In Vivo Tissue Distribution of Polystyrene or Mixed Polymer Microspheres and Metabolomic Analysis after Oral Exposure in Mice.

BACKGROUND: Global plastic use has consistently increased over the past century with several different types of plastics now being produced. Much of these plastics end up in oceans or landfills leading to a substantial accumulation of plastics in the environment. Plastic debris slowly degrades into microplastics (MPs) that can ultimately be inhaled or ingested by both animals and humans. A growing body of evidence indicates that MPs can cross the gut barrier and enter into the lymphatic and systemic circulation leading to accumulation in tissues such as the lungs, liver, kidney, and brain. The impacts of mixed MPs exposure on tissue function through metabolism remains largely unexplored. OBJECTIVES: This study aims to investigate the impacts of polymer microspheres on tissue metabolism in mice by assessing the microspheres ability to translocate across the gut barrier and enter into systemic circulation. Specifically, we wanted to examine microsphere accumulation in different organ systems, identify concentration-dependent metabolic changes, and evaluate the effects of mixed microsphere exposures on health outcomes. METHODS: To investigate the impact of ingested microspheres on target metabolic pathways, mice were exposed to either polystyrene (5µm) microspheres or a mixture of polymer microspheres consisting of polystyrene (5µm), polyethylene (1-4µm), and the biodegradability and biocompatible plastic, poly-(lactic-co-glycolic acid) (5µm). Exposures were performed twice a week for 4 weeks at a concentration of either 0, 2, or 4mg/week via oral gastric gavage. Tissues were collected to examine microsphere ingress and changes in metabolites. RESULTS: In mice that ingested microspheres, we detected polystyrene microspheres in distant tissues including the brain, liver, and kidney. Additionally, we report on the metabolic differences that occurred in the colon, liver, and brain, which showed differential responses that were dependent on concentration and type of microsphere exposure. DISCUSSION: This study uses a mouse model to provide critical insight into the potential health implications of the pervasive issue of plastic pollution. These findings demonstrate that orally consumed polystyrene or mixed polymer microspheres can accumulate in tissues such as the brain, liver, and kidney. Furthermore, this study highlights concentration-dependent and polymer type-specific metabolic changes in the colon, liver, and brain after plastic microsphere exposure. These results underline the mobility within and between biological tissues of MPs after exposure and emphasize the importance of understanding their metabolic impact. https://doi.org/10.1289/EHP13435.

Ethnic variation and structure-function analysis of tauopathy-associated PERK alleles.

EIF2AK3, also known as PERK, plays a pivotal role in cellular proteostasis, orchestrating the Unfolded Protein Response (UPR) and Integrated Stress Response (ISR) pathways. In addition to its central position in intracellular stress regulation, human GWAS identify EIF2AK3 as a risk factor in tauopathies, neurodegenerative diseases caused by aberrant tau protein accumulation. Guided by these genomic indicators, our investigation systematically analyzed human PERK variants, focusing on those with potential tauopathy linkages. We assembled a comprehensive data set of human PERK variants associated with Wolcott Rallison Syndrome (WRS), tauopathies, and bioinformatically predicted loss-of-function, referencing the gnomAD, Ensembl, and NCBI databases. We found extensive racial/ethnic variation in the prevalence of common PERK polymorphisms linked to tauopathies. Using SWISS-MODEL, we identified structural perturbations in the ER stress-sensing luminal domain dimers/oligomers of tauopathy-associated PERK variants, Haplotypes A and B, in combination with another tauopathy-linked R240H mutation. Recombinant expression of disease-associated variants in vitro revealed altered PERK signal transduction kinetics in response to ER stress compared to the predominant non-disease variant. In summary, our data further substantiates that human PERK variants identified in tauopathy genetic studies negatively impact PERK structure, function, and downstream signaling with significant variations in prevalence among different racial and ethnic groups.

FLASH Proton Radiation Therapy Mitigates Inflammatory and Fibrotic Pathways and Preserves Cardiac Function in a Preclinical Mouse Model of Radiation-Induced Heart Disease.

