Biology subjects

Nishida, N.

Publications and source records attributed to Nishida, N..

12 recordsLinked to original sources

bioRxiv2026

Modular mRNA platform for in vivo antigen-specific immune tolerance

Immune-mediated diseases such as autoimmunity and allergy arise from dysregulated antigen-specific responses, yet current therapies largely rely on broad immunosuppression and rarely re-establish durable, antigen-selective tolerance. Here we introduce Tol-mRNA, a modular tolerogenic mRNA platform designed to deliver multiple immune-regulatory signals in vivo to program antigen-specific tolerance. Tol-mRNA co-encodes an antigen together with two complementary regulatory cues, PD-L1 and TGF-{beta}, enabling coordinated antigen presentation and tolerogenic signaling that robustly induces antigen-specific Tregs and suppresses pathogenic effector responses. In murine disease models, Tol-mRNA conferred therapeutic benefit across distinct immune pathologies. In experimental autoimmune encephalomyelitis, Tol-mRNA ameliorated clinical disease and reduced inflammatory immune activation. In an ovalbumin-driven food allergy model, Tol-mRNA prevented allergic symptoms and dampened type 2 inflammation. Extending these findings to humans, Tol-mRNA demonstrated potent, antigen-dependent immunosuppressive activity and a strong capacity to promote antigen-specific Treg induction in human immune settings. Collectively, these results establish Tol-mRNA as a scalable and versatile approach to reprogram antigen-specific immunity and support its development as a next-generation therapeutic modality for immune-mediated diseases.

immunology↗

bioRxiv2025

Clinical and molecular characterisation of primary refractoriness to atezolizumab plus bevacizumab in patients with unresectable hepatocellular carcinoma.

BackgroundDespite improved outcomes with atezolizumab plus bevacizumab (A+B) in hepatocellular carcinoma (HCC), primary refractoriness (PRef), characterised by early progression or short-lived disease stabilisation following treatment, remains a significant and poorly understood clinical challenge. MethodsWe analysed 1296 patients with HCC and Child-Pugh A liver cirrhosis treated with frontline A+B (AB-real) and validated findings in 645 trial participants recruited to IMbrave150 and GO30140. PRef was defined by Society for the Immunotherapy of Cancer (SITC) criteria as progressive disease in the first 6 months after treatment initiation. Patients who achieved complete response, partial response or stable disease for [≥] 6 months were classified as responders. We performed a multi-parametric analysis of pre-treatment tumour tissue including machine learning-based quantification of tumour-infiltrating lymphocytes, imaging mass cytometry and RNA sequencing (RNAseq) to evaluate differences in the tumour microenvironment (TME) of PRef versus responding patients. We employed conditional inference tree analyses to provide a hierarchical organisation of determinants of PRef. ResultsAmong 677 AB-real and 378 trial patients evaluable by SITC criteria, PRef identified inferior median OS in comparison with responding patients (AB-real: 7.3 vs. 31.5 months, HR 3.7, 95%CI 2.8-8.5, p<0.001; Trials: 10.8 vs. NR, HR 4.6, 95%CI 3.3-6.3, p<0.001). PRef patients exhibited higher baseline systemic inflammation (neutrophil-to-lymphocyte ratio, NLR [≥]3), a distinctively immunosuppressive TME enriched in CD163+ tumour-associated macrophages and a higher Treg/Teff ratio. RNAseq of tumour tissue demonstrated lower intrinsic immunogenicity in PRef samples, characterised by repressed IFN-{gamma} and Teff signatures, with elevated myeloid infiltration. Conditional inference tree analysis identified IFN-{gamma} signature downregulation combined with NLR [≥]3 as the strongest contributor of PRef. ConclusionsPRef to A+B identifies a distinct biological entity characterised by unopposed systemic inflammation, myeloid cell infiltration and T-cell depletion. Targeting myeloid-mediated immunosuppression, particularly in patients with low IFN-{gamma} signature expression and elevated NLR might enhance responsiveness to A+B. HighlightsO_LIPrimary refractoriness to atezolizumab plus bevacizumab in hepatocellular carcinoma, as defined by SITC criteria, is associated with poor clinical outcomes. C_LIO_LITumour microenvironment profiling reveals an immunosuppressive phenotype characterized by high myeloid infiltration, reduced interferon-{gamma} signalling, and T-cell depletion. C_LIO_LIThe combination of systemic inflammation and low IFN-{gamma} signature expression strongly predicts primary refractoriness and may inform therapeutic decision-making. C_LI

