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Abstract Anti-DNA antibodies (Abs), serological hallmarks of systemic lupus erythematosus (SLE) and markers for diagnosis and disease activity, show a specificity for non-nucleic acid molecules, such as N-pyrrolated proteins (pyrP) containing N ε-pyrrole-L-lysine (pyrK) residues. However, the detailed mechanism for the binding of anti-DNA Abs to pyrP remains unknown. In the present study, to gain structural insights into the dual-specificity of anti-DNA Abs, we used phage display to obtain DNA-binding, single-chain variable fragments (scFvs) from SLE-prone mice and found that they also cross-reacted with pyrP. It was revealed that a variable heavy chain (VH) domain is sufficient for the recognition of DNA/pyrP. Identification of an antigenic sequence containing pyrK in pyrP suggested that the presence of both pyrK and multiple acidic amino acid residues plays important roles in the electrostatic interactions with the Abs. X-ray crystallography and computer-predicted simulations of the pyrK-containing peptide-scFv complexes identified key residues of Abs involved in the interaction with the antigens. These data provide a mechanistic insight into the molecular basis of the dual-specificity of the anti-DNA Abs and provide a basis for therapeutic intervention against SLE.

Background/Objectives: Brick dust molecules exhibit high melting points and ultralow solubility. Overcoming this solubility issue is challenging. Previously, we formulated a co-amorphous system for a neuropeptide Y5 receptor antagonist (NP) as a brick dust drug using sodium taurocholate (ST) to improve its dissolution profile. In this study, we have designed a ternary amorphous system involving polymer addition to further improve a co-amorphous system. Methods: The amorphous samples were prepared by the ball milling. The thermal and spectroscopic analyses were performed, and the isothermal crystallization and dissolution profiles were evaluated. Results: The ball milling of NPs, ST, and each of the three types of polymers successfully converted crystalline NPs to amorphous NPs. Thermal analysis confirmed the formation of a single amorphous phase. The infrared spectra revealed a specific interaction between an NP and ST in the co-amorphous system. Moreover, the intermolecular interactions of NP-ST were maintained in the ternary amorphous systems, suggesting the miscible dispersion of the co-amorphous system into the polymer via weak interactions as co-amorphous solid dispersions. The dissolution profile of co-amorphous NP-ST was 4.1- and 6.7-fold higher than that of crystalline NPs in pH 1.2 and 6.8 buffers, respectively. The drug concentration in the ternary amorphous system in pH 1.2 and 6.8 buffers became 1.1–1.2- and 1.4–2.7-fold higher than that seen in the co-amorphous system, respectively. Conclusions: Co-amorphous solid dispersion is a promising method for enhancing the solubility of brick dust molecules.

We introduce an quantum-classical hybrid algorithm, named automatic quantum circuit encoding (AQCE), that is designed to encode an arbitrary quantum state |Ψ〉 onto an optimal quantum circuit C[over ̂] composed of a finite set of single- and two-qubit quantum gates. The algorithm employs an objective function based on the absolute value of fidelity, F=〈0|C[over ̂]^{†}|Ψ〉, which is iteratively maximized to construct an optimal quantum circuit C[over ̂] with controlled accuracy. Here, |0〉 denotes a trivial product state in the computational basis of a quantum computer. The core of this algorithm lies in the sequential determination of an optimal set of two-qubit unitary operators, identified one by one through the singular value decomposition of the fidelity tensor. Once an optimal set of operators is determined, including the location of qubits on which each operator acts, elementary quantum gates are assigned algebraically to these two-qubit unitary operators. Importantly, these procedures are deterministic without assuming any quantum circuit ansatz and therefore eliminate the need for parameter optimization of parametrized quantum gates. Through noiseless numerical simulations, we demonstrate the effectiveness of the AQCE algorithm in encoding ground states of quantum many-body systems, including the spin-1/2 antiferromagnetic Heisenberg model and the spin-1/2 XY model. We also compare these results with quantum circuit encoding employing predefined circuit structures such as Trotter-like and MERA-like circuit ansatze. Furthermore, our algorithm extends to classical data, for instance, classical images represented as quantum states using amplitude encoding. The adaptability enables us to adjust the quantum resource requirement, i.e., the number of qubits, by partitioning classical data into multiple distinct segments. This feature holds potential for near-term applications in quantum machine learning, such as a state preparation of classical data for an input quantum state to be processed. Finally, using a real quantum device provided by IBM Quantum, we experimentally validate that a quantum circuit generated by the AQCE algorithm can indeed reasonably represent the original quantum state.

Tensor network (TN) states, including entanglement renormalization (ER), can encompass a wider variety of entangled states. When the entanglement structure of the quantum state of interest is nonuniform in real space, accurately representing the state with a limited number of degrees of freedom hinges on appropriately configuring the TN to align with the entanglement pattern. However, a proposal has yet to show a structural search of ER due to its high computational cost and the lack of flexibility in its algorithm. In this study, we conducted an optimal structural search of TN, including ER, based on the reconstruction of their local structures with respect to variational energy. First, we demonstrated that our algorithm for the spin-1/2 tetramer singlets model could calculate exact ground energy using the multiscale entanglement renormalization ansatz (MERA) structure as an initial TN structure. Subsequently, we applied our algorithm to the random XY models with the two initial structures: MERA and the suitable structure underlying the strong disordered renormalization group. We found that, in both cases, our algorithm achieves improvements in variational energy, fidelity, and entanglement entropy. The degree of improvement in these quantities is superior in the latter case compared to the former, suggesting that utilizing an existing TN design method as a preprocessing step is important for maximizing our algorithm's performance.

Polymorphic control is vital for the quality control of pharmaceutical crystals. Here, we investigated the relationship between the hydrate and anhydrate polymorphs of a monoacylglycerol acyltransferase 2 inhibitor (S-309309). Solvent evaporation and slurry conversion revealed two polymorphs, the hydrate and the solvate. The solvate was transformed into the hydrate by heating. X-ray powder diffraction demonstrated that the hydrate was transformed into an anhydrate via an intermediate state when heated. These crystal forms were confirmed under controlled humidity conditions; the presence of the anhydrate, the intermediate hydrate, or the hydrate depended on the relative humidity at 25 °C. The stoichiometry of S-309309 in water in the hydrate form was 4:1. The hydrates and anhydrates exhibited similar crystal structures and stability. The water of hydration in the intermediate hydrate was 0.1–0.15 mol according to the dynamic vapor sorption profile. The stability and dissolution profile of the anhydrate and hydrate showed no significant change due to similar crystal lattices and quick rehydration of the anhydrate. A mechanism for the reversible crystal transformation between the anhydrate and pseudo-polymorphs of the hydrate was discovered. We concluded that S-309309 causes a pseudo-polymorphic transformation; however, this is not a critical issue for pharmaceutical use.

