Search

Biodegradable tissue engineering scaffolds have garnered increasing interest for their role in providing mechanical support, promoting tissue regeneration, and eliminating the need for removal. However, the in vivo degradation processes remain challenging to track. Here, a novel biodegradable polymer, N‐methyldiethanolamine (MDEA) and Gadolinium(III) diethylenetriamine pentaacetate (Gd‐DTPA) modified biodegradable photoluminescent polymers (BPLPMGd), which combines near‐infrared (NIR) fluorescence and magnetic resonance (MR) dual‐modality imaging are introduced to monitor scaffold degradation in vivo. The chemical structure of BPLPMGd is characterized and its dual‐imaging properties in vitro are evaluated. Subsequently, non‐invasive dual‐modality imaging to track the degradation of implanted BPLPMGd scaffolds is performed in a rat model, comparing these results with histological data. This approach reveals that BPLPMGd enables reliable non‐invasive tracking of the degradation, where NIR fluorescence imaging offers a qualitative and quantitative analysis of scaffold mass loss, total volume and solid content changes, while magnetic resonance imaging (MRI) details structural and morphological changes, allowing for 5D monitoring of implant degradation, including 3D structure, location, mass, volume, and geometry. The combination of these imaging modalities provides a comprehensive view of scaffold degradation, where the synergistic use of both yields results greater than either modality alone, offering unprecedented 5D information of implantable devices. This innovative approach has potential applications in regenerative engineering and beyond. [ABSTRACT FROM AUTHOR]

Advancing stable and biocompatible photothermal agents has emerged as a pivotal focus in the field of tumor photothermal treatment. Here, we synthesized two aza-boron-dipyrromethene (Aza-BODIPY) derivatives (NN-BP and TP-BP) with donor–acceptor–donor (D-A-D) structures, which exhibited robust near-infrared (NIR) absorption. Following encapsulation within the amphiphilic polymer (DSPEG-PEG2000), both NN-BP and TP-BP underwent self-assembly into nanoparticles (NN-BP NPs and TP-BP NPs), which displayed emission extended to near-infrared II regions (1000 nm). In vitro studies revealed that the two nanoparticles showed outstanding biocompatibility and a strong photothermal effect. These properties allowed the NPs to successfully destroy tumor cells under laser exposure. The TP-BP molecules featured typical AIE structure characteristics because of the triphenylamine group on both sides. After administration of TP-BP NP through the tail vein, NIR-II fluorescence imaging was realized in vivo with 4T1 tumor-bearing mice as a model. Compared with other treatment groups, the combination of TP-BP NPs and laser irradiation exhibited high tumor suppression efficiency without observable side effects. Altogether, TP-BP NPs achieved promising NIR-II imaging and local photothermal treatment of tumors, which opened an avenue for the development of phototheranostics. [ABSTRACT FROM AUTHOR]

Styrylpyridinium dye bearing azadithia‐15‐crown‐5 ether receptor group SP and 4‐alkoxy‐1,8‐naphthalimide fluorophore were linked using copper‐catalyzed azide‐alkyne cycloaddition click reaction to afford dyad compound NI‐SP. Chemosensor NI‐SP exhibited selective ratiometric fluorescent response to the presence of Hg2+ in aqueous solution due to the interplay between resonance energy transfer (RET) and intramolecular charge transfer (ICT) processes occurred upon excitation. The observed switching of the ratio of emission intensities in the blue and red channels R was higher than in the case of monochromophoric styrylpyridine derivative SP showing ratiometric response based on ICT mechanism only. Biological studies revealed that NI‐SP penetrates into human lung adenocarcinoma A549 cells and accumulates in cytoplasm and lysosomes. When cells were pre‐incubated with mercury (II) perchlorate, the ratio R was increased 2.6 times, which enables detection of intracellular Hg2+ ions and their quantitative analysis in the 0.7–6.0 μM concentration range. [ABSTRACT FROM AUTHOR]

Two‐photon excited fluorescence is a process in which the excited fluorophore relaxes and emits fluorescence after two‐photon absorption. Combined with appropriate two‐photon fluorescent probes, two‐photon microscopy enables observation deep inside tissue without damage. Former reports showed that the two‐photon absorption cross sections of donor–acceptor (D–A) dipoles, as well as D‐π‐D and D–A–D quadrupoles can be strengthened by using strong D–A groups. In this study, a novel water‐soluble fluorescent probe with D‐π‐A structure containing pyridine‐2,6‐dicarboxylic acid unit was synthesized. The maximum excitation wavelength and emission wavelength of the probe in aqueous solution were 328 nm and 471 nm, respectively. The fluorescence emitted by the probe can be quenched by various metal cations. The pyridyl N and carboxyl O in the probe formed chelates with Cu2+, which resulted in a fluorescence quenching response, and showed a linear relationship between fluorescent intensity and Cu2+ concentration with wide linear range and low limit of detection. CCK‐8 tests showed the probe had no obvious toxicity to 4T1 cells in the range of 0.4–50 μmol L−1, and the cell survival rate was above 86 %. The probe had a large two‐photon absorption cross section ranging from 680 nm to 820 nm. The two‐photon absorption cross section reached 100 GM at 760 nm of excitation wavelength. By means of microscopic imaging of 4T1 cells stained with the probe, one‐photon fluorescence represented blue emission, while two‐photon fluorescence represented bright green emission. The probe was proved to have good cell permeability, and preferred to target nucleus, showing application potential in two‐photon fluorescence imaging technology. [ABSTRACT FROM AUTHOR]

Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive and lethal malignancy with extremely poor patient survival rates. A key reason for the poor prognosis is the lack of effective diagnostic tools to detect the disease at curable, premetastatic stages. Tumor surgical resection is PDAC's first‐line treatment, however distinguishing between cancerous and healthy tissue with current imaging tools remains a challenge. In this work, we report a DOTA‐based fluorescent probe targeting plectin‐1 for imaging PDAC with high specificity. To enable heterogeneous functionalization of the DOTA‐core with multiple targeting peptide units and the fluorophore, a novel, fully clickable synthetic route that proceeds in one pot was developed. Extensive validation of the probe set the stage for PDAC detection in mice and human tissue. Altogether, these findings may pave the way for improved clinical understanding and early detection of PDAC progression as well as more accurate resection criteria. [ABSTRACT FROM AUTHOR]

