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At least 271 records · Page 15

cTULIP: application of a human-based RNA-seq primary tumor classification tool for cross-species primary tumor classification in canine

The domestic dog, Canis familiaris, is quickly gaining traction as an advantageous model for use in the study of cancer, one of the leading causes of death worldwide. Naturally occurring canine cancers share clinical, histological, and molecular characteristics with the corresponding human diseases. In this study, we take a deep-learning approach to test how similar the gene expression profile of canine glioma and bladder cancer (BLCA) tumors are to the corresponding human tumors. We likewise develop a tool for identifying misclassified or outlier samples in large canine oncological datasets, analogous to that which was developed for human datasets. We test a number of machine learning algorithms and found that a convolutional neural network outperformed logistic regression and random forest approaches. We use a recently developed RNA-seq-based convolutional neural network, TULIP, to test the robustness of a human-data-trained primary tumor classification tool on cross-species primary tumor prediction. Our study ultimately highlights the molecular similarities between canine and human BLCA and glioma tumors, showing that protein-coding one-to-one homologs shared between humans and canines, are sufficient to distinguish between BLCA and gliomas. The results of this study indicate that using protein-coding one-to-one homologs as the features in the input layer of TULIP performs good primary tumor prediction in both humans and canines. Furthermore, our analysis shows that our selected features also contain the majority of features with known clinical relevance in BLCA and gliomas. Our success in using a human-data-trained model for cross-species primary tumor prediction also sheds light on the conservation of oncological pathways in humans and canines, further underscoring the importance of the canine model system in the study of human disease.

60 APPLIED LIFE SCIENCES↗

An Approach to Dynamic Human Reliability Analysis and Its Data Collection Framework

Human reliability analysis (HRA) is a method for evaluating human errors in a variety of complex systems such as nuclear power plants, military systems, aircraft, and chemical plants. Most HRA methods currently used by regulatory institutes or utilities are called static HRA and are carried out by simple worksheets or simple calculators. To date, there are many unsolved or intrinsic challenges in static HRA. For example, existing static HRA does not realistically model and evaluate human actions as they would be performed at actual systems. There is no method with HRA to objectively estimate the time required for human actions despite being essential to HRA processes. In addition, many HRA methods still rely on a dataset generated prior to the 1980s, from unrelated industry experience or simply from expert judgment. Accordingly, this study attempted to research how to overcome the challenges of existing HRA via dynamic risk assessment (a.k.a., simulation-based or computation-based risk assessment) techniques. First, this study developed a dynamic HRA method, named as PRocedure-based Investigation Method of EMRALD Risk Assessment – HRA (PRIMERA-HRA). The PRIMERA-HRA mainly concentrates on providing HRA analysts with specific guidelines on how to reasonably model human actions, assign human reliability data and evaluate output of simulation within a dynamic probabilistic risk assessment tool, called as Event Modeling Risk Assessment using Linked Diagrams (EMRALD). Second, this study also developed a module for performance shaping factors (i.e., the key concept in HRA quantification) applicable to dynamic HRA, then implemented it based on PRIMERA-HRA within the EMRALD tool. Third, this study developed an HRA data collection framework to support dynamic HRA, called as Simplified Human Error Experimental Program (SHEEP). Originally, the SHEEP study aimed to support static HRA and its data collection, but recently extended the scope to the new technologies such as dynamic HRA or HRA for advanced reactors. SHEEP focuses on the use of data collected from simplified simulators to complement—but not replace—data collection studies using full-scope simulators and actual operators. To date, many experiments were conducted under the SHEEP framework. Multiple analyses, such as human performance analysis, human error analysis, task complexity analysis, learning effect analysis and time distribution analysis, were also carried out using the collected data. Then, based on the major insights, an approach to inferring full-scope data based on simplified simulator data was proposed. The PRIMERA-HRA and SHEEP research are expected to evaluate human actions more realistically than existing static HRA, provide an opportunity to collect more HRA data with reasonable cost and labor, then contribute to enhance the quality of HRA.

