Red blood cell complement receptor one level varies with Knops blood group, α(+)thalassaemia and age among Kenyan children

Both the invasion of red blood cells (RBCs) by Plasmodium falciparum parasites and the sequestration of parasite-infected RBCs in the microvasculature are mediated in part by complement receptor one (CR1). RBC surface CR1 level can vary between individuals by more than 20-fold and may be associated with the risk of severe malaria. The factors that influence RBC CR1 level variation are poorly understood, particularly in African populations. We studied 3535 child residents of a malaria-endemic region of coastal Kenya and report, for the first time, that the CR1 Knops blood group alleles Sl2 and McC(b), and homozygous HbSS are positively associated with RBC CR1 level. Sickle cell trait and ABO blood group did not influence RBC CR1 level. We also confirm the previous observation that α(+)thalassaemia is associated with reduced RBC CR1 level, possibly due to small RBC volume, and that age-related changes in RBC CR1 expression occur throughout childhood. RBC CR1 level in malaria-endemic African populations is a complex phenotype influenced by multiple factors that should be taken into account in the design and interpretation of future studies on CR1 and malaria susceptibility

Docosahexaenoic acid liposomes for targeting chronic inflammatory diseases and cancer: an in vitro assessment

Amr Alaarg,1,2 Nan Yeun Jordan,1 Johan JF Verhoef,1 Josbert M Metselaar,2,3 Gert Storm,1,2 Robbert J Kok1 1Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Utrecht, 2Department of Biomaterials Science and Technology, Institute for Biomedical Technology and Technical Medicine (MIRA), University of Twente, Enschede, the Netherlands; 3Department of Experimental Molecular Imaging, University Clinic and Helmholtz Institute for Biomedical Engineering, RWTH-Aachen University, Aachen, Germany Abstract: Inflammation, oxidative stress, and uncontrolled cell proliferation are common key features of chronic inflammatory diseases, such as atherosclerosis and cancer. ω3 polyunsaturated fatty acids (PUFAs; also known as omega3 fatty acids or fish oil) have beneficial effects against inflammation upon dietary consumption. However, these effects cannot be fully exploited unless diets are enriched with high concentrations of fish oil supplements over long periods of time. Here, a nanomedicine-based approach is presented for delivering effective levels of PUFAs to inflammatory cells. Nanoparticles are internalized by immune cells, and hence can adequately deliver bioactive lipids into these target cells. The ω3 FA docosahexaenoic acid was formulated into liposomes (ω-liposomes), and evaluated for anti-inflammatory effects in different types of immune cells. ω-Liposomes strongly inhibited the release of reactive oxygen species and reactive nitrogen species from human neutrophils and murine macrophages, and also inhibited the production of the proinflammatory cytokines TNFα and MCP1. Moreover, ω-liposomes inhibited tumor-cell proliferation when evaluated in FaDu head and neck squamous carcinoma and 4T1 breast cancer cells in in vitro cultures. We propose that ω-liposomes are a promising nanonutraceutical formulation for intravenous delivery of fish oil FAs, which may be beneficial in the treatment of inflammatory disorders and cancer. Keywords: nanomedicine, PUFA, inflammation, cancer, fish oil, deliver

Applying nanomedicine in maladaptive inflammation and angiogenesis

Inflammation and angiogenesis drive the development and progression of multiple devastating diseases such as atherosclerosis, cancer, rheumatoid arthritis, and inflammatory bowel disease. Though these diseases have very different phenotypic consequences, they possess several common pathophysiological features in which monocyte recruitment, macrophage polarization, and enhanced vascular permeability play critical roles. Thus, developing rational targeting strategies tailored to the different stages of the journey of monocytes, from bone marrow to local lesions, and their extravasation from the vasculature in diseased tissues will advance nanomedicine. The integration of in vivo imaging uniquely allows studying nanoparticle kinetics, accumulation, clearance, and biological activity, at levels ranging from subcellular to an entire organism, and will shed light on the fate of intravenously administered nanomedicines. We anticipate that convergence of nanomedicines, biomedical engineering, and life sciences will help to advance clinically relevant therapeutics and diagnostic agents for patients with chronic inflammatory diseases. (C) 2017 Elsevier B.V. All rights reserve

Cells release subpopulations of exosomes with distinct molecular and biological properties.

