Cellular Uptake Of Nanoparticles Research Articles

A generic model based on the properties of nanoparticles and cells for predicting cellular uptake

Nanoparticles (NPs) are widely used in industry and technology due to their small size and versatility, which makes them easy to enter organisms and pose threats to human and ecological health. Given the particularity and complex structure of NPs, statistical models alone cannot reliably predict uptake.Hence, we developed a generic model for predicting the cellular uptake of NPs with organic coatings, based on physicochemical interactions underlying uptake. The model utilized the concentration, experimental conditions and properties of NPs viz. size, surface coating and coverage. These parameters were converted to surface energy components and surface potentials, and combined with the components and potential for a cell membrane. For NPs uptake, we constructed energetic profiles and barriers for adsorption and permeation onto/through cell membranes. The relationships derived were compared to experimental uptake data.The model provided accurate and robust uptake estimates for neutrally charged unhalogenated NPs and six different cell types. We envision that the model provides a reference for cellular accumulation of neutral NPs and (ecological/human) risk assessment of NPs or microparticles.

Development of Microfluidic, Serum-Free Bronchial Epithelial Cells-on-a-Chip to Facilitate a More Realistic In vitro Testing of Nanoplastics.

Most cell culture models are static, but the cellular microenvironment in the body is dynamic. Here, we established a microfluidic-based in vitro model of human bronchial epithelial cells in which cells are stationary, but nutrient supply is dynamic, and we used this system to evaluate cellular uptake of nanoparticles. The cells were maintained in fetal calf serum-free and bovine pituitary extract-free cell culture medium. BEAS-2B, an immortalized, non-tumorigenic human cell line, was used as a model and the cells were grown in a chip within a microfluidic device and were briefly infused with amorphous silica (SiO2) nanoparticles or polystyrene (PS) nanoparticles of similar primary sizes but with different densities. For comparison, tests were also performed using static, multi-well cultures. Cellular uptake of the fluorescently labeled particles was investigated by flow cytometry and confocal microscopy. Exposure under dynamic culture conditions resulted in higher cellular uptake of the PS nanoparticles when compared to static conditions, while uptake of SiO2 nanoparticles was similar in both settings. The present study has shown that it is feasible to grow human lung cells under completely animal-free conditions using a microfluidic-based device, and we have also found that cellular uptake of PS nanoparticles aka nanoplastics is highly dependent on culture conditions. Hence, traditional cell cultures may not accurately reflect the uptake of low-density particles, potentially leading to an underestimation of their cellular impact.

The effect of drug loading and multiple administration on the protein corona formation and brain delivery property of PEG-PLA nanoparticles

The presence of protein corona on the surface of nanoparticles modulates their physiological interactions such as cellular association and targeting property. It has been shown that α-mangostin (αM)-loaded poly(ethylene glycol)-poly(l-lactide) (PEG-PLA) nanoparticles (NP-αM) specifically increased low density lipoprotein receptor (LDLR) expression in microglia and improved clearance of amyloid beta (Aβ) after multiple administration. However, how do the nanoparticles cross the blood‒brain barrier and access microglia remain unknown. Here, we studied the brain delivery property of PEG-PLA nanoparticles under different conditions, finding that the nanoparticles exhibited higher brain transport efficiency and microglia uptake efficiency after αM loading and multiple administration. To reveal the mechanism, we performed proteomic analysis to characterize the composition of protein corona formed under various conditions, finding that both drug loading and multiple dosing affect the composition of protein corona and subsequently influence the cellular uptake of nanoparticles in b.End3 and BV-2 cells. Complement proteins, immunoglobulins, RAB5A and CD36 were found to be enriched in the corona and associated with the process of nanoparticles uptake. Collectively, we bring a mechanistic understanding about the modulator role of protein corona on targeted drug delivery, and provide theoretical basis for engineering brain or microglia-specific targeted delivery system.

