Lost in Rotation: How TiO2 and ZnO Nanoparticles Disrupt Coordinated Epithelial Cell Rotation.

Coordinated cell movement is a cardinal feature in tissue organization that highlights the importance of cells working together as a collective unit. Disruptions to this synchronization can have far-reaching pathological consequences, ranging from developmental disorders to tissue repair impairment. Herein, it is shown that metal oxide nanoparticles (NPs), even at low and non-toxic doses (1 and 10 µg mL-1), can perturb the coordinated epithelial cell rotation (CECR) in micropatterned human epithelial cell clusters via distinct nanoparticle-specific mechanisms. Zinc oxide (ZnO) NPs are found to induce significant levels of intracellular reactive oxygen species (ROS) to promote mitogenic activity. Generation of a new localized force field through changes in the cytoskeleton organization and an increase in cell density leads to the arrest of CECR. Conversely, epithelial cell clusters exposed to titanium dioxide (TiO2) NPs maintain their CECR directionality but display suppressed rotational speed in an autophagy-dependent manner. Thus, these findings reveal that nanoparticles can actively hijack the nano-adaptive responses of epithelial cells to disrupt the fundamental mechanics of cooperation and communication in a collective setting.

3D Airway Epithelial-Fibroblast Biomimetic Microfluidic Platform to Unravel Engineered Nanoparticle-Induced Acute Stress Responses as Exposome Determinants.

Insights into how biological systems respond to high- and low-dose acute environmental stressors are a fundamental aspect of exposome research. However, studying the impact of low-level environmental exposure in conventional in vitro settings is challenging. This study employed a three-dimensional (3D) biomimetic microfluidic lung-on-chip (µLOC) platform and RNA-sequencing to examine the effects of two model anthropogenic engineered nanoparticles (NPs): zinc oxide nanoparticles (Nano-ZnO) and copier center nanoparticles (Nano-CCP). The airway epithelium exposed to these NPs exhibited dose-dependent increases in cytotoxicity and barrier dysregulation (dominance of the external exposome). Interestingly, even nontoxic and low-level exposure (10 µg/mL) of the epithelium compartment to Nano-ZnO triggered chemotaxis of lung fibroblasts toward the epithelium. An increase in α smooth muscle actin (α-SMA) expression and contractile activity was also observed in these cells, indicating a bystander-like adaptive response (dominance of internal exposome). Further bioinformatics and network analysis showed that a low-dose Nano-ZnO significantly induced a robust transcriptomic response and upregulated several hub genes associated with the development of lung fibrosis. We propose that Nano-ZnO, even at a no observable effect level (NOEL) dose according to conventional standards, can function as a potent nanostressor to disrupt airway epithelium homeostasis. This leads to a cascade of profibrotic events in a cross-tissue compartment fashion. Our findings offer new insights into the early acute events of respiratory harm associated with environmental NPs exposure, paving the way for better exposomic understanding of this emerging class of anthropogenic nanopollutants.

Direct and Label-Free Cell Status Monitoring of Spheroids and Microcarriers Using Microfluidic Impedance Cytometry.

3D cellular spheroids/microcarriers (100 µm-1 mm) are widely used in biomanufacturing, and non-invasive biosensors are useful to monitor cell quality in bioprocesses. In this work, a novel microfluidic approach for label-free and continuous-flow monitoring of single spheroid/microcarrier (hydrogel and Cytodex) based on electrical impedance spectroscopy using co-planar Field's metal electrodes is reported. Through numerical simulation and experimental validation, two unique impedance signatures (|ZLF | (60 kHz), |ZHF | (1 MHz)) which are optimal for spheroid growth and viability monitoring are identified. Using a closed-loop recirculation system, it is demonstrated that |ZLF | increases with breast cancer (MCF-7) spheroid biomass, while higher opacity (impedance ratio |ZHF |/|ZLF |) indicates cell death due to compromised cell membrane. Anti-cancer drug (paclitaxel)-treated spheroids also exhibit lower |ZLF | with increased cell dissociation. Interestingly, impedance characterization of adipose-derived mesenchymal stem cell differentiation on Cytodex microcarriers reveals that adipogenic cells (higher intracellular lipid content) exhibit higher impedance than osteogenic cells (more conductive due to calcium ions) for both microcarriers and single cell level. Taken together, the developed platform offers great versatility for multi-parametric analysis of spheroids/microcarriers at high throughput (≈1 particle/s), and can be readily integrated into bioreactors for long-term and remote monitoring of biomass and cell quality.