A tumor-targeting fluorescent probe has attracted increasing interest in fluorescent imaging for the noninvasive detection of cancers in recent years. Sulfonamide-containing naphthalimide derivatives (SN-2NI, SD-NI) were synthesized by the incorporation of N-butyl-4-ethyldiamino-1,8-naphthalene imide (NI) into sulfonamide (SN) and sulfadiazine (SD) as the tumor-targeting groups, respectively. These derivatives were further characterized by mass spectrometry (MS), nuclear magnetic resonance spectroscopy (1H NMR), Fourier transform infrared spectroscopy (FT-IR), ultraviolet–visible spectroscopy (UV), and a fluorescence assay. In vitro properties, including cell cytotoxicity and the cell uptake of tumor cells, were also evaluated. Sulfonamide-containing naphthalimide derivatives possessed low cell cytotoxicity to B16F10 melanoma cells. Moreover, SN-2NI and SD-NI can be taken up highly by B16F10 cells and then achieve good green fluorescent images in B16F10 cells. Therefore, sulfonamide-containing naphthalimide derivatives can be considered to be the potential probes used to target fluorescent imaging in tumors. [ABSTRACT FROM AUTHOR]

[Display omitted] The development of bimodal imaging probes represents a hot topic of current research. Herein, we deal with developing an innovative bimodal contrast agent enabling fluorescence imaging (FI)/magnetic resonance imaging (MRI) and, simultaneously, consisting of biocompatible nanostructures. Optimized synthesis of advanced protein-embedded bimetallic (APEBM) nanocomposite containing luminescent gold nanoclusters (AuNC) and superparamagnetic iron oxide nanoparticles (SPION), suitable for in vivo dual-modal FI/MR imaging is reported. The APEBM nanocomposite was prepared by a specific sequential one-pot green synthetic approach that is optimized to increase metals (Au, Fe) content and, consequently, the imaging ability of the resulting nanostructures. The protein matrix, represented by serum albumin, was intentionally chosen, and used since it creates an efficient protein corona for both types of optically/magnetically-susceptible nanostructures (AuNC, SPION) and ensures biocompatibility of the resulting APEBM nanocomposite although it contains elevated metal concentrations (approx. 1 mg·mL −1 of Au, around 0.3 mg·mL −1 of Fe). In vitro and in vivo imaging was performed. Successful in vivo FI and MRI recorded in healthy mice corroborated the applicability of the APEBM nanocomposite and, simultaneously, served as a proof of concept concerning the potential future exploitation of this new FI/MRI bimodal contrast agent in preclinical and clinical practice. [ABSTRACT FROM AUTHOR]

Aristolochic acid I (AAI), a natural compound in aristolochia type Chinese medicinal herb, is generally acknowledged to have nephrotoxicity, which may be associated with mitophagy. Mitophagy is a cellular process with important functions that drive AAI-induced renal injury. Mitochondrial pH is currently measured by fluorescent probes in cell culture, but existing probes do not allow for in situ imaging of AAI-induced mitophagy in vivo. We developed a ratiometric fluorescent/PA dual-modal probe with a silicon rhodamine fluorophore and a pH-sensitive hemicyanine dye covalently linked via a short chain to obtain a FRET type probe. The probe was used to measure AAI-mediated mitochondrial acidification in live cells and in vivo. The Förster resonance energy transfer (FRET)-mediated ratiometric and bimodal method can efficiently eliminate signal variability associated with the commonly used one-emission and single detection mode by ratiometric two channels of the donor and acceptor. The probe has good water-solubility and low molecular weight with two positively charged, facilitating its precise targeting into renal mitochondria, where the fluorescent/PA changes in response to mitochondrial acidification, enabling dynamic and semi-quantitative mapping of subtle changes in mitochondrial pH in AAI-induced nephrotoxicity mouse model for the first time. Also, the joint use of L-carnitine could mitigate the mitophagy in AAI-induced nephrotoxicity.

[Display omitted] This work introduces non-covalent hydrophilic coating of NaGd 0.7 Eu 0.3 F 4 nanocrystals by polylysine (PL) and polyethyleneimine (PEI) for MRI and fluorescent imaging purposes. PL- and PEI-stabilized nanocrystals exhibit high colloidal stability, cell internalization, fluorescent contrasting of nuclei and cytoplasm, low cytotoxicity, and relaxivity (r 1 = 0.17 dm3 mmol–1 s–1 and r 1 = 0.23 dm3 mmol–1 s–1). [ABSTRACT FROM AUTHOR]

Abstract Carbon quantum dots (CQDs) have emerged as prominent contenders in the realm of luminescent nanomaterials over the past decade owing to their tunable optical properties, robust photostability, versatile surface functionalization and doping potential, low toxicity, and straightforward synthesis utilizing environmentally friendly precursors. In this review, we commence with a concise introduction, presenting both top‐down and bottom‐up strategies for the eco‐friendly synthesis of CQDs. Subsequently, we delve into a comprehensive examination of CQDs' structure and optical characteristics, encompassing their ultraviolet–visible absorption properties, surface confinement effects, and surface state emissions contributing to room‐temperature photoluminescence (PL). This review proceeds to elucidate recent advancements in modification strategies for CQDs, specifically focusing on surface oxidation, passivation, and the incorporation of heteroatoms. These strategies serve to afford control over the physicochemical properties, facilitating the enhancement of PL through the decoration of highly visible‐responsive CQDs. This enhancement is achieved by suppressing the nonradiative recombination of electron‐hole pairs, enabling red/blue shifts in CQDs for the generation of a full‐color emission spectrum, and regulating the band‐gap and surface states to broaden the photoabsorption range. Finally, we offer an overview of the most recent developments in the applications of fluorescent CQDs, emphasizing their utility in biomedicine, fluorescent sensors, lighting, and displays, as well as photocatalysis.

The bispidinone and bispidinol derivatives were synthesized using green synthetic methodologies and isolated in high yield without column chromatographic purification. The chemical structures of newly synthesized compounds were elucidated using spectroscopic techniques like IR, 1D & 2D NMR, and HRMS. The pivotal step, the reduction of the carbonyl group to alcohol, was developed by utilizing silica chloride and sodium borohydride. Their unique chemical structures offer versatility for functionalization, allowing for the introduction of specific moieties tailored to desired applications. Furthermore, the investigation into their anti-bacterial, anti-fungal, anti-oxidant, and cytotoxicity properties underscores their therapeutic potential. The compound 5n emerged as the best antimicrobial agent with low cytotoxicity. Subsequently, the green synthesis of bispidinones and bispidinols are valuable precursors to developing novel biomedical tools with diverse applications in imaging and cancer therapy, leveraging the unique properties of bispidine-derived compounds.

