Different coatings on magnetic nanoparticles dictate their degradation kinetics in vivo for 15 months after intravenous administration in mice.

BACKGROUND: The surface coating of iron oxide magnetic nanoparticle (MNPs) drives their intracellular trafficking and degradation in endolysosomes, as well as dictating other cellular outcomes. As such, we assessed whether MNP coatings might influence their biodistribution, their accumulation in certain organs and their turnover therein, processes that must be understood in vivo to optimize the design of nanoformulations for specific therapeutic/diagnostic needs. RESULTS: In this study, three different MNP coatings were analyzed, each conferring the identical 12 nm iron oxide cores with different physicochemical characteristics: 3-aminopropyl-triethoxysilane (APS), dextran (DEX), and dimercaptosuccinic acid (DMSA). When the biodistribution of these MNPs was analyzed in C57BL/6 mice, they all mainly accumulated in the spleen and liver one week after administration. The coating influenced the proportion of the MNPs in each organ, with more APS-MNPs accumulating in the spleen and more DMSA-MNPs accumulating in the liver, remaining there until they were fully degraded. The changes in the physicochemical properties of the MNPs (core size and magnetic properties) was also assessed during their intracellular degradation when internalized by two murine macrophage cell lines. The decrease in the size of the MNPs iron core was influenced by their coating and the organ in which they accumulated. Finally, MNP degradation was analyzed in the liver and spleen of C57BL/6 mice from 7 days to 15 months after the last intravenous MNP administration. CONCLUSIONS: The MNPs degraded at different rates depending on the organ and their coating, the former representing the feature that was fundamental in determining the time they persisted. In the liver, the rate of degradation was similar for all three coatings, and it was faster than in the spleen. This information regarding the influence of coatings on the in vivo degradation of MNPs will help to choose the best coating for each biomedical application depending on the specific clinical requirements.

Iron oxide and iron oxyhydroxide nanoparticles impair SARS-CoV-2 infection of cultured cells.

BACKGROUND: Coronaviruses usually cause mild respiratory disease in humans but as seen recently, some human coronaviruses can cause more severe diseases, such as the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the global spread of which has resulted in the ongoing coronavirus pandemic. RESULTS: In this study we analyzed the potential of using iron oxide nanoparticles (IONPs) coated with biocompatible molecules like dimercaptosuccinic acid (DMSA), 3-aminopropyl triethoxysilane (APS) or carboxydextran (FeraSpin™ R), as well as iron oxyhydroxide nanoparticles (IOHNPs) coated with sucrose (Venofer®), or iron salts (ferric ammonium citrate -FAC), to treat and/or prevent SARS-CoV-2 infection. At non-cytotoxic doses, IONPs and IOHNPs impaired virus replication and transcription, and the production of infectious viruses in vitro, either when the cells were treated prior to or after infection, although with different efficiencies. Moreover, our data suggest that SARS-CoV-2 infection affects the expression of genes involved in cellular iron metabolism. Furthermore, the treatment of cells with IONPs and IOHNPs affects oxidative stress and iron metabolism to different extents, likely influencing virus replication and production. Interestingly, some of the nanoparticles used in this work have already been approved for their use in humans as anti-anemic treatments, such as the IOHNP Venofer®, and as contrast agents for magnetic resonance imaging in small animals like mice, such as the FeraSpin™ R IONP. CONCLUSIONS: Therefore, our results suggest that IONPs and IOHNPs may be repurposed to be used as prophylactic or therapeutic treatments in order to combat SARS-CoV-2 infection.

Polyethylenimine-coated superparamagnetic iron oxide nanoparticles impair in vitro and in vivo angiogenesis.