Multitrack Compressed Sensing for Faster Hyperspectral Imaging.

Hyperspectral imaging (HSI) provides additional information compared to regular color imaging, making it valuable in areas such as biomedicine, materials inspection and food safety. However, HSI is challenging because of the large amount of data and long measurement times involved. Compressed sensing (CS) approaches to HSI address this, albeit subject to tradeoffs between image reconstruction accuracy, time and generalizability to different types of scenes. Here, we develop improved CS approaches for HSI, based on parallelized multitrack acquisition of multiple spectra per shot. The multitrack architecture can be paired up with either of the two compatible CS algorithms developed here: (1) a sparse recovery algorithm based on block compressed sensing and (2) an adaptive CS algorithm based on sampling in the wavelet domain. As a result, the measurement speed can be drastically increased while maintaining reconstruction speed and accuracy. The methods were validated computationally both in noiseless as well as noisy simulated measurements. Multitrack adaptive CS has a ∼10 times shorter measurement plus reconstruction time as compared to full sampling HSI without compromising reconstruction accuracy across the sample images tested. Multitrack non-adaptive CS (sparse recovery) is most robust against Poisson noise at the expense of longer reconstruction times.

Inflammation Increases Susceptibility of Human Small Airway Epithelial Cells to Pneumonic Nanotoxicity.

Exposure to inhaled anthropogenic nanomaterials (NM) with dimension <100 nm has been implicated in numerous adverse respiratory outcomes. Although studies have identified key NM physiochemical determinants of pneumonic nanotoxicity, the complex interactive and cumulative effects of NM exposure, especially in individuals with preexisting inflammatory respiratory diseases, remain unclear. Herein, the susceptibility of primary human small airway epithelial cells (SAEC) exposed to a panel of reference NM, namely, CuO, ZnO, mild steel welding fume (MSWF), and nanofractions of copier center particles (Nano-CCP), is examined in normal and tumor necrosis factor alpha (TNF-α)-induced inflamed SAEC. Compared to normal SAEC, inflamed cells display an increased susceptibility to NM-induced cytotoxicity by 15-70% due to a higher basal level of intracellular reactive oxygen species (ROS). Among the NM screened, ZnO, CuO, and Nano-CCP are observed to trigger an overcompensatory response in normal SAEC, resulting in an increased tolerance against subsequent oxidative insults. However, the inflamed SAEC fails to adapt to the NM exposure due to an impaired nuclear factor erythroid 2-related factor 2 (Nrf2)-mediated cytoprotective response. The findings reveal that susceptibility to pulmonary nanotoxicity is highly dependent on the interplay between NM properties and inflammation of the alveolar milieu.

Potent-By-Design: Amino Acids Mimicking Porous Nanotherapeutics with Intrinsic Anticancer Targeting Properties.

Exogenous sources of amino acids are essential nutrients to fuel cancer growth. Here, the increased demand for amino acid displayed by cancer cells is unconventionally exploited as a design principle to replete cancer cells with apoptosis inducing nanoscopic porous amino acid mimics (Nano-PAAM). A small library consisting of nine essential amino acids nanoconjugates (30 nm) are synthesized, and the in vitro anticancer activity is evaluated. Among the Nano-PAAMs, l-phenylalanine functionalized Nano-PAAM (Nano-pPAAM) has emerged as a novel nanotherapeutics with excellent intrinsic anticancer and cancer-selective properties. The therapeutic efficacy of Nano-pPAAM against a panel of human breast, gastric, and skin cancer cells could be ascribed to the specific targeting of the overexpressed human large neutral amino acid transporter SLC7A5 (LAT-1) in cancer cells, and its intracellular reactive oxygen species (ROS) inducing properties of the nanoporous core. At the mechanistic level, it is revealed that Nano-pPAAM could activate both the extrinsic and intrinsic apoptosis pathways to exert a potent "double-whammy" anticancer effect. The potential clinical utility of Nano-pPAAM is further investigated using an MDA-MB-231 xenograft in NOD scid gamma mice, where an overall suppression of tumor growth by 60% is achieved without the aid of any drugs or application of external stimuli.

