Multiple energy X-ray imaging of metal oxide particles inside gingival tissues.

BACKGROUND: Periodontal disease affects over 50% of the global population and is characterized by gingivitis as the initial sign. One dental health issue that may contribute to the development of periodontal disease is foreign body gingivitis (FBG), which can result from exposure to some kinds of foreign metal particles from dental products or food. OBJECTIVE: We design a novel, portable, affordable, multispectral X-ray and fluorescence optical microscopic imaging system dedicated to detecting and differentiating metal oxide particles in dental pathological tissues. A novel denoising algorithm is applied. We verify the feasibility and optimize the performance of the imaging system with numerical simulations. METHODS: The designed imaging system has a focused X-ray tube with tunable energy spectra and thin scintillator coupled with an optical microscope as detector. A simulated soft tissue phantom is embedded with 2-micron thick metal oxide discs as the imaged object. GATE software is used to optimize the systematic parameters such as energy bandwidth and X-ray photon number. We have also applied a novel denoising method, Noise2Sim with a two-layer UNet structure, to improve the simulated image quality. RESULTS: The use of an X-ray source operating with an energy bandwidth of 5 keV, X-ray photon number of 108, and an X-ray detector with a 0.5 micrometer pixel size in a 100 by 100-pixel array allowed for the detection of particles as small as 0.5 micrometer. With the Noise2Sim algorithm, the CNR has improved substantially. A typical example is that the Aluminum (Al) target's CNR is improved from 6.78 to 9.72 for the case of 108 X-ray photons with the Chromium (Cr) source of 5 keV bandwidth. CONCLUSIONS: Different metal oxide particles were differentiated using Contrast-to-Noise ratio (CNR) by utilizing four different X-ray spectra.

A multi-depth spiral milli fluidic device for whole mount zebrafish antibody staining.

Whole mount zebrafish antibody staining (ABS) is a common staining technique used to localize protein information in a zebrafish embryo or larva. Like most biological assays, the whole mount zebrafish ABS is still largely conducted manually through labor intensive and time-consuming steps which affect both consistency and throughput of the assay. In this work, we develop a milli fluidic device that can automatically trap and immobilize the fixed chorion-less zebrafish embryos for the whole mount ABS. With just a single loading step, the zebrafish embryos can be trapped by the milli fluidic device through a chaotic hydrodynamic trapping process. Moreover, a consistent body orientation (i.e., head point inward) for the trapped zebrafish embryos can be achieved without additional orientation adjustment device. Furthermore, we employed a consumer-grade SLA 3D printer assisted method for device prototyping which is ideal for labs with limited budgets. Notably, the milli fluidic device has enabled the optimization and successful implementation of whole mount zebrafish Caspase-3 ABS. We demonstrated our device can accelerate the overall procedure by reducing at least 50% of washing time in the standard well-plate-based manual procedure. Also, the consistency is improved, and manual steps are reduced using the milli fluidic device. This work fills the gap in the milli fluidic application for whole mount zebrafish immunohistochemistry. We hope the device can be accepted by the zebrafish community and be used for other types of whole mount zebrafish ABS procedures or expanded to more complicated in situ hybridization (ISH) procedure.

Role of Polysaccharides in Diatom Thalassiosira pseudonana and its Associated Bacteria in Hydrocarbon Presence.

Diatoms secrete a significant amount of polysaccharides, which can serve as a critical organic carbon source for bacteria. The 2010 Deepwater Horizon oil spill exposed the Gulf of Mexico to substantial amounts of oil that also impacted the phytoplankton community. Increased production of exopolymeric substances was observed after this oil spill. Polysaccharides make up a major fraction of exopolymeric substances; however, their physiological role during an oil spill remains poorly understood. Here, we analyzed the role of polysaccharides in the growth and physiology of the oil-sensitive diatom Thalassiosira pseudonana and how they shape the surrounding bacterial community and its activity in the presence of oil. We found that inhibition of chrysolaminarin synthesis had a negative effect on the growth of T pseudonana and intracellular monosaccharide accumulation, which in turn suppressed photosynthesis by feedback inhibition. In addition, by acting as a carbon reserve, chrysolaminarin helped in the recovery of T pseudonana in the presence of oil. Inhibition of chrysolaminarin synthesis also influenced the bacterial community in the free-living fraction but not in the phycosphere. Exposure to oil alone led to increased abundance of oil-degrading bacterial genera and the activity of exoenzyme lipase. Our data show that chrysolaminarin synthesis plays an important role in the growth and survival of T pseudonana in the presence of oil, and its inhibition can influence the composition and activity of the surrounding bacterial community.

