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Valproic acid (VPA), a clinically approved small molecule, has been reported to activate Wnt signalling that is critical for dorsal-ventral (DV) patterning of neural tube. However, little is known about the impact of VPA on DV patterning process. Here, we show that even though VPA has a negative impact on the early formation of human neural tube organoids (hNTOs), it significantly enhances the efficiency of ventrally patterned hNTOs, when VPA is added during the entire differentiation process. RNA sequencing and RT-qPCR analysis demonstrates VPA activates endogenous Wnt signalling in hNTOs. Surprisingly, transcriptome analysis also identifies upregulation of genes for degradation of GLI2 and GLI3 proteins, whose truncated fragment are transcriptional repressors of Shh signalling. The Western-blot analysis confirms the increase of GLI3R proteins after VPA treatment. Thus, VPA might enhance ventral patterning of hNTOs through both activating Wnt, which can antagonise Shh signalling by inducing GLI3 expression, and/or inhibiting Shh signalling by inducing GLI protein degradation. We further obtain results to show that VPA still increases patterning efficiency of hNTOs with a weak influence on their early formation when the initiation time of VPA is delayed and its duration is reduced. Taken together, this study demonstrates that VPA enhances the generation of more reproducible hNTOs with ventral patterning, opening the avenues for the applications of hNTOs in developmental biology and regenerative medicine., (© 2024 The Author(s). Cell Proliferation published by Beijing Institute for Stem Cell and Regenerative Medicine and John Wiley & Sons Ltd.)

Mounting evidence in animal experiments proves that early life stage exposure to organophosphate flame retardants (OPFRs) affects the locomotor behavior and changes the transcriptions of central nervous system genes. Unfortunately, their effect on human motor neuron (MN) development, which is necessary for body locomotion and survival, has not yet characterized. Here, we utilized a spinal cord MN differentiation model from human embryonic stem cells (ESCs) and adopted this model to test the effects of two typical OPFRs tris (2-butoxyethyl) phosphate (TBEP) and tris (2-chloroethyl) phosphate (TCEP), on MN development and the possible mechanisms underlying. Our findings revealed TBEP exerted a much more inhibitory effect on MN survival, while TCEP exhibited a stronger stimulatory effect on ESCs differentiation into MN, and thus TBEP exhibited a stronger inhibition on MN development than TCEP. RNA sequencing analysis identified TBEP and TCEP inhibited MN survival mainly by disrupting extracellular matrix (ECM)-receptor interaction. Focusing on the pathway guided MN differentiation, we found both TBEP and TCEP activated BMP signaling, whereas TCEP simultaneously downregulated Wnt signaling. Collectively, this is the first study demonstrated TBEP and TCEP disrupted human MN development by affecting their survival and differentiation, thereby raising concern about their potential harm in causing MN disorders., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Excessive exposure to light is a global issue. Artificial light pollution has been shown to disrupt the body's natural circadian rhythm. To investigate the impacts of light on metabolism, we studied Sprague-Dawley rats chronically exposed to red or blue light during daytime or nighttime. Rats in the experimental group were exposed to extended light for 4 hours during daytime or nighttime to simulate the effects of excessive light usage. Strikingly, we found systemic metabolic alterations only induced by blue light during daytime. Furthermore, we conducted metabolomic analyses of the cerebrospinal fluid, serum, heart, liver, spleen, adrenal, cerebellum, pituitary, prostate, spermatophore, hypothalamus and kidney from rats in the control and blue light exposure during daytime. Significant changes in metabolites have been observed in cerebrospinal fluid, serum, hypothalamus and kidney of rats exposed to blue light during daytime. Metabolic alterations observed in rats encompassing pyruvate metabolism, glutathione metabolism homocysteine degradation, phosphatidylethanolamine biosynthesis, and phospholipid biosynthesis, exhibit analogous patterns to those inherent in specific physiological processes, notably neurodevelopment, cellular injury, oxidative stress, and autophagic pathways. Our study provides insights into tissue-specific metabolic changes in rats exposed to blue light during the daytime and may help explain potential mechanisms of photopathogenesis., Competing Interests: Declaration of Competing Interest All authors declare that they have no conflicts of interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Studies have shown that bortezomib resistance in multiple myeloma (MM) is mediated by the abnormalities of various molecules and microenvironments. Exploring these resistance mechanisms will improve the therapeutic efficacy of bortezomib. In this study, bone marrow tissues from three patients with MM, both sensitive and resistant to bortezomib, were collected for circRNA high-throughput sequencing analysis. The relationship between circ_0000337, miR-98-5p, and target gene DNA2 was analyzed by luciferase detection and verified by RT-qPCR. We first found that circ_0000337 was significantly upregulated in bortezomib-resistant MM tissues and cells, and overexpression of circ_0000337 could promote bortezomib resistance in MM cells. circ_0000337 may act as a miR-98-5p sponge to upregulate DNA2 expression, regulate DNA damage repair, and induce bortezomib resistance. Furthermore, it was determined that the increased circ_0000337 level in bortezomib-resistant cells was due to an increased N6-methyladenosine (m6A) level, resulting in enhanced RNA stability. In conclusion, the m6A level of circ_0000337 and its regulation may be a new and potential therapeutic target for overcoming bortezomib resistance in MM., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Jiang, Gao, Li, Zhang, Zhang and Liu.)

Arginine methylation (ArgMet), as a post-translational modification, plays crucial roles in RNA processing, transcriptional regulation, signal transduction, DNA repair, apoptosis and liquid-liquid phase separation (LLPS). Since arginine methylation is associated with cancer pathogenesis and progression, protein arginine methyltransferases have gained interest as targets for anti-cancer therapy. Despite considerable process made to elucidate (patho)physiological mechanisms regulated by arginine methylation, there remains a lack of tools to visualize arginine methylation with high spatiotemporal resolution in live cells. To address this unmet need, we generated an ArgMet-sensitive genetically encoded, Förster resonance energy transfer-(FRET) based biosensor, called GEMS, capable of quantitative real-time monitoring of ArgMet dynamics. We optimized these biosensors by using different ArgMet-binding domains, arginine-glycine-rich regions and adjusting the linkers within the biosensors to improve their performance. Using a set of mammalian cell lines and modulators, we demonstrated the applicability of GEMS for monitoring changes in arginine methylation with single-cell and temporal resolution. The GEMS can facilitate the in vitro screening to find potential protein arginine methyltransferase inhibitors and will contribute to a better understanding of the regulation of ArgMet related to differentiation, development and disease., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

The increasing demand for rare earth elements (REEs) in modern applications has drawn significant attention. REEs can be introduced into the environment through REE-containing fertilizers, abandoned REE-rich equipment, and mining, persisting and impacting soil quality, nutrient cycles, and plant growth. Scientists have raised concerns about REEs entering the food chain from the environment and eventually accumulating in organisms. Decades of experimental evidence have shown that these effects include inhibited growth, impaired liver function, and alterations in children's intelligence quotients. However, there exists a paucity of research that has elucidated the metabolic-level biological impacts of REEs. In our study, Caenorhabditis elegans (C. elegans) was used as a model organism to investigate physiological and inherent metabolic changes under exposure to different concentrations of REEs. The diet bacteria of nematodes play a key role in their life and development. Therefore, we investigated the influence of bacterial activity on the nematodes' response to REE exposure. We observed a concentration-dependent accumulation of REEs in nematodes, which consequently led to a reduction in lifespan and alterations in body length. Exposure to a mixed solution of REEs, in comparison to a single REE solution, resulted in greater toxicity toward nematodes. The metabolic results showed that the above changes were closely related to REE-induced amino acid metabolism disorder, membrane disturbance, DNA damage, and oxidative stress. Of note, the presence of living bacteria elicits REE effects in C. elegans. These findings highlight the potential intrinsic metabolic changes occurring in nematodes under REE exposure. Our study raises awareness of the exposure risks associated with REEs, provides valuable insight into the metabolic-level biological impacts of REEs and contributes to the development of effective mitigation strategies to reduce potential risks to human health., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: POEMS syndrome is a rare and complex plasma cell disorder characterized by polyneuropathy, organomegaly, endocrinopathy, high M protein levels, and skin changes. Understanding of POEMS syndrome has advanced rapidly since the 21st century. This study aims to summarize and evaluate the research status of POEMS syndrome in the past 23 years through a bibliometric and visualization analysis, and identify research focuses and emerging hotspots for the future., Methods: POEMS syndrome-related articles published between January 1, 2000, and March 8, 2023, were systematically retrieved from the Web of Science Core Collection. Data processing and visualization analysis were carried out using a combination of R software, HistCite, VOSviewer, and CiteSpace., Results: Since entering the 21st century, 3677 authors from 1125 institutions in 68 countries/regions have published 830 original and review articles on POEMS syndrome in 408 journals so far, among which the USA, Japan, and China published the most articles, and Mayo Clinic, Udice French Research Universities, and Peking Union Medical College listed the top three most prolific institutions. However, collaborative research across countries and groups in the study of POEMS syndrome remain significantly limited. Angela Dispenzieri ranked first in POEMS syndrome research from every aspect of authors, producing the most papers and contributing the most-cited article, followed by Satoshi Kuwabara and Sonoko Misawa. Internal medicine was the most productive journal on POEMS syndrome. "endothelial growth factor" was the keyword with the highest occurrence except for "POEMS syndrome", and "bevacizumab", "lenalidomide", "dexamethasone", and "management" were recognized as emerging topics., Conclusion: This study utilized bibliometric and visualization analysis to systematically summarize the research of POEMS syndrome in the first two decades of the 21st century, offering a data-based and objective perspective on the field of POEMS syndrome and guiding researchers in the identification of novel research directions., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 The Authors.)

