Nanostructured Lipid Carriers Deliver Resveratrol, Restoring Attenuated Dilation in Small Coronary Arteries, via the AMPK Pathway.

Nanostructured lipid carriers (NLCs) are an emerging drug delivery platform for improved drug stability and the bioavailability of antihypertensive drugs and vasoprotective nutraceutical compounds, such as resveratrol (RV). The objective of this study was to ascertain NLCs' potential to deliver RV and restore attenuated dilator function, using an ex vivo model of acute hypertension. Trimyristin-triolein NLCs were synthesized and loaded with RV. The uptake of RV-NLCs by human coronary artery endothelial cells (HCAECs) maintained their viability and reduced both mitochondrial and cytosolic superoxide levels. Acute pressure elevation in isolated coronary arteries significantly attenuated endothelial-dependent dilator responses, which were reversed following incubation in RV-NLCs, superoxide dismutase or apocynin (p < 0.0001). RV-NLCs demonstrated a five-fold increase in potency in comparison to RV solution. At elevated pressure, in the presence of RV-NLCs, incubation with Nω-nitro-l-arginine (L-NNA) or indomethacin resulted in a significant reduction in the restored dilator component (p < 0.0001), whereas apamin and TRAM-34 had no overall effect. Incubation with the adenosine monophosphate-activated protein kinase (AMPK) inhibitor dorsomorphin significantly attenuated dilator responses (p < 0.001), whereas the SIRT-1 inhibitor EX-527 had no effect. RV-NLCs improved the impaired endothelial-dependent dilation of small coronary arteries, following acute pressure elevation, via NO and downstream COX elements, mediated by AMPK. We suggest that RV-NLCs are an effective delivery modality for improved potency and sustained drug release into the vasculature. Our findings have important implications for the future design and implementation of antihypertensive treatment strategies.

Tetramethoxystilbene-Loaded Liposomes Restore Reactive-Oxygen-Species-Mediated Attenuation of Dilator Responses in Rat Aortic Vessels Ex vivo.

The methylated analogue of the polyphenol resveratrol (RV), 2,3',4,5'-tetramethoxystilbene (TMS) displays potent antioxidant properties and is an effective cytochrome P450 (CYP) 1B1 inhibitor. The bioavailability of TMS is low. Therefore, the use of liposomes for the encapsulation of TMS is a promising delivery modality for enhanced uptake into tissues. We examined the effect of delivery of TMS in liposomes on the restoration of vasodilator responses of isolated aortic vessels after acute tension elevation ex vivo. Aortic vessels from young male Wistar rats were isolated, and endothelial-dependent (acetylcholine, ACh) and -independent (sodium nitroprusside, SNP) responses assessed. Acute tension elevation (1 h) significantly reduced ACh dilator responses, which were restored following incubation with superoxide dismutase or apocynin (an NADPH oxidase inhibitor). Incubation with TMS-loaded liposomes (mean diameter 157 ± 6 nm; PDI 0.097) significantly improved the attenuated dilator responses following tension elevation, which was sustained over a longer period (4 h) when compared to TMS solution. Endothelial denudation or co-incubation with L-NNA (Nω-nitro-l-arginine; nitric oxide synthase inhibitor) resulted in loss of dilator function. Our findings suggest that TMS-loaded liposomes can restore attenuated endothelial-dependent dilator responses induced by an oxidative environment by reducing NADPH-oxidase-derived ROS and potentiating the release of the vasodilator nitric oxide. TMS-loaded liposomes may be a promising therapeutic strategy to restore vasodilator function in vascular disease.

Titania coating of mesoporous silica nanoparticles for improved biocompatibility and drug release within blood vessels.

Blood vessel disease is a major contributor to cardiovascular morbidity and mortality and is hallmarked by dysfunction of the lining endothelial cells (ECs). These cells play a significant role in vascular homeostasis, through the release of mediators to control vessel diameter, hence tissue perfusion. Mesoporous silica nanoparticles (MSNs) can be used as potential drug delivery platforms for vasodilator drugs. Here, using an ex vivo model of vascular function, we examine the use of titania coating for improved biocompatibility and release dynamics of MSN loaded sodium nitroprusside (SNP). MSNs (95 ±â€¯23 nm diameter; pore size 2.7 nm) were synthesised and fully characterised. They were loaded with SNP and coated with titania (TiO2), using the magnetron sputtering technique. Pre-constricted aortic vessels were exposed to drug loaded MSNs (at 1.96 × 1012 MSN mL-1) and the time course of vessel dilation observed, in real time. Exposure of viable vessels to MSNs lead to their internalization into the cytoplasm of ECs, while TiMSNs were also observed in the elastic lamina and smooth muscle cell layers. We demonstrate that titania coating of MSNs significantly improves their biocompatibility and alters the dynamics of drug release. A slow and more sustained relaxation was evident after uptake of TiMSN-SNP, in comparison to uncoated MSN-SNP (rate of dilation was 0.08% per min over a 2.5 h period). The use of titania coated MSNs for drug delivery to the vasculature may be an attractive strategy for therapeutic clinical intervention in cardiovascular disease. STATEMENT OF SIGNIFICANCE: Cardiovascular disease is a major cause of mortality and morbidity worldwide, with a total global cost of over $918 billion, by 2030. Mesoporous silica nanoparticles (MSNs) have great potential for the delivery of drugs that can treat vessel disease. This paper provides the first description for the use of titania coated MSNs with increased vascular penetration, for the delivery of vasodilator drugs, without compromising overall vessel function. We demonstrate that titania coating of MSNs significantly improves their biocompatibility and uptake within aortic blood vessels and furthermore, enables a slower and more sustained release of the vasodilator drug, sodium nitroprusside within the vessel, thus making them an attractive strategy for the treatment of vascular disease.