Nicotine alters mucin rheological properties.

Tobacco smoke exposure, the major cause of chronic obstructive pulmonary disease (COPD), instigates a dysfunctional clearance of thick obstructive mucus. However, the mechanism underlying the formation of abnormally viscous mucus remains elusive. We investigated whether nicotine can directly alter the rheological properties of mucin by examining its physicochemical interactions with human airway mucin gels secreted from A549 lung epithelial cells. Swelling kinetics and multiple particle tracking were utilized to assess mucin gel viscosity change when exposed to nicotine. Herein we show that nicotine (≤50 nM) significantly hindered postexocytotic swelling and hydration of released mucins, leading to higher viscosity, possibly by electrostatic and hydrophobic interactions. Moreover, the close association of nicotine and mucins allows airway mucus to function as a reservoir for prolonged nicotine release, leading to correlated pathogenic effects. Our results provide a novel explanation for the maltransport of poorly hydrated mucus in smokers. More importantly, this study further indicates that even low-concentration nicotine can profoundly increase mucus viscosity and thus highlights the health risks of secondhand smoke exposure.

Impact of microplastics and nanoplastics on liver health: Current understanding and future research directions.

With continuous population and economic growth in the 21st century, plastic pollution is a major global issue. However, the health concern of microplastics/ nanoplastics (MPs/NPs) decomposed from plastic wastes has drawn public attention only in the recent decade. This article summarizes recent works dedicated to understanding the impact of MPs/NPs on the liver-the largest digestive organ, which is one of the primary routes that MPs/NPs enter human bodies. The interrelated mechanisms including oxidative stress, hepatocyte energy re-distribution, cell death and autophagy, as well as immune responses and inflammation, were also featured. In addition, the disturbance of microbiome and gut-liver axis, and the association with clinical diseases such as metabolic dysfunction-associated fatty liver disease, steatohepatitis, liver fibrosis, and cirrhosis were briefly discussed. Finally, we discussed potential directions in regard to this trending topic, highlighted current challenges in research, and proposed possible solutions.

Determine the quality of human embryonic stem colonies with laser light scattering patterns.

BACKGROUND: With the prompt developments of regenerative medicine, the potential clinical applications of human embryonic stem cells have attracted intense attention. However, the labor-intensive and complex manual cell selection processes required during embryonic stem cell culturing have seriously limited large-scale production and broad applications. Thus, availability of a label-free, non-invasive platform to replace the current cumbersome manual selection has become a critical need. RESULTS: A non-invasive, label-free, and time-efficient optical platform for determining the quality of human embryonic stem cell colonies was developed by analyzing the scattering signals from those stem cell colonies. Additionally, confocal microscopy revealed that the cell colony morphology and surface structures were correlated with the resulting characteristic light scattering patterns. Standard immunostaining assay (Oct-4) was also utilized to validate the quality-determination from this light scattering protocol. The platform developed here can therefore provide identification accuracy of up to 87% for colony determination. CONCLUSIONS: Our study here demonstrated that light scattering patterns can serve as a feasible alternative approach to replace conventional manual selection for human embryonic stem cell cultures.

Size-Dependent Diffusion and Dispersion of Particles in Mucin.

