BRCA1 protects cardiac microvascular endothelial cells against irradiation by regulating p21-mediated cell cycle arrest
Graphical abstract
Graphical abstract of the findings.
BRCA1 plays a protective role in radiation-induced heart disease by regulating vessel endothelial cell cycle arrest mediated by p21 signaling pathway. Abbreviation: CMECs, cardiac microvascular endothelial cells; RIHD, radiation-induced heart disease.
Introduction
Radiation-induced heart disease (RIHD) has become one of the major late side effects of thoracic radiotherapy with the increasing number of long-term cancer survivors [1]. Accumulated epidemiological studies have shown that a relative increase in the risk of RIHD was observed in cancer survivors who had received thoracic irradiation decades ago, such as in patients with (especially left-sided) breast cancer [2], Hodgkin's lymphoma (relative risks (RR) 2-7-fold) [3,4], childhood cancers [5] and other thoracic malignant tumors [6]. As a late effect, the symptoms of RIHD usually manifest several decades later. The pathologies of RIHD are chronic progressive processes. Previous studies have reported that endothelial cell damage, especially in microvascular cells, may play a vital role in RIHD [[7], [8], [9]]. Cardiac microvascular endothelial cells are more sensitive to radiation than other types of cells in heart tissue [10]. The DNA damage pathway and DNA repair pathway in normal tissue cells, including cell cycle-related pathways, would be activated after ionizing radiation [11,12]. However, the molecular mechanism underlying RIHD has not been fully defined.
BRCA1 is a tumor suppressor gene and plays an important role in DNA damage repair [13]. In addition, BRCA1 is able to directly mediate transcriptional activation of p21 [14]or form a large multi-subunit protein complex known as the BRCA1-associated genome surveillance complex (BASC) [15] in cancer cells. Furthermore, BRCA1 in complex with CDK, cyclin A and cyclin D participates in modulating the cell cycle [16]. Recently, some studies have illustrated that BRCA1 can also regulate cardiac endothelial cells. Praphulla C. Shukla and colleagues reported that BRCA1 in cardiomyocytes played an important role in protecting myocardial infarction patients against cardiac remodeling, poor ventricular function and higher mortality via DNA double-strand break repair and inactivation of p53-mediated pro-apoptotic signaling [17]. Accumulated studies found that p21 is a crucial determinant of DNA repair [18,19].
Until now, the role of BRCA1 in protecting endothelial cells from DNA damage has rarely been explored, especially in RIHD. Therefore, we proposed a hypothesis that BRCA1 protects radiation-induced heart damage by blocking the endothelial cell cycle via the activation of p21. In this study, we demonstrated the underlying mechanism and designed a radiation-induced heart damage model in C57BL/6 mice to confirm this observation both in vitro and in vivo. BRCA1 protected endothelial cells against irradiation by p21-mediated cell cycle arrest.
Section snippets
Cell culture and irradiation
HUVECs were obtained from the Key Laboratory of Molecular Medical in Jiangxi and cultured in endothelial growth medium (Gibco, Grand Island, USA) enriched with 10% fetal bovine serum (Gibco, Grand Island, NY, USA) at 37 °C in a 5% CO2 humidified atmosphere. Cells were irradiated with 2–20 Gy of X-rays (6 MeV, SSD = 100 cm) using a Varian linear accelerator (Varian Clinical 23EX, USA) at a dose rate of 600 cGy/min.
Transfection and RNA interference
The plasmids pSin-EF2-BRCA1 and pCDEF-BRC1-Flag were purchased from Ribobio
BRCA1 and p21 expression are up-regulated in vascular endothelial cells both in vitro and vivo after radiation exposure
To examine the mechanism and prove our hypothesis, we investigated the effect of the BRCA1 on the cellular level using HUVECs. BRCA1 and p21 protein expression levels were detected via Western blotting after the cells received 20 Gy X-ray at different times and ionizing radiation. We found that BRCA1 and p21 protein expression in HUVECs increased in a dose- and time-dependent manner after radiation exposure (Fig. 1A–D). In vivo, BRCA1 and p21 expression levels in mouse cardiac vascular
Discussion
Here, we demonstrated the protective role BRCA1 in RIHD by establishing an irradiation-induced heart damage model in C57BL/6 mice (Fig. 7). In this study, we first demonstrated that BRCA1 plays a role in the response of cardiac endothelial cells to radiation by regulating p21-cell cycle arrest and crisis in HUVEC and ultimately leading to endothelial cell apoptosis. In the wild mouse RIHD model, we confirmed that that the pathological process of RIHD presented as microvessel density decline,
Author contributions
Z-M Z, in charge of design of the work, performed cell and animal experiment, evaluated experimental data, and drafted the manuscript. H-Y D, L X, X-L Z performed part of cell and animal experiment. P-Z performed mouse radiation. H-Y D, X-L and J-C helped to analyze the data. L H and A-W L, the Corresponding author, was in charge of guidance of the design and analysis the whole research.