In this paper, we introduce a tensor network (TN) scheme into the entanglement augmentation process of the synergistic optimization framework by Rudolph et al. [arXiv:2208.13673] to build its process systematically for inhomogeneous systems. Our synergistic approach first embeds the variational optimal solution of the TN state with the entropic area law, which can be perfectly optimized in conventional (classical) computers, in a quantum variational circuit ansatz inspired by the TN state with the entropic volume law. Next, the framework performs a variational quantum eigensolver (VQE) process with embedded states as the initial state. We applied the synergistic to the ground-state analysis of the all-to-all coupled random transverse-field Ising, XYZ, Heisenberg model, employing the binary multiscale entanglement renormalization ansatz (MERA) state and branching MERA states as TN states with entropic area law and volume law, respectively. We then show that the synergistic accelerates VQE calculations in the three models without an initial parameter guess of the branching-MERA-inspired ansatz and can avoid a local solution trapped by a standard VQE with the ansatz in the Ising model. The improvement of optimizers for MERA in all-to-all coupled inhomogeneous systems, enhancement, and potential synergistic applications are also discussed.

Abstract Since late 2019, the outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and the resultant spread of COVID-19 have given rise to a worldwide health crisis that is posing great challenges to public health and clinical treatment, in addition to serving as a formidable threat to the global economy. To obtain an effective tool to prevent and diagnose viral infections, we attempted to obtain human antibody fragments that can effectively neutralize viral infection and be utilized for rapid virus detection. To this end, several human monoclonal antibodies were isolated by bio-panning a phage-displayed human antibody library, Tomlinson I. The selected clones were demonstrated to bind to the S1 domain of the spike glycoprotein of SARS-CoV-2. Moreover, clone A7 in Fab and IgG formats were found to effectively neutralize the binding of S protein to angiotensin-converting enzyme 2 in the low nM range. In addition, this clone was successfully converted to quench-based fluorescent immunosensors (Quenchbodies) that allowed antigen detection within a few minutes, with the help of a handy fluorometer.

A red fluorescent protein-based indicator of cyclic guanosine 3′, 5′-monophosphate (cGMP) allows for multi-colour imaging and optogenetic applications while monitoring cGMP activity in cells.

Background and purpose: Physicochemical properties of an amorphous solid dispersion (ASD) comprising an experimental grade of hydroxypropyl methylcellulose acetate succinate (HPMCAS-MX) with lower glass transition temperature have been previously investigated. This study aimed to evaluate applicability of HPMCAS-MX to hot-melt extrusion (HME) and dissolution-permeation performance of prepared ASDs using MicroFLUX. Review approach: A physical mixture of indomethacin (IMC) and HPMCAS-MX or -MG (a commercial grade with higher transition temperature) at 20:80 weight ratio was hot-melt extruded to prepare an ASD (IMC-MX and IMC-MG, respectively). The dissolution-permeation performance and the stability of the ASDs were measured. Key results: A torque reduction at 120 °C implied that IMC-MX transformed into an amorphous state at this temperature, but IMC-MG required around 170 °C. This result was supported by Raman mapping of the the HME samples. IMC-MG and IMC-MX remained in an amorphous state at 40 °C for three months. The initial dissolution rate and solubility of the ASDs were higher than that of crystalline IMC. The apparent permeability of IMC from IMC-MX and IMC-MG was comparable but was approximately two-fold higher than that from crystalline IMC. Conclusion: HPMCAS-MX enabled HME process at a lower temperature and improved the dissolution-permeation performance of indomethacin.

Fibromyalgia (FM) is a chronic pain condition characterized by widespread musculoskeletal pain and other frequent symptoms such as fatigue, sleep disturbance, cognitive impairment, and mood disorder. Based on the view that intermittent stress would be the most probable etiology for FM, intermittent cold- and intermittent psychological stress-induced generalized pain (ICGP and IPGP) models in mice have been developed and validated as FM-like pain models in terms of the patho-physiological and pharmacotherapeutic features that are shared with clinical versions. Both models show long-lasting and generalized pain and female-predominant sex differences after gonadectomy. Like many other neuropathic pain models, ICGP and IPGP were abolished in lysophosphatidic acid receptor 1 (LPAR1) knock-out mice or by LPAR1 antagonist treatments, although deciding the clinical importance of this mechanism depends on waiting for the development of a clinically available LPAR1 antagonist. On the other hand, the nonsteroidal anti-inflammatory drug diclofenac with morphine did not suppress hyperalgesia in these models, and this is consistent with the clinical findings. Pharmacological studies suggest that the lack of morphine analgesia is associated with opioid tolerance upon the stress-induced release of endorphins and subsequent counterbalance through anti-opioid NMDA receptor mechanisms. Regarding pharmacotherapy, hyperalgesia in both models was suppressed by pregabalin and duloxetine, which have been approved for FM treatment in clinic. Notably, repeated treatments with mirtazapine, an α2 adrenergic receptor antagonist-type antidepressant, and donepezil, a drug for treating Alzheimer’s disease, showed potent therapeutic actions in these models. However, the pharmacotherapeutic treatment should be carried out 3 months after stress, which is stated in the FM guideline, and many preclinical studies, such as those analyzing molecular and cellular mechanisms, as well as additional evidence using different animal models, are required. Thus, the ICGP and IPGP models have the potential to help discover and characterize new therapeutic medicines that might be used for the radical treatment of FM, although there are several limitations to be overcome.

Development of a rapid detection method for deoxycholic acid (DCA) is crucial for its diagnosis in the early stages of inflammation and cancer. In this study, we expressed a soluble recombinant anti-DCA single-chain variable fragment (scFv) in Escherichia coli. To convert scFv into a Quenchbody (Q-body), we labeled scFv using commercially available maleimide-linked fluorophores. The TAMRA-C5-maleimide-conjugated Q-body showed the highest response within a few minutes of DCA addition, indicating its applicability as a wash-free immunoassay probe for onsite DCA detection.

Abstract Quenchbody (Q-body) is a quench-based fluorescent immunosensor labeled with fluorescent dye(s) near the antigen-binding site of an antibody. Q-bodies can detect a range of target molecules rapidly and directly. However, because Q-bodies show different antigen responses depending on the antibody used, time-consuming optimization of the Q-body structure is often necessary, and a high-throughput screening method for discriminating and selecting good Q-bodies is required. Here, we aimed to develop a molecular display method of nanobody-based “mini Q-bodies” by combining yeast surface display and coiled-coil forming E4/K4 peptide-based fluorescence labeling. As a result, the yeast-displayed mini Q-body recognizing the anti-cancer agent methotrexate (MTX) showed significant quenching and MTX-dependent dequenching on cells. To demonstrate the applicability of the developed method to select highly responsive mini Q-bodies, a small nanobody library consisting of 30 variants that recognize human serum albumin was used as a model. The best variant, showing a 2.4-fold signal increase, was obtained through selection by flow cytometry. Furthermore, the same nanobody prepared from Escherichia coli also worked as a mini Q-body after dye labeling. The described approach will be applied to quickly obtain well-behaved Q-bodies and other fluorescent biosensors for various targets through directed evolutionary approaches.

