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The treatment of Parkinson's disease (PD) is still symptomatic since disease-modifying treatments for PD are not available. Oral levodopa is the gold standard for the treatment of PD motor symptoms. However, incomplete and fluctuating plasma exposure of levodopa leads to suboptimal treatment of the symptoms. The main objective of this study was to investigate to what extent increased carbidopa doses (50 and 100 mg) increase the plasma levels of 100-mg immediate-release (IR) levodopa compared to a 25-mg carbidopa dose with and without co-administration of 200 mg entacapone.A double-blind, placebo-controlled, randomized, crossover, phase I, pharmacokinetic study with 25 healthy volunteers was conducted. In addition, a semi-mechanistic pharmacokinetic model was built to theoretically evaluate the effect of inhibiting aromatic amino acid decarboxylase (AADC) and catechol-O-methyltransferase (COMT) mediated metabolism of levodopa on the exposure of levodopa.The effect of increased carbidopa doses 50 and 100 mg on the total exposure (AUC) of 100 mg IR levodopa was +29% and +36%, respectively, when entacapone was co-administered. Without entacapone, the corresponding increases were +13% and +17%. With entacapone co-administration, the increased carbidopa dose also clearly increased levodopa trough concentration. There was no significant effect on the peak concentrations of levodopa.Increasing carbidopa doses significantly increased the exposure and reduced the fluctuation of IR levodopa in plasma during simultaneous COMT inhibition with entacapone. Theoretical pharmacokinetic simulations suggested that the plasma profile of oral IR levodopa can be even further improved by optimizing AADC and COMT inhibition.

Background There is a need for effective and cost-effective interprofessional care models that support older people to maintain their quality of life (QoL) and physical performance to live longer independently in their own homes. Objectives The objectives were to evaluate effectiveness, QoL and physical performance, and cost-utility of a people-centred care model (PCCM), including the contribution of clinically trained pharmacists, compared with that of usual care in primary care. Methods A randomised controlled trial (RCT) with a two-year follow-up was conducted. The participants were multimorbid community-living older people, aged ≥75 years. The intervention comprised an at-home patient interview, health review, pharmacist-led clinical medication review, an interprofessional team meeting, and nurse-led care coordination and health support. At the baseline and at the 1-year and 2-year follow-ups, QoL (SF-36, 36-Item Short-Form Health Survey) and physical performance (SPPB, Short Performance Physical Battery) were measured. Additionally, a physical dimension component summary in the SF-36 was calculated. The SF-36 data were transformed into SF-6D scores to calculate quality-adjusted life-years (QALYs). Healthcare resource use were collected and transformed into costs. A healthcare payer perspective was adopted. Incremental cost-effectiveness ratio (ICER) was calculated, and one-way sensitivity analysis was performed. Results No statistically or clinically significant differences were observed between the usual care (n = 126) and intervention group (n = 151) patients in their QoL; at the 2-year follow-up the mean difference was −0.02, (95 % CI -0.07; 0.04,p = 0.56). While the mean difference between the groups in physical performance at the 2-year follow-up was −1.02, (−1.94;-0.10,p = 0.03), between the physical component summary scores it was −7.3, (−15.2; 0.6,p = 0.07). The ICER was −73 638€/QALY, hence, the developed PCCM dominated usual care, since it was more effective and less costly. Conclusions The cost-utility analysis showed that the PCCM including pharmacist-led medication review dominated usual care. However, it had no effect on QoL and the effect towards physical performance remained unclear.

The assessment of therapeutic synergy was calculated with FTV method. Observed FTV (mean tumor volume experimental)/(mean tumor volume control). Expected FTV (Mean FTV of Carnosine6K) x (Mean FTV of virus). A ratio>1 indicates a synergistic effect, and a ratio