With the expansion of ICP-MS application into the field of bioanalysis, there is an urgent need for novel element tags today. Here, we report the design of a dual-element Ir-Eu tag, opening the door to simultaneous fluorescent imaging and ICP-MS quantification. The ratio of 153Eu/193Ir may serve as a precision control of the labeling process, allowing internal validation of the quantitative results obtained. As for SIRPα and its host cell analysis exemplified here, the Ir-Eu tag demonstrated superior figures of ICP-MS quantification with the LOD (3σ) down to 0.5 (153Eu) and 1.1 (193Ir) pM SIRPα and 220 (153Eu) and 830 (193Ir) RAW264.7 cells more than 130 times more sensitive compared with the LOD (3σ) of 65.2 pM SIRPα at 612 nm using fluorometry. Not limited to these demonstrations, we believe that the design ideas of the dual Ir-Eu tags should be applicable to various cases of bioanalysis when dual optical profiling and ICP-MS quantification are indispensable. [ABSTRACT FROM AUTHOR]

Abnormal viscosity is closely related to the occurrence of many diseases, such as cancer. Therefore, real-time detection of changes in viscosity in living cells is of great importance. Fluorescent molecular rotors play a critical role in detecting changes in cellular viscosity. Developing red emission viscosity probes with large Stokes shifts and high sensitivity and specificity remains an urgent and important topic. Herein, a novel viscosity-sensitive fluorescent probe (TCF-VIS1) with a large stokes shift and red emission was prepared based on the 2-dicyanomethylene-3-cyano-4,5,5-trimethyl-2,5-dihydrofuran (TCF) skeleton. Due to intramolecular rotation, the probe itself does not fluorescence at low viscosity. With the increase in viscosity, the rotation of TCF-VIS1 is limited, and its fluorescence is obviously enhanced. The probe has the advantages of simple preparation, large Stokes shift, good sensitivity and selectivity, and low cytotoxicity, which make it successfully used for viscosity detection in living cells. Moreover, TCF-VIS1 showed its potential for cancer diagnosis at the cell level and in tumor-bearing mice by detecting viscosity. Therefore, the probe is expected to enrich strategies for the detection of viscosity in biological systems and offer a potential tool for cancer diagnosis. [ABSTRACT FROM AUTHOR]

Copyright of Advances in Biochemistry / Postepy Biochemii is the property of Polish Biochemical Society / Acta Biochimica Polonica and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Organic fluorophores are indispensable tools in cells, tissue and in vivo imaging, and have enabled much progress in the wide range of biological and biomedical fields. However, many available dyes suffer from insufficient performances, such as short absorption and emission wavelength, low brightness, poor stability, small Stokes shift, and unsuitable permeability, restricting their application in advanced imaging technology and complex imaging. Over the past two decades, many efforts have been made to improve these performances of fluorophores. Starting with the luminescence principle of fluorophores, this review clarifies the mechanisms of the insufficient performance for traditional fluorophores to a certain extent, systematically summarizes the modified approaches of optimizing properties, highlights the typical applications of the improved fluorophores in imaging and sensing, and indicates existing problems and challenges in this area. This progress not only proves the significance of improving fluorophores properties, but also provide a theoretical guidance for the development of high‐performance fluorophores. [ABSTRACT FROM AUTHOR]

Carbon quantum dots (CQDs) have emerged as prominent contenders in the realm of luminescent nanomaterials over the past decade owing to their tunable optical properties, robust photostability, versatile surface functionalization and doping potential, low toxicity, and straightforward synthesis utilizing environmentally friendly precursors. In this review, we commence with a concise introduction, presenting both top‐down and bottom‐up strategies for the eco‐friendly synthesis of CQDs. Subsequently, we delve into a comprehensive examination of CQDs' structure and optical characteristics, encompassing their ultraviolet–visible absorption properties, surface confinement effects, and surface state emissions contributing to room‐temperature photoluminescence (PL). This review proceeds to elucidate recent advancements in modification strategies for CQDs, specifically focusing on surface oxidation, passivation, and the incorporation of heteroatoms. These strategies serve to afford control over the physicochemical properties, facilitating the enhancement of PL through the decoration of highly visible‐responsive CQDs. This enhancement is achieved by suppressing the nonradiative recombination of electron‐hole pairs, enabling red/blue shifts in CQDs for the generation of a full‐color emission spectrum, and regulating the band‐gap and surface states to broaden the photoabsorption range. Finally, we offer an overview of the most recent developments in the applications of fluorescent CQDs, emphasizing their utility in biomedicine, fluorescent sensors, lighting, and displays, as well as photocatalysis. [ABSTRACT FROM AUTHOR]

Abstract Background Transmembrane 4 L six family member 5 (TM4SF5) translocates subcellularly and functions metabolically, although it is unclear how intracellular TM4SF5 translocation is linked to metabolic contexts. It is thus of interests to understand how the traffic dynamics of TM4SF5 to subcellular endosomal membranes are correlated to regulatory roles of metabolisms. Methods Here, we explored the metabolic significance of TM4SF5 localization at mitochondria‐lysosome contact sites (MLCSs), using in vitro cells and in vivo animal systems, via approaches by immunofluorescence, proximity labelling based proteomics analysis, organelle reconstitution etc. Results Upon extracellular glucose repletion following depletion, TM4SF5 became enriched at MLCSs via an interaction between mitochondrial FK506‐binding protein 8 (FKBP8) and lysosomal TM4SF5. Proximity labeling showed molecular clustering of phospho‐dynamic‐related protein I (DRP1) and certain mitophagy receptors at TM4SF5‐enriched MLCSs, leading to mitochondrial fission and autophagy. TM4SF5 bound NPC intracellular cholesterol transporter 1 (NPC1) and free cholesterol, and mediated export of lysosomal cholesterol to mitochondria, leading to impaired oxidative phosphorylation but intact tricarboxylic acid (TCA) cycle and β‐oxidation. In mouse models, hepatocyte Tm4sf5 promoted mitophagy and cholesterol transport to mitochondria, both with positive relations to liver malignancy. Conclusions Our findings suggested that TM4SF5‐enriched MLCSs regulate glucose catabolism by facilitating cholesterol export for mitochondrial reprogramming, presumably while hepatocellular carcinogenesis, recapitulating aspects for hepatocellular carcinoma metabolism with mitochondrial reprogramming to support biomolecule synthesis in addition to glycolytic energetics.

