Hepatic gluconeogenesis is essential for glucose homeostasis and also a therapeutic target for type 2 diabetes, but its mechanism is incompletely understood. Here, we report that Sam68, an ...RNA-binding adaptor protein and Src kinase substrate, is a novel regulator of hepatic gluconeogenesis. Both global and hepatic deletions of Sam68 significantly reduce blood glucose levels and the glucagon-induced expression of gluconeogenic genes. Protein, but not mRNA, levels of CRTC2, a crucial transcriptional regulator of gluconeogenesis, are >50% lower in Sam68-deficient hepatocytes than in wild-type hepatocytes. Sam68 interacts with CRTC2 and reduces CRTC2 ubiquitination. However, truncated mutants of Sam68 that lack the C- (Sam68
) or N-terminal (Sam68
) domains fails to bind CRTC2 or to stabilize CRTC2 protein, respectively, and transgenic Sam68
mice recapitulate the blood-glucose and gluconeogenesis profile of Sam68-deficient mice. Hepatic Sam68 expression is also upregulated in patients with diabetes and in two diabetic mouse models, while hepatocyte-specific Sam68 deficiencies alleviate diabetic hyperglycemia and improves insulin sensitivity in mice. Thus, our results identify a role for Sam68 in hepatic gluconeogenesis, and Sam68 may represent a therapeutic target for diabetes.
Renal tubulointerstitial fibrosis (TIF), characterized by epithelial-to-mesenchymal transition (EMT) of renal tubular epithelial cells, is the typical pathological alteration in diabetic nephropathy. ...Gentiopicroside (GPS), a natural compound with anti-inflammatory activity, has been demonstrated to alleviate glomerulosclerosis, whereas whether GPS inhibits TIF
via
regulating inflammation remains unclear. In this study, diabetic db/db mice and high glucose (HG)-stimulated renal tubular epithelial cells (NRK-52E) were applied to explore the effects and mechanisms of GPS on TIF. The results
in vivo
showed that GPS effectively improves glycolipid metabolism disorder, renal dysfunction, and TIF. In particular, GPS treatment reversed the abnormal expressions of EMT marker proteins including elevated α-smooth muscle actin and vimentin and decreased E-cadherin in the kidney of db/db mice. Moreover, GPS treatment also inhibited protein expressions of angiotensinⅡ type 1 receptor (AT1R) and CK2α and the activation of the NF-κB pathway. Importantly, the aforementioned effects of GPS acted
in vivo
were further observed
in vitro
in HG-stimulated NRK-52E cells, which were independent of its effects on glucose and lipid-lowering activity but were reversed by AT1R over-expression. Together, our results indicate that GPS that directly inhibits the CK2/NF-κB inflammatory signaling pathway
via
AT1R may also contribute to the amelioration of TIF in diabetes.
Berberine (BBR) has been demonstrated to exert cardiovascular protective effects by regulating gut microbiota. However, few studies examine the effect of BBR on the gut microbiota in hypertension. ...This study aims to investigate the role of BBR in regulating microbial alterations and vascular function in hypertension. C57BL/6 J mice were infused with Ang II (0.8 mg/kg/day)
osmotic minipumps and treated with BBR (150 mg/kg/day) or choline (1%) for 4 weeks. Blood pressure was detected by tail-cuff measurement once a week. Abdominal aorta pulse wave velocity (PWV) and endothelium dependent vasodilatation were measured to evaluate vascular function. Vascular remodeling was assessed by histological staining of aortic tissue. The fecal microbiota was profiled using 16S ribosomal DNA (rDNA) sequencing. Plasma trimethylamine (TMA)/trimethylamine-N-oxide (TMAO) and hepatic FMO3 expression were measured. We found that BBR treatment significantly alleviated the elevated blood pressure, vascular dysfunction, and pathological remodeling in Ang II-induced hypertensive mice, while choline treatment aggravated hypertension-related vascular dysfunction. 16S rDNA gene sequencing results showed that BBR treatment altered gut microbiota composition (reduced the Firmicutes/Bacteroidetes (F/B) ratio and increased the abundances of
). Moreover, BBR inhibited FMO3 expression and plasma TMA/TMAO production in hypertensive mice. TMAO treatment increased the apoptosis and oxidative stress of human aortic endothelial cells (HAECs) and aggravated Ang II-induced HAECs dysfunction
. These results indicate that the protective effect of BBR in hypertension might be attributed (at least partially) to the inhibition of TMAO production
regulating the gut microbiota.
