Construction and validation of a nomogram to predict left ventricular hypertrophy in low-risk patients with hypertension.

Electrocardiography (ECG) is an accessible diagnostic tool for screening patients with hypertensive left ventricular hypertrophy (LVH). However, its diagnostic sensitivity is low, with a high probability of false-negatives. Thus, this study aimed to establish a clinically useful nomogram to supplement the assessment of LVH in patients with hypertension and without ECG-LVH based on Cornell product criteria (low-risk hypertensive population). A cross-sectional dataset was used for model construction and divided into development (n = 2906) and verification (n = 1447) datasets. A multivariable logistic regression risk model and nomogram were developed after screening for risk factors. Of the 4353 low-risk hypertensive patients, 673 (15.4%) had LVH diagnosed by echocardiography (Echo-LVH). Eleven risk factors were identified: hypertension awareness, duration of hypertension, age, sex, high waist-hip ratio, education level, tea consumption, hypochloremia, and other ECG-LVH diagnostic criteria (including Sokolow-Lyon, Sokolow-Lyon products, and Peguero-Lo Presti). For the development and validation datasets, the areas under the curve were 0.724 (sensitivity = 0.606) and 0.700 (sensitivity = 0.663), respectively. After including blood pressure, the areas under the curve were 0.735 (sensitivity = 0.734) and 0.716 (sensitivity = 0.718), respectively. This novel nomogram had a good predictive ability and may be used to assess the Echo-LVH risk in patients with hypertension and without ECG-LVH based on Cornell product criteria.

Atorvastatin rescues vascular endothelial injury in hypertension by WWP2-mediated ubiquitination and degradation of ATP5A.

As a widely used lipid-lowering drug in clinical practice, atorvastatin is widely recognized for its role in protecting vascular endothelium in the cardiovascular system. However, a clear mechanistic understanding of its action is lacking. Here, we found that atorvastatin counteracted angiotensin II-induced vascular endothelial injury in mice with hypertension. Mechanistically, atorvastatin up-regulated WWP2, a E6AP C-terminus (HECT)-type E3 ubiquitin ligase with an essential role in regulating protein ubiquitination and various biological processes, thereby rescuing vascular endothelial injury. By ubiquitinating ATP5A (ATP synthase mitochondrial F1 complex subunit alpha), WWP2 degraded ATP5A via the proteasome pathway, stabilizing Bcl-2/Bax in the mitochondrial pathway of apoptosis. Moreover, atorvastatin further ameliorated death of vascular endothelial cells and improved vascular endothelial functions under WWP2 overexpression, whereas WWP2 knockout abrogated these beneficial effects of atorvastatin. Furthermore, we generated endothelial cell-specific WWP2 knockout mice, and this WWP2-mediated mechanism was faithfully recapitulated in vivo. Thus, we propose that activation of a WWP2-dependent pathway that is pathologically repressed in damaged vascular endothelium under hypertension is a major mechanism of atorvastatin. Our findings are also pertinent to develop novel therapeutic strategies for vascular endothelial injury-related cardiovascular diseases.

α-myosin heavy chain lactylation maintains sarcomeric structure and function and alleviates the development of heart failure.

