Publications by authors named "Kostas Kostarelos"

196 Publications

Transient reprogramming of postnatal cardiomyocytes to a dedifferentiated state.

PLoS One 2021 5;16(5):e0251054. Epub 2021 May 5.

Nanomedicine Lab, Faculty of Biology, Medicine and Health, The University of Manchester, Manchester, United Kingdom.

In contrast to mammals, lower vertebrates are capable of extraordinary myocardial regeneration thanks to the ability of their cardiomyocytes to undergo transient dedifferentiation and proliferation. Somatic cells can be temporarily reprogrammed to a proliferative, dedifferentiated state through forced expression of Oct3/4, Sox2, Klf4 and c-Myc (OSKM). Here, we aimed to induce transient reprogramming of mammalian cardiomyocytes in vitro utilising an OSKM-encoding non-integrating vector. Reprogramming factor expression in postnatal rat and mouse cardiomyocytes triggered rapid but limited cell dedifferentiation. Concomitantly, a significant increase in cell viability, cell cycle related gene expression and Ki67 positive cells was observed consistent with an enhanced cell cycle activation. The transient nature of this partial reprogramming was confirmed as cardiomyocyte-specific cell morphology, gene expression and contractile activity were spontaneously recovered by day 15 after viral transduction. This study provides the first evidence that adenoviral OSKM delivery can induce partial reprogramming of postnatal cardiomyocytes. Therefore, adenoviral mediated transient reprogramming could be a novel and feasible strategy to recapitulate the regenerative mechanisms of lower vertebrates.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0251054PLOS
May 2021

Nanoparticle-Enabled Enrichment of Longitudinal Blood Proteomic Fingerprints in Alzheimer's Disease.

ACS Nano 2021 04 17;15(4):7357-7369. Epub 2021 Mar 17.

Nanomedicine Lab, School of Health Sciences, Faculty of Biology, Medicine and Health, The University of Manchester, Manchester M13 9PT, United Kingdom.

Blood-circulating biomarkers have the potential to detect Alzheimer's disease (AD) pathology before clinical symptoms emerge and to improve the outcomes of clinical trials for disease-modifying therapies. Despite recent advances in understanding concomitant systemic abnormalities, there are currently no validated or clinically used blood-based biomarkers for AD. The extremely low concentration of neurodegeneration-associated proteins in blood necessitates the development of analytical platforms to address the "signal-to-noise" issue and to allow an in-depth analysis of the plasma proteome. Here, we aimed to discover and longitudinally track alterations of the blood proteome in a transgenic mouse model of AD, using a nanoparticle-based proteomics enrichment approach. We employed blood-circulating, lipid-based nanoparticles to extract, analyze and monitor AD-specific protein signatures and to systemically uncover molecular pathways associated with AD progression. Our data revealed the existence of multiple proteomic signals in blood, indicative of the asymptomatic stages of AD. Comprehensive analysis of the nanoparticle-recovered blood proteome by label-free liquid chromatography-tandem mass spectrometry resulted in the discovery of AD-monitoring signatures that could discriminate the asymptomatic phase from amyloidopathy and cognitive deterioration. While the majority of differentially abundant plasma proteins were found to be upregulated at the initial asymptomatic stages, the abundance of these molecules was significantly reduced as a result of amyloidosis, suggesting a disease-stage-dependent fluctuation of the AD-specific blood proteome. The potential use of the proposed nano-omics approach to uncover information in the blood that is directly associated with brain neurodegeneration was further exemplified by the recovery of focal adhesion cascade proteins. We herein propose the integration of nanotechnology with already existing proteomic analytical tools in order to enrich the identification of blood-circulating signals of neurodegeneration, reinvigorating the potential clinical utility of the blood proteome at predicting the onset and kinetics of the AD progression trajectory.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acsnano.1c00658DOI Listing
April 2021

Graphene oxide prevents lateral amygdala dysfunctional synaptic plasticity and reverts long lasting anxiety behavior in rats.

Biomaterials 2021 Apr 4;271:120749. Epub 2021 Mar 4.

Neuron Physiology and Technology Lab, International School for Advanced Studies (SISSA), Neuroscience, Via Bonomea 265, 34136, Trieste, Italy. Electronic address:

Engineered small graphene oxide (s-GO) sheets were previously shown to reversibly down-regulate glutamatergic synapses in the hippocampus of juvenile rats, disclosing an unexpected translational potential of these nanomaterials to target selective synapses in vivo. Synapses are anatomical specializations acting in the Central Nervous System (CNS) as functional interfaces among neurons. Dynamic changes in synaptic function, named synaptic plasticity, are crucial to learning and memory. More recently, pathological mechanisms involving dysfunctional synaptic plasticity were implicated in several brain diseases, from dementia to anxiety disorders. Hyper-excitability of glutamatergic neurons in the lateral nucleus of the amygdala complex (LA) is substantially involved in the storage of aversive memory induced by stressful events enabling post-traumatic stress disorder (PTSD). Here we translated in PTSD animal model the ability of s-GO, when stereotaxically administered to hamper LA glutamatergic transmission and to prevent the behavioral response featured in long-term aversive memory. We propose that s-GO, by interference with glutamatergic plasticity, impair LA-dependent memory retrieval related to PTSD.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.biomaterials.2021.120749DOI Listing
April 2021

Trends in Micro-/Nanorobotics: Materials Development, Actuation, Localization, and System Integration for Biomedical Applications.

