D-peptide-magnetic nanoparticles fragment tau fibrils and rescue behavioral deficits in a mouse model of Alzheimer's disease.

Amyloid fibrils of tau are increasingly accepted as a cause of neuronal death and brain atrophy in Alzheimer's disease (AD). Diminishing tau aggregation is a promising strategy in the search for efficacious AD therapeutics. Previously, our laboratory designed a six-residue, nonnatural amino acid inhibitor D-TLKIVW peptide (6-DP), which can prevent tau aggregation in vitro. However, it cannot block cell-to-cell transmission of tau aggregation. Here, we find D-TLKIVWC (7-DP), a d-cysteine extension of 6-DP, not only prevents tau aggregation but also fragments tau fibrils extracted from AD brains to neutralize their seeding ability and protect neuronal cells from tau-induced toxicity. To facilitate the transport of 7-DP across the blood-brain barrier, we conjugated it to magnetic nanoparticles (MNPs). The MNPs-DP complex retains the inhibition and fragmentation properties of 7-DP alone. Ten weeks of MNPs-DP treatment appear to reverse neurological deficits in the PS19 mouse model of AD. This work offers a direction for development of therapies to target tau fibrils.

How short peptides can disassemble ultra-stable tau fibrils extracted from Alzheimer's disease brain by a strain-relief mechanism.

Reducing fibrous aggregates of protein tau is a possible strategy for halting progression of Alzheimer's dis-ease (AD). Previously we found that in vitro the D-peptide D-TLKIVWC disassembles tau fibrils from AD brains (AD-tau) into benign segments with no energy source present beyond ambient thermal agitation. This disassembly by a short peptide was unexpected, given that AD-tau is sufficiently stable to withstand disas-sembly in boiling SDS detergent. To consider D peptide-mediated disassembly as a potential therapeutic for AD, it is essential to understand the mechanism and energy source of the disassembly action. We find as-sembly of D-peptides into amyloid-like fibrils is essential for tau fibril disassembly. Cryo-EM and atomic force microscopy reveal that these D-peptide fibrils have a right-handed twist and embrace tau fibrils which have a left-handed twist. In binding to the AD-tau fibril, the oppositely twisted D-peptide fibril produces a strain, which is relieved by the disassembly of both fibrils. This strain-relief mechanism appears to operate in other examples of amyloid fibril disassembly and provides a new direction for the development of first-in-class therapeutics for amyloid diseases.

How short peptides can disassemble ultra-stable tau fibrils extracted from Alzheimer's disease brain by a strain-relief mechanism.

Reducing fibrous aggregates of protein tau is a possible strategy for halting progression of Alzheimer's disease (AD). Previously we found that in vitro the D-peptide D-TLKIVWC disassembles tau fibrils from AD brains (AD-tau) into benign segments with no energy source present beyond ambient thermal agitation. This disassembly by a short peptide was unexpected, given that AD-tau is sufficiently stable to withstand disassembly in boiling SDS detergent. To consider D peptide-mediated disassembly as a potential therapeutic for AD, it is essential to understand the mechanism and energy source of the disassembly action. We find assembly of D-peptides into amyloid-like fibrils is essential for tau fibril disassembly. Cryo-EM and atomic force microscopy reveal that these D-peptide fibrils have a right-handed twist and embrace tau fibrils which have a left-handed twist. In binding to the AD-tau fibril, the oppositely twisted D-peptide fibril produces a strain, which is relieved by disassembly of both fibrils. This strain-relief mechanism appears to operate in other examples of amyloid fibril disassembly and provides a new direction for the development of first-in-class therapeutics for amyloid diseases.

Structural polymorphism of amyloid fibrils in ATTR amyloidosis revealed by cryo-electron microscopy.