The sarcomeric interaction of α-myosin heavy chain (α-MHC) with Titin is vital for cardiac structure and contraction. However, the mechanism regulating this interaction in normal and failing hearts remains unknown. Lactate is a crucial energy substrate of the heart. Here, we identify that α-MHC undergoes lactylation on lysine 1897 to regulate the interaction of α-MHC with Titin. We observed a reduction of α-MHC K1897 lactylation in mice and patients with heart failure. Loss of K1897 lactylation in α-MHC K1897R knock-in mice reduces α-MHC-Titin interaction and leads to impaired cardiac structure and function. Furthermore, we identified that p300 and Sirtuin 1 act as the acyltransferase and delactylase of α-MHC, respectively. Decreasing lactate production by chemical or genetic manipulation reduces α-MHC lactylation, impairs α-MHC-Titin interaction and worsens heart failure. By contrast, upregulation of the lactate concentration by administering sodium lactate or inhibiting the pivotal lactate transporter in cardiomyocytes can promote α-MHC K1897 lactylation and α-MHC-Titin interaction, thereby alleviating heart failure. In conclusion, α-MHC lactylation is dynamically regulated and an important determinant of overall cardiac structure and function. Excessive lactate efflux and consumption by cardiomyocytes may decrease the intracellular lactate level, which is the main cause of reduced α-MHC K1897 lactylation during myocardial injury. Our study reveals that cardiac metabolism directly modulates the sarcomeric structure and function through lactate-dependent modification of α-MHC.

Deacetylation of Septin4 by SIRT2 (Silent Mating Type Information Regulation 2 Homolog-2) Mitigates Damaging of Hypertensive Nephropathy.

BACKGROUND: Hypertension can lead to podocyte damage and subsequent apoptosis, eventually resulting in glomerulosclerosis. Although alleviating podocyte apoptosis has clinical significance for the treatment of hypertensive nephropathy, an effective therapeutic target has not yet been identified. The function of septin4, a proapoptotic protein and an important marker of organ damage, is regulated by post-translational modification. However, the exact role of septin4 in regulating podocyte apoptosis and its connection to hypertensive renal damage remains unclear. METHODS: We investigated the function and mechanism of septin4 in hypertensive nephropathy to discover a theoretical basis for targeted treatment. Mouse models including Rosa 26 (Gt(ROSA)26Sor)-SIRT2 (silent mating type information regulation 2 homolog-2)-Flag-TG (transgenic) (SIRT2-TG) mice SIRT2-knockout, and septin4-K174Q mutant mice, combined with proteomic and acetyl proteomics analysis, followed by multiple molecular biological methodologies, were used to demonstrate mechanisms of SIRT2-mediated deacetylation of septin4-K174 in hypertensive nephropathy. RESULTS: Using transgenic septin4-K174Q mutant mice treated with the antioxidant Tempol, we found that hyperacetylation of the K174 site of septin4 exacerbates Ang II (angiotensin II)- induced hypertensive renal injury resulting from oxidative stress. Proteomics and Western blotting assays indicated that septin4-K174Q activates the cleaved-PARP1 (poly [ADP-ribose] polymerase family, member 1)-cleaved-caspase3 pathway. In septin4-knockdown human renal podocytes, septin4-K174R, which mimics deacetylation at K174, rescues podocyte apoptosis induced by Ang II. Immunoprecipitation and mass spectrometry analyses identified SIRT2 as a deacetylase that interacts with the septin4 GTPase domain and deacetylates septin4-K174. In Sirt2-deficient mice and SIRT2-knockdown renal podocytes, septin4-K174 remains hyperacetylated and exacerbates hypertensive renal injury. By contrast, in Rosa26-Sirt2-Flag (SIRT2-TG) mice and SIRT2-knockdown renal podocytes reexpressing wild-type SIRT2, septin4-K174 is hypoacetylated and mitigates hypertensive renal injury. CONCLUSIONS: Septin4, when activated through acetylation of K174 (K174Q), promotes hypertensive renal injury. Septin4-K174R, which mimics deacetylation by SIRT2, inhibits the cleaved-PARP1-cleaved-caspase3 pathway. Septin4-K174R acts as a renal protective factor, mitigating Ang II-induced hypertensive renal injury. These findings indicate that septin4-K174 is a potential therapeutic target for the treatment of hypertensive renal injury.

The role of E3 ubiquitin ligase WWP2 and the regulation of PARP1 by ubiquitinated degradation in acute lymphoblastic leukemia.

