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DOE OSTI2025

Development of a military-specific mesh-type computational phantom library and its application to internal dosimetry and in-field radiological triage screening

Estimates of organ-absorbed and committed doses to individuals exposed to radioactive materials via acute inhalation often rely on internal dose coefficients and detector responses from reference human computational models. To achieve more accurate dose assessments to United States Armed Forces service members exposed in-field, computational models with varying morphometric parameters representative of this population are necessary. The International Commission on Radiological Protection (ICRP) Publication 145 provides detailed mesh reference computational phantoms (MRCPs) for adult males and females, with morphometric parameters matched to the 50th percentile. Previously, these phantoms were 2D and 3D scaled to match desired height, mass, and secondary anthropomorphic parameters in the creation of the University of Florida / Memorial Sloan Kettering (UF/MSK) computational phantom library. To achieve body fat percentage targets required for accession into the US Armed Forces, muscle and fat volumes were adjusted accordingly, thus, creating the UF/Department of Defence computational phantom library presented in this study. A comprehensive library of mesh-type computational human phantoms was created, including 57 adult males and 49 adult females with morphometric parameters aligned with United States Armed Forces service members. Phantoms were restricted to a body mass index between 19 and 27.5, with body fat percentages below 26% for males and 36% for females. Specific absorbed fractions were computed for selected source and target combinations, demonstrating how variations in height and body mass influence energy absorption in target regions relative to the ICRP MRCPs. Radiation detector responses were also computed, revealing that higher body masses resulted in decreased registered counts in the detection volume. These findings highlight the importance of incorporating morphometric variability in computational phantoms to achieve more accurate dose assessments and radiation detection responses for United States Armed Forces service members who inhale radioactive materials in-field.

computational phantoms↗

DOE OSTI2021

Let it rip: The mechanics of self-bisection in asexual planarians determines their population reproductive strategies

Asexual freshwater planarians reproduce by transverse bisection (binary fission) into two pieces. This process produces a head and a tail, which fully regenerate within 1-2 weeks. How planarians split into two offspring - using only their musculature and substrate traction – is a challenging biomechanics problem. We found that three different species, Dugesia japonica, Girardia tigrina and Schmidtea mediterranea, have evolved three different mechanical solutions to self-bisect. Using time lapse imaging of the fission process, we quantitatively characterize the main steps of division in the three species and extract the distinct and shared key features. Across the three species, planarians actively alter their body shape, regulate substrate traction, and use their muscles to generate tensile stresses large enough to overcome the ultimate tensile strength of the tissue. Moreover, we show that how each planarian species divides dictates how resources are split among its offspring. This ultimately determines offspring survival and reproductive success. Furthermore, heterospecific differences in the mechanics of self-bisection of individual worms explain the observed differences in the population reproductive strategies of different planarian species.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2024

Physiological Adaptations to Progressive Endurance Exercise Training in Adult and Aged Rats: Insights from the Molecular Transducers of Physical Activity Consortium (MoTrPAC)

While regular physical activity is a cornerstone of health, wellness, and vitality, the impact of endurance exercise training on molecular signaling within and across tissues remains to be delineated. The Molecular Transducers of Physical Activity Consortium (MoTrPAC) was established to characterize molecular networks underlying the adaptive response to exercise. Here, we describe the endurance exercise training studies undertaken by the Preclinical Animal Sites Studies component of MoTrPAC, in which we sought to develop and implement a standardized endurance exercise protocol in a large cohort of rats. To this end, Adult (6-mo) and Aged (18-mo) female (n = 151) and male (n = 143) Fischer 344 rats were subjected to progressive treadmill training (5 d/wk, ~70%–75% VO 2 max) for 1, 2, 4, or 8 wk; sedentary rats were studied as the control group. A total of 18 solid tissues, as well as blood, plasma, and feces, were collected to establish a publicly accessible biorepository and for extensive omics-based analyses by MoTrPAC. Treadmill training was highly effective, with robust improvements in skeletal muscle citrate synthase activity in as little as 1–2 wk and improvements in maximum run speed and maximal oxygen uptake by 4–8 wk. For body mass and composition, notable age- and sex-dependent responses were observed. This work in mature, treadmill-trained rats represents the most comprehensive and publicly accessible tissue biorepository, to date, and provides an unprecedented resource for studying temporal-, sex-, and age-specific responses to endurance exercise training in a preclinical rat model.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2020

