Stanford University

NSF Awards · FY 2024 · 2024-09

How new species are formed is a grand challenge across biology. Particular combinations of genes from different populations may not interact favorably during hybridization, creating unhealthy offspring due to genetic incompatibilities. The specific genes involved in this process are seldom identified, but genes involved in mitochondrial function are prime candidates and the focus of recent research. This project will investigate how different combinations of mitochondrial and nuclear genes create reproductive barriers in hybridizing swordtail fish (Xiphophorus), a model system for genetic incompatibilities. Genomic tools will be used to identify which specific combinations of nuclear and mitochondrial genes influence overall hybrid health. Specific aspects of mitochondrial function such as respiratory efficiency will also be investigated as a mechanistic basis for hybrid incompatibilities. Genetic incompatibilities will also be investigated in different environments and developmental stages because incompatibilities may only manifest in certain conditions. This work includes generating hybrids in the laboratory by targeted crossing experiments and examining natural populations with ongoing hybridization. These activities will be used to recruit students from diverse backgrounds to STEM research, especially in the opportunity-rich field of bioinformatics. Freshmen will be explicitly targeted though the development of a new program called “Power in the Powerhouses” as part of the University of Texas at Austin’s highly successful Freshman Research Initiative to recruit and train the next generation of STEM researchers. Coevolution between nuclear and cytoplasmic genomes can create coadapted genomes within a population that may be disrupted during hybridization, creating reproductive isolation and acting as a common mechanism of speciation. Under this hypothesis, selection during introgressive hybridization should act to maintain coadapted cytonuclear genotypes. To test this hypothesis, genome-wide patterns of ancestry will be generated from three naturally hybridizing pairs and three lab-generated hybrid pairs of swordtail fish species (genus Xiphophorus). Selection should especially favor matched ancestry between mitochondrial genomes and the subset of nuclear-encoded genes that interact with mitochondrial-encoded gene products. Mitonuclear incompatibilities will be identified through statistical associations between nuclear alleles and mitotypes in natural and lab-bred hybrids. Lethal mitonuclear incompatibilities have already been identified using this approach in one pair of hybridizing Xiphophorus. Mitochondrial- and nuclear-interacting genes should also show concordant clines in ancestry with geography. Compromised energetic phenotypes as well as reduced organismal fitness should result from incompatible combinations of mitochondrial and nuclear genes, likely in an environmentally-dependent context. Therefore, in addition to standard metrics of organismal health, whole-organism metabolic rates and mitochondrial DNA copy number will be assessed in parental Xiphophorus species and their hybrids in response to thermal and hypoxic stressors. Multiple respiratory phenotypes in isolated mitochondria will also be investigated, including those dependent on mitonuclear interactions. Phenotypes will be assessed in adults and embryos, as lethal mitonuclear incompatibilities can prevent embryos from developing. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NSF Awards · FY 2024 · 2024-09

This project focuses on the heart of one of the most imperiled ecosystems, coral reefs, where climate change and heat waves have killed many corals. Corals live in a fine-tuned symbiosis with single celled algae, and heatwaves severely disrupt this symbiosis: even a few degrees can cause the partnership to breakdown. The team's preliminary data suggest that this sensitivity arises from how well individual coral and algal partners interact with one another at the cellular level such that corals with better genetic matches to their symbiont seem to resist heat better. This project is identifying genes and regions of the genome that interact to result in a functional partnership and testing this relationship across a range of environmental conditions. The results of these experiments, in combination with mobile sequencing platforms that generate data in real time, are bolstering conservation efforts by finding optimal populations for restoration projects driven by local communities in Palau. This allows quick translation and scaling the results of the experiments into conservation strategies by quickly identifying colonies with optimal host-symbiont combinations, improving the efficiency and efficacy of local coral restoration projects in Palau. The project also provides research training for postdoctoral scientists, graduate students and a research associate, and public outreach through the production of video and multimedia products. For ecosystems dependent on symbioses, climate resilience depends on successful interactions between partners (i.e. GxGxE interactions). In corals, this variation has been investigated at the level of coral genetic variants (within or between species) or genus-level symbiont variants. Yet, despite a great deal of careful work, these data have not yet revealed strong impact of specific coral genes on bleaching variation. This project is 1) characterizing population-level variation in both the host and symbiont populations to identify pairs of genetic loci in the host and symbiont genome that are highly correlated (i.e. exhibit strong linkage disequilibrium), termed ‘matched’ loci; 2) testing for these matched loci in three species of Acropora from three locations in Palau that historically have exhibited dramatically different thermal profiles and identifying how matching relates to bleaching resistance; 3) reciprocally transplanting colonies from all three locations to test for local adaptation at the level of the holobiont, specifically focusing on whether the ‘matching’ profile changes in different environmental contexts; 4) identifying and characterizing colonies whose dominant symbiont partner switched after transplantation to elucidate the selective forces shaping symbiont genotypes in light of both their environmental and host-specific context; and 5) partnering with local governments and communities to implement restoration plots using rapid and mobile sequencing platforms to test if identifying and using genetically optimal host and symbiont partners in restoration efforts helps to improve conservation outcomes. This project is jointly funded by Biological Oceanography (GEO/OCE) and Organismal Response to Climate Change (BIO/IOS). This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NSF Awards · FY 2024 · 2024-09

Modern Earth is covered by lush tropical forests, extensive grasslands, and soaring redwoods—in striking contrast to landscapes through much of Earth’s early history that consisted largely of bare rock and microbial mats. Plants have dramatically altered Earth’s landscape and climate (like the shapes of rivers and patterns of rainfall). However, there is currently little consensus on how the development of plants, starting with the first ground-hugging mosses and liverworts around 470 million years ago, followed by the eventual rise of trees around 380 million years ago, affected nutrient and oxygen levels both on land and in the oceans. This research combines field, laboratory, and modeling approaches to examine the effects of early land plants on the Earth system. This study focuses on the Canadian Arctic Archipelago which contains some of the best-preserved sedimentary rocks chronicling this key time period of early plant evolution. The team of researchers are studying fossil plants, pollen, and spores and geochemical elements to understand how weathering changed on land, how plant material was delivered to the ocean, how the availability of critical nutrients like phosphorus changed on the land and in the oceans, and how oxygen and sulfur levels changed in the ocean. The broader impacts activities stemming from the research include educational and mentorship opportunities for students in middle-school through graduate school. Graduate students will be co-mentored by the Principal Investigators, and undergraduates will also be recruited to analyze collected samples. The Yale Peabody Museum and the Yale Pathways to Science program will provide platforms for community-oriented outreach efforts, including educational events fostering scientific literacy and engagement in local middle-school students. The team will also take advantage of the unique opportunity provided by recent Peabody renovations to develop a new public-facing exhibit on “Ecosystem Engineering” focused on land plants and their impacts on Earth’s landscapes and ecosystems. The University of California, Riverside’s Camp Highlander program is fostering local high-school student engagement with Earth sciences. Finally, field-conducted telepresence outreach through the new “Annals of the Arctic” program, integrated with existing summer programs at Stanford, Yale, and UCR, will provide public-facing exposure to day-by-day realities of geologic fieldwork in remote terrains. This will increase the accessibility of geologic research and provide a venue for direct illustration of geologic concepts, human experiences of the dynamic nature of polar ecosystems, and their vulnerability to ongoing environmental change. Reconstructing the biotic, biogeochemical and climatic impacts of the evolution of land plants has been hampered by the commonly fragmentary and disassociated records of geochemical and paleontological change across the lower-middle Paleozoic transition, and by the limited integration of empirical observations with the mechanistic framework that can be provided by biogeochemical and Earth-system models. To address these fundamental questions, we are generating new, high-resolution field-based geochemical data (biomarker, programmed pyrolysis, carbon isotope, lithium isotope, osmium isotope, phosphorus speciation and phosphate-oxygen isotope, iron speciation, and trace metal abundances) and sedimentological and paleontological (plant body fossils, palynomorphs, graptolite and conodont biostratigraphy) records from key sections in the Canadian Arctic to reconstruct first-order ecological and environmental changes—in both continental and marine settings—concurrent with the radiation of early land plants. The Silurian–Devonian transition is an under-characterized but key interval for both land plant evolution and marine redox state, and these data will be integrated with long-term records to distinguish perturbations from more permanent state shifts. These new empirical records will be coupled to biogeochemical modeling over a range of scales—from local critical zone and seafloor diagenetic processes to continental climate and ocean and atmospheric carbon-oxygen cycle modeling—to develop a more robust process-based understanding of plant-biogeochemical feedbacks and reconstruct the long-term consequences of early land plant evolution for the broader Earth system. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NIH Research Projects · FY 2025 · 2024-09

