Glioblastoma Cells Use an Integrin- and CD44-Mediated Motor-Clutch Mode of Migration in Brain Tissue.

Purpose: Glioblastoma (GBM) is an aggressive malignant brain tumor with 2 year survival rates of 6.7% (Stupp et al. in J Clin Oncol Off J Am Soc Clin Oncol 25:4127-4136, 2007; Mohammed et al. in Rep Pract Oncol Radiother 27:1026-1036, 2002). One key characteristic of the disease is the ability of glioblastoma cells to migrate rapidly and spread throughout healthy brain tissue (Lefranc et al. in J Clin Oncol Off J Am Soc Clin Oncol 23:2411-2422, 2005; Hoelzinger et al. in J Natl Cancer Inst 21:1583-1593, 2007). To develop treatments that effectively target cell migration, it is important to understand the fundamental mechanism driving cell migration in brain tissue. Several models of cell migration have been proposed, including the motor-clutch, bleb-based motility, and osmotic engine models. Methods: Here we utilized confocal imaging to measure traction dynamics and migration speeds of glioblastoma cells in mouse organotypic brain slices to identify the mode of cell migration. Results: We found that nearly all cell-vasculature interactions reflected pulling, rather than pushing, on vasculature at the cell leading edge, a finding consistent with a motor-clutch mode of migration, and inconsistent with an osmotic engine model or confined bleb-based migration. Reducing myosin motor activity, a key component in the motor-clutch model, was found to decrease migration speed at high doses for all cell types including U251 and 6 low-passage patient-derived xenograft lines (3 proneural and 3 mesenchymal subtypes). Variable responses were found at low doses, consistent with a motor-clutch mode of migration which predicts a biphasic relationship between migration speed and motor-to-clutch ratio. Targeting of molecular clutches including integrins and CD44 slowed migration of U251 cells. Conclusions: Overall we find that glioblastoma cell migration is most consistent with a motor-clutch mechanism to migrate through brain tissue ex vivo, and that both integrins and CD44, as well as myosin motors, play an important role in constituting the adhesive clutch. Supplementary Information: The online version contains supplementary material available at 10.1007/s12195-024-00799-x.

The OTX2 Gene Induces Tumor Growth and Triggers Leptomeningeal Metastasis by Regulating the mTORC2 Signaling Pathway in Group 3 Medulloblastomas.

Medulloblastoma (MB) encompasses diverse subgroups, and leptomeningeal disease/metastasis (LMD) plays a substantial role in associated fatalities. Despite extensive exploration of canonical genes in MB, the molecular mechanisms underlying LMD and the involvement of the orthodenticle homeobox 2 (OTX2) gene, a key driver in aggressive MB Group 3, remain insufficiently understood. Recognizing OTX2's pivotal role, we investigated its potential as a catalyst for aggressive cellular behaviors, including migration, invasion, and metastasis. OTX2 overexpression heightened cell growth, motility, and polarization in Group 3 MB cells. Orthotopic implantation of OTX2-overexpressing cells in mice led to reduced median survival, accompanied by the development of spinal cord and brain metastases. Mechanistically, OTX2 acted as a transcriptional activator of the Mechanistic Target of Rapamycin (mTOR) gene's promoter and the mTORC2 signaling pathway, correlating with upregulated downstream genes that orchestrate cell motility and migration. Knockdown of mTOR mRNA mitigated OTX2-mediated enhancements in cell motility and polarization. Analysis of human MB tumor samples (N = 952) revealed a positive correlation between OTX2 and mTOR mRNA expression, emphasizing the clinical significance of OTX2's role in the mTORC2 pathway. Our results reveal that OTX2 governs the mTORC2 signaling pathway, instigating LMD in Group 3 MBs and offering insights into potential therapeutic avenues through mTORC2 inhibition.

Paediatric emergency front-of-neck airway: issues of ethics, law, and philosophy.