ATTR amyloidosis is caused by the deposition of transthyretin in the form of amyloid fibrils in virtually every organ of the body, including the heart. This systemic deposition leads to a phenotypic variability that has not been molecularly explained yet. In brain amyloid conditions, previous studies suggest an association between clinical phenotype and the molecular structures of their amyloid fibrils. Here we investigate whether there is such an association in ATTRv amyloidosis patients carrying the mutation I84S. Using cryo-electron microscopy, we determined the structures of cardiac fibrils extracted from three ATTR amyloidosis patients carrying the ATTRv-I84S mutation, associated with a consistent clinical phenotype. We found that in each ATTRv-I84S patient, the cardiac fibrils exhibited different local conformations, and these variations can co-exist within the same fibril. Our finding suggests that one amyloid disease may associate with multiple fibril structures in systemic amyloidoses, calling for further studies.

Cryo-EM structures of the D290V mutant of the hnRNPA2 low-complexity domain suggests how D290V affects phase separation and aggregation.

Heterogeneous nuclear ribonucleoprotein A2 (hnRNPA2) is a human ribonucleoprotein that transports RNA to designated locations for translation via its ability to phase separate. Its mutated form, D290V, is implicated in multisystem proteinopathy known to afflict two families, mainly with myopathy and Paget's disease of bone. Here, we investigate this mutant form of hnRNPA2 by determining cryo-EM structures of the recombinant D290V low complexity domain. We find that the mutant form of hnRNPA2 differs from the WT fibrils in four ways. In contrast to the WT fibrils, the PY-nuclear localization signals in the fibril cores of all three mutant polymorphs are less accessible to chaperones. Also, the mutant fibrils are more stable than WT fibrils as judged by phase separation, thermal stability, and energetic calculations. Similar to other pathogenic amyloids, the mutant fibrils are polymorphic. Thus, these structures offer evidence to explain how a D-to-V missense mutation diverts the assembly of reversible, functional amyloid-like fibrils into the assembly of pathogenic amyloid, and may shed light on analogous conversions occurring in other ribonucleoproteins that lead to neurological diseases such as amyotrophic lateral sclerosis and frontotemporal dementia.

CT imaging of intrauterine devices (IUD): expected findings, unexpected findings, and complications.

Intrauterine devices (IUDs) are a commonly used form of long-acting reversible contraception, which either contain copper or levonorgestrel to prevent pregnancy. Although symptomatic patients with indwelling IUDs may first undergo ultrasound to assess for device malposition and complications, IUDs are commonly encountered on CT in patients undergoing evaluation for unrelated indications. Frequently, IUD malposition and complications may be asymptomatic or clinically unsuspected. For these reasons, it is important for the radiologist to carefully scrutinize the IUD on any study in which it is encountered. To do so, the radiologist must recognize that normally positioned IUDs are located centrally within the uterine cavity. IUDs are extremely effective in preventing pregnancy, though inadvertent pregnancy risk is higher with malpositioned IUDs. Presence of fibroids or Mullerian abnormalities may preclude proper IUD placement. Radiologists play an important role in identifying complications when they arise and special considerations when planning for an IUD placement. There is a wide range of IUD malposition, affecting IUDs differently depending on the type of IUD and its mechanism of action. IUD malposition is the most common complication, but embedment and/or partial perforation can and can lead to difficulty when removed. Retained IUD fragments can result in continued contraceptive effect. Perforated IUDs do not typically cause intraperitoneal imaging findings.

Fibril structures of TFG protein mutants validate the identification of TFG as a disease-related amyloid protein by the IMPAcT method.

We previously presented a bioinformatic method for identifying diseases that arise from a mutation in a protein's low-complexity domain that drives the protein into pathogenic amyloid fibrils. One protein so identified was the tropomyosin-receptor kinase-fused gene protein (TRK-fused gene protein or TFG). Mutations in TFG are associated with degenerative neurological conditions. Here, we present experimental evidence that confirms our prediction that these conditions are amyloid-related. We find that the low-complexity domain of TFG containing the disease-related mutations G269V or P285L forms amyloid fibrils, and we determine their structures using cryo-electron microscopy (cryo-EM). These structures are unmistakably amyloid in nature and confirm the propensity of the mutant TFG low-complexity domain to form amyloid fibrils. Also, despite resulting from a pathogenic mutation, the fibril structures bear some similarities to other amyloid structures that are thought to be nonpathogenic and even functional, but there are other factors that support these structures' relevance to disease, including an increased propensity to form amyloid compared with the wild-type sequence, structure-stabilizing influence from the mutant residues themselves, and double-protofilament amyloid cores. Our findings elucidate two potentially disease-relevant structures of a previously unknown amyloid and also show how the structural features of pathogenic amyloid fibrils may not conform to the features commonly associated with pathogenicity.