The activating protein-1 (AP-1) transcription factors (TFs) have been associated with many different cancer types and are promising therapeutic targets in logical malignancies. However, the mechanisms of their role in multiple myeloma (MM) remain elusive. The present study determined and compared the mRNA and protein expression levels of the AP-1 family member JunB in CD138 + mononuclear cells from MM patients and healthy donors. Herein, we investigated the effect of T-5224, an inhibitor of JUN/AP-1, on MM. We found that the cytotoxicity of T-5224 toward myeloma is due to its ability to induce cell apoptosis, inhibit proliferation, and induce cell cycle arrest by increasing the levels of cleaved caspase3/7 and concomitantly inhibiting the IRF4/MYC axis. We also noticed that siJunB-mediated deletion of JunB/AP-1 enhanced MM cell apoptosis and affected cell proliferation. The software PROMO was used in the present study to predict the AP-1 TF that may bind the promoter region of IRF4. We confirmed the correlation between JunB/AP-1 and IRF4. Given that bortezomib (BTZ) facilitates IRF4 degradation in MM cells, we applied combination treatment of BTZ with T-5224. T-5224 and BTZ exerted synergistic effects, and T-5224 reversed the effect of BTZ on CD138 + primary resistance in MM cells, in part due to suppression of the IRF4/MYC axis. Our results suggest that targeting AP-1 TFs is a promising therapeutic strategy for MM. Additionally, targeting both AP-1 and IRF4 with T-5224 may be a synergistic therapeutic strategy for this clinically challenging subset of MM., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Multiple myeloma (MM) is a malignant and incurable disease. Chemotherapy is currently the primary treatment option for MM. However, chemotherapeutic drugs can interrupt treatment because of serious side effects. Therefore, development of novel therapeutics for MM is essential. In this study, we designed and constructed an innovative nanoparticle-based drug delivery system, P-R@Ni 3 P-BTZ, and investigated its feasibility, effectiveness, and safety both in vitro and in vivo. P-R@Ni 3 P-BTZ is a nanocomposite that consists of two parts: (1) the drug carrier (Ni 3 P), which integrates photothermal therapy (PTT) with chemotherapy by loading bortezomib (BTZ); and (2) the shell (P-R), a CD38 targeting peptide P-modified red blood cell membrane nanovesicles. In vitro and in vivo, it was proven that P-R@Ni 3 P-BTZ exhibits remarkable antitumor effects by actively targeting CD38 + MM cells. P-R@Ni 3 P-BTZ significantly induces the accumulation of intracellular reactive oxygen species (ROS) and increases the apoptosis of MM cells, which underlies the primary mechanism of its antitumor effects. In addition, P-R@Ni 3 P exhibits good biocompatibility and biosafety, both in vitro and in vivo. Overall, P-R@Ni 3 P-BTZ is a specific and efficient MM therapeutic method., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

Nanotoxicology on plants has raised great concerns about the prevalence of nanoparticles (NPs) in ecosystems, and the most fundamental aspect is to assess the fate of NPs in plants. Nonetheless, the lack of high-sensitivity tracking techniques in vivo constraints intensive research on NP distribution in plants. Herein, we have overcome this limitation by first introducing persistent luminescent nanoparticles (PLNPs) as an imaging probe, which can detect the distribution of NPs in whole plants with high sensitivity by completely eliminating interference from autofluorescence. We synthesized two differently surface-charged PLNPs with excellent biosafety and then exposed these NPs to plants in the hydroponic medium. Persistent luminescence (PersL) images visually showed the disparate accumulation of PLNPs in plants. PersL signals of positively charged PLNPs were observed in the whole exposed portion of the plant roots, while negatively charged PLNPs were mainly in the root collars instead of the exposed portion. With prolonged exposure, the presence of PersL signals in leaves indicated the long-distance translocation of differently charged PLNPs from roots to leaves via hypocotyls. For further confirmation of imaging results, the distribution of NPs in plants was investigated using electron microscopy. Based on their unique optical properties, PLNPs provide a promising strategy for tracking NP's fate in plants.

Frontotemporal dementia (FTD) is a heterogeneous neurodegenerative disease with multiple histopathological subtypes. FTD patients share similar symptoms with Alzheimer's disease (AD). Hence, FTD patients are commonly misdiagnosed as AD, despite the consensus clinical diagnostic criteria. It is therefore of great clinical need to identify a biomarker that can distinguish FTD from AD and control individuals, and potentially further differentiate between FTD pathological subtypes. We conducted a metabolomic analysis on post-mortem human brain tissue from three regions: cerebellum, frontal cortex and occipital cortex from control, FTLD-TDP type A, type A-C9, type C and AD. Our results indicate that the brain subdivisions responsible for different functions show different metabolic patterns. We further explored the region-specific metabolic characteristics of different FTD subtypes and AD patients. Different FTD subtypes and AD share similar metabolic phenotypes in the cerebellum, but AD exhibited distinct metabolic patterns in the frontal and occipital regions compared to FTD. The identified brain region-specific metabolite biomarkers could provide a tool for distinguishing different FTD subtypes and AD and provide the first insights into the metabolic changes of FTLD-TDP type A, type A-C9, type C and AD in different regions of the brain. The importance of protein arginine methylation in neurodegenerative disease has come to light, so we investigated whether the arginine methylation level contributes to disease pathogenesis. Our findings provide new insights into the relationship between arginine methylation and metabolic changes in FTD subtypes and AD that could be further explored, to study the molecular mechanism of pathogenesis., Competing Interests: Declaration of Competing Interest The authors declare that this article was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: Clear cell sarcomas (CCSs) are translocated aggressive malignancies, most commonly affecting young adults with a high incidence of metastases and a poor prognosis. Research into the disease is more feasible when adequate models are available. By establishing CCS cell lines from a primary and metastatic lesion and isolating healthy fibroblasts from the same patient, the in vivo process is accurately reflected and aspects of clinical multistep carcinogenesis recapitulated., Methods: Isolated tumor cells and normal healthy skin fibroblasts from the same patient were compared in terms of growth behavior and morphological characteristics using light and electron microscopy. Tumorigenicity potential was determined by soft agar colony formation assay and in vivo xenograft applications. While genetic differences between the two lineages were examined by copy number alternation profiles, nuclear magnetic resonance spectroscopy determined arginine methylation as epigenetic features. Potential anti-tumor effects of a protein arginine N-methyltransferase type I (PRMT1) inhibitor were elicited in 2D and 3D cell culture experiments using cell viability and apoptosis assays. Statistical significance was calculated by one-way ANOVA and unpaired t-test., Results: The two established CCS cell lines named MUG Lucifer prim and MUG Lucifer met showed differences in morphology, genetic and epigenetic data, reflecting the respective original tissue. The detailed cell line characterization especially in regards to the epigenetic domain allows investigation of new innovative therapies. Based on the epigenetic data, a PRMT1 inhibitor was used to demonstrate the targeted antitumor effect; normal tissue cells isolated and immortalized from the same patient were not affected with the IC 50 used., Conclusions: MUG Lucifer prim, MUG Lucifer met and isolated and immortalized fibroblasts from the same patient represent an ideal in vitro model to explore the biology of CCS. Based on this cell culture model, novel therapies could be tested in the form of PRMT1 inhibitors, which drive tumor cells into apoptosis, but show no effect on fibroblasts, further supporting their potential as promising treatment options in the combat against CCS. The data substantiate the importance of tailored therapies in the advanced metastatic stage of CCS., (© 2023. The Author(s).)