Polyvinylpyrrolidone-coated gold nanoparticles inhibit endothelial cell viability, proliferation, and ERK1/2 phosphorylation and reduce the magnitude of endothelial-independent dilator responses in isolated aortic vessels.

BACKGROUND: Gold nanoparticles (AuNPs) demonstrate clinical potential for drug delivery and imaging diagnostics. As AuNPs aggregate in physiological fluids, polymer-surface modifications are utilized to allow their stabilization and enhance their retention time in blood. However, the impact of AuNPs on blood vessel function remains poorly understood. In the present study, we investigated the effects of AuNPs and their stabilizers on endothelial cell (EC) and vasodilator function. MATERIALS AND METHODS: Citrate-stabilized AuNPs (12±3 nm) were synthesized and surface-modified using mercapto polyethylene glycol (mPEG) and polyvinylpyrrolidone (PVP) polymers. Their uptake by isolated ECs and whole vessels was visualized using transmission electron microscopy and quantified using inductively coupled plasma mass spectrometry. Their biological effects on EC proliferation, viability, apoptosis, and the ERK1/2-signaling pathway were determined using automated cell counting, flow cytometry, and Western blotting, respectively. Endothelial-dependent and independent vasodilator functions were assessed using isolated murine aortic vessel rings ex vivo. RESULTS: AuNPs were located in endothelial endosomes within 30 minutes' exposure, while their surface modification delayed this cellular uptake over time. After 24 hours' exposure, all AuNPs (including polymer-modified AuNPs) induced apoptosis and decreased cell viability/proliferation. These inhibitory effects were lost after 48 hours' exposure (except for the PVP-modified AuNPs). Furthermore, all AuNPs decreased acetylcholine (ACh)-induced phosphorylation of ERK1/2, a key signaling protein of cell function. mPEG-modified AuNPs had lower cytostatic effects than PVP-modified AuNPs. Citrate-stabilized AuNPs did not alter endothelial-dependent vasodilation induced by ACh, but attenuated endothelial-independent responses induced by sodium nitroprusside. PVP-modified AuNPs attenuated ACh-induced dilation, whereas mPEG-modified AuNPs did not, though this was dose-related. CONCLUSION: We demonstrated that mPEG-modified AuNPs at a therapeutic dosage showed lower cytostatic effects and were less detrimental to vasodilator function than PVP-modified AuNPs, indicating greater potential as agents for diagnostic imaging and therapy.

The influence of silica nanoparticles on small mesenteric arterial function.

AIM: To determine the influence of silica nanoparticles (SiNPs) on small arterial function; both ex vivo and in vivo. METHODS: Mono-dispersed dye-encapsulated SiNPs (97.85 ± 2.26 nm) were fabricated and vasoconstrictor and vasodilator responses of mesenteric arteries assessed. RESULTS: We show that while exposure to SiNPs under static conditions, attenuated endothelial dependent dilator responses ex vivo, attenuation was only evident at lower agonist concentrations, when exposed under flow conditions or after intravenous administration in vivo. Pharmacological inhibition studies suggest that SiNPs may interfere with the endothelial dependent hyperpolarizing factor vasodilator pathway. CONCLUSION: The dosage dependent influence of SiNPs on arterial function will help identify strategies for their safe clinical administration.

Real-time observation of aortic vessel dilation through delivery of sodium nitroprusside via slow release mesoporous nanoparticles.