Endothelial cells are essential to tumor vascularization and impairing their activity can potentially limit tumor growth. Since polyethylenimine (PEI)-coated superparamagnetic iron oxide nanoparticles (SPIONs) are bioactive nanosystems that modulate inflammatory macrophage responses and limit tumor cell invasion, we evaluated their effects on endothelial cell angiogenesis. PEI-SPION triggered proinflammatory gene profiles in a murine endothelial cell line and in primary human umbilical cord vein endothelial cells (HUVECs). These nanoparticles impaired endothelial cell migration and inhibited HUVEC tube formation. Magnetically tumor-targeted PEI-SPIONs reduced tumor vessel numbers and promoted intratumor macrophage infiltration in a tumor xenograft model. PEI-SPION treatment impaired M2 macrophage-promoted tube formation and affected HUVEC cytoskeleton by limiting Src and Cortactin activation. These mechanisms could contribute to PEI-SPION in vitro and in vivo antiangiogenic potential. These data confirm that PEI-SPION administration and application of a localized magnetic field could offer an affordable anti-angiogenic anti-tumoral targeted treatment that would complement other therapies.

Superparamagnetic iron oxide nanoparticle uptake alters M2 macrophage phenotype, iron metabolism, migration and invasion.

Superparamagnetic iron oxide nanoparticles (SPIONs) have shown promise as contrast agents and nanocarriers for drug delivery. Their impact on M2-polarised macrophages has nonetheless not been well studied. Here we explored the effects of SPIONs coated with dimercaptosuccinic acid, aminopropyl silane or aminodextran in two M2 macrophage models (murine primary IL-4-activated bone marrow-derived macrophages and human M2-like differentiated THP-1 cells). All SPIONs were internalised and no cell toxicity was observed. SPION treatment produced reactive oxygen species and activated the extracellular signal-regulated kinase and AKT pathways. After 24-h SPION incubation, M2 macrophages switched their iron metabolism towards an iron-replete state. SPION treatment in both M2 macrophage models altered their M2 activation profiles, promoted IL-10 production, and stimulated protease-dependent invasion. These results highlight the need to evaluate the interactions between SPIONs and cells to take full advantage of the intrinsic properties of these nanoparticles in biological systems. FROM THE CLINICAL EDITOR: Superparamagnetic iron oxide nanoparticles (SPIONs) have been used as an MRI contrast agent in many experimental studies. The authors here investigated the effects of these nanoparticles on M2 macrophages after cellular uptake. The findings of cell activation further enhanced our current knowledge on the interaction of SPIONS with macrophages.

Validation of ZIP4 as a tumour-associated antigen for nanotargeting.

Pancreatic ductal adenocarcinoma remains a highly aggressive and untreatable cancer. There is a need to develop a new PDAC-associated antigen-targeting drug delivery system to tackle this disease. We validated choosing ZIP4 as a putative target in PDAC theranostics. We developed a nanosystem composed of a fluorescent polystyrene core coated with gold nanoparticles onto which a ZIP4-specific polyclonal antibody is attached. The polystyrene core's fluorescence properties allow the nanosystem tracking by intravital imaging. We also developed two ZIP4-expressing cell lines by stably transfecting HEK293 and RWP1 cells with a ZIP4-coding plasmid that simultaneously provides cells with puromycin resistance. We studied the cell internalisation of the as-synthesised nanoparticles and demonstrated that ZIP4-expressing HEK293 and ZIP4-expressing RWP1 cells tended to take up more ZIP4-targeting nanoparticles. Moreover, we observed that ZIP4-targeting nanoparticles accumulated more in ZIP4-expressing HEK293 and RWP1 tumours when injected intravenously in a subcutaneous xenograft and an orthotopic in vivo model, respectively. Furthermore, the administration of these nanoparticles did not induce any significant systemic toxicity as determined by histological analysis of all organs. Altogether, these results provide the first evidence of the feasibility of using a ZIP4-targeting nanosystem further to design efficient therapeutic and diagnostic tools for PDAC.

Interaction of Iron Oxide Nanoparticles with Macrophages Is Influenced Distinctly by "Self" and "Non-Self" Biological Identities.