Adv Mater 2021 Jan 4;33(4):e2002047. Epub 2020 Dec 4.

Department of Mechanical and Automation Engineering, The Chinese University of Hong Kong, Shatin N.T., Hong Kong, China.

Micro-/nanorobots (m-bots) have attracted significant interest due to their suitability for applications in biomedical engineering and environmental remediation. Particularly, their applications in in vivo diagnosis and intervention have been the focus of extensive research in recent years with various clinical imaging techniques being applied for localization and tracking. The successful integration of well-designed m-bots with surface functionalization, remote actuation systems, and imaging techniques becomes the crucial step toward biomedical applications, especially for the in vivo uses. This review thus addresses four different aspects of biomedical m-bots: design/fabrication, functionalization, actuation, and localization. The biomedical applications of the m-bots in diagnosis, sensing, microsurgery, targeted drug/cell delivery, thrombus ablation, and wound healing are reviewed from these viewpoints. The developed biomedical m-bot systems are comprehensively compared and evaluated based on their characteristics. The current challenges and the directions of future research in this field are summarized.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/adma.202002047DOI Listing
January 2021

Enhanced liquid phase exfoliation of graphene in water using an insoluble bis-pyrene stabiliser.

Faraday Discuss 2021 Apr 9;227:46-60. Epub 2020 Dec 9.

Department of Chemistry, University of Manchester, Oxford Road, Manchester, M13 9PL, UK.

Stabilisers, such as surfactants, polymers and polyaromatic molecules, offer an effective way to produce graphene dispersions in water by Liquid Phase Exfoliation (LPE) without degrading the properties of graphene. In particular, pyrene derivatives provide better exfoliation efficiency than traditional surfactants and polymers. A stabiliser is expected to be relatively soluble in order to disperse hydrophobic graphene in water. Here, we show that exfoliation can also be achieved with insoluble pyrene stabilisers if appropriately designed. In particular, bis-pyrene stabilisers (BPSs) functionalised with pyrrolidine provide a higher exfoliation efficiency and percentage of single layers compared to traditional pyrene derivatives under the same experimental conditions. This is attributed to the enhanced interactions between BPS and graphene, provided by the presence of two pyrene binding groups. This approach is therefore attractive not only to produce highly concentrated graphene, but also to use graphene to disperse insoluble molecules in water. The enhanced adsorption of BPS on graphene, however, is reflected in higher toxicity towards human epithelial bronchial immortalized cells, limiting the use of this material for biomedical applications.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1039/c9fd00114jDOI Listing
April 2021

Multiparametric Profiling of Engineered Nanomaterials: Unmasking the Surface Coating Effect.

Adv Sci (Weinh) 2020 Nov 11;7(22):2002221. Epub 2020 Oct 11.

Institute of Environmental Medicine Karolinska Institutet Stockholm 171 77 Sweden.

Despite considerable efforts, the properties that drive the cytotoxicity of engineered nanomaterials (ENMs) remain poorly understood. Here, the authors inverstigate a panel of 31 ENMs with different core chemistries and a variety of surface modifications using conventional in vitro assays coupled with omics-based approaches. Cytotoxicity screening and multiplex-based cytokine profiling reveals a good concordance between primary human monocyte-derived macrophages and the human monocyte-like cell line THP-1. Proteomics analysis following a low-dose exposure of cells suggests a nonspecific stress response to ENMs, while microarray-based profiling reveals significant changes in gene expression as a function of both surface modification and core chemistry. Pathway analysis highlights that the ENMs with cationic surfaces that are shown to elicit cytotoxicity downregulated DNA replication and cell cycle responses, while inflammatory responses are upregulated. These findings are validated using cell-based assays. Notably, certain small, PEGylated ENMs are found to be noncytotoxic yet they induce transcriptional responses reminiscent of viruses. In sum, using a multiparametric approach, it is shown that surface chemistry is a key determinant of cellular responses to ENMs. The data also reveal that cytotoxicity, determined by conventional in vitro assays, does not necessarily correlate with transcriptional effects of ENMs.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/advs.202002221DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7675037PMC
November 2020

Nanotools for Sepsis Diagnosis and Treatment.

Adv Healthc Mater 2021 01 25;10(1):e2001378. Epub 2020 Nov 25.

Nanomedicine Lab, Faculty of Biology, Medicine and Health, AV Hill Building, The University of Manchester, Manchester, M13 9PT, UK.

Sepsis is one of the leading causes of death worldwide with high mortality rates and a pathological complexity hindering early and accurate diagnosis. Today, laboratory culture tests are the epitome of pathogen recognition in sepsis. However, their consistency remains an issue of controversy with false negative results often observed. Clinically used blood markers, C reactive protein (CRP) and procalcitonin (PCT) are indicators of an acute-phase response and thus lack specificity, offering limited diagnostic efficacy. In addition to poor diagnosis, inefficient drug delivery and the increasing prevalence of antibiotic-resistant microorganisms constitute significant barriers in antibiotic stewardship and impede effective therapy. These challenges have prompted the exploration for alternative strategies that pursue accurate diagnosis and effective treatment. Nanomaterials are examined for both diagnostic and therapeutic purposes in sepsis. The nanoparticle (NP)-enabled capture of sepsis causative agents and/or sepsis biomarkers in biofluids can revolutionize sepsis diagnosis. From the therapeutic point of view, currently existing nanoscale drug delivery systems have proven to be excellent allies in targeted therapy, while many other nanotherapeutic applications are envisioned. Herein, the most relevant applications of nanomedicine for the diagnosis, prognosis, and treatment of sepsis is reviewed, providing a critical assessment of their potentiality for clinical translation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/adhm.202001378DOI Listing
January 2021

Intracerebral Injection of Graphene Oxide Nanosheets Mitigates Microglial Activation Without Inducing Acute Neurotoxicity: A Pilot Comparison to Other Nanomaterials.