Spherical mesoporous nanoparticles (MNPs) with a diameter of ∼100nm were synthesised via a sol-gel method in the presences of organic template (with and without fluorescein dye encapsulation). The template molecules were removed by acidic extraction to form a regular pore lattice structure. The nanoparticle size and morphology were analysed using transmission electron microscopy and dynamic light scattering analysis. The MNPs were further characterised by zeta potential, nitrogen adsorption measurements and infra-red spectroscopy. The interior pores had an average diameter of ∼3nm and were loaded with an endothelial-independent vasodilator, sodium nitroprusside (SNP). The optimal drug loading and drug release was determined in high potassium physiological salt solution using dialysis and atomic absorption spectroscopy. We demonstrate that the initial instantaneous release is due to the surface desorption of the drug followed by diffusion from the pores. Furthermore, these drug loaded MNPs (with and without fluorescein dye encapsulation) were added to viable aortic vessels and release in real-time was observed, ex vivo. MNPs and loaded with and without SNP were incubated with the vessel (at 1.96×10(12)NPmL(-1)) over a 3h time period. The real-time exposure to unloaded MNPs resulted in a small attenuation in constriction that occurred after approximately 1h. In contrast, MNPs loaded with SNP led to a rapid relaxation of aortic vessels that was sustained over the 3h period (p<0.001).

Attenuation of endothelial-dependent vasodilator responses, induced by dye-encapsulated silica nanoparticles, in aortic vessels.

AIM: To determine the influence of silica nanoparticle (SiNP) number, size and dye encapsulation on conduit arterial function, in vitro. MATERIALS & METHODS: Rhodamine B isothiocyanate (RBITC) dye molecules were encapsulated in a silica shell to produce nanoparticles (silica RBITC nanoparticles) smaller than 100 nm size. Their effects on endothelial-dependent (acetylcholine; 0.01-200 µM) and -independent (sodium nitroprusside; 0.001-10 µM) dilator responses were examined. RESULTS: When incubated with 1.96 × 10(12) nanoparticles/ml, both 30 and 70 nm SiNPs and silica RBITC nanoparticles significantly reduced endothelium-dependent, but not -independent, vasodilation. The degree of attenuation was related to nanoparticle surface area, rather than size, and influenced by dye encapsulation. Furthermore, attenuated dilation due to silica RBITC nanoparticles, but not SiNPs, could be partially restored using superoxide dismutase. CONCLUSION: Our results suggest that the mechanism of attenuated dilation is different for SiNPs and silica RBITC nanoparticles, which has implications for the future fabrication of biocompatible nanoparticles for imaging diagnostics.

Altered sensitivity to nitric oxide donors, induced by intravascular infusion of quantum dots, in murine mesenteric arteries.

Quantum dots (QDs) are utilised in imaging diagnostics, tissue engineering and medical therapeutics, however, their influence on vascular function is not ascertained. Here, we examined small mesenteric arterial responses after acute intravascular exposure to QDs. Incubation in mercaptoundecanoic acid (MUA)-coated QDs (at 15 µg/mL) had no influence on endothelial-dependent dilator responses (Acetylcholine; Ach) but led to an attenuated relaxation to the nitric oxide donor, sodium nitroprusside (SNP). Conversely, incubation in POSS-PCU coated QDs (at 15 µg/mL) led to attenuated Ach responses (10(-11)-10(-3)M; n=5, P<0.05), but had no influence on SNP-induced relaxation. At lower concentrations of POSS-PCU coated QDs (5 µg/mL), Ach responses were preserved. We demonstrate that acute exposure to QDs, can attenuate vasodilation but not vasoconstriction, and is dependent on their surface coatings. Our findings have implications in QD use for imaging diagnostics in disease states, where SNP based drugs are used in therapeutic intervention. FROM THE CLINICAL EDITOR: In this paper, the influence of quantum dots on vascular function is investigated---an important aspect to consider with the growing utility of quantum dots in imaging diagnostics, tissue engineering and medical therapeutics.

Biocompatibility of amorphous silica nanoparticles: Size and charge effect on vascular function, in vitro.

Synthetic amorphous silica is gaining popularity as the material of choice in the fabrication of nanoparticles for use in imaging diagnostics, medical therapeutics, and tissue engineering because of its biocompatible nature. However, recent evidence suggests that silica nanoparticles (SiNPs) show a concentration- and size-dependent toxic effect that is cell specific. We investigated the direct influence of SiNP uptake on the vasodilator responses of rat aortic vessels, in vitro, using fabricated SiNPs of defined size (97 ± 7.60 and 197 ± 7.50 nm) and charge (positive and nonmodified). Dilator responses to cumulative doses of endothelial-dependent [acetylcholine (Ach); 0.01 µM-1.0 mM] and endothelial-independent (sodium nitroprusside; 0.01-10 µM) agonists were determined before and 30 Min after incubation in SiNPs (at 1.1 × 10(11) nanoparticles/mL). Acute exposure to SiNPs led to their rapid uptake by the lining endothelial cells (as verified by transmission electron microscopy). SiNP uptake had no significant influence on dilator responses, although a greater degree of attenuation was evident after uptake of the 100 nm and positively charged SiNPs (significant at the highest 1.0 mM Ach concentration between positive and nonmodified 200 nm SiNPs; P < 0.05). In summary, our findings suggest that SiNP surface interactions, rather than mass, affect vasodilator function of aortic vessels.