Practitioners can face significant challenges when managing the airways of infants and neonates because of their unique anatomical and physiological features. The requirement for emergency airway management in this age group is rare. Details of emergency airway techniques in paediatric practice guidelines are missing or lack consensus, and it is known that outcomes for affected children can be poor. Ideally, these children should be managed by experienced paediatric airway practitioners working in a team. However, situations can arise where practitioners, unfamiliar and inexperienced with infants, find themselves in charge. So, what happens when such a practitioner encounters this life-or-death scenario and feels ill-equipped to act? The ethical and legal issues surrounding the management of this emergency are clearly defined, but they can be unknown or misunderstood by doctors. Compounding the extreme stress of the scenario is the moral and ethical dilemma of whether to act or not. The following discussion explores these issues and examines the philosophical and psychological perspectives.

Glioblastoma cells use an integrin- and CD44-mediated motor-clutch mode of migration in brain tissue.

Glioblastoma (GBM) is an aggressive malignant brain tumor with 2-year survival rates of 6.7% [1], [2]. One key characteristic of the disease is the ability of glioblastoma cells to migrate rapidly and spread throughout healthy brain tissue[3], [4]. To develop treatments that effectively target cell migration, it is important to understand the fundamental mechanism driving cell migration in brain tissue. Here we utilized confocal imaging to measure traction dynamics and migration speeds of glioblastoma cells in mouse organotypic brain slices to identify the mode of cell migration. Through imaging cell-vasculature interactions and utilizing drugs, antibodies, and genetic modifications to target motors and clutches, we find that glioblastoma cell migration is most consistent with a motor-clutch mechanism to migrate through brain tissue ex vivo, and that both integrins and CD44, as well as myosin motors, play an important role in constituting the adhesive clutch.

Disability rights and empowerment: Reflections on AJCP research and a call to action.

People living with physical, sensory, intellectual, and/or developmental disabilities experience complex social, environmental, political, and cultural challenges along with stigma and marginalization in education, employment, and community life. These multiple and complex barriers often hinder their full and effective participation in society. In this reflection, we curated articles on physical, sensory, intellectual, and/or developmental disabilities published in the American Journal of Community Psychology from 1973 to 2022. We reviewed titles and abstracts to identify themes that grouped manuscripts in relevant community psychology core concepts and values. From our analysis, five themes emerged: (a) promoting empowerment and advocacy; (b) promoting organizations and settings that support people with disabilities; (c) including people with disabilities in knowledge production; (d) promoting social justice in disability research, and (e) promoting support networks of families of people with disabilities. We conclude this reflection with a discussion of recommendations for future research, practice, and a call to action.

RAD-TGTs: high-throughput measurement of cellular mechanotype via rupture and delivery of DNA tension probes.

Mechanical forces drive critical cellular processes that are reflected in mechanical phenotypes, or mechanotypes, of cells and their microenvironment. We present here "Rupture And Deliver" Tension Gauge Tethers (RAD-TGTs) in which flow cytometry is used to record the mechanical history of thousands of cells exerting forces on their surroundings via their propensity to rupture immobilized DNA duplex tension probes. We demonstrate that RAD-TGTs recapitulate prior DNA tension probe studies while also yielding a gain of fluorescence in the force-generating cell that is detectable by flow cytometry. Furthermore, the rupture propensity is altered following disruption of the cytoskeleton using drugs or CRISPR-knockout of mechanosensing proteins. Importantly, RAD-TGTs can differentiate distinct mechanotypes among mixed populations of cells. We also establish oligo rupture and delivery can be measured via DNA sequencing. RAD-TGTs provide a facile and powerful assay to enable high-throughput mechanotype profiling, which could find various applications, for example, in combination with CRISPR screens and -omics analysis.

Closing the Information Gap: Making COVID-19 Information Accessible for People with Disabilities.

It is essential that people with disabilities have equitable access to COVID-19 communication resources to protect themselves, their families, and their communities. The Accessible Materials and Culturally Relevant Messages for Individuals with Disabilities project aimed to deliver essential COVID-19 information in braille, American Sign Language (ASL), simplified text, and other alternative formats, along with providing additional tools and trainings that people with disabilities and organizations that serve them can use to apply the COVID-19 guidance. Lessons learned from this project can be implemented in future public health emergencies as well as in general public health messaging for people with disabilities. This project, led by Georgia Tech's Center for Inclusive Design and Innovation (CIDI) and with technical assistance from the Centers for Disease Control and Prevention (CDC), was supported by the CDC Foundation, using funds from the CDC Foundation's COVID-19 Emergency Response Fund.