Microvilli Adhesion: An Alternative Route for Nanoparticle Cell Internalization.

The cellular uptake of nanoparticles (NPs) represents a critical step in nanomedicine and a crucial point for understanding the interaction of nanomaterials with biological systems. No specific mechanism of uptake has been identified so far, as the NPs are generally incorporated by the cells through one of the few well-known endocytotic mechanisms. Here, an alternative internalization route mediated by microvilli adhesion is demonstrated. This microvillus-mediated adhesion (MMA) has been observed using ceria and magnetite NPs with a dimension of <40 nm functionalized with polyacrylic acid but not using NPs with a neutral or positive functionalization. Such an adhesion was not cell specific, as it was demonstrated in three different cell lines. MMA was also reduced by modifications of the microvillus lipid rafts, obtained by depleting cholesterol and altering synthesis of sphingolipids. We found a direct relationship between MAA, cell cycle, and density of microvilli. The evidence suggests that MMA differs from the commonly described uptake mechanisms and might represent an interesting alternative approach for selective NP delivery.

Genotoxic Potential of Nanoparticles: Structural and Functional Modifications in DNA.

The rapid advancement of nanotechnology enhances the production of different nanoparticles that meet the demand of various fields like biomedical sciences, industrial, material sciences and biotechnology, etc. This technological development increases the chances of nanoparticles exposure to human beings, which can threaten their health. It is well known that various cellular processes (transcription, translation, and replication during cell proliferation, cell cycle, cell differentiation) in which genetic materials (DNA and RNA) are involved play a vital role to maintain any structural and functional modification into it. When nanoparticles come into the vicinity of the cellular system, chances of uptake become high due to their small size. This cellular uptake of nanoparticles enhances its interaction with DNA, leading to structural and functional modification (DNA damage/repair, DNA methylation) into the DNA. These modifications exhibit adverse effects on the cellular system, consequently showing its inadvertent effect on human health. Therefore, in the present study, an attempt has been made to elucidate the genotoxic mechanism of nanoparticles in the context of structural and functional modifications of DNA.

Sulfated alginate/polycaprolactone double-emulsion nanoparticles for enhanced delivery of heparin-binding growth factors in wound healing applications

Diabetic foot ulcers (DFUs) that are not effectively treated could lead to partial or complete lower limb amputations. The lack of connective tissue growth factor (CTGF) and insulin-like growth factor (IGF-I) in DFUs results in limited matrix deposition and poor tissue repair. To enhance growth factor (GF) availability in DFUs, heparin (HN)-mimetic alginate sulfate/polycaprolactone (AlgSulf/PCL) double emulsion nanoparticles (NPs) with high affinity and sustained release of CTGF and IGF-I were synthesized. The NPs size, encapsulation efficiency (EE), cytotoxicity, cellular uptake and wound healing capacity in immortalized primary human adult epidermal cells (HaCaT) were assessed. The sonication time and amplitude used for NPs synthesis enabled the production of particles with a minimum of 236 ± 25 nm diameter. Treatment of HaCaT cells with up to 50 μg mL−1 of NPs showed no cytotoxic effects after 72 h. The highest bovine serum albumin EE (94.6 %, P = 0.028) and lowest burst release were attained with AlgSulf/PCL. Moreover, cells treated with AlgSulf/CTGF (250 ng mL−1) exhibited the most rapid wound closure compared to controls while maintaining fibronectin synthesis. Double-emulsion NPs based on HN-mimetic AlgSulf represent a novel approach which can significantly enhance diabetic wound healing and can be expanded for applications requiring the delivery of other HN-binding GFs.

