Small Fibrils Research Articles

Engineered nanofibrillar collagen with tunable biophysical properties for myogenic, endothelial, and osteogenic cell guidance

A goal of regenerative engineering is the rational design of materials to restore the structure-function relationships that drive reparative programs in damaged tissues. Despite the widespread use of extracellular matrices for engineering tissues, their application has been limited by a narrow range of tunable features. The primary objective of this study is to develop a versatile platform for evaluating tissue-specific cellular interactions using Type I collagen scaffolds with highly tunable biophysical properties. The kinetics of collagen fibrillogenesis were modulated through a combination of varied shear rate and pH during neutralization, to achieve a broad range of fibril anisotropy, porosity, diameter, and storage modulus. The role that each of these properties play in guiding muscle, bone, and vascular cell types was comprehensively identified, and informed the in vitro generation of three distinct musculoskeletal engineered constructs. Myogenesis was highly regulated by smaller fibrils and larger storage moduli, endothelial inflammatory phenotype was predominantly guided by fibril anisotropy, and osteogenesis was enhanced by highly porous collagen with larger fibrils. This study introduces a novel approach for dynamically modulating Type I collagen materials and provides a robust platform for investigating cell-material interactions, offering insights for the future rational design of tissue-specific regenerative biomaterials. Statement of significanceThe biophysical properties of regenerative materials facilitate key cell-substrate interactions that can guide the morphology, phenotype, and biological response of cells. In this study, we describe the fabrication of an engineered collagen hydrogel that can be modified to exhibit control over a wide range of biophysical features, including fibril organization and size, nanoscale porosity, and mechanics. We identified the unique combination of collagen features that optimally promote regenerative muscle, bone, and vascular cell types while also delineating the properties that hinder these same cellular responses. This study presents a highly accessible method to control the biophysical properties of collagen hydrogels that can be adapted for a broad range of tissue engineering and regenerative applications.

Real-time single-molecule observation of incipient collagen fibrillogenesis and remodeling

The hierarchic assembly of fibrillar collagen into an extensive and ordered supramolecular protein fibril is critical for extracellular matrix function and tissue mechanics. Despite decades of study, we still know very little about the complex process of fibrillogenesis, particularly at the earliest stages where observation of rapidly forming, nanoscale intermediates challenges the spatial and temporal resolution of most existing microscopy methods. Using video rate scanning atomic force microscopy (VRS-AFM), we can observe details of the first few minutes of collagen fibril formation and growth on a mica surface in solution. A defining feature of fibrillar collagens is a 67-nm periodic banding along the fibril driven by the organized assembly of individual monomers over multiple length scales. VRS-AFM videos show the concurrent growth and maturation of small fibrils from an initial uniform height to structures that display the canonical banding within seconds. Fibrils grow in a primarily unidirectional manner, with frayed ends of the growing tip latching onto adjacent fibrils. We find that, even at extremely early time points, remodeling of growing fibrils proceeds through bird-caging intermediates and propose that these dynamics may provide a pathway to mature hierarchic assembly. VRS-AFM provides a unique glimpse into the early emergence of banding and pathways for remodeling of the supramolecular assembly of collagen during the inception of fibrillogenesis.

Effects of 32 d of spaceflight on tendon collagen fibril morphology and nanomechanics