Nuclear protein prothymosin α (ProTα) is a unique member of damage-associated molecular patterns (DAMPs)/alarmins. ProTα prevents neuronal necrosis by causing a cell death mode switch in serum-starving or ischemic/reperfusion models in vitro and in vivo. Underlying receptor mechanisms include Toll-like receptor 4 (TLR4) and Gi-coupled receptor. Recent studies have revealed that the mode of the fatal stress-induced extracellular release of nuclear ProTα from cortical neurons in primary cultures, astrocytes and C6 glioma cells has two steps: ATP loss-induced nuclear release and the Ca2+-mediated formation of a multiple protein complex and its extracellular release. Under the serum-starving condition, ProTα is diffused from the nucleus throughout the cell due to the ATP loss-induced impairment of importin α–mediated nuclear transport. Subsequent mechanisms are all Ca2+-dependent. They include the formation of a protein complex with ProTα, S100A13, p40 Syt-1 and Annexin A2 (ANXA2); the fusion of the protein complex to the plasma membrane via p40 Syt-1–Stx-1 interaction; and TMEM16F scramblase-mediated ANXA2 flop-out. Subsequently, the protein complex is extracellularly released, leaving ANXA2 on the outer cell surface. The ANXA2 is then flipped in by a force of ATP8A2 activity, and the non-vesicular release of protein complex is repeated. Thus, the ANXA2 flop-out could play key roles in a new type of non-vesicular and non-classical release for DAMPs/alarmins, which is distinct from the modes conducted via gasdermin D or mixed-lineage kinase domain-like pseudokinase pores.

Abstract Although Raman spectroscopy has been used for the quantitative analysis of samples in many fields, including material science, biomedical, and pharmaceutical research, its low sensitivity hindered the application of the analytical capability for high-throughput screening. Here, we developed a high-throughput Raman screening system that can analyze hundreds of specimens in a multiwell plate simultaneously. Multiple high numerical aperture (NA) lenses are assembled under each well in the multiwell plate to detect Raman scattering simultaneously with high sensitivity. The Raman spectrum of 192 samples loaded on a standard 384-well plate can be analyzed simultaneously. With the developed system, the throughput of Raman measurement was significantly improved (about 100 times) compared to conventional Raman instruments based on a single-point measurement. By using the developed system, we demonstrated high-throughput Raman screening to investigate drug polymorphism and identify a small-molecule binding site in a protein. Furthermore, the same system was used to demonstrate high-speed chemical mapping of a centimeter-sized pork slice.

Abstract Background Drug-induced pneumonia (d-pneumonia) and bacterial pneumonia (b-pneumonia) are often difficult to differentiate; therefore, this study examined the possibility of differentiating them using serum biomarkers. Methods The study included 22 and 16 patients diagnosed with b- and d-pneumonia, respectively, at our institution or affiliated institutions. For d-pneumonia, the causative drug was minocycline hydrochloride in four patients, gefitinib in two patients, nivolumab in two patients, pembrolizumab in two patients, sulfasalazine in two patients, loxoprofen in one patient, Bouiougitou in one patient, edoxaban tosilate hydrate in one patient, and abemaciclib in one patient. White blood cell (WBC), C-reactive protein (CRP), Krebs von den Lungen-6 (KL-6), surfactant protein (SP)-D, and SP-A levels were measured in each patient and compared between the groups. Results Significant differences were noted in the WBC and SP-D levels between the two groups (P

Students’ affective states describe their engagement, concentration, attitude, motivation, happiness, sadness, frustration, off-task behavior, and confusion level in learning. In online learning, students’ affective states are determinative of the learning quality. However, measuring various affective states and what influences them is exceedingly challenging for the lecturer without having real interaction with the students. Existing studies primarily use self-reported data to understand students’ affective states, while this paper presents a novel learning analytics system called MOEMO (Motion and Emotion) that could measure online learners’ affective states of engagement and concentration using emotion data. Therefore, the novelty of this research is to visualize online learners’ affective states on lecturers’ screens in real-time using an automated emotion detection process. In real-time and offline, the system extracts emotion data by analyzing facial features from the lecture videos captured by the typical built-in web camera of a laptop computer. The system determines online learners’ five types of engagement (“strong engagement”, “high engagement”, “medium engagement”, “low engagement”, and “disengagement”) and two types of concentration levels (“focused” and “distracted”). Furthermore, the dashboard is designed to provide insight into students’ emotional states, the clusters of engaged and disengaged students’, assistance with intervention, create an after-class summary report, and configure the automation parameters to adapt to the study environment.

Antibody applications in cancer immunotherapy involve diverse strategies, some of which redirect T cell-mediated immunity via engineered antibodies. Affinity is a trait that is crucial for these strategies, as optimal affinity reduces unwanted side effects while retaining therapeutic function. Antibody-antigen pairs possessing a broad affinity range are required to define optimal affinity and to investigate the affinity-associated functional profiles of T cell-engaging strategies such as bispecific antibodies and chimeric antigen receptor-engineered T cells. Here, we demonstrate the unique binding characteristic of the developed antibody clone MVR, which exhibits robust binding to B-lymphoid cell lines. Intriguingly, MVR specifically recognizes the highly polymorphic human leukocyte antigen (HLA)-DR complex and exhibits varying affinities that are dependent upon the HLA-DRB1 allele type. Remarkably, MVR binds to the conformational epitope that consists of two hypervariable regions. As an application of MVR, we demonstrate an MVR-engineered chimeric antigen receptor (CAR) that elicits affinity-dependent function in response to a panel of target cell lines that express different HLA-DRB1 alleles. This tool evaluates the effect of affinity on cytotoxic killing, polyfunctionality, and activation-induced cell death of CAR-engineered T cells. Collectively, MVR exhibits huge potential for the evaluation of the affinity-associated profile of T cells that are redirected by engineered antibodies.