Phospholipid metabolism, including phosphatidylcholine (PC) biosynthesis, is crucial for various biological functions and is associated with longevity. Phosphatidylethanolamine N -methyltransferase (PEMT) is a protein that catalyzes the biosynthesis of PC, the levels of which change in various organs such as the brain and kidneys during aging. However, the role of PEMT for systemic PC supply is not fully understood. To address how PEMT affects aging-associated energy metabolism in tissues responsible for nutrient absorption, lipid storage, and energy consumption, we employed NMR-based metabolomics to study the liver, plasma, intestine (duodenum, jejunum, and ileum), brown/white adipose tissues (BAT and WAT), and skeletal muscle of young (9-10 weeks) and old (91-132 weeks) wild-type (WT) and PEMT knockout (KO) mice. We found that the effect of PEMT-knockout was tissue-specific and age-dependent. A deficiency of PEMT affected the metabolome of all tissues examined, among which the metabolome of BAT from both young and aged KO mice was dramatically changed in comparison to the WT mice, whereas the metabolome of the jejunum was only slightly affected. As for aging, the absence of PEMT increased the divergence of the metabolome during the aging of the liver, WAT, duodenum, and ileum and decreased the impact on skeletal muscle. Overall, our results suggest that PEMT plays a previously underexplored, critical role in both aging and energy metabolism.

Active thermogenic adipocytes avidly consume energy substrates like fatty acids and glucose to maintain body temperature upon cold exposure. Despite strong evidence for the involvement of brown adipose tissue (BAT) in controlling systemic energy homeostasis upon nutrient excess, it is unclear how the activity of brown adipocytes is regulated in times of nutrient scarcity. Therefore, this study aimed to scrutinize factors that modulate BAT activity to balance thermogenic and energetic needs upon simultaneous fasting and cold stress. For an unbiased view, we performed transcriptomic and miRNA sequencing analyses of BAT from acutely fasted (24 h) mice under mild cold exposure. Combining these data with in-depth bioinformatic analyses and in vitro gain-of-function experiments, we define a previously undescribed axis of p53 inducing miR-92a-1-5p transcription that is highly upregulated by fasting in thermogenic adipocytes. p53, a fasting-responsive transcription factor, was previously shown to control genes involved in the thermogenic program and miR-92a-1-5p was found to negatively correlate with human BAT activity. Here, we identify fructose transporter Slc2a5 as one direct downstream target of this axis and show that fructose can be taken up by and metabolized in brown adipocytes. In sum, this study delineates a fasting-induced pathway involving p53 that transactivates miR-92a-1-5p, which in turn decreases Slc2a5 expression, and suggests fructose as an energy substrate in thermogenic adipocytes., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Reinisch, Klymiuk, Michenthaler, Moyschewitz, Galhuber, Krstic, Domingo, Zhang, Karbiener, Vujić, Kratky, Schreiber, Schupp, Lenihan-Geels, Schulz, Malli, Madl, Prokesch.)

Constructing hierarchical micro/nanostructures as anodes for sodium ion batteries is an important approach for exploiting efficient energy storage devices. Herein, sandwich structure hierarchical nanofibers composed of hollow carbon fibers as the substrate, and MoS 2 as the interlayer with Co and/or ZnS nanoparticles anchoring in carbon skeletons as the outer shell (carbon nanofiber/MoS 2 /Co-ZnS⊂NC) are prepared via a multistep reaction strategy. Profiting from the unique hierarchical structure, abundant migration channels of Na + , and multicomponent synergistic effects, the rapid diffusion kinetics are ensured and the utilization of active materials is maximized. The coaxial structure can evenly disperse volumetric strain, making structural stability guaranteed. Hierarchical nanofibers deliver a high reversible capacity of 352.3 mAh g -1 at 5.0 A g -1 over 3000 cycles. A discharge capacity of 182.5 mAh g -1 is retained even after 10 000 cycles at 10.0 A g -1 as well as a high rate capacity of 202.0 mAh g -1 up to 30 A g -1 . The optimal atomic ratio of Co element is further verified by the kinetic analysis. The full-cells assembled with Na 3 V 2 (PO 4 ) 3 cathode provide a high capacity of 179.2 mAh g -1 at 1.0 A g -1 for 500 cycles. Combining in situ and ex situ characterizations and theoretical calculations, possible sodium storage mechanisms and the origin of superior electrochemical properties are revealed., (© 2022 Wiley-VCH GmbH.)

Hepatitis B virus (HBV) infection is a worldwide health burden. Metabolomics analysis has revealed HBV-induced metabolism dysregulation in liver tissues and hepatocytes. However, as an infectious disease, the tissue-specific landscape of metabolic profiles of HBV infection remains unclear. To fill this gap, we applied untargeted nuclear magnetic resonance (NMR) metabolomic analysis of the heart, liver, spleen, lung, kidney, pancreas, and intestine (duodenum, jejunum, ileum) in HBV-transgenic mice and their wild-type littermates. Strikingly, we found systemic metabolic alterations induced by HBV in liver and extrahepatic organs. Significant changes in metabolites have been observed in most tissues of HBV-transgenic mice, except for ileum. The metabolic changes may provide novel therapeutic targets for the treatment of HBV infection. Moreover, tissue-specific metabolic profiles could speed up the study of HBV induced systemic metabolic reprogramming, which could help follow the progression of HBV infection and explain the underlying pathogenesis.

A fundamental step in developing a protein drug is the selection of a stable storage formulation that ensures efficacy of the drug and inhibits physiochemical degradation or aggregation. Here, we designed and evaluated a general workflow for screening of protein formulations based on small-angle X-ray scattering (SAXS). Our SAXS pipeline combines automated sample handling, temperature control, and fast data analysis and provides protein particle interaction information. SAXS, together with different methods including turbidity analysis, dynamic light scattering (DLS), and SDS-PAGE measurements, were used to obtain different parameters to provide high throughput screenings. Using a set of model proteins and biopharmaceuticals, we show that SAXS is complementary to dynamic light scattering (DLS), which is widely used in biopharmaceutical research and industry. We found that, compared to DLS, SAXS can provide a more sensitive measure for protein particle interactions, such as protein aggregation and repulsion. Moreover, we show that SAXS is compatible with a broader range of buffers, excipients, and protein concentrations and that in situ SAXS provides a sensitive measure for long-term protein stability. This workflow can enable future high-throughput analysis of proteins and biopharmaceuticals and can be integrated with well-established complementary physicochemical analysis pipelines in (biopharmaceutical) research and industry.

Energy metabolism, including alterations in energy intake and expenditure, is closely related to aging and longevity. Metabolomics studies have recently unraveled changes in metabolite composition in plasma and tissues during aging and have provided critical information to elucidate the molecular basis of the aging process. However, the metabolic changes in tissues responsible for food intake and lipid storage have remained unexplored. In this study, we aimed to investigate aging-related metabolic alterations in these tissues. To fill this gap, we employed NMR-based metabolomics in several tissues, including different parts of the intestine (duodenum, jejunum, ileum) and brown/white adipose tissues (BAT, WAT), of young (9-10 weeks) and old (96-104 weeks) wild-type (mixed genetic background of 129/J and C57BL/6) mice. We, further, included plasma and skeletal muscle of the same mice to verify previous results. Strikingly, we found that duodenum, jejunum, ileum, and WAT do not metabolically age. In contrast, plasma, skeletal muscle, and BAT show a strong metabolic aging phenotype. Overall, we provide first insights into the metabolic changes of tissues essential for nutrient uptake and lipid storage and have identified biomarkers for metabolites that could be further explored, to study the molecular mechanisms of aging.