Repurposing of Fruit Peel Waste as a Green Reductant for Recycling of Spent Lithium-Ion Batteries.

The development of environmentally benign hydrometallurgical processes to treat spent lithium-ion batteries (LIBs) is a critical aspect of the electronic-waste circular economy. Herein, as an alternative to the highly explosive H2O2, discarded orange peel powder (OP) is valorized as a green reductant for the leaching of industrially produced LIBs scraps in citric acid (H3Cit) lixiviant. The reductive potential of the cellulose- and antioxidant-rich OP was validated using the 3,5-dinitrosalicylic acid and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic) acid assays. Leaching parameters such as OP concentration (200 mg), processing temperature (100 °C), H3Cit concentration (1.5 M), reaction duration (4 h), and slurry density (25 g/mL) were systematically optimized to achieve 80-99% leaching efficiencies of Ni, Mn, Co, and Li from the LIB "black mass". Importantly, solid side-streams generated by the OP-enabled leaching displayed negligible cytotoxicity in three different human cell lines, suggesting that the process is environmentally safe. As a proof of concept, Co(OH)2 was selectively recovered from the green lixiviant and subsequently utilized to fabricate new batches of LiCoO2 (LCO) coin cell batteries. Galvanostatic charge-discharge test revealed that the regenerated batteries exhibited initial charge and discharge values of 120 and 103 mAh/g, respectively, which is comparable to the performance of commercial LCO batteries. The use of fruit peel waste to recover valuable metals from spent LIBs is an effective, ecofriendly, and sustainable strategy to minimize the environmental footprint of both waste types.

Interpenetrating Network of Alginate-Human Adipose Extracellular Matrix Hydrogel for Islet Cells Encapsulation.

Transplantation of microencapsulated islet cells holds great potential for the treatment of type 1 diabetes mellitus. However, its clinical translation is hampered by the peri-transplantation loss of islet viability and functionality in the microcapsules. In this work, a novel islet cells biomimetic microencapsulant material that is based on the interpenetrating networks of alginate and extracellular matrix (ECM) hydrogel composite (AEC) is presented. The ECM component is derived from human lipoaspirate. In situ encapsulation of pancreatic ß islet cells (MIN6 ß-cells) can be achieved via ionotropic gelation of the alginate matrix and thermal-induced gelation of the pepsin-solubilized ECM pre-gel. Due to the enhanced cell-matrix interaction, islets encapsulated within the AEC microcapsules (≈640 µm) display sevenfold increase in cell growth over 1 week of culture and characteristic glucose-stimulated insulin response in vitro. The results show that the AEC microcapsule is a potent platform to bioaugment the performance of islet cells.

Mechanoregulation of Cancer-Associated Fibroblast Phenotype in Three-Dimensional Interpenetrating Hydrogel Networks.

Tumor stromal residing cancer-associated fibroblasts (CAFs) are significant accomplices in the growth and development of malignant neoplasms. As cancer progresses, the stroma undergoes a dramatic remodeling and stiffening of its extracellular matrix (ECM). However, exactly how these biomechanical changes influence the CAF behavior and the functional paracrine crosstalk with the neighboring tumor cells in a 3-dimensional (3D) microenvironment remains elusive. Herein, a collagen and alginate interpenetrating network (CoAl-IPN) hydrogel system was employed as a 3D in vitro surrogate of the cancerous breast tissue stromal niche. In this study, the mechanical properties of CoAl-IPN were precisely fine-tuned with Young's modulus ( E) values of ∼108 and 898 Pa. The results revealed that the 3D polymeric network mechanics and microstructure are critical biophysical determinants of the human breast CAF (b-CAF) morphology, phenotype, and paracrine dialogue with MDA-MB-231 tumoroids. A compliant hydrogel network favors b-CAF spreading, nuclear translocation of the YAP/TAZ mechanosignaling protein, and upregulation of CAF hallmark transcripts. Conversely, a rigid and highly cross-linked hydrogel network imposed a physical entrapment effect on the b-CAFs that limited their spreading and phenotype in a manner that effectively muted their pro-tumorigenic paracrine activity. Collectively, the CoAl-IPN 3D culture system has proven to be a versatile platform in defining the 3D biophysical parameters that could either promote or restrain the protumorigenic activity of b-CAFs and sheds critical mechano-mediated light onto the phenotypic plasticity and corresponding specific bioactivity of b-CAFs in the 3D microenvironment.

