes, Endothelial Cells, and Fibroblasts from an Adult Rat Heart
Published on: May 19, 2017
Antiapoptotic effect of endothelin-1 in rat cardiomyocytes in vitro
Yukiyo Ogata1, Masafumi Takahashi, Shuichi Ueno
1Division of Cardiovascular Medicine, Department of Medicine, Jichi Medical School, Tochigi, Japan.
Insights
Endothelin-1 (ET-1) prevents cardiac myocyte apoptosis, a key feature in heart failure, by activating ETA receptors and a c-Src/Bcl-xL pathway. This discovery offers new insights into treating heart conditions.
Area of Science:
- Cardiology
- Molecular Biology
- Cell Biology
Background:
- Apoptosis of cardiac myocytes is implicated in heart failure and ischemic heart disease.
- Endothelin-1 (ET-1) is recognized for its significant role in these cardiovascular pathologies.
Purpose of the Study:
- To investigate the effect of ET-1 on cardiomyocyte apoptosis.
- To elucidate the molecular pathways involved in ET-1's action on cardiac myocytes.
Main Methods:
- Evaluated apoptosis in rat cardiomyocytes (H9c2, neonatal) using morphological criteria, DNA fragmentation, DAPI staining, and TUNEL assay.
- Utilized specific pharmacological inhibitors and adenovirus-mediated gene transfer of kinase-inactive (KI) c-Src.
- Assessed Bcl-xL upregulation using a luciferase-based reporter system.
Main Results:
- ET-1 dose-dependently prevented serum deprivation-induced apoptosis via ETA receptors, unlike angiotensin II.
- ET-1 inhibited cytochrome c release and its antiapoptotic effect was mediated by a tyrosine kinase pathway involving c-Src.
- ET-1 upregulated Bcl-xL expression, a process dependent on c-Src and Pyk2, but not other tyrosine kinases.
Conclusions:
- ET-1 exerts a protective effect against cardiac myocyte apoptosis.
- The antiapoptotic mechanism involves ETA receptor activation, leading to c-Src and Bcl-xL-dependent signaling.
- These findings highlight a novel therapeutic pathway for heart conditions involving ET-1 signaling.
Abstract:
Apoptosis of cardiac myocytes is thought to be a feature of many pathological disorders, including congestive heart failure (CHF) and ischemic heart disease (IHD). Because recent investigations indicate that endothelin-1 (ET-1) plays an important role in CHF and IHD, we investigated the effect of ET-1 on cardiomyocyte apoptosis. The presence of apoptosis in rat cardiomyocytes (H9c2 and neonatal) was evaluated by morphological criteria, electrophoresis of DNA fragments, 4',6'-diamidine-2'-phenylindole staining, and TUNEL analysis. ET-1, but not angiotensin II, prevented apoptosis induced by serum deprivation via ETA receptors in a dose-dependent manner (1 to 100 nmol/L). ET-1 also prevented cytochrome c release from mitochondria to the cytosol. The use of specific pharmacological inhibitors demonstrated that the antiapoptotic effect of ET-1 was mediated through a tyrosine kinase pathway (genistein and AG490) but not through protein kinase C (PKC; calphostin C), mitogen-activated protein kinases (PD98059 and SB203580), or PKA (KT5270) pathways. Adenovirus-mediated gene transfer of kinase-inactive (KI) c-Src reversed the antiapoptotic effect of ET-1. We further investigated whether Bcl-xL, an antiapoptotic molecule, would be upregulated by using a luciferase-based reporter system. ET-1 upregulated Bcl-xL, and this upregulation was inhibited by genistein or AG490 but not by calphostin C. The experiments with KI mutants for various tyrosine kinases revealed that c-Src and Pyk2 (but not JAK1, Jak2, Syk, and Tec) are involved in ET-1-induced upregulation of Bcl-xL expression. These findings suggest that ET-1 prevents apoptosis in cardiac myocytes through the ETA receptor and the subsequent c-Src/Bcl-xL-dependent pathway.