CoreCruncher : Fast and Robust Construction of Core Genomes in Large Prokaryotic Data Sets

The core genome represents the set of genes shared by all, or nearly all, strains of a given population or species of prokaryotes. Inferring the core genome is integral to many genomic analyses, however, most methods rely on the comparison of all the pairs of genomes; a step that is becoming increasingly difficult given the massive accumulation of genomic data. Here, we present CoreCruncher; a program that robustly and rapidly constructs core genomes across hundreds or thousands of genomes. CoreCruncher does not compute all pairwise genome comparisons and uses a heuristic based on the distributions of identity scores to classify sequences as orthologs or paralogs/xenologs. Although it is much faster than current methods, our results indicate that our approach is more conservative than other tools and less sensitive to the presence of paralogs and xenologs. CoreCruncher is freely available from: https://github.com/lbobay/CoreCruncher. CoreCruncher is written in Python 3.7 and can also run on Python 2.7 without modification. It requires the python library Numpy and either Usearch or Blast. Certain options require the programs muscle or mafft.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2022

Solutal-buoyancy-driven intertwining and rotation of patterned elastic sheets

Abstract The intertwining of strands into 3D spirals is ubiquitous in biology, enabling functions from information storage to maintenance of cell structure and directed locomotion. In synthetic systems, entwined fibers can provide superior mechanical properties and act as artificial muscle or structural reinforcements. Unlike structures in nature, the entwinement of synthetic materials typically requires application of an external stimulus, such as mechanical actuation, light, or a magnetic field. Herein, we use computational modeling to design microscale sheets that mimic biology by transducing chemical energy into mechanical action, and thereby self-organize and interlink into 3D spirals, which spontaneously rotate. These flexible sheets are immersed in a fluid-filled microchamber that encompasses an immobilized patch of catalysts on the bottom wall. The sheets themselves can be passive or active (coated with catalyst). Catalytic reactions in the solution generate products that occupy different volumes than the reactants. The resulting density variations exert a force on the fluid (solutal buoyancy force) that causes motion, which in turn drives the interlinking and collective swirling of the sheets. The individual sheets do not rotate; rotation only occurs when the sheets are interlinked. This level of autonomous, coordinated 3D structural organization, intertwining, and rotation is unexpected in synthetic materials systems operating without external controls. Using physical arguments, we identify dimensionless ratios that are useful in scaling these ideas to other systems. These findings are valuable for creating materials that act as “machines”, and directing soft matter to undergo self-sustained, multistep assembly that is governed by intrinsic chemical reactions.

Manna, Raj Kumar (ORCID:0000000322729487)↗

DOE OSTI2022

The Ahr2-Dependent wfikkn1 Gene Influences Zebrafish Transcriptome, Proteome, and Behavior