Cellular processing and destinies of artificial DNA nanostructures.

Since many bionanotechnologies are targeted at cells, understanding how and where their interactions occur and the subsequent results of these interactions is important. Changing the intrinsic properties of DNA nanostructures and linking them with interactions presents a holistic and powerful strategy for understanding dual nanostructure-biological systems. With the recent advances in DNA nanotechnology, DNA nanostructures present a great opportunity to understand the often convoluted mass of information pertaining to nanoparticle-biological interactions due to the more precise control over their chemistry, sizes, and shapes. Coupling just some of these designs with an understanding of biological processes is both a challenge and a source of opportunities. Despite continuous advances in the field of DNA nanotechnology, the intracellular fate of DNA nanostructures has remained unclear and controversial. Because understanding its cellular processing and destiny is a necessary prelude to any rational design of exciting and innovative bionanotechnology, in this review, we will discuss and provide a comprehensive picture relevant to the intracellular processing and the fate of various DNA nanostructures which have been remained elusive for some time. We will also link the unique capabilities of DNA to some novel ideas for developing next-generation bionanotechnologies.

Decoupling the Direct and Indirect Biological Effects of ZnO Nanoparticles Using a Communicative Dual Cell-Type Tissue Construct.

While matter at the nanoscale can be manipulated, the knowledge of the interactions between these nanoproducts and the biological systems remained relatively laggard. Current nanobiology study is rooted on in vitro study using conventional 2D cell culture model. A typical study employs monolayer cell culture that simplifies the real context of which to measure any nanomaterial effect; unfortunately, this simplification also demonstrated the limitations of 2D cell culture in predicting the actual biological response of some tissues. In fact, some of the characteristics of tissue such as spatial arrangement of cells and cell-cell interaction, which are simplified in 2D cell culture model, play important roles in how cells respond to a stimulus. To more accurately recapitulate the features and microenvironment of tissue for nanotoxicity assessments, an improved organotypic-like in vitro multicell culture system to mimic the kidney endoepithelial bilayer is introduced. Results showed that important nano-related parameters such as the diffusion, direct and indirect toxic effects of ZnO nanoparticles can be studied by combining this endoepithelial bilayer tissue model and traditional monolayer culture setting.

Understanding and exploiting nanoparticles' intimacy with the blood vessel and blood.

While the blood vessel is seldom the target tissue, almost all nanomedicine will interact with blood vessels and blood at some point of time along its life cycle in the human body regardless of their intended destination. Despite its importance, many bionanotechnologists do not feature endothelial cells (ECs), the blood vessel cells, or consider blood effects in their studies. Including blood vessel cells in the study can greatly increase our understanding of the behavior of any given nanomedicine at the tissue of interest or to understand side effects that may occur in vivo. In this review, we will first describe the diversity of EC types found in the human body and their unique behaviors and possibly how these important differences can implicate nanomedicine behavior. Subsequently, we will discuss about the protein corona derived from blood with foci on the physiochemical aspects of nanoparticles (NPs) that dictate the protein corona characteristics. We would also discuss about how NPs characteristics can affect uptake by the endothelium. Subsequently, mechanisms of how NPs could cross the endothelium to access the tissue of interest. Throughout the paper, we will share some novel nanomedicine related ideas and insights that were derived from the understanding of the NPs' interaction with the ECs. This review will inspire more exciting nanotechnologies that had accounted for the complexities of the real human body.