INTRODUCTION: Collagen is an essential component of connective tissue and plays a crucial role in elasticity, stability, and force distribution. While the adaptations of tendon to internal stimuli have been extensively researched, the impact of environmental factors (such as microgravity) on its structure and function at the nanoscopic scale (i.e., fibril level) remains largely unexplored. The purpose of this study was to determine the effects of 32 days (d) aboard the International Space Station (ISS) on murine tendon collagen fibril morphology and nanomechanics (Young’s Modulus, YM). We hypothesized there would be a decrease in fibril size and YM after spaceflight. Methods: Quadriceps tendon samples from female C57BL/6J mice from NASA Rodent Research-1 underwent chemical processing and mechanical isolation. Subsequently, collagen fibrils (n=56) were studied via atomic force microscopy (AFM; JPK NanoWizard 4a) to compare the morphology (diameter, height) and nanomechanics (average YM) between spaceflight (SF) and ground control (GC) specimens. The qp-BioAC-CB1 cantilever (spring constant: 0.3 N/m, tip radius: 10μm) was used under quantitative imaging mode (QI) for all force and morphological measurements. The data from 56 micrographs (SF n=30; GC n=26) was processed via the JPK Data Processing Software (Version 6.4.5.), and finally analyzed with Gwyddion (Version 2.62). A 2-way MANOVA compared all measurements. Results: More ‘small fibrils’ (height <50nm) were identified in the SF vs GC samples. SF fibrils averaged 20.3% smaller (width) compared to GC, but this difference was not significant (p>0.05). Additionally, while there was a trend for a lower YM in the SF samples, YM for SF specimens was not statistically different between groups (GC=3.26±1.94 MPa; SF= 2.76±1.44 MPa) (p>0.05). CONCLUSION: To our knowledge, this is the first investigation to use AFM QI-mode on tendon collagen fibrils after spaceflight. While there were no statistically significant changes in mean fibril size and YM between SF and GC, these results suggest that collagen fibrils may require more time to adapt to changes in their environment, and/or more rapid changes may take place at different levels of the collagen hierarchy. Our data provides valuable insight into mammalian tissue adaptations during spaceflight. Future research should continue these investigations with longer-duration missions to build a time-course of tendon adaptations to microgravity. FUNDING: CSUBIOTECH New Investigator Grant (JRB). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Comparison of Immunohistochemical Expression of Calretinin, Map2, S-100 and Glut1 in Rectal Biopsies from Suspected Hirschsprung’s Disease

Background: This study aimed to compare the immunohistochemical expression of calretinin, Map2, S-100, and Glut1 in rectal biopsy samples from patients suspected of Hirschsprung’s disease (HD).Methods and Results: Rectal biopsy samples from 40 patients with suspected HD were analyzed using hematoxylin and eosin (H&E) and immunohistochemistry (IHC). Immunohistochemical stains were assessed after previous routine histology interpretation, which was classified as “Positive or in favor for HD” and “Equivocal or negative for HD.” The staining patterns for calretinin, Map2, S-100, and Glut1 were analyzed regarding the following structures: lamina propria small nerve fibrils, submucosal small nerve fibrils,submucosal nerve fibers, and submucosal ganglia. The IHC stains for calretinin and Map2 were score-ranked as 0 – negative and 1 – positive. The IHC stain for S-100 was score-ranked as 0 – normal and 1 – hypertrophic. The IHC stain for Glut1 was ranked as 0 – normal perineural and 1 – conspicuous perineural accentuation Calretinin had 92% accuracy, the highest sensitivity (80%) and specificity (92.00%).Map2 also had the same accuracy as calretinin but lower sensitivity (46.67%). Regarding S-100 and Glut1, these two markers did not support a conclusive diagnosis. The accuracy for S-100 and Glut1 was 66.7% and 60%, respectively. Conclusion: Calretinin remains the currently most valuable single IHC marker in diagnosing difficult cases of HD.

Small-angle neutron scattering from cellulose solutions in phosphoric acid at different water content

Cellulose from biomass is an abundant and renewable alternative source for chemicals and fuels, yet its utilization by chemical or biological process requires pre-treatment in order to release the macromolecules from their tightly packed crystal structure. Phosphoric acid (PA) has been known for many years to be an efficient solvent for crystalline cellulose. It is also established that a certain quantity of water content in PA is required for efficient pretreatment. This study uses small-angle neutron scattering (SANS) measurements to evaluate cellulose dissolution in deuterated phosphoric acid (dPA), at different wt% dPA between 78 and 97 % (different D2O content). The SANS method is useful for this purpose due to the availability of deuterated dPA, its contrast in scattering length density towards cellulose, and its low incoherent scattering cross-section. The results indicate that most of the cellulose in 2 wt% solution is dissolved in PA as individual chains, at acid content of 81–94 wt% PA. Structural differences of the dissolved cellulose in PA of the various water compositions in this range are insignificant. At 78 % dPA the cellulose crystal still seem to be disrupted, yet the structure can be modeled as mass-surface fractals of small fibrils with irregular surface, possibly due to dissolved chain segments, which are aggregated as mass fractals of rods. At 97 % dPA evidence for a small content of undissolved fibrils is noted.