Mucus, composed significantly of glycosylated mucins, is a soft and rheologically complex material that lines respiratory, reproductive, and gastrointestinal tracts in mammals. Mucus may present as a gel, as a highly viscous fluid, or as a viscoelastic fluid. Mucus acts as a barrier to the transport of harmful microbes and inhaled atmospheric pollutants to underlying cellular tissue. Studies on mucin gels have provided critical insights into the chemistry of the gels, their swelling kinetics, and the diffusion and permeability of molecular constituents such as water. The transport and dispersion of micron and sub-micron particles in mucin gels and solutions, however, differs from the motion of small molecules since the much larger tracers may interact with microstructure of the mucin network. Here, using brightfield and fluorescence microscopy, high-speed particle tracking, and passive microrheology, we study the thermally driven stochastic movement of 0.5-5.0 µm tracer particles in 10% mucin solutions at neutral pH, and in 10% mucin mixed with industrially relevant dust; specifically, unmodified limestone rock dust, modified limestone, and crystalline silica. Particle trajectories are used to calculate mean square displacements and the displacement probability distributions; these are then used to assess tracer diffusion and transport. Complex moduli are concomitantly extracted using established microrheology techniques. We find that under the conditions analyzed, the reconstituted mucin behaves as a weak viscoelastic fluid rather than as a viscoelastic gel. For small- to moderately sized tracers with a diameter of lessthan 2 µm, we find that effective diffusion coefficients follow the classical Stokes-Einstein relationship. Tracer diffusivity in dust-laden mucin is surprisingly larger than in bare mucin. Probability distributions of mean squared displacements suggest that heterogeneity, transient trapping, and electrostatic interactions impact dispersion and overall transport, especially for larger tracers. Our results motivate further exploration of physiochemical and rheological mechanisms mediating particle transport in mucin solutions and gels.

Interactions between silica and titanium nanoparticles and oral and gastrointestinal epithelia: Consequences for inflammatory diseases and cancer.

Engineered nanoparticles (NPs) composed of elements such as silica and titanium, smaller than 100 nm in diameter and their aggregates, are found in consumer products such as cosmetics, food, antimicrobials and drug delivery systems, and oral health products such as toothpaste and dental materials. They may also interact accidently with epithelial tissues in the intestines and oral cavity, where they can aggregate into larger particles and induce inflammation through pathways such as inflammasome activation. Persistent inflammation can lead to precancerous lesions. Both the particles and lesions are difficult to detect in biopsies, especially in clinical settings that screen large numbers of patients. As diagnosis of early stages of disease can be lifesaving, there is growing interest in better understanding interactions between NPs and epithelium and developing rapid imaging techniques that could detect foreign particles and markers of inflammation in epithelial tissues. NPs can be labelled with fluorescence or radioactive isotopes, but it is challenging to detect unlabeled NPs with conventional imaging techniques. Different current imaging techniques such as synchrotron radiation X-ray fluorescence spectroscopy are discussed here. Improvements in imaging techniques, coupled with the use of machine learning tools, are needed before diagnosis of particles in biopsies by automated imaging could move usefully into the clinic.

X-ray projection imaging of metal oxide particles inside gingival tissues.

There is increasing recognition that oral health affects overall health and systemic diseases. Nonetheless it remains challenging to rapidly screen patient biopsies for signs of inflammation or the pathogens or foreign materials that elicit the immune response. This is especially true in conditions such as foreign body gingivitis (FBG), where the foreign particles are often difficult to detect. Our long term goal is to establish a method to determine if the inflammation of the gingival tissue is due to the presence of a metal oxide, with emphasis on elements that were previously reported in FBG biopsies, such as silicon dioxide, silica, and titanium dioxide whose persistent presence can be carcinogenic. In this paper, we proposed to use multiple energy X-ray projection imaging to detect and to differentiate different metal oxide particles embedded inside gingival tissues. To simulate the performance of the imaging system, we have used GATE simulation software to mimic the proposed system and to obtain images with different systematic parameters. The simulated parameters include the X-ray tube anode metal, the X-ray spectra bandwidth, the X-ray focal spot size, the X-ray photon number, and the X-ray dector pixel. We have also applied the de-noising algorithm to obtain better Contrast-to-noise ratio (CNR). Our results indicate that it is feasible to detect metal particles as small as 0.5 micrometer in diameter when we use a Chromium anode target with an energy bandwidth of 5 keV, an X-ray photon number of 10^8, and an X-ray detector with a pixel size of 0.5 micrometer and 100 by 100 pixels. We have also found that different metal particles could be differentiated from the CNR at four different X-ray anodes and spectra. These encouraging initial results will guide our future imaging system design.

Aggregation, dissolution, and stability of quantum dots in marine environments: importance of extracellular polymeric substances.