PURPOSE: Studies during the past 9 years suggest that delivering radiation at dose rates exceeding 40 Gy/s, known as "FLASH" radiation therapy, enhances the therapeutic index of radiation therapy (RT) by decreasing normal tissue damage while maintaining tumor response compared with conventional (or standard) RT. This study demonstrates the cardioprotective benefits of FLASH proton RT (F-PRT) compared with standard (conventional) proton RT (S-PRT), as evidenced by reduced acute and chronic cardiac toxicities. METHODS AND MATERIALS: Mice were imaged using cone beam computed tomography to precisely determine the heart's apex as the beam isocenter. Irradiation was conducted using a shoot-through technique with a 5-mm diameter circular collimator. Bulk RNA-sequencing was performed on nonirradiated samples, as well as apexes treated with F-PRT or S-PRT, at 2 weeks after a single 40 Gy dose. Inflammatory responses were assessed through multiplex cytokine/chemokine microbead assay and immunofluorescence analyses. Levels of perivascular fibrosis were quantified using Masson's Trichrome and Picrosirius red staining. Additionally, cardiac tissue functionality was evaluated by 2-dimensional echocardiograms at 8- and 30-weeks post-PRT. RESULTS: Radiation damage was specifically localized to the heart's apex. RNA profiling of cardiac tissues treated with PRT revealed that S-PRT uniquely upregulated pathways associated with DNA damage response, induction of tumor necrosis factor superfamily, and inflammatory response, and F-PRT primarily affected cytoplasmic translation, mitochondrion organization, and adenosine triphosphate synthesis. Notably, F-PRT led to a milder inflammatory response, accompanied by significantly attenuated changes in transforming growth factor ß1 and α smooth muscle actin levels. Critically, F-PRT decreased collagen deposition and better preserved cardiac functionality compared with S-PRT. CONCLUSIONS: This study demonstrated that F-PRT reduces the induction of an inflammatory environment with lower expression of inflammatory cytokines and profibrotic factors. Importantly, the results indicate that F-PRT better preserves cardiac functionality, as confirmed by echocardiography analysis, while also mitigating the development of long-term fibrosis.

In Vivo Tissue Distribution of Microplastics and Systemic Metabolomic Alterations After Gastrointestinal Exposure.

Global plastic use has consistently increased over the past century with several different types of plastics now being produced. Much of these plastics end up in oceans or landfills leading to a substantial accumulation of plastics in the environment. Plastic debris slowly degrades into microplastics (MPs) that can ultimately be inhaled or ingested by both animals and humans. A growing body of evidence indicates that MPs can cross the gut barrier and enter into the lymphatic and systemic circulation leading to accumulation in tissues such as the lungs, liver, kidney, and brain. The impacts of mixed MPs exposure on tissue function through metabolism remains largely unexplored. To investigate the impact of ingested MPs on target metabolomic pathways, mice were subjected to either polystyrene microspheres or a mixed plastics (5 µm) exposure consisting of polystyrene, polyethylene and the biodegradability and biocompatible plastic, poly-(lactic-co-glycolic acid). Exposures were performed twice a week for four weeks at a dose of either 0, 2, or 4 mg/week via oral gastric gavage. Our findings demonstrate that, in mice, ingested MPs can pass through the gut barrier, be translocated through the systemic circulation, and accumulate in distant tissues including the brain, liver, and kidney. Additionally, we report on the metabolomic changes that occur in the colon, liver and brain which show differential responses that are dependent on dose and type of MPs exposure. Lastly, our study provides proof of concept for identifying metabolomic alterations associated with MPs exposure and adds insight into the potential health risks that mixed MPs contamination may pose to humans.

Neurodegeneration risk factor, EIF2AK3 (PERK), influences tau protein aggregation.