A Scoping Review of the Pharmacy Curriculum Outcomes Assessment Literature.

Objective. To identify themes, gaps, and sources of evidence from the literature regarding the Pharmacy Curriculum Outcomes Assessment (PCOA) to inform practice and additional areas for research in pharmacy education.Findings. Nineteen articles describing the administration and use of PCOA were identified. Since PCOA was made a curricular requirement by the Accreditation Council of Pharmacy Education in 2016, the focus of literature related to the PCOA has shifted from administration practices (four articles published before 2016 vs two articles published since) to determining models that may predict student performance on the assessment (two vs five articles) or how the examination might be used to predict future performance (one vs seven articles), especially on the North American Pharmacist Licensing Examination. While there is a growing body of literature focused on the PCOA's utility for measuring performance, few variables have been consistently used.Summary. This review found no studies with objectives that aligned with the initial intended use of the PCOA as defined by the National Association of Boards of Pharmacy, which included tracking individual student performance throughout the curriculum, benchmarking programs against other programs, and evaluating whether a program was meeting their desired outcomes. Additionally, no consensus across the Academy was found as to the proper use of the PCOA, and a paucity of literature exists regarding how the PCOA informs schools and colleges about the effectiveness of their curriculum. There is a need for the Academy to establish a uniform application for the PCOA in pharmacy schools, assess the resources that programs need to administer this required assessment, and determine the utility of the PCOA to measure curricular effectiveness and/or student performance.

Failure of the "Gold Standard": The Role of a Mixed Methods Research Toolkit and Human-Centered Design in Transformative WASH.

From preventing cholera and diarrhea by reducing exposure to human waste, to reducing transmission of COVID-19 through handwashing, water, sanitation and hygiene (WASH) can save lives. Numerous global health initiatives have been created to combat the spread of infectious diseases. However, according to the Sanitation and Hygiene Fund, "decades of under investment in sanitation and hygiene have made this sector the weakest link in our efforts to achieve the [Sustainable Development Goals (SDGs)]." There appear to be various reasons for the lag in global attention to, funding toward, and innovation around WASH-related diseases. Firstly, WASH is comprised of three interrelated components, water, sanitation, and hygiene, each of which has its own subset of indicators, priorities, and infrastructure, thus making streamlined communications and impact measurement within the sector incredibly complex. Secondly, WASH is a field that bridges many sectors, and there has historically been a lack of understanding of where responsibility lies to consistently fund and execute WASH interventions, programming, and policymaking. Additionally, public health research and funding tend to favor evaluations using randomized controlled trials (RCTs), which are often referred to as the "gold standard." RCTs, like all single evaluative methods, have limitations which may not effectively capture the complexity of WASH interventions and their subsequent multi-sectoral outcomes. In some cases "it may be infeasible (or unethical) to randomize communities to a [WASH] intervention" which would prohibit the research from reaching the current "gold standard" threshold for academic rigor and subsequent funding. A new concept called "Transformative WASH" has recently emerged in the WASH sector as a result of three RCTs and calls for a "comprehensive package of WASH interventions" to effectively improve health and social outcomes. We believe that the current definition of the "gold standard" in academic research is failing the WASH sector and does not align with "Transformative WASH." Rather, the "gold standard" should instead be a mixed methods research toolkit that utilizes Human-Centered Design (HCD) practices, and proxy methods such as "participatory design" or "Behavior Centered Design theory" to better design and evaluate WASH interventions.

Smart Sanitation-Biosensors as a Public Health Tool in Sanitation Infrastructure.