Mechanistic Investigation of the Biological Effects of SiO2, TiO2, and ZnO Nanoparticles on Intestinal Cells.

Silicon dioxide (SiO2), titanium dioxide (TiO2), and zinc oxide (ZnO) are currently among the most widely used nanoparticles (NPs) in the food industry. This could potentially lead to unintended exposure of the gastrointestinal tract to these NPs. This study aims to investigate the potential side-effects of these food-borne NPs on intestinal cells and to mechanistically understand the observed biological responses. Among the panel of tested NPs, ZnO NPs are the most toxic. Consistently in all three tested intestinal cell models, ZnO NPs invoke the most inflammatory responses from the cells and induce the highest intracellular production of reactive oxygen species (ROS). The elevated ROS levels induce significant damage to the DNA of the cells, resulting in cell-cycle arrest and subsequently cell death. In contrast, both SiO2 and TiO2 NPs elicit minimum biological responses from the intestinal cells. Overall, the study showcases the varying capability of the food-borne NPs to induce a cellular response in the intestinal cells. In addition to physicochemical differences in the NPs, the genetic landscape of the intestinal cell models governs the toxicology profile of these food-borne NPs.

Biomimicry 3D gastrointestinal spheroid platform for the assessment of toxicity and inflammatory effects of zinc oxide nanoparticles.

Our current mechanistic understanding on the effects of engineered nanoparticles (NPs) on cellular physiology is derived mainly from 2D cell culture studies. However, conventional monolayer cell culture may not accurately model the mass transfer gradient that is expected in 3D tissue physiology and thus may lead to artifactual experimental conclusions. Herein, using a micropatterned agarose hydrogel platform, the effects of ZnO NPs (25 nm) on 3D colon cell spheroids of well-defined sizes are examined. The findings show that cell dimensionality plays a critical role in governing the spatiotemporal cellular outcomes like inflammatory response and cytotoxicity in response to ZnO NPs treatment. More importantly, ZnO NPs can induce different modes of cell death in 2D and 3D cell culture systems. Interestingly, the outer few layers of cells in 3D model could only protect the inner core of cells for a limited time and periodically slough off from the spheroids surface. These findings suggest that toxicological conclusions made from 2D cell models might overestimate the toxicity of ZnO NPs. This 3D cell spheroid model can serve as a reproducible platform to better reflect the actual cell response to NPs and to study a more realistic mechanism of nanoparticle-induced toxicity.

Toxicity profiling of water contextual zinc oxide, silver, and titanium dioxide nanoparticles in human oral and gastrointestinal cell systems.

Engineered nanoparticles (ENPs) are increasingly detected in water supply due to environmental release of ENPs as the by-products contained within the effluent of domestic and industrial run-off. The partial recycling of water laden with ENPs, albeit at ultra-low concentrations, may pose an uncharacterized threat to human health. In this study, we investigated the toxicity of three prevalent ENPs: zinc oxide, silver, and titanium dioxide over a wide range of concentrations that encompasses drinking water-relevant concentrations, to cellular systems representing oral and gastrointestinal tissues. Based on published in silico-predicted water-relevant ENPs concentration range from 100 pg/L to 100 µg/L, we detected no cytotoxicity to all the cellular systems. Significant cytotoxicity due to the NPs set in around 100 mg/L with decreasing extent of toxicity from zinc oxide to silver to titanium dioxide NPs. We also found that noncytotoxic zinc oxide NPs level of 10 mg/L could elevate the intracellular oxidative stress. The threshold concentrations of NPs that induced cytotoxic effect are at least two to five orders of magnitude higher than the permissible concentrations of the respective metals and metal oxides in drinking water. Based on these findings, the current estimated levels of NPs in potable water pose little cytotoxic threat to the human oral and gastrointestinal systems within our experimental boundaries.

Nanoparticles strengthen intracellular tension and retard cellular migration.

Nanoparticles can have profound effects on cell biology. Here, we show that after TiO2, SiO2, and hydroxyapatite nanoparticles treatment, TR146 epithelial cell sheet displayed slower migration. Cells after exposure to the nanoparticles showed increased cell contractility with significantly impaired wound healing capability however without any apparent cytotoxicity. We showed the mechanism is through nanoparticle-mediated massive disruption of the intracellular microtubule assembly, thereby triggering a positive feedback that promoted stronger substrate adhesions thus leading to limited cell motility.