Protein-bound arginine is commonly methylated in many proteins and regulates their function by altering the physicochemical properties, their interaction with other molecules, including other proteins or nucleic acids. This work presents an easily implementable protocol for quantifying arginine and its derivatives, including asymmetric and symmetric dimethylarginine (ADMA and SDMA, respectively) and monomethyl arginine (MMA). Following protein isolation from biological body fluids, tissues, or cell lysates, a simple method for homogenization, precipitation of proteins, and protein hydrolysis is described. Since the hydrolysates contain many other components, such as other amino acids, lipids, and nucleic acids, a purification step using solid-phase extraction (SPE) is essential. SPE can either be performed manually using centrifuges or a pipetting robot. The sensitivity for ADMA using the current protocol is about 100 nmol/L. The upper limit of detection for arginine is 3 mmol/L due to SPE saturation. In summary, this protocol describes a robust method, which spans from biological sample preparation to NMR-based detection, providing valuable hints and pitfalls for successful work when studying the arginine methylome.

Arginine-glycine(-glycine) (RG/RGG) regions are highly abundant in RNA-binding proteins and involved in numerous physiological processes. Aberrant liquid-liquid phase separation (LLPS) and stress granule (SGs) association of RG/RGG regions in the cytoplasm have been implicated in several neurodegenerative disorders. LLPS and SG association of these proteins is regulated by the interaction with nuclear import receptors, such as transportin-1 (TNPO1), and by post-translational arginine methylation. Strikingly, many RG/RGG proteins harbour potential phosphorylation sites within or close to their arginine methylated regions, indicating a regulatory role. Here, we studied the role of phosphorylation within RG/RGG regions on arginine methylation, TNPO1-binding and LLPS using the cold-inducible RNA-binding protein (CIRBP) as a paradigm. We show that the RG/RGG region of CIRBP is in vitro phosphorylated by serine-arginine protein kinase 1 (SRPK1), and discovered two novel phosphorylation sites in CIRBP. SRPK1-mediated phosphorylation of the CIRBP RG/RGG region impairs LLPS and binding to TNPO1 in vitro and interferes with SG association in cells. Furthermore, we uncovered that arginine methylation of the CIRBP RG/RGG region regulates in vitro phosphorylation by SRPK1. In conclusion, our findings indicate that LLPS and TNPO1-mediated chaperoning of RG/RGG proteins is regulated through an intricate interplay of post-translational modifications., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Lenard, Hutten, Zhou, Usluer, Zhang, Bourgeois, Dormann and Madl.)

Due to the complex structure and function of central nervous system (CNS), human CNS in vitro modeling is still a great challenge. Neurotoxicity testing of environmental chemicals mainly depends on the traditional animal models, which have various limitations such as species differences, expensive and time-consuming. Meanwhile, in vitro two-dimensional (2D) cultured cells or three-dimensional (3D) cultured neurospheres cannot fully simulate complex 3D structure of neural tissues. Recent advancements in neural organoid systems provides excellent models for the testing of environmental chemicals that affect the development of human CNS. Neural organoids derived from hPSCs not only can simulate the process of CNS development, including early stage neural tube formation, neuroepithelium differentiation and regional specification, but also its 3D structure, thus can be used to evaluate the effect of chemicals on differentiation and morphogenesis. Here, we provide a review of recent progress in the methods of culturing neural organoids and their applications in neurotoxicity testing of environmental chemicals. We conclude by highlighting challenge and future directions in neurotoxicity testing based on neural organoids., (© 2021. The Author(s).)

Sensitive, selective, rapid, and label-free detection of pathogenic bacteria with high generality is of great importance for clinical diagnosis, biosecurity, and public health. However, most traditional approaches, such as microbial cultures, are time-consuming and laborious. To circumvent these problems, surface-enhanced Raman spectroscopy (SERS) appears to be a powerful technique to characterize bacteria at the single-cell level. Here, by SERS, we report a strategy for the rapid and specific detection of 22 strains of common pathogenic bacteria. A novel and high-quality silver nanorod SERS substrate, prepared by the facile interface self-assembly method, was utilized to acquire the chemical fingerprint information of pathogens with improved sensitivity. We also applied the mathematical analysis methods, such as the t -test and receiver operating characteristic method, to determine the Raman features of these 22 strains and demonstrate the clear identification of most bacteria (20 strains) from the rest and also the reliability of this SERS sensor. This rapid and specific strategy for wide-range bacterial detection offers significant advantages over existing approaches and sets the base for automated and onsite detection of pathogenic bacteria in a complex real-life situation.

Intrinsically disordered proteins and proteins containing intrinsically disordered regions are highly abundant in the proteome of eukaryotes and are extensively involved in essential biological functions. More recently, their role in the organization of biomolecular condensates has become evident and along with their misregulation in several neurologic disorders. Currently, most studies involving these proteins are carried out in vitro and using purified proteins. Given that in cells, condensate-forming proteins are exposed to high, millimolar concentrations of cellular metabolites, we aimed to reveal the interactions of cellular metabolites and a representative condensate-forming protein. Here, using the arginine-glycine/arginine-glycine-glycine (RG/RGG)-rich cold inducible RNA binding protein (CIRBP) as paradigm, we studied binding of the cellular metabolome to CIRBP. We found that most of the highly abundant cellular metabolites, except nucleotides, do not directly bind to CIRBP. ATP, ADP, and AMP as well as NAD + , NADH, NADP + , and NADPH directly interact with CIRBP, involving both the folded RNA-recognition motif and the disordered RG/RGG region. ATP binding inhibited RNA-driven phase separation of CIRBP. Thus, it might be beneficial to include cellular metabolites in in vitro liquid-liquid phase separation studies of RG/RGG and other condensate-forming proteins in order to better mimic the cellular environment in the future., (© 2021 The Authors. Protein Science published by Wiley Periodicals LLC on behalf of The Protein Society.)

Quantitative information about the levels and dynamics of post-translational modifications (PTMs) is critical for an understanding of cellular functions. Protein arginine methylation (ArgMet) is an important subclass of PTMs and is involved in a plethora of (patho)physiological processes. However, because of the lack of methods for global analysis of ArgMet, the link between ArgMet levels, dynamics, and (patho)physiology remains largely unknown. We utilized the high sensitivity and robustness of nuclear magnetic resonance (NMR) spectroscopy to develop a general method for the quantification of global protein ArgMet. Our NMR-based approach enables the detection of protein ArgMet in purified proteins, cells, organoids, and mouse tissues. We demonstrate that the process of ArgMet is a highly prevalent PTM and can be modulated by small-molecule inhibitors and metabolites and changes in cancer and during aging. Thus, our approach enables us to address a wide range of biological questions related to ArgMet in health and disease., Competing Interests: The authors declare no competing interests., (© 2021 The Authors.)

Background: tRNA-derived fragments have been reported to be key regulatory factors in human tumors. However, their roles in the progression of multiple myeloma remain unknown., Results: This study employed RNA-sequencing to explore the expression profiles of tRFs/tiRNAs in new diagnosed MM and relapsed/refractory MM samples. The expression of selected tRFs/tiRNAs were further validated in clinical specimens and myeloma cell lines by qPCR. Bioinformatic analysis was performed to predict their roles in multiple myeloma progression.We identified 10 upregulated tRFs/tiRNAs and 16 downregulated tRFs/tiRNAs. GO enrichment and KEGG pathway analysis were performed to analyse the functions of 1 significantly up-regulated and 1 significantly down-regulated tRNA-derived fragments. tRFs/tiRNAs may be involved in MM progression and drug-resistance., Conclusion: tRFs/tiRNAs were dysregulated and could be potential biomarkers for relapsed/refractory MM.

