Determination of the combined effect of grape seed extract and cold atmospheric plasma on foodborne pathogens and their environmental stress knockout mutants.

The study aimed to explore the antimicrobial efficacy of grape seed extract (GSE) and cold atmospheric plasma (CAP) individually or in combination against L. monocytogenes and E. coli wild type (WT) and their isogenic mutants in environmental stress genes. More specifically, we examined the effects of 1% (wt/vol) GSE, 4 min of CAP treatment, and their combined effect on L. monocytogenes 10403S WT and its isogenic mutants ΔsigB, ΔgadD1, ΔgadD2, ΔgadD3, as well as E. coli K12 and its isogenic mutants ΔrpoS, ΔoxyR, and ΔdnaK. In addition, the sequence of the combined treatments was tested. A synergistic effect was achieved for all L. monocytogenes strains when exposure to GSE was followed by CAP treatment. However, the same effect was observed against E. coli strains, only for the reversed treatment sequence. Additionally, L. monocytogenes ΔsigB was more sensitive to the individual GSE and the combined GSE/CAP treatment, whereas ΔgadD2 was more sensitive to CAP, as compared to the rest of the mutants under study. Individual GSE exposure was unable to inhibit E. coli strains, and individual CAP treatment resulted in higher inactivation of E. coli in comparison to L. monocytogenes with the strain ΔrpoS appearing the most sensitive among all studied strains. Our findings provide a step toward a better understanding of the mechanisms playing a role in the tolerance/sensitivity of our model Gram-positive and Gram-negative bacteria toward GSE, CAP, and their combination. Therefore, our results contribute to the development of more effective and targeted antimicrobial strategies for sustainable decontamination.IMPORTANCEAlternative approaches to conventional sterilization are gaining interest from the food industry, driven by (i) the consumer demand for minimally processed products and (ii) the need for sustainable, environmentally friendly processing interventions. However, as such alternative approaches are milder than conventional heat sterilization, bacterial pathogens might not be entirely killed by them, which means that they could survive and grow, causing food contamination and health hazards. In this manuscript, we performed a systematic study of the impact of antimicrobials derived from fruit industry waste (grape seed extract) and cold atmospheric plasma on the inactivation/killing as well as the damage of bacterial pathogens and their genetically modified counterparts, for genes linked to the response to environmental stress. Our work provides insights into genes that could be responsible for the bacterial capability to resist/survive those novel treatments, therefore, contributing to the development of more effective and targeted antimicrobial strategies for sustainable decontamination.

Double Imprinted Nanoparticles for Sequential Membrane-to-Nuclear Drug Delivery.

Efficient and site-specific delivery of therapeutics drugs remains a critical challenge in cancer treatment. Traditional drug nanocarriers such as antibody-drug conjugates are not generally accessible due to their high cost and can lead to serious side effects including life-threatening allergic reactions. Here, these problems are overcome via the engineering of supramolecular agents that are manufactured with an innovative double imprinting approach. The developed molecularly imprinted nanoparticles (nanoMIPs) are targeted toward a linear epitope of estrogen receptor alfa (ERα) and loaded with the chemotherapeutic drug doxorubicin. These nanoMIPs are cost-effective and rival the affinity of commercial antibodies for ERα. Upon specific binding of the materials to ERα, which is overexpressed in most breast cancers (BCs), nuclear drug delivery is achieved via receptor-mediated endocytosis. Consequentially, significantly enhanced cytotoxicity is elicited in BC cell lines overexpressing ERα, paving the way for precision treatment of BC. Proof-of-concept for the clinical use of the nanoMIPs is provided by evaluating their drug efficacy in sophisticated three-dimensional (3D) cancer models, which capture the complexity of the tumor microenvironment in vivo without requiring animal models. Thus, these findings highlight the potential of nanoMIPs as a promising class of novel drug compounds for use in cancer treatment.

The microbial composition of pancreatic ductal adenocarcinoma: A systematic review of 16S rRNA gene sequencing.