Acute lymphoblastic leukemia (ALL) has been a huge threat for people's health and finding effective target therapy is urgent and important. WWP2, as one of E3 ubiquitin ligase, is involved in many biological processes by specifically binding to substrates. PARP1 plays a role in cell apoptosis and is considered as a therapeutic target of certain cancers. In this study, we firstly found that WWP2 expressed higher in newly diagnosed ALL patients comparing with complete remission (CR) ALL patients and normal control people, and WWP2 in relapse ALL patients expressed higher than normal control people. WWP2 expression was related with the FAB subtype of ALL and the proportion of blast cells in bone marrow blood tested by flow cytometry. We demonstrated knockout WWP2 inhibited the ALL growth and enhanced apoptosis induced by Dox in vitro and vivo for the first time. WWP2 negatively regulated and interacted with PARP1 and WWP2 mechanically degraded PARP1 through polyubiquitin-proteasome pathway in ALL. These findings suggested WWP2 played a role in ALL development as well as growth and apoptosis, and also displayed a regulatory pathway of PARP1, which provided a new potential therapeutic target for the treatment of ALL.

The regulatory roles of the E3 ubiquitin ligase NEDD4 family in DNA damage response.

E3 ubiquitin ligases, an important part of ubiquitin proteasome system, catalyze the covalent binding of ubiquitin to target substrates, which plays a role in protein ubiquitination and regulates different biological process. DNA damage response (DDR) is induced in response to DNA damage to maintain genome integrity and stability, and this process has crucial significance to a series of cell activities such as differentiation, apoptosis, cell cycle. The NEDD4 family, belonging to HECT E3 ubiquitin ligases, is reported as regulators that participate in the DDR process by recognizing different substrates. In this review, we summarize recent researches on NEDD4 family members in the DDR and discuss the roles of NEDD4 family members in the cascade reactions induced by DNA damage. This review may contribute to the further study of pathophysiology for certain diseases and pharmacology for targeted drugs.

DNA Polymerase Gamma Recovers Mitochondrial Function and Inhibits Vascular Calcification by Interacted with p53.

DNA polymerase gamma (PolG) is the major polymerase of mitochondrial DNA (mtDNA) and essential for stabilizing mitochondrial function. Vascular calcification (VC) is common senescence related degenerative pathology phenomenon in the end-stage of multiple chronic diseases. Mitochondrial dysfunction was often observed in calcified vessels, but the function and mechanism of PolG in the calcification process was still unknown. The present study found PolGD257A/D257A mice presented more severe calcification of aortas than wild type (WT) mice with vitamin D3 (Vit D3) treatment, and this phenomenon was also confirmed in vitro. Mechanistically, PolG could enhance the recruitment and interaction of p53 in calcification condition to recover mitochondrial function and eventually to resist calcification. Meanwhile, we found the mutant PolG (D257A) failed to achieve the same rescue effects, suggesting the 3'-5' exonuclease activity guarantee the enhanced interaction of p53 and PolG in response to calcification stimulation. Thus, we believed that it was PolG, not mutant PolG, could maintain mitochondrial function and attenuate calcification in vitro and in vivo. And PolG could be a novel potential therapeutic target against calcification, providing a novel insight to clinical treatment.

An unexpected role for BAG3 in regulating PARP1 ubiquitination in oxidative stress-related endothelial damage.

