Rewiring of 3D Chromatin Topology Orchestrates Transcriptional Reprogramming and the Development of Human Dilated Cardiomyopathy.

BACKGROUND: Transcriptional reconfiguration is central to heart failure, the most common cause of which is dilated cardiomyopathy (DCM). The effect of 3-dimensional chromatin topology on transcriptional dysregulation and pathogenesis in human DCM remains elusive. METHODS: We generated a compendium of 3-dimensional epigenome and transcriptome maps from 101 biobanked human DCM and nonfailing heart tissues through highly integrative chromatin immunoprecipitation (H3K27ac [acetylation of lysine 27 on histone H3]), in situ high-throughput chromosome conformation capture, chromatin immunoprecipitation sequencing, assay for transposase-accessible chromatin using sequencing, and RNA sequencing. We used human induced pluripotent stem cell-derived cardiomyocytes and mouse models to interrogate the key transcription factor implicated in 3-dimensional chromatin organization and transcriptional regulation in DCM pathogenesis. RESULTS: We discovered that the active regulatory elements (H3K27ac peaks) and their connectome (H3K27ac loops) were extensively reprogrammed in DCM hearts and contributed to transcriptional dysregulation implicated in DCM development. For example, we identified that nontranscribing NPPA-AS1 (natriuretic peptide A antisense RNA 1) promoter functions as an enhancer and physically interacts with the NPPA (natriuretic peptide A) and NPPB (natriuretic peptide B) promoters, leading to the cotranscription of NPPA and NPPB in DCM hearts. We revealed that DCM-enriched H3K27ac loops largely resided in conserved high-order chromatin architectures (compartments, topologically associating domains) and their anchors unexpectedly had equivalent chromatin accessibility. We discovered that the DCM-enriched H3K27ac loop anchors exhibited a strong enrichment for HAND1 (heart and neural crest derivatives expressed 1), a key transcription factor involved in early cardiogenesis. In line with this, its protein expression was upregulated in human DCM and mouse failing hearts. To further validate whether HAND1 is a causal driver for the reprogramming of enhancer-promoter connectome in DCM hearts, we performed comprehensive 3-dimensional epigenome mappings in human induced pluripotent stem cell-derived cardiomyocytes. We found that forced overexpression of HAND1 in human induced pluripotent stem cell-derived cardiomyocytes induced a distinct gain of enhancer-promoter connectivity and correspondingly increased the expression of their connected genes implicated in DCM pathogenesis, thus recapitulating the transcriptional signature in human DCM hearts. Electrophysiology analysis demonstrated that forced overexpression of HAND1 in human induced pluripotent stem cell-derived cardiomyocytes induced abnormal calcium handling. Furthermore, cardiomyocyte-specific overexpression of Hand1 in the mouse hearts resulted in dilated cardiac remodeling with impaired contractility/Ca2+ handling in cardiomyocytes, increased ratio of heart weight/body weight, and compromised cardiac function, which were ascribed to recapitulation of transcriptional reprogramming in DCM. CONCLUSIONS: This study provided novel chromatin topology insights into DCM pathogenesis and illustrated a model whereby a single transcription factor (HAND1) reprograms the genome-wide enhancer-promoter connectome to drive DCM pathogenesis.

Influence of FOSL1 Inhibition on Vascular Calcification and ROS Generation through Ferroptosis via P53-SLC7A11 Axis.

BACKGROUND: Vascular calcification during aging is highly prevalent in patients with cardiovascular disease; however, there is still no improvement in clarifying the development of vascular calcification. FOSL1 is a transcription regulator belonging to the AP-1 family, which has a unique function in vascular senescence, but its role in vascular calcification needs to be further explored. METHODS: Primary mouse vascular smooth muscle cells were isolated and used to construct a calcification model in vitro. Seven-week-old male C57BL/6 mice were used to build the vitD3-induced calcification model in vivo. qRT-PCR and western blot were used to verify the expression of FOSL1 and other genes expressed in vascular smooth muscle cells and aortas. The level of calcification was determined by Alizarin Red S (ARS) staining and the calcium content assay. The level of cellular GSH was detected by the GSH assay kit. RESULTS: Here, we report that FOSL1 was up-regulated after high-calcium/phosphate treatment in both the in vivo and in vitro vascular calcification models. Functional studies have shown that the reduction of FOSL1 attenuates ferroptosis and calcification in vascular smooth muscle cells, as indicated by ARS staining, calcium content assay, and western blot. The inhibition of FOSL1 downregulated the expression of bone-related molecules including Msh Homeobox 2 (MSX2) and tumor necrosis factor receptor superfamily, member 11b/osteoprotegerin (OPG), suggesting that FOSL1 promoted osteogenic differentiation of vascular smooth muscle cells. Furthermore, we found that the ferroptosis-inducing drug erastin can significantly accelerate calcification in the aortic ring while Ferrostatin-1 (fer-1), a drug to protect cells from ferroptosis, can alleviate calcification. Further experiments have shown that inhibiting FOSL1 can promote the expression of ferroptosis-related genes and attenuate calcification. Functionally, cellular GSH levels were increased after the reduction of FOSL1. CONCLUSIONS: In this study, we observed a significant protective effect when we reduced the expression of FOSL1 during vascular calcification, and this effect might regulate ferroptosis to a great extent.

Liquid-Liquid Phase Separation in Cardiovascular Diseases.

Liquid-liquid phase separation (LLPS) is a biochemical process in cells that can drive proteins, RNA, and other molecules to concentrate into droplets. These droplets do not have a lipid membrane but rather exist as distinct organelles relative to the surrounding environment, and act as biochemical reaction chambers. In recent years, significant progress has been made in the study of LLPS, especially in the neurodegenerative disease, cancer, and virology fields, but little is known about LLPS in cardiovascular disease (CVD). In this review, we discuss the current understanding of the mechanism and biological functions of LLPS, particularly its roles in regulating CVD.

The Novel LncRNA AK035396 Drives Cardiomyocyte Apoptosis Through Mterf1 in Myocardial Ischemia/Reperfusion Injury.

Background: Myocardial ischaemia/reperfusion (I/R) injury is still a major challenge in clinical treatment. The role of long non-coding RNA (lncRNA) in the regulation of myocardial I/R injury still needs to be elucidated. Methods: The primary isolated neonatal mousse cardiomyocytes and adult mice were used to construct a myocardial ischemia-reperfusion model. qRT-PCR is used to verify gene expression in myocardial tissue and myocardial cells. The effect of AK035396 in primary cardiomyocytes and mouse myocardium was confirmed by TUNEL staining and in vitro flow cytometry experiments. RNA pulldown and Western blot were used to identify AK035396 interacting proteins. The expression of apoptosis-related proteins was identified by qRT-PCR and Western blot. Results: In vivo and in vitro MIRI models, AK035396 was up-regulated after myocardial infarction. Functional studies have shown that knockdown of AK035396 reduces the apoptosis of primary cardiomyocytes and mouse myocardial tissue. AK035396 directly interacts with Mterf1 and inhibits the level of Mterf1. Further experiments have shown that inhibiting Mterf1 will promote the expression of mitochondrial genes COXII and CYTb and cause cell apoptosis. Conclusion: AK035396 plays an important role in myocardial ischaemia-reperfusion injury by regulating the Mterf1-COXII/CYTb pathway.