Oncolytic viruses are able to specifically replicate, infect, and kill only cancer cells. Their combination with chemotherapeutic drugs has shown promising results due to the synergistic action of virus and drugs; the combinatorial therapy is considered a potential clinically relevant approach for cancer. In this study, we optimized a strategy to absorb peptides on the viral capsid, based on electrostatic interaction, and used this strategy to deliver an active antitumor drug. We used L-carnosine, a naturally occurring histidine dipeptide with a significant antiproliferative activity. An ad hoc modified, positively charged L-carnosine was combined with the capsid of an oncolytic adenovirus to generate an electrostatic virus–carnosine complex. This complex showed enhanced antitumor efficacy in vitro and in vivo in different tumor models. In HCT-116 colorectal and A549 lung cancer cell lines, the complex showed higher transduction ratio and infectious titer compared with an uncoated oncolytic adenovirus. The in vivo efficacy of the complex was tested in lung and colon cancer xenograft models, showing a significant reduction in tumor growth. Importantly, we investigated the molecular mechanisms underlying the effects of complex on tumor growth reduction. We found that complex induces apoptosis in both cell lines, by using two different mechanisms, enhancing viral replication and affecting the expression of Hsp27. Our system could be used in future studies also for delivery of other bioactive drugs. Mol Cancer Ther; 15(4); 651–60. ©2016 AACR.

Most representative microscop photograph of human fibroblast cell line. (E) Cell viability was performed by MTS Assay on CCD- 112Sk cell line, in order to evaluate if complex affects negatively healthy cells. We have not observed toxic effect(incubation time 96 hrs) on fibroblast cultures, neither with complex nor virus alone and Carnosine6K alone (virus:100 VP/cell, Carnosine6K: 8mM).

Nanofibrillated cellulose (NFC) hydrogel is a versatile biomaterial suitable, for example, for three-dimensional (3D) cell spheroid culturing, drug delivery, and wound treatment. By freeze-drying NFC hydrogel, highly porous NFC structures can be manufactured. We freeze-dried NFC hydrogel and subsequently reconstituted the samples into a variety of concentrations of NFC fibers, which resulted in different stiffness of the material, i.e., different mechanical cues. After the successful freeze-drying and reconstitution, we showed that freeze-dried NFC hydrogel can be used for one-step 3D cell spheroid culturing of primary mesenchymal stem/stromal cells, prostate cancer cells (PC3), and hepatocellular carcinoma cells (HepG2). No difference was observed in the viability or morphology between the 3D cell spheroids cultured in the freeze-dried and reconstituted NFC hydrogel and fresh NFC hydrogel. Furthermore, the 3D cultured spheroids showed stable metabolic activity and nearly 100% viability. Finally, we applied a convolutional neural network (CNN)-based automatic nuclei segmentation approach to automatically segment individual cells of 3D cultured PC3 and HepG2 spheroids. These results provide an application to culture 3D cell spheroids more readily with the NFC hydrogel and a step towards automatization of 3D cell culturing and analysis.

Objective: Skin graft donor site management is a concern particularly for elderly patients and patients with poor wound healing competence, and also because donor sites are a source of pain and discomfort. Although different types of dressings exist, there is no consensus regarding optimal dressing type on donor site care to promote healing, reduce pain, and improve patients' comfort. Approach: This prospective, single-center clinical trial evaluated the performance of nanofibrillar cellulose (NFC) wound dressing (FibDex (R) by UPM-Kymmene Corporation) for treatment of donor sites compared with a polylactide-based copolymer dressing. The study enrolled 24 patients requiring skin grafting with mean age of 49 +/- 18. The primary outcome measure was wound healing time. Secondary outcomes, the epithelialization, subjective pain, the scar appearance assessed using the Patient and Observer Scar Assessment Scale (POSAS), and skin elasticity and transepidermal water loss (TEWL), were evaluated at 1 and 6 months postoperatively. Results: No statistically significant differences were observed between NFC and copolymer dressings regarding wound healing time, epithelialization, experience of pain, or TEWL. Significant differences were observed in the POSAS results for thickness and vascularity in the Observer score, in the favor of NFC over copolymer dressing. Moreover, skin elasticity was significantly improved with NFC dressing in terms of viscoelasticity and elastic modulus at 1 month postoperatively. Innovation: NFC dressing is a new, green sustainable product for wound treatment without animal or human-origin components. Conclusion: NFC dressing provides efficient wound healing at skin graft donor sites and is comparable or even preferable compared with the copolymer dressing.