Project Summary Alcohol Use Disorder (AUD) contributes to 88,000 deaths per year in the United States. Although several pharmacological treatments are available, adherence to these treatments is low and approximately 60% of individuals relapse within 6 months. Further, these treatments modulate the brain in a relatively global fashion. Evidence from neuroimaging studies of AUD patients has shown that elevated cortico-striatal activity in response to alcohol cues predicts relapse. Thus, there is an emerging interest in developing novel, neural-circuit specific therapeutic tools to enhance AUD treatment outcomes. Transcranial magnetic stimulation (TMS) is one such non-invasive, neural-circuit specific tool. Through electromagnetic induction, repetitive pulses of TMS can be applied to the cortex to change neural activity within a cortical circuit. Subsequently, TMS for AUD has been developed as a strategy to reduce brain reactivity to alcohol cues within the prefrontal cortex and its downstream targets in the striatum. I recently led an analysis of a randomized, double-blind clinical trial applying 10 days of TMS to the medial prefrontal cortex among AUD patients. Relative to sham, individuals who received real TMS were 3 times more likely to remain sober and experienced a reduction in average brain reactivity to alcohol cues. Despite these positive results, there was considerable variability in that approximately 1/3 of individuals who received real TMS relapsed to alcohol and reductions to striatal reactivity to alcohol cues were not uniform. My NIAAA-sponsored F31 demonstrated that individual patterns of brain response to alcohol cues are highly variable among AUD patients and often occur outside of the prefrontal cortex. Given this spatial variability, delivering TMS to a fixed point on the scalp leads to a situation in which TMS electrical fields do not always overlap with an indvidual’s brain response to alcohol cues. In a proof-of-concept retrospective analysis, I found that patients who had overlap between alcohol cue-reactivity patterns and TMS electrical fields had the best clinical outcomes. This K99/R00 proposal seeks to build upon this analysis by developing prospective functional connectivitiy - guided TMS (fcg-TMS) clinical trials for AUD. First, using an existing dataset, we aim to characterize the spatial topography of TMS cortical targets as defined by peak functional connectivity with the striatum during alcohol cue presentation (Aim 1, K99 phase). Then, we aim to evaluate the efficacy of one session of functional- connectivity guided TMS (fcg-TMS) on reducing striatal response to alcohol cues, a key biomarker of treatment efficacy (Aim 2, K99 Phase). These K99 aims will be complemented by a training plan that includes 4 domains: clinical trial design, fcg-TMS, laboratory leadership, and complex modeling of relapse and brain networks in AUD. Following these foundational trainings and experiments, Aim 3 (R00 phase) of this proposal will evaluate the efficacy of 15 sessions of fcg-TMS clinical trial in reducing drinking, relapse rates, and striatal response to alcohol cues. Cutting edge tools will be used to analyze the data presented in this proposal, including prospective, computational electrical field modeling, advanced functional connectivity analysis, and multivariate statistics.

NIH Research Projects · FY 2025 · 2024-09

There is considerable societal need to better understand neurobiological mechanisms, psychosocial processes, treatments, preventions, and policy in child and maternal pain. This application seeks funding for a T90/R90 institutional postdoctoral training program in clinical pain research, with a focus on maternal and childhood pain, at Stanford University. Our proposal describes a collaborative, interdisciplinary postdoctoral training program to bolster the clinical pain research workforce as outlined through the NIH HEAL Initiative Partnership to Advance Interdisciplinary (PAIN) Training in Clinical Pain Research. We venture to join with other T90/R90 postdoctoral training programs in clinical pain research to foster a valuable cohort experience for trainees to collaborate across institutions. This T90/R90 will reside within the Department of Anesthesiology, Perioperative, and Pain Medicine and be in partnership with the Maternal and Child Health Research Institute, the Department of Pediatrics, Psychiatry and Behavioral Sciences, and Biomedical Data Science at Stanford University School of Medicine. This program will provide mentorship to trainees to launch and maintain productive careers in the clinical pain research workforce. We propose training of five fellows per year who will spend 2-3 years cumulative time in research. Research opportunities are offered by the NIH- funded faculty mentors with proven records of success in the training of postdoctoral fellows, with various research programs ranging from data science, translational, clinical, and health services research. Core faculty research leaders will help match trainees to mentors within the HEAL clinical pain research content areas of: (1) bioinformatics, (2) pain across the lifespan, specifically child and maternal pain, (3) nonpharmacological (behavioral) interventions for pain, (4) prevention of the transition from acute to chronic pain and (5) health and wellness in the field of pain. Program mentors will provide training in clinical pain research, neuroscience, biomedical data science, behavioral health, epidemiology, maternal health, and health in children and adolescents. The training will (1) integrate the biopsychosocial model of pain, (2) use a team science approach through partnerships with psychiatry, pediatrics and biomedical data science, (3) center on whole person health, (4) incorporate patient participatory research methods to ensure lived experiences of pain inform the clinical research outputs, (5) provide advanced statistics and research methods, (6) involve grantsmanship skills, and (7) prepare for applying for and securing academic positions in clinical pain research. Lastly, the program will coordinate monthly webinars related to HEAL priority areas an annual Maternal and Child Pain School open to all cohort participants, and a robust didactic program to provide career development skills with seminars in stated training areas. Trainee progress will be monitored by the Assessment and Evaluation Team. The long-term goal of this T90/R90 is to train the next generation of interdisciplinary scientific leaders in maternal and/or child pain.

NIH Research Projects · FY 2025 · 2024-09

Project Summary The idea of evolutionary constraint is central to understanding of how populations have and will adapt(ed) in response to heterogeneous selective pressures. Constraints therefore determine how organisms with relevance to human health, like pathogens and cancers, adapt to different drugs, hosts, or environments. Constraints can be formed by strict trade-offs, when mutations adaptive in one environment have antagonistically pleiotropic costs in alternate environments, or by more dynamic processes, when the ability to select for hypothetically possible costless generalism is limited by mutational accessibility, speed of selection, or ecological opportunity for selection. These overlapping processes can make evolutionary constraint difficult to measure and these different processes difficult to disentangle. Despite the centrality of constraint to evolutionary theory, these difficulties mean that the processes forming constraints at different levels remain poorly understood— leaving open questions about how they affect evolution. In this proposal, I leverage high throughput microbial experimental evolution lineage tracking methods that provide unprecedented power to measure the distributions of fitness effects for mutations in multiple environments. These methods allow us to generate quantitative insights into the dynamics of evolutionary constraint. Aim 1 will ask whether constraints in laboratory experimental evolution resemble natural variation by measuring the joint distribution of fitness effects (JDFE) for natural isolates of S. cerevisiae in two nutrient environments mimicking natural conditions (synthetic wine must and synthetic beer wort) and then experimentally evolving a barcoded laboratory strain in these and an alternating media environment to determine whether the laboratory evolved JDFE resembles the naturally evolved JDFE. Aim 2 will explore how evolutionary constraints change over time by leveraging a rebarcoding system to experimentally evolve S. cerevisiae in the same conditions as Aim 1 for 5 evolutionary `steps' (~400 generations) and measure the JDFE at each `step'. Most laboratory experimental evolution examines shorter time scales, but we know from long-term experiments and natural observations that evolutionary dynamics change over time as populations near fitness peaks and/or change their environment. Thus, Aim 2 will allow us to quantitatively measure how constraints change over time. Through this work, I will build a powerful system for conducting high-throughput, long term evolution in multiple environments. The proposed research will significantly expand my training in the areas of quantitative genetics, population genetics, and molecular biology while encouraging increasing independence. It will therefore compliment my PhD training in evolutionary ecology to allow me to build an integrative research program that links genetic, evolutionary, and ecological processes to understand how constraints determine eco- evolutionary dynamics. Additionally, I will participate in professional development and outreach activities.