99 - GENERAL AND MISCELLANEOUS↗

A Typology for Characterizing Human Action in MultiSector Dynamics Models

The role of individual and collective human agency is increasingly recognized as a prominent and arguably paramount determinant in shaping the behavior, trajectory, and vulnerability of multisector systems. This human influence operates at multiple scales: from short-term (hourly to daily) to long-term (annually to centennial) timescales, and from the local to the global, pushing systems towards either desirable or undesirable outcomes. However, the effort to represent human systems in multisector models has been fragmented across philosophical, methodological, and disciplinary lines. To cohere insights across diverse modeling approaches, we present a new typology for classifying how human actors are represented in the broad suite of coupled human-natural system models that are applied in MultiSector Dynamics (MSD) research. The typology conceptualizes a “sector” as a system-of-systems that includes a diverse group of human actors, defined across individual to collective social levels, involved in governing, provisioning, and utilizing products, goods, or services towards some human end. We trace the salient features of modeled representations of human systems by organizing the typology around three key questions: 1) Who are the modeled actors in MSD systems? 2) What are their modeled actions? 3) How and for what purpose are these actors and actions operationalized in a computational model? The typology is used to critically examine existing models and chart the frontier of future human systems modeling for MSD research.

29 ENERGY PLANNING, POLICY, AND ECONOMY↗

Are humans still necessary? Expanding the discussion

The rise of automation, artificial intelligence (AI), and autonomous systems raises important questions about the future role of humans and the field of human factors/ergonomics in workplaces. This paper builds on Dr. Peter Hancock’s 2023 ‘Are Humans Still Necessary?’ article published in the Ergonomics journal. Using a multi-method approach that included a debate, opinion polling, roundtable discussions, and AI queries, the current effort examined the necessity of human involvement in future work environments. Debate team members presented arguments for and against the need for human workers, considering human factors, technology, and socioeconomic factors. Observations indicate that while AI may handle routine tasks, humans will likely remain essential for complex decision making, creativity, and ethical considerations. The paper advocates for viewing workplace dynamics as collaborative human-AI partnerships rather than competition, highlighting the need for a transdisciplinary approach in which human factors/ergonomics professionals play a vital role in enhancing these relationships.

99 - GENERAL AND MISCELLANEOUS↗

The Development of A Human Systems Simulation Laboratory: Strategic Direction

The Human System Simulation Laboratory (HSSL) at the Idaho National Laboratory is one of few facilities of its kind that allows human factors researchers to evaluate various aspects of human performance and human system interaction for proposed reactor designs and upgrades. A basic system architecture, physical configuration and simulation capability were established to enable human factors researchers to support multiple, simultaneous simulations and also different power plant technologies. Although still evolving in terms of its technical and functional architecture, the HSSL is already proving its worth in supporting current and future nuclear industry needs for light water reactor sustainability and small modular reactors. The evolution of the HSSL is focused on continual physical and functional refinement to make it a fully equipped, reconfigurable facility where advanced research, testing and validation studies can be conducted on a wider range of reactor technologies. This requires the implementation of additional plant models to produce empirical research data on human performance with emerging human-system interaction technologies. Additional beneficiaries of this information include system designers and HRA practitioners. To ensure that results of control room crew studies will be generalizable to the existing and evolving fleet of US reactors, future expansion of the HSSL may also include other SMR plant models, plant-specific simulators and a generic plant model aligned to the current generation of pressurized water reactors (PWRs) and future advanced reactor designs. Collaboration with industry partners is also proving to be a vital component of the facility as this helps to establish a formal basis for current and future human performance experiments to support nuclear industry objectives. A long-range Program Plan has been developed for the HSSL to ensure that the facility will support not only the Department of Energy’s Light Water Reactor Sustainability Program, but also to provide human factors guidance for all future developments of the nuclear industry.

46 INSTRUMENTATION RELATED TO NUCLEAR SCIENCE AND ↗

Identifying Requirements for Effective Human-Automation Teamwork

Previous studies have shown that poorly designed human-automation collaboration, such as poorly designed communication protocols, often leads to problems for the human operators, such as: lack of vigilance, complacency, and loss of skills. These problems often lead to suboptimal system performance. To address this situation, a considerable amount of research has been conducted to improve human-automation collaboration and to make automation function better as a “team player.” Much of this research is based on an understanding of what it means to be a good team player from the perspective of a human team. However, the research is often based on a simplified view of human teams and teamwork. In this study, we sought to better understand the capabilities and limitations of automation from the standpoint of human teams. We first examined human teams to identify the principles for effective teamwork. We next reviewed the research on integrating automation agents and human agents into mixed agent teams to identify the limitations of automation agents to conform to teamwork principles. This research resulted in insights that can lead to more effective human-automation collaboration by enabling a more realistic set of requirements to be developed based on the strengths and limitations of all agents.