The aryl hydrocarbon receptor (AHR) is required for vertebrate development and is also activated by exogenous chemicals, including polycyclic aromatic hydrocarbons (PAHs) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). AHR activation is well-understood, but roles of downstream molecular signaling events are largely unknown. From previous transcriptomics in 48 h postfertilization (hpf) zebrafish exposed to several PAHs and TCDD, we found wfikkn1 was highly coexpressed with cyp1a (marker for AHR activation). Thus, we hypothesized wfikkn1’s role in AHR signaling, and showed that wfikkn1 expression was Ahr2 (zebrafish ortholog of human AHR)-dependent in developing zebrafish exposed to TCDD. To functionally characterize wfikkn1, we made a CRISPR-Cas9 mutant line with a 16-bp deletion in wfikkn1’s exon, and exposed wildtype and mutants to dimethyl sulfoxide or TCDD. 48-hpf mRNA sequencing revealed over 700 genes that were differentially expressed (p < .05, log 2 FC > 1) between each pair of treatment combinations, suggesting an important role for wfikkn1 in altering both the 48-hpf transcriptome and TCDD-induced expression changes. Mass spectrometry-based proteomics of 48-hpf wildtype and mutants revealed 325 significant differentially expressed proteins. Functional enrichment demonstrated wfikkn1 was involved in skeletal muscle development and played a role in neurological pathways after TCDD exposure. Mutant zebrafish appeared morphologically normal but had significant behavior deficiencies at all life stages, and absence of Wfikkn1 did not significantly alter TCDD-induced behavior effects at all life stages. In conclusion, wfikkn1 did not appear to be significantly involved in TCDD’s overt toxicity but is likely a necessary functional member of the AHR signaling cascade.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2024

In vitro evidence that the vasorelaxant effects of 2‐nitro‐1‐phenyl‐1‐propanol on rat coronary arteries involve cyclic nucleotide pathways

Abstract The synthetic nitro‐alcohol 2‐nitro‐1‐phenyl‐1‐propanol (NPP) has endothelium‐independent relaxing properties in isolated preparations of rat aorta and mesenteric artery. In this study, we investigated whether the vasodilator effects occur in coronary vessels and explored whether hyperpolarization is involved in the underlying mechanism of NPP‐induced smooth muscle relaxation. The relaxing responses were studied in isolated preparations of the left anterior descending coronary (ADC) and the septal coronary (SC) arteries, which had been previously maintained under sustained contraction induced by the thromboxane A 2 analogue U‐46619. Administered cumulatively, NPP elicited concentration‐dependent vasorelaxation with similar potency in both vessels. The relaxant effect remained unaffected by the nitric oxide synthase inhibitor L‐NAME, the protein kinase C inhibitor bisindolylmaleimide IV and the Rho‐associated protein kinase inhibitor Y‐27632. However, it was significantly diminished by the adenylyl cyclase inhibitor MDL‐12,330A, the guanylyl cyclase inhibitor ODQ, as well as the K + channel inhibitors tetraethylammonium and CsCl. In ADC preparations impaled with intracellular micropipettes, NPP hyperpolarized the vascular preparation. When the isolated preparation was precontracted by 5‐hydroxytryptamine or 80 mM KCl, NPP‐induced relaxation with lower pharmacological potency compared to the vessels contracted by U‐46619. In conclusion, NPP exhibits vasorelaxant effects on rat coronary arteries, likely involving pathways that include cyclic nucleotide production and membrane hyperpolarization.

Vasconcelos‐Silva, Alfredo Augusto↗

DOE OSTI2020

Laser writing of nitrogen-doped silicon carbide for biological modulation

Conducting or semiconducting materials embedded in insulating polymeric substrates can be useful in biointerface applications; however, attainment of this composite configuration by direct chemical processes is challenging. Laser-assisted synthesis has evolved as a fast and inexpensive technique to prepare various materials, but its utility in the construction of biophysical tools or biomedical devices is less explored. Here, we use laser writing to convert portions of polydimethylsiloxane (PDMS) into nitrogen-doped cubic silicon carbide (3C-SiC). The dense 3C-SiC surface layer is connected to the PDMS matrix via a spongy graphite layer, facilitating electrochemical and photoelectrochemical activity. We demonstrate the fabrication of arbitrary two-dimensional (2D) SiC-based patterns in PDMS and freestanding 3D constructs. To establish the functionality of the laser-produced composite, we apply it as flexible electrodes for pacing isolated hearts and as photoelectrodes for local peroxide delivery to smooth muscle sheets.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2024

Stiffness anisotropy coordinates supracellular contractility driving long-range myotube-ECM alignment