To date, 14C tracer studies using accelerator mass spectrometry (AMS) have not yet resolved lipid-soluble analytes into individual lipoprotein density subclasses. The objective of this work was to develop a reliable method for lipoprotein separation and quantitative recovery for biokinetic modeling purposes. The novel method developed provides the means for use of small volumes (10–200 µL) of frozen plasma as a starting material for continuous isopycnic lipoprotein separation within a carbon- and pH-stable analyte matrix, which, following post-separation fraction clean up, created samples suitable for highly accurate 14C/12C isotope ratio determinations by AMS. Manual aspiration achieved 99.2 ± 0.41% recovery of [5-14CH3]-(2R, 4′R, 8′R)-α-tocopherol contained within 25 µL plasma recovered in triacylglycerol rich lipoproteins (TRL = Chylomicrons + VLDL), LDL, HDL, and infranatant (INF) from each of 10 different sampling times for one male and one female subject, n = 20 total samples. Small sample volumes of previously frozen plasma and high analyte recoveries make this an attractive method for AMS studies using newer, smaller footprint AMS equipment to develop genuine tracer analyses of lipophilic nutrients or compounds in all human age ranges. [ABSTRACT FROM AUTHOR]

The outcomes of unresectable gastric cancer (GC) are unfavorable even with chemotherapy; therefore, a new treatment modality is required. The combination of an oncolytic virus and photodynamic therapy can be one of the promising modalities to overcome this. Mammalian orthoreovirus (MRV) is an oncolytic virus that has been used in clinical trials for several cancers. In this study, we developed and evaluated a recombinant MRV strain type 3 Dearing (T3D) that expresses membrane-targeting KillerRed (KRmem), a phototoxic fluorescent protein that produces cytotoxic reactive oxygen species upon light irradiation. KRmem was fused in-frame to the 3′ end of the σ2 viral gene in the S2 segment using a 2A peptide linker, enabling the expression of multiple proteins from a single transcript. RNA electrophoresis, Western blotting, and immunofluorescence analyses confirmed functional insertion of KRmem into the recombinant virus. The growth activity of the recombinant virus was comparable to that of the wild-type MRV in a cultured cell line. The recombinant virus infected two GC cell lines (MKN45P and MKN7), and a significant cytocidal effect was observed in MKN45P cells infected with the recombinant virus after light irradiation. Thus, recombinant MRV-expressing KRmem has the potential to serve as a novel treatment tool for GC. [ABSTRACT FROM AUTHOR]

Chronic venous insufficiency with and without ulceration is a common complication of varicose veins. Heinz-Lippmann disease is a potentially underreported complication of chronic venous insufficiency that involves heterotopic ossification of the peri-wound that might contribute to the chronicity of venous leg ulcers. Most commonly, heterotopic ossification is associated with traumatic injury. We report the cases of three patients with chronic venous insufficiency and no history of trauma who were found to have subcutaneous calcifications consistent with Heinz-Lippmann disease. Osteomyelitis was confirmed in two patients. Physical examination and diagnostic imaging can be helpful in determining the diagnosis. Interventions can include biopsy to rule out osteomyelitis, with subsequent treatment, if needed.

Abstract Due to their inherent tunable spectrum, high brightness, excellent biostability and biocompatibility, and functionalization of surfaces, semiconducting polymer dots (Pdots) are now playing an essential role in fluorescent (FL) imaging and disease treatment through bioconjugation with peptides or biomimetic materials. In particular, biomimetic Pdots exhibit their capability in targeted imaging of lesion and increased efficacy for targeting disease treatment. This review will inspect the recent advances in the design and functionalization strategies of biomodified and biomimetic Pdots for enhanced disease detection and therapy. More importantly, the application of these two modifications in targeted FL imaging and cancer treatment is to be addressed in detail. Meanwhile, the main challenges and prospects of biomimetic and biomodified Pdots are to be discussed, which will pave a new avenue for improved disease detection and imaging‐guided treatment.

A tumor-targeting fluorescent probe has attracted increasing interest in fluorescent imaging for the noninvasive detection of cancers in recent years. Sulfonamide-containing naphthalimide derivatives (SN-2NI, SD-NI) were synthesized by the incorporation of N-butyl-4-ethyldiamino-1,8-naphthalene imide (NI) into sulfonamide (SN) and sulfadiazine (SD) as the tumor-targeting groups, respectively. These derivatives were further characterized by mass spectrometry (MS), nuclear magnetic resonance spectroscopy (1H NMR), Fourier transform infrared spectroscopy (FT-IR), ultraviolet–visible spectroscopy (UV), and a fluorescence assay. In vitro properties, including cell cytotoxicity and the cell uptake of tumor cells, were also evaluated. Sulfonamide-containing naphthalimide derivatives possessed low cell cytotoxicity to B16F10 melanoma cells. Moreover, SN-2NI and SD-NI can be taken up highly by B16F10 cells and then achieve good green fluorescent images in B16F10 cells. Therefore, sulfonamide-containing naphthalimide derivatives can be considered to be the potential probes used to target fluorescent imaging in tumors.

Heat shock proteins 70 kDa (HSP70) protect intracellular proteins from the damaging effects of stress factors of various natures. Moreover, HSP70 play an important role in the vital activity of cells under normal physiological conditions, performing chaperone functions. These functions are realized in the intracellular space; however, in some cases, these proteins are also found on the cell surface and in the extracellular environment. The causes and mechanisms of HSP70 translocation to the cell surface and secretion into the extracellular space have not yet been well understood, but such an unusual localization of HSP70 activates the immune system. The surface HSP70 and their extracellular pool stimulate the cytotoxic activity of NK cells. However, direct experimental evidence for the internalization of HSP70 molecules by NK cells has not yet been demonstrated. This paper presents the results of the interaction of the extracellular HSP70 pool with NK cells from the peripheral blood. The results demonstrated the confirmation of the internalization of exogenous HSP70 molecules by NK cells. To this end, fluorescently labeled recombinant stress-inducible human HSP70 were obtained. The electrophoretic data indicated the absence of protein degradation during the labeling process, the purity and stability of the modified protein. To assess the interaction of HSP70 with NK cells, the fluorescently labeled HSP70 was added to an in vitro culture of NK cells isolated by magnetic separation from the peripheral blood mononuclear fraction and analyzed by confocal microscopy. This analysis indicated that living NK cells internalize extracellular HSP70 with localization both in lysosomes and in phagosomes. Our experiments illustrated for the first time the process of penetration of the extracellular form of HSP70 into these cells. The results suggest that the activation of NK cells under the action of exogenous HSP70 could be associated with the internalization of these protein molecules.