Investigation of supramolecular structures in various aqueous solutions of an amyloid forming peptide using small-angle X-ray scattering.

Aggregation of peptide molecules into amyloid fibrils is a characteristic feature of several degenerative diseases. However, the details behind amyloid-formation, and other self-assembled peptide aggregates, remain poorly understood. In this study, we have used small-angle X-ray scattering (SAXS), static and dynamic light scattering (SLS and DLS) as well as cryogenic transmission electron microscopy (cryo-TEM) to determine the structural geometry of self-assembled peptide aggregates in various dilute aqueous solutions. Pramlintide was used as a model peptide to assess the aggregation behaviour of an amyloid-forming peptide. The effects of adding sodium chloride (NaCl), sodium thiocyanate (NaSCN), and sodium fluoride (NaF) and the co-solvent dimethyl sulfoxide (DMSO) on the aggregation behaviour were studied. Our scattering data analysis demonstrates that small oligomeric fibrils aggregate to form networks of supramolecular assemblies with fractal dimensions. The choice of anion in small amounts of added salt has a significant impact on the size of the fibrils as well as on the fractal dimensions of supramolecular clusters. In DMSO the fractal dimension decreased with increasing DMSO concentration, indicating the formation of a less compact structure of the supramolecular assemblies.

Ultrastructural Examination of Glomerular Fibrillary Deposits in Diabetic Nephropathy

Glomerular fibrillary deposits have occasionally been reported in diabetic nephropathy, but no large-scale, ultrastructural evaluation of these deposits has been reported so far. Here, we report our study of glomerular non-Congophilic, DnaJ homolog subfamily B member 9 negative fibrillary deposits in diabetic nephropathy as characterized by transmission electron microscopy. Clinical data from 55 patients with biopsy-confirmed diabetic nephropathy and 18 healthy living donors were reviewed, and their biopsies were evaluated by light microscopy, immunofluorescence, and electron microscopy. Small fibrillary structures with a diameter of 10 ± 1 nm were present in all cases with diabetic nephropathy, regardless of the histologic class. In addition, glomerular fibrillary structures with a diameter of 23 ± 5 nm or 30 ± 7 nm were present in 35 cases. Interestingly, especially the small- and medium-sized fibrils, usually without apparent organization, were comparable with fibrils in fibrillary glomerulopathy. We conclude that glomerular fibrillary deposits occur far more commonly in renal biopsies of patients with diabetic nephropathy than generally considered. This is an important finding because their similarity to fibrils in fibrillary glomerulonephritis may complicate the histologic diagnostic process, especially in cases of overlapping clinical manifestations. Therefore, when encountering fibrillary deposits on electron microscopy, it is important to consider diabetic nephropathy as an alternative diagnosis.

Sustainable Lithium Ferro-Phosphate Cathode Manufacturing: A Semi-Dry Approach with Water-Based Processing and Polytetrafluorethylene Binders

A novel water-based lithium ferro-phosphate (LFP) cathode manufacturing process characterized by a significant reduction in the amount of solvent has been developed (semi-dry). To establish and validate this new process, Polytetrafluorethylene (PTFE) is used as a binder, with a binder content of 1 wt.%, minimizing the amount of inactive material within the electrode. Extrusion screws with multiple kneading zones stress the PTFE more intensively and thus produce more and smaller fibrils. The resulting extent of fibrillation is quantified by melting enthalpy as well as mechanical electrode properties. The degree of fibrillation of the binder in an electrode is known to influence the conductive electric and ionic pathways, which in turn affect the discharge capacity. It is shown that this process provides a flexible cathode layer that achieves a specific capacitance of 155 mAh g−1 in initial cycling tests at 0.1 C. Compared to a conventionally processed LFP cathode, the discharge capacity and overall energy output are significantly increased, and the overall energy consumption decreases for the semi-dry processed LFP cathodes.