The inhibition of retinal ischemia-induced damage by post-ischemic prothymosin alpha (ProTα) was not affected in toll-like receptor 4 knockout (TLR4−/−) mice but blocked by the pretreatment with antibody against F0/F1 ATPase α- or β-subunit, novel candidate for ProTα-receptor. In addition to the previous observation of ProTα-induced ATP release from cells, the present study showed a ProTα-induced enhancement of ATP hydrolysis activity of recombinant ATP5A1/5B complex. As the protection of retinal function by post-ischemic ProTα was abolished by anti-P2Y12 antibody, the activation of F0/F1 ATPase and subsequent P2Y12 receptor system may play roles in beneficial actions by post-ischemic ProTα.

Quenchbodies (Q-bodies), a type of biosensor, are antibodies labeled with a fluorescent dye near the antigen recognition site. In the absence of an antigen, the dye is quenched by tryptophans in the antibody sequence; however, in its presence, the dye is displaced and therefore de-quenched. Although scFv and Fab are mainly used to create Q-bodies, this is the first report where a single-domain heavy chain VH from a semi-synthetic human antibody library formed the basis. To create a proof of concept “mini Q-body”, a human anti-lysozyme single-domain VH antibody C3 was used. Mini Q-bodies were successfully developed using seven dyes. Different responses were observed depending on the dye and linker length; it was concluded that the optimal linker length for the TAMRA dye was C5, and rhodamine 6G was identified as the dye with the largest de-quenching response. Three single-domain antibodies with sequences similar to that of the C3 antibody were chosen, and the results confirmed the applicability of this method in developing mini Q-bodies. In summary, mini Q-bodies are an easy-to-use and time-saving method for detecting proteins.

Prothymosin alpha (ProTα) was discovered to be a necrosis inhibitor from the conditioned medium of a primary culture of rat cortical neurons under starved conditions. This protein carries out a neuronal cell-death-mode switch from necrosis to apoptosis, which is, in turn, suppressed by a variety of neurotrophic factors (NTFs). This type of NTF-assisted survival action of ProTα is reproduced in cerebral and retinal ischemia–reperfusion models. Further studies that used a retinal ischemia–reperfusion model revealed that ProTα protects retinal cells via ecto-F1 ATPase coupled with the Gi-coupled P2Y12 receptor and Toll-like receptor 4 (TLR4)/MD2 coupled with a Toll–IL-1 receptor domain-containing adaptor inducing IFN-β (TRIF). In cerebral ischemia–reperfusion models, ProTα has additional survival mechanisms via an inhibition of matrix metalloproteases in microglia and vascular endothelial cells. Heterozygous or conditional ProTα knockout mice show phenotypes of anxiety, memory learning impairment, and a loss of neurogenesis. There are many reports that ProTα has multiple intracellular functions for cell survival and proliferation through a variety of protein–protein interactions. Overall, it is suggested that ProTα plays a key role as a brain guardian against ischemia stress through a cell-death-mode switch assisted by NTFs and a role of neurogenesis.

The tree tensor network (TTN) provides an essential theoretical framework for the practical simulation of quantum many-body systems, where the network structure defined by the connectivity of the isometry tensors plays a crucial role in improving its approximation accuracy. In this paper, we propose a TTN algorithm that enables us to automatically optimize the network structure by local reconnections of isometries to suppress the bipartite entanglement entropy on their legs. The algorithm can be seamlessly implemented to such a conventional TTN approach as the density-matrix renormalization group. We apply the algorithm to the inhomogeneous antiferromagnetic Heisenberg spin chain, having a hierarchical spatial distribution of the interactions. We then demonstrate that the entanglement structure embedded in the ground state of the system can be efficiently visualized as a perfect binary tree in the optimized TTN. Possible improvements and applications of the algorithm are also discussed.

Brick dust molecules are usually poorly soluble in water and lipoidal components, making it difficult to formulate them in dosage forms that provide efficient pharmacological effects. A co-amorphous system is an effective strategy to resolve these issues. However, their glass transition temperatures (Tg) are relatively lower than those of polymeric amorphous solid dispersions, suggesting the instability of the co-amorphous system. This study aimed to formulate a stable co-amorphous system for brick dust molecules by utilizing sodium taurocholate (NaTC) with a higher Tg. A novel neuropeptide Y5 receptor antagonist (AntiY5R) and NaTC with Tg of 155 °C were used as the brick dust model and coformer, respectively. Ball milling formed a co-amorphous system for AntiY5R and NaTC (AntiY5R-NaTC) at various molar ratios. Deviation from the theoretical Tg value and peak shifts in Fourier-transform infrared spectroscopy indicated intermolecular interactions between AntiY5R and NaTC. AntiY5R-NaTC at equal molar ratios resulting in an 8.5-fold increase in AntiY5R solubility over its crystalline form. The co-amorphous system remained amorphous for 1 month at 25 °C and 40 °C. These results suggest that the co-amorphous system formed by utilizing NaTC as a coformer could stably maintain the amorphous state and enhance the solubility of brick dust molecules.

The protein–protein interaction assay is a key technology in various fields, being applicable in drug screening as well as in diagnosis and inspection, wherein the stability of assays is important. In a previous study, we developed a unique protein–protein interaction assay “FlimPIA” based on the functional complementation of mutant firefly luciferases (Fluc). The catalytic step of Fluc was divided into two half steps: D-luciferin was adenylated in the first step, while adenylated luciferin was oxidized in the second step. We constructed two mutants of Fluc from Photinus pyralis (Ppy); one mutant named Donor is defective in the second half reaction, while the other mutant named Acceptor exhibited low activity in the first half reaction. To date, Ppy has been used in the system; however, its thermostability is low. In this study, to improve the stability of the system, we applied Fluc from thermostabilized Luciola lateralis to FlimPIA. We screened suitable mutants as probes for FlimPIA and obtained Acceptor and Donor candidates. We detected the interaction of FKBP12-FRB with FlimPIA using these candidates. Furthermore, after the incubation of the probes at 37°C for 1 h, the luminescence signal of the new system was 2.4-fold higher than that of the previous system, showing significant improvement in the stability of the assay.

Abstract A fundamental but controversial question in information coding of moving visual target is which of ‘motion’ or ‘position’ signal is employed in the brain for producing quick motor reactions. Prevailing theory assumed that visually guided reaching is driven always via target position representation influenced by various motion signals (e.g., target texture and surroundings). To rigorously examine this theory, we manipulated the nature of the influence of internal texture motion on the position representation of the target in reaching correction tasks. By focusing on the difference in illusory position shift of targets with the soft- and hard-edges, we succeeded in extracting the temporal development of an indirect effect only ascribed to changes in position representation. Our data revealed that the onset of indirect effect is significantly slower than the adjustment onset itself. This evidence indicates multi-stream processing in visuomotor control: fast and direct contribution of visual motion for quick action initiation, and relatively slow contribution of position representation updated by relevant motion signals for continuous action regulation. The distinctive visuomotor mechanism would be crucial in successfully interacting with time-varying environments in the real world.

