scholarly journals siRNA nanoparticles targeting CaMKIIγ in lesional macrophages improve atherosclerotic plaque stability in mice

2020 ◽  
Vol 12 (553) ◽  
pp. eaay1063 ◽  
Author(s):  
Wei Tao ◽  
Arif Yurdagul ◽  
Na Kong ◽  
Wenliang Li ◽  
Xiaobo Wang ◽  
...  
Keyword(s):  
Preclinical Model ◽  
Dependent Protein Kinase ◽  
Cardiovascular Research ◽  
Plaque Stability ◽  
Fibrous Cap ◽  
Atherosclerotic Lesions ◽  
Dependent Protein ◽  
Fibrous Cap Thickness ◽  
Interfering Rna ◽  
Human And Mouse

Atherosclerotic lesional macrophages express molecules that promote plaque progression, but lack of mechanisms to therapeutically target these molecules represents a major gap in translational cardiovascular research. Here, we tested the efficacy of a small interfering RNA (siRNA) nanoparticle (NP) platform targeting a plaque-destabilizing macrophage molecule—Ca2+/calmodulin-dependent protein kinase γ (CaMKIIγ). CaMKIIγ becomes activated in advanced human and mouse plaque macrophages and drives plaque necrosis by suppressing the expression of the efferocytosis receptor MerTK. When macrophage-targeted siCamk2g NPs were administered to Western diet–fed Ldlr−/− mice, the atherosclerotic lesions showed decreased CaMKIIγ and increased MerTK expression in macrophages, improved phagocytosis of apoptotic cells (efferocytosis), decreased necrotic core area, and increased fibrous cap thickness—all signs of increased plaque stability—compared with mice treated with control siRNA NPs. These findings demonstrate that atherosclerosis-promoting genes in plaque macrophages can be targeted with siRNA NPs in a preclinical model of advanced atherosclerosis.

Atherosclerotic plaque disruption induced by stress and lipopolysaccharide in apolipoprotein E knockout mice

2009 ◽  
Vol 296 (5) ◽  
pp. H1598-H1606 ◽  
Author(s):  
Mei Ni ◽  
Yan Wang ◽  
Mei Zhang ◽  
Peng Fei Zhang ◽  
Shi Fang Ding ◽  
...  
Keyword(s):  
Apolipoprotein E ◽  
High Fat Diet ◽  
Morphological Analysis ◽  
Fibrous Cap ◽  
Atherosclerotic Lesions ◽  
Plaque Disruption ◽  
Apolipoprotein E Knockout Mice ◽  
Fibrous Cap Thickness ◽  
And Control ◽  
Disruption Rate

To establish an animal model with disruptions of atherosclerotic plaques, 96 male apolipoprotein E knockout (apoE−/−) mice were randomly divided into stress, lipopolysaccharide (LPS), stress+LPS, and control groups ( n = 24 each). All mice were fed a high-fat diet throughout the experiment, and carotid atherosclerotic lesions were induced by placement of a constrictive perivascular collar. Four weeks after surgery, mice in the LPS and stress+LPS groups were intraperitoneally injected with LPS (1 mg/kg twice per week for 8 wk). Eight weeks after surgery, mice in the stress and stress+LPS groups were treated with intermittent physical stress (electric foot shock and noise stimulation) for 4 wk. Morphological analysis revealed a plaque disruption rate of 16.7% in control, 34.8% in LPS, 54.2% in stress, and 60.9% in stress+LPS groups. The disruption rates in stress and stress+LPS groups were both significantly higher than those of controls ( P = 0.007 and P = 0.002, respectively). Luminal thrombosis secondary to plaque disruption was observed only in the stress+LPS group. Both stress and LPS stimulation significantly decreased fibrous cap thickness and increased macrophage and lipid contents in plaques. Moreover, the combination of stress and LPS stimulation further lowered cap thickness and enhanced accumulation of macrophages and expression of inflammatory cytokines and matrix metalloproteinases. Stress activated the sympathetic nervous system, as manifested by increased blood pressure and flow velocity. Plasma fibrinogen levels were remarkably elevated in the stress and stress+LPS groups. In conclusion, stress- and LPS-costimulated apoE−/− mice provide a useful model for studies of plaque vulnerability and interventions.