99 GENERAL AND MISCELLANEOUS↗

Considerations for Regulation and Practical Implementation of Analyses of Inadvertent Human Intrusion - 20169

Recommendations from the International Commission for Radiological Protection (ICRP) and the International Atomic Energy Agency (IAEA) identify the need to consider potential inadvertent human intrusion into a near-surface radioactive waste disposal facility when developing limits for radionuclide concentrations that can be disposed. The recommendations also provide perspective on expectations for performance criteria and general assumptions for such analyses. Inadvertent human intrusion analyses are conducted to address the potential failure of institutional controls and loss of memory of a disposal facility. The analyses consider potential consequences of human actions that result in unknowingly bringing waste to the surface. Inadvertent human intrusion scenarios consider potential impacts to someone involved in an activity that actually disturbs the waste (e.g., drilling or excavation of a basement) or that exposes the person to emanations from the waste (e.g., radioactive gases) and also for the case where there may be longer term exposures to waste that has been brought to the surface. In practice, inadvertent human intrusion analyses have proven to be a critical consideration for the development of waste classification systems and for the determination of disposal requirements and limitations on wastes that can be disposed at shallow depths. The IAEA implemented a project on Human Intrusion in the context of Disposal of Radioactive waste (HIDRA) to further explore the regulatory expectations and practical implementation of analyses of potential inadvertent human intrusion. The paper begins with a discussion of international recommendations related to consideration of inadvertent human intrusion and then summarizes examples of some topical considerations related to addressing inadvertent human intrusion that are being discussed as part of the second phase of the HIDRA project. The topical considerations are planned to be addressed in a formal IAEA publication. All information in this paper simply reflects on-going project discussions and no formal recommendations are implied for these topical considerations at this time. (authors)

12 MANAGEMENT OF RADIOACTIVE AND NON-RADIOACTIVE W↗

Monoacylglycerol O-acyltransferase 1 is regulated by peroxisome proliferator-activated receptor γ in human hepatocytes and increases lipid accumulation

Monoacylglycerol O-acyltransferase (MGAT) is an enzyme that is involved in triglyceride synthesis by catalyzing the formation of diacylglycerol from monoacylglycerol and fatty acyl CoAs. Recently, we reported that MGAT1 has a critical role in hepatic TG accumulation and that its suppression ameliorates hepatic steatosis in a mouse model. However, the function of MGAT enzymes in hepatic lipid accumulation has not been investigated in humans. Unlike in rodents, MGAT3 as well as MGAT1 and MGAT2 are present in humans. In this study, we evaluated the differences between MGAT subtypes and their association with peroxisome proliferator-activated receptor γ (PPARγ), a regulator of mouse MGAT1 expression. In human primary hepatocytes, basal expression of MGAT1 was lower than that of MGAT2 or MGAT3, but was strongly induced by PPARγ overexpression. A luciferase assay as well as an electromobility shift assay revealed that human MGAT1 promoter activity is driven by PPARγ by direct binding to at least two regions of the promoter in 293T and HepG2 cells. Moreover, siRNA-mediated suppression of MGAT1 expression significantly attenuated lipid accumulation by PPARγ overexpression in HepG2 cells, as evidenced by oil-red-O staining. These results suggest that human MGAT1 has an important role in fatty liver formation as a target gene of PPARγ, and blocking MGAT1 activity could be an efficient therapeutic way to reduce nonalcoholic fatty liver diseases in humans. - Highlights: • PPARγ promotes MGAT1 expression in human primary hepatocytes. • PPARγ directly regulates MGAT1 promoter activity. • Human MGAT1 promoter has at least two PPARγ-binding elements. • Inhibition of MGAT1 expression attenuates hepatic lipid accumulation in humans.