The health of individuals and communities is more interconnected than ever, and emergent technologies have the potential to improve public health monitoring at both the community and individual level. A systematic literature review of peer-reviewed and gray literature from 2000-present was conducted on the use of biosensors in sanitation infrastructure (such as toilets, sewage pipes and septic tanks) to assess individual and population health. 21 relevant papers were identified using PubMed, Embase, Global Health, CDC Stacks and NexisUni databases and a reflexive thematic analysis was conducted. Biosensors are being developed for a range of uses including monitoring illicit drug usage in communities, screening for viruses and diagnosing conditions such as diabetes. Most studies were nonrandomized, small-scale pilot or lab studies. Of the sanitation-related biosensors found in the literature, 11 gathered population-level data, seven provided real-time continuous data and 14 were noted to be more cost-effective than traditional surveillance methods. The most commonly discussed strength of these technologies was their ability to conduct rapid, on-site analysis. The findings demonstrate the potential of this emerging technology and the concept of Smart Sanitation to enhance health monitoring at the individual level (for diagnostics) as well as at the community level (for disease surveillance).

Regulation and dynamics of force transmission at individual cell-matrix adhesion bonds.

Integrin-based adhesion complexes link the cytoskeleton to the extracellular matrix (ECM) and are central to the construction of multicellular animal tissues. How biological function emerges from the tens to thousands of proteins present within a single adhesion complex remains unclear. We used fluorescent molecular tension sensors to visualize force transmission by individual integrins in living cells. These measurements revealed an underlying functional modularity in which integrin class controlled adhesion size and ECM ligand specificity, while the number and type of connections between integrins and F-actin determined the force per individual integrin. In addition, we found that most integrins existed in a state of near-mechanical equilibrium, a result not predicted by existing models of cytoskeletal force transduction. A revised model that includes reversible cross-links within the F-actin network can account for this result and suggests one means by which cellular mechanical homeostasis can arise at the molecular level.

Innate Immunity in the C. elegans Intestine Is Programmed by a Neuronal Regulator of AWC Olfactory Neuron Development.

Olfactory neurons allow animals to discriminate nutritious food sources from potential pathogens. From a forward genetic screen, we uncovered a surprising requirement for the olfactory neuron gene olrn-1 in the regulation of intestinal epithelial immunity in Caenorhabditis elegans. During nematode development, olrn-1 is required to program the expression of odorant receptors in the AWC olfactory neuron pair. Here, we show that olrn-1 also functions in AWC neurons in the cell non-autonomous suppression of the canonical p38 MAPK PMK-1 immune pathway in the intestine. Low activity of OLRN-1, which activates the p38 MAPK signaling cassette in AWC neurons during larval development, also de-represses the p38 MAPK PMK-1 pathway in the intestine to promote immune effector transcription, increased clearance of an intestinal pathogen, and resistance to bacterial infection. These data reveal an unexpected connection between olfactory receptor development and innate immunity and show that anti-pathogen defenses in the intestine are developmentally programmed.

Redirection of SKN-1 abates the negative metabolic outcomes of a perceived pathogen infection.

Early host responses toward pathogens are essential for defense against infection. In Caenorhabditis elegans, the transcription factor, SKN-1, regulates cellular defenses during xenobiotic intoxication and bacterial infection. However, constitutive activation of SKN-1 results in pleiotropic outcomes, including a redistribution of somatic lipids to the germline, which impairs health and shortens lifespan. Here, we show that exposing C. elegans to Pseudomonas aeruginosa similarly drives the rapid depletion of somatic, but not germline, lipid stores. Modulating the epigenetic landscape refines SKN-1 activity away from innate immunity targets, which alleviates negative metabolic outcomes. Similarly, exposure to oxidative stress redirects SKN-1 activity away from pathogen response genes while restoring somatic lipid distribution. In addition, activating p38/MAPK signaling in the absence of pathogens, is sufficient to drive SKN-1-dependent loss of somatic fat. These data define a SKN-1- and p38-dependent axis for coordinating pathogen responses, lipid homeostasis, and survival and identify transcriptional redirection, rather than inactivation, as a mechanism for counteracting the pleiotropic consequences of aberrant transcriptional activity.

Raman Spectroscopy for Rapid Evaluation of Surgical Margins during Breast Cancer Lumpectomy.