Intracellular delivery of anti-BCR/ABL antibody by PLGA nanoparticles suppresses the oncogenesis of chronic myeloid leukemia cells

BackgroundThe pathogenesis of chronic myeloid leukemia (CML) is the formation of the BCR/ABL protein, which is encoded by the bcr/abl fusion gene, possessing abnormal tyrosine kinase activity. Despite the wide application of tyrosine kinase inhibitors (TKIs) in CML treatment, TKIs drug resistance or intolerance limits their further usage in a subset of patients. Furthermore, TKIs inhibit the tyrosine kinase activity of the BCR/ABL oncoprotein while failing to eliminate the pathologenic oncoprotein. To develop alternative strategies for CML treatment using therapeutic antibodies, and to address the issue that antibodies cannot pass through cell membranes, we have established a novel intracellular delivery of anti-BCR/ABL antibodies, which serves as a prerequisite for CML therapy.MethodsAnti-BCR/ABL antibodies were encapsulated in poly(d, l-lactide-co-glycolide) nanoparticles (PLGA NPs) by a double emulsion method, and transferrin was labeled on the surface of the nanoparticles (Ab@Tf-Cou6-PLGA NPs). The characteristics of nanoparticles were measured by dynamic light scattering (DLS) and transmission electron microscopy (TEM). Cellular uptake of nanoparticles was measured by flow cytometry (FCM). The effect of nanoparticles on the apoptosis and proliferation of CML cells was testified by FCM and CCK-8 assay. In addition, the anti-cancer impact of nanoparticles was evaluated in mouse models of CML.ResultsThe results demonstrated that the Ab@Tf-Cou6-PLGA NPs functioned as an intracellular deliverer of antibodies, and exhibited an excellent effect on degrading BCR/ABL oncoprotein in CML cells via the Trim-Away pathway. Treatment with Ab@Tf-Cou6-PLGA NPs inhibited the proliferation and induced the apoptosis of CML cells in vitro as well as impaired the oncogenesis ability of CML cells in vivo.ConclusionsIn conclusion, our study indicated that this approach achieved safe and efficient intracellular delivery of antibodies and degraded BCR/ABL oncoprotein via the Trim-Away pathway, which provides a promising therapeutic strategy for CML patients, particularly those with TKI resistance.

Theoretical Investigation of Photoacoustics From Cancer Cells: Modified Models

Photoacoustic (PA) technology has wide applications in cell study. However, photoacoustics from cancer cells lacks adequate understanding. Here, we establish two modified models for theoretical investigation of photoacoustics from cancer cells. The first model is based on endogenous absorbers of cancer cells, which can be measured by label-free PA microscopy. Different from previous studies about PA for cancer cells that are limited to melanoma cells, our model considers general cancer cells. Further, the feasibility of cancer cell cycle analysis via PA spectrum analysis is demonstrated. The second model is based on exogenous absorbers, e.g., nanoparticles (NPs), in cancer cells. The interactions between NPs and cells in PA signal generation are considered. The nucleus-to-cytoplasm ratio, refractive index, cellular uptake of NPs, NPs size heterogeneity and cell size are modeled to evaluate the contribution of each factor to the change of PA signal amplitudes, which is valuable to guide experiments for cancer cell identification by utilizing different PA signal amplitudes between normal cells and cancer cells. In conclusion, our study comprehensively investigates photoacoustics from cancer cells, which lays theoretical groundwork for future research on PA techniques for cancer cells and is expected to be useful for cancer diagnosis, treatment, and prognosis.

Suitability of the In Vitro Cytokinesis-Block Micronucleus Test for Genotoxicity Assessment of TiO2 Nanoparticles on SH-SY5Y Cells.