Upon contact with biological fluids like serum, a protein corona (PC) complex forms on iron oxide nanoparticles (IONPs) in physiological environments and the proteins it contains influence how IONPs act in biological systems. Although the biological identity of PC-IONP complexes has often been studied in vitro and in vivo, there have been inconsistent results due to the differences in the animal of origin, the type of biological fluid, and the physicochemical properties of the IONPs. Here, we identified differences in the PC composition when it was derived from the sera of three species (bovine, murine, or human) and deposited on IONPs with similar core diameters but with different coatings [dimercaptosuccinic acid (DMSA), dextran (DEX), or 3-aminopropyl triethoxysilane (APS)], and we assessed how these differences influenced their effects on macrophages. We performed a comparative proteomic analysis to identify common proteins from the three sera that adsorb to each IONP coating and the 10 most strongly represented proteins in PCs. We demonstrated that the PC composition is dependent on the origin of the serum rather than the nature of the coating. The PC composition critically affects the interaction of IONPs with macrophages in self- or non-self identity models, influencing the activation and polarization of macrophages. However, such effects were more consistent for DMSA-IONPs. As such, a self biological identity of IONPs promotes the activation and M2 polarization of murine macrophages, while a non-self biological identity favors M1 polarization, producing larger quantities of ROS. In a human context, we observed the opposite effect, whereby a self biological identity of DMSA-IONPs promotes a mixed M1/M2 polarization with an increase in ROS production. Conversely, a non-self biological identity of IONPs provides nanoparticles with a stealthy character as no clear effects on human macrophages were evident. Thus, the biological identity of IONPs profoundly affects their interaction with macrophages, ultimately defining their biological impact on the immune system.

Cationic Polyethyleneimine (PEI)-Gold Nanocomposites Modulate Macrophage Activation and Reprogram Mouse Breast Triple-Negative MET-1 Tumor Immunological Microenvironment.

Nanomedicines based on inorganic nanoparticles have grown in the last decades due to the nanosystems' versatility in the coating, tuneability, and physical and chemical properties. Nonetheless, concerns have been raised regarding the immunotropic profile of nanoparticles and how metallic nanoparticles affect the immune system. Cationic polymer nanoparticles are widely used for cell transfection and proved to exert an adjuvant immunomodulatory effect that improves the efficiency of conventional vaccines against infection or cancer. Likewise, gold nanoparticles (AuNPs) also exhibit diverse effects on immune response depending on size or coatings. Photothermal or photodynamic therapy, radiosensitization, and drug or gene delivery systems take advantage of the unique properties of AuNPs to deeply modify the tumoral ecosystem. However, the collective effects that AuNPs combined with cationic polymers might exert on their own in the tumor immunological microenvironment remain elusive. The purpose of this study was to analyze the triple-negative breast tumor immunological microenvironment upon intratumoral injection of polyethyleneimine (PEI)-AuNP nanocomposites (named AuPEI) and elucidate how it might affect future immunotherapeutic approaches based on this nanosystem. AuPEI nanocomposites were synthesized through a one-pot synthesis method with PEI as both a reducing and capping agent, resulting in fractal assemblies of about 10 nm AuNPs. AuPEI induced an inflammatory profile in vitro in the mouse macrophage-like cells RAW264.7 as determined by the secretion of TNF-α and CCL5 while the immunosuppressor IL-10 was not increased. However, in vivo in the mouse breast MET-1 tumor model, AuPEI nanocomposites shifted the immunological tumor microenvironment toward an M2 phenotype with an immunosuppressive profile as determined by the infiltration of PD-1-positive lymphocytes. This dichotomy in AuPEI nanocomposites in vitro and in vivo might be attributed to the highly complex tumor microenvironment and highlights the importance of testing the immunogenicity of nanomaterials in vitro and more importantly in vivo in relevant immunocompetent mouse tumor models to better elucidate any adverse or unexpected effect.

Multiphoton imaging of melanoma 3D models with plasmonic nanocapsules.