Tauopathies are neurodegenerative diseases caused by pathologic misfolded tau protein aggregation in the nervous system. Population studies implicate EIF2AK3 (eukaryotic translation initiation factor 2 alpha kinase 3), better known as PERK (protein kinase R-like endoplasmic reticulum kinase), as a genetic risk factor in several tauopathies. PERK is a key regulator of intracellular proteostatic mechanisms-unfolded protein response and integrated stress response. Previous studies found that tauopathy-associated PERK variants encoded functional hypomorphs with reduced signaling in vitro. But, it remained unclear how altered PERK activity led to tauopathy. Here, we chemically or genetically modulated PERK signaling in cell culture models of tau aggregation and found that PERK pathway activation prevented tau aggregation, whereas inhibition exacerbated tau aggregation. In primary tauopathy patient brain tissues, we found that reduced PERK signaling correlated with increased tau neuropathology. We found that tauopathy-associated PERK variants targeted the endoplasmic reticulum luminal domain; and two of these variants damaged hydrogen bond formation. Our studies support that PERK activity protects against tau aggregation and pathology. This may explain why people carrying hypomorphic PERK variants have increased risk for developing tauopathies. Finally, our studies identify small-molecule augmentation of PERK signaling as an attractive therapeutic strategy to treat tauopathies by preventing tau pathology.

Network biology analysis of P23H rhodopsin interactome identifies protein and mRNA quality control mechanisms.

Rhodopsin is essential for phototransduction, and many rhodopsin mutations cause heritable retinal degenerations. The P23H rhodopsin variant generates a misfolded rhodopsin protein that photoreceptors quickly target for degradation by mechanisms that are incompletely understood. To gain insight into how P23H rhodopsin is removed from rods, we used mass spectrometry to identify protein interaction partners of P23H rhodopsin immunopurified from RhoP23H/P23H mice and compared them with protein interaction partners of wild-type rhodopsin from Rho+/+ mice. We identified 286 proteins associated with P23H rhodopsin and 276 proteins associated with wild-type rhodopsin. 113 proteins were shared between wild-type and mutant rhodopsin protein interactomes. In the P23H rhodopsin protein interactome, we saw loss of phototransduction, retinal cycle, and rhodopsin protein trafficking proteins but gain of ubiquitin-related proteins when compared with the wild-type rhodopsin protein interactome. In the P23H rhodopsin protein interactome, we saw enrichment of gene ontology terms related to ER-associated protein degradation, ER stress, and translation. Protein-protein interaction network analysis revealed that translational and ribosomal quality control proteins were significant regulators in the P23H rhodopsin protein interactome. The protein partners identified in our study may provide new insights into how photoreceptors recognize and clear mutant rhodopsin, offering possible novel targets involved in retinal degeneration pathogenesis.

A stromal Integrated Stress Response activates perivascular cancer-associated fibroblasts to drive angiogenesis and tumour progression.

Bidirectional signalling between the tumour and stroma shapes tumour aggressiveness and metastasis. ATF4 is a major effector of the Integrated Stress Response, a homeostatic mechanism that couples cell growth and survival to bioenergetic demands. Using conditional knockout ATF4 mice, we show that global, or fibroblast-specific loss of host ATF4, results in deficient vascularization and a pronounced growth delay of syngeneic melanoma and pancreatic tumours. Single-cell transcriptomics of tumours grown in Atf4Δ/Δ mice uncovered a reduction in activation markers in perivascular cancer-associated fibroblasts (CAFs). Atf4Δ/Δ fibroblasts displayed significant defects in collagen biosynthesis and deposition and a reduced ability to support angiogenesis. Mechanistically, ATF4 regulates the expression of the Col1a1 gene and levels of glycine and proline, the major amino acids of collagen. Analyses of human melanoma and pancreatic tumours revealed a strong correlation between ATF4 and collagen levels. Our findings establish stromal ATF4 as a key driver of CAF functionality, malignant progression and metastasis.

An Overview of Online Resources for Medical Spanish Education for Effective Communication with Spanish-Speaking Patients.

