Mapping enzyme activity in living systems by real-time mid-infrared photothermal imaging of nitrile chameleons.

Simultaneous spatial mapping of the activity of multiple enzymes in a living system can elucidate their functions in health and disease. However, methods based on monitoring fluorescent substrates are limited. Here, we report the development of nitrile (C≡N)-tagged enzyme activity reporters, named nitrile chameleons, for the peak shift between substrate and product. To image these reporters in real time, we developed a laser-scanning mid-infrared photothermal imaging system capable of imaging the enzymatic substrates and products at a resolution of 300 nm. We show that when combined, these tools can map the activity distribution of different enzymes and measure their relative catalytic efficiency in living systems such as cancer cells, Caenorhabditis elegans, and brain tissues, and can be used to directly visualize caspase-phosphatase interactions during apoptosis. Our method is generally applicable to a broad category of enzymes and will enable new analyses of enzymes in their native context.

Droplet Transportation on Liquid-Infused Asymmetrically Structured Surfaces by Mechanical Oscillation and Viscosity Control.

The transportation of droplets on solid surfaces has received significant attention owing to its importance in biochemical analysis and microfluidics. In this study, we propose a novel strategy for controlling droplet motion by combining an asymmetric structure and infused lubricating oil on a vibrating substrate. The transportation of droplets with volumes ranging from 10 to 90 µL was realized, and the movement speed could be adjusted from 1.45 to 10.87 mm/s. Typical droplet manipulations, including droplet transportation along a long trajectory and selective movement of multiple droplets, were successfully demonstrated. Through experimental exploration and theoretical analysis, we showed that the adjustment of droplet transport velocity involves an intricate interaction among the Ohnesorge number, droplet volume, and input amplitude. It can potentially be used for the more complex manipulation of liquid droplets in microfluidic and biochemical analysis systems.

High acoustic numerical aperture photoacoustic microscopy with improved sensitivity.

Limited by the numerical aperture of ultrasonic detection, optical resolution photoacoustic microscopy (OR-PAM) has not achieved optimal sensitivity. To address this problem, we have developed a high acoustic numerical aperture ($ {\sim} 0.74 $∼0.74) OR-PAM (HNA-OR-PAM). Via engineering the acoustic lens, we implement the highest acoustic numerical aperture that a spherical concave lens can achieve. The sensitivity of HNA-OR-PAM is improved to around 160%-the state-of-the-art OR-PAM. Without averaging, the new system can image oxygen saturation in vivo with only 10-nJ pulse energy. The improved sensitivity allows us to image weaker absorbers, penetrate deeper, and reduce nonlinear effects induced by high pulse energy. Moreover, the photoacoustic view angle is augmented to 51.8 deg and makes tilted features more visible. We validate the improved view angle in both a phantom study and brain imaging.

Theoretical and Experimental Study of Heterodyne Phase-Sensitive Dispersion Spectroscopy with an Injection-Current-Modulated Quantum Cascade Laser.

We report the theoretical and experimental study of calibration-free heterodyne phase-sensitive dispersion spectroscopy (HPSDS) in the mid-infrared using a direct current modulated mid-infrared quantum cascade laser (QCL). The modulation of QCL current at several hundred MHz or higher generates the synchronous frequency and intensity modulation of the QCL emission. An analytical model of the phase of the beat note signal in HPSDS is derived by considering the absorption and dispersion processes and incorporating the QCL modulation parameters. In the experiment, a 4.5 µm QCL modulated at 350 MHz was used to measure N2O at 200 Torr in a 10 cm gas cell. The N2O concentrations inferred from the analytical model were compared with the nominal values to show good agreement over the concentration range of 189-805 ppm with a standard deviation <3%. When the QCL wavelength was locked at the line-center of the molecular transition, it was of interest to find that the theoretical model was simplified to that used for near-infrared HPSDS with an electro-optical modulator for laser modulation.

Comparative genomic analysis for nucleotide, codon, and amino acid usage patterns of mycoplasmas.