Distinguishing the precise phases of cells in the cell cycle is important for elucidation of their regulatory mechanisms and exploiting cell‐cycle‐targeted drugs. Here, a simple but efficient strategy is established based on a peptide‐modified aggregation‐induced emission luminogen (AIEgen) probe PKK–TTP by utilizing its differential cellular uptake amounts for specific cell cycle recognition and targeted therapy. After incubating the probe in cells with different phases, maximal fluorescence and reactive oxygen species (ROS) generation are observed in S phase, middle in G0/G1 phase, and the minimum in G2/M phase. The cell‐cycle‐dependent fluorescent intensity is proven to be related to variable uptake routes of PKK–TTP. The maximized uptake of PKK–TTP in S phase relies on three simultaneously working endocytic routes, including macro‐pinocytosis and clathrin‐ and caveolin‐dependent endocytosis, while decreased uptake in the G0/G1 and G2/M phases mainly depends on the single route. Furthermore, the uptake of PKK–TTP is further verified in normal cells and drug‐resistant cancer cells (S). Bright fluorescence of PKK–TTP is observed in drug‐resistant cancer cells in S phase, while a little fluorescence is observed in paclitaxel (PTX)‐treated cancer cells due to G2/M phase. Collectively, PKK–TTP can be applied to distinguish cancer cells, evaluating the drug resistance and exerting its therapeutic efficiency downstream. [ABSTRACT FROM AUTHOR]

• Proteins of interest such as PTEN and αPD-L1 can be site-specifically labeled or conjugated using amber-suppression mediated genetic incorporation of unnatural amino acid (UAA). • Human PTEN intracellular activity can be tracked using UAA-based fluorescent imaging. • The in vivo biodistribution of the immune checkpoint protein PD-L1 can be mapped using UAA-based positron emission tomography (PET). This work features the use of amber suppression-mediated unnatural amino acid (UAA) incorporation into proteins for various imaging purposes. The site-specific incorporation of the UAA, p-azido-L-phenylalanine (pAzF), provides an azide handle that can be used to complete the strain promoted azide-alkyne click cycloaddition (SPAAC) reaction to introduce an imaging modality such as a fluorophore or a positron emission tomography (PET) tracer on the protein of interest (POI). Such methodology can be pursued directly in mammalian cell lines or on proteins expressed in vitro , thereby conferring a homogeneous pool of protein conjugates. A general procedure for UAA incorporation to use with a site-specific protein labeling method is provided allowing for in vitro and in vivo imaging applications based on the representative proteins PTEN and PD-L1. This approach would help elucidate the cellular or in vivo biological activities of the POI. [ABSTRACT FROM AUTHOR]

Polydopamine (PDA), the synthetic counterpart of melanin, is a widely investigated bio-inspired material for its chemical and photophysical properties, and in the last few years, bio-application of PDA and PDA-based materials have had a dramatic increase. In this review, we described PDA application in optical biosensing, exploring its multiple roles as a nanomaterial. In optical sensing, PDA can not only be used for its intrinsic fluorescent and photoacoustic properties as a probe: in some cases, a sample optical signal can be derived by melanin generation in situ or it can be enhanced in another material thanks to PDA modification. The various possibilities of PDA use coupled with its biocompatibility will indeed widen even more its application in optical bioimaging. [ABSTRACT FROM AUTHOR]

Monitoring the microenvironment within specific cellular regions is crucial for a comprehensive understanding of life events. Fluorescent probes working in different ranges of pH regions have been developed for the local imaging of different pH environments. Especially, rhodamine-based fluorescent pH probes have been of great interest due to their ON/OFF fluorescence depending on the spirolactam ring's opening/closure. By introducing the N-alkyl-hydroxamic acid instead of the alkyl amines in the spirolactam of rhodamine, we were able to tune the pH range where the ring opening and closing of the spirolactam occurs. This six-membered cyclic hydroxamate spirolactam ring of rhodamine B proved to be highly fluorescent in acidic pH environments. In addition, we could monitor pH changes of lysosomes in live cells and zebrafish. [ABSTRACT FROM AUTHOR]

Traditional molecular biology tools have elucidated the identities and functions of RNA molecules, which are essential to the understanding of gene transcription and protein translation. Deepening this research field would further require the direct visualization of RNA dynamics such as the DNA‐RNA interactions and RNA‐protein interactions. Towards this goal, the rise of RNA imaging tools over the past 15 years has reformed how we looked at these processes. In this Emerging Topic, we first highlighted recent advances on three main RNA imaging tools based on the species of interacting molecules: RNA‐RNA pairing, RNA‐protein binding, and small molecule‐RNA complex. We introduced the advantages of these tools from a technical viewpoint, including binding affinity, fluorescent turn‐on ratio, stability, and impacts on targeted RNA. Next, we discussed new rising opportunities and future directions, echoing the state‐of‐the‐art imaging tools in the fields of fluorescent proteins and small fluorescent molecules. Together, we believe this emerging field will bring new insights on how we study RNA biology in living systems. [ABSTRACT FROM AUTHOR]

Traditional molecular biology tools have elucidated the identities and functions of RNA molecules, which are essential to the understanding of gene transcription and protein translation. Deepening this research field would further require the direct visualization of RNA dynamics such as the DNA‐RNA interactions and RNA‐protein interactions. Towards this goal, the rise of RNA imaging tools over the past 15 years has reformed how we looked at these processes. In this Emerging Topic, we first highlighted recent advances on three main RNA imaging tools based on the species of interacting molecules: RNA‐RNA pairing, RNA‐protein binding, and small molecule‐RNA complex. We introduced the advantages of these tools from a technical viewpoint, including binding affinity, fluorescent turn‐on ratio, stability, and impacts on targeted RNA. Next, we discussed new rising opportunities and future directions, echoing the state‐of‐the‐art imaging tools in the fields of fluorescent proteins and small fluorescent molecules. Together, we believe this emerging field will bring new insights on how we study RNA biology in living systems. [ABSTRACT FROM AUTHOR]

Archaea are key players in many critical ecological processes. In comparison to eukaryotes and bacteria, however, our understanding of both the cell biology and diversity of archaea remains limited. While archaea inhabit a wide range of environmental conditions, many species are extremophiles, surviving in extreme temperature, salt or pH conditions, making their cell biology hard to study. Recently, our understanding of archaeal cell biology has been advanced significantly by the advent of live cell imaging in extremis as well as the development of genetic tools to exogenously express fluorescent proteins in some mesophilic archaeal model systems, e.g., Haloferax volcanii. However, for most archaeal species, especially thermophilic species or emerging model systems without well characterized genetic tools, live cell imaging remains dependent on fluorescent chemical probes to label and track the dynamics of living cells. While a wide range of fluorescent stains and markers that label different components of the cell are available commercially, their use has usually been optimized for use in a small number of eukaryotic cell systems. Here we report the successes and failures of the application of membrane, DNA, S-layer and cytoplasm markers in live cell imaging of archaea, as well as the optimization of fixation and immunolabelling approaches. We have applied these markers to the thermoacidophilic archaeon Sulfolobus acidocaldarius, but expect some to work in other archaeal species. Furthermore, those procedures that failed in S. acidocaldarius may still prove useful for imaging archaea that grow at a more neutral pH and/or at a less extreme temperature. [ABSTRACT FROM AUTHOR]