BACKGROUND: Pancreatic cancer, specifically pancreatic ductal adenocarcinoma (PDAC), continues to pose a significant clinical and scientific challenge. The most significant finding of recent years is that PDAC tumours harbour their specific microbiome, which differs amongst tumour entities and is distinct from healthy tissue. This review aims to evaluate and summarise all PDAC studies that have used the next-generation technique, 16S rRNA gene amplicon sequencing within each bodily compartment. As well as establishing a causal relationship between PDAC and the microbiome. MATERIALS AND METHODS: This systematic review was carried out according to the Preferred Reporting Items for Systematic Reviews and Meta-analysis (PRISMA) guidelines. A comprehensive search strategy was designed, and 1727 studies were analysed. RESULTS: In total, 38 studies were selected for qualitative analysis and summarised significant PDAC bacterial signatures. Despite the growing amount of data provided, we are not able to state a universal 16S rRNA gene microbial signature that can be used for PDAC screening. This is most certainly due to the heterogeneity of the presentation of results, lack of available datasets and the intrinsic selection bias between studies. CONCLUSION: Several key studies have begun to shed light on causality and the influence the microbiome constituents and their produced metabolites could play in tumorigenesis and influencing outcomes. The challenge in this field is to shape the available microbial data into targetable signatures. Making sequenced data readily available is critical, coupled with the coordinated standardisation of data and the need for consensus guidelines in studies investigating the microbiome in PDAC.

Single-Cell Untargeted Lipidomics Using Liquid Chromatography and Data-Dependent Acquisition after Live Cell Selection.

We report the development and validation of an untargeted single-cell lipidomics method based on microflow chromatography coupled to a data-dependent mass spectrometry method for fragmentation-based identification of lipids. Given the absence of single-cell lipid standards, we show how the methodology should be optimized and validated using a dilute cell extract. The methodology is applied to dilute pancreatic cancer and macrophage cell extracts and standards to demonstrate the sensitivity requirements for confident assignment of lipids and classification of the cell type at the single-cell level. The method is then coupled to a system that can provide automated sampling of live, single cells into capillaries under microscope observation. This workflow retains the spatial information and morphology of cells during sampling and highlights the heterogeneity in lipid profiles observed at the single-cell level. The workflow is applied to show changes in single-cell lipid profiles as a response to oxidative stress, coinciding with expanded lipid droplets. This demonstrates that the workflow is sufficiently sensitive to observing changes in lipid profiles in response to a biological stimulus. Understanding how lipids vary in single cells will inform future research into a multitude of biological processes as lipids play important roles in structural, biophysical, energy storage, and signaling functions.

Chemotherapy Assessment in Advanced Multicellular 3D Models of Pancreatic Cancer: Unravelling the Importance of Spatiotemporal Mimicry of the Tumor Microenvironment.

Pancreatic ductal adenocarcinoma (PDAC) is a challenge for global health with very low survival rate and high therapeutic resistance. Hence, advanced preclinical models for treatment screening are of paramount importance. Herein, chemotherapeutic (gemcitabine) assessment on novel (polyurethane) scaffold-based spatially advanced 3D multicellular PDAC models is carried out. Through comprehensive image-based analysis at the protein level, and expression analysis at the mRNA level, the importance of stromal cells is confirmed, primarily activated stellate cells in the chemoresistance of PDAC cells within the models. Furthermore, it is demonstrated that, in addition to the presence of activated stellate cells, the spatial architecture of the scaffolds, i.e., segregation/compartmentalization of the cancer and stromal zones, affect the cellular evolution and is necessary for the development of chemoresistance. These results highlight that, further to multicellularity, mapping the tumor structure/architecture and zonal complexity in 3D cancer models is important for better mimicry of the in vivo therapeutic response.

Unravelling the impact of fat content on the microbial dynamics and spatial distribution of foodborne bacteria in tri-phasic viscoelastic 3D models.