Feasibility pilot study of a Japanese teaching kitchen program.

Background: This pilot study examined the feasibility of a new lifestyle modification program involving a "Teaching Kitchen" in Japan. Our goal was to explore (1) feasibility of the program; (2) acceptability for class frequency (weekly vs. bi-weekly); and (3) changes in biometrics, dietary intakes, and lifestyle factors. Methods: A total of 24 employees with obesity in a Japanese company were recruited. Participants were randomly divided into two groups (weekly or bi-weekly group), each attending the program consisting of four two-hour classes (lectures on nutrition, exercise, mindfulness, and culinary instructions). Participants were observed for changes in dietary intakes, biometrics, and health related quality of life over the subsequent 3 months. We tested the between-group differences in changes using linear mixed-effect models. Results: The program completion rates were 83.3% in total (91.7% for weekly group and 75.0% for bi-weekly group). From baseline to post-intervention, significant decreases were observed in weight (p < 0.001), body mass index (p < 0.001), diastolic blood pressure (p = 0.03), body fat mass (p < 0.001), and dietary intakes in total fat (p = 0.03) and sodium (p = 0.008) among 17 participants who were available for measurements. Improvements in biometrics remained significant 1 month after the intervention (all p ≤ 0.03 in 14 participants). Participants' health related quality of life was significantly improved in bodily pain, general health, vitality, and mental component score (all p ≤ 0.047). Conclusions: The new Japanese Teaching Kitchen program is feasible with high program completion rates in Japanese office workers with obesity. While this was a small feasibility study, significant multiple improvements in dietary intakes, biometrics, and health related quality of life suggest that this line of inquiry warrants further exploration to address obesity and obesity-related diseases in Japan.

Racemic Peptides from Amyloid ß and Amylin Form Rippled ß-Sheets Rather Than Pleated ß-Sheets.

The rippled ß-sheet was theorized by Pauling and Corey in 1953 as a structural motif in which mirror image peptide strands assemble into hydrogen-bonded periodic arrays with strictly alternating chirality. Structural characterization of the rippled ß-sheet was limited to biophysical methods until 2022 when atomic resolution structures of the motif were first obtained. The crystal structural foundation is restricted to four model tripeptides composed exclusively of aromatic residues. Here, we report five new rippled sheet crystal structures derived from amyloid ß and amylin, the aggregating toxic peptides of Alzheimer's disease and type II diabetes, respectively. Despite the variation in peptide sequence composition, all five structures form antiparallel rippled ß-sheets that extend, like a fibril, along the entire length of the crystalline needle. The long-range packing of the crystals, however, varies. In three of the crystals, the sheets pack face-to-face and exclude water, giving rise to cross-ß architectures grossly resembling the steric zipper motif of amyloid fibrils but differing in fundamental details. In the other two crystals, the solvent is encapsulated between the sheets, yielding fibril architectures capable of host-guest chemistry. Our study demonstrates that the formation of rippled ß-sheets from aggregating racemic peptide mixtures in three-dimensional (3D) assemblies is a general phenomenon and provides a structural basis for targeting intrinsically disordered proteins.

Low grade endometrial stromal sarcoma in pregnancy: A case in support of safe abortion access.

Background: Endometrial stromal sarcoma (ESS) is extremely rare in pregnancy. It shares clinical and imaging features with more common pregnancy findings such as leiomyoma and molar gestations, which makes diagnosis challenging. Case: A 36-year-old patient presented at 8 weeks and 1 day gestation for vaginal bleeding. An intrauterine pregnancy with an appropriately sized embryo with heart motion and a 9.5 cm complex uterine mass was found on ultrasound. MRI showed an 11.4 cm cystic mass with nodular septations causing mass effect on the endometrial cavity. After extensive counseling, the patient underwent a gravid abdominal hysterectomy and bilateral salpingectomy. Final pathology showed low grade ESS. Conclusion: This case highlights the importance of evaluating suspicious uterine masses in pregnancy and the necessity for safe abortion access.