Declaration of competing interest
The authors report no conflict of interest.
Acknowledgements
This work was supported by the National Nature Science Foundation [grant number 81560509, 81760566 and 81960571], Key Research and Development Project of Jiangxi province [grant number 20171ACB20034, 20181ACG70011 and r20192ACB70013], Science and Technology Innovation Platform of Jiangxi Province [grant number 20171BCD40022 and 20192BCBL23023]. We thank all the other members in the Zhejiang key laboratory of radiation oncology for supporting our work. Thank Caifeng Xie, Associate Professor of
References (51)
- et al.
Cancer survivorship: cardiotoxic therapy in the adult cancer patient; cardiac outcomes with recommendations for patient management
Semin. Oncol.
(2013) - et al.
Morbidity and mortality in long-term survivors of Hodgkin lymphoma: a report from the Childhood Cancer Survivor Study
Blood.
(2011) - et al.
Is cardiac toxicity a relevant issue in the radiation treatment of esophageal cancer?
Radiother. Oncol.
(2015) - et al.
Late proliferating and inflammatory effects on murine microvascular heart and lung endothelial cells after irradiation
Radiother. Oncol.
(2015) - et al.
Ionizing radiation-induced endothelial cell senescence and cardiovascular diseases
Radiat. Res.
(2016) - et al.
ATM, ATR, and DNA-PK: the trinity at the heart of the DNA damage response
Mol. Cell
(2017) - et al.
Function of BRCA1 in the DNA damage response is mediated by ADP-ribosylation
Cancer Cell
(2013) - et al.
Multiple functions of p21 in cell cycle, apoptosis and transcriptional regulation after DNA damage
DNA Repair.
(2016) - et al.
Irradiation induced modest changes in murine cardiac function despite progressive structural damage to the myocardium and microvasculature
Radiother. Oncol.
(2012) - et al.
Pathogenetic mechanisms in radiation fibrosis
Radiother. Oncol.
(2010)
Chronic intermittent hypobaric hypoxia attenuates radiation induced heart damage in rats
Life Sci.
Ionising radiation induces persistent alterations in the cardiac mitochondrial function of C57BL/6 mice 40 weeks after local heart exposure
Radiother. Oncol.
Mechanisms of normal tissue injury from irradiation
Semin. Radiat. Oncol.
The radiotherapeutic injury–a complex ‘wound’
Radiother. Oncol.
Targeting DNA repair in cancer: beyond PARP inhibitors
Cancer Discov.
Molecular determinants of radiosensitivity in normal and tumor tissue: a bioinformatic approach
Cancer Lett.
Role of cell cycle in mediating sensitivity to radiotherapy
Int. J. Radiat. Oncol. Biol. Phys.
Risk of ischemic heart disease in women after radiotherapy for breast cancer
N. Engl. J. Med.
Review of the cardiac long-term effects of therapy for Hodgkin lymphoma
Br. J. Haematol.
The childhood cancer survivor study: a National Cancer Institute–supported resource for outcome and intervention research
J. Clin. Oncol. Off. J. Am. Soc. Clin. Oncol.
Potential markers and metabolic processes involved in the mechanism of radiation-induced heart injury
Can. J. Physiol. Pharmacol.
Pathology and biology of radiation-induced cardiac disease
J. Radiat. Res.
The DNA-damage response in human biology and disease
Nature.
BRCA1, FANCD2 and Chk1 are potential molecular targets for the modulation of a radiation-induced DNA damage response in bystander cells
Cancer Lett.
Arrest of the cell cycle by the tumour-suppressor BRCA1 requires the CDK-inhibitor p21WAF1/CiP1
Nature
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