The evolutionary factors in influencing the genetic characteristics of nucleotide, synonymous codon, and amino acid usage of 18 mycoplasma species were analyzed. The nucleotide usage at the 1st and 2nd codon position which determines amino acid composition of proteins has a significant correlation with the total nucleotide composition of gene population of these mycoplasma species, however, the nucleotide usage at the 3rd codon position which affects synonymous codon usage patterns has a slight correlation with either the total nucleotide composition or the nucleotide usage at the 1st and 2nd codon position. Other evolutionary factors join in the evolutionary process of mycoplasma apart from mutation pressure caused by nucleotide usage constraint based on the relationships between effective number of codons/codon adaptation index and nucleotide usage at the 3rd codon position. Although nucleotide usage of gene population in mycoplasma dominates in forming the overall codon usage trends, the relative abundance of codon with nucleotide context and amino acid usage pattern show that translation selection involved in translation accuracy and efficiency play an important role in synonymous codon usage patterns. In addition, synonymous codon usage patterns of gene population have a bigger power to represent genetic diversity among different species than amino acid usage. These results suggest that although the mycoplasmas reduce its genome size during the evolutionary process and shape the form, which is opposite to their hosts, of AT usages at high levels, this kind organism still depends on nucleotide usage at the 1st and 2nd codon positions to control syntheses of the requested proteins for surviving in their hosts and nucleotide usage at the 3rd codon position to develop genetic diversity of different mycoplasma species. This systemic analysis with 18 mycoplasma species may provide useful clues for further in vivo genetic studies on the related species.

National serosurvey of encephalomyocarditis virus in healthy people and pigs in China.

Encephalomyocarditis virus (EMCV) is a zoonotic pathogen that has a wide spectrum of host range. The virus has been discovered on swine farms worldwide and can cause acute fatal myocarditis in piglets and reproductive disorders in sows. Although EMCV infection has been documented in farmed pigs in China, seroprevalence in humans has not been reported. In this study, we conducted nationwide serological surveys for EMCV in humans and farmed pigs in China in 2013, by the use of a double antigen sandwich ELISA method. A total of 3305 serum samples from healthy people were obtained from seven geographical regions in China, of which 1010 samples (30.56%) were positive for EMCV antibodies. The overall seroprevalence for EMCV in the age groups of 0-20, 21-40, 41-60 and >60 years were 13.5%, 30.25%, 36.83% and 38.71% respectively, showing a tendency of increasing with age (P = 0.000). A total of 3470 serum samples from farmed pigs were collected and tested for antibodies to EMCV. A high seroprevalence of 77% was recorded, and significant regional differences were observed. It was concluded that people and pigs in China were commonly infected by EMCV. In addition, in order to characterize changes of seroprevalence during natural EMCV infection in pigs, 240 serial serum samples were collected from 30 pigs (at 0, 15, 30, 60, 75, 90, 120, and 150 days of age) in a farrow-to-finish farm in China. The data showed that there were two EMCV antibody peaks: the first peak appeared at day 30, followed by a decrease in EMCV antibody titer, and the second occurred after day 75. Thus, the most susceptible period of pigs for EMCV infection was between day 30 and day 75 of age.

[Prevalence of blindness and moderate and severe visual impairment among adults aged 50 years or above in Shuangcheng City of Heilongjiang Province: the China Nine-Province Survey].