Cells release nano-sized membrane vesicles that are involved in intercellular communication by transferring biological information between cells. It is generally accepted that cells release at least three types of extracellular vesicles (EVs): apoptotic bodies, microvesicles and exosomes. While a wide range of putative biological functions have been attributed to exosomes, they are assumed to represent a homogenous population of EVs. We hypothesized the existence of subpopulations of exosomes with defined molecular compositions and biological properties. Density gradient centrifugation of isolated exosomes revealed the presence of two distinct subpopulations, differing in biophysical properties and their proteomic and RNA repertoires. Interestingly, the subpopulations mediated differential effects on the gene expression programmes in recipient cells. In conclusion, we demonstrate that cells release distinct exosome subpopulations with unique compositions that elicit differential effects on recipient cells. Further dissection of exosome heterogeneity will advance our understanding of exosomal biology in health and disease and accelerate the development of exosome-based diagnostics and therapeutics

Design and development of nanomedicines to treat atherosclerosis: A cross platform head-to-head theranostic study

Background: Atherosclerosis is a chronic inflammatory disease of the large arteries and a leading cause of death worldwide. Macrophages are key players in the progression of atherosclerosis and are a compelling target for disease management [1]. Statins, HMG-CoA reductase inhibitors, exhibit anti-inflammatory and antiproliferative pleiotropic effects [2]. Using a nanomedicine approach these effects can be amplified [3]. Here, we systematically study three different statin-loaded nanoparticles in atherosclerotic apoE -/- mice. Since the nanomedicines also contain diagnostic probes in addition to the encapsulated drug, we can use powerful imaging modalities like positron emission tomography with computed tomography (PET/CT) and near-infrared fluorescence (NIRF) imaging to follow the fate of the theranostics nanoparticles. Methods: Simvastatin loaded high-density lipoprotein ([S]-rHDL), polymeric nanoparticles ([S]-PN), and liposomes ([S]-lip.) were developed. The three formulations were characterized by transmission electron microscopy (TEM), dynamic light scattering (DLS) for mean size and polydispersity (PDI), as well as for drug entrapment efficiency (EE%) by high performance liquid chromatography (HPLC). The effect of the three formulations on RAW 264.7 macrophage viability was evaluated in vitro. For multimodal imaging, Cy5.5 phospholipid (for NIRF imaging) and desferrioxamine (DFO)-phospholipid (for 89Zr-labeling and PET imaging), were included in the [S]-rHDL and [S]-lip. formulations, while for [S]-PN, Cy5.5 and DFO were covalently conjugated to the polymeric nanoparticles. In apoE -/- mice, the nanoparticles were injected i.v. 24 hours before performing in vivo PET/CT. The radioactivity and dye distribution were assessed by gamma counting, autoradiography and NIRF imaging ex vivo. Results: The [S] formulations were successfully prepared and had average sizes 60% (Fig.1B and C). [S]-PN shows a more potent effect on RAW 264.7 cell viability with IC50 of 5 μM, compared to IC50 of 10 μM for [S]-HDL and >25 μM for [S]-L (Fig. 1D). Distinct biodistribution profiles for the three nanoparticles were observed by PET/CT imaging, which was corroborated by ex vivo gamma counting (Fig 1E and F). [S]-PN accumulates to a higher degree in spleen and liver compared to [S]-lip. and [S]-rHDL, while the latter [S]-rHDL accumulates to a higher extent in the kidneys. [S]-lip. shows slightly higher concentration in blood at 24 hours. Radioactivity concentration in the aortas was similar for the three nanoparticles. Ex vivo NIRF imaging and autoradiography demonstrated co-localization of 89Zr with Cy5.5 signal in the aortas (Fig. 1G and H). Conclusions: We have developed three [S]-loaded nanoparticle platforms and labeled them for imaging with PET and NIRF. This allowed us to non-invasively visualize their distinct biodistribution profiles and to assess their plaque targeting ability. Studies are ongoing to evaluate the impact of platforms on the therapeutic outcomes