Small 2020 Nov 10:e2004029. Epub 2020 Nov 10.

Nanomedicine Lab, School of Health Sciences, Faculty of Biology, Medicine and Health, The University of Manchester, Manchester, M13 9PL, UK.

Carbon-based nanomaterials (CNMs) are being explored for neurological applications. However, systematic in vivo studies investigating the effects of CNM nanocarriers in the brain and how brain cells respond to such nanomaterials are scarce. To address this, functionalized multiwalled carbon nanotubes and graphene oxide (GO) sheets are injected in mice brain and compared with charged liposomes. The induction of acute neuroinflammatory and neurotoxic effects locally and in brain structures distant from the injection site are assessed up to 1 week postadministration. While significant neuronal cell loss and sustained microglial cell activation are observed after injection of cationic liposomes, none of the tested CNMs induces either neurodegeneration or microglial activation. Among the candidate nanocarriers tested, GO sheets appear to elicit the least deleterious neuroinflammatory profile. At molecular level, GO induces moderate activation of proinflammatory markers compared to vehicle control. At histological level, brain response to GO is lower than after vehicle control injection, suggesting some capacity for GO to reduce the impact of stereotactic injection on brain. While these findings are encouraging and valuable in the selection and design of nanomaterial-based brain delivery systems, they warrant further investigations to better understand the mechanisms underlying GO immunomodulatory properties in brain.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/smll.202004029DOI Listing
November 2020

The biomolecule corona of lipid nanoparticles contains circulating cell-free DNA.

Nanoscale Horiz 2020 11 27;5(11):1476-1486. Epub 2020 Aug 27.

Nanomedicine Lab, Faculty of Biology, Medicine and Health, The University of Manchester, AV Hill Building, Manchester, UK.

The spontaneous adsorption of biomolecules onto the surface of nanoparticles (NPs) in complex physiological biofluids has been widely investigated over the last decade. Characterisation of the protein composition of the 'biomolecule corona' has dominated research efforts, whereas other classes of biomolecules, such as nucleic acids, have received no interest. Scarce, speculative statements exist in the literature about the presence of nucleic acids in the biomolecule corona, with no previous studies attempting to describe the contribution of genomic content to the blood-derived NP corona. Herein, we provide the first experimental evidence of the interaction of circulating cell-free DNA (cfDNA) with lipid-based NPs upon their incubation with human plasma samples, obtained from healthy volunteers and ovarian carcinoma patients. Our results also demonstrate an increased amount of detectable cfDNA in patients with cancer. Proteomic analysis of the same biomolecule coronas revealed the presence of histone proteins, suggesting an indirect, nucleosome-mediated NP-cfDNA interaction. The finding of cfDNA as part of the NP corona, offers a previously unreported new scope regarding the chemical composition of the 'biomolecule corona' and opens up new possibilities for the potential exploitation of the biomolecule corona for the enrichment and analysis of blood-circulating nucleic acids.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1039/d0nh00333fDOI Listing
November 2020

Nitric oxide-dependent biodegradation of graphene oxide reduces inflammation in the gastrointestinal tract.

Nanoscale 2020 Aug;12(32):16730-16737

Institute of Environmental Medicine, Karolinska Institutet, Stockholm, Sweden.

Understanding the biological fate of graphene-based materials such as graphene oxide (GO) is crucial to assess adverse effects following intentional or inadvertent exposure. Here we provide first evidence of biodegradation of GO in the gastrointestinal tract using zebrafish as a model. Raman mapping was deployed to assess biodegradation. The degradation was blocked upon knockdown of nos2a encoding the inducible nitric oxide synthase (iNOS) or by pharmacological inhibition of NOS using l-NAME, demonstrating that the process was nitric oxide (NO)-dependent. NO-dependent degradation of GO was further confirmed in vitro by combining a superoxide-generating system, xanthine/xanthine oxidase (X/XO), with an NO donor (PAPA NONOate), or by simultaneously producing superoxide and NO by decomposition of SIN-1. Finally, by using the transgenic strain Tg(mpx:eGFP) to visualize the movement of neutrophils, we could show that inhibition of the degradation of GO resulted in increased neutrophil infiltration into the gastrointestinal tract, indicative of inflammation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1039/d0nr03675gDOI Listing
August 2020

The challenge of recognising sepsis: Future nanotechnology solutions.

J Intensive Care Soc 2020 Aug 18;21(3):241-246. Epub 2019 Dec 18.

Department of Critical Care, Salford Royal Foundation Trust, Salford, UK.