We report the synthesis of plasmonic nanocapsules and the cellular responses they induce in 3D melanoma models for their perspective use as a photothermal therapeutic agent. The wall of the nanocapsules is composed of polyelectrolytes. The inner part is functionalized with discrete gold nanoislands. The cavity of the nanocapsules contains a fluorescent payload to show their ability for loading a cargo. The nanocapsules exhibit simultaneous two-photon luminescent, fluorescent properties and X-ray contrasting ability. The average fluorescence lifetime (τ) of the nanocapsules measured with FLIM (0.3 ns) is maintained regardless of the intracellular environment, thus proving their abilities for bioimaging of models such as 3D spheroids with a complex architecture. Their multimodal imaging properties are exploited for the first time to study tumorspheres cellular responses exposed to the nanocapsules. Specifically, we studied cellular uptake, toxicity, intracellular fate, generation of reactive oxygen species, and effect on the levels of hypoxia by using multi-photon and confocal laser scanning microscopy. Because of the high X-ray attenuation and atomic number of the gold nanostructure, we imaged the nanocapsule-cell interactions without processing the sample. We confirmed maintenance of the nanocapsules' geometry in the intracellular milieu with no impairment of the cellular ultrastructure. Furthermore, we observed the lack of cellular toxicity and no alteration in oxygen or reactive oxygen species levels. These results in 3D melanoma models contribute to the development of these nanocapsules for their exploitation in future applications as agents for imaging-guided photothermal therapy. STATEMENT OF SIGNIFICANCE: The novelty of the work is that our plasmonic nanocapsules are multimodal. They are responsive to X-ray and to multiphoton and single-photon excitation. This allowed us to study their interaction with 2D and 3D cellular structures and specifically to obtain information on tumor cell parameters such as hypoxia, reactive oxygen species, and toxicity. These nanocapsules will be further validated as imaging-guided photothermal probes.

The surface coating of iron oxide nanoparticles drives their intracellular trafficking and degradation in endolysosomes differently depending on the cell type.

Magnetic nanoparticles (MNPs) are potential theranostic tools that are biodegraded through different endocytic pathways. However, little is known about the endolysosomal network through which MNPs transit and the influence of the surface coating in this process. Here, we studied the intracellular transit of two MNPs with identical iron oxide core size but with two distinct coatings: 3-aminopropyl-trietoxysilane (APS) and dimercaptosuccinic acid (DMSA). Using endolysosomal markers and a high throughput analysis of the associated proteome, we tracked the MNPs intracellularly in two different mouse cell lines, RAW264.7 (macrophages) and Pan02 (tumor cells). We did not detect differences in the MNP trafficking kinetics nor in the MNP-containing endolysosome phenotype in Pan02 cells. Nonetheless, DMSA-MNPs transited at slower rate than APS-MNPs in macrophages as measured by MNP accumulation in Rab7+ endolysosomes. Macrophage DMSA-MNP-containing endolysosomes had a higher percentage of lytic enzymes and catalytic proteins than their APS-MNP counterparts, concomitantly with a V-type ATPase enrichment, suggesting an acidic nature. Consequently, more autophagic vesicles are induced by DMSA-MNPs in macrophages, enhancing the expression of iron metabolism-related genes and proteins. Therefore, unlike Pan02 cells, the MNP coating appears to influence the intracellular trafficking rate and the endolysosome nature in macrophages. These results highlight how the MNP coating can determine the nanoparticle intracellular fate and biodegradation in a cell-type bias.

The Use of Iron Oxide Nanoparticles to Reprogram Macrophage Responses and the Immunological Tumor Microenvironment.

The synthesis and functionalization of iron oxide nanoparticles (IONPs) is versatile, which has enhanced the interest in studying them as theranostic agents over recent years. As IONPs begin to be used for different biomedical applications, it is important to know how they affect the immune system and its different cell types, especially their interaction with the macrophages that are involved in their clearance. How immune cells respond to therapeutic interventions can condition the systemic and local tissue response, and hence, the final therapeutic outcome. Thus, it is fundamental to understand the effects that IONPs have on the immune response, especially in cancer immunotherapy. The biological effects of IONPs may be the result of intrinsic features of their iron oxide core, inducing reactive oxygen species (ROS) and modulating intracellular redox and iron metabolism. Alternatively, their effects are driven by the nanoparticle coating, for example, through cell membrane receptor engagement. Indeed, exploiting these properties of IONPs could lead to the development of innovative therapies. In this review, after a presentation of the elements that make up the tumor immunological microenvironment, we will review and discuss what is currently known about the immunomodulatory mechanisms triggered by IONPs, mainly focusing on macrophage polarization and reprogramming. Consequently, we will discuss the implications of these findings in the context of plausible therapeutic scenarios for cancer immunotherapy.