Design and development of nanomedicines to treat atherosclerosis: A cross platform head-to-head theranostic study

Background: Atherosclerosis is a chronic inflammatory disease of the large arteries and a leading cause of death worldwide. Macrophages are key players in the progression of atherosclerosis and are a compelling target for disease management [1]. Statins, HMG-CoA reductase inhibitors, exhibit anti-inflammatory and antiproliferative pleiotropic effects [2]. Using a nanomedicine approach these effects can be amplified [3]. Here, we systematically study three different statin-loaded nanoparticles in atherosclerotic apoE -/- mice. Since the nanomedicines also contain diagnostic probes in addition to the encapsulated drug, we can use powerful imaging modalities like positron emission tomography with computed tomography (PET/CT) and near-infrared fluorescence (NIRF) imaging to follow the fate of the theranostics nanoparticles. Methods: Simvastatin loaded high-density lipoprotein ([S]-rHDL), polymeric nanoparticles ([S]-PN), and liposomes ([S]-lip.) were developed. The three formulations were characterized by transmission electron microscopy (TEM), dynamic light scattering (DLS) for mean size and polydispersity (PDI), as well as for drug entrapment efficiency (EE%) by high performance liquid chromatography (HPLC). The effect of the three formulations on RAW 264.7 macrophage viability was evaluated in vitro. For multimodal imaging, Cy5.5 phospholipid (for NIRF imaging) and desferrioxamine (DFO)-phospholipid (for 89Zr-labeling and PET imaging), were included in the [S]-rHDL and [S]-lip. formulations, while for [S]-PN, Cy5.5 and DFO were covalently conjugated to the polymeric nanoparticles. In apoE -/- mice, the nanoparticles were injected i.v. 24 hours before performing in vivo PET/CT. The radioactivity and dye distribution were assessed by gamma counting, autoradiography and NIRF imaging ex vivo. Results: The [S] formulations were successfully prepared and had average sizes 60% (Fig.1B and C). [S]-PN shows a more potent effect on RAW 264.7 cell viability with IC50 of 5 μM, compared to IC50 of 10 μM for [S]-HDL and >25 μM for [S]-L (Fig. 1D). Distinct biodistribution profiles for the three nanoparticles were observed by PET/CT imaging, which was corroborated by ex vivo gamma counting (Fig 1E and F). [S]-PN accumulates to a higher degree in spleen and liver compared to [S]-lip. and [S]-rHDL, while the latter [S]-rHDL accumulates to a higher extent in the kidneys. [S]-lip. shows slightly higher concentration in blood at 24 hours. Radioactivity concentration in the aortas was similar for the three nanoparticles. Ex vivo NIRF imaging and autoradiography demonstrated co-localization of 89Zr with Cy5.5 signal in the aortas (Fig. 1G and H). Conclusions: We have developed three [S]-loaded nanoparticle platforms and labeled them for imaging with PET and NIRF. This allowed us to non-invasively visualize their distinct biodistribution profiles and to assess their plaque targeting ability. Studies are ongoing to evaluate the impact of platforms on the therapeutic outcomes

A systematic comparison of clinically viable nanomedicines targeting HMG-CoA reductase in inflammatory atherosclerosis