OBJECTIVE: To investigate the prevalence of blindness and moderate and severe visual impairment among adults aged 50 years or above in Shuangcheng City of Heilongjiang Province, China. METHODS: It was a population-based cross-section study. Geographically defined cluster sampling was used in randomly selecting 5 841 individuals aged 50 years or above in 28 basic sample units in Shuangcheng City from September to December 2006. The survey was preceded by a pilot study where operational methods were refined and quality assurance evaluation was carried out. All participants were enumerated through village registers followed door-to-door visits.Eligible individuals were invited to receive visual acuity measurement and eye examination. Statistical analyses were performed using Stata/SE Statistical Software, release 9.0. Chi-square test was used to investigate the association of age, gender and education with presenting and best corrected visual acuity. RESULTS: Five thousands four hundreds and eighty-one individuals were enumerated and 5 047 persons were examined, the response rate was 92.08%. Based on the criteria of World Health Organization visual impairment classification in 1973, the prevalence of blindness and moderate and severe visual impairment defined as best corrected visual acuity was 1.72% (87/5 047) and 4.69% (237/5 047) respectively. The prevalence of blindness and moderate and severe visual impairment defined as presenting visual acuity was 1.90% (96/5 047) and 7.23% (365/5 047) respectively. The prevalence of blindness and moderate and severe visual impairment was higher in aged (trend χ(2)=674.44, P < 0.01), female (χ(2)=39.61, P < 0.01) and illiterate (trend χ(2)=142.82, P < 0.01) persons. Cataract (52.70%) was still the first leading cause of blindness and visual impairment. The percentage of the retinal diseases, including age-related macular degeneration, high myopic retinopathy and diabetic retinopathy was 11.31% among the eyes with blindness and moderate and severe visual impairment. Un-corrected refractive error(13.38%) also was the main cause of visual impairment. CONCLUSIONS: The prevalence of blindness and moderate and severe visual impairment in Shuangcheng City is relatively higher in China Nine Province Survey. Cataract, retinal diseases and un-corrected refractive error are the main causes of the blindness and moderate and severe visual impairment.

Controlled Growth Lateral/Vertical Heterostructure Interface for Lithium Storage.

Artificial heterostructures with structural advancements and customizable electronic interfaces are fundamental for achieving high-performance lithium-ion batteries (LIBs). Here, a design idea for a covalently bonded lateral/vertical black phosphorus (BP)-graphdiyne oxide (GDYO) heterostructure achieved through a facile ball-milling approach, is designed. Lateral heterogeneity is realized by the sp2-hybridized mode P-C bonds, which connect the phosphorus atoms at the edges of BP with the carbon atoms of the terminal acetylene in GDYO. The vertical connection of the heterojunction of BP and GDYO is connected by P-O-C bond. Experimental and theoretical studies demonstrate that BP-GDYO incorporates interfacial and structural engineering features, including built-in electric fields, chemical bond interactions, and maximized nanospace confinement effects. Therefore, BP-GDYO exhibits improved electrochemical kinetics and enhanced structural stability. Moreover, through ex- and in-situ studies, the lithiation mechanism of BP-GDYO, highlighting that the introduction of GDYO inhibits the shuttle/dissolution effect of phosphorus intermediates, hinders volume expansion, provides more reactive sites, and ultimately promotes reversible lithium storage, is clarified. The BP-GDYO anode exhibits lithium storage performance with high-rate capacity and long-cycle stability (602.6 mAh g-1 after 1 000 cycles at 2.0 A g-1). The proposed interfacial and structural engineering is universal and represents a conceptual advance in building high-performance LIBs electrode.

Synthesis of P-Containing Polycyclic Aromatic Hydrocarbons from Alkynyl-phosphonium Salts.

This work presents a straightforward method for synthesizing a series of phosphorus-containing polycyclic aromatic hydrocarbons (P-PAHs) featuring an internal ylidic bond. The method involves anion exchange, alkyne annulation, and deprotonation reactions, enabling the efficient production of cyclic phosphonium salts, which serve as pivotal intermediates in the synthesis of P-PAHs. The alkyne annulation reaction exhibits high regioselectivity, ensuring the successful synthesis of λ5-phosphaphenanthrene isomers. Additionally, the incorporation of electron-withdrawing groups effectively stabilizes the internal ylidic bond of P-PAHs.

Mid-Infrared Photoacoustic Stimulation of Neurons through Vibrational Excitation in Polydimethylsiloxane.

Photoacoustic (PA) emitters are emerging ultrasound sources offering high spatial resolution and ease of miniaturization. Thus far, PA emitters rely on electronic transitions of absorbers embedded in an expansion matrix such as polydimethylsiloxane (PDMS). Here, it is shown that mid-infrared vibrational excitation of C─H bonds in a transparent PDMS film can lead to efficient mid-infrared photoacoustic conversion (MIPA). MIPA shows 37.5 times more efficient than the commonly used PA emitters based on carbon nanotubes embedded in PDMS. Successful neural stimulation through MIPA both in a wide field with a size up to a 100 µm radius and in single-cell precision is achieved. Owing to the low heat conductivity of PDMS, less than a 0.5 °C temperature increase is found on the surface of a PDMS film during successful neural stimulation, suggesting a non-thermal mechanism. MIPA emitters allow repetitive wide-field neural stimulation, opening up opportunities for high-throughput screening of mechano-sensitive ion channels and regulators.