Hepatocellular carcinoma (HCC) is a common malignancy with poor prognosis, high morbidity and mortality concerning with lack of effective diagnosis and high postoperative recurrence. Similar with other cancers, HCC cancer cells have to alter their metabolism to adapt to the changing requirements imposed by the environment of the growing tumor. In less vascularized regions of tumor, cancer cells experience hypoxia and nutrient starvation. Here, we show that HCC undergoes a global metabolic reprogramming during tumor growth. A combined proteomics and metabolomics analysis of paired peritumoral and tumor tissues from 200 HCC patients revealed liver-specific metabolic reprogramming and metabolic alterations with increasing tumor sizes. Several proteins and metabolites associated with glycolysis, the tricarboxylic acid cycle and pyrimidine synthesis were found to be differentially regulated in serum, tumor and peritumoral tissue with increased tumor sizes. Several prognostic metabolite biomarkers involved in HCC metabolic reprogramming were identified and integrated with clinical and pathological data. We built and validated this combined model to discriminate against patients with different recurrence risks. An integrated and comprehensive metabolomic analysis of HCC is provided by our present work. Metabolomic alterations associated with the advanced stage of the disease and poor clinical outcomes, were revealed. Targeting cancer metabolism may deliver effective therapies for HCC.

Background: Multiple myeloma (MM) is incurable in spite of recent treatment improvements, highlighting the development of new therapies. Chimeric antigen receptor (CAR) T-cell therapy has dramatically changed the therapeutic effectiveness in high-risk B-cell malignancies. For relapsed/refractory multiple myeloma (RRMM), preclinical evaluations of CAR-T therapy have shown promising efficacy, thus various active clinical trials are under way. Herein, we conducted this review to summarize efficacy and safety of CAR-T therapy and provide more evidence to guide clinical treatments. Method: We systematically searched literature based on databases (PubMed, EMBASE, Cochrane Central Register of Controlled Trials), and conference abstracts reported from American Society of Hematology (ASH), European Hematology Association (EHA) and American Society of Clinical Oncology (ASCO), in addition to other sources (www.clinicaltrials.gov, article citations). Data assessed efficacy and safety of CAR-T therapy in patients with RRMM were extracted and evaluated, and then systematically analyzed by Comprehensive Meta-analysis 3.0 (CMA 3.0). Results: A total of 23 studies including 350 participants from different countries, diagnosed as RRMM and treated with CAR-T therapy (containing 7 antigens targeted by CARs) were combined. In summary, we discovered the pooled overall response rate (77%), complete response rate (37%) and minimal residual disease (MRD) negativity rate within responders (78%). Furthermore, the pooled relapse rate of responders was 38% and median progression-free survival was 8 months. The pooled survival rate was 87% at last follow-up (median, 12 months). In addition, the pooled grade 3-4 rates of cytokine release syndrome (CRS) and neurologic toxicities (NT) were 14% and 13%, respectively. Conclusion: Our study suggests that CAR-T therapy has demonstrated efficacy and safety in RRMM patients. BCMA-targeted CAR-T and anti-BCMA contained regimen have shown better efficacy., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

The dysregulation of cellular metabolism is a hallmark of ageing. To understand the metabolic changes that occur as a consequence of the ageing process and to find biomarkers for age-related diseases, we conducted metabolomic analyses of the brain, heart, kidney, liver, lung and spleen in young (9-10 weeks) and old (96-104 weeks) wild-type mice [mixed genetic background of 129/J and C57BL/6] using NMR spectroscopy. We found differences in the metabolic fingerprints of all tissues and distinguished several metabolites to be altered in most tissues, suggesting that they may be universal biomarkers of ageing. In addition, we found distinct tissue-clustered sets of metabolites throughout the organism. The associated metabolic changes may reveal novel therapeutic targets for the treatment of ageing and age-related diseases. Moreover, the identified metabolite biomarkers could provide a sensitive molecular read-out to determine the age of biologic tissues and organs and to validate the effectiveness and potential off-target effects of senolytic drug candidates on both a systemic and tissue-specific level.

The photodynamic therapy (PDT) as a promising antitumor therapy technique is greatly hampered by the low tissue penetration of light and the photothermal effect of prolonged irradiation. Near-infrared (NIR) persistent luminescence nanoparticles (NPLNPs) possess the potential for application in next-generation PDT. However, owing to the low re-excitation efficiency of NPLNPs in deep tissue, the current PDT nanoplatform based on NPLNPs is faced with the disadvantage of decreased PDT efficiency induced by persistent luminescence (PersL) decay at the lesion site. Herein, NPLNPs, Zn 1.3 Ga 1.4 Sn 0.3 O 4 :Cr 3+ (ZGS), with small particle sizes and excellent optical properties are synthesized via a simple acetylacetonate combustion method. The ZGS can be repeatedly excited by the biological window (659 nm) light to produce a strong NIR (700 nm) PersL. The response efficiency of ZGS to the wavelength in the biological window has been greatly improved by doping Sn 4+ into the ZnGa 2 O 4 matrix, which is 55 times higher than that of traditional ZnGa 2 O 4 :Cr 3+ . We further develop a PDT nanoplatform by modifying a photosensitizer on its surface. The PDT experiments show that the developed nanoplatform can achieve continuous and efficient tumor PDT with a depth of up to 3 cm by repeated excitation using a 659 nm LED. The NPLNPs largely solve the problem of the low re-excitation efficiency after NIR PersL decay of traditional NPLNPs in deep tissue applications and will further promote the application of NIR PLNPs in the biomedical field.

Endothelial lipase (EL) is a strong determinant of structural and functional properties of high-density lipoprotein (HDL). We examined whether the antioxidative capacity of HDL is affected by EL. EL-modified HDL (EL-HDL) and control EV-HDL were generated by incubation of HDL with EL- overexpressing or control HepG2 cells. As determined by native gradient gel electrophoresis, electron microscopy, and small-angle X-ray scattering EL-HDL is smaller than EV-HDL. Mass spectrometry revealed an enrichment of EL-HDL with lipolytic products and depletion of phospholipids and triacylglycerol. Kinetics of conjugated diene formation and HPLC-based malondialdehyde quantification revealed that EL-HDL exhibited a significantly higher resistance to copper ion-induced oxidation and a significantly higher capacity to protect low-density lipoprotein (LDL) from copper ion-induced oxidation when compared to EV-HDL. Depletion of the lipolytic products from EL-HDL abolished the capacity of EL-HDL to protect LDL from copper ion-induced oxidation, which could be partially restored by lysophosphatidylcholine enrichment. Proteomics of HDL incubated with oxidized LDL revealed significantly higher levels of methionine 136 sulfoxide in EL-HDL compared to EV-HDL. Chloramine T (oxidizes methionines and modifies free thiols), diminished the difference between EL-HDL and EV-HDL regarding the capacity to protect LDL from oxidation. In absence of LDL small EV-HDL and EL-HDL exhibited higher resistance to copper ion-induced oxidation when compared to respective large particles. In conclusion, the augmented antioxidative capacity of EL-HDL is primarily determined by the enrichment of HDL with EL-generated lipolytic products and to a lesser extent by the decreased HDL particle size and the increased activity of chloramine T-sensitive mechanisms., (Copyright © 2019 The Author(s). Published by Elsevier B.V. All rights reserved.)

Clinical outcomes of patients with multiple myeloma (MM) have almost doubled the overall survival over the last decade owing to the use of proteasome inhibitor such as bortezomib (BTZ). However, some patients with MM develop primary resistance to BTZ, whereas others develop resistance after treatment. In this study, we investigated relationships between BTZ resistance and dysfunction of long non-coding RNAs (lncRNAs) in patients with MM. Bone marrow samples were collected from patients with MM and healthy donors for lncRNA microarray and survival analyses. To investigate functions and underlying mechanisms of lncRNA-mediated BTZ resistance in MM, we performed CCK-8 assays, flow cytometry analyses, dual luciferase report gene assays, and RNA pulldown assays with samples from nude mice carrying tumor xenografts and in clinical samples. Differentially expressed lncRNA myocardial infarction associated transcripts (MIAT) were highly expressed in patients with MM compared with healthy controls, and were predictive of poor survival outcomes. Moreover, MIAT expression was significantly increased in BTZ-resistant patients with MM compared with newly diagnosed patients with MM, and was identified as a BTZ-inducible lncRNA. Specifically, BTZ upregulated MIAT expression through increased stat1 phosphorylation. Silencing of MIAT inhibited MM cell growth and sensitized MM cells to BTZ by negatively regulating miR-29b. Our data demonstrated the utility of MIAT as a tool for overcoming BTZ resistance in patients with MM.