Mmp14 is required for matrisome homeostasis and circadian rhythm in fibroblasts

The circadian clock in tendon regulates the daily rhythmic synthesis of collagen-I and the appearance and disappearance of small-diameter collagen fibrils in the extracellular matrix. How the fibrils are assembled and removed is not fully understood. Here, we first showed that the collagenase, membrane type I-matrix metalloproteinase (MT1-MMP, encoded by Mmp14), is regulated by the circadian clock in postnatal mouse tendon. Next, we generated tamoxifen-induced Col1a2-Cre-ERT2::Mmp14 KO mice (Mmp14 conditional knockout (CKO)). The CKO mice developed hind limb dorsiflexion and thickened tendons, which accumulated narrow-diameter collagen fibrils causing ultrastructural disorganization. Mass spectrometry of control tendons identified 1195 proteins of which 212 showed time-dependent abundance. In Mmp14 CKO mice 19 proteins had reversed temporal abundance and 176 proteins lost time dependency. Among these, the collagen crosslinking enzymes lysyl oxidase-like 1 (LOXL1) and lysyl hydroxylase 1 (LH1; encoded by Plod2) were elevated and had lost time-dependent regulation. High-pressure chromatography confirmed elevated levels of hydroxylysine aldehyde (pyridinoline) crosslinking of collagen in CKO tendons. As a result, collagen-I was refractory to extraction. We also showed that CRISPR-Cas9 deletion of Mmp14 from cultured fibroblasts resulted in loss of circadian clock rhythmicity of period 2 (PER2), and recombinant MT1-MMP was highly effective at cleaving soluble collagen-I but less effective at cleaving collagen pre-assembled into fibrils. In conclusion, our study shows that circadian clock-regulated Mmp14 controls the rhythmic synthesis of small diameter collagen fibrils, regulates collagen crosslinking, and its absence disrupts the circadian clock and matrisome in tendon fibroblasts.

Tunable Stress Relaxing Biomimetic Matrices: Hyaluronan/Hydroxyapatite Hybridization Mediates Assembly of Collagen Fibrils.

The alluring correlations of cellular behaviors with viscoelastic extracellular matrices have driven increasing endeavors directed toward the understanding of mechanical cues on cell growth and differentiation via preparing biomimetic scaffolds/gels with viscoelastic controllability. Indeed, systematic investigations, especially into calcium phosphate-containing biomimetics, are relatively rare. Here, oxidized hyaluronic acid/hydroxyapatite hybrids (OHAHs) were synthesized by hyaluronan-mediated biomimetic mineralization with confined ion diffusion and subsequent oxidization treatment. The collagen self-assembly was applied to fabricate tunable stress relaxing fibrillar matrices in the presence of OHAHs in which the incorporated hyaluronic acid with aldehyde groups acted to improve the component compatibility as well as to supplement the molecular interactions with the occurrence of a Schiff-base reaction. With the addition of varying OHAH contents, the self-assembly behavior of collagen was altered, and the obtained collagen-hybrid (CH) matrices presented a heterogeneous fibrillar structure interspersed with OHAHs, characterized by large fibrillar bundles coexisting with small fibrils. The OHAHs improved the hydrogel stability of pure collagen, and according to rheological and nanoindentation measurements, CH matrices also exhibited tunable stress relaxation rates, following an OHAH concentration-dependent fashion. The proliferation and spreading of MC3T3-E1 cells cultured onto such CH matrices were further found to increase with the stress relaxing rate of the matrices. The present study showed that the introduction of hydroxyapatite incorporated with active hyaluronic acid during collagen reconstitution was a simple and effective strategy to realize the preparation of tunable stress relaxing biomimetic matrices potentially used for further appraising the regulation of mechanical cues on cell behaviors.

Knockdown of biglycan reveals an important role in maintenance of structural and mechanical properties during tendon aging.