The aim of the current study is to develop and characterise novel complex multi-phase in vitro 3D models, for advanced microbiological studies. More specifically, we enriched our previously developed bi-phasic polysaccharide (Xanthan Gum)/protein (Whey Protein) 3D model with a fat phase (Sunflower Oil) at various concentrations, i.e., 10%, 20%, 40% and 60% (v/v), for better mimicry of the structural and biochemical composition of real food products. Rheological, textural, and physicochemical analysis as well as advanced microscopy imaging (including spatial mapping of the fat droplet distribution) of the new tri-phasic 3D models revealed their similarity to industrial food products (especially cheese products). Furthermore, microbial growth experiments of foodborne bacteria, i.e., Listeria monocytogenes, Escherichia coli, Pseudomonas aeruginosa and Lactococcus lactis on the surface of the 3D models revealed very interesting results, regarding the growth dynamics and distribution of cells at colony level. More specifically, the size of the colonies formed on the surface of the 3D models, increased substantially for increasing fat concentrations, especially in mid- and late-exponential growth phases. Furthermore, colonies formed in proximity to fat were substantially larger as compared to the ones that were located far from the fat phase of the models. In terms of growth location, the majority of colonies were located on the protein/polysaccharide phase of the 3D models. All those differences at microscopic level, that can directly affect the bacterial response to decontamination treatments, were not captured by the macroscopic kinetics (growth dynamics), which were unaffected from changes in fat concentration. Our findings demonstrate the importance of developing structurally and biochemically complex 3D in vitro models (for closer proximity to industrial products), as well as the necessity of conducting multi-level microbial analyses, to better understand and predict the bacterial behaviour in relation to their biochemical and structural environment. Such studies in advanced 3D environments can assist a better/more accurate design of industrial antimicrobial processes, ultimately, improving food safety.

Bile Microbiome Signatures Associated with Pancreatic Ductal Adenocarcinoma Compared to Benign Disease: A UK Pilot Study.

Pancreatic ductal adenocarcinoma (PDAC) has a very poor survival. The intra-tumoural microbiome can influence pancreatic tumourigenesis and chemoresistance and, therefore, patient survival. The role played by bile microbiota in PDAC is unknown. We aimed to define bile microbiome signatures that can effectively distinguish malignant from benign tumours in patients presenting with obstructive jaundice caused by benign and malignant pancreaticobiliary disease. Prospective bile samples were obtained from 31 patients who underwent either Endoscopic Retrograde Cholangiopancreatography (ERCP) or Percutaneous Transhepatic Cholangiogram (PTC). Variable regions (V3-V4) of the 16S rRNA genes of microorganisms present in the samples were amplified by Polymerase Chain Reaction (PCR) and sequenced. The cohort consisted of 12 PDAC, 10 choledocholithiasis, seven gallstone pancreatitis and two primary sclerosing cholangitis patients. Using the 16S rRNA method, we identified a total of 135 genera from 29 individuals (12 PDAC and 17 benign). The bile microbial beta diversity significantly differed between patients with PDAC vs. benign disease (Permanova p = 0.0173). The separation of PDAC from benign samples is clearly seen through unsupervised clustering of Aitchison distance. We found three genera to be of significantly lower abundance among PDAC samples vs. benign, adjusting for false discovery rate (FDR). These were Escherichia (FDR = 0.002) and two unclassified genera, one from Proteobacteria (FDR = 0.002) and one from Enterobacteriaceae (FDR = 0.011). In the same samples, the genus Streptococcus (FDR = 0.033) was found to be of increased abundance in the PDAC group. We show that patients with obstructive jaundice caused by PDAC have an altered microbiome composition in the bile compared to those with benign disease. These bile-based microbes could be developed into potential diagnostic and prognostic biomarkers for PDAC and warrant further investigation.

On the evaluation of the antimicrobial effect of grape seed extract and cold atmospheric plasma on the dynamics of Listeria monocytogenes in novel multiphase 3D viscoelastic models.