The ability of cells to organize into tissues with proper structure and function requires the effective coordination of proliferation, migration, polarization, and differentiation across length scales. Skeletal muscle is innately anisotropic; however, few biomaterials can emulate mechanical anisotropy to determine its influence on tissue patterning without introducing confounding topography. Here, we demonstrate that substrate stiffness anisotropy coordinates contractility-driven collective cellular dynamics resulting in C2C12 myotube alignment over millimeter-scale distances. When cultured on mechanically anisotropic liquid crystalline polymer networks (LCNs) lacking topography, C2C12 myoblasts collectively polarize in the stiffest direction. Cellular coordination is amplified through reciprocal cell-ECM dynamics that emerge during fusion, driving global myotube-ECM ordering. Conversely, myotube alignment was restricted to small local domains with no directional preference on mechanically isotropic LCNs of the same chemical formulation. These findings provide valuable insights for designing biomaterials that mimic anisotropic microenvironments and underscore the importance of stiffness anisotropy in orchestrating tissue morphogenesis.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2023

Bioadhesive polymer semiconductors and transistors for intimate biointerfaces

The use of bioelectronic devices relies on direct contact with soft biotissues. For transistor-type bioelectronic devices, the semiconductors that need to have direct interfacing with biotissues for effective signal transduction do not adhere well with wet tissues, thereby limiting the stability and conformability at the interface. Here we report a bioadhesive polymer semiconductor through a double-network structure formed by a bioadhesive brush polymer and a redox-active semiconducting polymer. The resulting semiconducting film can form rapid and strong adhesion with wet tissue surfaces together with high charge-carrier mobility of ~1 square centimeter per volt per second, high stretchability, and good biocompatibility. Further fabrication of a fully bioadhesive transistor sensor enabled us to produce high-quality and stable electrophysiological recordings on an isolated rat heart and in vivo rat muscles.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

DOE OSTI2023

Pharmacological Profiling of a Brugia malayi Muscarinic Acetylcholine Receptor as a Putative Antiparasitic Target

The diversification of anthelmintic targets and mechanisms of action will help ensure the sustainable control of nematode infections in response to the growing threat of drug resistance. G protein-coupled receptors (GPCRs) are established drug targets in human medicine but remain unexploited as anthelmintic substrates despite their important roles in nematode neuromuscular and physiological processes. Bottlenecks in exploring the druggability of parasitic nematode GPCRs include a limited helminth genetic toolkit and difficulties establishing functional heterologous expression. In an effort to address some of these challenges, we profile the function and pharmacology of muscarinic acetylcholine receptors in the human parasite Brugia malayi, an etiological agent of human lymphatic filariasis. While acetylcholine-gated ion channels are intensely studied as targets of existing anthelmintics, comparatively little is known about metabotropic receptor contributions to parasite cholinergic signaling. Using multivariate phenotypic assays in microfilariae and adults, we show that nicotinic and muscarinic compounds disparately affect parasite fitness traits. We identify a putative G protein-linked acetylcholine receptor of B. malayi (Bma-GAR-3) that is highly expressed across intramammalian life stages and adapt spatial RNA in situ hybridization to map receptor transcripts to critical parasite tissues. Tissue-specific expression of Bma-gar-3 in Caenorhabditis elegans (body wall muscle, sensory neurons, and pharynx) enabled receptor deorphanization and pharmacological profiling in a nematode physiological context. Finally, we developed an image-based feeding assay as a reporter of pharyngeal activity to facilitate GPCR screening in parasitized strains. We expect that these receptor characterization approaches and improved knowledge of GARs as putative drug targets will further advance the study of GPCR biology across medically important nematodes.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2023

New insights into the natural history of bronchopulmonary dysplasia from proteomics and multiplexed immunohistochemistry