Abstract 149: Pluripotency Factors Krüppel-Like Factor 4 and Octamer-binding Transcription Factor 4 Alter Indices of Plaque Stability in Long Term Western Diet Fed Mice by Regulating Smooth Muscle Cell Phenotypes

2013 ◽  
Vol 33 (suppl_1) ◽  
Author(s):  
Laura S Shankman ◽  
Olga A Cherepanova ◽  
Delphine Gomez ◽  
Gary K Owens
Keyword(s):  
Smooth Muscle ◽  
Foam Cell ◽  
Lineage Tracing ◽  
Phenotypic Switching ◽  
Marker Genes ◽  
Plaque Stability ◽  
Pluripotency Factors ◽  
Fibrous Cap ◽  
Atherosclerotic Lesions ◽  
Life Threatening

The bulk of life threatening thrombotic events have been associated with disruption of the fibrous cap, an atheroprotective layer of smooth muscle α-actin positive (ACTA2+) cells that form around the plaque, and the presence of a large foam cell-laden necrotic core within the plaque. Despite the overwhelming research demonstrating that ACTA2+ cells are beneficial for plaque stability, and cells positive for macrophage-markers are detrimental, there are major ambiguities regarding the origins of these cells, and their role in lesion stability. To clearly define the contribution of smooth muscle cells (SMCs) within atherosclerotic lesions, we generated SMC specific lineage tracing Apoe-/- mice containing a SM myosin heavy chain ( Myh11 ) tamoxifen-inducible cre-recombinase gene and a floxed STOP ROSA eYFP gene ( Myh11 YFP ApoE-/- mice) thus allowing activation of eYFP exclusively in fully differentiated SMCs before the onset of atherosclerosis and subsequent determination of the fate of these cells and their progeny irrespective of continued expression of MYH11 or other SMC marker genes. Remarkably, our results reveal that 86% of SMCs cannot be identified using traditional SMC markers, such as ACTA2, and 23% of presumed macrophages (LGALS3+ cells) are derived from SMC origins. The last finding was confirmed in human coronary atheromas using the ISH-PLA approach. SMC specific knockout (KO) of the pluripotency factor Klf4 in Myh11 YFP ApoE-/- mice did not alter the frequency of phenotypically modulated (ACTA2-eYFP+) SMCs within atherosclerotic lesions of mice fed a high fat diet for 18 weeks, however, decreased the number of ACTA2-eYFP+ SMCs that expressed LGALS3, and increased several indices of plaque stability, suggesting a detrimental role for KLF4 in SMCs within atherosclerotic lesions. Conversely, SMC specific Oct4 KO resulted in a dramatic reduction in the number of ACTA2-eYFP+ SMCs within the lesion with marked decreases in indices of plaque stability. In summary results show that the majority of SMC-derived cells within advanced atherosclerotic lesions cannot be identified using conventional SMC marker genes, and that phenotypic switching of SMC during atherogenesis is differentially regulated by the pluripotency factors KLF4 and OCT4.

scholarly journals Allosteric MAPKAPK2 inhibitors improve plaque stability in advanced atherosclerosis

PLoS ONE ◽  
2021 ◽  
Vol 16 (5) ◽  
pp. e0246600
Author(s):  
Lale Ozcan ◽  
Canan Kasikara ◽  
Arif Yurdagul ◽  
George Kuriakose ◽  
Brian Hubbard ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Vascular Disease ◽  
Atherosclerotic Plaque ◽  
Plasma Cholesterol ◽  
Mitogen Activated Protein Kinase ◽  
Preclinical Model ◽  
High Fat ◽  
Atherosclerotic Vascular Disease ◽  
Plaque Stability ◽  
Fibrous Cap