Vote2Cap-DETR++: Decoupling Localization and Describing for End-to-End 3D Dense Captioning.

3D dense captioning requires a model to translate its understanding of an input 3D scene into several captions associated with different object regions. Existing methods adopt a sophisticated "detect-then-describe" pipeline, which builds explicit relation modules upon a 3D detector with numerous hand-crafted components. While these methods have achieved initial success, the cascade pipeline tends to accumulate errors because of duplicated and inaccurate box estimations and messy 3D scenes. In this paper, we first propose Vote2Cap-DETR, a simple-yet-effective transformer framework that decouples the decoding process of caption generation and object localization through parallel decoding. Moreover, we argue that object localization and description generation require different levels of scene understanding, which could be challenging for a shared set of queries to capture. To this end, we propose an advanced version, Vote2Cap-DETR++, which decouples the queries into localization and caption queries to capture task-specific features. Additionally, we introduce the iterative spatial refinement strategy to vote queries for faster convergence and better localization performance. We also insert additional spatial information to the caption head for more accurate descriptions. Without bells and whistles, extensive experiments on two commonly used datasets, ScanRefer and Nr3D, demonstrate Vote2Cap-DETR and Vote2Cap-DETR++ surpass conventional "detect-then-describe" methods by a large margin. We have made the code available at https://github.com/ch3cook-fdu/Vote2Cap-DETR.

Overtone photothermal microscopy for high-resolution and high-sensitivity vibrational imaging.

Photothermal microscopy is a highly sensitive pump-probe method for mapping nanostructures and molecules through the detection of local thermal gradients. While visible photothermal microscopy and mid-infrared photothermal microscopy techniques have been developed, they possess inherent limitations. These techniques either lack chemical specificity or encounter significant light attenuation caused by water absorption. Here, we present an overtone photothermal (OPT) microscopy technique that offers high chemical specificity, detection sensitivity, and spatial resolution by employing a visible probe for local heat detection in the C-H overtone region. We demonstrate its capability for high-fidelity chemical imaging of polymer nanostructures, depth-resolved intracellular chemical mapping of cancer cells, and imaging of multicellular C. elegans organisms and highly scattering brain tissues. By bridging the gap between visible and mid-infrared photothermal microscopy, OPT establishes a new modality for high-resolution and high-sensitivity chemical imaging. This advancement complements large-scale shortwave infrared imaging approaches, facilitating multiscale structural and chemical investigations of materials and biological metabolism.

Click-free imaging of carbohydrate trafficking in live cells using an azido photothermal probe.

Real-time tracking of intracellular carbohydrates remains challenging. While click chemistry allows bio-orthogonal tagging with fluorescent probes, the reaction permanently alters the target molecule and only allows a single snapshot. Here, we demonstrate click-free mid-infrared photothermal (MIP) imaging of azide-tagged carbohydrates in live cells. Leveraging the micromolar detection sensitivity for 6-azido-trehalose (TreAz) and the 300-nm spatial resolution of MIP imaging, the trehalose recycling pathway in single mycobacteria, from cytoplasmic uptake to membrane localization, is directly visualized. A peak shift of azide in MIP spectrum further uncovers interactions between TreAz and intracellular protein. MIP mapping of unreacted azide after click reaction reveals click chemistry heterogeneity within a bacterium. Broader applications of azido photothermal probes to visualize the initial steps of the Leloir pathway in yeasts and the newly synthesized glycans in mammalian cells are demonstrated.

[Drug dynamics research of salvia miltiorrhiza in rabbit plasma and aqueous humor after intravenous injection].