NIH Research Projects · FY 2026 · 2024-09

PROJECT SUMMARY Glaucoma is the leading cause of irreversible blindness and along with other optic neuropathies is characterized by the loss of retinal ganglion cells (RGCs). Increased intraocular pressure (IOP) management is the current standard of care for glaucoma patients, but fails to stop the irreversible loss of RGCs and progressive visual dysfunction. Glaucomatous RGC death was recently found to be driven by reactive optic nerve head (ONH) astrocytes, suggesting targeting these (and other) glial populations in the retina may be a viable strategy to protect RGCs. We recently discovered nuclear and cytoplasmic pools of cAMP, dependent on expression of soluble adenylyl cyclase (sAC), and associated with the stress-induced cell cycle inhibitor p21Cip, differentially regulate reactive astrocyte proliferation, microglial/macrophage activation, and RGC survival after traumatic optic nerve injury. Here, using a newly developed and reversible model of glaucoma in mice, we will establish the molecular, cellular, and transcriptional mechanisms that confer specificity to neurotoxic and protective astrocyte reactivity regulated by compartmented cAMP, sAC, and p21Cip in ONH astrocytes and Muller glia. An exciting element of our proposal is the use of state-of-the-art single cell RNA sequencing and cut-and-tag assays to identify the transcriptional and (epi)genetic changes induced by compartmented cAMP manipulation, and link those changes to reactive phenotypes in astrocytes and downstream effects on RGC survival and microglial/macrophage infiltration. All of this will be accomplished using novel AAV viral vectors to specifically target ONH astrocytes and Muller glia. These experiments will lead to the discovery of new biological pathways that regulate glial reactivity in neurodegenerative disease, and serve in the development of gliotheraputics for the treatment of glaucoma and other optic neuropathies.

NIH Research Projects · FY 2025 · 2024-09

PROJECT SUMMARY Night or rotating schedules under shiftwork are unavoidable, especially in healthcare work. Shiftwork can disturb circadian rhythms due to unusual light exposure and has been linked to many long-term negative health outcomes such as cardiovascular and cardiometabolic disorders. Studying disturbances in circadian rhythms in shift workers has proven challenging due to limitations in measuring circadian rhythms in humans under real- world conditions. However, with recent developments in wearable sensors, we can now capture continuous, remote, and longitudinal measurements of physiological signals (e.g., heart rate, breathing rate, biomechanics), behavioral patterns (e.g., activity tracking), and other external stimuli (e.g., light exposure), enabling the characterization of circadian disturbances and supporting identifying actionable strategies for shift workers to regain and maintain internal synchrony. This proposal seeks to examine the impact of the coordination of central and peripheral body clocks internally and with the external environment in the real-world. We hypothesize that internal and external synchrony of the body’s clocks is critical for regaining stability after perturbations such as shiftwork. In this proposal, we seek to develop a quantitative framework for assessing body clock synchrony in the real-world using wearable sensors. In the long term, we seek to leverage this work to interrogate the impact of the coordination of body clocks on health state and develop timely interventions to regain stability. To do this, in Aim 1, I will improve circadian phase prediction algorithms using real-world light diets to enable the study of circadian rhythms outside of the laboratory and in real-world settings. In Aim 2, I will assess internal synchrony leveraging continuous, ambulatory monitoring of physiological signals and estimation of central clock phase to develop a quantitative framework for measuring the coordination of central and peripheral body clocks. This will enable deeper understanding of their coordination and contribution to recovery from disturbances. In Aim 3, I will develop predictive algorithms using machine learning to enable the evaluation of light diets to improve synchrony in shift workers to translate tools to empower them to maintain synchrony and reduce risk of long- term negative health outcomes. Completing these aims and working in collaboration with my sponsor, Dr. Jamie Zeitzer, and co-sponsor, Dr. Todd Coleman, will enable me to develop the skillset necessary to study disease dynamics in the real-world using wearable sensors and pursue a future career as a robust, independent, and interdisciplinary researcher. My sponsor and co-sponsor are well equipped to provide training in measuring complex human physiological signals, developing engineering skills in statistical signal processing and estimation theory, and communication and leadership skills to facilitate the translation of my work to a wider audience. The support of my mentors as well as the plentiful resources and collaborative environment of the Bioengineering PhD program and Stanford University are opportune for the successful completion of this work and complementary training goals.

NIH Research Projects · FY 2024 · 2024-09

PROJECT SUMMARY/ABSTRACT One of the fundamental challenges in modern biology is to decode the functionalities of human genome sequence. Over the past decade, genome-wide association studies (GWAS) have generated a wealth of new information, including the genotype–phenotype associations in various diseases and traits. Despite clear successes in identifying novel disease susceptibility genes and in translating these findings into clinical care, GWAS has been criticized for the fact that most association signals reflect variants and genes with no direct biological relevance to phenotype. The development of large language model (LLM) has been the main driving force behind many recent breakthroughs in artificial intelligence. Research into the “genomic LLM” therefore has the potential to significantly advance our understanding of how the genetics variants lead to the changes in phenotypes by disrupting the underlying regulatory syntax of DNA. The Research Training Plan will first develop and improve the core technologies of genomic LLMs to deepen our understanding on understanding the complex regulatory mechanisms in gene regulation (Aim 1). The developed genomic LLMs will then be applied in imaging genetics studies where imaging traits are used as phenotypes (Aim 2) and the development of new machine learning (ML) approaches for Alzheimer’s disease diagnosis (Aim 3). In Aim 1, the applicant Dr. Qiao Liu will develop new genomic LLM techniques and provide biological model interpretation with special focus on how transcription factor (TF) binds DNA recognition sites in genomic regulatory regions to control genomic transcription and affect epigenomic signals in a context-specific manner. The proposed genomic LLMs will serve as solid foundation for both Aim 2 and Aim 3. In Aim 2, Dr. Liu will focus on the imaging genetics studies, which can be considered as GWAS of imaging phenotypes, for linking genetic variants/genes to structural or functional imaging features through the mediation of genomic LLMs. Genomic LLMs thus will bridge the gap between personal genetics and radiomics. In Aim 3 during the R00 phase, Dr. Liu will develop new ML approaches on AD diagnosis by considering the causal genetic-imaging-clinical pathways and leveraging the power from the genomic LLM. To succeed in these aims, a Career Development Plan is tailored to enable Dr. Liu to gain new knowledge and skills in radiomics, neuroimaging, and Alzheimer’s disease, as well as career skills through practice and coursework with the support of the outstanding mentoring team and scientific advisory committee. Stanford University is an ideal environment, providing all of the facilities needed for the proposed research and a rich interdisciplinary environment for collaborative studies. In summary, the strong mentoring team and scientific advisory committee, as well as the training plan are anticipated to fully prepare Dr. Liu to launch his independent career. The proposed studies promise to offer mechanistic insights into both genetics and radiomics, and may help uncover important genetic-imaging-clinical pathways for better understanding complex diseases.