Bronchopulmonary dysplasia (BPD) is a disease of prematurity related to the arrest of normal lung development. The objective of this study was to better understand how proteome modulation and cell-type shifts are noted in BPD pathology. Pediatric human donors aged 1–3 yr were classified based on history of prematurity and histopathology consistent with “healed” BPD (hBPD, n = 3) and “established” BPD (eBPD, n = 3) compared with respective full-term born (n = 6) age-matched term controls. Proteins were quantified by tandem mass spectroscopy with selected Western blot validations. Multiplexed immunofluorescence (MxIF) microscopy was performed on lung sections to enumerate cell types. Protein abundances and MxIF cell frequencies were compared among groups using ANOVA. Cell type and ontology enrichment were performed using an in-house tool and/or EnrichR. Proteomics detected 5,746 unique proteins, 186 upregulated and 534 downregulated, in eBPD versus control with fewer proteins differentially abundant in hBPD as compared with age-matched term controls. Cell-type enrichment suggested a loss of alveolar type I, alveolar type II, endothelial/capillary, and lymphatics, and an increase in smooth muscle and fibroblasts consistent with MxIF. Histochemistry and Western analysis also supported predictions of upregulated ferroptosis in eBPD versus control. Finally, several extracellular matrix components mapping to angiogenesis signaling pathways were altered in eBPD. Despite clear parsing by protein abundance, comparative MxIF analysis confirms phenotypic variability in BPD. This work provides the first demonstration of tandem mass spectrometry and multiplexed molecular analysis of human lung tissue for critical elucidation of BPD trajectory-defining factors into early childhood.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2023

DT-HYDRO

The software solves the time dependent, one dimensional (1D) coupled mass and momentum balance equations governing the elastic flow of water through the penstock, turbine and draft tube in a hydroelectric facility using a high order finite volume based method. The numerical method is based on the Kurganov-Tadmor central method paired with the Monotonic Upstream-centered Scheme for Conservation Laws (MUSCL). This solution method accurately resolves the fast transient behavior of the flow, including water hammer. Additionally, the software estimates the full 3D flow field within the turbine chamber in real time, a feat that is made possible by leveraging pre-computed CFD results by utilizing a reduced order modeling method based on an efficient randomized singular value decomposition (SVD) driven proper orthogonal decomposition (POD) with POD-mode weight regression. The reduced order model of the 3D flow is directly coupled to the 1D elastic flow model so the entire flow field through the penstock and turbine system is resolved quickly and with high fidelity.

Gurecky, William [Oak Ridge National Laboratory (O↗

DOE OSTI2024

An FDA-approved drug structurally and phenotypically corrects the K210del mutation in genetic cardiomyopathy models

Dilated cardiomyopathy (DCM) due to genetic disorders results in decreased myocardial contractility, leading to high morbidity and mortality rates. There are several therapeutic challenges in treating DCM, including poor understanding of the underlying mechanism of impaired myocardial contractility and the difficulty of developing targeted therapies to reverse mutation-specific pathologies. In this report, we focused on K210del, a DCM-causing mutation, due to 3-nucleotide deletion of sarcomeric troponin T (TnnT), resulting in loss of Lysine210. We resolved the crystal structure of the troponin complex carrying the K210del mutation. K210del induced an allosteric shift in the troponin complex resulting in distortion of activation Ca 2+ -binding domain of troponin C (TnnC) at S69, resulting in calcium discoordination. Next, we adopted a structure-based drug repurposing approach to identify bisphosphonate risedronate as a potential structural corrector for the mutant troponin complex. Cocrystallization of risedronate with the mutant troponin complex restored the normal configuration of S69 and calcium coordination. Risedronate normalized force generation in K210del patient-induced pluripotent stem cell–derived (iPSC-derived) cardiomyocytes and improved calcium sensitivity in skinned papillary muscles isolated from K210del mice. Systemic administration of risedronate to K210del mice normalized left ventricular ejection fraction. Collectively, these results identify the structural basis for decreased calcium sensitivity in K210del and highlight structural and phenotypic correction as a potential therapeutic strategy in genetic cardiomyopathies.