Structure-based design of nanobodies that inhibit seeding of Alzheimer's patient-extracted tau fibrils.

Despite much effort, antibody therapies for Alzheimer's disease (AD) have shown limited efficacy. Challenges to the rational design of effective antibodies include the difficulty of achieving specific affinity to critical targets, poor expression, and antibody aggregation caused by buried charges and unstructured loops. To overcome these challenges, we grafted previously determined sequences of fibril-capping amyloid inhibitors onto a camel heavy chain antibody scaffold. These sequences were designed to cap fibrils of tau, known to form the neurofibrillary tangles of AD, thereby preventing fibril elongation. The nanobodies grafted with capping inhibitors blocked tau aggregation in biosensor cells seeded with postmortem brain extracts from AD and progressive supranuclear palsy (PSP) patients. The tau capping nanobody inhibitors also blocked seeding by recombinant tau oligomers. Another challenge to the design of effective antibodies is their poor blood-brain barrier (BBB) penetration. In this study, we also designed a bispecific nanobody composed of a nanobody that targets a receptor on the BBB and a tau capping nanobody inhibitor, conjoined by a flexible linker. We provide evidence that the bispecific nanobody improved BBB penetration over the tau capping inhibitor alone after intravenous administration in mice. Our results suggest that the design of synthetic antibodies that target sequences that drive protein aggregation may be a promising approach to inhibit the prion-like seeding of tau and other proteins involved in AD and related proteinopathies.

Standard Patient History Can Be Augmented Using Ethnographic Foodlife Questions.

The relationship between what and how individuals eat and their overall and long-term health is non-controversial. However, for decades, food and nutrition discussions have often been highly medicalized. Given the significant impact of poor nutrition on health, broader discussions about food should be integrated into routine patient history taking. We advocate for an expansion of the current, standard approach to patient history taking in order to include questions regarding patients' 'foodlife' (total relationship to food) as a screening and baseline assessment tool for referrals. We propose that healthcare providers: (1) routinely engage with patients about their relationship to food, and (2) recognize that such dialogues extend beyond nutrition and lifestyle questions. Mirroring other recent revisions to medical history taking-such as exploring biopsychosocial risks-questions about food relationships and motivators of eating may be essential for optimal patient assessment and referrals. We draw on the novel tools of 'foodlife' ethnography (developed by co-author ethnographer J.J.L., and further refined in collaboration with the co-authors who contributed their clinical experiences as a former primary care physician (D.M.E.), registered dietitian (J.W.M.), and diabetologist (H.Z.)) to model a set of baseline questions for inclusion in routine clinical settings. Importantly, this broader cultural approach seeks to augment and enhance current food intake discussions used by registered dietitian nutritionists, endocrinologists, internists, and medical primary care providers for better baseline assessments and referrals. By bringing the significance of food into the domain of routine medical interviewing practices by a range of health professionals, we theorize that this approach can set a strong foundation of trust between patients and healthcare professionals, underscoring food's vital role in patient-centered care.

Characteristics of Current Teaching Kitchens: Findings from Recent Surveys of the Teaching Kitchen Collaborative.

Teaching kitchens are physical and virtual forums that foster practical life skills through participation in experiential education. Given the well-supported connection between healthy eating patterns and the prevention and management of chronic diseases, both private and public organizations are building teaching kitchens (TKs) to enhance the health and wellness of patients, staff, youth, and the general community. Although implementation of TKs is becoming more common, best practices for starting and operating programs are limited. The present study aims to describe key components and professionals required for TK operations. Surveys were administered to Teaching Kitchen Collaborative (TKC) members and questions reflected seven primary areas of inquiry: (1) TK setting(s), (2) audiences served, (3) TK model(s), (4) key lines of operations, (5) team member who manages or directs the TK, (6) team member(s) who performs key operations and other professionals or partnerships that may be needed, and (7) the primary funding source(s) to build and operate the TK (among various other topics). Findings were used to articulate recommendations for organizations seeking to establish a successful TK as well as for TKs to expand their collective reach, research capacity, and impact.