Abnormal viscosity is closely related to the occurrence of many diseases, such as cancer. Therefore, real-time detection of changes in viscosity in living cells is of great importance. Fluorescent molecular rotors play a critical role in detecting changes in cellular viscosity. Developing red emission viscosity probes with large Stokes shifts and high sensitivity and specificity remains an urgent and important topic. Herein, a novel viscosity-sensitive fluorescent probe (TCF-VIS1) with a large stokes shift and red emission was prepared based on the 2-dicyanomethylene-3-cyano-4,5,5-trimethyl-2,5-dihydrofuran (TCF) skeleton. Due to intramolecular rotation, the probe itself does not fluorescence at low viscosity. With the increase in viscosity, the rotation of TCF-VIS1 is limited, and its fluorescence is obviously enhanced. The probe has the advantages of simple preparation, large Stokes shift, good sensitivity and selectivity, and low cytotoxicity, which make it successfully used for viscosity detection in living cells. Moreover, TCF-VIS1 showed its potential for cancer diagnosis at the cell level and in tumor-bearing mice by detecting viscosity. Therefore, the probe is expected to enrich strategies for the detection of viscosity in biological systems and offer a potential tool for cancer diagnosis.

Archaea are key players in many critical ecological processes. In comparison to eukaryotes and bacteria, however, our understanding of both the cell biology and diversity of archaea remains limited. While archaea inhabit a wide range of environmental conditions, many species are extremophiles, surviving in extreme temperature, salt or pH conditions, making their cell biology hard to study. Recently, our understanding of archaeal cell biology has been advanced significantly by the advent of live cell imaging in extremis as well as the development of genetic tools to exogenously express fluorescent proteins in some mesophilic archaeal model systems, e.g., Haloferax volcanii. However, for most archaeal species, especially thermophilic species or emerging model systems without well characterized genetic tools, live cell imaging remains dependent on fluorescent chemical probes to label and track the dynamics of living cells. While a wide range of fluorescent stains and markers that label different components of the cell are available commercially, their use has usually been optimized for use in a small number of eukaryotic cell systems. Here we report the successes and failures of the application of membrane, DNA, S-layer and cytoplasm markers in live cell imaging of archaea, as well as the optimization of fixation and immunolabelling approaches. We have applied these markers to the thermoacidophilic archaeon Sulfolobus acidocaldarius, but expect some to work in other archaeal species. Furthermore, those procedures that failed in S. acidocaldarius may still prove useful for imaging archaea that grow at a more neutral pH and/or at a less extreme temperature.

Background: We performed a systematic review that identified at least 9,000 scientific papers on PubMed that include immunofluorescent images of cells from the central nervous system (CNS). These CNS papers contain tens of thousands of immunofluorescent neural images supporting the findings of over 50,000 associated researchers. While many existing reviews discuss different aspects of immunofluorescentmicroscopy, such as image acquisition and staining protocols, few papers discuss immunofluorescent imaging from an image-processing perspective. We analyzed the literature to determine the image processing methods that were commonly published alongside the associated CNS cell, microscopy technique, and animal model, and highlight gaps in image processing documentation and reporting in the CNS research field. Methods: We completed a comprehensive search of PubMed publications using Medical Subject Headings (MeSH) terms and other general search terms for CNS cells and common fluorescent microscopy techniques. Publications were found on PubMed using a combination of column description terms and row description terms. We manually tagged the comma-separated values file (CSV) metadata of each publication with the following categories: animal or cell model, quantified features, threshold techniques, segmentation techniques, and image processing software. Results: Of the almost 9,000 immunofluorescent imaging papers identified in our search, only 856 explicitly include image processing information. Moreover, hundreds of the 856 papers are missing thresholding, segmentation, and morphological feature details necessary for explainable, unbiased, and reproducible results. In our assessment of the literature, we visualized current image processing practices, compiled the image processing options from the top twelve software programs, and designed a road map to enhance image processing. We determined that thresholding and segmentation methods were often left out of publications and underreported or underutilized for quantifying CNS cell research. Discussion: Less than 10% of papers with immunofluorescent images include image processing in their methods. A few authors are implementing advanced methods in image analysis to quantify over 40 different CNS cell features, which can provide quantitative insights in CNS cell features that will advance CNS research. However, our review puts forward that image analysis methods will remain limited in rigor and reproducibility without more rigorous and detailed reporting of image processing methods. Conclusion: Image processing is a critical part of CNS research that must be improved to increase scientific insight, explainability, reproducibility, and rigor. [ABSTRACT FROM AUTHOR]

Effective and non-invasive inspection method toward viscosity is expected to ease the burden of continued increased health problems worldwide. Herein, we proposed that viscosity is one of the most important markers in the liquid microenvironment. Then, a natural product was further developed from resveratrol derivate, polydatin, as an activatable molecular sensor for monitoring liquid micro-environmental viscosity alterations for metamorphic extent determination. This natural molecular sensor was constructed by the single double bond alternating conjugate structure, which can rotate freely in the low viscosity media, and in the highly viscous media, bright fluorescent signals are specifically activated (around 383 nm, 53-fold), endowing the natural sensor with the capacity to avoid autofluorescence from the additives and achieving high signal-to-noise ratio imaging. With the lower detection limit 1.125 cP, the sensor polydatin exhibits higher viscosity sensitive coefficient (0.60), and 75.5 nm of Stokes shift was found in the distilled water. Afterward, polydatin achieved non-invasively identifying the thickening effects of food thickeners. In addition, the capability of polydatin to evaluate the deterioration extent of liquids with viscosity as a robust marker was explored. It was found that the viscosity level in microenvironments is highly dependent on the liquid food metamorphic period. The fluorescent intensity increment and viscosity value changes were fitted into a linear equation. We expected that this approach would reinvigorate the fluorescent analytical methods toward liquid safety inspection. [ABSTRACT FROM AUTHOR]