Tumor-Selective Immune-Active Mild Hyperthermia Associated with Chemotherapy in Colon Peritoneal Metastasis by Photoactivation of Fluorouracil-Gold Nanoparticle Complexes.

Peritoneal metastasis (PM) is considered as the terminal stage of metastatic colon cancer, with still poor median survival rate even with the best recent chemotherapy treatment. The current PM treatment combines cytoreductive surgery, which consists of resecting all macroscopic tumors, with hyperthermic intraperitoneal chemotherapy (HIPEC), which uses mild hyperthermia to boost the diffusion and cytotoxic effect of chemotherapeutic drugs. As HIPEC is performed via a closed circulation of a hot liquid containing chemotherapy, it induces uncontrolled heating and drug distribution in the whole peritoneal cavity with important off-site toxicity and a high level of morbidity. Here, we propose a safer precision strategy using near-infrared (NIR) photoactivated gold nanoparticles (AuNPs) coupled to the chemotherapeutic drug 5-fluorouracil (5-FU) to enable a spatial and temporal control of mild chemo-hyperthermia targeted to the tumor nodules within the peritoneal cavity. Both the 16 nm AuNPs and the corresponding complex with 5-FU (AuNP-5-FU) were shown as efficient NIR photothermal agents in the microenvironment of subcutaneous colon tumors as well as PM in syngeneic mice. Noteworthy, NIR photothermia provided additional antitumor effects to 5-FU treatment. A single intraperitoneal administration of AuNP-5-FU resulted in their preferential accumulation in tumor nodules and peritoneal macrophages, allowing light-induced selective hyperthermia, extended tumor necrosis, and activation of a pro-inflammatory immune response while leaving healthy tissues without any damage. From a translational standpoint, the combined and tumor-targeted photothermal and chemotherapy mediated by the AuNP-drug complex has the potential to overcome the current off-target toxicity of HIPEC in clinical practice.

The Intrinsic Biological Identities of Iron Oxide Nanoparticles and Their Coatings: Unexplored Territory for Combinatorial Therapies.

Over the last 20 years, iron oxide nanoparticles (IONPs) have been the subject of increasing investigation due to their potential use as theranostic agents. Their unique physical properties (physical identity), ample possibilities for surface modifications (synthetic identity), and the complex dynamics of their interaction with biological systems (biological identity) make IONPs a unique and fruitful resource for developing magnetic field-based therapeutic and diagnostic approaches to the treatment of diseases such as cancer. Like all nanomaterials, IONPs also interact with different cell types in vivo, a characteristic that ultimately determines their activity over the short and long term. Cells of the mononuclear phagocytic system (macrophages), dendritic cells (DCs), and endothelial cells (ECs) are engaged in the bulk of IONP encounters in the organism, and also determine IONP biodistribution. Therefore, the biological effects that IONPs trigger in these cells (biological identity) are of utmost importance to better understand and refine the efficacy of IONP-based theranostics. In the present review, which is focused on anti-cancer therapy, we discuss recent findings on the biological identities of IONPs, particularly as concerns their interactions with myeloid, endothelial, and tumor cells. Furthermore, we thoroughly discuss current understandings of the basic molecular mechanisms and complex interactions that govern IONP biological identity, and how these traits could be used as a stepping stone for future research.

Endocytosis-driven gold nanoparticle fractal rearrangement in cells and its influence on photothermal conversion.