Biglycan, a small leucine-rich proteoglycan (SLRP), is involved in collagen fibrillogenesis and also acts as a signaling molecule. Although decorin has been considered as the primary SLRP in developing and maintaining tendon structure and mechanics, more recent work using inducible knockdown models suggests that biglycan is involved in tendon homeostasis. The purpose of the study was to determine the role of biglycan in tendon homeostasis to maintain mechanical and structural integrity in aged mice. Aged (485 days old) female Bgn+/+ control (wild type [WT], n = 16) and 16 bitransgenic conditional Bgnflox/flox mice (I-Bgn-/- , n = 16) with a tamoxifen-inducible Cre (driven by ROSA) were utilized. After biglycan knockdown, the transgenic model demonstrated effective knockdown of the target gene without any compensation from other SLRPs or type I collagen. Patellar tendon cellularity was not modified after biglycan knockdown. However, biglycan knockdown had an impact on collagen fibrillogenesis with a higher percentage of small diameter fibrils (25-45 nm) and a lower percentage of medium size fibrils (150-165 nm) in I-Bgn-/- tendons. Biglycan knockdown also induced a reduction in the midsubstance modulus and maximum stress compared to WT. Stress relaxation was reduced at 4% strain in I-Bgn-/- tendons but no changes were observed in dynamic modulus and tan delta. As in mature tendons (120 days old), this study showed significant effects of biglycan knockdown on mechanical and structural properties of aged tendons only 30 days after knockdown. These data suggest that biglycan has a major role in maintaining homeostasis in aged tendon.

Effect of fibrillation degree of empty fruit bunch fiber on sandwich composite incorporate with spent mushroom substrate

Sandwich composite panel for heat insulation application were fabricated from two types of agricultural biomass waste, the oil palm empty fruit bunch and spent mushroom substrate. These agricultural biomasses are abundant, renewable, and without proper disposal management, might cause massive environmental pollution. This study investigated the effect of fibrillation degree on EFB fiber through the morphology modification to enhance the properties and thermal characteristic of the sandwich composite panel. Fibrillation is a physical changes that occurred on the surface and internal structure of fiber after undergo excessive physical modification by using refiner machine. Higher fibrillation degree could be achieve by narrowing the space area between two disks of refiner which also known as refining gap. In this research, a composite panel was developed from (SMS) and (EFB) fiber via sandwich hot pressing method. One composite sample using raw EFB fiber and three composite sample at fibrillation degree 191.40%, 211.70% and 271.68% were made at density 0.8 g/cm3. Based on result, the morphological structure of EFB fiber improved in term of formation of small fibril formation, larger surface area, optimum fiber length, optimum fiber diameter and better fiber distribution in composite panel. The mechanical properties of composite was obtained at range 2.77–7.21 MPa for tensile strength, 16.61–18.59 MPa for flexural strength, 2.06–3.18 MPa for internal bond and 4.35–15.79 kJ/m2 for impact strength. For physical properties, value of water absorption and thickness swelling were obtained at range 90.34–142.61% and 18.01–44.80%, respectively. Last but not least, the thermal conductivity value of sandwich composite was obtained at range 0.234–0.282 W/m.K. Overall result found that increasing fibrillation degree on EFB fiber at 211.70% able to contribute in improving the mechanical properties, physical properties and thermal characteristic of sandwich composite panel. This research finding suggested that enhancement of fiber morphology using fibrillation degree approach is considered as an alternative eco-friendly method that could be implemented to improve the properties and thermal characteristic of sandwich composite panel.

Constructing a desired nanofibril network morphology for stretchable polymer films by weakening the intermolecular interaction of a conjugated polymer in an elastomer matrix and extending the film-forming time

Low Ra(b) solvent o-DCB weakens intermolecular interactions of N2200 in the SEBS matrix, leading to the formation of a nanofibril network morphology with a small fibril size (<45 nm), which greatly improves the stretchability of the blend film.

Lignin-Containing Cellulose Nanofibrils from TEMPO-Mediated Oxidation of Date Palm Waste: Preparation, Characterization, and Reinforcing Potential.