Extracellular vesicles (EVs) are a complex and heterogeneous population of nanoparticles involved in cell-to-cell communication. Recently, numerous studies have indicated the potential of EVs as therapeutic agents, drug carriers and diagnostic tools. However, the results of these studies are often difficult to evaluate, since different characterization methods are used to assess the purity, physical and biochemical characteristics of the EV samples. In this study, we compared four methods for the EV sample characterization and purity assessment: i) the particle-to-protein ratio based on particle analyses with nanoparticle tracking and protein concentration by bicinchoninic acid assay, ii) Western Blot analysis for specific EV biomarkers, iii) two spectroscopic lipid-to protein ratios by either the attenuated total reflection Fourier transform infrared (ATR-FTIR) or Raman spectroscopy. The results confirm the value of Raman and ATR-FTIR spectroscopy as robust, fast and operator independent tools that require only a few microliters of EV sample. We propose that the spectroscopic lipid-to protein (Li/Pr) ratios are reliable parameters for the purity assessment of EV preparations. Moreover, apart from determining protein concentrations, we show that ATR-FTIR spectroscopy can also be used for indirect measurements of EV concentrations. Nevertheless, the Li/Pr ratios do not represent full characterization of the EV preparations. For a complete characterization of selected EV preparations, we recommend also additional use of particle size distribution and EV biomarker analysis.

Biomaterial aerogel fabrication by freeze-drying must be further improved to reduce the costs of lengthy freeze-drying cycles and to avoid the formation of spongy cryogels and collapse of the aerogel structures. Residual water content is a critical quality attribute of the freeze-dried product, which can be monitored in-line with near-infrared (NIR) spectroscopy. Predictive models of NIR have not been previously applied for biomaterials and the models were mostly focused on the prediction of only one formulation at a time. We recorded NIR spectra of different nanofibrillated cellulose (NFC) hydrogel formulations during the secondary drying and set up a partial least square regression model to predict their residual water contents. The model can be generalized to measure residual water of formulations with different NFC concentrations and the excipients, and the NFC fiber concentrations and excipients can be separated with the principal component analysis. Our results provide valuable information about the freeze-drying of biomaterials and aerogel fabrication, and how NIR spectroscopy can be utilized in the optimization of residual water content.

Funding Information: V.G. and P.V. acknowledge Academy of Finland’s Project No. 308931. J.J.V.D. and M.Ö. acknowledge Academy of Finland’s Flagship Programme under Project Nos. 318890 and 318891 (Competence Center for Materials Bioeconomy, FinnCERES). M.Y. acknowledges Academy of Finland’s Flagship Programme under Project No. 337430 (Gene, Cell and Nano Therapy Competence Cluster for the Treatment of Chronic Diseases, GeneCellNano). Publisher Copyright: © 2021 The Authors. Published by American Chemical Society. Bingel cyclopropanation between Buckminster fullerene and a heteroarmed malonate was utilized to produce a hexakis-functionalized C60 core, with azide and tetrazine units. This orthogonally bifunctional C60 scaffold can be selectively one-pot functionalized by two pericyclic click reactions, that is, inverse electron-demand Diels-Alder and azide-alkyne cycloaddition, which with appropriate ligands (monosaccharides, a peptide and oligonucleotides tested) allows one to control the assembly of heteroantennary bioconjugates.

Advanced 3D in vitro models are laborious to prepare and susceptible to unintentional design errors due to culture adaptations, cell immaturity, xenofactors or yet incomplete knowledge of the dynamics within tissues or materials. In order to acquire cost-efficient research material with intact in vivo composition, we developed novel tissue culture method with plant-derived scaffolding.Human skin-, foreskin- and glioblastoma multiforme biopsies were dissected mechanically and cultivated for 28 days in plant-derived nanofibrillar cellulose hydrogel. Comparative cultures were done using mouse sarcoma tumor –derived Matrigel™. Long-term preservation of cultivated tissues was evaluated against typical immunohistochemical biomarkers for each tissue type: skin tissues for cytokeratins 5/6, E-cadherin and vimentin for sustained tissue structures, and brain neoplasia for Olig2, S100, Nestin, NOTCH1, MAP2 and GFAP for preserved disease profile.Histological analysis from both culture conditions showed that until day 28, all cultivated biopsy types were able to sustain their characteristic protein expressions without signs of necrosis. We here conclude a novel tissue culture model in xeno-free 3D scaffolding, that can enable long-term sample storage in vitro, studies of human tumor tissues and their non-neoplastic microenvironment, and innovations in personalized medicine research.