Oxidative stress-associated endothelial damage is the initiation factor of cardiovascular disease, and protein posttranslational modifications play critical roles in this process. Bcl-2-associated athanogene 3 (BAG3) is a molecular chaperone regulator of the BAG family, which interacts with various proteins and influences cell survival by activating multiple pathways. BAG3 undergoes posttranslational modifications; however, research evaluating BAG3 acetylation and its regulatory mechanism is lacking. In addition, the interacting protein and regulatory mechanism of BAG3 in oxidative stress-associated endothelial damage remain unclear. Here, key molecular interactions and protein modifications of BAG3 were identified in oxidative stress-associated endothelial damage. Endothelial-specific BAG3 knockout in the mouse model starkly enhances oxidative stress-associated endothelial damage and vascular remodeling, while BAG3 overexpression in mice significantly relieves this process. Mechanistically, poly(ADP-ribose) polymerase 1 (PARP1), causing oxidative stress, was identified as a novel physiological substrate of BAG3. Indeed, BAG3 binds to PARP1's BRCT domain to promote its ubiquitination (K249 residue) by enhancing the E3 ubiquitin ligase WWP2, which leads to proteasome-induced PARP1 degradation. Furthermore, we surprisingly found that BAG3 represents a new substrate of the acetyltransferase CREB-binding protein (CBP) and the deacetylase Sirtuin 2 (SIRT2) under physiological conditions. CBP/SIRT2 interacted with BAG3 and acetylated/deacetylated BAG3's K431 residue. Finally, deacetylated BAG3 promoted the ubiquitination of PARP1. This work reveals a novel regulatory system, with deacetylation-dependent regulation of BAG3 promoting PARP1 ubiquitination and degradation via enhancing WWP2, which is one possible mechanism to decrease vulnerability of oxidative stress in endothelial cells.

Deacetylation-dependent regulation of PARP1 by SIRT2 dictates ubiquitination of PARP1 in oxidative stress-induced vascular injury.

Poly(ADP-ribose) polymerase 1 (PARP1) has a major regulatory role in cardiovascular disease. However, inhibiting PARP1 activity does not significantly improve clinical outcomes of cardiovascular disease, which suggests that the regulatory mechanism of PARP1 in cardiovascular disease is unclear. Here, we focused on deacetylation regulatory mechanisms of PARP1 and crosstalk of PARP1 post-translational modifications. We uncovered the crucial molecular interactions and protein modifications of deacetylase Sirtuin 2 (SIRT2) and PARP1 in vascular damage. The results showed that SIRT2 was involved in this process and oxidative stress damage factor PARP1 was a novel physiological substrate of SIRT2. SIRT2 interacted with PARP1 at the PARP-A-helical domain and deacetylated the K249 residue of PARP1. Furthermore, SIRT2 promoted ubiquitination of the K249 residue of PARP1 via mobilization of the E3 ubiquitin ligase WW domain-containing protein 2 (WWP2), which led to proteasome-mediated degradation of PARP1. Knockout of SIRT2 in mice and cells increased PARP1 acetylation and decreased PARP1 ubiquitination, which in turn aggravated oxidative stress-induced vascular injury and remodeling. Conversely, overexpression of SIRT2 in mice and cells decreased PARP1 acetylation, increased PARP1 ubiquitination, and relieved oxidative stress-induced vascular injury and remodeling. Overall, this study revealed a previously unrecognized mechanistic link between SIRT2 and PARP1 in the regulation of oxidative stress-induced vascular injury.

Septin4 promotes cardiomyocytes apoptosis by enhancing the VHL-mediated degradation of HIF-1α.

Septin4, a protein localized at mitochondrion, can promote cells apoptosis mainly by binding XIAP (X-linked inhibitors of apoptosis), however, nothing is known about the role and mechanism of Septin4 in cardiomyocytes apoptosis. Here in the current study, we report that HIF-1α (hypoxia-inducible factor 1 alpha) is a novel interacting protein with Septin4 at Septin4-GTPase domain. In addition, Septin4 enhances the binding between HIF-1α and the E3 ubiquitin ligase VHL (von Hippel-Lindau protein) to down-regulate HIF-1α, and by reducing cardio-protective factor HIF-1α levels, Septin4 aggravated the hypoxia-induced cardiomyocytes apoptosis. We believe these findings will be beneficial to provide effective strategies for clinical treatment of myocardial ischemia and the subsequent injury caused by myocardial hypoxia.

The role of SIRT2 in vascular-related and heart-related diseases: A review.