Standard toxicity tests might not be fully adequate for evaluating nanomaterials since their unique features are also responsible for unexpected interactions. The in vitro cytokinesis-block micronucleus (CBMN) test is recommended for genotoxicity testing, but cytochalasin-B (Cyt-B) may interfere with nanoparticles (NP), leading to inaccurate results. Our objective was to determine whether Cyt-B could interfere with MN induction by TiO2 NP in human SH-SY5Y cells, as assessed by CBMN test. Cells were treated for 6 or 24 h, according to three treatment options: co-treatment with Cyt-B, post-treatment, and delayed co-treatment. Influence of Cyt-B on TiO2 NP cellular uptake and MN induction as evaluated by flow cytometry (FCMN) were also assessed. TiO2 NP were significantly internalized by cells, both in the absence and presence of Cyt-B, indicating that this chemical does not interfere with NP uptake. Dose-dependent increases in MN rates were observed in CBMN test after co-treatment. However, FCMN assay only showed a positive response when Cyt-B was added simultaneously with TiO2 NP, suggesting that Cyt-B might alter CBMN assay results. No differences were observed in the comparisons between the treatment options assessed, suggesting they are not adequate alternatives to avoid Cyt-B interference in the specific conditions tested.

Lethal effects and ultrastructure of cellular uptake of ingested gold nanoparticles in the freshwater rotifer Brachionus calyciflorus (Monogononta: Brachionidae)

Much of the recent literature concerning the threat posed by anthropogenic microscopic pollution has focussed on marine organisms although freshwater environments face the same degree of pollution and therefore risk. Although several studies have documented the ingestion of nanoparticles (NPs) in species of the pelagic freshwater rotifer genus Brachionus, unambiguous evidence for its cellular uptake in this group remains lacking. We therefore used transmission electron microscopy (TEM) of ultrathin sections through the digestive tract of individuals of Brachionus calyciflorus exposed in vitro to citrate stabilized gold nanoparticles (AuCit NPs) in their culture medium to provide the first concrete evidence for the cellular uptake of NPs in rotifers, a group of organisms that comprise an important part of the zooplankton community. Using this method, AuCit NPs with average diameters of 8.5 ± 1.4 nm and 12.5 ± 1.5 nm could be localized clearly within large vacuoles within the stomach cells. Moreover, the occasional presence of pits containing AuCit NPs in the outer membranes of these cells hints that the particles are taken up by some form of endocytosis. In all cases, the ingestion of AuCit NPs showed lethal effects after only one day with virtually no individuals surviving more than two days of exposure. Combined with the TEM evidence above, we hypothesize that death might derive from some form of lysosomal overload. In total, our results document the potential threat that microscopic pollution also poses for freshwater organisms. Through this, we hope that additional emphasis in this context will be directed toward freshwater environments and the potential for such pollution both to enter as well as to move up the food chain via trophic transfer events.

The system of self-consistent models for the uptake of nanoparticles in PaCa2 cancer cells

Quantitative structure-property/activity relationships (QSPRs/QSARs) are an important component of modern science. Validation of the QSPR/QSAR is the basis for applying. The system of self-consistent models is a new approach to validate QSPR/QSAR. The principle ‘QSAR is a random event’ means that an approach may be recognized as robust only if the statistical characteristics of models obtained by this approach for different splits (training/test) are reproduced. The above principle applies to the case of the nano-QSAR, also. Here, the cellular uptake of nanoparticles in pancreatic cancer cells examines as the endpoint. Groups of models for different splits (training/test) are compared. This comparison gives the possibility to formulate the system of self-consistent models as a way to assess the predictive potential for an arbitrary QSPR/QSAR and/or nano-QSPR/QSAR. The correlation intensity index (CII) has been tested as a tool to improve the quality of models for the cellular uptake of nanoparticles in pancreatic cancer cells (PaCa2). It has shown, that the CII can be useful, but only incorporating with the Index of ideality of correlation (IIC).

Regulating the uptake of poly(N-(2-hydroxypropyl) methacrylamide)-based micelles in cells cultured on micropatterned surfaces.