Studies of extracellular vesicles (EVs), their trafficking and characterization often employ fluorescent labelling. Unfortunately, little attention has been paid thus far to a thorough evaluation of the purification of EVs after labelling, although the presence of an unbound dye may severely compromise the results or even lead to wrong conclusions on EV functionality. Here, we systematically studied five dyes for passive EV labelling and meticulously compared five typical purification methods: ultracentrifugation (UC), ultracentrifugation with discontinuous density gradient (UCG), ultrafiltration (UF), size exclusion chromatography (SEC), and anion exchange chromatography (AEC). A general methodology for evaluation of EV purification efficiency after the labelling was developed and tested to select the purification methods for the chosen dyes. Firstly, we found that some methods initially lead to high EV losses even in the absence of the dye. Secondly, the suitable purification method needs to be found for each particular dye and depends on the physical and chemical properties of the dye. Thirdly, we demonstrated that the developed parameter

Urinary extracellular vesicles (uEV) are a topical source of non-invasive biomarkers for health and diseases of the urogenital system. However, several challenges have become evident in the standardization of uEV pipelines from collection of urine to biomarker analysis. Here, we studied the effect of pre-analytical variables and developed means of quality control for uEV isolates to be used in transcriptomic biomarker research. We included urine samples from healthy controls and individuals with type 1 or type 2 diabetes and normo-, micro- or macroalbuminuria and isolated uEV by ultracentrifugation. We studied the effect of storage temperature (-20 degrees C vs. -80 degrees C), time (up to 4 years) and storage format (urine or isolated uEV) on quality of uEV by nanoparticle tracking analysis, electron microscopy, Western blotting and qPCR. Urinary EV RNA was compared in terms of quantity, quality, and by mRNA or miRNA sequencing. To study the stability of miRNA levels in samples isolated by different methods, we created and tested a list of miRNAs commonly enriched in uEV isolates. uEV and their transcriptome were preserved in urine or as isolated uEV even after long-term storage at -80 degrees C. However, storage at -20 degrees C degraded particularly the GC-rich part of the transcriptome and EV protein markers. Transcriptome was preserved in RNA samples extracted with and without DNAse, but read distributions still showed some differences in e.g. intergenic and intronic reads. MiRNAs commonly enriched in uEV isolates were stable and concordant between different EV isolation methods. Analysis of never frozen uEV helped to identify surface characteristics of particles by EM. In addition to uEV, qPCR assays demonstrated that uEV isolates commonly contained polyoma viruses. Based on our results, we present recommendations how to store and handle uEV isolates for transcriptomics studies that may help to expedite standardization of the EV biomarker field.

The diversity and safety of nanofibrillated cellulose (NFC) hydrogels have gained a vast amount of interest at the pharmaceutical site in recent years. Moreover, this biomaterial has a high potential to be utilized as a protective matrix during the freeze-drying of heat-sensitive pharmaceuticals and biologics to increase their properties for long-term storing at room temperature and transportation. Since freeze-drying and subsequent reconstitution have not been optimized for this biomaterial, we must find a wider understanding of the process itself as well as the molecular level interactions between the NFC hydrogel and the most suitable lyoprotectants. Herein we optimized the reconstitution of the freeze-dried NFC hydrogel by considering critical quality attributes required to ensure the success of the process and gained insights of the obtained experimental data by simulating the effects of the used lyoprotectants on water and NFC. We discovered the correlation between the measured characteristics and molecular dynamics simulations and obtained successful freeze-drying and subsequent reconstitution of NFC hydrogel with the presence of 300 mM of sucrose. These findings demonstrated the possibility of using the simulations together with the experimental measurements to obtain a more comprehensive way to design a successful freeze-drying process, which could be utilized in future pharmaceutical applications.