Abstract Background Lysophosphatidic acid (LPA), generated by autotaxin (ATX), is a bioactive lipid mediator that binds to the receptors (LPA1–6), and serves as an important mediator in inflammation. Previous studies have demonstrated that LPA-LPA1 cascade contributes to arthritis and skin sclerosis. In this study, we examined the role of LPA signals in murine Candida albicans water-soluble fraction (CAWS)-induced vasculitis. Methods ATX and LPA receptor expressions were analyzed by immunohistochemistry and quantitative reverse transcription-polymerase chain reaction. Effects of LPA1 inhibition on CAWS-induced vasculitis were evaluated in LPA1-deficient mice or using an LPA1 antagonist, LA-01. Migration activity was assessed using a chemotaxis chamber. The number of migrated fluorescently labeled neutrophils, which were transferred into the vasculitis mice, was counted in the aortic wall. CXCL1 and IL-8 concentrations were determined by enzyme-linked immunosorbent assay. Results ATX and LPA1 were highly expressed in the inflamed region of CAWS-induced vasculitis. Severity of the vasculitis in LPA1-deficient mice was suppressed. The LPA1 antagonist, LA-01, also ameliorated the CAWS-induced vasculitis. LPA induced neutrophil migration, which was inhibited by LA-01 in vitro. Infiltration of transferred neutrophils from LPA1-deficient mice into the coronary arteries was suppressed. LA-01 also inhibited the infiltration of wild-type neutrophils. Expression of CXCL1 and IL-8 in human endothelial cells was enhanced by LPA, but was inhibited by LA-01. ATX and LPA1 expression levels were higher in the affected skin region of vasculitis patients than in healthy controls. Conclusions These results suggest that LPA-LPA1 signaling contributes to the development of vasculitis via chemoattractant production from endothelial cells followed by neutrophil recruitment. Thus, LPA1 has potential as a novel target for vasculitis therapies.

Abstract It is a common practice to issue a summary of a learner’s learning achievements in form of a transcript or certificate. However, detailed information on the depth of learning and how learning or teachings were conducted is not present in the transcript of scores. This work presents the first practical implementation of a new platform for keeping track of learning achievements beyond transcripts and certificates. This is achieved by maintaining digital hashes of learning activities and managing access rights through the use of smart contracts on the blockchain. The blockchain of learning logs (BOLL) is a platform that enable learners to move their learning records from one institution to another in a secure and verifiable format. This primarily solves the cold-start problem faced by learning data analytic platforms when trying to offer personalized experience to new learners. BOLL enables existing learning data analytic platforms to access the learning logs from other institutions with the permission of the learners and/or institution who originally have ownership of the logs. The main contribution of this paper is to investigate how learning records could be connected across institutions using BOLL. We present an overview of how the implementation has been carried out, discuss resource requirements, and compare the advantages BOLL has over other similar tools.

Tyrosine-arginine (kyotorphin), an opioid analgesic dipeptide, was discovered more than 40 years ago in 1979. The evidence accumulated during this period has established the physiological significance of kyotorphin as a neuromodulating peptide, and pharmacological applications. Some of the following important findings have been discussed in this review: (1) kyotorphin is unevenly distributed in the brain; it is found in high concentrations in the pain pathway, which involves the regions associated with morphine analgesia; (2) kyotorphin is subcellularly localized in the synaptosome fraction or nerve-ending particles; (3) a specific synthetase generates kyotorphin from tyrosine and arginine; (4) kyotorphin may be also processed from calpastatin by a novel calcium-activated neutral protease or calpain; (5) kyotorphin preloaded into the synaptosome is released by high K+ depolarization in a Ca2+-dependent manner; (6) kyotorphin has a specific G protein coupled receptor, which mediates the activation of phospholipase C (PLC) and inhibition of adenylyl cyclase through Gi; (7) leucine-arginine works as a specific kyotorphin receptor antagonist; 8) membrane-bound aminopeptidase or excretion through a peptide transporter, PEPT2, may contribute to the inactivation of kyotorphin; and (9) kyotorphin causes increased Met-enkephalin release from brain and spinal slices. It is also known that the opening of plasma membrane Ca2+ channels through a conformational coupling of the InsP3 receptor with the transient receptor potential C1, which is downstream of the kyotorphin receptor-mediated activation of Gi and PLC, could be a potential underlying mechanism of Met-enkephalin release. Considering these findings, translational research is an exciting domain that can be explored in the future. As kyotorphin is a small molecule, we could design function-added kyotorphin derivatives. These studies would include not only the brain-permeable kyotorphin derivatives but also hybrid kyotorphin derivatives conjugated with small compounds that have additional pharmacological actions. Further, since there are reports of kyotorphin being involved in either the etiology or treatment of Alzheimer's disease, epilepsy, inflammation, and chronic pain, studies on the beneficial effects of kyotorphin derivatives should also be expected in the future.

Fibromyalgia (FM), a disease of unknown etiology characterized by chronic generalized pain, is partly recapitulated in an animal model induced by repeated acid saline injections into the gastrocnemius muscle. Here, we attempted to investigate the sex difference in pain hypersensitivity (mechanical allodynia and hypersensitivity to electrical stimulation) in the repeated acid saline-induced FM-like generalized pain (AcGP) model. The first unilateral acid injection into gastrocnemius muscle at day 0/D0 and second injection at D5 (post day 0, P0) induced transient and long-lasting mechanical allodynia, respectively, on both sides of male and female mice. The pretreatment with gonadectomy did not affect the first injection-induced allodynia in both sexes, but gradually reversed the second injection-induced allodynia in male but not female mice. Moreover, the AcGP in male mice was abolished by intracerebroventricular minocycline treatments during D4–P4 or P5–P11, but not by early treatments during D0–D5 in male but not female mice, suggesting that brain microglia are required for AcGP in late-onset and sex-dependent manners. We also found that the intravenous treatments of splenocytes derived from male but not female mice treated with AcGP caused allodynia in naive mice. In addition, the purified CD4+ T cells derived from splenocytes of acid-treated male mice retained the ability to cause allodynia in naive mice. These findings suggest that FM-like AcGP has multiple sexual dimorphic mechanisms.

