Cardiac hypertrophy is initiated as an adaptive response to sustained overload but progresses pathologically as heart failure ensues. Here we report that genetic loss of APJ, a G-protein-coupled ...receptor, confers resistance to chronic pressure overload by markedly reducing myocardial hypertrophy and heart failure. In contrast, mice lacking apelin (the endogenous APJ ligand) remain sensitive, suggesting an apelin-independent function of APJ. Freshly isolated APJ-null cardiomyocytes exhibit an attenuated response to stretch, indicating that APJ is a mechanosensor. Activation of APJ by stretch increases cardiomyocyte cell size and induces molecular markers of hypertrophy. Whereas apelin stimulates APJ to activate Gαi and elicits a protective response, stretch signals in an APJ-dependent, G-protein-independent fashion to induce hypertrophy. Stretch-mediated hypertrophy is prevented by knockdown of β-arrestins or by pharmacological doses of apelin acting through Gαi. Taken together, our data indicate that APJ is a bifunctional receptor for both mechanical stretch and the endogenous peptide apelin. By sensing the balance between these stimuli, APJ occupies a pivotal point linking sustained overload to cardiomyocyte hypertrophy.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, KISLJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The proteasome is a vital cellular machine that maintains protein homeostasis, which is of particular importance in multiple myeloma and possibly other cancers. Targeting of proteasome 20S peptidase ...activity with bortezomib and carfilzomib has been widely used to treat myeloma. However, not all patients respond to these compounds, and those who do eventually suffer relapse. Therefore, there is an urgent and unmet need to develop new drugs that target proteostasis through different mechanisms. We identified quinoline-8-thiol (8TQ) as a first-in-class inhibitor of the proteasome 19S subunit Rpn11. A derivative of 8TQ, capzimin, shows >5-fold selectivity for Rpn11 over the related JAMM proteases and >2 logs selectivity over several other metalloenzymes. Capzimin stabilized proteasome substrates, induced an unfolded protein response, and blocked proliferation of cancer cells, including those resistant to bortezomib. Proteomic analysis revealed that capzimin stabilized a subset of polyubiquitinated substrates. Identification of capzimin offers an alternative path to develop proteasome inhibitors for cancer therapy.
BACKGROUND—Long-term inhibition of nitric oxide synthase by L-arginine analogues such as N-nitro-L-arginine methyl ester (L-NAME) has been shown to induce senescence in vitro and systemic ...hypertension and arteriosclerosis in vivo. We previously reported that plasminogen activator inhibitor-1 (PAI-1)–deficient mice (PAI-1) are protected against L-NAME-induced pathologies. In this study, we investigated whether a novel, orally active PAI-1 antagonist (TM5441) has a similar protective effect against L-NAME treatment. Additionally, we studied whether L-NAME can induce vascular senescence in vivo and investigated the role of PAI-1 in this process.
METHODS AND RESULTS—Wild-type mice received either L-NAME or L-NAME and TM5441 for 8 weeks. Systolic blood pressure was measured every 2 weeks. We found that TM5441 attenuated the development of hypertension and cardiac hypertrophy compared with animals that had received L-NAME alone. Additionally, TM5441-treated mice had a 34% reduction in periaortic fibrosis relative to animals on L-NAME alone. Finally, we investigated the development of vascular senescence by measuring p16 expression and telomere length in aortic tissue. We found that L-NAME increased p16 expression levels and decreased telomere length, both of which were prevented with TM5441 cotreatment.
CONCLUSIONS—Pharmacological inhibition of PAI-1 is protective against the development of hypertension, cardiac hypertrophy, and periaortic fibrosis in mice treated with L-NAME. Furthermore, PAI-1 inhibition attenuates the arterial expression of p16 and maintains telomere length. PAI-1 appears to play a pivotal role in vascular senescence, and these findings suggest that PAI-1 antagonists may provide a novel approach in preventing vascular aging and hypertension.
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NAD+ is a crucial cellular factor that plays multifaceted roles in wide ranging biological processes. Low levels of NAD+ have been linked to numerous diseases including metabolic ...disorders, cardiovascular disease, neurodegeneration, and muscle wasting disorders. A novel strategy to boost NAD+ is to activate nicotinamide phosphoribosyltransferase (NAMPT), the putative rate-limiting step in the NAD+ salvage pathway. We previously showed that NAMPT activators increase NAD+ levels in vitro and in vivo. Herein we describe the optimization of our NAMPT activator prototype (SBI-0797812) leading to the identification of 1-(4-((4-chlorophenyl)sulfonyl)phenyl)-3-(oxazol-5-ylmethyl)urea (34) that showed far more potent NAMPT activation and improved oral bioavailability.