Atherosclerotic vascular disease resulting from unstable plaques is the leading cause of morbidity and mortality in subjects with type 2 diabetes (T2D), and thus a major therapeutic goal is to discover T2D drugs that can also promote atherosclerotic plaque stability. Genetic or pharmacologic inhibition of mitogen-activated protein kinase-activated protein kinase-2 (MAPKAPK2 or MK2) in obese mice improves glucose homeostasis and enhances insulin sensitivity. We developed two novel orally active small-molecule inhibitors of MK2, TBX-1 and TBX-2, and tested their effects on metabolism and atherosclerosis in high-fat Western diet (WD)-fed Ldlr-/- mice. Ldlr-/- mice were first fed the WD to allow atherosclerotic lesions to become established, and the mice were then treated with TBX-1 or TBX-2. Both compounds improved glucose metabolism and lowered plasma cholesterol and triglyceride, without an effect on body weight. Most importantly, the compounds decreased lesion area, lessened plaque necrosis, and increased fibrous cap thickness in the aortic root lesions of the mice. Thus, in a preclinical model of high-fat feeding and established atherosclerosis, MK2 inhibitors improved metabolism and also enhanced atherosclerotic plaque stability, suggesting potential for further clinical development to address the epidemic of T2D associated with atherosclerotic vascular disease.

scholarly journals Inositol Trisphosphate Receptors and Nuclear Calcium in Atrial Fibrillation

2020 ◽  
Author(s):  
Xiao Yan Qi ◽  
Faezeh Vahdahi Hassani ◽  
Dennis Hoffmann ◽  
Jiening Xiao ◽  
Feng Xiong ◽  
...  
Keyword(s):  
Atrial Fibrillation ◽  
Inositol Trisphosphate ◽  
Luciferase Assay ◽  
Dependent Protein Kinase ◽  
Expression Ratio ◽  
Inositol Trisphosphate Receptor ◽  
Dependent Protein ◽  
Trisphosphate Receptor ◽  
Interfering Rna

Rationale: The mechanisms underlying atrial fibrillation (AF), the most common clinical arrhythmia, are poorly understood. Nucleoplasmic Ca 2+ regulates gene-expression, but the nature and significance of nuclear Ca 2+ -changes in AF are largely unknown. Objective: To elucidate mechanisms by which AF alters atrial cardiomyocyte (CM) nuclear Ca 2+ ([Ca 2+ ] Nuc ) and Ca 2+ /calmodulin-dependent protein kinase-II (CaMKII)-related signaling. Methods and Results: Atrial CMs were isolated from control and AF-dogs (kept in AF by atrial tachypacing [600 bpm x 1 week]). [Ca 2+ ] Nuc and cytosolic [Ca 2+ ] (Ca 2+ ] Cyto ) were recorded via confocal microscopy. Diastolic [Ca 2+ ] Nuc was greater than [Ca 2+ ] Cyto under control conditions, while resting [Ca 2+ ] Nuc was similar to [Ca 2+ ] Cyto ; both diastolic and resting [Ca 2+ ] Nuc increased with AF. Inositol-trisphosphate-receptor (IP 3 R) stimulation produced larger [Ca 2+ ] Nuc increases in AF versus control CMs, and IP 3 R-blockade suppressed the AF-related [Ca 2+ ] Nuc -differences. AF upregulated nuclear protein-expression of IP 3 R-type 1 (IP 3 R1) and of phosphorylated CaMKII (immunohistochemistry and immunoblot), while decreasing the nuclear/cytosolic expression-ratio for histone deacetylase type-4 (HDAC4). Isolated atrial CMs tachypaced at 3 Hz for 24 hours mimicked AF-type [Ca 2+ ] Nuc changes and L-type calcium current (ICaL) decreases versus 1-Hz-paced CMs; these changes were prevented by IP3R knockdown with short-interfering RNA directed against IP 3 R1. Nuclear/cytosolic HDAC4 expression-ratio was decreased by 3-Hz pacing, while nuclear CaMKII and HDAC4 phosphorylation were increased. Either CaMKII-inhibition (by autocamtide-2-related peptide) or IP 3 R-knockdown prevented the CaMKII-hyperphosphorylation and nuclear-to-cytosolic HDAC4 shift caused by 3-Hz pacing. In human atrial CMs from AF patients, nuclear IP 3 R1-expression was significantly increased, with decreased nuclear/non-nuclear HDAC4 ratio. MicroRNA-26a was predicted to target ITPR1 (confirmed by Luciferase assay) and was downregulated in AF atrial CMs; microRNA-26a silencing reproduced AF-induced IP3R1 upregulation and nuclear diastolic Ca 2+ -loading. Conclusions: AF increases atrial CM nucleoplasmic Ca 2+ -handling by IP 3 R1-upregulation involving miR-26a, leading to enhanced IP 3 R1-CaMKII-HDAC4 signaling and I CaL -downregulation.