CRISPR screens in iPSC-derived neurons reveal principles of tau proteostasis.

A hallmark of age-associated neurodegenerative diseases is the aggregation of proteins. Aggregation of the protein tau defines tauopathies, which include Alzheimer's disease and frontotemporal dementia. Specific neuronal subtypes are selectively vulnerable to the accumulation of tau aggregates, and subsequent dysfunction and death. The mechanisms underlying cell type-selective vulnerability are unknown. To systematically uncover the cellular factors controlling the accumulation of tau aggregates in human neurons, we conducted a genome-wide CRISPRi-based modifier screen in iPSC-derived neurons. The screen uncovered expected pathways, including autophagy, but also unexpected pathways including UFMylation and GPI anchor synthesis, that control tau oligomer levels. We identify the E3 ubiquitin ligase CUL5 as a tau interactor and potent modifier of tau levels. In addition, disruption of mitochondrial function increases tau oligomer levels and promotes proteasomal misprocessing of tau. These results reveal new principles of tau proteostasis in human neurons and pinpoint potential therapeutic targets for tauopathies.

Perspective: Teaching Kitchens: Conceptual Origins, Applications and Potential for Impact within Food Is Medicine Research.

There is a need to identify innovative strategies whereby individuals, families, and communities can learn to access and prepare affordable and nutritious foods, in combination with evidence-based guidance about diet and lifestyle. These approaches also need to address issues of equity and sustainability. Teaching Kitchens (TKs) are being created as educational classrooms and translational research laboratories to advance such strategies. Moreover, TKs can be used as revenue-generating research sites in universities and hospitals performing sponsored research, and, potentially, as instruments of cost containment when placed in accountable care settings and self-insured companies. Thus, TKs can be considered for inclusion in future health professional training programs, and the recently published Biden-Harris Administration Strategy on Hunger, Nutrition and Health echoes this directive. Recent innovations in the ability to provide TK classes virtually suggest that their impact may be greater than originally envisioned. Although the impact of TK curricula on behaviors, outcomes and costs of health care is preliminary, it warrants the continued attention of medical and public health thought leaders involved with Food Is Medicine initiatives.

Low complexity domains of the nucleocapsid protein of SARS-CoV-2 form amyloid fibrils.

The self-assembly of the Nucleocapsid protein (NCAP) of SARS-CoV-2 is crucial for its function. Computational analysis of the amino acid sequence of NCAP reveals low-complexity domains (LCDs) akin to LCDs in other proteins known to self-assemble as phase separation droplets and amyloid fibrils. Previous reports have described NCAP's propensity to phase-separate. Here we show that the central LCD of NCAP is capable of both, phase separation and amyloid formation. Within this central LCD we identified three adhesive segments and determined the atomic structure of the fibrils formed by each. Those structures guided the design of G12, a peptide that interferes with the self-assembly of NCAP and demonstrates antiviral activity in SARS-CoV-2 infected cells. Our work, therefore, demonstrates the amyloid form of the central LCD of NCAP and suggests that amyloidogenic segments of NCAP could be targeted for drug development.

Heavy Menstrual Bleeding Treatment With a Levonorgestrel 52-mg Intrauterine Device.