The urgent need to start anti-infective therapeutic interventions in suspected sepsis, and the lack of specific time-critical diagnostic information often lead to the widespread administration of broad-spectrum antimicrobial therapies, increasing the risk of unwanted patient harms and contributing to rising pathogen antimicrobial resistance. Nanotechnology, which involves engineering at the nanoscale, allows for the bespoke development of diagnostic solutions with multi-functionality and high sensitivity that has the potential to help provide time-critical information to make more accurate diagnoses and treatment decisions for sepsis. Nanotechnologies also have the potential to improve upon the current strategies used for novel biomarker discovery. Here we describe some of the current limitations to identifying sepsis and explore the potential role for nanotechnology solutions.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1177/1751143719896554DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7401438PMC
August 2020

Splenic Capture and Intracellular Biodegradation of Biological-Grade Graphene Oxide Sheets.

ACS Nano 2020 08 28;14(8):10168-10186. Epub 2020 Jul 28.

Nanomedicine Lab, National Graphene Institute and Faculty of Biology, Medicine & Health, The University of Manchester, Manchester, M13 9PT, United Kingdom.

Carbon nanomaterials, including 2D graphene-based materials, have shown promising applicability to drug delivery, tissue engineering, diagnostics, and various other biomedical areas. However, to exploit the benefits of these materials in some of the areas mentioned, it is necessary to understand their possible toxicological implications and long-term fate . We previously demonstrated that following intravenous administration, 2D graphene oxide (GO) nanosheets were largely excreted the kidneys; however, a small but significant portion of the material was sequestered in the spleen. Herein, we interrogate the potential consequences of this accumulation and the fate of the spleen-residing GO over a period of nine months. We show that our thoroughly characterized GO materials are not associated with any detectable pathological consequences in the spleen. Using confocal Raman mapping of tissue sections, we determine the sub-organ biodistribution of GO at various time points after administration. The cells largely responsible for taking up the material are confirmed using immunohistochemistry coupled with Raman spectroscopy, and transmission electron microscopy (TEM). This combination of techniques identified cells of the splenic marginal zone as the main site of GO bioaccumulation. In addition, through analyses using both bright-field TEM coupled with electron diffraction and Raman spectroscopy, we reveal direct evidence of intracellular biodegradation of GO sheets with ultrastructural precision. This work offers critical information about biological processing and degradation of thin GO sheets by normal mammalian tissue, indicating that further development and exploitation of GO in biomedicine would be possible.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acsnano.0c03438DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7458483PMC
August 2020

Size-Dependent Pulmonary Impact of Thin Graphene Oxide Sheets in Mice: Toward Safe-by-Design.

Adv Sci (Weinh) 2020 Jun 7;7(12):1903200. Epub 2020 May 7.

Nanomedicine Lab Faculty of Biology, Medicine and Health University of Manchester Manchester Academic Health Science Centre Manchester M13 9PT UK.

Safety assessment of graphene-based materials (GBMs) including graphene oxide (GO) is essential for their safe use across many sectors of society. In particular, the link between specific material properties and biological effects needs to be further elucidated. Here, the effects of lateral dimensions of GO sheets in acute and chronic pulmonary responses after single intranasal instillation in mice are compared. Micrometer-sized GO induces stronger pulmonary inflammation than nanometer-sized GO, despite reduced translocation to the lungs. Genome-wide RNA sequencing also reveals distinct size-dependent effects of GO, in agreement with the histopathological results. Although large GO, but not the smallest GO, triggers the formation of granulomas that persists for up to 90 days, no pulmonary fibrosis is observed. These latter results can be partly explained by Raman imaging, which evidences the progressive biotransformation of GO into less graphitic structures. The findings demonstrate that lateral dimensions play a fundamental role in the pulmonary response to GO, and suggest that airborne exposure to micrometer-sized GO should be avoided in the production plant or applications, where aerosolized dispersions are likely to occur. These results are important toward the implementation of a safer-by-design approach for GBM products and applications, for the benefit of workers and end-users.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/advs.201903200DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7312279PMC
June 2020

Graphene oxide nanosheets modulate spinal glutamatergic transmission and modify locomotor behaviour in an in vivo zebrafish model.

Nanoscale Horiz 2020 08 19;5(8):1250-1263. Epub 2020 Jun 19.

Department of Neuroscience, Psychology and Behaviour, College of Life Sciences, University of Leicester, Leicester, LE1 7RH, UK.

Graphene oxide (GO), an oxidised form of graphene, is widely used for biomedical applications, due to its dispersibility in water and simple surface chemistry tunability. In particular, small (less than 500 nm in lateral dimension) and thin (1-3 carbon monolayers) graphene oxide nanosheets (s-GO) have been shown to selectively inhibit glutamatergic transmission in neuronal cultures in vitro and in brain explants obtained from animals injected with the nanomaterial. This raises the exciting prospect that s-GO can be developed as a platform for novel nervous system therapeutics. It has not yet been investigated whether the interference of the nanomaterial with neurotransmission may have a downstream outcome in modulation of behaviour depending specifically on the activation of those synapses. To address this problem we use early stage zebrafish as an in vivo model to study the impact of s-GO on nervous system function. Microinjection of s-GO into the embryonic zebrafish spinal cord selectively reduces the excitatory synaptic transmission of the spinal network, monitored in vivo through patch clamp recordings, without affecting spinal cell survival. This effect is accompanied by a perturbation in the swimming activity of larvae, which is the locomotor behaviour generated by the neuronal network of the spinal cord. Such results indicate that the impact of s-GO on glutamate based neuronal transmission is preserved in vivo and can induce changes in animal behaviour. These findings pave the way for use of s-GO as a modulator of nervous system function.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1039/c9nh00777fDOI Listing
August 2020

Stable, concentrated, biocompatible, and defect-free graphene dispersions with positive charge.