Transmembrane protein integrins play a key role in cell adhesion. Cell-biomaterial interactions are affected by integrin expression and conformation, which are actively controlled by cells. Although integrin structure and function have been studied in detail, quantitative analyses of integrin-mediated cell-biomaterial interactions are still scarce. Here, we have used atomic force spectroscopy to study how integrin distribution and activation (via intracellular mechanisms in living cells or by divalent cations) affect the interaction of human pluripotent stem cells (WA07) and human hepatocarcinoma cells (HepG2) with promising biomaterials-human recombinant laminin-521 (LN-521) and cellulose nanofibrils (CNF). Cell adhesion to LN-521-coated probes was remarkably influenced by cell viability, divalent cations, and integrin density in WA07 colonies, indicating that specific bonds between LN-521 and activated integrins play a significant role in the interactions between LN-521 and HepG2 and WA07 cells. In contrast, the interactions between CNF and cells were nonspecific and not influenced by cell viability or the presence of divalent cations. These results shed light on the underlying mechanisms of cell adhesion, with direct impact on cell culture and tissue engineering applications.

International audience; Freeze-drying is the most widespread method to preserve protein drugs and vaccines in a dry form facilitating their storage and transportation without the laborious and expensive cold chain. Extending this method for the preservation of natural biomaterials and cells in a dry form would provide similar benefits, but most results in the domain are still below expectations. In this review, rather than consider freeze-drying as a traditional black box we "break it" through a detailed process thinking approach. We discuss freeze-drying from process thinking aspects, introduce the chemical, physical, and mechanical environments important in this process, and present advanced biophotonic process analytical technology. In the end, we review the state of the art in the freezedrying of the biomaterials, extracellular vesicles, and cells. We suggest that the rational design of the experiment and implementation of advanced biophotonic tools are required to successfully preserve the natural biomaterials and cells by freeze-drying. We discuss this change of paradigm with existing literature and elaborate on our perspective based on our new unpublished results.

An azide-functionalized 12-armed Buckminster fullerene has been monosubstituted in organic media with a substoichiometric amount of cyclooctyne-modified oligonucleotides. Exposing the intermediate products then to the same reaction (i.e., strain-promoted alkyne-azide cycloaddition, SPAAC) with an excess of slightly different oligonucleotide constituents in an aqueous medium yields molecularly defined monofunctionalized spherical nucleic acids (SNAs). This procedure offers a controlled synthesis scheme in which one oligonucleotide arm can be functionalized with labels or other conjugate groups (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid, DOTA, and Alexa-488 demonstrated), whereas the rest of the 11 arms can be left unmodified or modified by other conjugate groups in order to decorate the SNAs' outer sphere. Extra attention has been paid to the homogeneity and authenticity of the C60-azide scaffold used for the assembly of full-armed SNAs. publishedVersion

Hydrogels, natural and synthetic origin, are actively studied for their use for implants and payload carriers. These biomaterials for delivery systems have enormous potential in basic biomedical research, drug development, and long-term delivery of biologics. Nanofibrillated cellulose (NFC) hydrogels, both natural and anionic (ANFC) ones, allow drug loading for immediate and controlled release via the slow drug dissolution of solid drug crystals into hydrogel and its subsequent release. This property makes NFC originated hydrogels an interesting non-toxic and non-human origin material as drug reservoir for long-term controlled release formulation or implant for patient care. A compelling tool for studying NFC hydrogels is Raman spectroscopy, which enables to resolve the chemical structures of different molecules in a high-water content like hydrogels, since Raman spectroscopy is insensitive to water molecules. That offers real time investigation of label-free drugs and their release in high-water-content media. Despite the huge potential of Raman spectroscopy in bio-pharmaceutical applications, the strong fluorescence background of many drug samples masking the faint Raman signal has restricted the widespread use of it. In this study we used a Raman spectrometer capable of suppressing the unpleasant fluorescence background by combining a pulsed laser and time-resolved complementary metal-oxide-semiconductor (CMOS) single-photon avalanche diode (SPAD) line sensor for the label-free investigation of Metronidazole and Vitamin C diffusivities in ANFC. The results show the possibility to modulate the ANFC-based implants and drug delivery systems, when the release rate needs to be set to a desired value. More importantly, the now developed label free real-time method is universal and can be adapted to any hydrogel/drug combination for producing reliable drug diffusion coefficient data in complex and heterogeneous systems, where traditional sampling-based methods are cumbersome to use. The wide temporal range of the time-resolved CMOS SPAD sensors makes it possible to capture also the fluorescence decay of samples, giving rise to a combined time-resolved Raman and fluorescence spectroscopy, which provides additional information on the chemical, functional and structural changes in samples. publishedVersion