Waste-to-resource: Extraction and transformation of aquatic biomaterials for regenerative medicine.

The fisheries and aquaculture industry are known for generating substantial waste or by-products, often underutilized, or relegated to low-value purposes. However, this overlooked segment harbors a rich repository of valuable bioactive materials of which have a broad-spectrum of high-value applications. As the blue economy gains momentum and fisheries expand, sustainable exploitation of these aquatic resources is increasingly prioritized. In this review, we present a comprehensive overview of technology-enabled methods for extracting and transforming aquatic waste into valuable biomaterials and their recent advances in regenerative medicine applications, focusing on marine collagen, chitin/chitosan, calcium phosphate and bioactive-peptides. We discuss the inherent bioactive qualities of these "waste-to-resource" aquatic biomaterials and identify opportunities for their use in regenerative medicine to advance healthcare while achieving the Sustainable Development Goals.

Label-Free Single Microparticles and Cell Aggregates Sorting in Continuous Cell-Based Manufacturing.

There is a paradigm shift in biomanufacturing toward continuous bioprocessing but cell-based manufacturing using adherent and suspension cultures, including microcarriers, hydrogel microparticles, and 3D cell aggregates, remains challenging due to the lack of efficient in-line bioprocess monitoring and cell harvesting tools. Herein, a novel label-free microfluidic platform for high throughput (≈50 particles/sec) impedance bioanalysis of biomass, cell viability, and stem cell differentiation at single particle resolution is reported. The device is integrated with a real-time piezo-actuated particle sorter based on user-defined multi-frequency impedance signatures. Biomass profiling of Cytodex-3 microcarriers seeded with adipose-derived mesenchymal stem cells (ADSCs) is first performed to sort well-seeded or confluent microcarriers for downstream culture or harvesting, respectively. Next, impedance-based isolation of microcarriers with osteogenic differentiated ADSCs is demonstrated, which is validated with a twofold increase of calcium content in sorted ADSCs. Impedance profiling of heterogenous ADSCs-encapsulated hydrogel (alginate) microparticles and 3D ADSC aggregate mixtures is also performed to sort particles with high biomass and cell viability to improve cell quality. Overall, the scalable microfluidic platform technology enables in-line sample processing from bioreactors directly and automated analysis of cell quality attributes to maximize cell yield and improve the control of cell quality in continuous cell-based manufacturing.

Attenuating Epithelial-to-Mesenchymal Transition in Cancer through Angiopoietin-Like 4 Inhibition in a 3D Tumor Microenvironment Model.

Epithelial-to-mesenchymal transition (EMT) plays a crucial role in metastatic cancer progression, and current research, which relies heavily on 2D monolayer cultures, falls short in recapitulating the complexity of a 3D tumor microenvironment. To address this limitation, a transcriptomic meta-analysis is conducted on diverse cancer types undergoing EMT in 2D and 3D cultures. It is found that mechanotransduction is elevated in 3D cultures and is further intensified during EMT, but not during 2D EMT. This analysis reveals a distinct 3D EMT gene signature, characterized by extracellular matrix remodeling coordinated by angiopoietin-like 4 (Angptl4) along with other canonical EMT regulators. Utilizing hydrogel-based 3D matrices with adjustable mechanical forces, 3D cancer cultures are established at varying physiological stiffness levels. A YAP:EGR-1 mediated up-regulation of Angptl4 expression is observed, accompanied by an upregulation of mesenchymal markers, at higher stiffness during cancer EMT. Suppression of Angptl4 using antisense oligonucleotides or anti-cAngptl4 antibodies leads to a dose-dependent abolishment of EMT-mediated chemoresistance and tumor self-organization in 3D, ultimately resulting in diminished metastatic potential and stunted growth of tumor xenografts. This unique programmable 3D cancer cultures simulate stiffness levels in the tumor microenvironment and unveil Angptl4 as a promising therapeutic target to inhibit EMT and impede cancer progression.