Research & Experimental Medicine↗

DOE OSTI2022

Hip Abductor Strength Asymmetry: Relationship to Upper Extremity Injury in Professional Baseball Players

Background: Hip strength is an important factor for control of the lumbo-pelvic-hip complex. Deficits in hip strength may affect throwing performance and contribute to upper extremity injuries. Hypothesis: Deficits in hip abduction isometric strength would be greater in those who sustained an upper extremity injury and hip strength would predict injury incidence. Study Design: Prospective cohort study. Level of Evidence: Level 3. Methods: Minor League baseball players (n = 188, age = 21.5 ± 2.2 years; n = 98 pitchers; n = 90 position players) volunteered. Hip abduction isometric strength was assessed bilaterally with a handheld dynamometer in side-lying position, expressed as torque using leg length (N·m). Hip abduction strength asymmetry was represented by [(trail leg/lead leg) × 100]. Overuse or nontraumatic throwing arm injuries were prospectively tracked. Poisson regression models were used to estimate relative risk ratios associated with hip asymmetry; confounders, including history of prior overuse injury in the past year, were included. Results: Hip abduction asymmetry ranged from 0.05% to 57.5%. During the first 2 months of the season, 18 players (n = 12 pitchers) sustained an upper extremity injury. In pitchers, for every 5% increase in hip abduction asymmetry, there was a 1.24 increased risk of sustaining a shoulder or elbow injury. No relationship between hip abduction strength and injury was observed for position players. Conclusion: Hip abduction asymmetry in pitchers was related to subsequent upper extremity injuries. The observed risk ratio indicates that hip abduction asymmetry may contribute a significant but small increased risk of injury. Clinical Relevance: Hip abduction muscle deficits may affect pitching mechanics and increase arm stress. Addressing hip asymmetry deficits that exceed 5% may be beneficial in reducing upper extremity injury rates in pitchers.

Sport Sciences↗

DOE OSTI2021

Global transcriptome profile of the developmental principles of in vitro iPSC-to-motor neuron differentiation

Abstract Background Human induced pluripotent stem cells (iPSC) have opened new avenues for regenerative medicine. Consequently, iPSC-derived motor neurons have emerged as potentially viable therapies for spinal cord injuries and neurodegenerative disorders including Amyotrophic Lateral Sclerosis. However, direct clinical application of iPSC bears in itself the risk of tumorigenesis and other unforeseeable genetic or epigenetic abnormalities. Results Employing RNA-seq technology, we identified and characterized gene regulatory networks triggered by in vitro chemical reprogramming of iPSC into cells with the molecular features of motor neurons (MNs) whose function in vivo is to innervate effector organs. We present meta-transcriptome signatures of 5 cell types: iPSCs, neural stem cells, motor neuron progenitors, early motor neurons, and mature motor neurons. In strict response to the chemical stimuli, along the MN differentiation axis we observed temporal downregulation of tumor growth factor-β signaling pathway and consistent activation of sonic hedgehog, Wnt/β-catenin, and Notch signaling. Together with gene networks defining neuronal differentiation (neurogenin 2, microtubule-associated protein 2, Pax6, and neuropilin-1), we observed steady accumulation of motor neuron-specific regulatory genes, including Islet-1 and homeobox protein HB9. Interestingly, transcriptome profiling of the differentiation process showed that Ca 2+ signaling through cAMP and LPC was downregulated during the conversion of the iPSC to neural stem cells and key regulatory gene activity of the pathway remained inhibited until later stages of motor neuron formation. Pathways shaping the neuronal development and function were well-represented in the early motor neuron cells including, neuroactive ligand-receptor interactions, axon guidance, and the cholinergic synapse formation. A notable hallmark of our in vitro motor neuron maturation in monoculture was the activation of genes encoding G-coupled muscarinic acetylcholine receptors and downregulation of the ionotropic nicotinic acetylcholine receptors expression. We observed the formation of functional neuronal networks as spontaneous oscillations in the extracellular action potentials recorded on multi-electrode array chip after 20 days of differentiation. Conclusions Detailed transcriptome profile of each developmental step from iPSC to motor neuron driven by chemical induction provides the guidelines to novel therapeutic approaches in the re-construction efforts of muscle innervation.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2024

Cellular response of keratinocytes to the entry and accumulation of nanoplastic particles