Inflammatory activation and immune infiltration play important roles in the pathologic process of heart failure (HF). The current study is designed to investigate the immune infiltration and identify ...related biomarkers in heart failure patients due to ischemic cardiomyopathy.
Expression data of HF due to ischemic cardiomyopathy (CM) samples and non-heart failure (NF) samples were downloaded from gene expression omnibus (GEO) database. Differentially expressed genes (DEGs) between CM and NF samples were identified. Single sample gene set enrichment analysis (ssGSEA) was performed to explore the landscape of immune infiltration. Weighted gene co-expression network analysis (WGCNA) was applied to screen the most relevant module associated with immune infiltration. The diagnostic values of candidate genes were evaluated by receiver operating curves (ROC) curves. The mRNA levels of potential biomarkers in the peripheral blood mononuclear cells (PBMCs) isolated from 10 CM patients and 10 NF patients were analyzed to further assess their diagnostic values.
A total of 224 DEGs were identified between CM and NF samples in GSE5406, which are mainly enriched in the protein processing and extracellular matrix related biological processes and pathways. The result of ssGSEA showed that the abundance of dendritic cells (DC), mast cells, natural killer (NK) CD56dim cells, T cells, T follicular helper cells (Tfh), gammadelta T cells (Tgd) and T helper 2 (Th2) cells were significantly higher, while the infiltration of eosinophils and central memory T cells (Tcm) were lower in CM samples compared to NF ones. Correlation analysis revealed that Calumenin (CALU) and palladin (PALLD) were negatively correlated with the abundance of DC, NK CD56dim cells, T cells, Tfh, Tgd and Th2 cells, but positively correlated with the level of Tcm. More importantly, CALU and PALLD were significantly lower in PBMCs from CM patients compared to NF ones.
Our study revealed that CALU and PALLD are potential biomarkers associated with immune infiltration in heart failure due to ischemic cardiomyopathy.
The conserved long non-coding RNA (lncRNA) myocardial infarction associate transcript (
) was identified for its multiple single-nucleotide polymorphisms that are strongly associated with ...susceptibility to MI, but its role in cardiovascular biology remains elusive. Here we investigated whether
regulates cardiac response to pathological hypertrophic stimuli.
Both an angiotensin II (Ang II) infusion model and a transverse aortic constriction (TAC) model were used in adult WT and
-null knockout (
-KO) mice to induce pathological cardiac hypertrophy. Heart structure and function were evaluated by echocardiography and histological assessments. Gene expression in the heart was evaluated by RNA sequencing (RNA-seq), quantitative real-time RT-PCR (qRT-PCR), and Western blotting. Primary WT and
-KO mouse cardiomyocytes were isolated and used in Ca
transient and contractility measurements.
Continuous Ang II infusion for 4 weeks induced concentric hypertrophy in WT mice, but to a lesser extent in
-KO mice. Surgical TAC for 6 weeks resulted in decreased systolic function and heart failure in WT mice but not in
-KO mice. In both models,
-KO mice displayed reduced heart-weight to tibia-length ratio, cardiomyocyte cross-sectional area, cardiomyocyte apoptosis, and cardiac interstitial fibrosis and a better-preserved capillary density, as compared to WT mice. In addition, Ang II treatment led to significantly reduced mRNA and protein expression of the Ca
cycling genes Sarcoplasmic/endoplasmic reticulum Ca
ATPase 2a (SERCA2a) and ryanodine receptor 2 (RyR2) and a dramatic increase in global RNA splicing events in the left ventricle (LV) of WT mice, and these changes were largely blunted in
-KO mice. Consistently, cardiomyocytes isolated from
-KO mice demonstrated more efficient Ca
cycling and greater contractility.
