Histone methyltransferase MLL4 protects against pressure overload-induced heart failure via a THBS4-mediated protection in ER stress.

Pressure overload-induced pathological cardiac hypertrophy eventually leads to heart failure (HF). Unfortunately, lack of effective targeted therapies for HF remains a challenge in clinical management. Mixed-lineage leukemia 4 (MLL4) is a member of the SET family of histone methyltransferase enzymes, which possesses histone H3 lysine 4 (H3K4)-specific methyltransferase activity. However, whether and how MLL4 regulates cardiac function is not reported in adult HF. Here we report that MLL4 is required for endoplasmic reticulum (ER) stress homeostasis of cardiomyocytes and protective against pressure overload-induced cardiac hypertrophy and HF. We observed that MLL4 is increased in the heart tissue of HF mouse model and HF patients. The cardiomyocyte-specific deletion of Mll4 (Mll4-cKO) in mice leads to aggravated ER stress and cardiac dysfunction following pressure overloading. MLL4 knockdown neonatal rat cardiomyocytes (NRCMs) also display accelerated decompensated ER stress and hypertrophy induced by phenylephrine (PE). The combined analysis of Cleavage Under Targets and Tagmentation sequencing (CUT&Tag-seq) and RNA sequencing (RNA-seq) data reveals that, silencing of Mll4 alters the chromatin landscape for H3K4me1 modification and gene expression patterns in NRCMs. Interestingly, the deficiency of MLL4 results in a marked reduction of H3K4me1 and H3K27ac occupations on Thrombospondin-4 (Thbs4) gene loci, as well as Thbs4 gene expression. Mechanistically, MLL4 acts as a transcriptional activator of Thbs4 through mono-methylation of H3K4 and further regulates THBS4-dependent ER stress response, ultimately plays a role in HF. Our study indicates that pharmacologically targeting MLL4 and ER stress might be a valid therapeutic approach to protect against cardiac hypertrophy and HF.

Evaluation of time-dependent phenotypes of myocardial ischemia-reperfusion in mice.

BACKGROUND: A mouse model of myocardial ischemia-reperfusion (I/R) is widely used to study myocardial ischemia-reperfusion injury (I/RI). However, few studies focus on the direct comparison of the extent of pathological events resulting from variant durations of ischemia and reperfusion process. METHODS: A mouse model of I/RI was established by ligation and perfusion of the left anterior descending coronary artery (LAD), and the dynamic changes were recorded by electrocardiogram at different stages of I/R. Subsequently, reperfusion duration was used as a variable to directly compare the phenotypes of different myocardial injury degrees induced by 3 h, 6 h and 24 h reperfusion from myocardial infarct size, myocardial apoptosis, myocardial enzyme, and inflammatory cytokine levels. RESULTS: All mice subjected to myocardial I/R surgery showed obvious myocardial infarction, extensive myocardial apoptosis, dynamic changes in serum myocardial enzyme and inflammatory cytokines, at least for the first 24 h of reperfusion. The infarct size and apoptosis rates gradually increased with the extension of reperfusion time. The peaks of serum myocardial enzyme and inflammatory cytokines occurred at 6 h and 3 h of reperfusion, respectively. We also established I/R mice models with 30 and 60 mins of ischemia. After 21 days of remodeling, longer periods of ischemia increased the degree of fibrosis and reduced cardiac function. CONCLUSIONS: In summary, we conclude that reperfusion durations of 3 h, 6 h, and 24 h induces different injury phenotypes in ischemia-reperfusion mouse model. At the same time, the ischemia duration before reperfusion also affects the degree of cardiac remodeling.

Loss of Histone Methyltransferase KMT2D Attenuates Angiogenesis in the Ischemic Heart by Inhibiting the Transcriptional Activation of VEGF-A.