There is an increasing concern that a considerable fraction of engineered nanoparticles (ENs), including quantum dots (QDs), will eventually find their way into the marine environment and have negative impacts on plankton. As ENs enter the ocean, they will encounter extracellular polymeric substances (EPS) from microbial sources before directly interacting with plankton cells. In this study, EPS harvested from four phytoplankton species, Amphora sp., Dunaliella tertiolecta, Phaeocystis globosa, and Thalassiosira pseudonana, were examined for potential interactions with CdSe nonfunctionalized and functionalized (carboxyl- and amine-) QDs in artificial seawater. Our results show that EPS do not reduce the solubility of QDs but rather decrease their stability. The degradation rate of QDs was positively correlated to the protein composition of EPS (defined by the ratio of protein/carbohydrate). Two approaches showed significant inhibition to the degradation of carboxyl-functionalized QDs: (1) the presence of an antioxidant, such as N-acetyl cysteine, and (2) absence of light. Owing to the complexity in evaluating integrated effects of QDs intrinsic properties and the external environmental factors that control the stability of QDs, conclusions must be based on a careful consideration of all these factors when attempting to evaluate the bioavailability of QDs and other ENs in the marine environments.

A mixture of anatase and rutile TiO2 nanoparticles induces histamine secretion in mast cells.

BACKGROUND: Histamine released from mast cells, through complex interactions involving the binding of IgE to FcεRI receptors and the subsequent intracellular Ca²âº signaling, can mediate many allergic/inflammatory responses. The possibility of titanium dioxide nanoparticles (TiO2 NPs), a nanomaterial pervasively used in nanotechnology and pharmaceutical industries, to directly induce histamine secretion without prior allergen sensitization has remained uncertain. RESULTS: TiO2 NP exposure increased both histamine secretion and cytosolic Ca²âº concentration ([Ca²âº]C) in a dose dependent manner in rat RBL-2H3 mast cells. The increase in intracellular Ca²âº levels resulted primarily from an extracellular Ca²âº influx via membrane L-type Ca²âº channels. Unspecific Ca²âº entry via TiO2 NP-instigated membrane disruption was demonstrated with the intracellular leakage of a fluorescent calcein dye. Oxidative stress induced by TiO2 NPs also contributed to cytosolic Ca²âº signaling. The PLC-IP3-IP3 receptor pathways and endoplasmic reticulum (ER) were responsible for the sustained elevation of [Ca²âº]C and histamine secretion. CONCLUSION: Our data suggests that systemic circulation of NPs may prompt histamine release at different locales causing abnormal inflammatory diseases. This study provides a novel mechanistic link between environmental TiO2 NP exposure and allergen-independent histamine release that can exacerbate manifestations of multiple allergic responses.

Temperature Effects on Effluent Microgel Formation.

Wastewater treatment plant effluent is considered an important hotspot of dissolved organic matter. The behavior and transformation of dissolved effluent organic matter (dEfOM) regulate particle sedimentation, pollutant fate, microbial attachment, and biofilm formation. However, studies have so far focused on the transformation of marine and riverine organic matter, and the current knowledge of dEfOM behavior is still limited. Fluctuations in water conditions, especially temperature, may directly alter the size, assembly speed, and structure of microgels, thereby potentially disturbing fate and the transportation of organic matter. In this study, we firstly investigated the effects of temperature on the behavior and capacity of dEfOM assembly into microgels and the possible mechanism. The microgel size and granularity of dEfOM were monitored by flow cytometry. Our results suggest that, with regard to microgels, a higher temperature leads to a higher assembly capacity but also a decrease in the size distribution. By contrast, assembly at 4 °C reduces the relative assembly capacity but increases the microgel size and granularity. The size distribution of the formed microgels at the various temperatures was ordered as follows: 4 °C > 20 °C > 35 °C. The size reduction in dEfOM assembly may be closely tied to the enhancement of hydrophobic interactions. The reduction in microgel granularity in warm conditions (35 °C) in terms of the effluent water may be caused by thermally induced condensation. Overall, the findings demonstrate the effects of temperature on dEfOM assembly and can facilitate further relevant studies on aquatic organic particle formation during current global warming scenarios.

Marine microplastics in the surface waters of "pristine" Kuroshio.