NSF Awards · FY 2024 · 2024-09

Tropical peatlands are believed to play an outsized role in the global carbon cycle. Despite their relatively small extent, these ecosystems act as some of the strongest long-term terrestrial carbon sinks on Earth. They are also believed to constitute some of the largest natural methane sources. However, these claims are anchored in very little field data. In South America, which may encompass the largest area of tropical peatlands in the world, we do not know where peatlands are, what controls the rate of peat formation, which conditions constrain methane emissions, and whether these ecosystems are resilient to climate change. This project focuses on the PanAm peatlands, which we define as the natural lowland peatlands found across the tropical region of the American continent, from 20 degrees N to 20 degrees S. The selected approximately 80 study sites encompass broad environmental gradients and include distinct climate areas such as the Caribbean coast and the Amazon basin. The main approach involves using a novel field sampling kit, which will allow for consistent field data collection at the continental scale. The ultimate goals of this research are to gain new insights into peatland environments, and better constrain regional and global carbon budgets. Training the next generation of scientists is also a central component of this project. In addition to involving U.S. postdoctoral researchers, graduate students and undergraduate students, in-person field training will be offered to local scientists (including students) through workshops that will be held in the Caribbean and the Amazon. High-quality videos showing how to deploy instruments and take measurements using our field kit will also be developed in English and Spanish, and made available to all. Data analysis training will be offered to peatland scientists and students through hybrid workshops. Overall, this work will contribute to U.S. expertise in Earth System Science, increase capacity through student training, advance the peatland, wetland, and carbon cycling communities’ research agenda, and guide policy and land management decisions. Because tropical peatlands these ecosystems are being lost at a rate about three times faster than forests, it is critical to collect baseline information about their structure and function. This research undertakes a large number of systematic field measurements that will improve mechanistic understanding of plant-soil-water-nutrient-carbon interactions. This information is needed to build holistic representations of the different types of tropical peatlands that exist. These field data and statistical models will allow for predictive assessments of tropical peatland carbon input and output, with an emphasis on methane emissions. Ultimately, the project seeks to (1) identify the hydrological thresholds needed for peatland formation across the tropics, (2) quantify the key constraints on the rate of peat formation over space and time, and (3) document patterns of methane production, consumption, and emission at the ecosystem scale. This field-based research may lead to the development of new theoretical foundations needed to improve multiple modeling efforts. Recurring engagement with modelers is planned to help integrate these field data into peatland mapping algorithms, process-based peatland ecosystem models, and land surface models. A database that combines the new results with a literature synthesis of existing and ongoing measurements will be generated and archived in the Environmental Data Initiative repository. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NIH Research Projects · FY 2025 · 2024-09

PROJECT SUMMARY/ABSTRACT There is considerable societal need to better understand neurobiological mechanisms, psychosocial processes, treatments, preventions, and policy in child and maternal pain. This application seeks funding for a T90/R90 institutional postdoctoral training program in clinical pain research, with a focus on maternal and childhood pain, at Stanford University. Our proposal describes a collaborative, interdisciplinary postdoctoral training program to bolster the clinical pain research workforce as outlined through the NIH HEAL Initiative Partnership to Advance Interdisciplinary (PAIN) Training in Clinical Pain Research. We venture to join with other T90/R90 postdoctoral training programs in clinical pain research to foster a valuable cohort experience for trainees to collaborate across institutions. This T90/R90 will reside within the Department of Anesthesiology, Perioperative, and Pain Medicine and be in partnership with the Maternal and Child Health Research Institute, the Department of Pediatrics, Psychiatry and Behavioral Sciences, and Biomedical Data Science at Stanford University School of Medicine. This program will provide mentorship to trainees to launch and maintain productive careers in the clinical pain research workforce. We propose training of five fellows per year who will spend 2-3 years cumulative time in research. Research opportunities are offered by the NIH- funded faculty mentors with proven records of success in the training of postdoctoral fellows, with various research programs ranging from data science, translational, clinical, and health services research. Core faculty research leaders will help match trainees to mentors within the HEAL clinical pain research content areas of: (1) bioinformatics, (2) pain across the lifespan, specifically child and maternal pain, (3) nonpharmacological (behavioral) interventions for pain, (4) prevention of the transition from acute to chronic pain and (5) health and wellness in the field of pain. Program mentors will provide training in clinical pain research, neuroscience, biomedical data science, behavioral health, epidemiology, maternal health, and health in children and adolescents. The training will (1) integrate the biopsychosocial model of pain, (2) use a team science approach through partnerships with psychiatry, pediatrics and biomedical data science, (3) center on whole person health, (4) incorporate patient participatory research methods to ensure lived experiences of pain inform the clinical research outputs, (5) provide advanced statistics and research methods, (6) involve grantsmanship skills, and (7) prepare for applying for and securing academic positions in clinical pain research. Lastly, the program will coordinate monthly webinars related to HEAL priority areas an annual Maternal and Child Pain School open to all cohort participants, and a robust didactic program to provide career development skills with seminars in stated training areas. Trainee progress will be monitored by the Assessment and Evaluation Team. The long-term goal of this T90/R90 is to train the next generation of interdisciplinary scientific leaders in maternal and/or child pain.

NIH Research Projects · FY 2026 · 2024-09

PROJECT SUMMARY/ABSTRACT Delirium exposes patients to a greater risk of developing Alzheimer’s disease and dementia, and patients with Alzheimer’s disease and dementia are more likely to develop delirium. Delirium among elderly patients is dangerous and common, yet it is underdiagnosed and undertreated. Especially common in patients with Alzheimer’s disease, delirium is a strong predictor of poor outcomes including post-operative cognitive decline (POCD) and mortality. The key to reducing the burden associated with delirium is early identification of those at high risk for it, especially prior to surgery, which is known to be a major risk factor for delirium. However, currently, there are no reliable tools available to predict the risk of post-operative delirium (POD). Also, there is no clear understanding of the pathophysiological mechanism of delirium. The long-term goal of this project is to investigate the role of epigenetics to better understand the molecular mechanism of pathogenesis of POD and to identify potential biomarkers associated with POD and POCD. We will test for a specific type of epigenetic modification, DNA methylation (DNAm), which is known to be induced by environmental factors. Previous searches for biomarkers of delirium in humans led to the proposal that inflammation and pro-inflammatory cytokines play a key role in the pathophysiology of delirium. Yet, how the aging process enhances cytokine release remains unclear. Aging is known to have a strong effect on gene expression, and such changes in gene expression are tightly controlled by DNAm. Thus, it is suspected that DNAm changes over the course of aging, leading to enhanced cytokine release due to altered control of gene expression, and that such DNAm modification contributes to delirium. This project will be the first to investigate the role of epigenetics associated with POD in humans, and the objective of the project is to identify potential biomarkers for elevated risk of POD and subsequent patient outcomes related to delirium such as POCD and dementia. The proposed research will accomplish its objective by following a large sample of elderly patients going through hip fracture surgery throughout their hospital stay, comparing differences in genome-wide DNAm between those who develop POD and those who do not. Lastly, the project will test if DNAm is predictive of patient outcomes including survival and long-term memory problem after surgery. This research will enhance our understanding of the epigenetic mechanisms in the pathogenesis of delirium and its interplay with the progression of dementia, and may identify biomarkers predictive of their risk. The identification of epigenetic markers of delirium would hopefully lead to improvement in medical and surgical practice.