We propose a divide-and-conquer method for the quantum-classical hybrid algorithm to solve larger problems with small-scale quantum computers. Specifically, we concatenate a variational quantum eigensolver (VQE) with a reduction in the system dimension, where the interactions between divided subsystems are taken as an effective Hamiltonian expanded by the reduced basis. Then the effective Hamiltonian is further solved by the VQE, which we call deep VQE. Deep VQE allows us to apply quantum-classical hybrid algorithms on small-scale quantum computers to large systems with strong intrasubsystem interactions and weak intersubsystem interactions, or strongly correlated spin models on large regular lattices. As proof-of-principle numerical demonstrations, we use the proposed method for quasi-one-dimensional models, including one-dimensionally coupled 12-qubit Heisenberg antiferromagnetic models on kagome lattices as well as two-dimensional Heisenberg antiferromagnetic models on square lattices. The largest problem size of 64 qubits is solved by simulating 20-qubit quantum computers with a reasonably good accuracy approximately a few %. The proposed scheme enables us to handle the problems of >1000 qubits by concatenating VQEs with a few tens of qubits. While it is unclear how accurate ground-state energy can be obtained for such a large system, our numerical results on a 64-qubit system suggest that deep VQE provides a good approximation (discrepancy within a few percent) and has room for further improvement. Therefore, deep VQE provides us a promising pathway to solve practically important problems on noisy intermediate-scale quantum computers.

We investigated the visuomotor strategies of baseball batting, in particular, the relationship between eye and body (head and hip) movements during batting for a wide range of ball speeds. Nine college baseball players participated in the experiment and hit balls projected by a pitching machine operating at four different ball speeds (80, 100, 120, 140 km/h). Eye movements were measured with a wearable eye tracker, and body movements were measured with an optical motion capture system. In the early period of the ball's flight, batters foveated the ball with overshooting head movements in the direction of the ball's flight while compensating for the overshooting head movements with eye movements for the two slower ball speeds (80 and 100 km/h) and only head rotations for the two faster ball speeds (120 and 140 km/h). After that, batters made a predictive saccade and a quick head rotation to the future ball position before the angular velocity of the ball drastically increased. We also found that regardless of the ball speed, the onsets of the predictive saccade and the quick head movement were temporally aligned with the bat-ball contact and rotation of the hip (swing motion), but were not correlated with the elapsed time from the ball's release or the ball's location. These results indicate that the gaze movements in baseball batting are not solely driven by external visual information (ball position or velocity) but are determined in relation to other body movements.

Homozygosity of the p.Arg204Trp variation in the Pleckstrin homology and RhoGEF domain containing G2 (PLEKHG2) gene, which encodes a Rho family-specific guanine nucleotide-exchange factor, is responsible for microcephaly with intellectual disability. However, the role of PLEKHG2 during neurodevelopment remains unknown. In this study, we analyzed mouse Plekhg2 function during cortical development, both in vitro and in vivo. The p.Arg200Trp variant in mouse (Plekhg2-RW), which corresponds to the p.Arg204Trp variant in humans, showed decreased guanine nucleotide-exchange activity for Rac1, Rac3, and Cdc42. Acute knockdown of Plekhg2 using in utero electroporation-mediated gene transfer did not affect the migration of excitatory neurons during corticogenesis. On the other hand, silencing Plekhg2 expression delayed dendritic arbor formation at postnatal day 7 (P7), perhaps because of impaired Rac/Cdc42 and p21-activated kinase 1 signaling pathways. This phenotype was rescued by expressing an RNAi-resistant version of wildtype Plekhg2, but not of Plekhg2-RW. Axon pathfinding was also impaired in vitro and in vivo in Plekhg2-deficient cortical neurons. At P14, knockdown of Plekhg2 was observed to cause defects in dendritic spine morphology formation. Collectively, these results strongly suggest that PLEKHG2 has essential roles in the maturation of axon, dendrites, and spines. Moreover, impairment of PLEKHG2 function is most likely to cause defects in neuronal functions that lead to neurodevelopmental disorders.

Lysophosphatidic acid (LPA) and LPA1 receptor signaling play a crucial role in the initiation of peripheral nerve injury-induced neuropathic pain through the alternation of pain-related genes/proteins expression and demyelination. However, LPA and its signaling in the brain are still poorly understood. In the present study, we revealed that the LPA5 receptor expression in corpus callosum elevated after the initiation of demyelination, and the hyperalgesia through Aδ-fibers following cuprizone-induced demyelination was mediated by LPA5 signaling. These data suggest that LPA5 signaling may play a key role in the mechanisms underlying neuropathic pain following demyelination in the brain. Keywords: LPA5, Neuropathic pain, Multiple sclerosis

RNA synthesis inhibitors and protein synthesis inhibitors are useful for investigating whether biological events with unknown mechanisms require transcription or translation; however, the dependence of RNA synthesis has been difficult to verify because many RNA synthesis inhibitors cause adverse events that trigger a p53 response. In this study, we screened a library containing 9600 core compounds and obtained STK160830 that shows anti-apoptotic effects in irradiated wild-type-p53-bearing human T-cell leukemia MOLT-4 cells and murine thymocytes. In many of the p53-impaired cells and p53-knockdown cells tested, STK160830 did not show a remarkable anti-apoptotic effect, suggesting that the anti-apoptotic activity is p53-dependent. In the expression analysis of p53, p53-target gene products, and reference proteins by immunoblotting, STK160830 down-regulated the expression of many of the proteins examined, and the downregulation correlated strongly with its inhibitory effect on cell death. mRNA expression analyses by qPCR and nascent RNA capture kit revealed that STK160830 showed a decreased mRNA expression, which was similar to that induced by the RNA synthesis inhibitor actinomycin D but differed to some extent. Furthermore, unlike other RNA synthesis inhibitors such as actinomycin D, p53 accumulation by STK160830 alone was negligible, and a DNA melting-curve analysis showed very weak DNA-intercalating activity, indicating that STK160830 is a useful inhibitor for RNA synthesis without triggering p53-mediated damage responses.

With the widespread application of recombinant DNA technology, many useful substances are produced by bioprocesses. For the monitoring of the recombinant protein production process, most of the existing technologies are those for the culture environment (pH, O2, etc.). However, the production status of the target protein can only be known after the subsequent separation and purification process. To speed up the monitoring of the production process and screening of the higher-yield target protein variants, here we developed an antibody-based His-tag sensor Quenchbody (Q-body), which can quickly detect the C-terminally His-tagged recombinant protein produced in the culture medium. Compared with single-chain Fv-based Q-body having one dye, the Fab-based Q-body having two dyes showed a higher response. In addition, not only was fluorescence response improved but also detection sensitivity by the mutations of tyrosine to tryptophan in the heavy chain CDR region. Moreover, the effect of the mutations on antigen-binding was successfully validated by molecular docking simulation by CDOCKER. Finally, the constructed Q-body was successfully applied to monitor the amount of anti-SARS CoV-2 nanobody secreted into the Brevibacillus culture media.