PHENOMENON: Despite the rapid development of virtual medical Spanish educational materials, online resources lack transparency and a peer-review process. The purpose of this interdisciplinary study was to provide a critical inventory of virtual resources for medical Spanish education, thereby providing a panorama of the current state of online medical Spanish. APPROACH: Research team members conducted iterative searches to identify medical Spanish online resources, which were then screened for predetermined inclusion/exclusion criteria. Between June and August 2020, a panel of medical and language experts then adapted and applied a previously published evaluation tool to determine whether resources that met study criteria would help learners achieve medical Spanish core competencies and to what extent each resource incorporated communicative language activities. Consensus meetings were conducted to resolve disagreements and identify gaps in online education. FINDINGS: Out of 465 resources, 127 were further screened, and eight were selected for evaluation. Medical and language specialists independently scored each resource and, following discussions, achieved consensus. Overall, no resource met suitability criteria for all five medical Spanish learner competencies or cultural elements, and only one was suitable for achieving the self-assessment competency. INSIGHTS: Interdisciplinary consensus meetings provide an important avenue for resolving differences of opinion and for integrating both language and medical perspectives into the evaluation process. Existing online resources should be used in conjunction with other materials to ensure that all core competencies for medical Spanish education are addressed. This study revealed important gaps in online resources, including a need to target advanced Spanish learners, apply authentic communicative activities, include assessment opportunities, and integrate culture in the learning program. Based on the current state of online medical Spanish, we offer recommendations for future resources.

ATF6 is required for efficient rhodopsin clearance and retinal homeostasis in the P23H rho retinitis pigmentosa mouse model.

Retinitis Pigmentosa (RP) is a blinding disease that arises from loss of rods and subsequently cones. The P23H rhodopsin knock-in (P23H-KI) mouse develops retinal degeneration that mirrors RP phenotype in patients carrying the orthologous variant. Previously, we found that the P23H rhodopsin protein was degraded in P23H-KI retinas, and the Unfolded Protein Response (UPR) promoted P23H rhodopsin degradation in heterologous cells in vitro. Here, we investigated the role of a UPR regulator gene, activating transcription factor 6 (Atf6), in rhodopsin protein homeostasis in heterozygous P23H rhodopsin (Rho+/P23H) mice. Significantly increased rhodopsin protein levels were found in Atf6-/-Rho+/P23H retinas compared to Atf6+/-Rho+/P23H retinas at early ages (~ P12), while rhodopsin mRNA levels were not different. The IRE1 pathway of the UPR was hyper-activated in young Atf6-/-Rho+/P23H retinas, and photoreceptor layer thickness was unchanged at this early age in Rho+/P23H mice lacking Atf6. By contrast, older Atf6-/-Rho+/P23H mice developed significantly increased retinal degeneration in comparison to Atf6+/-Rho+/P23H mice in all retinal layers, accompanied by reduced rhodopsin protein levels. Our findings demonstrate that Atf6 is required for efficient clearance of rhodopsin protein in rod photoreceptors expressing P23H rhodopsin, and that loss of Atf6 ultimately accelerates retinal degeneration in P23H-KI mice.

Elucidating the effect of enzymatic polymerized polysaccharide particle morphology on emulsion properties.

Exploiting the shape of Pickering stabilizers offers the ability to unlock the full potential of nanoparticle-stabilized emulsions for applications in enhanced oil recovery, pharmaceuticals, cosmetics, and coatings. In this work, we utilize engineered polysaccharide particles derived from the enzymatic polymerization of glucose from sucrose with controlled shape for the stabilization of dodecane-in-water emulsions. Altering the particle shape (spherical aggregates, fibrids, or platelets), while maintaining a neutral surface charge allows for a systematic examination of the role of particle shape in the stabilization of emulsions. We find that platelet-shaped particles reduce the interfacial tension and result in the smallest droplet size, while emulsions stabilized by aggregates and fibrids are governed by a network of particles in the continuous phase. Exploiting the synergy between these particles allowed for the tuning of their microstructure and rheological signature which allows us to map and tailor these emulsions for a wider variety of applications.

Feasibility of measuring pericoronary fat from precontrast scans: Effect of iodinated contrast on pericoronary fat attenuation.