Nanoscale 2020 Jun 3;12(23):12383-12394. Epub 2020 Jun 3.

Department of Chemistry, University of Manchester, Oxford Road, Manchester, UK.

The outstanding properties of graphene offer high potential for biomedical applications. In this framework, positively charged nanomaterials show better interactions with the biological environment, hence there is strong interest in the production of positively charged graphene nanosheets. Currently, production of cationic graphene is either time consuming or producing dispersions with poor stability, which strongly limit their use in the biomedical field. In this study, we made a family of new cationic pyrenes, and have used them to successfully produce water-based, highly concentrated, stable, and defect-free graphene dispersions with positive charge. The use of different pyrene derivatives as well as molecular dynamics simulations allowed us to get insights on the nanoscale interactions required to achieve efficient exfoliation and stabilisation. The cationic graphene dispersions show outstanding biocompatibility and cellular uptake as well as exceptional colloidal stability in the biological medium, making this material extremely attractive for biomedical applications.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1039/d0nr02689aDOI Listing
June 2020

Protein corona fingerprinting to differentiate sepsis from non-infectious systemic inflammation.

Nanoscale 2020 May;12(18):10240-10253

Nanomedicine Lab, Faculty of Biology, Medicine & Health, AV Hill Building, The University of Manchester, Manchester, M13 9PT, UK.

Rapid and accurate diagnosis of sepsis remains clinically challenging. The lack of specific biomarkers that can differentiate sepsis from non-infectious systemic inflammatory diseases often leads to excessive antibiotic treatment. Novel diagnostic tests are urgently needed to rapidly and accurately diagnose sepsis and enable effective treatment. Despite investment in cutting-edge technologies available today, the discovery of disease-specific biomarkers in blood remains extremely difficult. The highly dynamic environment of plasma restricts access to vital diagnostic information that can be obtained by proteomic analysis. Here, we employed clinically used lipid-based nanoparticles (AmBisome®) as an enrichment platform to analyze the human plasma proteome in the setting of sepsis. We exploited the spontaneous interaction of plasma proteins with nanoparticles (NPs) once in contact, called the 'protein corona', to discover previously unknown disease-specific biomarkers for sepsis diagnosis. Plasma samples obtained from non-infectious acute systemic inflammation controls and sepsis patients were incubated ex vivo with AmBisome® liposomes, and the resultant protein coronas were thoroughly characterised and compared by mass spectrometry (MS)-based proteomics. Our results demonstrate that the proposed nanoparticle enrichment technology enabled the discovery of 67 potential biomarker proteins that could reproducibly differentiate non-infectious acute systemic inflammation from sepsis. This study provides proof-of-concept evidence that nanoscale-based 'omics' enrichment technologies have the potential to substantially improve plasma proteomics analysis and to uncover novel biomarkers in a challenging clinical setting.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1039/d0nr02788jDOI Listing
May 2020

Nanoscale nights of COVID-19.

Nat Nanotechnol 2020 05;15(5):343-344

Nanomedicine Lab, The University of Manchester, Manchester, UK.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41565-020-0687-4DOI Listing
May 2020

Next-Generation Sequencing Reveals Differential Responses to Acute versus Long-Term Exposures to Graphene Oxide in Human Lung Cells.

Small 2020 05 29;16(21):e1907686. Epub 2020 Mar 29.

Nanosafety and Nanomedicine Laboratory, Division of Molecular Toxicology, Institute of Environmental Medicine, Karolinska Institutet, Stockholm, 171 77, Sweden.

Numerous studies have addressed the biological impact of graphene-based materials including graphene oxide (GO), yet few have focused on long-term effects. Here, RNA sequencing is utilized to unearth responses of human lung cells to GO. To this end, the BEAS-2B cell line derived from normal human bronchial epithelium is subjected to repeated, low-dose exposures of GO (1 or 5 µg mL ) for 28 days or to the equivalent, cumulative amount of GO for 48 h. Then, samples are analyzed by using the NovaSeq 6000 sequencing system followed by pathway analysis and gene ontology enrichment analysis of the differentially expressed genes. Significant differences are seen between the low-dose, long-term exposures and the high-dose, short-term exposures. Hence, exposure to GO for 48 h results in mitochondrial dysfunction. In contrast, exposure to GO for 28 days is characterized by engagement of apoptosis pathways with downregulation of genes belonging to the inhibitor of apoptosis protein (IAP) family. Validation experiments confirm that long-term exposure to GO affects the apoptosis threshold in lung cells, accompanied by a loss of IAPs. These studies reveal the sensitivity of RNA-sequencing approaches and show that acute exposure to GO is not a good predictor of the long-term effects of GO.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/smll.201907686DOI Listing
May 2020

Thin graphene oxide nanoflakes modulate glutamatergic synapses in the amygdala cultured circuits: Exploiting synaptic approaches to anxiety disorders.

Nanomedicine 2020 06 5;26:102174. Epub 2020 Mar 5.