Ablation of
attenuates pathological hypertrophy and heart failure, in part, by enhancing cardiomyocyte contractility.
The majority of current cardiovascular cell therapy trials use bone marrow progenitor cells (BM PCs) and achieve only modest efficacy; the limited potential of these cells to differentiate into ...endothelial-lineage cells is one of the major barriers to the success of this promising therapy. We have previously reported that the E2F transcription factor 1 (E2F1) is a repressor of revascularization after ischemic injury.
We sought to define the role of E2F1 in the regulation of BM PC function.
Ablation of E2F1 (E2F1 deficient) in mouse BM PCs increases oxidative metabolism and reduces lactate production, resulting in enhanced endothelial differentiation. The metabolic switch in E2F1-deficient BM PCs is mediated by a reduction in the expression of pyruvate dehydrogenase kinase 4 and pyruvate dehydrogenase kinase 2; overexpression of pyruvate dehydrogenase kinase 4 reverses the enhancement of oxidative metabolism and endothelial differentiation. Deletion of E2F1 in the BM increases the amount of PC-derived endothelial cells in the ischemic myocardium, enhances vascular growth, reduces infarct size, and improves cardiac function after myocardial infarction.
Our results suggest a novel mechanism by which E2F1 mediates the metabolic control of BM PC differentiation, and strategies that inhibit E2F1 or enhance oxidative metabolism in BM PCs may improve the effectiveness of cell therapy.
Background:
Lacidipine, a third-generation calcium channel blocker, exerts beneficial effects on the endothelium of hypertensive patients in addition to blood pressure lowering. However, the detailed ...mechanism underlying Lacidipine-related endothelial protection is still elusive.
Methods:
Sixteen spontaneous hypertensive rats (SHRs) were randomly divided into two groups: Lacidipine-treated SHR group and saline-treated control group. Tail systolic blood pressure was monitored for four consecutive weeks. Endothelial cells (ECs) were pretreated with Lacidipine prior to being stimulated with H
2
O
2
, bleomycin, or Lipopolysaccharides (LPS)
in vitro
. Then, cell activity, migration, and senescence were measured by Cell Counting Kit-8 assay, transwell assay, and β-galactosidase staining, respectively. The fluorescent probe 2′, 7′-dichlorofluorescein diacetate (DCFH-DA) was used to assess the intracellular reactive oxygen species (ROS). Related protein expression was detected by Western blotting and immunofluorescence.
Results:
Our data showed that Lacidipine treatment lowered the blood pressure of SHRs accompanied by the elevation of CXCR7 expression and suppression of P38 and CCAAT/enhancer-binding protein beta (C/EBP-β) compared with the control group.
In vitro
experiments further demonstrated that Lacidipine increased the cell viability and function of ECs under oxidative stress, cell senescence, and inflammatory activation
via
the CXCR7/P38/signaling pathway.
Conclusions:
Our results suggested that Lacidipine plays a protective role in EC senescence, oxidative stress, and inflammatory injury through the regulation of CXCR7/P38/C/EBP-β signaling pathway.
Hypertension is characterized by endothelial dysfunction and arterial stiffness, which contribute to the pathogenesis of atherosclerotic cardiovascular diseases. Nicotinamide adenine dinucleotide ...(NAD+) is an indispensable cofactor in all living cells that is involved in fundamental biological processes. However, in hypertensive patients, alterations in NAD+ levels and their relation with blood pressure (BP) elevation and vascular damage have not yet been studied. Here we reported that hypertensive patients exhibited lower NAD+ levels, as detected by high-performance liquid chromatography-mass spectrometry (HPLC-MS), in both peripheral blood mononuclear cells (PBMCs) and aortas, which was parallel to vascular dysfunction. NAD+ boosting therapy with nicotinamide mononucleotide (NMN) supplement reduced BP and ameliorated vascular dysfunction in hypertensive patients (NCT04903210) and AngII-induced hypertensive mice. Upregulation of CD38 in endothelial cells led to endothelial NAD+ exhaustion by reducing NMN bioavailability. Pro-inflammatory macrophages infiltration and increase in IL-1β generation derived from pro-inflammatory macrophages resulted in higher CD38 expression by activating JAK1-STAT1 signaling pathway. CD38 KO, CD38 inhibitors treatment, or adeno-associated virus (AAV)-mediated endothelial CD38 knockdown lowered BP and improved vascular dysfunction in AngII-induced hypertensive mice. The present study demonstrated for the first time that endothelial CD38 activation and subsequently accelerated NAD+ degradation due to enhanced macrophage-derived IL-1β production was responsible for BP elevation and vascular damage in hypertension. NAD+ boosting therapy can be used as a novel therapeutic strategy for the management of hypertensive patients.