Plastic accumulation in the environment is rapidly increasing, and nanoplastics (NP), byproducts of environmental weathering of bulk plastic waste, pose a significant public health risk. Particles may enter the human body through many possible routes such as ingestion, inhalation, and skin absorption. However, studies on NP penetration and accumulation in human skin are limited. Loss or reduction of the keratinized skin barrier may enhance the skin penetration of NPs. The present study investigated the entry of NPs into a human skin system modeling skin with compromised barrier functions and cellular responses to the intracellular accumulations of NPs. Two in vitro models were employed to simulate human skin lacking keratinized barriers. The first model was an ex vivo human skin culture with the keratinized dermal layer (stratum corneum) removed. The second model was a 3D keratinocyte/dermal fibroblast cell co-culture model with stratified keratinocytes on the top and a monolayer of skin fibroblast cells co-cultured at the bottom. The penetration and accumulation of the NPs in different cell types were observed using fluorescent microscopy, confocal microscopy, and cryogenic electron microscopy (cryo-EM). The cellular responses of keratinocytes and dermal fibroblast cells to stress induced by NPs stress were measured. The genetic regulatory pathway of keratinocytes to the intracellular NPs was identified using transcript analyses and KEGG pathway analysis. The cellular uptake of NPs by skin cells was confirmed by imaging analyses. Transepidermal transport and penetration of NPs through the skin epidermis were observed. According to the gene expression and pathway analyses, an IL-17 signaling pathway was identified as the trigger for cellular responses to internal NP accumulation in the keratinocytes. The transepidermal NPs were also found in co-cultured dermal fibroblast cells and resulted in a large-scale transition from fibroblast cells to myofibroblast cells with enhanced production of α-smooth muscle actin and pro-Collagen Ia. The upregulation of inflammatory factors and cell activation may result in skin inflammation and ultimately trigger immune responses.

36 MATERIALS SCIENCE↗

DOE OSTI2024

Morphological and molecular characterization of a Sarcocystis bovifelis-like sarcocyst in American beef

Abstract Background Parasites in the apicomplexan genusSarcocystisinfect cattle worldwide. Assessing the economic importance of each such parasite species requires proper diagnosis.Sarcocystiscruzi,a thin-walled species, infects virtually all cattle. The prevalence of the other thin-walled parasite,Sarcocystisheydorni, remains less well established. The remaining six species all have thick (> 3 µm) cyst walls (Sarcocystishirsuta,S.hominis,S.bovifelis,S.bovini,S.sigmoideus, andS.rommeli). Thick-walled sarcocysts often induce inflammation in striated muscles (causing bovine eosinophilic myositis), leading to condemnation of carcasses at slaughter. One of these,S.hirsuta, can be seen macroscopically and lead to condemnation of beef. TwoSarcocystisspecies,S.hominisandS.heydorni, are zoonotic. AlthoughS.hominishas been reported as prevalent in Europe, the occurrence of thick-walled species in the US remains poorly known. Here, for the first time to our knowldge, we characterize a thick-walledSarcocystisspecies from a sample of beef from a local grocery store in Maryland. By morphological and genetic criteria, it closely, but not perfectly, resembles parasites previously ascribed toS.bovifelis. Methods Beef samples were examined forSarcocystisinfection, using acid-pepsin digestion to search for bradyzoites, microscopically by compression between a glass slide and coverslip, by histology of paraffin embedded sections stained with hematoxylin and eosin, and by transmission electron microscopy (TEM). Molecular characterization was attempted employing genetic markers:18SrRNA,28SrRNA,cox1,ITS1,gapdh1,ron3, andrpoB. Results Molecular evaluation revealed 100% identity withS.bovifelis-like sarcocysts from naturally infected cattle from Germany and Argentina; although the condition of the frozen material precludes complete characterization by TEM, we noted morphological features which differed from theS.bovifelisoriginally described from experimentally infected cattle from Germany. Conclusions A novelSarcocystisspecies is described from beef from the USA but not named until further evaluation. Graphical Abstract

Parasitology↗