International School for Advanced Studies, SISSA, Trieste, Italy. Electronic address:

Anxiety disorders (ADs) are nervous system maladies involving changes in the amygdala synaptic circuitry, such as an upregulation of excitatory neurotransmission at glutamatergic synapses. In the field of nanotechnology, thin graphene oxide flakes with nanoscale lateral size (s-GO) have shown outstanding promise for the manipulation of excitatory neuronal transmission with high temporal and spatial precision, thus they were considered as ideal candidates for modulating amygdalar glutamatergic transmission. Here, we validated an in vitro model of amygdala circuitry as a screening tool to target synapses, towards development of future ADs treatments. After one week in vitro, dissociated amygdalar neurons reconnected forming functional networks, whose development recapitulated that of the tissue of origin. When acutely applied to these cultures, s-GO flakes induced a selective modification of excitatory activity. This type of interaction between s-GO and amygdalar neurons may form the basis for the exploitation of alternative approaches in the treatment of ADs.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.nano.2020.102174DOI Listing
June 2020

Grouping all carbon nanotubes into a single substance category is scientifically unjustified.

Nat Nanotechnol 2020 03;15(3):164

University of Manchester, Manchester, UK.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41565-020-0654-0DOI Listing
March 2020

Hampering brain tumor proliferation and migration using peptide nanofiber:si/ complexes.

Nanomedicine (Lond) 2019 12;14(24):3127-3142

Nanomedicine Lab, Faculty of Biology, Medicine & Health, The University of Manchester, AV Hill Building, Manchester, M13 9PT, UK.

To develop a nonviral tool for the delivery of siRNA to brain tumor cells using peptide nanofibers (PNFs). Uptake of PNFs was evaluated by confocal microscopy and flow cytometry. Gene silencing was determined by RT-qPCR and cell invasion assay. PNFs enter phagocytic (BV-2) and nonphagocytic (U-87 MG) cells via endocytosis and passive translocation. si delivered using PNFs reduced the expression of polo-like kinase 1 mRNA and induced cell death in a panel of immortalized and glioblastoma-derived stem cells. Moreover, targeting MMP2 using PNF:si reduced the invasion capacity of U-87 MG cells. We show that stereotactic intra-tumoral administration of PNF:si significantly extends the survival of tumor bearing mice comparing with the untreated tumor bearing animals. Our results suggest that this nanomedicine-based RNA interference approach deserves further investigation as a potential brain tumor therapeutic tool.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.2217/nnm-2019-0298DOI Listing
December 2019

Thermal monitoring during photothermia: hybrid probes for simultaneous plasmonic heating and near-infrared optical nanothermometry.

Theranostics 2019 25;9(24):7298-7312. Epub 2019 Sep 25.

CIC biomaGUNE and CIBER-BBN, Paseo de Miramón 182, 20014 Donostia-San Sebastián, Spain.

The control of temperature during photothermal therapy is key to preventing unwanted damage in surrounding tissue or post-treatment inflammatory responses. Lack of accurate thermal control is indeed one of the main limitations that hyperthermia techniques present to allow their translation into therapeutic applications. We developed a nanoprobe that allows controlled local heating, combined with nanothermometry. The design of the probe follows a practical rationale that aims at simplifying experimental requirements and exploits exclusively optical wavelengths matching the first and second biological windows in the near-infrared. Hybrid nanostructures were chemically synthesized, and combine gold nanostars (photothermal agents) with CaF:Nd,Y nanoparticles (luminescent nanothermometers). Both components were simultaneously excited in the near-infrared range, at 808 nm. Following the goal of simplifying the thermal monitoring technique, the luminescent signal was recorded with a portable near-infrared detector. The performance of the probes was tested in 3D tumor spheroids from a human glioblastoma (U87MG) cell line. The location of the beads within the spheroids was determined measuring Nd emission in a commercial Lightsheet microscope, modified in-house to be able to select the required near-infrared wavelengths. The temperature achieved inside the tumor spheroids was deduced from the luminescence of Nd, following a protocol that we developed to provide reliable thermal readings. The choice of materials was shown to work as an optically excited hybrid probe. Depending on the illumination parameters, temperature can be controlled in a range between 37 ºC and 100 ºC. The near-infrared emission of nanothermometers also allows microscopic tracking of the hybrid nanostructures, confirming that the probes can penetrate deeper into the spheroid mass. We observed that, application of optical thermometry in biological environments requires often neglected considerations, since the optical signal changes along the optical path. Accordingly, we developed data analysis protocols that guarantee reliable thermal readings. The prepared hybrid probes are internalized in 3D tumor spheroids and can be used to induce cell death through photothermal effects, while simultaneously measuring the local temperature . We show that luminescent thermometry in biomedical applications requires the development of protocols that guarantee accurate readings. Regarding photothermal treatments, we observe a sharp thermal threshold at around 55 ºC (for 10 min treatments) that separates high survival ratio from complete cell death.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.7150/thno.38091DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6831289PMC
September 2020

Selective Liposomal Transport through Blood Brain Barrier Disruption in Ischemic Stroke Reveals Two Distinct Therapeutic Opportunities.

ACS Nano 2019 11 6;13(11):12470-12486. Epub 2019 Nov 6.

Nanomedicine Lab, Faculty of Biology, Medicine and Health, AV Hill Building , The University of Manchester , Manchester M13 9PT , United Kingdom.