NSF Awards · FY 2024 · 2024-09

For six decades, the semiconductor industry has relied on reducing the sizes of transistors, memory, and wires from microns to nanometers. The conventional approach of “cramming more components” onto the same chip area, leading to greater functionality and lower cost per function over time, is approaching its physical limits. However, a seismic shift in the future of semiconductors can be enabled by building three-dimensional (3D) integrated circuits (ICs) with much higher density than have been possible through conventional, lateral scaling. This project will develop the fundamental steps necessary to accomplish such a vision through an inter-disciplinary approach from nanomaterials to systems, in a manner that will accelerate their lab-to-fab translation. The project will pursue the integration of atomically thin materials (e.g. one- and two-dimensional semiconductors like carbon nanotubes and MoS2) into individual circuit layers, which will then be stacked in 3D to achieve high-density circuits. The research will also advance the materials science and reliability of 3D integration, areas of inquiry that have received limited attention to date. The 3D integrated circuits which this research could enable, are expected to be much more energy-efficient (over 50x) than today’s technology due to the tight integration of logic and memory, with potentially massive impact from low-power wearables to large energy-hungry data centers. This project will also educate the future US semiconductor workforce in a unique way with much broader exposure (from materials-to-systems) than a traditional education which “silos” the various components (e.g. materials vs. circuits). The team will build on a strong track record of mentoring students who have gone into the semiconductor industry, and their educational efforts will be scaled up through online learning. These efforts could have a large societal impact, as contributions to US semiconductors are seen as major growth opportunities for industry, as important for our quality of life, and as critical to our economy and national security. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NIH Research Projects · FY 2025 · 2024-09

PROJECT SUMMARY/ ABSTRACT The brain and spinal cord are filled with and surrounded by a complex fluid, the cerebrospinal fluid (CSF). CSF directly contacts brain progenitors to act as a stem cell niche that provides buoyancy, ionic and osmotic balance, and health- and growth- promoting factors. Pathological deviations in CSF volume and composition are associated with congenital, neuropsychiatric, infectious, and geriatric diseases, as well as injury. As the brain matures during development, CSF composition changes profoundly. We recently discovered that CSF ion concentrations also change dramatically across development, including a ~2.5-fold drop in CSF [K+] during the first postnatal week in rodents (from ~10 mM to ~3.2 mM). This large natural shift in CSF [K+] has the potential to affect key processes in brain development including progenitor maintenance, neurogenesis, and physiology. Our lab has the tools and expertise to directly control CSF [K+] and assess neurodevelopmental outcomes. Extracellular K+ is a fundamental signal for proliferation, survival, and cellular migration. K+ is also a key ion regulating cellular physiology, excitability, and ion co-transport. It is therefore crucial to understand how developmentally dynamic CSF ions contribute to brain generation and maturation. A major tissue source of CSF ions is the choroid plexus. We found that choroid plexus-restricted knockdown or overexpression of the sodium- potassium-chloride cotransporter NKCC1 (Slc12a2) delays or accelerates the drop in CSF [K+], respectively. It is now possible to directly test hypotheses that stage-specific CSF ions support neural progenitors and immature neurons to drive long-term brain function. Here, we propose to answer fundamental, yet transformative questions of whether CSF ions are necessary and sufficient to support brain development. Here we adapt explant manipulation and in vivo AAV gene delivery to investigate how the higher [K+] that we observe in early CSF specifically supports early neurodevelopment (aim 1); how the lower [K+] that we observed in postnatal CSF specifically supports neural maturation (aim 2); and test whether the shift in CSF [K+] alters the Cl- and K+ shunting that occurs as part of the developmental GABA switch (aim 3). This multi-tiered approach will yield widely applicable information and tools for testing hypotheses of CSF ion function over development, and in health and disease. Each component builds on my unique expertise to facilitate a new research program investigating how CSF supports the maturation of neurons and circuits underlying psychiatric disease. This innovative research program will fundamentally change our understanding of brain development and reveal roles for CSF ions in supporting brain generation and physiology. The CSF is an accessible avenue for CNS surveillance or supplementation, even in humans (e.g. intranasal spray, intrathecal injections). Therefore, outcomes will guide efforts to harness CSF to provide a supportive developmental environment for the brain, reduce neurologic symptoms, and may provide therapeutic strategies for tackling a range of disease.

NSF Awards · FY 2024 · 2024-09

Meteoroids are small rocky and icy bodies, ranging from dust particles to boulders in size, that are formed from the collision and interaction of asteroids and other larger objects throughout the solar system. Approximately 100,000 kg of extraterrestrial matter, primarily composed of meteoroids, enters the Earth's atmosphere every day. As meteoroids burn up in the atmosphere, they create a plasma, or ionized gas, known as a meteor or a shooting star. The PI plans to study meteors through analysis of meteor observations using radar and developing simulations to capture the physics of the meteoroid shedding material and how that material interacts with the atmosphere. Understanding the physics of the upper atmosphere where meteors are formed is crucial to NSFs mission of progressing science and securing national defense. This research will provide information that can help predict atmospheric effects on spacecraft operations, including potential collisions and failures, reduced satellite lifetime, or communication outages. The development of new simulation techniques has application in many other fields including spacecraft design and analysis, parachute dynamics, and hypersonic reentry vehicles. Involvement of two graduate students is planned in all aspects of the modeling and simulation. The objective of this proposal is to study the E region through the analysis of high-resolution data from High-Power Large-Aperture (HPLA) radars associated with the smallest, and most numerous, meteoroids. Meteor activity is the primary source of metals in the MLT region and gives origin to the upper atmospheric metallic and ion layers. It is anticipated that the increasing numbers of satellites will lead to a drastic increase in metallic mass deposition in the upper atmosphere as they de-orbit. The mass deposition rate, which depends on impactor properties and background conditions, though critical for modeling the upper atmospheric chemical processes remains poorly understood. The work will lead to the development of a combined Direct Simulation Monte Carlo – Finite Element Method (DSMC-FEM) model, the first of its kind, and modification of an existing Particle-In-Cell (PIC) algorithm to determine plasma formation and expansion. The researchers will then apply a Finite-Difference Time-Domain (FDTD) model to map radar signal strength to plasma density. This research, which will contribute to the National Space Weather Program’s goal of understanding the evolution of ionospheric irregularities, will answer the following scientific questions: (a) How do meteoroids deposit mass within the thermosphere and create localized regions of high-density extraterrestrial material? (b) What kinds of small-scale atmospheric neutral density structures exist at a single geographic location, and can we correlate neutral density to trail echo onset time? And (c) What are the long-term effects of increased material on the MLT region? This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NIH Research Projects · FY 2025 · 2024-09

PROJECT SUMMARY Aging is a complex process characterized by many hallmarks, such as DNA instability, epigenetic changes, and loss of protein homeostasis. Cellular rejuvenation, which aims to restore cells to a youthful state, offers hope to counteract aging and its associated diseases. Recent advances in epigenetic reprogramming using Yamanaka factors (a set of four transcription factors) have rejuvenated aged cells to youthful states and extended lifespan in mice. However, the clinical use of Yamanaka factors is limited due to their tumorigenic risk and full reprogramming potential. Therefore, it is important to find new rejuvenating transcription factors that are safer and more potent than the Yamanaka factors. The Li lab has recently developed a systematic approach and identified ~30 potential rejuvenation transcription factors. They have identified ~30 transcription factors that can restore youthful gene expression patterns in aged human fibroblasts in vitro. Critically, they also validated a few top hits with cellular and molecular phenotyping of aging hallmarks. However, the underlying mechanisms by which these transcription factors/chromatin modifiers rejuvenate aged cells, and the ability of these transcription factors to rejuvenate other types of aged cells (such as post-mitotic cells) are unknown. To better prepare me for such research, I propose to continue my training in cell/molecular biology by investigating how BRWD3 regulates DNA replication and epigenetic modifications (F99 Aims). This will not only enhance my comprehensive skill set for mechanistic studies, but also deepen my understanding and investigative strategies for chromatin-associated proteins, which are central to my proposed F00 aging research. During the K00 phase, I will leverage my molecular research expertise and the Li lab's system-level approaches to advance our understanding of cellular rejuvenation. I propose to dissect the mechanism of a top rejuvenation candidate and identify its key targets responsible for rejuvenation (K00 Aim 2.1). I will also use the induced neurons with the system-level approaches developed by the Li lab to explore the rejuvenation potential of the top 30 transcription factors in aged neurons (post-mitotic). Collectively, these experiments will not only deepen our understanding of the rejuvenation mechanism, but also shed light on rejuvenation approaches for overlooked post-mitotic cells, leading to the discovery of safer and more universal rejuvenation solutions for both mitotic and post-mitotic cells.