At present, cardiovascular disease is one of the important factors of human death, and there are many kinds of proteins involved. Sirtuins family proteins are involved in various physiological and pathological activities of the human body. Among them, there are more and more studies on the relationship between sirtuin2 (SIRT2) protein and cardiovascular diseases. SIRT2 can effectively inhibit pathological cardiac hypertrophy. The effect of SIRT2 on ischaemia-reperfusion injury has different effects under different conditions. SIRT2 can reduce the level of reactive oxygen species (ROS), which may help to reduce the severity of diabetic cardiomyopathy. SIRT2 can affect a variety of cardiovascular diseases, energy metabolism and the ageing of cardiomyocytes, thereby affecting heart failure. SIRT2 also plays an important role in vascular disease. For endothelial cell damage used by oxidative stress, the role of SIRT2 is bidirectional, which is related to the degree of oxidative stress stimulation. When the degree of stimulation is small, SIRT2 plays a protective role, and when the degree of stimulation increases to a certain level, SIRT2 plays a negative role. In addition, SIRT2 is also involved in the remodelling of blood vessels and the repair of skin damage.

The Role of BRG1 in Antioxidant and Redox Signaling.

Redox homeostasis is regulated by critical molecules that modulate antioxidant and redox signaling (ARS) within the cell. Imbalances among these molecules can lead to oxidative stress and damage to cell functions, causing a variety of diseases. Brahma-related gene 1 (BRG1), also known as SMARCA4, is the central ATPase catalytic subunit of the switch/sucrose nonfermentable (SWI/SNF) chromatin remodeling complex, which plays a core role in DNA replication, repair, recombination, and transcriptional regulation. Numerous recent studies show that BRG1 is involved in the regulation of various cellular processes associated with ARS. BRG1, as a major factor in chromatin remodeling, is essential for the repair of oxidative stress-induced DNA damage and the activation of antioxidant genes under oxidative stress. Consequently, a comprehensive understanding of the roles of BRG1 in redox homeostasis is crucial to understand the normal functioning as well as pathological mechanisms. In this review, we summarized and discussed the role of BRG1 in the regulation of ARS.

E3 Ubiquitin ligase NEDD4 family­regulatory network in cardiovascular disease.

Protein ubiquitination represents a critical modification occurring after translation. E3 ligase catalyzes the covalent binding of ubiquitin to the protein substrate, which could be degraded. Ubiquitination as an important protein post-translational modification is closely related to cardiovascular disease. The NEDD4 family, belonging to HECT class of E3 ubiquitin ligases can recognize different substrate proteins, including PTEN, ENaC, Nav1.5, SMAD2, PARP1, Septin4, ALK1, SERCA2a, TGFßR3 and so on, via the WW domain to catalyze ubiquitination, thus participating in multiple cardiovascular-related disease such as hypertension, arrhythmia, myocardial infarction, heart failure, cardiotoxicity, cardiac hypertrophy, myocardial fibrosis, cardiac remodeling, atherosclerosis, pulmonary hypertension and heart valve disease. However, there is currently no review comprehensively clarifying the important role of NEDD4 family proteins in the cardiovascular system. Therefore, the present review summarized recent studies about NEDD4 family members in cardiovascular disease, providing novel insights into the prevention and treatment of cardiovascular disease. In addition, assessing transgenic animals and performing gene silencing would further identify the ubiquitination targets of NEDD4. NEDD4 quantification in clinical samples would also constitute an important method for determining NEDD4 significance in cardiovascular disease.

WWP2 regulates SIRT1-STAT3 acetylation and phosphorylation involved in hypertensive angiopathy.