The nanoplatform of synergistic chemo-photothermal therapy has superior advantages on antitumor. It is urgently needed to explore novel nanocarrier for improving photothermal performance in drug delivery process. Herein, we synthesized polydopamine doped mesoporous silica-coated reduced graphene oxide (rGO/MSN/PDA) by simply adding dopamine hydrochloride into the oil-water biphasic reaction system as a multifunctional drug carrier for anticancer treatment, which combines chemotherapy and photothermal therapy. The rGO/MSN/PDA showed nearly twice the photothermal conversion efficiency of mesoporous silica-coated graphene oxide (GO/MSN) due to the reduction of GO and doping with PDA. In addition, the rGO/MSN/PDA showed pH-response DOX release abilities, which means higher release of DOX in tumor cells. The cell experiments in vitro proved that rGO/MSN/PDA with better biocompatibility compare to GO/MSN might offer a promising tool for improving the therapeutic effects of hepatocellular carcinoma cells through synergistic chemo-photothermal therapy., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Sorafenib (So) is a multi-target kinase inhibitor extensively used in clinic for hepatocellular carcinoma therapy. It demonstrated strong inhibition both in tumor proliferation and tumor angiogenesis, while hampered by associated cutaneous side-effect and drug resistance. The knockdown of miR-21 with antisense oligonucleotides (antimiRNA21) was regarded as an efficient strategy for increasing tumor sensibility to chemotherapy, which could be employed to appreciate the efficacy of So. Herein, we successfully formulated a dual-targeting delivery system for enhanced hepatocellular carcinoma therapy by encapsulating So and antimiRNA21 in RGD pentapeptide-modified reconstituted high-density lipoprotein (RGD-rHDL/So/antimiRNA21). The RGD and apolipoprotein A-I (ApoA-I) on nanoparticles (NPs) could drive the system simultaneously to tumor neovascular and parenchyma by binding to the overexpressed ανβ3-integrin and SR-B1 receptors, achieving precise delivery of therapeutics to maximize the efficacy. A series in vitro and in vivo experiments revealed that co-delivery of So and antimiRNA21 by RGD-rHDL significantly strengthened the anti-tumor and anti-angiogenic effect of So with negligible toxicity towards major organs, reversed drug-resistance and was capable of remodeling tumor environments. The constructed RGD-rHDL/So/antimiRNA21 with improved efficacy and excellent tumor targeting ability provided new idea for chemo-gene combined therapy in hepatocellular carcinoma., Statement of Significance: Sorafenib (So) is a multi-target kinase inhibitor which was approved by FDA as first-line drug for hepatocellular carcinoma (HCC) therapy. However, long term application of So in clinic was hampered by serious dermal toxicity and drug resistance. Although numerous researchers were devoted to finding alternatives or therapies as combination treatments with So to reach more desired therapeutic efficacy, the therapeutic options were still limited. The present study prepares RGD pentapeptide decorated biomimic reconstituted high-density lipoprotein (rHDL) loaded with So and antimiRNA21 (RGD-rHDL/So/antimiRNA21) for enhanced HCC therapy. The RGD-rHDL/So/antimiRNA21 NPs offer an effective platform for anti-tumor and anti-angiogenesis therapy in HCC and provide new approach to reverse drug-resistance of So for feasible clinical application., (Copyright © 2018 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Increasing understanding in tumor pathology reveals that tumor microenvironment (TME), which supports tumor progression and poses barriers for available therapies, takes a great responsibility in inefficient treatment and poor prognosis. In recent years, the versatile nanotechnology employed in TME regulation has made great progress. The nanoparticles (NPs) can be tailored as needed to accurately target TME components by distinguishing healthy tissues from malignancy, and to regulate TME to promote tumor regression. Meanwhile, the emerging microRNAs (miRNAs) demonstrate great potentials for TME regulation, but are regrettably restricted by quick degradation. NPs systems enable the successful delivery of miRNA to TME without the limitation, expanding the application of nucleic acid drug. In this review, we summarized recent NPs-based strategies aiming at regulating TME in different ways, including anti-angiogenesis, extracellular matrix (ECM) remodeling, tumor-associated fibroblasts (TAFs) treatment and tumor-associated macrophages (TAMs) treatment, along with the miRNAs-loaded NPs for TME regulation. Catching and utilizing the features of TME for NPs design can contribute to reversing drug-resistance, optimized drug distribution, and eventually more efficient cancer therapy., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Drug resistance of bacteria has become a global health problem, as it makes conventional antibiotics less efficient. It is urgently needed to explore novel antibacterial materials and develop effective treatment strategies to overcome the drug resistance of antibiotics. Herein, we successfully synthesized silver decorated sandwich-like mesoporous silica/reduced graphene oxide nanosheets (rGO/MSN/Ag) as a novel antibacterial material through facile method. The rGO and Ag nanoparticles can be reduced in the reaction system without adding any other reductants. In addition, the rGO/MSN/Ag showed higher photothermal conversion capacity due to the modification of silver nanoparticles and exhibited excellent antibacterial activities against Pseudomonas putida, Escherichia coli and Rhodococcus at relatively low dosages, which was confirmed by the minimum inhibitory concentration (MIC) test. Meanwhile, the E. coli with a high concentration was selected for exposure using an 808 nm laser, and the antibacterial effect was obviously enhanced by the near-infrared irradiation induced photothermal effect. Moreover, the hepatocyte LO2 were used for the cytotoxicity evaluation, and the rGO/MSN/Ag showed low toxicity and were without detectable cytotoxicity at the antimicrobial dose. As the prepared rGO/MSN/Ag nanosheets have the advantages of low-cost and high antibacterial activity, they might be of promising and useful antibacterial agents for different applications.

Multi-walled carbon nanotube (MWNT) with its versatility has exhibited tremendous superiority in drug delivery. Despite plenty of researches on MWNT based delivery systems, precision-guided assistances to maximize their profitable properties are still lacking in substantive progress. We developed here a dual-targeting and co-delivery system based on MWNT for antiangiogenesis therapy in lung cancer which aimed at renin-angiotensin system (RAS) dysregulation by synergistically conducting angiotensin II type 1 receptor (AT 1 R) and type 2 receptor (AT 2 R) pathway. In this work, iRGD peptide connected to polyethyleneimine (PEI) was linked to MWNT skeleton, accompanying with candesartan (CD) conjugated to MWNT mediated by cystamine (SS). The functionalized MWNT is assembled with plasmid AT 2 (pAT 2 ) to form iRGD-PEI-MWNT-SS-CD/pAT 2 complexes. iRGD and CD act as pilots for complexes to dually target symbolic ανβ3-integrin and AT 1 R both overexpressed on tumor angiogenic endothelium and lung cancer cell. CD as chemotherapy showed synergistic downregulation of VEGF when combining of pAT 2 and efficiently inhibited angiogenesis. iRGD-PEI-MWNT-SS-CD/pAT 2 complexes greatly appreciated drug activities by changing drug distribution and exhibited remarkable tumor growth suppression in A549 xenograft nude mice. Our work presents that such dual-targeting strategy highly improves the delivery performance of MWNT and open a new avenue for RAS related lung cancer therapy., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Single-walled carbon nanotubes (SWNTs) with unique physicochemical properties have exhibited promising biomedical applications as drug and gene carriers. In this study, polyethylenimine (PEI)-modified SWNT conjugates linked with candesartan (CD) were developed to deliver vascular endothelial growth factor (VEGF)-targeted siRNA (siVEGF) for the synergistic and targeted treatment of tumor angiogenesis. The characterization results revealed that SWNT-PEI-CD conjugates were successfully synthesized and exhibited desirable dispersibility and superior stability. Confocal laser scanning microscopy (CLSM) and flow cytometry (FCM) results showed that SWNT-PEI-CD/siVEGF complexes could achieve high cellular uptake and specific intracellular distribution of siRNA in AT 1 R overexpressed PANC-1 cells. Strong down-regulation of VEGF was also verified by qualitative real-time polymerase chain reaction, enzyme-linked immunosorbent assay, and Western blot in complex-treated PANC-1 cells. The in vitro angiogenesis assay showed that SWNT-PEI-CD/siVEGF complexes highly inhibited tube formation of human umbilical vein endothelial cells. Furthermore, in vivo observation in PANC-1 xenografted nude mice demonstrated that SWNT-PEI-CD/siVEGF complexes exhibited significant distribution at tumor sites and caused obvious inhibition of tumor growth and tumor-associated angiogenesis repression induced by the drug combination of CD and siVEGF. Finally, a WST-1 assay indicated that SWNT-PEI-CD possessed low cytotoxicity, and a hemolysis test showed good biocompatibility of SWNT-PEI-CD. Hematological and histological analyses confirmed that SWNT-PEI-CD/siVEGF complexes did not cause any obvious toxic effects to blood and major organs. These findings suggested that the SWNT-PEI-CD/siVEGF co-delivery system with tumor-targeting specificity, improved endosomal escaping properties, and collaboration of angiogenesis inhibition could be a prospective method for efficient tumor antiangiogenic therapy.