BACKGROUND: Pericoronary adipose tissue (PCAT) attenuation has been identified as a marker for cardiovascular risk. The effect of contrast enhancement on fat attenuation is unknown. We aim to compare precontrast coronary scans to postcontrast CCTA for quantification of pericoronary fat volume and attenuation. METHODS: Thin slice pre- and post-contrast studies obtained at 120 kVp, heart rate <60, with no plaque or artifact in the right coronary artery (RCA) were selected. Analysis was limited to pixels -30 Hounsfield units (HU) to -190 HU and from 10 mm to 50 mm distal to the RCA origin at a radial distance equal to the vessel diameter. A subgroup with no plaque across all coronaries was also analyzed. RESULTS: Of 119 study pairs, the average RCA diameter was highly correlated at 3.85 mm (postcontrast) and 3.84 mm (precontrast), r = 0.97, p < 0.0001. The mean attenuation of pre- and postcontrast images was also highly correlated at -87.02 ±â€¯7.15 HU and -82.74 ±â€¯6.54 HU, respectively (r = 0.65, p < 0.0001). Pericoronary fat volume in the -190 to -30 HU range was 396 mm³ lower in the post contrast versus pre-contrast, consistent with higher attenuation (less negative) voxels postcontrast (p < 0.0001). Inter- and intra-reader agreement ranged 95-100% and 90% for precontrast and 85-90% for postcontrast studies, respectively. Subgroup analysis revealed precontrast attenuation -85.59 ±â€¯7.53 HU and postcontrast -82.21 ±â€¯7.15 HU were highly correlated r = 0.67, p < 0.0001. CONCLUSION: Pericoronary fat enhances with iodinated contrast, potentially explaining some of its risk-predictive capabilities. Fat attenuation and volume can be reliably measured from precontrast calcium scans, with volume quantification showing particularly strong correlation. Excellent inter- and intra-reader agreement is also demonstrated.

Multiexon deletion alleles of ATF6 linked to achromatopsia.

Achromatopsia (ACHM) is an autosomal recessive disease that results in severe visual loss. Symptoms of ACHM include impaired visual acuity, nystagmus, and photoaversion starting from infancy; furthermore, ACHM is associated with bilateral foveal hypoplasia and absent or severely reduced cone photoreceptor function on electroretinography. Here, we performed genetic sequencing in 3 patients from 2 families with ACHM, identifying and functionally characterizing 2 mutations in the activating transcription factor 6 (ATF6) gene. We identified a homozygous deletion covering exons 8-14 of the ATF6 gene from 2 siblings from the same family. In another patient from a different family, we identified a heterozygous deletion covering exons 2 and 3 of the ATF6 gene found in trans with a previously identified ATF6 c.970C>T (p.Arg324Cys) ACHM disease allele. Recombinant ATF6 proteins bearing these exon deletions showed markedly impaired transcriptional activity by qPCR and RNA-Seq analysis compared with WT-ATF6. Finally, RNAscope revealed that ATF6 and the related ATF6B transcripts were expressed in cones as well as in all retinal layers in normal human retina. Overall, our data identify loss-of-function ATF6 disease alleles that cause human foveal disease.

Coarse-grained molecular dynamics simulations of α-1,3-glucan.

In this paper we present a computational study of aggregation in aqueous solutions of α-1,3-glucan captured using a coarse-grained (CG) model that can be extended to other polysaccharides. This CG model captures atomistic geometry (i.e., relative placement of the hydrogen bonding donors and acceptors within the monomer) of the α-1,3-glucan monomer, the directional interactions due to the donor-acceptor hydrogen bonds, and their effect on aggregation of multiple α-1,3-glucan chains without the extensive computational resources needed for simulations with atomistic models. Using this CG model, we conduct molecular dynamics simulations to assess the effect of varying α-1,3-glucan chain length and hydrogen bond interaction strengths on the aggregation of multiple chains at finite concentrations in implicit solvent. We quantify the hydrogen bonding strength needed for multiple chains to aggregate, the distribution of inter- and intra-chain hydrogen bonds within the aggregate and in some cases, the shapes of the aggregate. We also explore the effect of substitution/silencing of some randomly selected or specific hydrogen bonding sites in the chain on the aggregation and aggregate structure. In the unmodified α-1,3-glucan solution, the inter-chain hydrogen bonds cause the chains to aggregate into sheets. Random silencing of hydrogen bonding donor sites only increases the hydrogen bond strength needed for aggregation but retains the same aggregate structure as the unmodified chains. Specific silencing of the hydrogen-bonding site on the C6 carbon leads to the chains aggregating into planar sheets that then fold over to form hollow cylinders at intermediate hydrogen bond strength - 4.7 to 5.3 kcal mol-1. These cylindrical aggregates assemble end-to-end to form larger aggregates at higher hydrogen bond strengths.