Cellular uptake of nanoparticles plays a crucial role in cell-targeted biomedical applications. Despite abundant studies trying to understand the interaction between nanoparticles and cells, the influence of cell geometry traits such as cell spreading area and cell shape on the uptake of nanoparticles remains unclear. In this study, poly(vinyl alcohol) is micropatterned on polystyrene cell culture plates using ultraviolet photolithography to control the spreading area and shape of individual cells. The effects of these factors on the cellular uptake of poly(N-(2-hydroxypropyl)methacrylamide)-based micelles were investigated at a single-cell level. Human carcinoma MCF-7 and A549 cells as well as normal Hs-27 and MRC-5 fibroblasts were cultured on micropatterned surfaces. MCF-7 and A549 cells, both with larger sizes, had a higher total micelle uptake. However, the uptake of Hs-27 and MRC-5 cells decreased with increasing spreading area. In terms of cell shapes, MCF-7 and A549 cells with round shapes showed a higher micelle uptake, while those with a square shape had a lower cellular uptake. On the other hand, Hs-27 and MRC-5 cells showed opposite behaviors. The results indicate that the geometry of cells can influence the nanoparticle uptake and may shed light on the design of functional nanoparticles.

Cell Cycle Control of Nanoplastics Internalization in Phytoplankton.

Nanoparticles (NPs) for delivering chemotherapeutic drugs are now in clinical trials, and cellular uptake of NPs plays an important role in determining the drug delivery efficiency. Herein, we reported that the bioaccumulation and internalization of NPs were governed by the cell cycle. Specifically, we found that the bioaccumulation of NPs was more favored in the G2/M stages, followed by the S and G0/G1 stages. We demonstrated that three key parameters-clathrin-mediated endocytosis capacity, algal cell membrane permeability, and exopolymer substance (EPS) thickness-were critical in the bioaccumulation of NPs during the cell cycling process. Over the 24-h average duration of cell cycle, clathrin-mediated endocytosis capacity was much higher at the S stage than that at the G0/G1 and G2/M stages. Besides, cell membrane permeability was measured to be higher in S and G2/M stages while the lowest in G0/G1 stage. We have also identified the change of EPS thickness during the 24-h cell cycle. Transition from G0/G1 to S and G2/M induced the attenuation in EPS thickness, and the thinnest EPS was found at the end of mitosis. The cell cycle control NPs internalization were further verified by exposing Ag nanoparticles to algae at different cell cycle stages, confirming the important roles of EPS thickness and cell cycle control in the dynamic internalization processes. The present study highlights the important roles of cell cycle controlling the NPs bioaccumulation and internalization, with possible implications in maximizing NPs internalization efficiency while reducing the cost.

Supramolecular Nanosubstrate-Mediated Delivery for CRISPR/Cas9 Gene Disruption and Deletion.

The clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein 9 (CRISPR/Cas9) is an efficient and precise gene-editing technology that offers a versatile solution for establishing treatments directed at genetic diseases. Currently, CRISPR/Cas9 delivery into cells relies primarily on viral vectors, which suffer from limitations in packaging capacity and safety concerns. These issues with a nonviral delivery strategy are addressed, where Cas9•sgRNA ribonucleoprotein (RNP) complexes can be encapsulated into supramolecular nanoparticles (SMNP) to form RNP⊂SMNPs, which can then be delivered into targeted cells via supramolecular nanosubstrate-mediated delivery. Utilizing the U87 glioblastoma cell line as a model system, a variety of parameters for cellular-uptake of the RNP-laden nanoparticles are examined. Dose- and time-dependent CRISPR/Cas9-mediated gene disruption is further examined in a green fluorescent protein (GFP)-expressing U87 cell line (GFP-U87). The utility of an optimized SMNP formulation in co-delivering Cas9 protein and two sgRNAs that target deletion of exons 45-55 (708 kb) of the dystrophin gene is demonstrated. Mutations in this region lead to Duchenne muscular dystrophy, a severe genetic muscle wasting disease. Efficient delivery of these gene deletion cargoes is observed in a human cardiomyocyte cell line (AC16), induced pluripotent stem cells, and mesenchymal stem cells.

Sonoporation Using Nanoparticle-Loaded Microbubbles Increases Cellular Uptake of Nanoparticles Compared to Co-Incubation of Nanoparticles and Microbubbles.