Near‐infrared fluorescence imaging is vital for exploring the biological world. The short emissions (<650 nm) and small Stokes shifts (<30 nm) of current xanthene dyes obstruct their biological applications since a long time. Recently, a potent and universal THQ structural modification technique that shifts emission to the NIR‐I/II range and enables a substantial Stokes shift (>100 nm) for THQ‐modified xanthene dyes is established. Thus, a timely discussion of THQ–xanthene and its applications is extensive. Hence, the advent, working principles, development trajectory, and biological applications of THQ–xanthene dyes, especially in the fields of fluorescence probe‐based sensing and imaging, cancer theranostics, and super‐resolution imaging, are introduced. It is envisioned that the THQ modification tactic is a simple yet exceptional approach to upgrade the performance of conventional xanthene dyes. THQ–xanthene will advance the strides of xanthene‐based potentials in early fluorescent diagnosis of diseases, cancer theranostics, and imaging‐guided surgery. [ABSTRACT FROM AUTHOR]

The hydrophobicity has been known as a pivotal factor limiting the therapeutic capacity of cancer drugs. Biocompatible nanocarriers as a potential platform for drug delivery can increase the solubility of hydrophobic drugs and prolong drug half-life, therefore improving therapeutic efficacies. Herein, we use the treated red blood cell membrane (RBM) as a nanocarrier to delivery hydrophobic carbon dot and paclitaxel (RBM-CD–PTX) for imaging-guided chemotherapy. The CD in RBM-CD with good biosafety endows them with excellent optical properties for fluorescent imaging from visible light to near-infrared light in cellular and animal models. The RBM-CD exhibited a high PTX loading capacity (loading capacity, 30.23%). The RBM-CD–PTX can be easily delivered and efficiently accumulated in tumor tissues, releasing PTX molecules to inhibit tumor growth. Such a nanoplatform manifests great potential to deliver clinical reagents for imaging-guided cancer therapy. [ABSTRACT FROM AUTHOR]

Developing effective intelligent nanocarriers is highly desirable for fluorescence imaging and therapeutic applications but remains challenging. Using a vinyl-grafted BMMs (bimodal mesoporous SiO2 materials) as a core and PAN ((2-aminoethyl)-6-(dimethylamino)-1H-benzo[de]isoquinoline-1,3(2H)-dione))-dispersed dual pH/thermal-sensitive poly(N-isopropylacrylamide-co-acrylic acid) as a shell, PAN@BMMs with strong fluorescence and good dispersibility were prepared. Their mesoporous features and physicochemical properties were extensively characterized via XRD patterns, N2 adsorption–desorption analysis, SEM/TEM images, TGA profiles, and FT-IR spectra. In particular, their mass fractal dimension (dm) features based on SAXS patterns combined with fluorescence spectra were successfully obtained to evaluate the uniformity of the fluorescence dispersions, showing that the dm values increased from 2.49 to 2.70 with an increase of the AN-additive amount from 0.05 to 1%, along with the red shifting of their fluorescent emission wavelength from 471 to 488 nm. The composite (PAN@BMMs-I-0.1) presented a densification trend and a slight decrease in peak (490 nm) intensity during the shrinking process. Its fluorescent decay profiles confirmed two fluorescence lifetimes of 3.59 and 10.62 ns. The low cytotoxicity obtained via in vitro cell survival assay and the efficient green imaging performed via HeLa cell internalization suggested that the smart PAN@BMM composites are potential carriers for in vivo imaging and therapy. [ABSTRACT FROM AUTHOR]

Hybrid emission filters, comprising an interference filter and an absorption filter, exhibit high excitation light rejection performance and can act as lensless fluorescent devices. However, it has been challenging to produce them in large batches over a large area. In this study, we propose and demonstrate a method for transferring a Si substrate, on which the hybrid filter is deposited, onto an image sensor by attaching it to the sensor and removing the substrate via plasma etching. Through this method, we can transfer uniform filters onto fine micrometer-sized needle devices and millimeter-sized multisensor chips. Optical evaluation reveals that the hybrid filter emits light in the 500 to 560 nm range, close to the emission region of green fluorescent protein (GFP). Furthermore, by observing the fluorescence emission from the microbeads, a spatial resolution of 12.11 μm is calculated. In vitro experiments confirm that the fabricated device is able to discriminate GFP emission patterns from brain slices. [ABSTRACT FROM AUTHOR]

Amino acid is the main transport form of reduced nitrogen in plants. To investigate the uptake and source–sink translocation process of plants to help understand their physiological roles and transport mechanisms, we designed and synthesized three fluorescent-dye-labeled amino acids as tools to visualize amino acid transportation in Arabidopsis thaliana; these amino acids consist of amino acids linked to the fluorophore nitrobenzoxadiazole (NBD) with excellent optical properties. Furthermore, we incubated Arabidopsis thaliana with these NBD fluorescent-dye-labeled amino acids for real-time imaging along with fluorescence enhancement for 24 h. The results showed that Arabidopsis thaliana could absorb them directly from the roots to the leaves. Therefore, our fluorescent-dye-labeled amino acids provide a de novo tool and strategy for visualizing amino acid absorption and transportation in plants. [ABSTRACT FROM AUTHOR]

Superoxide anion (O2•−) is an important reactive oxygen species (ROS), and plays critical roles in biological systems. ER stress has close relation with many metabolic diseases, and could lead to the abnormal production of ROS including O2•−. Herein, we present an ER-targeting probe (ER-Tf) for the detection of O2•− in living cells. The probe ER-Tf used triflate as the response site for O2•−, and employed p-methylbenzenesulfonamide as ER-targeting moiety. In response to O2•−, the triflate of the probe ER-Tf converted to hydroxyl group, providing strong blue emission under the excitation of ultraviolet light. The probe ER-Tf exhibited high sensitivity and selectivity to O2•−. Bioimaging experiments showed that the probe ER-Tf can be applied to detect O2•− at ER, and also demonstrated that rotenone could increase the generation of O2•− in living cells, while the O2•− level at ER showed no remarkable change during ferroptosis. [ABSTRACT FROM AUTHOR]

In this study, we report a fluorescent polydopamine nanoparticle-incorporated hydrogel (FPNP-hydrogel) for fluorescence imaging-guided photothermal cancer therapy. The FPNP-hydrogel was prepared by combining fluorescent polydopamine nanoparticles (FPNPs) with polyvinyl alcohol (PVA) using a simple method. The unique optical properties of the FPNPs facilitated the FPNP-hydrogel the excellent biocompatibility, strong fluorescence, and high photostability, showing an availability of fluorescence imaging in BALB/c nude mice. When combined with near-infrared laser irradiation, the FPNP-hydrogel noticeably reduced tumor volumes in 4T1 tumor-bearing mice within 14 days without inducing the inflammatory response in major organs. In conclusion, the results suggest that multifunctional hydrogels with excellent fluorescence properties and high photothermal conversion efficiency could be useful for designing novel therapeutic methods for cancer treatment. [ABSTRACT FROM AUTHOR]