Real-time label-free techniques are used to profile G protein-coupled receptor (GPCR) signaling pathways in living cells. However, interpreting the label-free signal responses is challenging, and previously reported methods do not reliably separate pathways from each other. In this study, a continuous angular-scanning surface plasmon resonance (SPR) technique is utilized for measuring label-free GPCR signal profiles. We show how the continuous angular-scanning ability, measuring up to nine real-time label-free parameters simultaneously, results in more information-rich label-free signal profiles for different GPCR pathways, providing a more accurate pathway separation. For this, we measured real-time full-angular SPR response curves for Gs, Gq, and Gi signaling pathways in living cells. By selecting two of the most prominent label-free parameters: the full SPR curve angular and intensity shifts, we present how this analysis approach can separate each of the three signaling pathways in a straightforward single-step analysis setup, without concurrent use of signal inhibitors or other response modulating compounds.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Biomaterials of different nature have been and are widely studied for various biomedical applications. In many cases, biomaterial assemblies are designed to mimic biological systems. Although biomaterials have been thoroughly characterized in many aspects, not much quantitative information on the molecular level interactions between different biomaterials is available. That information is very important, on the one hand, to understand the properties of biological systems and, on the other hand, to develop new composite biomaterials for special applications. This work presents a systematic, quantitative analysis of self- and cross-interactions between films of collagen I (Col I), collagen IV (Col IV), laminin (LN-521), and cellulose nanofibrils (CNF), that is, biomaterials of different nature and structure that either exist in biological systems (e.g., extracellular matrices) or have shown potential for 3D cell culture and tissue engineering. Direct surface forces and adhesion between biomaterials-coated spherical micro-particles and flat substrates were measured in phosphate-buffered saline using an atomic force microscope and the colloidal probe technique. Different methods (Langmuir-Schaefer deposition, spin-coating, or adsorption) were applied to completely coat the flat substrates and the spherical micro particles with homogeneous biomaterial films. The adhesion between biomaterials films increased with the time that the films were kept in contact. The strongest adhesion was observed between Col IV films, and between Col IV and LN-521 films after 30 s contact time. In contrast, low adhesion was measured between CNF films, as well as between CNF and LN-521 films. Nevertheless, a good adhesion between CNF and collagen films (especially Col I) was observed. These results increase our understanding of the structure of biological systems and can support the design of new matrices or scaffolds where different biomaterials are combined for diverse biological or medical applications.

Correction for ‘Addressing challenges in the removal of unbound dye from passively labelled extracellular vesicles’ by Kaisa Rautaniemi et al., Nanoscale Adv., 2022, DOI: 10.1039/d1na00755f.

The soluble oligomeric form of the amyloid beta (Aβ) peptide is the major causative agent in the molecular pathogenesis of Alzheimer's disease (AD). We have previously developed a pyrroline-nitroxyl fluorene compound (SLF) that blocks the toxicity of Aβ. Here we introduce the multi-parametric surface plasmon resonance (MP-SPR) approach to quantify SLF binding and effect on the self-association of the peptide via a label-free, real-time approach. Kinetic analysis of SLF binding to Aβ and measurements of layer thickness alterations inform on the mechanism underlying the ability of SLF to inhibit Aβ toxicity and its progression towards larger oligomeric assemblies. Depending on the oligomeric state of Aβ, distinct binding affinities for SLF are revealed. The Aβ monomer and dimer uniquely possess sub-nanomolar affinity for SLF via a non-specific mode of binding. SLF binding is weaker in oligomeric Aβ, which displays an affinity for SLF on the order of 100 μM. To complement these experiments we carried out molecular docking and molecular dynamics simulations to explore how SLF interacts with the Aβ peptide. The MP-SPR results together with in silico modeling provide affinity data for the SLF-Aβ interaction and allow us to develop a new general method for examining protein aggregation.