The demand for products that are minimally processed and produced in a sustainable way, without the use of chemical preservatives or antibiotics have increased over the last years. Novel non-thermal technologies such as cold atmospheric plasma (CAP) and natural antimicrobials such as grape seed extract (GSE) are attractive alternatives to conventional food decontamination methods as they can meet the above demands. The aim of this study was to investigate the microbial inactivation potential of GSE, CAP (in this case, a remote air plasma with an ozone-dominated RONS output) and their combination against L. monocytogenes on five different 3D in vitro models of varying rheological, structural, and biochemical composition. More specifically, we studied the microbial dynamics, as affected by 1 % (w/v) GSE, CAP or their combination, in three monophasic Xanthan Gum (XG) based 3D models of relatively low viscosity (1.5 %, 2.5 % and 5 % w/v XG) and in a biphasic XG/Whey Protein (WPI) and a triphasic XG/WPI/fat model. A significant microbial inactivation (comparable to liquid broth) was achieved in presence of GSE on the surface of all monophasic models regardless of their viscosity. In contrast, the GSE antimicrobial effect was diminished in the multiphasic systems, resulting to only a slight disturbance of the microbial growth. In contrast, CAP showed better antimicrobial potential on the surface of the complex multiphasic models as compared to the monophasic models. When combined, in a hurdle approach, GSE/CAP showed promising microbial inactivation potential in all our 3D models, but less microbial inactivation in the structurally and biochemically complex multiphasic models, with respect to the monophasic models. The level of inactivation also depended on the duration of the exposure to GSE. Our results contribute towards understanding the antimicrobial efficacy of GSE, CAP and their combination as affected by robustly controlled changes of rheological and structural properties and of the biochemical composition of the environment in which bacteria grow. Therefore, our results contribute to the development of sustainable food safety strategies.

Single-Cell Lipidomics Using Analytical Flow LC-MS Characterizes the Response to Chemotherapy in Cultured Pancreatic Cancer Cells.

In this work, we demonstrate the development and first application of nanocapillary sampling followed by analytical flow liquid chromatography-mass spectrometry for single-cell lipidomics. Around 260 lipids were tentatively identified in a single cell, demonstrating remarkable sensitivity. Human pancreatic ductal adenocarcinoma cells (PANC-1) treated with the chemotherapeutic drug gemcitabine can be distinguished from controls solely on the basis of their single-cell lipid profiles. Notably, the relative abundance of LPC(0:0/16:0) was significantly affected in gemcitabine-treated cells, in agreement with previous work in bulk. This work serves as a proof of concept that live cells can be sampled selectively and then characterized using automated and widely available analytical workflows, providing biologically relevant outputs.

A Step-by-Step Methodological Guide for Developing Zonal Multicellular Scaffold-Based Pancreatic Cancer Models.

The tumor microenvironment (TME), a complex heterogeneous mixture of various cellular, physical, and biochemical components and signals, is a major player in the process of tumor growth and its response to therapeutic methods. In vitro 2D monocellular cancer models are unable to mimic the complex in vivo characteristics of cancer TME involving cellular heterogeneity, presence of extracellular matrix (ECM) proteins, as well as spatial orientation and organization of different cell types forming the TME. In vivo animal-based studies have ethical concerns, are expensive and time-consuming, and involve models of non-human species. In vitro 3D models are capable of tiding over several issues associated with both 2D in vitro and in vivo animal models. We have recently developed a novel zonal multicellular 3D in vitro model for pancreatic cancer involving cancer cells, endothelial cells, and pancreatic stellate cells. Our model (i) can provide long-term culture (up to 4 weeks), (ii) can control the ECM biochemical configuration in a cell specific manner, (iii) shows large amounts of collagen secretion by the stellate cells mimicking desmoplasia, and (iv) expresses cell-specific markers throughout the whole culture period. This chapter describes the experimental methodology to form our hybrid multicellular 3D model for pancreatic ductal adenocarcinoma, including the immunofluorescence staining on the cell culture.