cancer biology↗

bioRxiv2024

Pelvic Ring Fractures: Does Spinopelvic Alignment Affect Sacral and Lumbopelvic Fixation Stability?

Background ContextPelvic ring is a complex boney and ligamentous complex and its structural integrity is paramount to be able to withstand the high forces. Pelvic ring fractures, which can result from low-energy impacts or high-velocity injuries and falls from great heights, can severely impair the structural integrity of the pelvic ring. These injuries can be fatal, especially in geriatric patients, with mortality rates ranging between 10% and 16%. Pelvic ring injuries require immediate stabilization, typically performed through internal fixation, to maintain circulatory balance and achieve anatomical restoration of the pelvis. MethodsA previously validated FE model of the spine, pelvis, and femur model was used to evaluate how different fixation techniques for pelvic ring injury affect spinopelvic biomechanics. The SS, PI, PT and LL of the normal, intact model was modified to generate two more SS models for this study. One modified model with an SS of 20{o} was considered the low SS model (SSD), while the second modified model with an SS of 32{o} was termed the high SS model (SSI). A unilateral pelvic ring fracture was simulated by resecting the left side of the sacrum and pelvis. The fractures were stabilized using two different posterior stabilization techniques 1) The fractures were stabilized using two different posterior stabilization techniques, and 2) L5-Ilium posterior screw fixation without cross connector (L5_PF_WO_CC). ResultsSSI demonstrated the higher range of motion, followed by Normal model and least by SSD model at right Sacroiliac Joint. No differences were seen between with and without cross connector model for range of motion at right SIJ. For L5-S1 motion models treated with cross connectors demonstrated the least motion compared to those treated without cross connectors for all configurations and loading conditions. For the sacrum fracture, models with high sacral slopes recorded the least horizontal displacement, followed by decreased sacral slope models for both configurations and loading conditions.

bioengineering↗

bioRxiv2024

Biomechanical Implications of Spinopelvic Alignment on Femoral Head Cartilage and the Proximal Femoral Physis in Slipped Capital Femoral Epiphysis: A Theoretical Finite Element Analysis

BackgroundSlipped capital femoral epiphysis (SCFE) is a prevalent pediatric hip disorder. Recent studies suggest the spines sagittal profile may influence the proximal femoral growth plates slippage, an aspect not extensively explored. This study utilizes finite element analysis to investigate how different spinopelvic alignments affect shear stress and potential slippage at the growth plate. MethodsA finite element model was developed from CT scans of a healthy adult male lumbar spine, pelvis, and femurs. The model was subjected to various sagittal alignments through rotational boundary conditions. Simulations of two-leg stance, one-leg stance, walking heel strike, ascending stairs heel strike, and descending stairs heel strike were conducted. Parameters measured included hip joint contact area, stress, and maximum Tresca (shear) stress on the growth plate. FindingsPosterior pelvic tilt cases indicated larger shear stresses compared to the anterior pelvic tilt variants except in two leg stance. Two leg stance resulted in decreases in the posterior tilted pelvi variants compared to anterior tilted pelvi, however a combination of posterior pelvic tilt and high pelvic incidence indicated larger shear stresses on the growth plate. One leg stance and heal strike resulted in higher shear stress on the growth plate in posterior pelvic tilt variants compared to anterior pelvic tilt, with a combination of posterior pelvic tilt and high pelvic incidence resulting in the largest shear stress. InterpretationOur findings suggest that posterior pelvic tilt and high pelvic incidence can lead to increased shear stress at the growth plate. Activities performed in patients with these alignments may predispose to biomechanical loading that shears the growth plate, potentially causing slippage.