Therapeutic agents can benefit from encapsulation in nanoparticles, due to improved pharmacokinetics and biodistribution, protection from degradation, increased cellular uptake and sustained release. Microbubbles in combination with ultrasound have been shown to improve the delivery of nanoparticles and drugs to tumors and across the blood-brain barrier. Here, we evaluate two different microbubbles for enhancing the delivery of polymeric nanoparticles to cells in vitro: a commercially available lipid microbubble (Sonazoid) and a microbubble with a shell composed of protein and nanoparticles. Various ultrasound parameters are applied and confocal microscopy is employed to image cellular uptake. Ultrasound enhanced cellular uptake depending on the pressure and duty cycle. The responsible mechanisms are probably sonoporation and sonoprinting, followed by uptake, and to a smaller degree enhanced endocytosis. The use of commercial Sonazoid microbubbles leads to significantly lower uptake than when using nanoparticle-loaded microbubbles, suggesting that proximity between cells, nanoparticles and microbubbles is important, and that mainly nanoparticles in the shell are taken up, rather than free nanoparticles in solution.

Cellular uptake of nanoparticles: Involvement of caveolae?

&lt;img src=” https://s3.amazonaws.com/production.scholastica/article/22201/large/1416290_background_image.jpg?1618497820”&gt; Here we discuss some pitfalls and challenges briefly when investigating which endocytic mechanisms are involved in the cellular uptake of nanoparticles. We specifically discuss some common misunderstandings regarding studies claimed to demonstrate uptake via caveolae. Scientists in the nanomedicine field should be aware that reducing the membrane content of cholesterol by adding methyl-β-cyclodextrin removes caveolae and inhibits other uptake mechanisms, such as macropinocytosis as well. Furthermore, the general tyrosine kinase inhibitor genistein is not a specific inhibitor of uptake from caveolae. Moreover, one can still see that scientists in the field write that they want to direct transport of their particles to caveolae and caveosomes to avoid lysosomal degradation. However, caveosomes are artifacts caused by overexpression of caveolin-1 constructs, and ligands or particles taken up by caveolae are transported to endosomes and lysosomes as reported for other types of endocytosis.

Transportan Peptide Stimulates the Nanomaterial Internalization into Mammalian Cells in the Bystander Manner through Macropinocytosis.

Covalent coupling with cell-penetrating peptides (CPPs) has been a common strategy to facilitate the cell entry of nanomaterial and other macromolecules. Though efficient, this strategy requires chemical modifications on nanomaterials, which is not always desired for their applications. Recent studies on a few cationic CPPs have revealed that they can stimulate the cellular uptake of nanoparticles (NPs) simply via co-administration (bystander manner), which bypasses the requirement of chemical modification. In this study, we investigated the other classes of CPPs and discovered that transportan (TP) peptide, an amphiphilic CPP, also exhibited such bystander activities. When simply co-administered, TP peptide enabled the cells to engulf a variety of NPs, as well as common solute tracers, while these payloads had little or no ability to enter the cells by themselves. This result was validated in vitro and ex vivo, and TP peptide showed no physical interaction with co-administered NPs (bystander cargo). We further explored the cell entry mechanism for TP peptide and its bystander cargo, and showed that it was mediated by a receptor-dependent macropinocytosis process. Together, our findings improve the understanding of TP-assisted cell entry, and open up a new avenue to apply this peptide for nanomaterial delivery.

