GLP-1 receptor (GLP-1R) agonists may improve endothelial function (EF) via metabolic improvement and direct vascular action. The current study determined the effect of GLP-1R agonist exenatide on postprandial EF in type 2 diabetes and the mechanisms underlying GLP-1R agonist–mediated vasodilation. Two crossover studies were conducted: 36 participants with type 2 diabetes received subcutaneous exenatide or placebo for 11 days and EF, and glucose and lipid responses to breakfast and lunch were determined; and 32 participants with impaired glucose tolerance (IGT) or diet-controlled type 2 diabetes had EF measured before and after intravenous exenatide, with or without the GLP-1R antagonist exendin-9. Mechanisms of GLP-1R agonist action were studied ex vivo on human subcutaneous adipose tissue arterioles and endothelial cells. Subcutaneous exenatide increased postprandial EF independent of reductions in plasma glucose and triglycerides. Intravenous exenatide increased fasting EF, and exendin-9 abolished this effect. Exenatide elicited eNOS activation and NO production in endothelial cells, and induced dose-dependent vasorelaxation and reduced high-glucose or lipid-induced endothelial dysfunction in arterioles ex vivo. These effects were reduced with AMPK inhibition. In conclusion, exenatide augmented postprandial EF in subjects with diabetes and prevented high-glucose and lipid-induced endothelial dysfunction in human arterioles. These effects were largely direct, via GLP-1R and AMPK activation.

The hexanucleotide repeat expansion GGGGCC (G 4 C 2) n in the C9orf72 gene is the most common genetic abnormality associated with amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Recent findings suggest that dysfunction of nuclear-cytoplasmic trafficking could affect the transport of RNA binding proteins in C9orf72 ALS/FTD. Here, we provide evidence that the RNA editing enzyme Adenosine Deaminase Acting on RNA 2 (ADAR2) is mislocalized in C9orf72 repeat expansion mediated ALS/FTD. ADAR2 is responsible for Adenosine (A) to Inosine (I) editing of double stranded RNA, and its function has been shown to be essential for survival. Here we show the mislocalization of ADAR2 in human induced pluripotent stem cellderived motor neurons (hiPSC-MNs) from C9orf72 patients, in mice expressing (G 4 C 2) 149 , and in C9orf72 ALS/FTD patient postmortem tissue. As a consequence of this mislocalization we observe alterations in RNA editing in our model systems and across multiple brain regions. Analysis of editing at 408,580 known RNA editing sites indicates that there are vast RNA A to I editing aberrations in C9orf72-mediated ALS/FTD. These RNA editing aberrations are found in many cellular pathways, such as the ALS pathway and the crucial EIF2 signaling pathway. Our findings suggest that the mislocalization of ADAR2 in C9orf72 mediated ALS/FTD is responsible

We tested whether nanoliposomes containing phosphatidylcholine, cholesterol and phosphatidic acid (NLPA) prevent bamyloid 1-42 (Ab42) fibrillation and Ab42-induced human arteriole endothelial dysfunction. NLPA abolished Ab42 fibril formation (thioflavin-T fluorescence/electron microscopy). In ex-vivo human adipose and leptomeningeal arterioles, Ab42 impaired dilator response to acetylcholine that was reversed by NLPA; this protection was abolished by L-NGnitroarginine methyl ester. Ab42 reduced human umbilical vein endothelial cell NO production that was restored by NLPA. Nanoliposomes prevented Ab42 amyloid formation, reversed Ab42-induced human microvascular endothelial dysfunction and may be useful in Alzheimer's disease.

Cardiovascular (CV) disease is the leading cause of mortality and morbidity in patients with type 2 diabetes mellitus (T2DM). However, improving glycaemic control alone has not decreased CV events. Therapies that improve glycaemic control, CV disease risk factors and CV function are more likely to be successful. Dipeptidyl peptidase-4 (DPP-4) inhibitors prevent breakdown of incretin hormones glucagon-like peptide-1(GLP-1) and glucose-dependent insulinotropic peptide and improve glycaemic control in patients with T2DM. DPP-4 acts on other substrates, many associated with cardioprotection. Thus, inhibition of DPP-4 may lead to elevations in these potentially beneficial substrates. Data from animal studies and small observational studies in humans suggest that DPP-4 inhibitors may potentially reduce CV risk. However, recently completed CV outcome trials in patients with T2DM and CV disease or at high risk of adverse CV events have shown that the DPP-4 inhibitors saxagliptin and alogliptin neither increased nor decreased major adverse CV events.

BackgroundLight chain amyloidosis (AL) is associated with high mortality, especially in patients with advanced cardiovascular involvement. It is caused by toxicity of misfolded light chain proteins (LC) in vascular, cardiac, and other tissues. There is no treatment to reverse LC tissue toxicity. We tested the hypothesis that nanoliposomes composed of monosialoganglioside, phosphatidylcholine, and cholesterol (GM1 ganglioside–containing nanoliposomes [NLGM1]) can protect against LC‐induced human microvascular dysfunction and assess mechanisms behind the protective effect.Methods and ResultsThe dilator responses of ex vivo abdominal adipose arterioles from human participants without AL to acetylcholine and papaverine were measured before and after exposure to LC (20 μg/mL) with or without NLGM1 (1:10 ratio for LC:NLGM1 mass). Human umbilical vein endothelial cells were exposed for 18 to 20 hours to vehicle, LC with or without NLGM1, or NLGM1 and compared for oxidative and nitrative stress response and cellular viability. LC impaired arteriole dilator response to acetylcholine, which was restored by co‐treatment with NLGM1. LC decreased endothelial cell nitric oxide production and cell viability while increasing superoxide and peroxynitrite; these adverse effects were reversed by NLGM1. NLGM1 increased endothelial cell protein expression of antioxidant enzymes heme oxygenase 1 and NAD(P)H quinone dehydrogenase 1 and increased nuclear factor, erythroid 2 like 2 (Nrf‐2) protein. Nrf‐2 gene knockdown reduced antioxidant stress response and reversed the protective effects of NLGM1.ConclusionsNLGM1 protects against LC‐induced human microvascular endothelial dysfunction through increased nitric oxide bioavailability and reduced oxidative and nitrative stress mediated by Nrf‐2–dependent antioxidant stress response. These findings point to a potential novel therapeutic approach for light chain amyloidosis.