Cellular endocytosis and intracellular trafficking of nanoparticles induce dynamic rearrangements that profoundly modify the physical properties of nanoparticle and govern their biological outcomes when activated by external fields. The precise structure, organization, distribution, and density of gold nanoparticles (AuNPs) confined within intracellular compartments such as lysosomes have not been studied comprehensively, hampering the derivation of predictive models of their therapeutic activity within the cells of interest. By using transmission electron microscopy and small-angle X-ray scattering, we have determined that canonical spherical citrate-coated AuNPs in the 3-30 nm size range form fractal clusters in endolysosomes of macrophages, endothelial cells, and colon cancer cells. Statistical analysis revealed that the cluster size and endolysosome size are correlated but do not depend on the size of AuNPs unless larger preformed aggregates of AuNPs are internalized. Smaller AuNPs are confined in greater numbers in loose aggregates covering a higher fraction of the endolysosomes compared to the largest AuNPs. The fractal dimensions of intracellular clusters increased with the particle size, regardless of the cell type. We thus analyzed how these intracellular structure parameters of AuNPs affect their optical absorption and photothermal properties. We observed that a 2nd plasmon resonance band was shifted to the near-infrared region when the nanoparticle size and fractal dimensions of the intracellular cluster increased. This phenomenon of intracellular plasmon coupling is not directly correlated to the size of the intralysosomal cluster or the number of AuNPs per cluster but rather to the compacity of the cluster and the size of the individual AuNPs. The intracellular plasmon-coupling phenomenon translates to an efficient heating efficiency with the excitation of the three cell types at 808 nm, transforming the NIR-transparent canonical AuNPs with sizes below 30 nm into NIR-absorbing clusters in the tumor microenvironment. Harnessing the spontaneous clustering of spherical AuNPs by cells might be a more valuable strategy for theranostic purposes than deploying complex engineering to derive NIR-absorbent nanostructures out of their environment. Our paper sheds light on AuNP intracellular reorganization and proposes a general method to link their intracellular fates to their in situ physical properties exploited in medical applications.

Polyethyleneimine-assisted one-pot synthesis of quasi-fractal plasmonic gold nanocomposites as a photothermal theranostic agent.

Gold nanoparticles have been thoroughly used in designing thermal ablative therapies and in photoacoustic imaging in cancer treatment owing to their unique and tunable plasmonic properties. While the plasmonic properties highly depend on the size and structure, controllable aggregation of gold nanoparticles can trigger a plasmonic coupling of adjacent electronic clouds, henceforth leading to an increase of light absorption within the near-infrared (NIR) window. Polymer-engraftment of gold nanoparticles has been investigated to achieve the plasmonic coupling phenomenon, but complex chemical steps are often needed to accomplish a biomedically relevant product. An appealing and controllable manner of achieving polymer-based plasmon coupling is a template-assisted Au+3 reduction that ensures in situ gold reduction and coalescence. Among the polymers exploited as reducing agents are polyethyleneimines (PEI). In this study, we addressed the PEI-assisted synthesis of gold nanoparticles and their further aggregation to obtain fractal NIR-absorbent plasmonic nanoaggregates for photothermal therapy and photoacoustic imaging of colorectal cancer. PEI-assisted Au+3 reduction was followed up by UV-visible light absorption, small-angle X-ray scattering (SAXS), and photo-thermal conversion. The reaction kinetics, stability, and the photothermal plasmonic properties of the as-synthesized nanocomposites tightly depended on the PEI : Au ratio. We defined a PEI-Au ratio range (2.5-5) for the one-pot synthesis of gold nanoparticles that self-arrange into fractal nanoaggregates with demonstrated photo-thermal therapeutic and imaging efficiency both in vitro and in vivo in a colorectal carcinoma (CRC) animal model.

Disturbance of adhesomes by gold nanoparticles reveals a size- and cell type-bias.