OBJECTIVES: To assess the permeability of Danshensu at blood-ocular barrier and its characteristics of pharmacokinetics by respectively measuring the concentrations of Danshensu in blood plasma and aqueous humor of the rabbit with the high performance liquid chromatography(HPLC). METHODS: It was an experimental study. Seventy-two white rabbits were split into three groups: control group (6 rabbits), plasma group (6 rabbits) and aqueous humor group (60 rabbits). After 0.85% salt water (control group)or salvia miltiorrhiza (1.0 g/kg) (plasma and aqueous group) was injected into the vein of auris-edge. Samples of blood and aqueous humor were obtained for analysis . The analytical column was a BDS C18 stainless steed column(5 µm, 4.6 mm× 250 mm); Precolumn:YWGC18; the mobile phase consists of acetonitrile and 0.01 mol/L KH2PO4 (8:92, adjusted to pH = 2.8 with phosphoric acid). The UV detector was set at 279 nm; the flow rate was 1.0 ml/min;and the column temperature was ordinary temperature. RESULTS: The plasma concentration-time curves of Salvia miltiorrhiza fitted three-compartment model. t1/2 ß (elimination half time): 5.661 min; Cmax (peak concentration): 727.29 mg/L; Tmax(peak time ): 0 min. The aqueous humor concentration-time curves fitted two-compartment model. t1/2 ß: 147.663 min; Cmax: 38.62 mg/L; Tmax: 25 min. CONCLUSIONS: HPLC is a sensitive, specific and accurate method that can be used in pharmacokinetics research on ocular tissue of rabbit for Danshensu, Salvia miltiorrhiza that is dissolvable in water can pass through the blood-ocular barrier after intravenous injection with a relatively stable concentration of danshensu in aqueous humor resulting from a slow rate of removal.

A high-sensitivity high-resolution intravascular photoacoustic catheter through mode cleaning in a graded-index fiber.

Intravascular photoacoustic imaging has been developed to evaluate the possibility of plaque rupture in atherosclerosis by high spatial resolution imaging of lipid. However, the detection sensitivity and spatial resolution are compromised by the poor focusing caused by a multimode fiber. In this work, we report an intravascular photoacoustic catheter with mode self-cleaning in a graded-index fiber to improve the beam quality and the sensitivity for lipid detection. Compared with the higher-order modes in a step-index multimode fiber, the lower-order modes generated by the self-cleaning effect in the graded-index fiber greatly enhanced the photoacoustic spatial resolution and detection sensitivity. The dominant ringing artifact caused by laser absorption of the ultrasound transducer was further reduced by using stripe suppression. A lipid plaque mimicking phantom was imaged for evaluation. Lipid particles with a small diameter of 75.7 µm were clearly observed.

Ecotoxicological and biochemical effects of di(2-ethylhexyl)phthalate on wheat (Jimai 22, Triticum aestivum L.).

Di(2-ethylhexyl)phthalate esters (DEHP) has attracted widespread attention due to its ecotoxicological effects on organisms. In this study, wheat seedlings were exposed to DEHP- contaminated soil with 4 concentration gradients (0, 1, 10, and 100 mg kg-1, respectively) for 30 days. The growth index, physiological index, oxidative damage system, and gene expression of wheat seedlings were comprehensively measured and analyzed. The results revealed that DEHP could reduce the germination rate of wheat. Only the 100 mg kg-1 treatment group significantly inhibited root length, but no effect on plant height. At the biochemical level, photosynthetic pigments of wheat seedlings were promoted first and then inhibited, while the soluble sugar content presented a trend of "inhibition - activation - inhibition". The antioxidant enzymes (SOD and POD) presented an approximate parabolic trend, while it was opposite for CAT. Whereas the corresponding antioxidant enzyme genes were up-regulated, and the Hsp70 heat-shock protein gene was down-regulated. Finally, integrated biological response index (IBR) analysis showed that the DEHP toxicity to wheat seedlings was dose dependent. Molecular docking indicated that DEHP could stably bind to GBSS and GST by intermolecular force. Overall, this study provided constructive insights for a comprehensive assessment of the toxicity risk of DEHP to wheat.

Ecological risk assessment of environmentally relevant concentrations of propofol on zebrafish (Danio rerio) at early life stage: Insight into physiological, biochemical, and molecular aspects.