OBJECTIVETo test the hypothesis that apelin protects against angiotensin II (Ang II)-induced cardiovascular fibrosis and vascular remodeling.
METHODS AND RESULTSWild-type mice administered apelin or ...apelin along with Ang II exhibited less cardiovascular fibrosis and decreased plasminogen activator inhibitor type-1 (PAI-1) gene expression than mice receiving Ang II, N-nitro-L-arginine methyl ester (L-NAME), apelin plus L-NAME or apelin plus Ang II plus L-NAME. In-vitro analysis using a luciferase construct driven by 3.1 kb of the human PAI-1 promoter revealed that apelin blocked Ang II-mediated PAI-1 gene expression. Immunoblotting for phosphorylated myosin phosphatase subunit and myosin light chain revealed that apelin blocked Ang II activation of the Rho kinase pathway, which is associated with induction of PAI-1 gene expression by Ang II. In addition, treatment of human aortic smooth muscle cells with apelin reduced PAI-1 mRNA and protein production in the presence and absence of Ang II. Conversely, L-NAME treatment attenuated the downregulation of PAI-1 by apelin in cells.
CONCLUSIONApelin protects against cardiac fibrosis and vascular remodeling through direct regulation of PAI-1 gene expression. This protective effect is mediated through the synergistic inhibition of Ang II signaling and increased production of nitric oxide by apelin. Our data extend previous findings and provide new insight into the molecular mechanisms by which apelin elicits a cardioprotective effect.
Aging drives the occurrence of numerous diseases, including cardiovascular disease (CVD). Recent studies indicate that blood from young mice reduces age-associated pathologies. However, the ...“anti-aging” factors in juvenile circulation remain poorly identified. Here, we characterize the role of the apelinergic axis in mammalian aging and identify apelin as an anti-aging factor. The expression of apelin (apln) and its receptor (aplnr) exhibits an age-dependent decline in multiple organs. Reduced apln signaling perturbs organismal homeostasis; mice harboring genetic deficiency of aplnr or apln exhibit enhanced cardiovascular, renal, and reproductive aging. Genetic or pharmacological abrogation of apln signaling also induces cellular senescence mediated, in part, by the activation of senescence-promoting transcription factors. Conversely, restoration of apln in 15-month-old wild-type mice reduces cardiac hypertrophy and exercise-induced hypertensive response. Additionally, apln-restored mice exhibit enhanced vigor and rejuvenated behavioral and circadian phenotypes. Hence, a declining apelinergic axis promotes aging, whereas its restoration extends the murine healthspan.
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•Senescence downregulates the endogenous apelinergic axis•Aplnr−/− and apln−/− mice and aplnkd cells exhibit accelerated senescence•Apln reduces cardiac hypertrophy and exercise-induced BP in 15-month-old mice
Rai et al. identify an anti-aging role of the endogenous apelinergic axis. They show that the apelin-apelin receptor axis is downregulated with age and that its absence accelerates the onset and progression of aging. Additionally, restoration of apelin extends the murine healthspan.
Fractalkine (FKN) is a membrane‐bound chemokine that can be cleaved by proteases such as ADAM 10, ADAM 17, and cathepsin S to generate soluble fragments. Studies using different forms of the soluble ...FKN yield conflicting results in vivo. These observations prompted us to investigate the function and pharmacology of two commonly used isoforms of FKN, a human full‐length soluble FKN (sFKN), and a human chemokine domain only FKN (cdFKN). Both are prevalent in the literature and are often assumed to be functionally equivalent. We observed that recombinant sFKN and cdFKN exhibit similar potencies in a cell‐based cAMP assay, but binding affinity for CX3CR1 was modestly different. There was a 10‐fold difference in potency between sFKN and cdFKN when assessing their ability to stimulate β‐arrestin recruitment. Interestingly, high concentrations of FKN, regardless of cleavage variant, were ineffective at reducing pro‐inflammatory microglial activation and may induce a pro‐inflammatory response. This effect was observed in mouse and rat primary microglial cells as well as microglial cell lines. The inflammatory response was exacerbated in aged microglia, which is known to exhibit age‐related inflammatory phenotypes. We observed the same effects in Cx3cr1−/− primary microglia and therefore speculate that an alternative FKN receptor may exist. Collectively, these data provide greater insights into the function and pharmacology of these common FKN reagents, which may clarify conflicting reports and urge greater caution in the selection of FKN peptides for use in in vitro and in vivo studies and the interpretation of results obtained using these differing peptides.