Gold nanoparticles (AuNP) have been thoroughly studied as multifunctional theranosis agents for cell imaging and cancer therapy as well as sensors due to their tunable physical and chemical properties. Although AuNP have proved to be safe in a wide concentration range, yet other important biological effects can arise in the sublethal window of treatment. This is especially pivotal to understand how AuNP can affect cell biology when labeling steps are needed for cell tracking in vivo, as nanoparticle loading can affect cell migratory/invasion ability, a function mediated by filamentous actin-rich nanometric structures collectively called adhesomes. It is noteworthy that, although numerous research studies have addressed the cell response to AuNP loading, yet none of them focuses on adhesome dynamics as a target of intracellular pathways affected by AuNP. We intend to study the collective dynamics of adhesive F-actin rich structures upon AuNP treatment as an approach to understand the complex AuNP-triggered modulation of migration/invasion related cellular functions. We demonstrated that citrate-coated spherical AuNP of different sizes (3, 11, 16, 30 and 40 nm) disturbed podosome-forming rosettes and the resulting extracellular matrix (ECM) degradation in a murine macrophage model depending on core size. This phenomenon was accompanied by a reduction in metalloproteinase MMP2 and an increment in metalloproteinase inhibitors, TIMP-1/2 and SerpinE1. We also found that AuNP treatment has opposite effects on focal adhesions (FA) in endothelial and mesenchymal stem cells. While endothelial cells reduced their mature FA number and ECM degradation rate upon AuNP treatment, mouse mesenchymal stem cells increased the number and size of mature FA and, therefore, the ECM degradation rate. Overall, AuNP appear to disturb adhesive structures and therefore migratory/invasive cell functions measured as ECM degradation ability, providing new insights into AuNP-cell interaction depending on cell type.

PI3K p85 ß regulatory subunit deficiency does not affect NK cell differentiation and increases NKG2D-mediated activation.

Activation of NK cells depends on a balance between activating and inhibitory signals. Class Ia PI3K are heterodimeric proteins with a catalytic and a regulatory subunit and have a central role in cell signaling by associating with tyrosine kinase receptors to trigger signaling cascades. The regulatory p85 subunit participates in signaling through NKG2D, one of the main activating receptors on NK cells, via its interaction with the adaptor protein DAP10. Although the effects of inhibiting catalytic subunits or deleting the regulatory p85α subunit have been studied, little attention has focused on the role of the p85ß subunit in NK cells. Using p85ß knockout mice, we found that p85ß deficiency does not alter NK cell differentiation and maturation in spleen or bone marrow. NK cells from p85ß-/- mice nonetheless produced more IFN-γ and degranulated more effectively when stimulated with anti-NKG2D antibody. These cells also degranulated and killed NKG2D ligand-expressing target cells more efficiently. We show that p85ß deficiency impaired NKG2D internalization, which could contribute to the activated phenotype. Decreasing p85ß subunit protein levels might thus constitute a therapeutic target to promote NK cell activity toward NKG2D ligand-expressing cells.

Polyethylenimine-coated SPIONs trigger macrophage activation through TLR-4 signaling and ROS production and modulate podosome dynamics.

Polyethylenimine (PEI) is widely used as transfection agent in preclinical studies, both in vitro and in vivo. Due to their unique chemical and physical properties, SPIONs (superparamagnetic iron oxide nanoparticles) have been thoroughly studied as nanocarriers. PEI appears to activate different immune cells to an inflammatory response (M1/TH1), whereas the SPION-induced response seems to be context-dependent; the immunogenicity of the combination of these components has not been studied. Here we show that PEI-coated SPIONs (PMag) activate macrophages, as determined by measuring IL-12 secretion into culture medium and upregulation of several genes linked to the M1 phenotype. PMag-induced phosphorylation of p38 MAPK, p44/p42 MAPK and JNK, and upregulation of CD40, CD80, CD86 and I-A/I-E activation markers. PMag-induced macrophage activation depended partially on TLR4 (Toll-like receptor 4) and ROS (reactive oxygen species) signaling. Comparison of these responses with the LPS (lipopolysaccharide)-induced phenotype showed differences in gene expression profiling. PMag positively modulated podosome formation in murine macrophages, but hampered gelatin degradation by these cells. In conclusion, PMag induced an M1-like phenotype that was partially dependent on both TLR4 and ROS. These results show the adjuvant potential of PMag and suggest their use in vaccination schedules.

Polyethylenimine-coated SPION exhibits potential intrinsic anti-metastatic properties inhibiting migration and invasion of pancreatic tumor cells.