Background and objectives: The number of laparoscopic surgeries performed is increasing every year and in most cases the pneumoperitoneum method is used. One alternative is the abdominal wall lifting method and this study was undertaken to evaluate changes of functional residual capacity during the abdominal wall lift procedure. Methods: From January to April 2013, 20 patients underwent laparoscopic cholecystectomy at a single institution. All patients were anesthetized using propofol, remifentanil and rocuronium. FRC was measured automatically by Engstrom Carestation before the abdominal wall lift and again 15 minutes after the start of the procedure. Results: After abdominal wall lift, there was a significant increase in functional residual capacity values (before abdominal wall lift 1.48 × 103 mL, after abdominal wall lift 1.64 × 103 mL) (p

Treatment for fibromyalgia is an unmet medical need and its pathogenesis is still poorly understood. The present study demonstrated that intermittent psychological stress (IPS), or empathy causes generalized chronic abnormal pain with female predominance. The persistence of the pain phenotype was dependent on the unpredictability of the stressor. The pain was reversed by pregabalin (PGB), duloxetine (DLX) or mirtazapine (Mir), but not by diclofenac or morphine. Differential administration of these existing medicines revealed that the sites of PGB and Mir actions exist in the brain, but not in the spinal cord, while that of DLX is preferentially in the spinal cord. It is interesting to note that the intracerebroventricular injection of PGB or Mir showed potent analgesia for 24 h or longer, though systemic injection of these medicines shows anti-hyperalgesia just for several hours. These results indicate that initial intense actions in the target brain may prevent the forthcoming development of pain memory. IPS-induced abnormal pain was prevented in mice deficient of lysophosphatidic acid receptor 1 (LPA1) gene, and completely cured by the repeated intrathecal treatments with LPA1 antagonist, AM966, which did not show acute action. All these results suggest that IPS model is an experimental animal model, which mimics the pathophysiology and pharmacotherapy in fibromyalgia in clinic, and LPA1 signaling plays crucial roles in the IPS-induced fibromyalgia-like abnormal pain. Keywords: Fibromyalgia, Empathy, Psychological stress, Mirtazapine, Morphine, Lysophosphatidic acid

Migration is a widespread but highly diverse component of many animal life histories. Fish migrate throughout the world's oceans, within lakes and rivers, and between the two realms, transporting matter, energy, and other species (e.g., microbes) across boundaries. Migration is therefore a process responsible for myriad ecosystem services. Many human populations depend on the presence of predictable migrations of fish for their subsistence and livelihoods. Although much research has focused on fish migration, many questions remain in our rapidly changing world. We assembled a diverse team of fundamental and applied scientists who study fish migrations in marine and freshwater environments to identify pressing unanswered questions. Our exercise revealed questions within themes related to understanding the migrating individual's internal state, navigational mechanisms, locomotor capabilities, external drivers of migration, the threats confronting migratory fish including climate change, and the role of migration. In addition, we identified key requirements for aquatic animal management, restoration, policy, and governance. Lessons revealed included the difficulties in generalizing among species and populations, and in understanding the levels of connectivity facilitated by migrating fishes. We conclude by identifying priority research needed for assuring a sustainable future for migratory fishes.

We developed a mouse model for central post-stroke pain (CPSP), a centrally-originated neuropathic pain (NeuP). In this mode, mice were first injected with Rose Bengal, followed by photo-irradiation of left middle cerebral artery (MCA) to generate thrombosis. Although the MCA thrombosis was soon dissolved, the reduced blood flow remained for more than 24 h due to subsequent occlusion of microvessels. This photochemically induced thrombosis (PIT) model showed a hypersensitivity to the electrical stimulation of both sides of paw, but did not show any abnormal pain in popular thermal or mechanical nociception tests. When tissue-type plasminogen activator (tPA) was injected 6 h after the PIT stress, tPA-dependent hypersensitivity to the electrical paw stimulation and stable thermal and mechanical hyperalgesia on both sides for more than 17 or 18 days after the PIT treatment. These hyperalgesic effects were abolished in lysophosphatidic acid receptor 1 (LPA1)- and lysophosphatidic acid receptor 3 (LPA3)-deficient mice. When Ki-16425, an LPA1 and LPA3 antagonist was treated twice daily for 6 days consecutively, the thermal and mechanical hyperalgesia at day 17 and 18 were significantly reversed. The liquid chromatography–mass spectrometry (LC–MS/MS) analysis revealed that there is a significant increase in several species of LPA molecules in somatosensory S-I and medial dorsal thalamus (MD), but not in striatum or ventroposterior thalamus. All these results suggest that LPA1 and LPA3 signaling play key roles in the development and maintenance of CPSP. Keywords: Central poststroke pain, Lysophosphatidic acid, LC–MS/MS, Photochemically induced thrombosis, tPA

Prothymosin alpha (ProTα) and S100A13 are released from C6 glioma cells under serum-free conditions via membrane tethering mediated by Ca2+-dependent interactions between S100A13 and p40 synaptotagmin-1 (Syt-1), which is further associated with plasma membrane syntaxin-1 (Stx-1). The present study revealed that S100A13 interacted with annexin A2 (ANXA2) and this interaction was enhanced by Ca2+ and p40 Syt-1. Amlexanox (Amx) inhibited the association between S100A13 and ANXA2 in C6 glioma cells cultured under serum-free conditions in the in situ proximity ligation assay. In the absence of Amx, however, the serum-free stress results in a flop-out of ANXA2 through the membrane, without the extracellular release. The intracellular delivery of anti-ANXA2 antibody blocked the serum-free stress-induced cellular loss of ProTα, S100A13, and Syt-1. The stress-induced externalization of ANXA2 was inhibited by pretreatment with siRNA for P4-ATPase, ATP8A2, under serum-free conditions, which ablates membrane lipid asymmetry. The stress-induced ProTα release via Stx-1A, ANXA2 and ATP8A2 was also evidenced by the knock-down strategy in the experiments using oxygen glucose deprivation-treated cultured neurons. These findings suggest that starvation stress-induced release of ProTα, S100A13, and p40 Syt-1 from C6 glioma cells is mediated by the ANXA2-flop-out via energy crisis-dependent recovery of membrane lipid asymmetry.