A Systematic Quantitative Determination of the Antimicrobial Efficacy of Grape Seed Extract against Foodborne Bacterial Pathogens.

Concerns regarding the role of antimicrobial resistance (AMR) in disease outbreaks are growing due to the excessive use of antibiotics. Moreover, consumers are demanding food products that are minimally processed and produced in a sustainable way, without the use of chemical preservatives or antibiotics. Grape seed extract (GSE) is isolated from wine industry waste and is an interesting source of natural antimicrobials, especially when aiming to increase sustainable processing. The aim of this study was to obtain a systematic understanding of the microbial inactivation efficacy/potential of GSE against Listeria monocytogenes (Gram-positive), Escherichia coli and Salmonella Typhimurium (Gram-negative) in an in vitro model system. More specifically, for L. monocytogenes, the effects of the initial inoculum concentration, bacterial growth phase and absence of the environmental stress response regulon (SigB) on the GSE microbial inactivation potential were investigated. In general, GSE was found to be highly effective at inactivating L. monocytogenes, with higher inactivation achieved for higher GSE concentrations and lower initial inoculum levels. Generally, stationary phase cells were more resistant/tolerant to GSE as compared to exponential phase cells (for the same inoculum level). Additionally, SigB appears to play an important role in the resistance of L. monocytogenes to GSE. The Gram-negative bacteria under study (E. coli and S. Typhimurium) were less susceptible to GSE as compared to L. monocytogenes. Our findings provide a quantitative and mechanistic understanding of the impact of GSE on the microbial dynamics of foodborne pathogens, assisting in the more systematic design of natural antimicrobial-based strategies for sustainable food safety.

Deodorants and antiperspirants: New trends in their active agents and testing methods.

Sweating is the human body's thermoregulation system but also results in unpleasant body odour which can diminish the self-confidence of people. There has been continued research in finding solutions to reduce both sweating and body odour. Sweating is a result of increased sweat flow and malodour results from certain bacteria and ecological factors such as eating habits. Research on deodorant development focuses on inhibiting the growth of malodour-forming bacteria using antimicrobial agents, whereas research on antiperspirant synthesis focuses on technologies reducing the sweat flow, which not only reduces body odour but also improves people's appearance. Antiperspirant's technology is based on the use of aluminium salts which can form a gel plug at sweat pores, obstructing the sweat fluid from arising onto the skin surface. In this paper, we perform a systematic review on the recent progress in the development of novel antiperspirant and deodorant active ingredients that are alcohol-free, paraben-free, and naturally derived. Several studies have been reported on the alternative class of actives that can potentially be used for antiperspirant and body odour treatment including deodorizing fabric, bacterial, and plant extracts. However, a significant challenge is to understand how the gel-plugs of antiperspirant actives are formed in sweat pores and how to deliver long-lasting antiperspirant and deodorant benefits.

La transpiration est le système de thermorégulation de l'organisme, mais elle entraîne également une odeur corporelle désagréable qui peut diminuer la confiance en soi. Des nombreuses recherches ont été menées afin de trouver des solutions pour réduire à la fois la transpiration et l'odeur corporelle. La transpiration est le résultat de l'augmentation du flux de sueur, et les mauvaises odeurs sont dues à certaines bactéries et à certains facteurs écologiques tels que les habitudes alimentaires. Les recherches sur le développement des déodorants se concentrent sur l'inhibition de la croissance des bactéries responsables des mauvaises odeurs à l'aide d'agents antimicrobiens, tandis que les recherches sur la synthèse des anti-transpirants se concentrent sur les technologies diminuant le flux de sueur, ce qui réduire non seulement les odeurs corporelles, mais améliore également l'apparence des personnes. La technologie des anti-transpirants repose sur l'utilisation de sels d'aluminium qui peuvent former un bouchon de gel au niveau des pores sudoripares, empêchant le liquide sudoral d'apparaître à la surface de la peau. Dans cet article, nous effectuons une revue systématique des progrès récents réalisés dans le développement de nouveaux principes actifs anti-transpirants et déodorants qui sont sans alcool, sans parabène et d'origine naturelle. Plusieurs études ont été rapportées sur la classe alternative de principes actifs qui peuvent potentiellement être utilisés pour le traitement anti-transpirant et des odeurs corporelles, y compris les tissus désodorisants, les bactéries et les extraits végétaux. Cependant, un défi important consiste à comprendre comment les bouchons de gel des actifs anti-transpirants se forment au niveau des pores sudoripares, et comment offrir des effets anti-transpirants et déodorants durables.