60 APPLIED LIFE SCIENCES↗

Activation of human natural killer cells by the soluble form of cellular prion protein

Cellular prion protein (PrP{sup C}) is widely expressed in various cell types, including cells of the immune system. However, the specific roles of PrP{sup C} in the immune system have not been clearly elucidated. In the present study, we investigated the effects of a soluble form of recombinant PrP{sup C} protein on human natural killer (NK) cells. Recombinant soluble PrP{sup C} protein was generated by fusion of human PrP{sup C} with the Fc portion of human IgG{sub 1} (PrP{sup C}-Fc). PrP{sup C}-Fc binds to the surface of human NK cells, particularly to CD56{sup dim} NK cells. PrP{sup C}-Fc induced the production of cytokines and chemokines and the degranulation of granzyme B from NK cells. In addition, PrP{sup C}-Fc facilitated the IL-15-induced proliferation of NK cells. PrP{sup C}-Fc induced phosphorylation of ERK-1/2 and JNK in NK cells, and inhibitors of the ERK or the JNK pathways abrogated PrP{sup C}-Fc-induced cytokine production in NK cells. In conclusion, the soluble form of recombinant PrP{sup C}-Fc protein activates human NK cells via the ERK and JNK signaling pathways. - Highlights: • Recombinant soluble PrP{sup C} (PrP{sup C}-Fc) was generated by fusion of human PrP{sup C} with IgG1 Fc portion. • PrP{sup C}-Fc protein induces the production of cytokines and degranulation from human NK cells. • PrP{sup C}-Fc protein enhances the IL-15-induced proliferation of human NK cells. • PrP{sup C}-Fc protein activates human NK cells via the ERK and JNK signaling pathways.

60 APPLIED LIFE SCIENCES↗

LncRNA EPIC1 protects human osteoblasts from dexamethasone-induced cell death

Highlights: • Dexamethasone (Dex) increases Lnc-EPIC1 expression in human osteoblasts. • Lnc-EPIC1 expression is elevated in necrotic femoral head tissues of Dex-taking patients. • Ectopic Lnc-EPIC1 overexpression protects human osteoblasts from Dex. • Lnc-EPIC1 siRNA potentiates Dex-induced cytotoxicity in human osteoblasts. • Myc is the primary target of Lnc-EPIC1 in human osteoblasts. Dexamethasone (Dex) can induce injury to human osteoblasts. Long non-coding RNA (LncRNA) EPIC1 (Lnc-EPIC1) is a novel Myc-interacting LncRNA. Its effect on Dex-treated human osteoblasts is studied here. In OB-6 osteoblastic cells and primary human osteoblasts, treatment with Dex increased expression of Lnc-EPIC1. Its expression is also elevated in the necrotic femoral head tissues of Dex-taking patients. Ectopic overexpression of Lnc-EPIC1 inhibited Dex-induced apoptosis and programmed necrosis in OB-6 cells and primary human osteoblasts. Reversely, Lnc-EPIC1 silencing by targeted siRNA potentiated Dex-induced cytotoxicity. Myc is the target of Lnc-EPIC1 in osteoblasts. Exogenous overexpression of Myc protected OB-6 cells from Dex. Conversely, Myc knockout by CRISPR-Cas-9 method abolished Lnc-EPIC1-induced OB-6 cytoprotection against Dex. Together, Lnc-EPIC1 expression protects human osteoblasts from Dex possible via regulation of Myc.

60 APPLIED LIFE SCIENCES↗

Bile acid-induced necrosis in primary human hepatocytes and in patients with obstructive cholestasis