Stabilin-1 is required for the endothelial clearance of small anionic nanoparticles

Clearance of nanoparticles (NPs) after intravenous injection – mainly by the liver – is a critical barrier for the clinical translation of nanomaterials. Physicochemical properties of NPs are known to influence their distribution through cell-specific interactions; however, the molecular mechanisms responsible for liver cellular NP uptake are poorly understood. Liver sinusoidal endothelial cells and Kupffer cells are critical participants in this clearance process. Here we use a zebrafish model for liver-NP interaction to identify the endothelial scavenger receptor Stabilin-1 as a non-redundant receptor for the clearance of small anionic NPs. Furthermore, we show that physiologically, Stabilin-1 is required for the removal of bacterial lipopolysaccharide (LPS/endotoxin) from circulation and that Stabilin-1 cooperates with its homolog Stabilin-2 in the clearance of larger (~100 nm) anionic NPs. Our findings allow optimization of anionic nanomedicine biodistribution and targeting therapies that use Stabilin-1 and -2 for liver endothelium-specific delivery.

Nanoencapsulation as a novel delivery approach for therapeutic applications of gla-rich protein (GRP)

Introduction Gla rich protein (GRP) is a vitamin K dependent protein, shown to function as an inhibitor of pathological calcification and as an anti-inflammatory agent, with potential therapeutic use for age-related diseases such as osteoarthritis (OA) [1,2]. OA is a leading cause of disability and morbidity in the older population and constitutes a major worldwide challenge for our health system. Presently, there are no drugs approved that can prevent, stop, or even restrain progression of OA. GRP has been shown to be able to lower inflammation and mineralisation processes in the articular tissue. Chitosan/tripolyphosphate (TPP) nanoparticles were selected for this study due to their biocompatibility, biodegradability and capacity to overcome the problem of low solubility of GRP in physiological conditions. This study aims to produce and characterise chitosan/TPP nanoparticles as GRP-delivery vehicles and test its anti-inflammatory potential in human macrophages. Materials and methods Nanoparticles of fluorescein-labelled chitosan/TPP with and without GRP (NG and NP, respectively) were prepared by ionic gelation [3]. Resulting NP and NG were characterised by dynamic light scattering, transmission electron microscopy (TEM) and flow cytometry. The anti-inflammatory activity of NP and NG was assessed in THP-1 cells differentiated to macrophages. Mac-THP-1 cells were pre-treated with both NP and NG, followed by LPS stimulation. Cell viability was assessed by the MTS cell proliferation assay, and levels of TNFα released to cell culture media were determined by ELISA. Results The average size determined for NG was increased relatively to the NP, while flow cytometry and TEM analysis indicate the presence of GRP in NG, suggesting an effective incorporation of human recombinant GRP. Flow cytometry studies confirmed the cellular uptake of nanoparticles by macrophages. The GRP-loaded nanoparticles were able to reduce the production of TNFα in LPS-stimulated macrophages. Discussion and conclusions The results confirm that chitosan/TPP nanoparticles are excellent drug delivery vehicles for GRP in macrophages and predict a wider therapeutic application in chronic inflammation-related diseases. GRP-containing nanoparticles will be further used in OA functional assays and the results will bring new knowledge on the role of GRP in the interplay between inflammation and mineralisation events associated with OA.

Sub-10 nm Substrate Roughness Promotes the Cellular Uptake of Nanoparticles by Upregulating Endocytosis-Related Genes.

Nanosubstrate engineering is an established approach for modulating cellular responses, but it remains infrequently exploited to facilitate the intracellular delivery of nanoparticles (NPs). We report nanoscale roughness of the extracellular environment as a critical parameter for regulating the cellular uptake of NPs. After seeding cells atop a substrate that contains randomly immobilized gold NPs (termed AuNP-S) with sub-10 nm surface roughness, we demonstrate that such cells internalize up to ∼100-fold more poly(ethylene glycol)-coated AuNPs (Au@PEG NPs) than those cells seeded on a conventional flat culture plate. Our result is generalizable to 4 different cell types and Au@PEG NPs modified with 13 different hydrocarbyl functional groups. Conditioning cells to AuNP-S not only leads to upregulation of clathrin- and integrin-related genes, but also supports elevated uptake of Au@PEG NPs via clathrin-mediated endocytosis. Our data suggest a simple and robust method for boosting the intracellular delivery of nanomedicines by nanosubstrate engineering.