Due to its aggressive behavior, pancreatic cancer is one of the principal causes of cancer-related deaths. The highly metastatic potential of pancreatic tumor cells demands the development of more effective anti-metastatic approaches for this disease. Although polyethylenimine-coated superparamagnetic iron oxide nanoparticles (PEI-coated SPIONs) have been studied for their utility as transfection agents, little is known of their effect on tumor cell biology. Here we demonstrated that PEI-coated SPIONs have potent inhibitory effects on pancreatic tumor cell migration/invasion, through inhibition of Src kinase and decreased expression of MT1-MMP and MMP2 metalloproteinases. When treated with PEI-coated SPIONs, the pancreatic tumor cell line Pan02 showed reduced invadosome density and thus, a decrease in their ability to invade through basement membrane. These nanoparticles temporarily downmodulated microRNA-21, thereby upregulating the cell migration inhibitors PTEN, PDCD4 and Sprouty-1. PEI-coated SPIONs thus show intrinsic, possibly anti-metastatic properties for modulating pancreatic tumor cell migration machinery, which indicates their potential as anti-metastatic agents for treatment of pancreatic cancer.

Comparación de la actividad de 2 factores estimuladores de colonias de granulocitos de producción nacional: Hebervital y Leukocim / Comparison of the activity of 2 domestic- production factors for granulocyte colonies: Hebervital and Leukocim

En 35 pacientes que debían recibir terapia celular regenerativa se evaluó el efecto estimulante de 2 factores estimuladores de colonias de granulocitos de producción cubana: Hebervital y Leukocim, para la movilización de células madre hematopoyéticas hacia la sangre periférica. Los pacientes se seleccionaron de forma aleatoria en 2 grupos de tratamiento y se les suministró por vía subcutánea una dosis total de 40 mg/kg distribuidos en 4 subdosis de 10 mg/kg administrados cada 12 horas. Se realizó el conteo de las células mononucleares y células CD34+ en sangre periférica mediante citometría de flujo, antes y después de la estimulación. No se encontraron diferencias estadísticamente significativas en los conteos de las células CD34+ obtenidas posestimulación con el Hebervital y el Leukocim(AU)

In 35 patients who should have received regenerative cell therapy, it was evaluated the effect of 2 domestic production granulocyte colony stimulating factors: Leukocim and Hebervital for the mobilization of hematopoietic stem cells into peripheral blood. Patients were randomly selected into 2 treatment groups and were given a total dose of 40 mg/kg in 4 sub-doses of 10 mg/kg subcutaneously administered every 12 hours. Count was performed for mononuclear cells and CD34+ cells in peripheral blood by flow cytometry before and after stimulation. There were no statistically significant differences in the counts of CD34+ cells obtained after stimulation with Hebervital and Leukocim(AU)

Comparación de la actividad de 2 factores estimuladores de colonias de granulocitos de producción nacional: Hebervital y Leukocim / Comparison of the activity of 2 domestic- production factors for granulocyte colonies: Hebervital and Leukocim

En 35 pacientes que debían recibir terapia celular regenerativa se evaluó el efecto estimulante de 2 factores estimuladores de colonias de granulocitos de producción cubana: Hebervital y Leukocim, para la movilización de células madre hematopoyéticas hacia la sangre periférica. Los pacientes se seleccionaron de forma aleatoria en 2 grupos de tratamiento y se les suministró por vía subcutánea una dosis total de 40 mg/kg distribuidos en 4 subdosis de 10 mg/kg administrados cada 12 horas. Se realizó el conteo de las células mononucleares y células CD34+ en sangre periférica mediante citometría de flujo, antes y después de la estimulación. No se encontraron diferencias estadísticamente significativas en los conteos de las células CD34+ obtenidas posestimulación con el Hebervital y el Leukocim

In 35 patients who should have received regenerative cell therapy, it was evaluated the effect of 2 domestic production granulocyte colony stimulating factors: Leukocim and Hebervital for the mobilization of hematopoietic stem cells into peripheral blood. Patients were randomly selected into 2 treatment groups and were given a total dose of 40 mg/kg in 4 sub-doses of 10 mg/kg subcutaneously administered every 12 hours. Count was performed for mononuclear cells and CD34+ cells in peripheral blood by flow cytometry before and after stimulation. There were no statistically significant differences in the counts of CD34+ cells obtained after stimulation with Hebervital and Leukocim