bioengineering↗

bioRxiv2020

Phase transition of prion protein initiates proteinase K resistant amyloid formation

Prion diseases are characterized by accumulation of amyloid fibrils. The causative agent is an infectious amyloid that is comprised solely of misfolded prion protein (PrPSc). Prions can convert PrPC to proteinase-resistant PrP (PrP-res) in vitro; however, the intermediate steps involved in the spontaneous conversion remain unknown. We investigated whether recombinant prion protein (rPrP) can directly convert into PrP-res via liquid-liquid phase separation in the absence of PrPSc. We found that rPrP underwent liquid-liquid phase separation at the interface of the aqueous two-phase system (ATPS) of polyethylene glycol (PEG) and dextran, whereas single-phase conditions were not inducible. Fluorescence recovery assay after photobleaching revealed that the liquid-solid phase transition occurred within a short time. The aged rPrP-gel acquired proteinase-resistant amyloid accompanied by {beta}-sheet conversion, as confirmed by western blotting, Fourier transform infrared spectroscopy, and Congo red staining. The reactions required both the N-terminal region of rPrP (amino acids 23-89) and kosmotropic salts, suggesting that the kosmotropic anions may interact with the N-terminal region of rPrP to promote liquid-liquid phase separation. Thus, structural conversion via liquid-liquid phase separation and liquid-solid phase transition are intermediate steps in the conversion of prions.

biochemistry↗

bioRxiv2019

Molecular dynamics simulation of local structural models of PrPSc reveals how codon 129 polymorphism affects propagation of PrPSc.

Prions are unconventional pathogen without nucleotide genome and their pathogenic properties are defined by the primary structure and the conformation of the constituent abnormal isoform (PrPSc) of prion protein (PrP). A polymorphic codon 129 of human PrP that is valine (V129) or methionine (M129) is particularly influential on properties of PrPSc, affecting transmission efficiencies and clinicopathological features. However, how the single residue is so influential has not been elucidated because the detailed structures of PrPSc have not been determined yet due to its incompatibility with high-resolution structural analysis. Previously we created an in-register parallel {beta}-sheet local structural model of human PrPSc encompassing residues 107 to 143 that seemed more compatible with V129 than M129, based on knowledge from -synuclein amyloids and an NMR-based model of the amyloid of Y145Stop mutant of PrP in the literature. Here, we created an M129-compatible local structural model of PrPSc. Severe destabilization of the model by G127V mutation was consistent with the protective effects of V127 polymorphism of human PrP against prions. It was highly sensitive to the length of the hydrophobic side chain of codon 129 and replacement of M129 with leucine or valine destabilized the structures. Interestingly, the U-shaped {beta}-arch which comprises M129 flexibly changed hydrophobic interaction networks inside the {beta}-arch depending on the interactions with the surrounding structures, whereas the previous model with V129 maintained the similar network patterns irrespective of the surroundings. The differences between the two models may explain influences of the codon 129 polymorphism on transmissions and properties of human prions.

molecular biology↗

bioRxiv2019

Structural basis for two-way communication between dynein and microtubules

The movements of cytoplasmic dynein on microtubule (MT) tracks is achieved by two-way communication between the microtubule-binding domain (MTBD) and the ATPase domain of dynein via an a-helical coiled-coil stalk, but the structural basis of this communication remains elusive. Here, we regulated MTBD either in high-affinity or low-affinity states by introducing a disulfide bond between the coiled-coils and analyzed the resulting structures by NMRand cryo-EM. In the MT-unbound state, the affinity changes of MTBD were achieved by sliding of the N-terminal -helix by one half-turn, which suggests that structural changes propagate from the ATPase-domain to MTBD. In addition, cryo-EM analysis showed that MT binding induced further sliding of the N-terminal -helix even without the disulfide bond, which suggests the MT-induced conformational changes propagate toward the ATPase domain. Based on differences in the MT-binding surface between the high- and low-affinity states, we propose a potential mechanism for the directional bias of dynein movement on MT tracks.