Nanocapillary sampling coupled to liquid chromatography mass spectrometry delivers single cell drug measurement and lipid fingerprints.

This work describes the development of a new approach to measure drug levels and lipid fingerprints in single living mammalian cells. Nanocapillary sampling is an approach that enables the selection and isolation of single living cells under microscope observation. Here, live single cell nanocapillary sampling is coupled to liquid chromatography for the first time. This allows molecular species to be separated prior to ionisation and improves measurement precision of drug analytes. The efficiency of transferring analytes from the sampling capillary into a vial was optimised in this work. The analysis was carried out using standard flow liquid chromatography coupled to widely available mass spectrometry instrumentation, highlighting opportunities for widespread adoption. The method was applied to 30 living cells, revealing cell-to-cell heterogeneity in the uptake of different drug molecules. Using this system, we detected 14-158 lipid features per single cell, revealing the association between bedaquiline uptake and lipid fingerprints.

Enhanced effect of X-rays in the presence of a static magnetic field within a 3D pancreatic cancer model.

OBJECTIVE: To evaluate the impact of static magnetic field (SMF) presence on the radiation response of pancreatic cancer cells in polyurethane-based highly macro-porous scaffolds in hypoxic (1% O2) and normoxic (21% O2) conditions, towards understanding MR-guided radiotherapy, shedding light on the potential interaction phenomenon between SMF and radiation in a three-dimensional (3D) microenvironment. METHODS: Pancreatic cancer cells (PANC-1, ASPC-1) were seeded into fibronectin-coated highly porous polyethene scaffolds for biomimicry and cultured for 4 weeks in in vitro normoxia (21% O2) followed by a 2-day exposure to either in vitro hypoxia (1% O2) or maintenance in in vitro normoxia (21% O2). The samples were then irradiated with 6 MV photons in the presence or absence of a 1.5 T field. Thereafter, in situ post-radiation monitoring (1 and 7 days post-irradiation treatment) took place via quantification of (i) live dead and (ii) apoptotic profiles. RESULTS: We report: (i) pancreatic ductal adenocarcinoma hypoxia-associated radioprotection, in line with our previous findings, (ii) an enhanced effect of radiation in the presence of SMFin in vitro hypoxia (1% O2) for both short- (1 day) and long-term (7 days) post -radiation analysis and (iii) an enhanced effect of radiation in the presence of SMF in in vitro normoxia (21% O2) for long-term (7 days) post-radiation analysis within a 3D pancreatic cancer model. CONCLUSION: With limited understanding of the potential interaction phenomenon between SMF and radiation, this 3D system allows combination evaluation for a cancer in which the role of radiotherapy is still evolving. ADVANCES IN KNOWLEDGE: This study examined the use of a 3D model to investigate MR-guided radiotherapy in a hypoxic microenvironment, indicating that this could be a useful platform to further understanding of SMF influence on radiation.

The Development of a Three-Dimensional Platform for Patient-Derived Ovarian Cancer Tissue Models: A Systematic Literature Review.