Lignin-containing cellulose nanofibrils (LCNFs) have emerged as a new class of nanocelluloses where the presence of residual lignin is expected to impart additional attributes such as hydrophobicity or UV-absorption. In the present work, LCNFs with a lignin content between 7 and 15 wt% were prepared via a TEMPO-mediated oxidation as chemical pretreatment followed by high-pressure homogenization. The impact of the carboxyl content (CC) on the properties of the resulting LCNF gel, in terms of lignin content, colloidal properties, morphology, crystallinity, and thermal stability, were investigated. It was found that lignin content was significantly decreased at increasing CC. In addition, CC had a positive effect on colloidal stability and water contact angle, as well as resulting in smaller fibrils. This lower size, together with the lower lignin content, resulted in a slightly lower thermal stability. The reinforcing potential of the LCNFs when incorporated into a ductile polymer matrix was also explored by preparing nanocomposite films with different LCNF contents that were mechanically tested under linear and non-linear regimes by dynamic mechanical analysis (DMA) and tensile tests. For comparison purposes, the reinforcing effect of the LCNFs with lignin-free CNFs was also reported based on literature data. It was found that lignin hinders the network-forming capacity of LCNFs, as literature data shows a higher reinforcing potential of lignin-free CNFs. Nonetheless, the tensile strength of the acrylic matrix was enhanced by 10-fold at 10 wt% of LCNF content.

Rational Generation of Monoclonal Antibodies Selective for Pathogenic Forms of Alpha-Synuclein.

Misfolded toxic forms of alpha-synuclein (α-Syn) have been implicated in the pathogenesis of synucleinopathies, including Parkinson’s disease (PD), dementia with Lewy bodies (DLB), and multiple system atrophy (MSA). The α-Syn oligomers and soluble fibrils have been shown to mediate neurotoxicity and cell-to-cell propagation of pathology. To generate antibodies capable of selectively targeting pathogenic forms of α-Syn, computational modeling was used to predict conformational epitopes likely to become exposed on oligomers and small soluble fibrils, but not on monomers or fully formed insoluble fibrils. Cyclic peptide scaffolds reproducing these conformational epitopes exhibited neurotoxicity and seeding activity, indicating their biological relevance. Immunization with the conformational epitopes gave rise to monoclonal antibodies (mAbs) with the desired binding profile showing selectivity for toxic α-Syn oligomers and soluble fibrils, with little or no reactivity with monomers, physiologic tetramers, or Lewy bodies. Recognition of naturally occurring soluble α-Syn aggregates in brain extracts from DLB and MSA patients was confirmed by surface plasmon resonance (SPR). In addition, the mAbs inhibited the seeding activity of sonicated pre-formed fibrils (PFFs) in a thioflavin-T fluorescence-based aggregation assay. In neuronal cultures, the mAbs protected primary rat neurons from toxic α-Syn oligomers, reduced the uptake of PFFs, and inhibited the induction of pathogenic phosphorylated aggregates of endogenous α-Syn. Protective antibodies selective for pathogenic species of α-Syn, as opposed to pan α-Syn reactivity, are expected to provide enhanced safety and therapeutic potency by preserving normal α-Syn function and minimizing the diversion of active antibody from the target by the more abundant non-toxic forms of α-Syn in the circulation and central nervous system.

Prediction of Aggregation of Biologically-Active Peptides with the UNRES Coarse-Grained Model.

The UNited RESidue (UNRES) model of polypeptide chains was applied to study the association of 20 peptides with sizes ranging from 6 to 32 amino-acid residues. Twelve of those were potentially aggregating hexa- or heptapeptides excised from larger proteins, while the remaining eight contained potentially aggregating sequences, functionalized by attaching larger ends rich in charged residues. For 13 peptides, the experimental data of aggregation were used. The remaining seven were synthesized, and their properties were measured in this work. Multiplexed replica-exchange simulations of eight-chain systems were conducted at 12 temperatures from 260 to 370 K at concentrations from 0.421 to 5.78 mM, corresponding to the experimental conditions. The temperature profiles of the fractions of monomers and octamers showed a clear transition corresponding to aggregate dissociation. Low simulated transition temperatures were obtained for the peptides, which did not precipitate after incubation, as well as for the H-GNNQQNY-NH2 prion–protein fragment, which forms small fibrils. A substantial amount of inter-strand β-sheets was found in most of the systems. The results suggest that UNRES simulations can be used to assess peptide aggregation except for glutamine- and asparagine-rich peptides, for which a revision of the UNRES sidechain–sidechain interaction potentials appears necessary.