biophysics↗

bioRxiv2019

Conformation-dependent influences of hydrophobic amino acids in two in-register parallel β-sheet amyloids, an α-synuclein amyloid and a local structural model of PrPSc

Prions are unconventional pathogens that encode the pathogenic information in conformations of the constituent abnormal isoform of prion protein (PrPSc), independently of the nucleotide genome. Therefore, conformational diversity of PrPSc underlies the existence of many prion strains and species barriers of prions, although the conformational information is extremely limited. Interestingly, differences between polymorphic or species-specific residues responsible for the species/strain barriers are often caused by conservative replacements between hydrophobic amino acids. This implies that subtle differences among hydrophobic amino acids are significant for PrPSc structures. Here, we analyzed the influence of different hydrophobic residues on the structures of an in-register parallel {beta}-sheet amyloid of -synuclein (Syn) using molecular dynamics (MD) simulation, and applied the knowledge from the Syn amyloid to modeling a local structure of human PrPSc encompassing residues 107-143. We found that mutations equivalent to polymorphisms that cause transmission barriers substantially affect the stabilities; for example, the G127V mutation, which makes the host resistant to various human prion diseases, greatly destabilized the model amyloid. Our study demonstrates specifically how and in what structures hydrophobic residues can exert unique effects on in-register parallel {beta}-sheet amyloids and provides insights into the molecular mechanism of the strain diversity of prions and other pathogenic amyloids. For Table of Contents Only O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=111 SRC="FIGDIR/small/758938v4_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@1e55372org.highwire.dtl.DTLVardef@c98952org.highwire.dtl.DTLVardef@1566c06org.highwire.dtl.DTLVardef@ce1945_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2019

In silico evidence of unique behaviors of methionine in an in-register parallel beta-sheet amyloid, suggestive of possible contribution to strain diversity of amyloids.

Mechanism of strain diversity of prions is a long-standing conundrum, because prions consist solely of abnormal isoform of prion protein (PrPSc) devoid of genetic material. Pathogenic properties of prions are determined by conformations of the constituent PrPSc according to the protein-only hypothesis, and alterations to even a single residue can drastically change the properties when the residue is located at a critical position for the structure of PrPSc. Interestingly, methionine (Met) is often recognized as the polymorphic or species-specific residues responsible for species/strain barriers of prions, implying its unique influences on the structures of PrPSc. However, how it is unique is difficult to demonstrate due to lack of the detailed structures of PrPSc. Here we analyzed influences of Met substitutions on structures of an in-register parallel {beta}-sheet amyloid of -synuclein (Syn) by molecular dynamics (MD) simulation, to extrapolate the results to PrPSc. The MD simulation revealed that Met uniquely stabilized a U-shaped {beta}-arch of the Greek-key Syn amyloid, whereas other hydrophobic amino acids tended to destabilize it. The stabilizing effect of Met was attributable to the long side chain without C{beta} branching. Our findings exemplify specifically how and in what structure of an in-register parallel {beta}-sheet amyloid Met can uniquely behave, and are suggestive of its influences on structures of PrPSc and strain diversity of prions. We also discuss about relations between -helix propensity and local structures of in-register parallel amyloids.

molecular biology↗

bioRxiv2018

Mechanisms of strain diversity of disease-associated in-register parallel β-sheet amyloids and implications about prion strains.

The mechanism of strain diversity of prions still remains unsolved, because the investigation of inheritance and diversification of the protein-based pathogenic information demands identification of the detailed structures of abnormal isoform of prion protein (PrPSc), while it is difficult to purify for analysis without affecting the infectious nature. On the other hand, the similar prion-like properties are recognized also in other disease-associated in-register parallel {beta}-sheet amyloids including Tau and -synuclein (Syn) amyloids. Investigations into structures of those amyloids by solid-state nuclear magnetic resonance spectroscopy and cryo-electron microscopy recently made remarkable advances, because of their relatively small sizes and lack of post-translational modifications. We review the advances on those pathogenic amyloids, particularly Tau and Syn, and discuss their implications about strain diversity mechanisms of prion/PrPSc from the viewpoint that PrPSc is an in-register parallel {beta}-sheet amyloid. We also present our recent data of molecular dynamics simulations of Syn amyloid, which suggest significance of compatibility between {beta}-sheet propensities of the substrate and local structures of the template for stability of the amyloid structures. Detailed structures of the Syn and Tau amyloids are good surrogate models of pathogenic amyloids including PrPSc to elucidate not only the strain diversity but also their pathogenic mechanisms.