Background Evidence point to vascular dysfunction and hypoperfusion as early abnormalities in Alzheimer's disease (AD); probing their mechanistic bases can lead to new therapeutic approaches. We tested the hypotheses that β-amyloid peptide induces endothelial dysfunction and oxidative stress in human microvasculature and that response will be similar between peripheral adipose and brain leptomeningeal arterioles. New Method Abdominal subcutaneous arterioles from living human subjects (n=17) and cadaver leptomeningeal arterioles (n=6) from rapid autopsy were exposed to Aβ1-42 (Aβ) for 1-hour and dilation response to acetylcholine/papaverine were measured and compared to baseline response. Adipose arteriole reactive oxygen species (ROS) production and nitrotyrosine content were measured. Comparison with existing methods Methods described allow direct investigation of human microvessel functional response that cannot be replicated by human noninvasive imaging or post-mortem histology. Results Adipose arterioles exposed to 2 μM Aβ showed impaired dilation to acetylcholine that was reversed by antioxidant polyethylene glycol superoxide dismutase (PEG-SOD) (Aβ-60.9±6%, control-93.2±1.8%, Aβ+PEGSOD-84.7±3.9%, both p<0.05 vs. Aβ). Aβ caused reduced dilation to papaverine. Aβ increased adipose arteriole ROS production and increased arteriole nitrotyrosine content. Leptomeningeal arterioles showed similar impaired response to acetylcholine when exposed to Aβ (43.0±6.2% versus 81.1±5.7% control, p<0.05). Conclusion Aβ exposure induced adipose arteriole endothelial and non-endothelial dysfunction and oxidative stress that were reversed by antioxidant treatment. Aβ-induced endothelial dysfunction was similar between peripheral adipose and leptomeningeal arterioles. Ex vivo living adipose and cadaver leptomeningeal arterioles are viable, novel and practical human tissue models to study Alzheimer's vascular pathophysiology.

Context A newly-recognized pathogenic mechanism underlying light chain amyloidosis (AL) involves endothelial dysfunction and cell injury caused by misfolded light chain proteins (LC). Nanoliposomes (NL) are artificial phospholipid vesicles that could attach to misfolded proteins and reduce tissue injury. Objective To test whether co-treatment with NL reduce LC-induced endothelial dysfunction and cell death. Methods Abdominal subcutaneous adipose arterioles from 14 non-AL subjects were cannulated; dilator response to acetylcholine and papaverine were measured at baseline and following 1-hour exposure to LC (20 μg/mL, 2 purified from AL subjects’ urine, 1 from human recombinant LC [AL-09]) ± NL (phosphatidylcholine/cholesterol/phosphatidic acid 70/25/5 molar ratio) or NL alone. Human aortic artery endothelial cells (HAEC) were exposed to Oregon Green-labeled LC±NL for 24 hours and intracellular LC and apoptosis (Hoechst stain) were measured. Circular dichroism spectroscopy was performed on AL-09 LC±NL to follow changes in secondary structure and protein thermal stability. Results LC caused impaired dilation to acetylcholine that was restored by NL (control-94.0±1.8%, LC-65.0±7.1%, LC+NL-95.3±1.8%, p≤0.001 LC vs. control or LC+NL). NL protection was inhibited by L-NG-nitroarginine methyl ester. NL increased the beta sheet structure of LC, reduced endothelial cell internalization of LC and protected against LC-induced endothelial cell death. Conclusions LC induced human adipose arteriole endothelial dysfunction and endothelial cell death, which were reversed by co-treatment with NL. This protection may partly be due to enhancing LC protein structure and reducing LC internalization. Nanoliposomes represent a promising new class of agents to ameliorate tissue injury from protein misfolding diseases such as AL.

Misfolded immunoglobulin light chain proteins (LC) in light chain amyloidosis (AL) are toxic to vascular tissues. We tested the hypothesis that chaperone protein clusterin preserves endothelial function and cell survival during LC exposure. Methods LC (20 μg/mL) were given to human aortic endothelial cells (EC) for 24-hours and clusterin protein/gene expression and secretion were measured. DNA fragmentation was measured with/without recombinant clusterin (Clu, 300 ng/mL). Adipose arterioles (non-AL subjects) were tested for dilator responses to acetylcholine/papaverine at baseline and after 1-hour of LC±Clu. Results LC reduced EC clusterin secretion, protein and gene expression while increasing DNA fragmentation. Clu attenuated LC-induced DNA fragmentation and restored dilator response to acetylcholine (logEC50: control −7.05±0.2, LC+Clu −6.53±0.4, LC −4.28±0.7, p<0.05 vs. control, LC+Clu). Conclusions LC induced endothelial cell death and dysfunction while reducing clusterin protein/gene expression and secretion. Exogenous clusterin attenuated LC toxicity. This represents a new pathobiologic mechanism and therapeutic target for AL amyloidosis.