Nanoliposomes protect against AL amyloid light chain protein-induced endothelial injury

Abstract

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.

Adipose and leptomeningeal arteriole endothelial dysfunction induced by β-amyloid peptide: A practical human model to study Alzheimer's disease vasculopathy

“…Living subject adipose arterioles were isolated, cannulated and pressurized to 60 mm Hg similar to previous protocol (Franco et al, 2012; Migrino et al, 2011; Truran et al, 2014). Baseline (control) dilator response was measured by videomicrometer after preconstriction with endothelin-1 followed by administration of acetylcholine (for endothelium-dependent dilation, 10 −9 -10 −4 M) and papaverine (for smooth muscle-dependent dilation, 10 −4 M), similar to methods validated by other investigators (Dharmashankar et al, 2012; Kizhakekuttu et al, 2012; Suboc et al, 2013).…”

Nanoliposomes protect against human arteriole endothelial dysfunction induced by β-amyloid peptide

“…Our group previously showed that NLPA altered the properties of AL amyloid light chain proteins by reducing the amount of partially folded states and shifting the equilibrium towards the folded state, 7 and atomic force microscopy imaging demonstrated physicochemical interaction between nanoliposome and light chain proteins (unpublished data). Our current data suggest that NLPA similarly alters the properties of Ab42 leading to inhibition of fibril and amyloid formation, potentially useful as a strategy to reduce amyloid deposition that is the hallmark of advanced and irreversible AD.…”

“…3,12 The dose of NLPA was chosen because this was similar to the dose that restored endothelial function in adipose arterioles exposed to AL amyloid light chain proteins. 7 In additional arterioles, Ab42 and NLPA were co-treated with L-NG-nitroarginine methyl ester (L-NAME, 5 mmol), an inhibitor of nitric oxide (NO) synthase (NOS).…”

“…10 NLPA was prepared according to prior methods. 7 In brief, phosphatidylcholine, cholesterol and phosphatidic acid (Avanti Polar Lipids, Alabaster AL) were mixed at 70:25:5 molar ratios (20 mg lipid/ mL). The lipid mixture was dissolved in chloroform followed by removal of the organic solvent by rotary evaporator.…”

Monosialoganglioside‐Containing Nanoliposomes Restore Endothelial Function Impaired by AL Amyloidosis Light Chain Proteins

“…They may be the ideal type of nanoparticles for disease treatment because their compositions can be varied to produce desired physiological effects, and they can be loaded with therapeutic cargo; in addition to this structural versatility, they have the advantage of being nonimmunogenic and fully biodegradable and have low cytotoxicity . We recently showed that phosphatidic acid–containing nanoliposomes restored endothelial function in microvessels exposed to LC and preserved endothelial cell viability, effects that parallel beneficial protection in human microvessels exposed to β‐amyloid peptides . Monosialogangliosides (GM1 ganglioside) are glycosphingolipids found in plasma membranes and may have a potential advantage over phosphatidic acid in AL because they were found to have antioxidant properties in brain and cardiac tissues and were shown to reduce myocardial ischemia–reperfusion injury, attributes that could be beneficial in reversing LC‐induced oxidative stress and cell injury.…”