molecular biology↗

bioRxiv2018

Molecular dynamics simulation reveals that switchable combinations of β-sheets underlie the prion-like properties of α-synuclein amyloids.

Diversity of prion strains is one of the most mysterious traits of prions because they are mere aggregates of abnormally-folded forms of single protein species, prion protein (PrPSc), without genome. Although the strain-specific properties are hypothesized to be enciphered in the strain-specific structures of PrPSc instead of nucleotide genome, specifically what structure can code the information remains an enigma due to the incompatibility of PrPSc with structural analyses. Although the strain diversity was regarded as unique to prions, recently other disease-associated amyloids of -synuclein (Syn) or tau are also reported to have \"strains\". As detailed structures of Syn amyloid are already identified and the properties of mutant Syn associated with familial Parkinsons diseases, e.g. A53T, H50Q, and G51D, have been characterized, structure-phenotype relations of this type of amyloid could be investigated by using the Syn amyloid as a model. Here we intensively investigated the mutant Syn amyloids by molecular dynamics simulation to characterize influences of mutations on the structures of homo- or hetero-oligomer stacks of the amyloid. The simulations revealed directionality of the amyloid stack, remote effects of the mutations on distant {beta}-sheets, existence of at least two switchable interfaces/amyloid cores, and distinct effects of hetero-oligomerization depending on mutation types. Collectively, those findings implied a possible mechanism of the strain diversity of the amyloids which have multiple in-register parallel {beta}-sheets side-by-side, and support the view that their prion-like properties are inherent in the characteristic structures. We expect that the notion is also applicable to PrPSc.

molecular biology↗

bioRxiv2017

PrPSc-induced conformational changes and strain-specific structures of PrPSc revealed by Disulfide-crosslink scanning

There exist many phenotypically-varied prion strains, like viruses, despite the absence of conventional genetic material which codes the phenotypic information. As prion is composed solely of the pathological isoform (PrPSc) of prion protein (PrP), the strain-specific traits are hypothesized to be enciphered in the structural details of PrPSc. Identification of the structures of PrPSc is therefore vital for the understanding of prion biology, though they remain unidentified due to the incompatibility of PrPSc with conventional high-resolution structural analyses. Based on our previous hypothesis that the region between the first and the second -helix (H1[~]H2) and the distal region of the third helix (Ctrm) of the cellular isoform of PrP (PrPC) have important roles for efficient interactions with PrPSc, we created series of mutant PrPs with two cysteine substitutions (C;C-PrP) which were systematically designed to form an intramolecular disulfide crosslink between H1[~]H2 and Ctrm and assessed their conformational changes by prions: Specifically, a cysteine substitution in H1[~]H2 from 165 to 169 was combined with cysteine-scanning along Ctrm from 220 to 229. C;C-PrPs with the crosslinks were expressed normally with the similar glycosylation patterns and subcellular localization as the wild-type PrP albeit with varied expression levels. Interestingly, some of the C;C-PrPs converted to the protease-resistant isoforms in the N2a cells persistently infected with 22L prion strain, whereas the same mutants did not convert in the cells infected with another prion strain Fukuoka1, indicating that local structures of PrPSc in these regions vary among prion strains and contribute to prion-strain diversity. Moreover, patterns of the crosslinks of the convertible C;C-PrPs implied drastic changes in positional relations of H1[~]H2 and Ctrm in the PrPSc-induced conformational changes by 22L prion. Thus, disulfide-crosslink scanning is a useful approach for investigation of strain-specific structures of PrPSc, and would be applicable to other types of amyloids as well.

molecular biology↗