Main Points
Soluble CX3CL1 can act in an anti‐inflammatory and pro‐inflammatory fashion.
A portion of this activity is maintained in CX3CR1 knock‐out mouse microglia, thus there is a novel receptor for fractalkine that regulates the proinflammatory response.
Although anti-vascular endothelial growth factor (VEGF) treatments reduce pathological neovascularization in the eye and in tumors, the regression is often not sustainable or is incomplete. We ...investigated whether vascular endothelial cells circumvent anti-VEGF therapies by activating the unfolded protein response (UPR) to override the classic extracellular VEGF pathway. Exposure of endothelial cells to VEGF, high glucose, or H2 O2 up-regulated the X-box binding protein-1/inositol-requiring protein-1 (IRE1) α and activating transcription factor 6 (ATF6) arms of the UPR compared with untreated cells. This was associated with increased expression in α-basic crystallin (CRYAB), which has previously bound VEGF. siRNA knockdown or pharmacological blockade of IRE1α, ATF6, or CRYAB increased intracellular VEGF degradation and decreased full-length intracellular VEGF. Inhibition of IRE1α, ATF6, or CRYAB resulted in an approximately 40% reduction of in vitro angiogenesis, which was further reduced in combination with a neutralizing antibody against extracellular VEGF. Blockade of IRE1α or ATF6 in the oxygen-induced retinopathy or choroidal neovascularization mouse models caused an approximately 35% reduction in angiogenesis. However, combination therapy of VEGF neutralizing antibody with UPR inhibitors or siRNAs reduced retinal/choroidal neovascularization by a further 25% to 40%, and this inhibition was significantly greater than either treatment alone. In conclusion, activation of the UPR sustains angiogenesis by preventing degradation of intracellular VEGF. The IRE1α/ATF6 arms of the UPR offer a potential therapeutic target in the treatment of pathological angiogenesis.
Neovascularization is the pathological driver of blinding eye diseases such as retinopathy of prematurity, proliferative diabetic retinopathy, and wet age-related macular degeneration. The loss of ...vision resulting from these diseases significantly impacts the productivity and quality of life of patients, and represents a substantial burden on the health care system. Current standard of care includes biologics that target vascular endothelial growth factor (VEGF), a key mediator of neovascularization. While anti-VGEF therapies have been successful, up to 30% of patients are non-responsive. Therefore, there is a need for new therapeutic targets, and small molecule inhibitors of angiogenesis to complement existing treatments. Apelin and its receptor have recently been shown to play a key role in both developmental and pathological angiogenesis in the eye. Through a cell-based high-throughput screen, we identified 4-aminoquinoline antimalarial drugs as potent selective antagonists of APJ. The prototypical 4-aminoquinoline, amodiaquine was found to be a selective, non-competitive APJ antagonist that inhibited apelin signaling in a concentration-dependent manner. Additionally, amodiaquine suppressed both apelin-and VGEF-induced endothelial tube formation. Intravitreal amodaiquine significantly reduced choroidal neovascularization (CNV) lesion volume in the laser-induced CNV mouse model, and showed no signs of ocular toxicity at the highest doses tested. This work firmly establishes APJ as a novel, chemically tractable therapeutic target for the treatment of ocular neovascularization, and that amodiaquine is a potential candidate for repurposing and further toxicological, and pharmacokinetic evaluation in the clinic.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Neurotensin receptor 1 (NTR1) is a G protein coupled receptor that is widely expressed throughout the central nervous system where it acts as a neuromodulator. Neurotensin receptors have been ...implicated in a wide variety of CNS disorders, but despite extensive efforts to develop small molecule ligands there are few reports of such compounds. Herein we describe the optimization of a quinazoline based lead to give 18 (SBI-553), a potent and brain penetrant NTR1 allosteric modulator.
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