Insufficient neovascularization, characterized by poor endothelial cell (EC) growth, contributes to the pathogenesis of ischemic heart disease and limits cardiac tissue preservation and regeneration. ...The E2F family of transcription factors are critical regulators of the genes responsible for cell-cycle progression and growth; however, the specific roles of individual E2Fs in ECs are not well understood. Here we investigated the roles of E2F2 and E2F3 in EC growth, angiogenesis, and their functional impact on myocardial infarction (MI). An endothelial-specific E2F3-deficient mouse strain VE-Cre; E2F3(fl/fl) was generated, and MI was surgically induced in VE-Cre; E2F3(fl/fl) and E2F2-null (E2F2 KO) mice and their wild-type (WT) littermates, VE-Cre; E2F3(+/+) and E2F2 WT, respectively. The cardiac function, infarct size, and vascular density were significantly better in E2F2 KO mice and significantly worse in VE-Cre; E2F3(fl/fl) mice than in their WT littermates. The loss of E2F2 expression was associated with an increase in the proliferation of ECs both in vivo and in vitro, while the loss of E2F3 expression led to declines in EC proliferation. Thus, E2F3 promotes while E2F2 suppresses ischemic cardiac repair through corresponding changes in EC proliferation; and differential targeting of specific E2F members may provide a novel strategy for therapeutic angiogenesis of ischemic heart disease.
Exosomes transport biologically active cargo (e.g., proteins and microRNA) between cells, including many of the paracrine factors that mediate the beneficial effects associated with stem-cell ...therapy. Stem cell derived exosomes, in particular mesenchymal stem cells (MSCs), have been shown previously to largely replicate the therapeutic activity associated with the cells themselves, which suggests that exosomes may be a useful cell-free alternative for the treatment of cardiovascular disorders. However, the mechanisms that govern how exosomes home to damaged cells and tissues or the uptake and distribution of exosomal cargo are poorly characterized, because techniques for distinguishing between exosomal proteins and proteins in the targeted tissues are lacking. Here, we report the development of an
model that enabled the visualization, tracking, and quantification of proteins from systemically administered MSC exosomes. The model uses bioorthogonal chemistry and cell-selective metabolic labeling to incorporate the non-canonical amino acid azidonorleucine (ANL) into the MSC proteome. ANL incorporation is facilitated via expression of a mutant (L274G) methionyl-tRNA-synthetase (MetRS
) and subsequent incubation with ANL-supplemented media; after which ANL can be covalently linked to alkyne-conjugated reagents (e.g., dyes and resins) via click chemistry. Our results demonstrate that when the exosomes produced by ANL-treated, MetRS
-expressing MSCs were systemically administered to mice, the ANL-labeled exosomal proteins could be accurately and reliably identified, isolated, and quantified from a variety of mouse organs, and that myocardial infarction (MI) both increased the abundance of exosomal proteins and redistributed a number of them from the membrane fraction of intact hearts to the cytosol of cells in infarcted hearts. Additionally, we found that Desmoglein-1c is enriched in MSC exosomes and taken up by ischemic myocardium. Collectively, our results indicate that this newly developed bioorthogonal system can provide crucial insights into exosome homing, as well as the uptake and biodistribution of exosomal proteins.
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