scholarly journals Allosteric MAPKAPK2 Inhibitors Improve Plaque Stability in Advanced Atherosclerosis

2021 ◽  
Author(s):  
Lale Ozcan ◽  
Canan Kasikara ◽  
Arif Yurdagul ◽  
George Kuriakose ◽  
Brian Hubbard ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Vascular Disease ◽  
Atherosclerotic Plaque ◽  
Plasma Cholesterol ◽  
Mitogen Activated Protein Kinase ◽  
Preclinical Model ◽  
High Fat ◽  
Atherosclerotic Vascular Disease ◽  
Plaque Stability ◽  
Fibrous Cap

ABSTRACTAtherosclerotic vascular disease resulting from unstable plaques is the leading cause of morbidity and mortality in subjects with type 2 diabetes (T2D), and thus a major therapeutic goal is to discover T2D drugs that can also promote atherosclerotic plaque stability. Genetic or pharmacologic inhibition of mitogen-activated protein kinase-activated protein kinase-2 (MK2) in obese mice improves glucose homeostasis and enhances insulin sensitivity. We developed two novel orally active small-molecule inhibitors of MK2, TBX-1 and TBX-2, and tested their effects on metabolism and atherosclerosis in high-fat Western diet (WD)-fed Ldlr-/- mice. Ldlr-/- mice were first fed the WD to allow atherosclerotic lesions to become established, and the mice were then treated with TBX-1 or TBX-2. Both compounds improved glucose metabolism and lowered plasma cholesterol and triglyceride, without an effect on body weight. Most importantly, the compounds decreased lesion area, lessened plaque necrosis, and increased fibrous cap thickness in the aortic root lesions of the mice. Thus, in a preclinical model of high-fat feeding and established atherosclerosis, MK2 inhibitors improved metabolism and also enhanced atherosclerotic plaque stability, suggesting potential for further clinical development to address the epidemic of T2D associated with atherosclerotic vascular disease.

scholarly journals RNA Interference Screening Reveals Host CaMK4 as a Regulator of Cryptococcal Uptake and Pathogenesis

2017 ◽  
Vol 85 (12) ◽  
Author(s):  
Deepa Srikanta ◽  
Camaron R. Hole ◽  
Matthew Williams ◽  
Shabaana A. Khader ◽  
Tamara L. Doering
Keyword(s):  
Host Cells ◽  
Wild Type ◽  
Dependent Protein Kinase ◽  
Content Type ◽  
Calcium Calmodulin Dependent ◽  
Dependent Protein ◽  
Opportunistic Fungal Pathogen ◽  
High Throughput Imaging ◽  
Interfering Rna ◽  
Host Genes