There is an unmet biomedical need for ex vivo tumour models that would predict drug responses and in turn help determine treatment regimens and potentially predict resistance before clinical studies. Research has shown that three dimensional models of ovarian cancer (OvCa) are more realistic than two dimensional in vitro systems as they are able to capture patient in vivo conditions in more accurate manner. The vast majority of studies aiming to recapitulate the ovarian tumour morphology, behaviors, and study chemotherapy responses have been using ovarian cancer cell lines. However, despite the advantages of utilising cancer cell lines to set up a platform, they are not as informative as systems applying patient derived cells, as cell lines are not able to recapitulate differences between each individual patient characteristics. In this review we discussed the most recent advances in the creation of 3D ovarian cancer models that have used patient derived material, the challenges to overcome and future applications.

Current strategies with implementation of three-dimensional cell culture: the challenge of quantification.

From growing cells in spheroids to arranging them on complex engineered scaffolds, three-dimensional cell culture protocols are rapidly expanding and diversifying. While these systems may often improve the physiological relevance of cell culture models, they come with technical challenges, as many of the analytical methods used to characterize traditional two-dimensional (2D) cells must be modified or replaced to be effective. Here we review the advantages and limitations of quantification methods based either on biochemical measurements or microscopy imaging. We focus on the most basic of parameters that one may want to measure, the number of cells. Precise determination of this number is essential for many analytical techniques where measured quantities are only meaningful when normalized to the number of cells (e.g. cytochrome p450 enzyme activity). Thus, accurate measurement of cell number is often a prerequisite to allowing comparisons across different conditions (culturing conditions or drug and treatment screening) or between cells in different spatial states. We note that this issue is often neglected in the literature with little or no information given regarding how normalization was performed, we highlight the pitfalls and complications of quantification and call for more accurate reporting to improve reproducibility.

Pericytes' Circadian Clock Affects Endothelial Cells' Synchronization and Angiogenesis in a 3D Tissue Engineered Scaffold.

Angiogenesis, the formation of new capillaries from existing ones, is a fundamental process in regenerative medicine and tissue engineering. While it is known to be affected by circadian rhythms in vivo, its peripheral regulation within the vasculature and the role it performs in regulating the interplay between vascular cells have not yet been investigated. Peripheral clocks within the vasculature have been described in the endothelium and in smooth muscle cells. However, to date, scarce evidence has been presented regarding pericytes, a perivascular cell population deeply involved in the regulation of angiogenesis and vessel maturation, as well as endothelial function and homeostasis. More crucially, pericytes are also a promising source of cells for cell therapy and tissue engineering. Here, we established that human primary pericytes express key circadian genes and proteins in a rhythmic fashion upon synchronization. Conversely, we did not detect the same patterns in cultured endothelial cells. In line with these results, pericytes' viability was disproportionately affected by circadian cycle disruption, as compared to endothelial cells. Interestingly, endothelial cells' rhythm could be induced following exposure to synchronized pericytes in a contact co-culture. We propose that this mechanism could be linked to the altered release/uptake pattern of lactate, a known mediator of cell-cell interaction which was specifically altered in pericytes by the knockout of the key circadian regulator Bmal1. In an angiogenesis assay, the maturation of vessel-like structures was affected only when both endothelial cells and pericytes did not express Bmal1, indicating a compensation system. In a 3D tissue engineering scaffold, a synchronized clock supported a more structured organization of cells around the scaffold pores, and a maturation of vascular structures. Our results demonstrate that pericytes play a critical role in regulating the circadian rhythms in endothelial cells, and that silencing this system disproportionately affects their pro-angiogenic function. Particularly, in the context of tissue engineering and regenerative medicine, considering the effect of circadian rhythms may be critical for the development of mature vascular structures and to obtain the maximal reparative effect.

Biocompatibility and Physiological Thiolytic Degradability of Radically Made Thioester-Functional Copolymers: Opportunities for Drug Release.