NIH Research Projects · FY 2025 · 2024-09

PROJECT SUMMARY Our Science Technology and Research at Stanford (STARS) program aims to equip high school students with the academic knowledge, creative problem-solving, and experiential skills necessary to succeed in high-demand, high-skill careers in the oral healthcare space. We will achieve this objective through two avenues: 1) career awareness activities coupled with school-year apprenticeships that provide students with meaningful exposure to the real-world applications of dental research; and 2) through paid, 8-week summer research internships in which participants will gain hands-on technical training and mentor-guided academic knowledge in oral healthcare. Integral to both components of STARS is a focus on communication, teamwork, problem-solving, critical thinking and adaptability abilities. After completing STARS, our students will graduate with a blend of technical skills, academic grounding, and professional behaviors enabling them to ultimately compete for high-skill careers in Dentistry and beyond. STARS is a partnership with Mountain View Los Altos (MVLA) school district. In collaboration with MVLA teachers in their Career Technical Education (CTE) and Advancement via Individual Determination (AVID) programs, 11 Stanford mentors will provide school year activities that help students discover career options related to in-demand healthcare professions. From this cohort of students, we will select participants for the summer research component of STARS. Selection will be based on a combination of interest, potential, and preparation. The 8-week summer internship is modeled after our highly successful 22-year-old summer program. The goal is to combine biomedical training and academic knowledge, which together create a well-rounded educational experience. Mentors hold DDS, PhD, combined DDS/PhD degrees, and certificates in dental hygiene: Prof. Jill Helms DDS/PhD (periodontist) leads a NIDCR-funded research lab, has taught undergraduates for 25 years, and has served for two decades as the Director of a hands-on summer internship for high school and community college students. Dr. Dyani Gaudilliere DDS, MS, MPH is the Chief of Stanford’s Dental Medicine & Surgery and brings clinical expertise while Bo Liu, DDS/PhD brings state-of-the-art research expertise to explore questions of clinical relevance in craniofacial biology and dental regenerative medicine. Fabiana Aellos DDS, MS (periodontist) brings teaching, mentoring, and clinical skills-building to the group. In addition to hands-on laboratory skills that advance NIDCR- funded research initiatives, students participate in community-building social activities created in partnership with local dental schools. An end-of-summer research symposium provides an opportunity to hone participants’ communication skills and offer networking opportunities with community healthcare providers. The STARS program is defined by a rigorous evaluation plan that regularly measures the impact of each programmatic component, through both pre- and post-surveys and outcomes tracking e.g., college enrollment rates and career decision-making by participants. Together, these empirical data enable rapid evolution of STARS to better serve participants, and to ensure that we are responsible stewards of NIDCR resources.

NIH Research Projects · FY 2025 · 2024-09

Project Summary/Abstract To treat wounds that won’t heal for patients with Recessive Dystrophic Epidermolysis Bullosa, we have developed induced pluripotent cell (iPS)-cell derived cGMP compatible, scaleable induced skin composite (iSC) differentiation system that requires multi-lineage interaction to generate graftable surface ectoderm (SE), mesoderm/dermis and melanocytes. Work from ARO73170, now in its 5th year, endeavors to fill gaps in knowledge about the chromatin dynamics and interactions between the lineages to enhance manufacturing efficacy and reduce line-to-line variability. ARO73170 identified transcription factors (TFs) TFAP2 and GRHL2 that pattern early SE differentiation. In addition GATA3 and the product of the Xia-Gibbs Syndrome locus Gibbin induce Gibbin-dependent mesoderm (GDM) that matures the SE stratification program through cross-regulatory signaling. For SE differentiation, GRHL2 activity is required for TFAP2A chromatin binding and in turn, TFAP2A restricts inappropriate GRHL2 binding to ectodermal disease-associated loci, but the mechanism of interaction, and how network signaling modifies its output, remains a gap in knowledge. For GDM differentiation, we found that wild type GDM can rescue mutant ectoderm, reinforcing the need to identity the cross-regulatory stratification factors that enhance the graftability of manufactured tissue. While GATA3 binding sites are known, a gap in knowledge exists how Gibbin accumulates on key developmentally regulated promoters to activate them. Overall our preliminary data supports the intriguing hypothesis that TFAP2-GRHL2 ectoderm signaling, with reciprocal GATA3-Gibbin-GDM induction and signaling, allows proper iPS- derived graftable tissue manufacturing. We will focus on key gaps in knowledge as we dissect spatial and temporal TFAP2/GRHL2-mediated surface ectodermal commitment by dissecting the ectoderm- mesoderm lineage commitment trajectory, refining spatial constraints to optimize surface ectoderm/mesoderm commitment, interrogating GRHL2-TFAP2 mutual regulatory interactions, and modulating the GRHL2-TFAP2 target gene network to improve ectoderm commitment; and by elucidating GDM functions during skin differentiation through elucidating how Gibbin localizes to promoters of key developmental regulators, dissecting Gibbin-Dependent mesoderm signaling during mouse ectodermal development, and validating GDM-dependent factors during iPS-derived skin manufacturing. Successful completion of this proposal will provide deep mechanistic insights into the chromatin dynamics of multi-lineage tissue differentiation, highlight candidate in-process biomarkers for clinical manufacturing of skin, and establish a flexible manufacturing platform for novel cell therapy for a previously untreatable diseases.

NIH Research Projects · FY 2025 · 2024-09

PROJECT SUMMARY Aging is the greatest risk factor for vascular dementia, the second most common cause of dementia after Alzheimer disease (AD). Vascular dementia comprises a large proportion of AD-related dementias, or ADRD. Vascular dementia is the result of poor vascular perfusion of the brain and can occur as a result of ischemic stroke, particularly in elderly individuals. The mechanisms underlying the greater risk of vascular dementia in elderly stroke patients are unclear thus limiting discovery of therapeutic approaches. Studies have shown a significant post-stroke inflammatory response in the aging ischemic brain that can persist and contribute to cognitive decline. However, the nature of this persistent immune response, particularly the adaptive T cell component, is not well characterized and the functions of brain-infiltrating T cells are not well established, especially in the context of aging. Therefore, this proposal will characterize late T cell responses to stroke in a mouse model of age-related post-stroke dementia. Aged stroke mice demonstrate a significant cognitive impairment that is accompanied by T cell infiltration in the post-stroke brain. As T cells undergo vast changes in immune repertoire and function with aging, it is likely that T cells that traffic to and reside in the post-stroke brain will have distinct functions that contribute to age-related differences in long-term stroke outcomes. This research proposal aims to (1) Use single cell T cell receptor and immune response gene sequencing to characterize infiltrating T cells in the aging ischemic hemisphere (Aim 1); (2) Utilize Seahorse bioenergetics, metabolomics, and T cell activation assays to identify age-related differences in T cell metabolism which fuel T cell activation and T cell function (Aim 2); (3) Test the extent to which excessive innate immune responses characteristic of aging stroke contribute to T cell-mediated cognitive decline, and determine the therapeutic potential of acutely inhibiting innate immunity to confer a more favorable adaptive immune response (Aim 3). Understanding how age-related changes in T cells contribute to chronic stroke outcomes will ultimately provide us with novel therapeutics for elderly patients of vascular dementia.