Angiogenesis occurred after myocardial infarction (MI) protects heart failure (HF). The aim of our study was to explore function of histone methyltransferase KMT2D (MLL4, mixed-lineage leukemia 4) in angiogenesis post-MI. Western blotting showed that KMT2D protein expression was elevated in MI mouse myocardial. Cardiomyocyte-specific Kmt2d-knockout (Kmt2d-cKO) mice were generated, and echocardiography and immunofluorescence staining detected significantly attenuated cardiac function and insufficient angiogenesis following MI in Kmt2d-cKO mice. Cross-talk assay suggested that Kmt2d-KO H9c2-derived conditioned medium attenuates EA.hy926 EC function. ELISA further identified that VEGF-A released from Kmt2d-KO H9c2 was significantly reduced. CUT&Tag and RT-qPCR revealed that KMT2D deficiency reduced Vegf-a mRNA expression and enrichment of H3K4me1 on the Vegf-a promoter. Moreover, KMT2D silencing in ECs also suppressed endothelial function. Our study indicates that KMT2D depletion in both cardiomyocytes and ECs attenuates angiogenesis and that loss of KMT2D exacerbates heart failure after MI in mice.

Copper promotes cardiac functional recovery via suppressing the transformation of fibroblasts to myofibroblasts in ischemia-infarcted monkey hearts.

Myocardial ischemia leads to cardiac fibrosis along with copper (Cu) loss. Cu repletion diminishes myocardial fibrosis and improves cardiac function. The transformation of fibroblasts to myofibroblasts is highly responsible for the pathogenesis of cardiac fibrosis. This study was undertaken to test the hypothesis that Cu inhibition of cardiac fibrosis results from suppression of myofibroblasts. Rhesus monkeys 4-5 years old were subjected to coronary artery ligation to induce myocardial infarction (MI). At the end of the fourth week after the surgery, an ultrasound-directed Cu-albumin microbubble organ-specific Cu delivery technique was used to treat the ischemia-infarcted monkey hearts twice a week for 4 weeks. This treatment increased Cu concentrations in the infarct area, loosened the collagen cross-linking network, restored blood vessel density, and improved cardiac contractility. Total fibroblasts labeled with vimentin were increased in the infarct area, and Cu repletion did not alter this increase. Myofibroblasts, dually labeled with vimentin and α-smooth muscle actin (α-SMA), were also significantly increased in the infarct area but were significantly reduced by Cu repletion. Correspondingly, the products of myofibroblasts, type I and III collagens and inhibitors of collagenases were significantly reduced. In contrast, metalloproteinase-1 (MMP-1) and MMP-1 producing fibroblasts (vimentin+ and MMP-1+ cells) were significantly increased. These results suggest that Cu inhibits the transformation of fibroblasts to myofibroblasts, leading to a pro-fibrinolytic switch and an improvement in cardiac function.

Immunosuppressants or corticosteroids compared with supportive therapy: a systematic review and meta-analysis on the efficacy and safety for IgA nephropathy treatment.

Background: Corticosteroids or immunosuppressants and supportive treatment in reducing the risk of proteinuria and end-stage kidney disease (ESKD) in immunoglobulin A (IgA) nephropathy (IgAN) patients were still controversial. The purpose of this meta-analysis was to evaluate the efficacy and safety of immunosuppressants or corticosteroids compared with supportive therapy for treatment of IgAN in order to provide guidance for clinical practice. Methods: We conducted an online search in PubMed, Embase, Cochrane Library, and Web of Science databases to collect randomized control trials (RCTs) about the efficacy and safety of immunosuppressants or corticosteroids compared with supportive therapy for treatment of IgA for relevant literature published from the databases' inception to August 21, 2021. The Cochrane risk assessment tool was used to assess the risk of bias in the included studies and analyzed by Revman 5.4 software, and Stata 15.0 statistical software was adopted for meta-analysis. Results: A total of 10,622 related studies were retrieved, and 11 RCTs were finally included in the meta-analysis, with a total sample size of 809 cases. The primary outcome measures for immunosuppressants or corticosteroids were better than those for supportive therapy: proteinuria [weighted mean difference (WMD) =-0.54, 95% confidence interval (CI): -0.63, -0.44, Z =10.79, P<0.001] and ESKD [relative risk (RR) =0.189, 95% CI: 0.059, 0.605, Z =2.81, P=0.005]. The secondary outcome measures were also better than that for supportive treatment: glomerular filtration rate [standardized mean difference (SMD) =0.32, 95% CI: 0.09, 0.54, Z =2.48, P=0.013]. The incidence of adverse reactions was consistent with that of supportive treatment, and the difference was not statistically significant (RR =1.06, 95% CI: 0.71, 1.59, Z =0.28, P=0.777). Discussion: Current evidence shows that immunosuppressants and corticosteroids can significantly reduce the risk of proteinuria and ESKD in IgAN patients. Due to limited quality and quantity of the included studies, more high-quality studies are need to verify above conclusion. In addition, we hope that more rationally designed multicenter RCTs that are not limited to short-term treatment outcomes will be conducted in the future.