Being nondegradable, vinyl polymers have limited biomedical applicability. Unfortunately, backbone esters incorporated through conventional radical ring-opening methods do not undergo appreciable abiotic hydrolysis under physiologically relevant conditions. Here, PEG acrylate and di(ethylene glycol) acrylamide-based copolymers containing backbone thioesters were prepared through the radical ring-opening copolymerization of the thionolactone dibenzo[c,e]oxepin-5(7H)-thione. The thioesters degraded fully in the presence of 10 mM cysteine at pH 7.4, with the mechanism presumed to involve an irreversible S-N switch. Degradations with N-acetylcysteine and glutathione were reversible through the thiol-thioester exchange polycondensation of R-SC(═O)-polymer-SH fragments with full degradation relying on an increased thiolate/thioester ratio. Treatment with 10 mM glutathione at pH 7.2 (mimicking intracellular conditions) triggered an insoluble-soluble switch of a temperature-responsive copolymer at 37 °C and the release of encapsulated Nile Red (as a drug model) from core-degradable diblock copolymer micelles. Copolymers and their cysteinolytic degradation products were found to be noncytotoxic, making thioester backbone-functional polymers promising for drug delivery applications.

Interfacial Superassembly of Light-Responsive Mechanism-Switchable Nanomotors with Tunable Mobility and Directionality.

Mechanism-switchable nanomotors are expected to exhibit high adaptability and wide applicability. Herein, for the first time, we report a flask-shaped carbon@Pt@fatty-acid nanomotor with a light-induced switch between nonionic self-diffusiophoresis and bubble propulsion. This nanomotor is fabricated through superassembly of platinum nanoparticles on the surface of carbon nanobottles, and fatty acids are infused into the cavity of carbon nanobottles to serve as a light-sensitive switch. Such a nanomotor can be propelled via catalytic decomposition of H2O2 by platinum nanoparticles, exhibiting self-diffusiophoresis with opening-forward migration. Upon 980 nm laser irradiation, the fatty acids melt due to the photothermal effect and are released from the cavity, switching the dominant operational mechanism to bubble propulsion with bottom-forward migration. Compared with self-diffusiophoresis, bubble propulsion shows higher mobility and better directionality due to the hindered self-rotation. Simulation results further reveal that the confinement effect of the cavity, which facilitates the nucleation of nanobubbles, leads to the switch to bubble propulsion. This study offers an insight into the relationship between nanostructures, fundamental nanomotor operational mechanisms, and apparent propulsion performance, as well as provides a novel strategy for the regulation of movement, which is instructive for both the design and applications of nanomotors.

A Systematic Comparative Assessment of the Response of Ovarian Cancer Cells to the Chemotherapeutic Cisplatin in 3D Models of Various Structural and Biochemical Configurations-Does One Model Type Fit All?

Epithelial Ovarian Cancer (EOC) is a silent, deadly and aggressive gynaecological disease with a relatively low survival rate. This has been attributed, to some extent, to EOC's high recurrence rate and resistance to currently available platinum-based chemotherapeutic treatment methods. Multiple groups have studied and reported the effect of chemotherapeutic agents on various EOC 3D in vitro models. However, there are very few studies wherein a direct comparative study has been carried out between the different in vitro 3D models of EOC and the effect of chemotherapy within them. Herein, we report, for the first time, a direct comprehensive systematic comparative study of three different 3D in vitro platforms, namely (i) spheroids, (ii) synthetic PeptiGels/hydrogels of various chemical configurations and (iii) polymeric scaffolds with coatings of various extracellular matrices (ECMs) on the cell growth and response to the chemotherapeutic (Cisplatin) for ovary-derived (A2780) and metastatic (SK-OV-3) EOC cell lines. We report that all three 3D models are able to support the growth of EOC, but for different time periods (varying from 7 days to 4 weeks). We have also reported that chemoresistance to Cisplatin, in vitro, observed especially for metastatic EOC cells, is platform-dependent, in terms of both the structural and biochemical composition of the model/platform. Our study highlights the importance of selecting an appropriate 3D platform for in vitro tumour model development. We have demonstrated that the selection of the best platform for producing in vitro tumour models depends on the cancer/cell type, the experimental time period and the application for which the model is intended.