1. Plasma concentrations of curcumin-O-glucuronide (COG) and curcumin-O-sulfate (COS) significantly increased after Sprague-Dawley rats dealt with the Oatp inhibitor rifampicin, with the C max ascending 2.9 and 6.7 times, and the AUC 0-∞ ascending 4.4 and 10.8 times, respectively. When pretreated with the Oat inhibitor probenecid, the C max increased 4.4 and 20 times, and the AUC 0-∞ increased 3.2 and 13.9 times, respectively. The results suggested that COG and COS may be the substrates of Oatp and Oat. 2. The accumulation of curcumin significantly increased in organic anion transporting polypeptide (OATP)- and organic anion transporter (OAT)-transfected human embryonic kidney (HEK) 293 systems, which suggested that curcumin was a substrate of OATP1B1, OATP1B3, OATP2B1, OAT1, and OAT3; and COG was a substrate of OATP1B1, OATP1B3, and OAT3. 3. Inhibition study using rosuvastatin as the substrate in OATP1B1- and OATP1B3-transfected cells indicated that curcumin was an OATP1B1 and 1B3 inhibitor, with IC 50 at 5.19 ± 0.05 and 3.68 ± 0.05 μM, respectively; the data for COG were 1.04 ± 0.01 and 1.08 ± 0.02 μM, respectively. COS was speculated to be an inhibitor of hepatic OATP1B1 as calculated using the ADMET Predictor. 4. COG and COS are substrates and inhibitors of OATP/Oatp. Co-administration of curcumin significantly increased rosuvastatin concentration in rat and dog plasma.

Multidrug resistance (MDR) is a major obstacle to cancer chemotherapy due to the overexpression of P-glycoprotein (P-gp). Herein, etoposide (ETO) was loaded onto oxidized carbon nanohorns (oxCNHs), which were modified by polyethylene glycol (PEG) and further functionalized with the targeting ligand P-gp monoclonal antibody (PA) in an attempt to overcome MDR. The obtained drug delivery system (ETO@oxCNHs/PEG-PA) showed high drug loading efficiency, enhanced drug release under laser irradiation, improved cellular uptake and increased therapeutic effect both in vitro and in vivo. In addition, NIR laser irradiation had a synergistic effect on overcoming MDR. The MDR-overcoming mechanism could be the efficient cellular uptake, enhanced drug release and reduced drug efflux by P-gp. These results demonstrated that ETO@oxCNHs/PEG-PA could be a promising drug delivery system for cancer MDR reversion.

In the past decades, reconstituted high density lipoprotein (rHDL) has been successfully developed as a drug carrier since the enhanced HDL-lipids uptake is demonstrated in several human cancers. In this paper, rHDL, for the first time, was utilized to co-encapsulate two hydrophobic drugs: an anticancer drug, paclitaxel (PTX), and a new reversal agent for P-gp (P-glycoprotein)-mediated multidrug resistance (MDR) of cancer, N-cyano-1-[(3,4-dimethoxyphenyl)methyl]-3,4-dihydro-6,7-dimethoxy-N'-octyl-2(1H)-isoquinoline-carboximidamide (HZ08). We proposed this drug co-delivery strategy to reverse PTX resistance. The study aimed to develop a biomimetic nanovector, reconstituted high density lipoprotein (rHDL), mediating targeted PTX-HZ08 delivery for cancer therapy. Using sodium cholate dialysis method, we successfully formulated dual-agent co-delivering rHDL nanoparticles (PTX-HZ08-rHDL NPs) with a typical spherical morphology, well-distributed size (~100nm), high drug encapsulation efficiency (approximately 90%), sustained drug release properties and exceptional stability even after storage for 1month or incubation in 10% fetal bovine serum (FBS) DMEM for up to 2days. Results demonstrated that PTX-HZ08-rHDL NPs significantly enhanced anticancer efficacy in vitro, including higher cytotoxicity and better ability to induce cell apoptosis against both PTX-sensitive and -resistant MCF-7 human breast cancer cell lines (MCF-7 and MCF-7/PTX cells). Mechanism studies demonstrated that these improvements could be correlated with increased cellular uptake of PTX mediated by scavenger receptor class B type I (SR-BI) as well as prolonged intracellular retention of PTX due to the HZ08 mediated drug-efflux inhibition. In addition, in vivo investigation showed that the PTX-HZ08-rHDL NPs were substantially safer, have higher tumor-targeted capacity and have stronger antitumor activity than the corresponding dosage of paclitaxel injection. These findings suggested that rHDL NPs could be an ideal tumor-targeted nanovector for simultaneous transfer of insoluble anticancer drug and drug resistance reversal agents. The PTX-HZ08-rHDL NPs co-delivery system might be a new promising strategy to overcome tumor drug resistance., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Mucuna pruriens, an ancient Indian herbal medicine containing levodopa, is widely used for Parkinson's disease. In order to simultaneously determine levodopa and 1,1-dimethyl-3-carboxy-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline (MD01) in rat plasma, an improved LC-MS/MS method was developed and validated for a pharmacokinetic study in rats orally administered levodopa or Mucuna pruriens extract (MPE). Elimination of matrix effect and improvement of extraction recovery were achieved through systematic optimization of reversed-phase and hydrophilic interaction chromatographic conditions together with sample clean-up procedures. A satisfactory chromatographic performance was obtained with a Thermo Aquasil C18 column (50 × 2.1 mm, 3 µm) using acetonitrile and water containing 0.2% formic acid as mobile phases. Futhermore, sodium metabisulfite and formic acid were used as stabilizers in neat solutions as well as rat plasma. The method was validated in a dynamic range of 20.0-10,000 ng/mL for levodopa and MD01; the intra- and inter-day precision and accuracy were acceptable. The method was successfully utilized to determine the levodopa level in plasma samples of rats administered levodopa or MPE. Pharmacokinetic results showed that an increase in the AUC of levodopa was observed in rats following oral administration of multiple doses of MPE. Copyright © 2016 John Wiley & Sons, Ltd., (Copyright © 2016 John Wiley & Sons, Ltd.)

Aim: Amitriptyline is a widely used tricyclic antidepressant, but the metabolic studies were conducted almost 20 years ago using high-performance liquid chromatography coupled with ultraviolet detector or radiolabeled methods., Results: First, multiple ion monitoring (MIM)- enhanced product ion (EPI) scan was used to obtain the diagnostic ions or neutral losses in human liver microsome incubations with amitriptyline. Subsequently, predicted multiple reaction monitoring (MRM)-EPI scan was used to identify the metabolites in human urine with the diagnostic ions or neutral losses. Finally, product ion filtering and neutral loss filtering were used as the data mining tools to screen metabolites. Consequently, a total of 28 metabolites were identified in human urine after an oral administration using LC-MS/MS., Conclusion: An integrated workflow using LC-MS/MS was developed to comprehensively profile the metabolites of amitriptyline in human urine, in which five N-acetyl-l-cysteine conjugates were characterized as tentative biomarkers for idiosyncratic toxicity.