Propofol is an intravenous anesthetic injection extensively used in clinic, which has been proved to be neurotoxic in humans. Improper use and disposal of propofol may lead to its release into the aquatic environment, but the potential ecological risk of propofol to aquatic organisms remains poorly understood. For this study, we comprehensively explored the ecotoxicological effects and potential mechanisms of propofol (0.04, 0.2 and 2 mg L-1) on 120 hpf zebrafish (Danio rerio) embryos from physiological, biochemical, and molecular perspectives. The results showed that propofol has moderate toxicity on zebrafish embryos (96 h LC50 = 4.260 mg L-1), which could significantly reduce the hatchability and delay the development. Propofol can trigger reactive oxygen species (ROS) generation, lipid peroxidation (Malondialdehyde, MDA) and DNA damage (8-hydroxy-2-deoxyguanosine, 8-OHdG). The glutathione peroxidase (GPX) activity of zebrafish embryos in 0.04 and 0.2 mg L-1 propofol treatment group was activated in response to oxidative damage, while activities of superoxide dismutase (SOD), catalase (CAT) and GPX in zebrafish treated with 2 mg L-1 was significant inhibited compared with the control group (p＜0.05). Moreover, the expression of antioxidant genes and related pathways was inhibited. Apoptosis was investigated at genes level and histochemistry. Molecular docking confirmed that propofol could change in the secondary structure of acetylcholinesterase (AChE) and competitively inhibited acetylcholine (ACh) binding to AChE, which may disturb the nervous system. These results described toxic response and molecular mechanism in zebrafish embryos, providing multiple aspects about ecological risk assessment of propofol in water environment.

Evaluation of the biological response of propofol in zebrafish (Danio rerio): Focusing on biochemical, transcriptional, and molecular level.

Propofol, one of the most widely used intravenous anesthetic in clinical practice, has been reported to impair cognitive and memory function. However, the toxicological effects of propofol on aquatic organisms are still poorly understood. This study explored the toxic effects of chronic propofol exposure (0.008, 0.04, and 0.2 mg L-1) on adult zebrafish from biochemical, transcriptional, and molecular level after 7, 14, 21 and 28 days of exposure. Results indicated that the reactive oxygen species (ROS) levels were significantly upregulated during the 28 days exposure period, and excessive ROS caused lipid peroxidation, resulting in increased malondialdehyde (MDA) contents in the zebrafish brain. In order to relieve the oxidative damage induced by the excessive ROS, the activities of antioxidant enzymes (superoxide dismutase (SOD), catalase (CAT)) were significantly activated, and detoxification enzyme (glutathione S-transferase, GST) activities showed an "activation-inhibition" trend. However, the antioxidant enzymes and detoxification enzyme system could not eliminate the excessive ROS in time and thus caused DNA damage in zebrafish brain. The olive tail moment (OTM) values displayed a "dose-response" relationship with propofol concentrations. Meanwhile, the transcription of related genes of Nrf2-Keap1 pathway was activated. Further molecular simulation experiments suggested that propofol could directly combine with SOD/CAT to change the activity of its biological enzyme. These findings indicated that zebrafish could regulate antioxidant capacity to combat oxidative stress at the early exposure stage, but the activity of antioxidant enzymes were significantly inhibited with the increase of propofol exposure time. Our results are of great importance for understanding toxicological effects of propofol on aquatic organisms.

Mapping Enzyme Activity in Living Systems by Real-Time Mid-Infrared Photothermal Imaging of Nitrile Chameleons.

Enzymes are vital components in a variety of physiological and biochemical processes. Participation of various enzyme species are required for many biological events and signaling networks. Thus, spatially mapping the activity of multiple enzymes in a living system is significant for elucidating enzymatic functions in health and connections to diseases. Here, we report the development of nitrile (C≡N)-tagged enzyme activity reporters, named nitrile chameleons for the shifted peak between substrate and product. By real-time mid-infrared photothermal imaging of the enzymatic substrates and products at 300 nm resolution, our approach can map the activity distribution of different enzymes and quantitate the relative catalytic efficiency in living cancer cells, C. elegans, and brain tissues. An important finding is the direct visualization of caspase-phosphatase cooperation during apoptosis. Our method is generally applicable to a broad category of enzymes and will advance the discovery of potential targets for diagnosis and drug development.