Direct Demonstration of Seed Size-Dependent α-Synuclein Amyloid Amplification.

The size of amyloid seeds is known to modulate their autocatalytic amplification and cellular toxicity. However, the seed size-dependent secondary nucleation mechanism, toxicity, and disease-associated biological processes mediated by α-synuclein (α-Syn) fibrils are largely unknown. Using the cellular model and in vitro reconstitution, we showed that the size of α-Syn fibril seeds dictates not only their cellular internalization and associated cell death but also the distinct mechanisms of fibril amplification pathways involved in the pathological conformational change of α-Syn. Specifically, small fibril seeds showed elongation possibly through monomer addition at the fibril termini, whereas longer fibrils template the fibril amplification by surface-mediated nucleation as demonstrated by super-resolution microscopy. The distinct mechanism of fibril amplification and cellular uptake along with toxicity suggest that breakage of fibrils into seeds of different sizes determines the underlying pathological outcome of synucleinopathies.

Cryo-EM structure of RNA-induced tau fibrils reveals a small C-terminal core that may nucleate fibril formation

In neurodegenerative diseases including Alzheimer’s and amyotrophic lateral sclerosis, proteins that bind RNA are found in aggregated forms in autopsied brains. Evidence suggests that RNA aids nucleation of these pathological aggregates; however, the mechanism has not been investigated at the level of atomic structure. Here, we present the 3.4-Å resolution structure of fibrils of full-length recombinant tau protein in the presence of RNA, determined by electron cryomicroscopy (cryo-EM). The structure reveals the familiar in-register cross-β amyloid scaffold but with a small fibril core spanning residues Glu391 to Ala426, a region disordered in the fuzzy coat in all previously studied tau polymorphs. RNA is bound on the fibril surface to the positively charged residues Arg406 and His407 and runs parallel to the fibril axis. The fibrils dissolve when RNase is added, showing that RNA is necessary for fibril integrity. While this structure cannot exist simultaneously with the tau fibril structures extracted from patients’ brains, it could conceivably account for the nucleating effects of RNA cofactors followed by remodeling as fibrils mature.

Strain rate induced toughening of individual collagen fibrils.

The nonlinear mechanical behavior of individual nanoscale collagen fibrils is governed by molecular stretching and sliding that result in a viscous response, which is still not fully understood. Toward this goal, the in vitro mechanical behavior of individual reconstituted mammalian collagen fibrils was quantified in a broad range of strain-rates, spanning roughly six orders of magnitude, from 10-4 to 35 s-1. It is shown that the nonlinear mechanical response is strain rate sensitive with the tangent modulus in the linear deformation regime increasing monotonically from 214 ± 8 to 358 ± 11 MPa. More pronounced is the effect of the strain rate on the ultimate tensile strength that is found to increase monotonically by a factor of four, from 42 ± 6 to 160 ± 14 MPa. Importantly, fibril strengthening takes place without a reduction in ductility, which results in equivalently large increase in toughness with the increasing strain rate. This experimental strain rate dependent mechanical response is captured well by a structural constitutive model that incorporates the salient features of the collagen microstructure via a process of gradual recruitment of kinked tropocollagen molecules, thus giving rise to the initial "toe-heel" mechanical behavior, followed by molecular stretching and sustained intermolecular slip that is initiated at a strain rate dependent stress threshold. The model shows that the fraction of tropocollagen molecules undergoing straightening increases continuously during loading, whereas molecular sliding is initiated after a small fibril strain (1%-2%) and progressively increases with applied strain.

Monomer recruitment by α-Synuclein fibrils relies on monomer N-terminal interaction

Aggregation of α-Synuclein (αS) into oligomers or mature fibrils in Lewy Bodies in neuronal cells is a hallmark of Parkinson’s Disease (PD). With no effective treatments or cures for PD, the disease outlook is both chronic and progressive. Short αS fibrils and oligomers may propagate cell-to-cell via a seeding mechanism, which involves the healthy cells taking up small fibril fragments that act as templates for further growth. This templated growth requires the recruitment of free monomers, but the forces behind the monomer - fibril interaction are largely unknown.