The development of effective therapies for stroke continues to face repeated translational failures. Brain endothelial cells form paracellular and transcellular barriers to many blood-borne therapies, and the development of efficient delivery strategies is highly warranted. Here, in a mouse model of stroke, we show selective recruitment of clinically used liposomes into the ischemic brain that correlates with biphasic blood brain barrier (BBB) breakdown. Intravenous administration of liposomes into mice exposed to transient middle cerebral artery occlusion took place at early (0.5 and 4 h) and delayed (24 and 48 h) time points, covering different phases of BBB disruption after stroke. Using a combination of real-time imaging and histological analysis we show that selective liposomal brain accumulation coincides with biphasic enhancement in transcellular transport followed by a delayed impairment to the paracellular barrier. This process precedes neurological damage in the acute phase and maintains long-term liposomal colocalization within the neurovascular unit, which could have great potential for neuroprotection. Levels of liposomal uptake by glial cells are similarly selectively enhanced in the ischemic region late after experimental stroke (2-3 days), highlighting their potential for blocking delayed inflammatory responses or shifting the polarization of microglia/macrophages toward brain repair. These findings demonstrate the capability of liposomes to maximize selective translocation into the brain after stroke and identify two windows for therapeutic manipulation. This emphasizes the benefits of selective drug delivery for efficient tailoring of stroke treatments.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acsnano.9b01808DOI Listing
November 2019

Enhanced Intraliposomal Metallic Nanoparticle Payload Capacity Using Microfluidic-Assisted Self-Assembly.

Langmuir 2019 10 2;35(41):13318-13331. Epub 2019 Oct 2.

Nanomedicine Lab, Division of Pharmacy and Optometry, Faculty of Biology, Medicine and Health , University of Manchester , Av Hill Building , Manchester M13 9PT , U.K.

Hybrids composed of liposomes (L) and metallic nanoparticles (NPs) hold great potential for imaging and drug delivery purposes. However, the efficient incorporation of metallic NPs into liposomes using conventional methodologies has so far proved to be challenging. In this study, we report the fabrication of hybrids of liposomes and hydrophobic gold NPs of size 2-4 nm (Au) using a microfluidic-assisted self-assembly process. The incorporation of increasing amounts of AuNPs into liposomes was examined using microfluidics and compared to L-AuNP hybrids prepared by the reverse-phase evaporation method. Our microfluidics strategy produced L-AuNP hybrids with a homogeneous size distribution, a smaller polydispersity index, and a threefold increase in loading efficiency when compared to those hybrids prepared using the reverse-phase method of production. Quantification of the loading efficiency was determined by ultraviolet spectroscopy, inductively coupled plasma mass spectroscopy, and centrifugal field flow fractionation, and qualitative validation was confirmed by transmission electron microscopy. The higher loading of gold NPs into the liposomes achieved using microfluidics produced a slightly thicker and more rigid bilayer as determined with small-angle neutron scattering. These observations were confirmed using fluorescent anisotropy and atomic force microscopy. Structural characterization of the liposomal-NP hybrids with cryo-electron microscopy revealed the coexistence of membrane-embedded and interdigitated NP-rich domains, suggesting AuNP incorporation through hydrophobic interactions. The microfluidic technique that we describe in this study allows for the automated production of monodisperse liposomal-NP hybrids with high loading capacity, highlighting the utility of microfluidics to improve the payload of metallic NPs within liposomes, thereby enhancing their application for imaging and drug delivery.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acs.langmuir.9b00579DOI Listing
October 2019

Exposure to graphene oxide sheets alters the expression of reference genes used for real-time RT-qPCR normalization.

Sci Rep 2019 08 29;9(1):12520. Epub 2019 Aug 29.

Nanomedicine Lab, Faculty of Biology, Medicine and Health, AV Hill Building, The University of Manchester, Manchester, M13 9PT, UK.

Studies unraveling the interactions between graphene oxide (GO) and the biological milieu, including cells and tissues, are multiplying quickly as the biomedical applications of this and other 2D materials continue to be explored. Many of such studies rely on real-time RT-qPCR as a powerful yet simple technique to assess gene expression. However, a systematic investigation of potential GO-induced changes in the expression of reference genes, crucial for appropriate qPCR data normalization, is still lacking. We aimed to cover this gap investigating the stability of the expression of ten candidate reference genes upon exposure to increasing, but subtoxic, GO concentrations, with two established algorithms (Bestkeeper and NormFinder). The study was performed in a human cancer cell line (MCF7) and in mouse, non-cancerous, primary cells (mouse embryonic fibroblasts, MEFs), to assess different behaviors between cell types. Both algorithms evidenced significant deviations in the expression of various reference genes. Ribosomal proteins scored among the most significantly dysregulated in both cell types. ACTB and GAPDH, the most frequent calibrators in real-time RT-qPCR, were also affected, although differences existed between cell lines. This study illustrates the need to validate reference genes for appropriate real-time RT-qPCR normalization, according to specific experimental conditions, when GO-cell interactions occur.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-019-48970-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6715796PMC
August 2019

Graphene oxide: A growth factor delivery carrier to enhance chondrogenic differentiation of human mesenchymal stem cells in 3D hydrogels.

Acta Biomater 2019 09 17;96:271-280. Epub 2019 Jul 17.