BackgroundWe performed a systematic review that identified at least 9,000 scientific papers on PubMed that include immunofluorescent images of cells from the central nervous system (CNS). These CNS papers contain tens of thousands of immunofluorescent neural images supporting the findings of over 50,000 associated researchers. While many existing reviews discuss different aspects of immunofluorescent microscopy, such as image acquisition and staining protocols, few papers discuss immunofluorescent imaging from an image-processing perspective. We analyzed the literature to determine the image processing methods that were commonly published alongside the associated CNS cell, microscopy technique, and animal model, and highlight gaps in image processing documentation and reporting in the CNS research field.MethodsWe completed a comprehensive search of PubMed publications using Medical Subject Headings (MeSH) terms and other general search terms for CNS cells and common fluorescent microscopy techniques. Publications were found on PubMed using a combination of column description terms and row description terms. We manually tagged the comma-separated values file (CSV) metadata of each publication with the following categories: animal or cell model, quantified features, threshold techniques, segmentation techniques, and image processing software.ResultsOf the almost 9,000 immunofluorescent imaging papers identified in our search, only 856 explicitly include image processing information. Moreover, hundreds of the 856 papers are missing thresholding, segmentation, and morphological feature details necessary for explainable, unbiased, and reproducible results. In our assessment of the literature, we visualized current image processing practices, compiled the image processing options from the top twelve software programs, and designed a road map to enhance image processing. We determined that thresholding and segmentation methods were often left out of publications and underreported or underutilized for quantifying CNS cell research.DiscussionLess than 10% of papers with immunofluorescent images include image processing in their methods. A few authors are implementing advanced methods in image analysis to quantify over 40 different CNS cell features, which can provide quantitative insights in CNS cell features that will advance CNS research. However, our review puts forward that image analysis methods will remain limited in rigor and reproducibility without more rigorous and detailed reporting of image processing methods.ConclusionImage processing is a critical part of CNS research that must be improved to increase scientific insight, explainability, reproducibility, and rigor.

Abstract Near‐infrared fluorescence imaging is vital for exploring the biological world. The short emissions (100 nm) for THQ‐modified xanthene dyes is established. Thus, a timely discussion of THQ–xanthene and its applications is extensive. Hence, the advent, working principles, development trajectory, and biological applications of THQ–xanthene dyes, especially in the fields of fluorescence probe‐based sensing and imaging, cancer theranostics, and super‐resolution imaging, are introduced. It is envisioned that the THQ modification tactic is a simple yet exceptional approach to upgrade the performance of conventional xanthene dyes. THQ–xanthene will advance the strides of xanthene‐based potentials in early fluorescent diagnosis of diseases, cancer theranostics, and imaging‐guided surgery.

Herein, a novel fluorescent probe RhoDCM was developed for monitoring the cysteine (Cys) dynamics. For the first time, the Cys-triggered implement was applied in relatively complete diabetic mice models. The response of RhoDCM towards Cys suggested advantages including practical sensitivity, high selectivity, rapid reaction, and steadiness in various pH and temperature conditions. RhoDCM could basically monitor the intracellular Cys level, both exogenous and endogenous. It could further monitor the glucose level via detecting consumed Cys. Furthermore, the diabetic mice models including the no diabetic control group, the induced model groups by streptozocin (STZ) or alloxan, and the treatment groups induced by STZ and treated with vildagliptin (Vil), dapagliflozin (DA), or metformin (Metf) were constructed. The models were checked by oral glucose tolerance test and significant liver-related serum indexes. Based on the models, the in vivo imaging and penetrating depth fluorescence imaging both indicated that RhoDCM could characterize the status of the development and treatment in the diabetic process via monitoring the Cys dynamics. Consequently, RhoDCM seemed beneficial for inferring the order of severity in the diabetic process and evaluating the potency of therapeutic schedules, which might be informatic for correlated investigations.

To date, 14C tracer studies using accelerator mass spectrometry (AMS) have not yet resolved lipid-soluble analytes into individual lipoprotein density subclasses. The objective of this work was to develop a reliable method for lipoprotein separation and quantitative recovery for biokinetic modeling purposes. The novel method developed provides the means for use of small volumes (10–200 µL) of frozen plasma as a starting material for continuous isopycnic lipoprotein separation within a carbon- and pH-stable analyte matrix, which, following post-separation fraction clean up, created samples suitable for highly accurate 14C/12C isotope ratio determinations by AMS. Manual aspiration achieved 99.2 ± 0.41% recovery of [5-14CH3]-(2R, 4′R, 8′R)-α-tocopherol contained within 25 µL plasma recovered in triacylglycerol rich lipoproteins (TRL = Chylomicrons + VLDL), LDL, HDL, and infranatant (INF) from each of 10 different sampling times for one male and one female subject, n = 20 total samples. Small sample volumes of previously frozen plasma and high analyte recoveries make this an attractive method for AMS studies using newer, smaller footprint AMS equipment to develop genuine tracer analyses of lipophilic nutrients or compounds in all human age ranges.

The outcomes of unresectable gastric cancer (GC) are unfavorable even with chemotherapy; therefore, a new treatment modality is required. The combination of an oncolytic virus and photodynamic therapy can be one of the promising modalities to overcome this. Mammalian orthoreovirus (MRV) is an oncolytic virus that has been used in clinical trials for several cancers. In this study, we developed and evaluated a recombinant MRV strain type 3 Dearing (T3D) that expresses membrane-targeting KillerRed (KRmem), a phototoxic fluorescent protein that produces cytotoxic reactive oxygen species upon light irradiation. KRmem was fused in-frame to the 3′ end of the σ2 viral gene in the S2 segment using a 2A peptide linker, enabling the expression of multiple proteins from a single transcript. RNA electrophoresis, Western blotting, and immunofluorescence analyses confirmed functional insertion of KRmem into the recombinant virus. The growth activity of the recombinant virus was comparable to that of the wild-type MRV in a cultured cell line. The recombinant virus infected two GC cell lines (MKN45P and MKN7), and a significant cytocidal effect was observed in MKN45P cells infected with the recombinant virus after light irradiation. Thus, recombinant MRV-expressing KRmem has the potential to serve as a novel treatment tool for GC.