Accumulation of bile acids is a major mediator of cholestatic liver injury. Recent studies indicate bile acid composition between humans and rodents is dramatically different, as humans have a higher percent of glycine conjugated bile acids and increased chenodeoxycholate content, which increases the hydrophobicity index of bile acids. This increase may lead to direct toxicity that kills hepatocytes, and promotes inflammation. To address this issue, this study assessed how pathophysiological concentrations of bile acids measured in cholestatic patients affected primary human hepatocytes. Individual bile acid levels were determined in serum and bile by UPLC/QTOFMS in patients with extrahepatic cholestasis with, or without, concurrent increases in serum transaminases. Bile acid levels increased in serum of patients with liver injury, while biliary levels decreased, implicating infarction of the biliary tracts. To assess bile acid-induced toxicity in man, primary human hepatocytes were treated with relevant concentrations, derived from patient data, of the model bile acid glycochenodeoxycholic acid (GCDC). Treatment with GCDC resulted in necrosis with no increase in apoptotic parameters. This was recapitulated by treatment with biliary bile acid concentrations, but not serum concentrations. Marked elevations in serum full-length cytokeratin-18, high mobility group box 1 protein (HMGB1), and acetylated HMGB1 confirmed inflammatory necrosis in injured patients; only modest elevations in caspase-cleaved cytokeratin-18 were observed. These data suggest human hepatocytes are more resistant to human-relevant bile acids than rodent hepatocytes, and die through necrosis when exposed to bile acids. These mechanisms of cholestasis in humans are fundamentally different to mechanisms observed in rodent models. - Highlights: • Cholestatic liver injury is due to cytoplasmic bile acid accumulation in hepatocytes. • Primary human hepatocytes are resistant to BA-induced injury compared to rodents. • Primary human hepatocytes largely undergo necrosis in response to BA toxicity. • Cholestatic liver injury in vivo is predominantly necrotic with minor apoptosis. • Rodent models of bile acid toxicity may not recapitulate the injury in man.

60 APPLIED LIFE SCIENCES↗

Characterization of a 2016-2017 Human Seasonal H3 Influenza A Virus Spillover Now Endemic to U.S. Swine

In 2017, the Iowa State University Veterinary Diagnostic Laboratory detected a reverse-zoonotic transmission of a human seasonal H3 influenza A virus into swine (IAV-S) in Oklahoma. Pairwise comparison between the recently characterized human seasonal H3 IAV-S (H3.2010.2) hemagglutinin (HA) sequences detected in swine and the most similar 2016–2017 human seasonal H3 revealed 99.9% nucleotide identity. To elucidate the origin of H3.2010.2 IAV-S, 45 HA and 27 neuraminidase (NA) sequences from 2017 to 2020 as well as 11 whole-genome sequences (WGS) were genetically characterized. Time to most recent common human ancestor was estimated between August and September 2016. The N2 NA was of human origin in all but one strain from diagnostic submissions with NA sequences, and the internal gene segments from WGS consisted of matrix genes originating from the 2009 pandemic H1N1 and another 5 internal genes of triple reassortant swine origin (TTTTPT). Pigs experimentally infected with H3.2010.2 demonstrated efficient nasal shedding and replication in the lungs, mild pneumonia, and minimal microscopic lung lesions and transmitted the virus to indirect contact swine. Antigenically, H3.2010.2 viruses were closer to a human seasonal vaccine strain, A/Hong Kong/4801/2014, than to the H3.2010.1 human seasonal H3 viruses detected in swine in 2012. This was the second sustained transmission of a human seasonal IAV into swine from the 2010 decade after H3.2010.1. Monitoring the spillover and detection of novel IAV from humans to swine may help vaccine antigen selection and could impact pandemic preparedness.

59 BASIC BIOLOGICAL SCIENCES↗

Lessons Learned from Performing a Human Factors Engineering Validation of an Upgraded Digital Control System in a Nuclear Power Plant Control Room

The United States' fleet of nuclear power plants (NPPs) have exceeded their original 40-year licenses and many are seeking license renewals to continue operations into the future. To support continued operation the Department of Energy's (DOE) Light Water Reactor Sustainability Program (LWRS) provides support for utilities to maintain and upgrade their power production capabilities. Control room modernization is one focus area, which aims to modernization the analog control rooms with digital controls and human-system interfaces (HSIs). The Turbine Control System (TCS) is a non-safety related system undergoing modernization efforts at multiple plants and utilities. Considerable human factors research efforts, comprised of expert reviews and operator in the loop studies were performed to develop human factors compliant and usable TCS. This report documents the final phase of the human factors engineering program used to design and validate the new digital TCS. According to the NUREG-0711 review guide, one of the last steps of human factors engineering program prior to implementation is human factors validation. This report details the method, results and conclusions from the human factors validation study of a collaborating utilities TCS slated to be implemented in one of their plants. One control room team consisting of two control room supervisors and two reactor operators participated in the validation. All team members had participated in the BNP TCS training program foregoing the TCS validation. Performance measures included task performance, situation awareness and workload, expert observation of any negative influences of human-system interface features on control room team performance, and interviews with the participating operators. Performance criteria were based on the performance scores and the topics identified by interviews and expert observations. The results showed that the crew using the new TCS clearly could achieve their overall objectives in the test scenarios with sufficient situation awareness. The new TCS compared to the old system provided clear improvements of the control room work regarding shell and chest warming and regarding valve and other system testing. However, a number of Human Engineering Discrepancies (HEDs) were identified. HEDs were prioritized (level 1, 2 or 3) according to potential safety, plant availability, operability or personnel performance consequences. Since no priority 1 HEDs were identified there is no HFE reason not to continue implementation of the TCS. The TCS upgrade is acceptable if the Priority 2 HEDs are resolved and dispositioned in a satisfactory way. Priority 3 HEDs should be noted and considered for future modifications.