NIH Research Projects · FY 2025 · 2024-09

Training the next generation of translational scholars who can innovate, think creatively, and apply rigorous and responsible research principles to major problems is society's best chance to recoup its enormous investment in biomedical research. The Stanford CTSA's Mentored Research Career Development Program (K12) sits at the center of a dynamic Stanford and Silicon Valley ecosystem of biomedical research at all stages, from basic discovery to practical implementation. Our KL2 program has a 15-year history of successfully training Scholars to become exceptional scientists, leaders, and mentors. Our Scholars have over 2,749 publications (average 58 each) cited over 45,000 times since graduation, and all who have graduated since 2021 in the last two cycles have received subsequent independent research funding. That success has rested on a philosophy of practical, individualized training tailored to the Scholars and focused on the established characteristics of successful translational scientists. We will provide tailored mentoring and career development to four instructors and junior faculty from varied disciplinary backgrounds each year. Our alliance with the VA early career and Center for Digital Health training programs brings new colleagues into our scholarly community. Thematic emphases include leveraging developments in informatics to accelerate translational research, interdisciplinary teams, and enhancing the rigor and reproducibility of clinical research to ensure that translated products will truly benefit patients. The science of translation brings that rigor to the study of moving research across the translational spectrum and is embedded throughout our program. We have a strong recruitment plan and rigorous evaluation process to ensure that we attract the best Scholars. We structure our program around four specific aims: mentorship, education and professional development, evaluation, and impact. We have assembled a world-class faculty committed to training and mentoring our Scholars who together have a research program funded in excess of $350 million. The MPIs are both award-winning mentors with strong experience in leading large research projects and synergistic strengths. We have also assembled an outstanding team of mentors specific to community engagement and analytic methods, as well as a strong emphasis on peer mentoring. The education and professional development aim covers not just the domains of translational research and the science of translation, but lectures and experiences designed to imbue Scholars with the practical and leadership skills they will need to succeed. Our talented External Advisory Board of nationally renowned educators and scientists will, together with the Community Advisory Board, give the program strategic guidance. A robust evaluation and a long-term outcomes tracking plan ensures that we will continuously improve this program and maximize impact.

NIH Research Projects · FY 2025 · 2024-09

Myocardial infarction (MI) is a leading cause of death in the United States, affecting over 800,000 people annually. Numerous protein therapies have been developed to treat MI, but effective delivery of therapeutics to the heart remains a formidable challenge. Systemic, intravenous (IV) delivery of therapeutics results in low tissue specificity and rapid loss of function, necessitating high dosing, repeated administration, and/or long treatment periods. Direct injection of therapeutics into the myocardium commonly results in rapid clearance due to the heart’s contractility. To be successful as therapies for MI, protein drugs need new delivery methods that allow localized delivery in a sustained, controlled manner with minimal cargo loss. Here, I propose the development of injectable liposome nanoparticle crosslinked (LINC) hydrogels designed for sustained protein therapeutic delivery in the myocardium. These hydrogels are formed by crosslinking hyaluronic acid (HA) with functionalized liposomes, forming HA-LINC hydrogels, through strong yet reversible dynamic covalent chemistry (DCC) bonds. The Heilshorn Group has shown that HA-based hydrogels crosslinked through DCCs are injectable, retained in the myocardium, and cyto-compatible. To evaluate their performance in a preclinical setting, I will use HA-LINC hydrogels to deliver the promising MI protein therapy neuregulin-1β (NRG1) in a rat model of MI. In Aim 1, I will synthesize a library of distinct HA-LINC hydrogels by tuning liposome functionalization, HA functionalization, and HA concentration. The resulting gels will be analyzed for viscoelasticity, in vitro hand injectability using a syringe pump, and toxicity when cultured with primary cardiomyocytes. In Aim 2, I will systematically tune the degree of liposome internal stabilization and evaluate the effects on liposome structure, hydrogel viscoelasticity, and cargo release rates. To determine the bioactivity of released NRG1, it will be delivered from HA-LINC hydrogels to primary cardiomyocytes. Cardiomyocytes will be examined for viability, proliferation, and morphology in a hypoxia challenge representing MI. Additionally, I will evaluate the ability of released NRG1 to rescue the phenotype of hydrogen peroxide-treated cardiac fibroblasts. In Aim 3, the HA-LINC formulation with the highest stiffness and lowest required injection force (Aim 1) and most sustained NRG1 release profile (Aim 2) will be evaluated in vivo. Following induction of MI through ligation of the left anterior descending (LAD) artery, HA- LINC hydrogels encapsulating NRG1 will be injected. NRG1 plasma concentration over time will be used to create a pharmacokinetic model of release. I will evaluate effects on cardiac function, tissue remodeling, gel retention, cardiomyocyte survival, and angiogenesis. The proposed HA-LINC hydrogels will provide localized, long-lasting, controllable protein delivery to treat MI. This work will be completed in the Heilshorn lab at Stanford University in collaboration with Profs. Wu and Appel, experts in cardiology and pharmacokinetics. I will be directly mentored by Prof. Heilshorn and my collaborators, take courses on drug delivery, cardiac regenerative medicine, and bioethics, and continue mentoring undergraduates in the lab.

NIH Research Projects · FY 2025 · 2024-09

PROJECT SUMMARY Diploid organisms, including humans, create haploid gametes for sexual reproduction through the specialized cell division program of meiosis. For meiosis to occur successfully, Crossover (CO) repair of DNA double- strand breaks (DSBs) is essential, as it creates temporary connections between homologous chromosome pairs that promote orderly homolog segregation at the first division. Failure to form COs results in aneuploidy, the leading cause of miscarriage and birth defects in humans. Yet, CO formation relies on the creation of DSBs, which pose a significant danger to genomic integrity. Meiotic cells must therefore balance the risks imposed by DSBs with the requirement for COs to promote orderly chromosome segregation. Further, CO formation is highly limited and constrained: In many organisms, the majority of DSBs are repaired as non-COs (which restore genomic integrity but do not connect homologous chromosomes), whereas CO recombination occurs at only a single DSB site per chromosome (or per chromosome arm). This restriction of CO formation necessitates that each site selected to become a CO must reliably mature as a CO. The paradox of requiring, yet limiting, CO recombination leads to several major outstanding questions: How are a subset of DSBs selected to become COs? What are the mechanisms that operate to ensure and limit CO formation? How is robustness conferred to the process of CO maturation? The proposed research will investigate the mechanisms that promote and ensure reliable formation of meiotic COs, exploiting genetic and cytological features of the C. elegans experimental system that make it especially amenable to addressing these issues. The strategy will focus on COSA-2 (CO-Site Associated), a newly-discovered intrinsically-disordered protein that I identified as a critical component of the CO machinery. COSA-2 exhibits striking localization to CO- designated recombination sites and is required for the retention/accumulation of additional CO-promoting factors at these sites. My preliminary data have led to the hypothesis that COSA-2 promotes CO formation by acting as a hub protein to concentrate CO-promoting factors, stabilize CO-site architecture, and promote formation of a protected spatial compartment to ensure CO-specific repair of DSBs. I will test this hypothesis by: (1) using innovative cytological methodologies and super-resolution microscopy to probe interrelationships between COSA-2 and known CO factors, CO-site architecture, and meiosis-specific chromosome structures; (2) identifying COSA-2 protein partners and features of the COSA-2 protein that contribute to its CO-promoting activity, and (3) using temporally-controlled protein degradation and ectopic recruitment assays to test the mechanistic role of COSA-2 in CO maturation at natural CO sites and to evaluate whether COSA-2 can drive formation of CO-site-like compartments and/or stimulate CO-specific repair. Experiments in this proposal will illuminate fundamental mechanisms that promote reliable formation of the COs needed to direct orderly chromosome segregation and ensure the faithful inheritance of genomes.