Atherosclerosis is a leading cause of worldwide morbidity and mortality whose management could benefit from novel targeted therapeutics. Nanoparticles are emerging as targeted drug delivery systems in chronic inflammatory disorders. To optimally exploit nanomedicines, understanding their biological behavior is crucial for further development of clinically relevant and efficacious nanotherapeutics intended to reduce plaque inflammation. Here, three clinically relevant nanomedicines, i.e., high-density lipoprotein ([S]-HDL), polymeric micelles ([S]-PM), and liposomes ([S]-LIP), that are loaded with the HMG-CoA reductase inhibitor simvastatin [S], were evaluated in the apolipoprotein E-deficient (Apoe(-/-)) mouse model of atherosclerosis. We systematically employed quantitative techniques, including in vivo positron emission tomography imaging, gamma counting, and flow cytometry to evaluate the biodistribution, nanomedicines' uptake by plaque-associated macrophages/monocytes, and their efficacy to reduce macrophage burden in atherosclerotic plaques. The three formulations demonstrated distinct biological behavior in Apoe(-/-)mice. While [S]-PM and [S]-LIP possessed longer circulation half-lives, the three platforms accumulated to similar levels in atherosclerotic plaques. Moreover, [S]-HDL and [S]-PM showed higher uptake by plaque macrophages in comparison to [S]-LIP, while [S]-PM demonstrated the highest uptake by Ly6C(high) monocytes. Among the three formulations, [S]-PM displayed the highest efficacy in reducing macrophage burden in advanced atherosclerotic plaques. In conclusion, our data demonstrate that [S]-PM is a promising targeted drug delivery system, which can be advanced for the treatment of atherosclerosis and other inflammatory disorders in the clinical settings. Our results also emphasize the importance of a thorough understanding of nanomedicines' biological performance, ranging from the whole body to the target cells, as well drug retention in the nanoparticles. Such systematic investigations would allow rational applications of nanomaterials', beyond cancer, facilitating the expansion of the nanomedicine horizo

Multiple pathway assessment to predict anti-atherogenic efficacy of drugs targeting macrophages in atherosclerotic plaques

BACKGROUND: Macrophages play a central role in atherosclerosis development and progression, hence, targeting macrophage activity is considered an attractive therapeutic. Recently, we documented nanomedicinal delivery of the anti-inflammatory compound prednisolone to atherosclerotic plaque macrophages in patients, which did however not translate into therapeutic efficacy. This unanticipated finding calls for in-depth screening of drugs intended for targeting plaque macrophages. METHODS AND RESULTS: We evaluated the effect of several candidate drugs on macrophage activity, rating overall performance with respect to changes in cytokine release, oxidative stress, lipid handling, endoplasmic reticulum (ER) stress, and proliferation of macrophages. Using this in vitro approach, we observed that the anti-inflammatory effect of prednisolone was counterbalanced by multiple adverse effects on other key pathways. Conversely, pterostilbene, T0901317 and simvastatin had an overall anti-atherogenic effect on multiple pathways, suggesting their potential for liposomal delivery. CONCLUSION: This dedicated assay setup provides a framework for high-throughput assessment. Further in vivo studies are warranted to determine the predictive value of this macrophage-based screening approach and its potential value in nanomedicinal drug development for cardiovascular patients

Liposomal prednisolone promotes macrophage lipotoxicity in experimental atherosclerosis

Atherosclerosis is a lipid-driven inflammatory disease, for which nanomedicinal interventions are under evaluation. Previously, we showed that liposomal nanoparticles loaded with prednisolone (LN-PLP) accumulated in plaque macrophages, however, induced proatherogenic effects in patients. Here, we confirmed in low-density lipoprotein receptor knockout (LDLr(-/-)) mice that LN-PLP accumulates in plaque macrophages. Next, we found that LN-PLP infusions at 10mg/kg for 2weeks enhanced monocyte recruitment to plaques. In follow up, after 6weeks of LN-PLP exposure we observed (i) increased macrophage content, (ii) more advanced plaque stages, and (iii) larger necrotic core sizes. Finally, in vitro studies showed that macrophages become lipotoxic after LN-PLP exposure, exemplified by enhanced lipid loading, ER stress and apoptosis. These findings indicate that liposomal prednisolone may paradoxically accelerate atherosclerosis by promoting macrophage lipotoxicity. Hence, future (nanomedicinal) drug development studies are challenged by the multifactorial nature of atherosclerotic inflammation