21 SPECIFIC NUCLEAR REACTORS AND ASSOCIATED PLANTS↗

Simulator Data Analysis to Inform Digitalized Environment Impacts on Human Reliability

The U.S. Nuclear Regulatory Commission (NRC) has developed a human reliability analysis (HRA) method, termed the Integrated Human Event Analysis System for Event and Condition Assessment (IDHEAS-ECA), in order to estimate human error probabilities (HEPs) in risk-informed regulatory applications. To update the quantification part of IDHEAS-ECA, the NRC required human performance and error data from fully digitalized main control rooms (MCRs); therefore, it requested that Idaho National Laboratory (INL) revisit previous data collection studies and investigate how the following three factors impact human reliability: self-checking, peer-checking, and automation. The HRA data collection studies revisited were the Human Reliability Data Extraction (HuREX) project, developed by the Korea Atomic Energy Research Institute (KAERI), and the Simplified Human Error Experimental Program (SHEEP), developed by INL. HuREX is a representative HRA data collection study that collects human reliability data from full-scope simulators staffed by licensed operators. SHEEP, on the other hand, has been proposed to complement such full-scope studies by collecting data via simplified simulators staffed by non-licensed student operators. In the HuREX study, KAERI collected HRA data from fully digitalized MCRs for the Advanced Power Reactor (APR)–1400. The SHEEP data were obtained from simplified simulators that partially mimicked the features of digitalized MCRs. The present report mainly discusses how the impacts of the aforementioned three factors on human errors were derived from these two data collection studies.

99 GENERAL AND MISCELLANEOUS↗

Long non-coding RNA THOR promotes human osteosarcoma cell growth in vitro and in vivo

Highlights: • LncRNA THOR (“Lnc-THOR”) is expressed in human osteosarcoma (OS) cells and tissues. • Lnc-THOR siRNA inhibits human OS cell survival and proliferation. • Lnc-THOR over-expression promotes human OS cell survival and proliferation. • Lnc-THOR knockout inhibits U2OS cell growth in vitro and in vivo Long non-coding RNA (LncRNA) dysregulation is associated with human osteosarcoma (OS) cell progression. Recent studies have characterized a novel but ultra-conserved LncRNA THOR (“Lnc-THOR”) as a cancer-specific LncRNA, mediating cell growth. In the current study, we show that Lnc-THOR is expressed in established and primary human OS cells. It is also detected in human OS tissues, but not in the surrounding normal bone tissues. siRNA-induced knockdown or CRSIPR/Cas9-mediated knockout Lnc-THOR significantly inhibited human OS cell survival and proliferation. Insulin-like growth factor 2 mRNA-binding protein 1 (IGF2BP1) target mRNAs, including IGF2, GLI1 and CD44, were downregulated in Lnc-THOR-silenced OS cells as well. Conversely, forced over-expression of Lnc-THOR enhanced IGF2BP1 target mRNA expression, promoting OS cell survival and proliferation. In vivo, xenograft tumors of Lnc-THOR-knockout U2OS cells grew significantly slower than the control U2OS tumors. Together, these results show that Lnc-THOR expression is essential for human OS cell growth. Lnc-THOR could be a novel therapeutic target and/or diagnosis marker for human OS.