Microplastics (MPs) are ubiquitous in oceans. Their transboundary transport and fate have aroused global attention. Taiwan is located close to the western boundary current-Kuroshio, is an excellent location to study of MP mobility in the global current and Pacific Garbage Patch. This study is the first investigation to understand the microplastic contamination from Taiwan to the Kuroshio. MP concentrations in the area varied from N.D. to 0.15 items m-3, with an average concentration of 0.05 ± 0.03 items m-3. The majority of MPs were polypropylene, polyethylene, polyethylene and terephthalate. We found two MP hotspots near the coastal zone. One additional hotspot was also identified in the "pristine" Kuroshio suggesting rivers and local currents may play critical roles in transporting or injecting MPs from Taiwan into the North Pacific Gyre. These findings suggest that marine environments are altered by anthropogenic disposal and provide needed data for modelling and prediction of MPs.

Self-assembled Camptothecin derivatives - Curcuminoids conjugate for combinatorial chemo-photodynamic therapy to enhance anti-tumor efficacy.

Camptothecin (CPT), an alkaloid, was first discovered from plants and has potent anti-tumor activity. Since then, CPT analogs (namely Irinotecan and Topotecan) have been approved by the FDA for cancer treatments. Curcumin, on the other hand, is a widely used photosensitizer in photodynamic therapy (PDT) treatment. In our previous work, we have reported a straightforward strategy to construct a drug self-delivery system in which two-molecular species Irinotecan and Curcumin can self-assembly into a complex of ion pairs, namely ICN, through intermolecular non-covalent interactions. We found that ICN has slightly better chemotherapy efficacy than its individual components with much fewer side effects. In this paper, we aim to combine the chemotherapy and the PDT of ICN to further improve its anti-tumor performance. The efficient cellular uptake of ICNs was observed by confocal microscopy. Dichloro-dihydro-fluorescein diacetate (DCFH-DA) assay was used to detect the generation of singlet oxygen species. We found that the cell viability was 9% with both chemotherapy and PDT, and 31% with chemotherapy alone for the case with an ICN concentration of 10 µM, which demonstrated that the anti-tumor efficacy against the HT-29 cancer cell line was enhanced substantially with the combination therapy strategy. The study with an in vivo mouse model has further verified that the chemo-PDT dual therapy can inhibit tumor growth by 84% and 18.8% comparing with the control group and the chemotherapy group, respectively. Our results demonstrated that the new strategy using self-assembly and carrier-free nanoparticles with their chemo-PDT dual therapy may provide new opportunities to develop future combinatorial therapy methods in treating cancer.

Marine Gel Interactions with Hydrophilic and Hydrophobic Pollutants.

Microgels play critical roles in a variety of processes in the ocean, including element cycling, particle interactions, microbial ecology, food web dynamics, air-sea exchange, and pollutant distribution and transport. Exopolymeric substances (EPS) from various marine microbes are one of the major sources for marine microgels. Due to their amphiphilic nature, many types of pollutants, especially hydrophobic ones, have been found to preferentially associate with marine microgels. The interactions between pollutants and microgels can significantly impact the transport, sedimentation, distribution, and the ultimate fate of these pollutants in the ocean. This review on marine gels focuses on the discussion of the interactions between gel-forming EPS and pollutants, such as oil and other hydrophobic pollutants, nanoparticles, and metal ions.

From Nano-Gels to Marine Snow: A Synthesis of Gel Formation Processes and Modeling Efforts Involved with Particle Flux in the Ocean.

Marine gels (nano-, micro-, macro-) and marine snow play important roles in regulating global and basin-scale ocean biogeochemical cycling. Exopolymeric substances (EPS) including transparent exopolymer particles (TEP) that form from nano-gel precursors are abundant materials in the ocean, accounting for an estimated 700 Gt of carbon in seawater. This supports local microbial communities that play a critical role in the cycling of carbon and other macro- and micro-elements in the ocean. Recent studies have furthered our understanding of the formation and properties of these materials, but the relationship between the microbial polymers released into the ocean and marine snow remains unclear. Recent studies suggest developing a (relatively) simple model that is tractable and related to the available data will enable us to step forward into new research by following marine snow formation under different conditions. In this review, we synthesize the chemical and physical processes. We emphasize where these connections may lead to a predictive, mechanistic understanding of the role of gels in marine snow formation and the biogeochemical functioning of the ocean.

Stickiness of extracellular polymeric substances on different surfaces via magnetic tweezers.