Ultrasonographic study of hemodynamics and contrast-enhanced ultrasound in the rhesus monkey kidney.

Nonhuman primates share many developmental similarities with humans. As the world has recognized the rhesus monkey as a standard experimental monkey, studies of rhesus monkey are very important and essential. The purpose of this study was to use gray-scale ultrasound, color Doppler flow imaging (CDFI), and contrast-enhanced ultrasound (CEUS) to study the ultrasound appearance of adult healthy rhesus monkey kidneys and to investigate the relationship between renal ultrasound manifestations and body weight, gender, and the left and right kidneys. Thirty adult healthy rhesus monkeys were studied in the experiments. The size of the kidney and the length and diameter of the renal artery were measured. The peak systolic velocity (PSV), end diastolic velocity (EDV), and resistance index (RI) of the renal artery and intrarenal arteries were measured by CDFI. In CEUS, the time-intensity curve (TIC) was used to obtain microvascular perfusion parameters. There were significant differences in renal size, diameter and length of the renal artery, and hemodynamics of the renal arteries between the different weight groups. In CEUS, there were significant differences in area under curve (AUC), time from peak to one half (THP), intensity peak (PI), time to peak (TTP), mean transit time (MTT), and wash-in-slope (WIS) between the different weight groups. There were no statistical differences between genders or the left and right kidneys. Our study provides valuable reference data for the studies of the kidney and indicates that CEUS can be used to evaluate renal perfusion in rhesus monkeys.

Atherosclerotic lesion-specific copper delivery suppresses atherosclerosis in high-cholesterol-fed rabbits.

Dietary cholesterol supplements cause hypercholesterolemia and atherosclerosis along with a reduction of copper concentrations in the atherosclerotic wall in animal models. This study was to determine if target-specific copper delivery to the copper-deficient atherosclerotic wall can block the pathogenesis of atherosclerosis. Male New Zealand white rabbits, 10-weeks-old and averaged 2.0 kg, were fed a diet containing 1% (w/w) cholesterol or the same diet without cholesterol as control. Twelve weeks after the feeding, the animals were injected with copper-albumin microbubbles and subjected to ultrasound sonication specifically directed at the atherosclerotic lesions (Cu-MB-US) for target-specific copper delivery, twice a week for four weeks. This regiment was repeated 3 times with a gap of two weeks in between. Two weeks after the last treatment, the animals were harvested for analyses of serum and aortic pathological changes. Compared to controls, rabbits fed cholesterol-rich diet developed atherosclerotic lesion with a reduction in copper concentrations in the lesion tissue. Cu-MB-US treatment significantly increased copper concentrations in the lesion, and reduced the size of the lesion. Furthermore, copper repletion reduced the number of apoptotic cells as well as the content of cholesterol and phospholipids in the atherosclerotic lesion without a disturbance of the stability of the lesion. The results thus demonstrate that target-specific copper supplementation suppresses the progression of atherosclerosis at least in part through preventing endothelial cell death, thus reducing lipid infiltration in the atherosclerotic lesion.

Copper Preserves Vasculature Structure and Function by Protecting Endothelial Cells from Apoptosis in Ischemic Myocardium.

The present study was undertaken to investigate whether Cu protects vasculatures from ischemic injury in the heart. C57/B6 mice were introduced to myocardial ischemia (MI) by permanent ligation of the left anterior descending (LAD) coronary artery. Two hours post-LAD ligation, mice were intravenously injected with a Cu-albumin (Cu-alb) solution, or saline as control. At 1, 4, or 7 days post-MI, hearts were collected for further analysis. A dramatic decrease in CD31-positive endothelial cells concomitantly with abundant apoptosis, along with obstruction of blood flow, was observed in ischemic myocardium 1 day post-MI. The early Cu-alb treatment protected CD31-positive cells from apoptosis, along with a preservation of micro-vessels and a decrease in infarct size. This early vasculature preservation ensured myocardial blood perfusion and protected cardiac contractile function until 28 days post-MI. This strategy of Cu-alb treatment immediately following MI would help develop a therapeutic approach for acute heart attack patients in a clinical setting.