OBJECTIVE: To evaluate heavy menstrual bleeding treatment outcomes with levonorgestrel 52-mg intrauterine device (IUD) use in participants without body mass index (BMI) or parity restrictions. METHODS: Investigators included participants aged 18-50 years with no pelvic or systemic pathology causing heavy menstrual bleeding at 29 U.S. centers in a prospective trial. Participants had up to three screening cycles with menstrual product collection for alkaline hematin blood-loss measurements. Investigators enrolled those with two menses with blood loss of 80 mL or more (values averaged for baseline blood loss), placed the IUD, and followed participants for up to six 28-day cycles. Participants collected any menstrual products used during cycles 3 and 6 for blood-loss measurement. We evaluated outcomes in participants with at least one follow-up assessment for the primary outcome of median absolute blood-loss change and, secondarily, treatment success , defined as the proportion with a final measured blood loss less than 80 mL and at least 50% reduction from baseline. We evaluated exploratory outcomes of differences in blood-loss changes by BMI and parity using Wilcoxon rank sum test. RESULTS: Of 105 enrolled participants, 47 (44.8%) had obesity (BMI 30.0 or higher) and 29 (27.6%) were nulliparous. Baseline mean blood loss ranged from 73 to 520 mL (median 143 mL, interquartile range 112-196 mL). Eighty-nine (84.8%) had at least one evaluable follow-up evaluation. Participants had median (interquartile range) absolute blood-loss decreases at cycles 3 (n=86) and 6 (n=81) of 93.3% (86.1-97.7%) and 97.6% (90.4-100%), respectively. At cycle 6, participants without obesity (n=43) and with obesity (n=38) had similar median [interquartile range] decreases (97.6% [91.8-100%] and 97.5% [90.3-100%], respectively; P =.89), with comparable findings for nulliparous (n=25) and parous (n=56) participants (97.0% [91.7-99.1%] and 98.1% [89.9-100%], respectively; P =.43). Treatment success occurred in 81.8% (95% CI 74.2-89.4%) of 99 participants, excluding those with no outcomes due to lost to follow-up or consent withdrawal, and did not vary by BMI or parity. The most common adverse events leading to discontinuation were bleeding or cramping (n=6 [5.7%]) and expulsion (n=5 [4.8%]). CONCLUSION: This levonorgestrel 52-mg IUD reduces blood loss by more than 90% over 6 months compared with baseline for most users with heavy menstrual bleeding. FUNDING SOURCE: Medicines360. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov , NCT03642210.

Small molecules disaggregate alpha-synuclein and prevent seeding from patient brain-derived fibrils.

The amyloid aggregation of alpha-synuclein within the brain is associated with the pathogenesis of Parkinson's disease (PD) and other related synucleinopathies, including multiple system atrophy (MSA). Alpha-synuclein aggregates are a major therapeutic target for treatment of these diseases. We identify two small molecules capable of disassembling preformed alpha-synuclein fibrils. The compounds, termed CNS-11 and CNS-11g, disaggregate recombinant alpha-synuclein fibrils in vitro, prevent the intracellular seeded aggregation of alpha-synuclein fibrils, and mitigate alpha-synuclein fibril cytotoxicity in neuronal cells. Furthermore, we demonstrate that both compounds disassemble fibrils extracted from MSA patient brains and prevent their intracellular seeding. They also reduce in vivo alpha-synuclein aggregates in C. elegans. Both compounds also penetrate brain tissue in mice. A molecular dynamics-based computational model suggests the compounds may exert their disaggregating effects on the N terminus of the fibril core. These compounds appear to be promising therapeutic leads for targeting alpha-synuclein for the treatment of synucleinopathies.

Amyloid nomenclature 2022: update, novel proteins, and recommendations by the International Society of Amyloidosis (ISA) Nomenclature Committee.

The Nomenclature Committee of the International Society of Amyloidosis met at the XVIII International Symposium on Amyloidosis in September and virtually in October 2022 with discussions resulting in this upgraded nomenclature recommendation. The nomenclature principles remain unchanged but there is an ongoing discussion regarding the importance and varying nature of intracellular protein aggregates, particularly those associated with neurodegenerative diseases. Six novel proteins were added to the list of human amyloid fibril proteins. Of these, three are polypeptide hormones and two currently utilised peptide drugs, making the number of known iatrogenic amyloid forms four, all appearing as subcutaneous nodules at the injection site. The sixth novel amyloid fibril protein is the transmembrane 106B protein, forming intracellular amyloid fibrils in disorders associated with frontotemporal dementia. The number of known human amyloid fibril proteins is now 42.