Failure to precisely distinguish malignant from healthy tissue has severe implications for breast cancer surgical outcomes. Clinical prognoses depend on precisely distinguishing healthy from malignant tissue during surgery. Laser Raman spectroscopy (LRS) has been previously shown to differentiate benign from malignant tissue in real time. However, the cost, assembly effort, and technical expertise needed for construction and implementation of the technique have prohibited widespread adoption. Recently, Raman spectrometers have been developed for non-medical uses and have become commercially available and affordable. Here we demonstrate that this current generation of Raman spectrometers can readily identify cancer in breast surgical specimens. We evaluated two commercially available, portable, near-infrared Raman systems operating at excitation wavelengths of either 785 nm or 1064 nm, collecting a total of 164 Raman spectra from cancerous, benign, and transitional regions of resected breast tissue from six patients undergoing mastectomy. The spectra were classified using standard multivariate statistical techniques. We identified a minimal set of spectral bands sufficient to reliably distinguish between healthy and malignant tissue using either the 1064 nm or 785 nm system. Our results indicate that current generation Raman spectrometers can be used as a rapid diagnostic technique distinguishing benign from malignant tissue during surgery.

Modeling distributed forces within cell adhesions of varying size on continuous substrates.

Cell migration and traction are essential to many biological phenomena, and one of their key features is sensitivity to substrate stiffness, which biophysical models, such as the motor-clutch model and the cell migration simulator can predict and explain. However, these models have not accounted for the finite size of adhesions, the spatial distribution of forces within adhesions. Here, we derive an expression that relates varying adhesion radius ( R) and spatial distribution of force within an adhesion (described by s) to the effective substrate stiffness ( κsub ), as a function of the Young's modulus of the substrate ( E Y ), which yields the relation, κsub=RsEY , for two-dimensional cell cultures. Experimentally, we found that a cone-shaped force distribution ( s = 1.05) can describe the observed displacements of hydrogels deformed by adherent U251 glioma cells. Also, we found that the experimentally observed adhesion radius increases linearly with the cell protrusion force, consistent with the predictions of the motor-clutch model with spatially distributed clutches. We also found that, theoretically, the influence of one protrusion on another through a continuous elastic environment is negligible. Overall, we conclude cells can potentially control their own interpretation of the mechanics of the environment by controlling adhesion size and spatial distribution of forces within an adhesion.

The fatty acid oleate is required for innate immune activation and pathogen defense in Caenorhabditis elegans.

Fatty acids affect a number of physiological processes, in addition to forming the building blocks of membranes and body fat stores. In this study, we uncover a role for the monounsaturated fatty acid oleate in the innate immune response of the nematode Caenorhabditis elegans. From an RNAi screen for regulators of innate immune defense genes, we identified the two stearoyl-coenzyme A desaturases that synthesize oleate in C. elegans. We show that the synthesis of oleate is necessary for the pathogen-mediated induction of immune defense genes. Accordingly, C. elegans deficient in oleate production are hypersusceptible to infection with diverse human pathogens, which can be rescued by the addition of exogenous oleate. However, oleate is not sufficient to drive protective immune activation. Together, these data add to the known health-promoting effects of monounsaturated fatty acids, and suggest an ancient link between nutrient stores, metabolism, and host susceptibility to bacterial infection.

The nuclear hormone receptor NHR-86 controls anti-pathogen responses in C. elegans.

Nuclear hormone receptors (NHRs) are ligand-gated transcription factors that control adaptive host responses following recognition of specific endogenous or exogenous ligands. Although NHRs have expanded dramatically in C. elegans compared to other metazoans, the biological function of only a few of these genes has been characterized in detail. Here, we demonstrate that an NHR can activate an anti-pathogen transcriptional program. Using genetic epistasis experiments, transcriptome profiling analyses and chromatin immunoprecipitation-sequencing, we show that, in the presence of an immunostimulatory small molecule, NHR-86 binds to the promoters of immune effectors to activate their transcription. NHR-86 is not required for resistance to the bacterial pathogen Pseudomonas aeruginosa at baseline, but activation of NHR-86 by this compound drives a transcriptional program that provides protection against this pathogen. Interestingly, NHR-86 targets immune effectors whose basal regulation requires the canonical p38 MAPK PMK-1 immune pathway. However, NHR-86 functions independently of PMK-1 and modulates the transcription of these infection response genes directly. These findings characterize a new transcriptional regulator in C. elegans that can induce a protective host response towards a bacterial pathogen.