The transformation of a crystalline drug into an amorphous form is a promising way to enhance the oral bioavailability of poorly water-soluble drugs. Blending of a carrier, such as a hydrophilic polymer, with an amorphous drug is a widely used method to produce a solid dispersion and inhibit crystallization. This study investigates an experimental grade of hydroxypropyl methylcellulose acetate succinate, HPMCAS-MX (MX), as a solid dispersion carrier. Enhancement of thermal stability and reduction of the glass transition temperature (Tg) of MX compared with those of the conventional grade were evaluated through thermogravimetric analysis and differential scanning calorimetry (DSC). The formation of a homogeneous amorphous solid dispersion between MX and indomethacin was confirmed by X-ray powder diffraction analysis, DSC, and Raman mapping. It was observed that 10–30% MX did not act as an anti-plasticizer, but the utilization of >40% MX caused an increase in Tg and reduction of molecular mobility. This could be explained by a change in intermolecular interactions, inferred from infrared spectroscopy combined with principal component analysis. HPMCAS-MX exhibited similar performance to that of conventional-grade, HPMCAS-MG. Although HPMCAS-MX has thermal properties different from those of conventional-grade HPMCAS-MG, it retains its ability as a solid dispersion carrier.

The detection of viruses, disease biomarkers, physiologically active substances, drugs, and chemicals is of great significance in many areas of our lives. Immunodetection technology is based on the specificity and affinity of antigen–antibody reactions. Compared with other analytical methods such as liquid chromatography coupled with mass spectrometry, which requires a large and expensive instrument, immunodetection has the advantages of simplicity and good selectivity and is thus widely used in disease diagnosis and food/environmental monitoring. Quenchbody (Q-body), a new type of fluorescent immunosensor, is an antibody fragment labeled with fluorescent dyes. When the Q-body binds to its antigen, the fluorescence intensity increases. The detection of antigens by changes in fluorescence intensity is simple, easy to operate, and highly sensitive. This review comprehensively discusses the principle, construction, application, and current progress related to Q-bodies.

Contextual factors in which learning occurs are crucial aspects that learning analytics and related disciplines aim to understand for optimizing learning and the environments in which learning occurs. In foreign vocabulary development, taking the notes or memos of learning contexts along with other factors, play an essential role in quick memorization and reflection. However, conventional tools fail to automate the learning contexts generation process as learners still need to take memos or e-notes to describe their vocabulary learning contexts. This paper presents the Image Understanding Project (hereafter IUEcosystem) that could produce smartly-generated learning contexts primarily in a learner’s target languages. The IUEcosystem uses visual content analysis of lifelogging images as the sensor data to produce smartly-generated learning contexts that could be used as an alternative to handwritten memos or electronic notes. The IUEcosystem uses applied artificial intelligence to produce smartly-generated learning contexts. This intelligent learning environment collects a learner’s learning satisfaction and interaction data and, later on, analyzes them to produce time-based notifications for enhancing retention. Furthermore, a new learning design is presented that aims to map a learner’s prior vocabulary knowledge with new learning vocabularies to be learned. This learning design would help learners to review and recall prior knowledge while learning new vocabulary.

Lysophosphatidic acid (LPA) initiates demyelination following peripheral nerve injury, which causes neuropathic pain. Our previous in vivo and ex vivo studies using mice have demonstrated that LPA-induced demyelination of spinal dorsal roots is attributed by the LPA1-type receptor-mediated down-regulation of myelin-related molecules, such as MBP and MPZ. In this study using S16 mature-type Schwann cells, we found that LPA-induced down-regulation of myelin-related genes is attributed by the activation of LPA1 receptor, Rho kinase, and p300, leading to an acetylation of NFκB, which down-regulates the transcription of Sox10, MBP and MPZ genes.

Prothymosin alpha (ProTα) suppresses stress-induced necrosis of cultured cortical neurons. As neuroprotection alone could not explain the long-lasting protective actions against cerebral ischemia by ProTα, we further examined whether ProTα, in addition to neuroprotective effects, has other anti-ischemic activities. When recombinant mouse ProTα (rmProTα) at 0.3 mg/kg was intravenously (i.v.) given 2 h after the start of tMCAO, all mice survived for more than 14 days. In evaluation of CD31- and tomato lectin-labeling as well as IgG and Evans blue leakage, rmProTα treatment (0.1 mg/kg) largely blocked ischemia-induced vascular damages. Therefore, rmProTα has novel beneficial effects against ischemia-induced brain damage through vascular mechanisms.

Pregabalin (PGB) is a valuable therapeutic drug against chronic pain. Here we attempted to perform the combinatorial drug therapy with P-glycoprotein (P-gp) inhibitors to lower therapeutic dosage of PGB in the intermittent cold stress-induced fibromyalgia-like pain model. Single intracerebroventricular (i.c.v.) PGB injection exerted long-lasting anti-hyperalgesic effects for 72 h, while the effect of PGB given intraperitoneally (i.p.) disappeared within 3 h. Importantly, the pretreatment with P-gp inhibitors markedly prolonged the PGB (i.p.) effects, which lasted for 72 h. These results suggest that the combinatorial treatment with P-gp inhibitor enables the prolongation of dose-interval for PGB.

Lysophosphatidic acid (LPA) signaling is known to play key roles in the initiation and maintenance of various chronic pain models. Here we examined whether LPA signaling is also involved in diabetes-induced abnormal pain behaviors. The high-fat diet (HFD) showing elevation of blood glucose levels and body weight caused thermal, mechanical hyperalgesia, hypersensitivity to 2000 or 250 Hz electrical-stimulation and hyposensitivity to 5 Hz stimulation to the paw in wild-type (WT) mice. These HFD-induced abnormal pain behaviors and body weight increase, but not elevated glucose levels were abolished in LPA1−/− and LPA3−/− mice. Repeated daily intrathecal (i.t.) treatments with LPA1/3 antagonist AM966 reversed these abnormal pain behaviors. Similar abnormal pain behaviors and their blockade by daily AM966 (i.t.) or twice daily Ki16425, another LPA1/3 antagonist was also observed in db/db mice which show high glucose levels and body weight. Furthermore, streptozotocin-induced similar abnormal pain behaviors, but not elevated glucose levels or body weight loss were abolished in LPA1−/− and LPA3−/− mice. These results suggest that LPA1 and LPA3 play key roles in the development of both type I and type II diabetic neuropathic pain.

The problem of illicit drug use and addiction is an escalating issue worldwide. As such, fast and precise detection methods are needed to help combat the problem. Herein, the synthesis method for an anti-methamphetamine Quenchbody (Q-body), a promising sensor for use in simple and convenient assays, has been described. The fluorescence intensity of the Q-body generated by two-site labeling of Escherichia coli produced anti-methamphetamine antigen-binding fragment (Fab) with TAMRA-C2-maleimide dyes increased 5.1-fold over background in the presence of a hydroxyl methamphetamine derivative, 3-[(2S)-2-(methylamino)propyl]phenol. This derivative has the closest structure to methamphetamine of the chemicals available for use in a laboratory. Our results indicate the potential use of this Q-body as a novel sensor for the on-site detection of methamphetamine, in such occasions as drug screening at workplace, suspicious substance identification, and monitoring patients during drug rehabilitation.