Extracellular vesicles (EVs) have the ability to function as molecular vehicles and could therefore be harnessed to deliver drugs to target cells in diseases such as cancer. The composition of EVs determines their function as well as their interactions with cells, which consequently affects the cell uptake efficacy of EVs. In this study, we present two novel label-free approaches for studying EVs; characterization of EV composition by time-gated surface-enhanced Raman spectroscopy (TG-SERS) and monitoring the kinetics and amount of cellular uptake of EVs by surface plasmon resonance (SPR) in real-time. Using these methods, we characterized the most abundant EVs of human blood, red blood cell (RBC)- and platelet (PLT)-derived EVs and studied their interactions with prostate cancer cells. Complementary studies were performed with nanoparticle tracking analysis for concentration and size determinations of EVs, zeta potential measurements for surface charge analysis, and fluorophore-based confocal imaging and flow cytometry to confirm EV uptake. Our results revealed distinct biochemical features between the studied EVs and demonstrated that PLT-derived EVs were more efficiently internalized by PC-3 cells than RBC-derived EVs. The two novel label-free techniques introduced in this study were found to efficiently complement conventional techniques and paves the way for further use of TG-SERS and SPR in EV studies.

Extracellular vesicles (EVs) have been showcased as auspicious candidates for delivering therapeutic cargo, including oncolytic viruses for cancer treatment. Delivery of oncolytic viruses in EVs could provide considerable advantages, hiding the viruses from the immune system and providing alternative entry pathways into cancer cells. Here we describe the formation and viral cargo of EVs secreted by cancer cells infected with an oncolytic adenovirus (IEVs, infected cell-derived EVs) as a function of time after infection. IEVs were secreted already before the lytic release of virions and their structure resembled normally secreted EVs, suggesting that they were not just apoptotic fragments of infected cells. IEVs were able to carry the viral genome and induce infection in other cancer cells. As such, the role of EVs in the life cycle of adenoviruses may be an important part of a successful infection and may also be harnessed for cancer- and gene therapy.

The exploitation of curcumin for oral disease treatment is limited by its low solubility, poor bioavailability, and low stability. Surface-functionalized poly-lactic-co-glycolic acid (PLGA) nanoparticles (NPs) have shown promising results to ameliorate selective delivery of drugs to the gastro-intestinal tract. In this study, curcumin-loaded PLGA NPs (C-PLGA NPs) of about 200 nm were surface-coated with chitosan (CS) for gastro-intestinal mucosa adhesion, wheat germ agglutinin (WGA) for colon targeting or GE11 peptide for tumor colon targeting. Spectrometric and zeta potential analyses confirmed the successful functionalization of the C-PLGA NPs. Real-time label-free assessment of the cell membrane-NP interactions and NP cell uptake were performed by quartz crystal microbalance coupled with supported lipid bilayers and by surface plasmon resonance coupled with living cells. The study showed that CS-coated C-PLGA NPs interact with cells by the electrostatic mechanism, while both WGA- and GE11-coated C-PLGA NPs interact and are taken up by cells by specific active mechanisms. In vitro cell uptake studies corroborated the real-time label-free assessment by yielding a curcumin cell uptake of 7.3 ± 0.3, 13.5 ± 1.0, 27.3 ± 4.9, and 26.0 ± 1.3 μg per 104 HT-29 cells for noncoated, CS-, WGA-, and GE11-coated C-PLGA NPs, respectively. Finally, preliminary in vivo studies showed that the WGA-coated C-PLGA NPs efficiently accumulate in the colon after oral administration to healthy Balb/c mice. In summary, the WGA- and GE11-coated C-PLGA NPs displayed high potential for application as active targeted carriers for anticancer drug delivery to the colon.