NIH Research Projects · FY 2025 · 2024-08

Pulmonary arterial hypertension (PAH) is a life-threatening disease characterized by abnormally elevated pulmonary pressures and right heart failure resulting in high morbidity and mortality. The pathologic hallmark of PAH is progressive loss and obstructive remodeling of lung microvessels unresponsive to available therapies. Studies by our group and others have shown that pulmonary microvascular endothelial cells (PMVECs) derived from PAH patients are highly susceptible to apoptosis and have a lower capacity to form blood vessels (i.e., angiogenesis). Fatty acid oxidation (FAO) is an essential energy source for PMVECs that requires metabolic coupling of mitochondria and endoplasmic reticulum (ER). Metabolic reprogramming characterized by high glycolysis, reduced FAO, and mitochondrial/ER dysfunction is a key pathological feature of PAH PMVECs linked to oxidative stress, endothelial dysfunction, and reduced angiogenesis. Our group has shown that reduced activity of bone morphogenetic protein receptor 2 (BMPR2), the most common genetic cause of hereditary and sporadic PAH, promotes metabolic reprogramming but, given the low penetrance of BMPR2 mutations, alterations in other genes (i.e., “second hit”) are likely necessary for PAH development. In this proposal, we will show that carboxylesterase 1 (CES1), a lipolytic enzyme responsible for releasing free fatty acids from the ER to the mitochondria, is required for FAO and metabolic homeostasis in PMVECs. Our preliminary studies show that CES1 knockdown in healthy PMVECs results in 1) high glycolysis, 2) reduced FAO, 3) mitochondrial/ER dysfunction, and 4) oxidative stress. Furthermore, loss of CES1 exacerbates metabolic reprogramming associated with BMPR2 insufficiency, and restoring CES1 expression improves PAH PMVEC functional status. Based on our preliminary studies, we hypothesize that loss of CES1 in PAH leads to endothelial dysfunction in PAH through metabolic reprogramming, lipotoxicity, and oxidative stress. To test this, we propose the following aims: (1): Determine the mechanisms by which loss of CES1 results in metabolic reprogramming and lipotoxicity in PMVECs, (2) Determine whether loss of CES1 promotes the development and severity of pulmonary hypertension and vasculopathy in mice, and (3) Determine the contribution of BMPR2 insufficiency and epigenetic repression to reduced CES1 expression in PAH PMVECs. Using the proposed approach, we will demonstrate that CES1 is essential for properly maintaining and repairing the pulmonary endothelium and acts as a key modifier of BMPR2 signaling. Given the limited capacity of current therapies to reverse endothelial dysfunction and prevent small vessel loss, therapeutic interventions that can restore CES1 expression could serve as a novel treatment approach for PAH.

NIH Research Projects · FY 2025 · 2024-08

Over 95 percent of maternal and child deaths globally occur in low- and middle-income countries (LMICs) where reliable death registries are mostly unavailable and other dependable data are scarce. Moreover, within LMICs, the most disadvantaged and remote communities tend to have the highest mortality rates but the least reliable data. Knowledge on the state of maternal and child health (MCH) in LMICs relies mostly on household surveys. These expensive and time-consuming surveys cover merely a small minority (usually <2%) of all communities in a country and are (at best) carried out only every couple of years. We propose a new approach to measuring MCH indicators that would provide up-to-date estimates at a very high geographic resolution and at little to no cost. Specifically, we hypothesize that machine learning in geotagged “big data” sources, with a key source being satellite imagery, can accurately estimate critical MCH indicators for each village and neighborhood in a country. Satellite imagery is a key data source in this project because it is updated frequently (at least monthly), covers all areas of a country, and is available free of charge. We will pursue three specific aims: 1) determining whether machine learning in satellite imagery and other publicly available geotagged data can accurately estimate key indicators of MCH status in a village or neighborhood at a snapshot in time and longitudinally over time; 2) determining whether this approach can also accurately estimate coverage with critical MCH services at a snapshot in time and over time; and 3) achieving sufficient interpretability of our machine learning models to inform on data needed to improve predictions, which interventions to target where, and generalizability. The approach to achieving our aims is to use household surveys that are geocoded at the village and neighborhood level as the “ground truth” against which we will train machine learning models in satellite images and publicly available geotagged data. Our extensive preliminary data, along with high-impact publications (e.g., in Science and Nature) by our team on using satellite images to predict important determinants of MCH in LMICs (such as community-level poverty, surface water quality, water and sanitation infrastructure, crop yields, travel time to the nearest healthcare facility, and air pollution), demonstrate that this approach is feasible. Crucially, our predictions will improve over time as the size and quality (e.g., the resolution of satellite imagery) of our data continue to increase. In addition to a unique dataset as well as open-access code for our machine learning models, this project will provide maps with an extremely high geographic resolution of key MCH indicators to inform policymakers and researchers on the current state of MCH. This project is significant because it can inform the geographically precise planning and targeting of MCH interventions, may enable the evaluation of past interventions and policies, and can serve as a blueprint for extending a “precision public health” approach to health outcomes beyond MCH.

NIH Research Projects · FY 2025 · 2024-08

PROJECT SUMMARY Congenital heart disease occurs in about 1% of births, making it the most common type of birth defect and leading cause of infant mortality in the United States. Congenital aortic valve disease, one category of congenital heart disease, frequently requires surgery. Despite generally positive results, undesirable outcomes remain common, and measures of success are typically based on an empirical, retrospective “guess and check” approach. Due to heterogeneous disease, clinical trials are difficult to conduct. Two promising procedures are bicuspidization repair, in which the surgeon constructs a two leaflet valve, and bioprinted valve replacement, a new type of highly experimental prosthetic valve that can potentially grow with the patient. The optimal valve geometry for both pro- cedures remains under debate or unknown. Thus, there is an unmet clinical need for engineering design tools to optimize postoperative valve geometry and improve outcomes. Simulations provide a controllable and efficient means to predict optimal surgical procedures, and after validation, translate guidelines to the clinic. Central to this work is a novel and robust modeling framework for simulating heart valves developed by the applicant, Dr. Alexander D. Kaiser. This research will leverage his modeling methods combined with a multimodal approach including in vitro and human studies to efficiently translate scientific knowledge to the clinic. Dr. Kaiser pro- poses to (1) optimize the postoperative leaflet geometry of bicuspidization repair and bioprinted replacement with simulation-guided design, (2) confirm and validate optimal valve performance in vitro and (3) translate guidelines to the clinic to advise bicuspidization repairs. Dr. Kaiser has extensive previous training in applied mathematics, numerical methods and computational modeling of heart valves. His career development plan includes training in congenital heart disease pathophysiology and methods of surgical treatment, in vitro experimental methods and medical imaging. The Department of Cardiothoracic Surgery at Stanford University will provide an outstanding interdisciplinary environment to enable Dr. Kaiser’s transition from primarily researching applied mathematics to being an interdisciplinary, medical investigator. Mentor Michael Ma is a leading, innovative pediatric cardiac surgeon eager to incorporate new scientific information into his surgical practice. Co-mentor Alison Marsden is a renowned expert in computational modeling of the cardiovascular system. Complementary expertise will be offered by advisors Drs. Ennis (MRI, in vitro testing), Skylar-Scott (bioprinting), Feinstein (pediatric cardiology) and Woo (cardiac surgeon). Dr. Kaiser will receive extensive mentoring, guidance and resources to transition to independence. To conclude, the mentoring, training, research experience and clinical research environment will prepare Dr. Kaiser to become an independent, interdisciplinary medical investigator. Ultimately, the proposed research will benefit patients with congenital heart disease who will have better operative outcomes via improved heart valve function, and thus lead longer, healthier lives.