ABSTRACTCryptococcus neoformans, the causative agent of cryptococcosis, is an opportunistic fungal pathogen that kills over 200,000 individuals annually. This yeast may grow freely in body fluids, but it also flourishes within host cells. Despite extensive research on cryptococcal pathogenesis, host genes involved in the initial engulfment of fungi and subsequent stages of infection are woefully understudied. To address this issue, we combined short interfering RNA silencing and a high-throughput imaging assay to identify host regulators that specifically influence cryptococcal uptake. Of 868 phosphatase and kinase genes assayed, we discovered 79 whose silencing significantly affected cryptococcal engulfment. For 25 of these, the effects were fungus specific, as opposed to general alterations in phagocytosis. Four members of this group significantly and specifically altered cryptococcal uptake; one of them encoded CaMK4, a calcium/calmodulin-dependent protein kinase. Pharmacological inhibition of CaMK4 recapitulated the observed defects in phagocytosis. Furthermore, mice deficient in CaMK4 showed increased survival compared to wild-type mice upon infection withC. neoformans. This increase in survival correlated with decreased expression of pattern recognition receptors on host phagocytes known to recognizeC. neoformans. Altogether, we have identified a kinase that is involved inC. neoformansinternalization by host cells and in host resistance to this deadly infection.

scholarly journals How Critical Is Fibrous Cap Thickness to Carotid Plaque Stability?

Stroke ◽  
2006 ◽  
Vol 37 (5) ◽  
pp. 1195-1199 ◽  
Author(s):  
Zhi-Yong Li ◽  
Simon P.S. Howarth ◽  
Tjun Tang ◽  
Jonathan H. Gillard
Keyword(s):  
Carotid Plaque ◽  
Plaque Stability ◽  
Fibrous Cap ◽  
Fibrous Cap Thickness

scholarly journals Specific NLRP3 Inhibition Protects Against Diabetes-Associated Atherosclerosis

2020 ◽  
Author(s):  
Ada Admin ◽  
Arpeeta Sharma ◽  
Judy S.Y. Choi ◽  
Nada Stefanovic ◽  
Annas-Al Sharea ◽  
...  
Keyword(s):  
Nlrp3 Inflammasome ◽  
Vascular Function ◽  
Vascular Complications ◽  
Fold Increase ◽  
Low Grade ◽  
Murine Bone Marrow ◽  
Fibrous Cap ◽  
Atherosclerotic Lesions ◽  
Persistent Inflammation ◽  
Fibrous Cap Thickness

Low-grade persistent inflammation is a feature of diabetes-driven vascular complications, in particular activation of the NLRP3-inflammasome to trigger the maturation and release of the inflammatory cytokine interleukin-1β (IL-1β). We investigated whether inhibiting the NLRP3-inflammasome, through the use of the specific small-molecule NLRP3 inhibitor, MCC950, could reduce inflammation, improve vascular function and protect against diabetes-associated atherosclerosis in the streptozotocin (STZ)-induced diabetic Apolipoprotein knockout (ApoE<sup>-/-</sup>) mouse. Diabetes led to a ~4-fold increase in atherosclerotic lesions throughout the aorta, which were significantly attenuated with MCC950 (<i>P</i><0.001). This reduction in lesions was associated with decreased monocyte-macrophage content, reduced necrotic core, attenuated inflammatory gene expression (Il-1β, TNFα, ICAM-1, MCP-1, <i>P</i><0.05) and reduced oxidative stress, whilst maintaining fibrous cap thickness. Additionally, vascular function was improved in diabetic vessels of mice treated with MCC950 (<i>P</i><0.05). In a range of cell lines (murine bone marrow-derived macrophages (BMDMs), human monocytic THP-1 cells, PMA-differentiated human macrophages and diabetic human aortic smooth muscle cells (AoSMCs)), MCC950 significantly reduced IL-1β and/or caspase-1 secretion and attenuated leukocyte-SMC interactions under high glucose or LPS conditions. In summary, MCC950 reduces plaque development, promotes plaque stability and improves vascular function, suggesting that targeting NLRP3-mediated inflammation is a novel therapeutic strategy to improve diabetes-associated vascular disease.

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