Division of Cell Matrix Biology and Regenerative Medicine, School of Biological Sciences, Faculty of Biology, Medicine and Health, University of Manchester, Manchester Academic Health Science Centre, Manchester, M13 9PT, UK; NIHR Manchester Biomedical Research Centre, Manchester University NHS Foundation Trust, Manchester Academic Health Science Centre, Grafton St, M13 9WU Manchester, UK. Electronic address:

Cartilage engineering with stem cells in 3D scaffolds is a promising future therapy to treat cartilage defects. One challenge in the field is to design carriers to efficaciously deliver biological factors in 3D scaffolds containing stem cells to appropriately guide differentiation of these cells in same scaffolds and promote specific tissue synthesis. Graphene-based 2D nanomaterials have recently attracted extensive interest for their biomedical applications as they can adsorb a plethora of biological molecules, thus offering high potential as delivery carriers. This study utilized graphene oxide (GO) flakes to adsorb transforming growth factor β3 (TGF-β3), which were then incorporated into a collagen hydrogel. Human mesenchymal stem cells (hMSCs) were encapsulated in the same gel and chondrogenic differentiation assessed. The study showed GO flakes adsorbed > 99% TGF-β3 with <1.7% release. Adsorbed TGF-β3 retained a similar conformation to its dissolved counterpart (free protein) but importantly demonstrated greater conformational stability. Smad2 phosphorylation was promoted, and higher chondrogenic gene expression and cartilage-specific extracellular matrix deposition were achieved compared to exogenously delivering TGF-β3 in culture media. Effects were sustained in long-term 28-day culture. The results demonstrate GO flakes as highly-efficient for delivering GFs in 3D to guide cells in the same scaffold and induce tissue formation. The ability of GO flakes to provide sustained local delivery makes this material attractive for tissue engineering strategies, in particular for regionally-specific MSC differentiation (e.g. osteochondral tissue engineering). STATEMENT OF SIGNIFICANCE: Cartilage engineering involving stem cells in 3D scaffolds is a promising future therapy to treat cartilage defects which can lead to debilitating conditions such as osteoarthritis. However, this field faces the challenge to design delivery carriers to efficaciously deliver biological factors inside these 3D cell-containing scaffolds for appropriately-guided cell differentiation. Graphene-based 2D nanomaterials offer high potential as delivery carriers, but to date studies using them to deliver biological factors have been restricted to 2D substrates, non-scaffold cell masses, or acellular 3D scaffolds. Our study for the first time demonstrated simultaneously incorporating both human mesenchymal stem cells (hMSCs) and GO (graphene oxide)-adsorbed growth factor TGFβ3 into a 3D scaffold, where GO-adsorbed TGFβ3 enhanced chondrogenic differentiation of hMSCs and cartilage-tissue synthesis throughout the scaffold without needing to repeatedly supply TGFβ3 exogenously.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.actbio.2019.07.027DOI Listing
September 2019

Graphene oxide as a 2D platform for complexation and intracellular delivery of siRNA.

Nanoscale 2019 Aug 12;11(29):13863-13877. Epub 2019 Jul 12.

Nanomedicine Lab, Faculty Biology, Medicine and Health, AV Hill Building, The University of Manchester, Manchester M13 9PT, UK. and National Graphene Institute, The University of Manchester, Booth Street E, Manchester M13 9PL, UK.

The development of efficient and safe nucleic acid delivery vectors remains an unmet need holding back translation of gene therapy approaches to the bedside. Graphene oxide (GO) could help bypass such bottlenecks, thanks to its large surface area, versatile chemistry and biocompatibility, which could overall enhance transfection efficiency while abolishing some of the limitations linked to the use of viral vectors. Here, we aimed to assess the capacity of bare GO, without any further surface modification, to complex a short double-stranded nucleic acid of biological relevance (siRNA) and mediate its intracellular delivery. GO formed stable complexes with siRNA at 10 : 1, 20 : 1 and 50 : 1 GO : siRNA mass ratios. Complexation was further corroborated by atomistic molecular dynamics simulations. GO : siRNA complexes were promptly internalized in a primary mouse cell culture, as early as 4 h after exposure. At this time point, intracellular siRNA levels were comparable to those provided by a lipid-based transfection reagent that achieved significant gene silencing. The time-lapse tracking of internalized GO and siRNA evidenced a sharp decrease of intracellular siRNA from 4 to 12 h, while GO was sequestered in large vesicles, which may explain the lack of biological effects (i.e. gene silencing) achieved by GO : siRNA complexes. This study underlines the potential of non-surface modified GO flakes to act as 2D siRNA delivery platforms, without the need for cationic functionalization, but warrants further vector optimization to allow the effective release of the nucleic acid and achieve efficient gene silencing.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1039/c9nr02301aDOI Listing
August 2019

Biocompatibility and biodegradability of 2D materials: graphene and beyond.

Chem Commun (Camb) 2019 May;55(39):5540-5546

University of Strasbourg, CNRS, Immunology, Immunopathology and Therapeutic Chemistry, 67000 Strasbourg, France.

The potential risks associated with two-dimensional (2D) nanomaterials may cause serious concerns about their real applications and impact in biological systems. In addition, the demonstration of biodegradability of these flat nanomaterials is essential in living organisms. Here, we summarise the state-of-the-art in the field of biocompatibility and biodegradability of graphene-related materials (such as 2D materials like MoS2, BN or WS2). The impact of chemical functionalisation on the potential control of the biodegradability profile of these structures is also discussed.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1039/c9cc01205bDOI Listing
May 2019