60 APPLIED LIFE SCIENCES↗

Cell adhesion properties of human defensins

Highlights: • Human α-Defensin 5 and Human Neutrophil Peptide 1 directly bind to fibronectin. • Human α-Defensin 5 and Human Neutrophil Peptide 1 modulate cell adhesion and cellto-cell interactions. • These properties do not extend to other human defensin peptides. Effector peptides of innate immunity play an important role in host defense. They act directly by inactivating microbes but also link innate to adaptive immunity. A variety of innate immune functions has been described for these peptides, including chemoattraction and cytokine release. In this study, we describe the effect on cell morphology and cell adhesion of human defensins. We find that Human Defensin 5, the major product of specialized gut epithelial cells, causes changes in cell morphology. HD-5 induces cell adhesion, binds to fibronectin and facilitates binding of T cells to intestinal epithelial cells. These effects were found also for a second prominent defensing, termed Human Neutrophil peptide-1, but not for other human defensins.

60 APPLIED LIFE SCIENCES↗

Potential transfer of neurotoxic amino acid β-N-methylamino-alanine (BMAA) from mother to infant during breast-feeding: Predictions from human cell lines

β-N-methylamino-alanine (BMAA) is a non-protein amino acid produced by cyanobacteria, diatoms and dinoflagellates. BMAA has potential to biomagnify in a terrestrial food chain, and to bioaccumulate in fish and shellfish. We have reported that administration of [{sup 14}C]L-BMAA to lactating mice and rats results in a mother to off-spring transfer via the milk. A preferential enantiomer-specific uptake of [{sup 14}C]L-BMAA has also been demonstrated in differentiated murine mammary epithelium HC11 cells. These findings, together with neurotoxic effects of BMAA demonstrated both in vitro and in vivo, highlight the need to determine whether such transfer could also occur in humans. Here, we used four cell lines of human origin to examine and compare the transport of the two BMAA enantiomers in vitro. The uptake patterns of [{sup 14}C]L- and [{sup 14}C]D-BMAA in the human mammary MCF7 cell line were in agreement with the results in murine HC11 cells, suggesting a potential secretion of BMAA into human breast milk. The permeability coefficients for both [{sup 14}C]L- and [{sup 14}C]D-BMAA over monolayers of human intestinal Caco2 cells supported an efficient absorption from the human intestine. As a final step, transport experiments confirmed that [{sup 14}C]L-and [{sup 14}C]D-BMAA can be taken up by human SHSY5Y neuroblastoma cells and even more efficiently by human U343 glioblastoma cells. In competition experiments with various amino acids, the ASCT2 specific inhibitor benzylserine was the most effective inhibitor of [{sup 14}C]L-BMAA uptake tested here. Altogether, our results suggest that BMAA can be transferred from an exposed mother, via the milk, to the brain of the nursed infant. - Highlights: • Transport of BMAA in human intestinal, mammary and CNS cell lines was examined. • The transport of L-BMAA over intestinal cell monolayers was unidirectional. • Enantiomer-selective uptake of L-BMAA in breast, neuron and glia cells was evident. • Competition experiments indicate that L-BMAA uptake involved several transporters. • A potential for mother to infant transfer of BMAA is proposed.

60 APPLIED LIFE SCIENCES↗

Differential Misfolding Properties of Glaucoma-Associated Olfactomedin Domains from Humans and Mice

Mutations in myocilin, predominantly within its olfactomedin (OLF) domain, are causative for the heritable form of open angle glaucoma in humans. Surprisingly, mice expressing Tyr423His mutant myocilin, corresponding to a severe glaucoma-causing mutation (Tyr437His) in human subjects, exhibit a weak, if any, glaucoma phenotype. To address possible protein-level discrepancies between mouse and human OLFs, which might lead to this outcome, biophysical properties of mouse OLF were characterized for comparison with those of human OLF. Here, the 1.55 Å resolution crystal structure of mouse OLF reveals an asymmetric 5-bladed β-propeller that is nearly indistinguishable from previous structures of human OLF. Wild-type and selected mutant mouse OLFs mirror thermal stabilities of their human OLF counterparts, including characteristic stabilization in the presence of calcium. Mouse OLF forms thioflavin T-positive aggregates with a similar end-point morphology as human OLF, but amyloid aggregation kinetic rates of mouse OLF are faster than human OLF. Simulations and experiments support the interpretation that kinetics of mouse OLF are faster because of a decreased charge repulsion arising from more neutral surface electrostatics. Taken together, phenotypic differences observed in mouse and human studies of mutant myocilin could be a function of aggregation kinetics rates, which would alter the lifetime of putatively toxic protofibrillar intermediates.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