Regulation of gene expression using p53 is a promising strategy for treatment of numerous cancers, and chemotherapeutic drug dichloroacetate (DCA) induces apoptosis and growth inhibition in tumor, without apparent toxicity in normal tissues. Combining DCA and p53 gene could be an effective way to treat tumors. The progress towards broad applications of DCA/p53 combination requires the development of safe and efficient vectors that target to specific cells. In this study, we developed a DSPE-PEG-AA (1,2-distearoryl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethyleneglycol-2000)] ammonium salt-anisamide) modified reconstituted high-density lipoprotein-based DCA/p53-loaded nanoparticles (DSPE-PEG-AA/rHDL/DCA-PEI/p53 complexes), which was fabricated as a drug/gene dual-targeting co-delivery system for potential cancer therapy. Here, DCA-PEI was utilized to effectively condense the p53 plasmid, to incorporate the plasmid into rHDL and to act as an antitumor drug to inhibit tumor cell growth. The DSPE-PEG-AA/rHDL/DCA-PEI/p53 complexes exhibited desirable and homogenous particle size, neutral surface charge and low cytotoxicity for normal cells in vitro. The results of confocal laser scanning microscopy (CLSM) and flow cytometry confirmed that the scavenger receptor class B type I (SR-BI) and sigma receptor mediated dual-targeting function of the complexes inducing efficient cytoplasmic drug delivery and gene transfection in human lung adenocarcinoma cell line A549. And in vivo investigation on nude mice bearing A549 tumor xenografts revealed that DSPE-PEG-AA/rHDL/DCA-PEI/p53 complexes possessed specific tumor targeting and strong antitumor activity. The work described here demonstrated that the DSPE-PEG-AA/rHDL/DCA-PEI/p53 complexes might offer a promising tool for effective cancer therapy., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Chemotherapy, a traditional method, plays an important role in tumor therapy. Currently, common clinical antitumor drugs have several defects like poor efficacy, side effects, etc. Furthermore, developing new antitumor drugs takes a long time and requires many resources. Recent studies have found that oldies are newbies for the oncologist, such as flavonoid, metformin, aspirin, etc. These non-traditional antitumor drugs (NTADs) are widely used in management of non-cancer diseases, which gained FDA approval for treatment of patients. Increasingly, studies about antitumor action of NTADs have attracted many researchers' interests. A giant amount of studies showed a decrease in cancer incidence in NTAD-treated patients. Several reports outlined a direct inhibitory effect of NTADs on cancer cell growth and antitumoral actions. This review summarized the research progress on antitumor effects of ten NTADs. Retrospective and meta-analyses of trials also showed that these NTADs had preventive effects against cancer in vitro and in vivo. These drugs represent a promising option for cancer treatment, which have clear benefits including clinical safety, obvious curative effect, and saving medical and health resources. Judged from previous reports, future studies will yield valuable data about the profitable effects of these drugs. With a better understanding of its mechanisms of antitumor activity, NTADs may become available for combination with chemotherapy or targeted therapy in clinic.

Background: MiRNAs has been shown to be implicated in the pathogenesis of many human diseases including cancer. Dysregulation of miR-744 is common in a number of cancers, indicating miR-774 might be closely correlated with the tumorigenesis process. However, the role and clinical significance of miR-774 in nasopharyngeal carcinoma (NPC) is poorly known. Thus the aim of this study is to investigate whether there was any clinical value of serum miR-744 in detecting and predicting the prognosis of NPC., Materials and Methods: Real-time PCR was used to examine the expression level of serum miR-744 in patients with NPC and the healthy volunteers. The changes in serum miR-744 expression level of NPC patients after receiving chemo-radiotherapy were also evaluated. The association between pre-treatment serum miR-744 expression level and NPC clinicopathological parameters was investigated. Finally we employed Kaplan-Meier method and Cox proportional hazards model to evaluate the clinical value of serum miR-744 in predicting the prognosis of NPC., Results: Our study showed the expression level of serum miR-744 was significant higher in patients with NPC in comparison with healthy controls (P<0.01). The serum miR-744 expression level was down-regulated significantly in NPC patients after receiving chemo-radiotherapy (P<0.01). The Pre-treatment Serum miR-744 expression level was correlated with various important NPC clinicopathological parameters including N stage, clinical stage and grade. In addition, NPC patients with higher serum miR-744 expression had poorer 5 year overall survival rate and relapse-free survival rate. What was more, serum miR-744 was showed to be an independent factor for predicting the prognosis of NPC., Conclusion: Serum miR-744 was up-regulated in NPC patients. Higher expression level of serum miR-744 was closely correlated with was associated with poor prognosis in NPC and it might be employed as a potential biomarker for predicting the clinical outcome of NPC patients.

The study is aimed to develop a versatile reticular polyethylenimine (PEI) derivative eprosartan-g-PEI (ESP) conjugate-mediated targeted drug and gene codelivery system for tumor therapy. Eprosartan (ES), an angiotensin II type 1 receptor blocker (ARB), which has been proven to exert beneficial effects on tumor progression, vascularization, and metastasis as the conventional antihypertensive drug, was conjugated with PEI-1.8K chains into ESP via a bis-amide bond of pH-sensitivity to overcome high cytotoxicity and nontargeted gene delivery of PEI-25K. P53 gene was encapsulated in the ESP to form the codelivery system of ESP/p53 complexes, and this system was comprehensively characterized. In vitro ESP/p53 complexes had a significant effect on inhibiting angiogenesis by reducing the expression and secretion of VEGF. In vivo the effective antitumor activity of ESP/p53 complexes was observed on nude mice bearing PANC-1 xenografts, and the microvessel density (MVD) examination demonstrated that ESP/p53 complex-produced antitumor efficacy was closely correlated with the efficient angiogenesis repression. These findings disclosed that the multifunctional ESP/p53 complexes might be a promising dual anticancer drug and gene codelivery system.

Overexpression of intracellular Notch plays an important role in the generation of human acute lymphoblastic T cell leukemia (T-ALL). In mouse models, it was shown that Notch-dependent T-ALL required pre-TCR signaling. Here we show that pre-TCR signaling is required to condition mice for Notch-dependent transformation but that it is not required to sustain malignant growth of T-ALL. In contrast to previous studies, we found that disease development does not require pre-TCR but that it can be accelerated in Rag2(-/-) mice by transient mimicking of pre-TCR signals.

MPI encodes phosphomannose isomerase, which interconverts fructose 6-phosphate and mannose 6-phosphate (Man-6-P), used for glycoconjugate biosynthesis. MPI mutations in humans impair protein glycosylation causing congenital disorder of glycosylation Ib (CDG-Ib), but oral mannose supplements normalize glycosylation. To establish a mannose-responsive mouse model for CDG-Ib, we ablated Mpi and provided dams with mannose to rescue the anticipated defective glycosylation. Surprisingly, although glycosylation was normal, Mpi(-/-) embryos died around E11.5. Mannose supplementation even hastened their death, suggesting that man-nose was toxic. Mpi(-/-) embryos showed growth retardation and placental hyperplasia. More than 90% of Mpi(-/-) embryos failed to form yolk sac vasculature, and 35% failed chorioallantoic fusion. We generated primary embryonic fibroblasts to investigate the mechanisms leading to embryonic lethality and found that mannose caused a concentration- and time-dependent accumulation of Man 6-P in Mpi(-/-) fibroblasts. In parallel, ATP decreased by more than 70% after 24 h compared with Mpi(+/+) controls. In cell lysates, Man-6-P inhibited hexokinase (70%), phosphoglucose isomerase (65%), and glucose-6-phosphate dehydrogenase (85%), but not phosphofructokinase. Incubating intact Mpi(-/-) fibroblasts with 2-[(3)H]deoxyglucose confirmed mannose-dependent hexokinase inhibition. Our results in vitro suggest that mannose toxicity in Mpi(-/-) embryos is caused by Man-6-P accumulation, which inhibits glucose metabolism and depletes intracellular ATP. This was confirmed in E10.5 Mpi(-/-) embryos where Man-6-P increased more than 10 times, and ATP decreased by 50% compared with Mpi(+/+) littermates. Because Mpi ablation is embryonic lethal, a murine CDG-Ib model will require hypomorphic Mpi alleles.