Organic particle dynamics in the surface ocean plays a critical part in the marine carbon cycle. Aggregation of marine organic particles drives their downward transport to support various marine organisms on their transit to the sediments. Extracellular polymeric substances (EPS) from various microbes are a major contributor to the oceanic organic particle pool. The stickiness of EPS is expected to play a determining role in the aggregation process of particles; however, stickiness parameters are usually indirectly estimated through data fitting without direct assessment. Here a magnetic tweezer method was developed to quantitatively assess the stickiness of three model EPS produced by: Amphora sp., (diatom), Emiliania huxleyi (coccolithophore), and Sagittula stellata (bacteria), under different in vitro environmental conditions (salinity or EDTA complexed cations) and surface matrices (EPS-EPS and bare glass). Our results showed the stickiness of three microbial EPS decreasing for S. stellata > E. huxleyi > Amphora sp., in line with their decreasing protein-to-carbohydrate (P/C) ratios (related to their relative hydrophobicity). The data not only emphasize the importance of hydrophobicity on EPS stickiness, but also demonstrates that salinity and the nature of the substrate surface can influence the stickiness. Furthermore, we investigated stickiness between various types of EPS, and the observed selective stickiness of EPS between species may shed light on the interactions among heterogeneous marine microorganisms. Overall, this newly developed system provides a platform to assess the EPS stickiness to advance our understanding of the aggregation and sedimentation process of organic particles that are critical for the fate of organic carbon as well as for biofilm formation and microbial colonization of surfaces in the ocean.

A new role for bicarbonate in mucus formation.

The impact of small anions on the physical properties of gel-forming mucin has been almost overlooked relative to that of cations. Recently, based on the coincident abnormalities in HCO(3)(-) secretion and abnormal mucus formed in the hereditary disease cystic fibrosis (CF), HCO(3)(-) was hypothesized to be critical in the formation of normal mucus by virtue of its ability to sequester Ca(2+) from condensed mucins being discharged from cells. However, direct evidence of the impact of HCO(3)(-) on mucus properties is lacking. Herein, we demonstrate for the first time that mucin diffusivity (∼1/viscosity) increases as a function of [HCO(3)(-)]. Direct measurements of exocytosed mucin-swelling kinetics from airway cells showed that mucin diffusivity increases by ∼300% with 20 mM extracellular HCO(3)(-) concentration. Supporting data indicate that HCO(3)(-) reduces free Ca(2+) concentration and decreases the amount of Ca(2+) that remains associated with mucins. The results demonstrate that HCO(3)(-) enhances mucin swelling and hydration by reducing Ca(2+) cross-linking in mucins, thereby decreasing its viscosity and likely increasing its transportability. In addition, HCO(3)(-) can function as a Ca(2+) chelator like EGTA to disperse mucin aggregates. This study indicates that poor HCO(3)(-) availability in CF may explain why secreted mucus remains aggregated and more viscous in affected organs. These insights bear on not only the fundamental pathogenesis in CF, but also on the process of gel mucus formation and release in general.

Efficient Nonviral Stable Transgenesis Mediated by Retroviral Integrase.

Efficient transgene delivery is critical for genetic manipulation and therapeutic intervention of target cells. Two well-characterized integrative systems have been described that rely on viral and nonviral vectors. However, use of viral vectors for gene therapy has been associated with several safety concerns. Here, we report a virus-free method for stable transgenesis based on the reaction of retroviral integrase. We constructed a gateway cloning compatible vector containing two truncated long terminal repeat (LTR) sequences (dLTR) that flank the transgene cassette. Notably, 5'-ACTG-3' and blunt-end restriction cutting sites were also embedded at the end of dLTR to be recognized by HIV-1 integrase. When performing coinjection of transgene cassette and integrase mRNA into zebrafish embryos at one cell stage, there were 50% to 55% of injected embryos expressing a marker gene in a desired pattern. When applying our method in mammalian cells, there were 42% of cultured human epithelial cell lines showing stable integration. These results demonstrated that our method can successfully insert an exogenous gene into the host genome with highly efficient integration. Importantly, this system operates without most of the viral components while retaining effective stable transgenesis. We anticipate this method will provide a convenient, safe, and highly efficient way for applications in transgenesis and gene therapy.