WWP2 is a HECT-type E3 ubiquitin ligase that regulates various physiological and pathological activities by binding to different substrates, but its function and regulatory mechanism in vascular smooth muscle cells (VSMCs) are still unknown. Here, we clarified the role of WWP2 in the regulation of SIRT1-STAT3 and the impact of this regulatory process in VSMCs. We demonstrated that WWP2 expression was significantly increased in angiotensin II-induced VSMCs model. Knockdown of WWP2 significantly inhibited angiotensin II-induced VSMCs proliferation, migration and phenotypic transformation, whereas overexpression of WWP2 had opposite effects. In vivo experiments showed that vascular smooth muscle-specific WWP2 knockout mice significantly relieved angiotensin II-induced hypertensive angiopathy. Mechanistically, mass spectrometry and co-immunoprecipitation assays identified that WWP2 is a novel interacting protein of SIRT1 and STAT3. Moreover, WWP2 formed a complex with SIRT1-STAT3, inhibiting the interaction between SIRT1 and STAT3, then reducing the inhibitory effect of SIRT1 on STAT3, ensuing promoting STAT3-K685 acetylation and STAT3-Y705 phosphorylation in angiotensin II-induced VSMCs and mice. In conclusion, WWP2 modulates hypertensive angiopathy by regulating SIRT1-STAT3 and WWP2 suppression in VSMCs can alleviate hypertensive angiopathy vitro and vivo. These findings provide new insights into the treatment of hypertensive vascular diseases.

Septin4 as an autophagy modulator regulates Angiotensin-II mediated VSMCs proliferation and migration.

Vascular smooth muscle cells (VSMCs) proliferation and migration play a fundamental role during the process of hypertensive angiopathy. Angiotensin-II (Ang-II) is one of the robust phenotype-modulating agents, which changes VSMCs to efficiently proliferate and migrate. The mechanism of the proliferation and migration is not well understood yet. Septin4, as a member of GTP binding protein family, is widely expressed in the eukaryotic cells and considered to be an essential component of the cytoskeleton which is involved in many important physiological processes. We approved that Septin4 expression was upregulated in mouse aorta by continuous infusion of Ang-II and in cultured VSMCs treated with Ang-II. Overexpression of Septin4 led to lower level of autophagy and decreased capacity of proliferation and migration. In order to identify the mechanism by which Septin4 interacts with these processes, we blocked autophagy by chloroquine (CQ). After inhibiting the autophagy, the ability of proliferation and migration was further restrained in the Septin4 overexpression VSMCs. In conclusion, our results indicated that during the process of VSMCs proliferation and migration induced by Ang-II, Septin4 modulated autophagy and thus regulated the activity of proliferation and migration.

Septin4 Prevents PDGF-BB-induced HAVSMC Phenotypic Transformation, Proliferation and Migration by Promoting SIRT1-STAT3 Deacetylation and Dephosphorylation.

SIRT1 and STAT3 are key to human aortic vascular smooth muscle cells (HAVSMCs) proliferation, migration and phenotypic transformation, but the regulatory mechanism of SIRT1-STAT3 in this process is still unclear. Septin4 is a cytoskeleton-related protein that regulates oxidative stress-vascular endothelial injury. However, the role and underlying mechanism of Septin4 in atherosclerosis remains unknown. Here, we revealed the role and mechanism of Septin4 in regulating SIRT1-STAT3 in atherosclerosis. We determined that the expression of Septin4 were markedly increased in Apoe-/- atherosclerosis mice and PDGF-BB-induced HAVSMCs. Knockdown of Septin4 significantly increased PDGF-BB-induced HAVSMCs proliferation, migration and phenotypic transformation, while overexpression of Septin4 had the opposite effects. Mechanically, co-immunoprecipitation results demonstrated that Septin4 was a novel interacting protein of STAT3 and SIRT1. Septin4 formed a complex with SIRT1-STAT3, enhancing the interaction between SIRT1 and STAT3, ensuing promoting SIRT1-regulated STAT3-K685 deacetylation and STAT3-Y705 dephosphorylation, which inhibited PDGF-BB-induced HAVSMCs proliferation, migration and phenotype transformation. Therefore, our findings provide novel insights into the prevention and treatment of atherosclerosis.