Biology subjects

Dong, E.

Publications and source records attributed to Dong, E..

9 recordsLinked to original sources

bioRxiv2026

Prenatal Stress Differentially Shapes Adult Behavior in Male and Female Offspring

BackgroundPrenatal stress (PNS) is a well-established risk factor for neuropsychiatric vulnerability, yet its sex-specific behavioral consequences remain incompletely defined. Because males and females follow distinct neurodevelopmental trajectories, clarifying how early-life stress differentially shapes behavior is essential for developing targeted interventions. However, few preclinical studies directly compare male and female offspring within the same experimental framework, limiting the ability to identify true sex-dependent effects. MethodsUsing a validated mouse model of gestational restraint stress, we conducted a comprehensive, within-study assessment of sex-dependent behavioral outcomes in adult offspring. Behavioral domains included locomotor activity, anxiety-like behavior, sociability, fear learning and extinction, recognition memory, and alcohol-related responses (ethanol preference and behavioral sensitivity), all measured using identical paradigms across sexes. ResultsPNS broadly disrupted behavior and cognition in both sexes, increasing locomotor activity and anxiety-like behavior, impairing fear extinction and recognition memory, and altering behavioral sensitivity to ethanols sedative effects. Direct comparison revealed distinct sex-dependent vulnerabilities: males showed reduced social interaction, whereas females exhibited numerically greater impairment in fear extinction and a significantly stronger ethanol preference. Baseline fear responses, total fluid intake, and sucrose consumption were unaffected. ConclusionPrenatal stress programs neurobehavioral trajectories in a sex-dependent manner, conferring vulnerability to anxiety-related behavior, cognitive disruption, and alcohol use. By directly comparing males and females within the same experimental design, this study provides one of the most integrated evaluations of sex-specific PNS outcomes to date and offers a robust framework for investigating the biological mechanisms underlying divergent pathways to stress-related psychopathology.

animal behavior and cognition↗

bioRxiv2024

A Drug-Elicitable Alternative-Splicing Module (DreAM) for Tunable AAV Expression and Controlled Myocardial Regeneration

Adeno-associated virus (AAV) is a major vector for gene therapy. A technique to fine-tune the time and level of AAV expression is lacking, which greatly restricts indication choice, therapeutic efficacy and safety. To solve this problem, here we developed a drug-elicitable alternative-splicing module (DreAM) responsive to risdiplam, an FDA-approved alternative splicing modulator. Risdiplam activates DreAM-regulated AAV expression in a dose-dependent manner with an over 2000-fold inducible change depending on the inducer dosage and the organ of interests. With a temporally resolution of a couple of days, DreAM could repeatedly activate AAV expression, determined by the frequency and duration of risdiplam administration. As an example of DreAM-realized gene therapy, this technique was used to control AAV-based cardiac delivery of YAP5SA, a potent cardiomyocyte regeneration factor. DreAM transiently activated AAV-YAP5SA, establishing the dedifferentiation-proliferation-redifferentiation cycle in cardiomyocytes. Consequently, DreAM fulfilled the regenerative capacity of AAV-YAP5SA in myocardial infarction while circumventing toxicity associated with prolonged AAV-YAP5SA expression. Together, these data demonstrated a tremendous potential of DreAM to enhance the efficacy, safety and applicable scope of gene therapy.

bioengineering↗

bioRxiv2024

Vasoactive Intestinal Peptide Amphiphile Micelle Material Properties Influence Their Cell Association and Internalization

Vasoactive intestinal peptide (VIP) is a promising anti-inflammatory peptide therapeutic that is known to induce biological effects by interacting with its cognate receptor (i.e., VPAC) on the surface of antigen presenting cells (APCs). For VIP-based drug delivery technologies like nano- and microparticles, little is known about the effect VPAC targeting has on APC behavior. This is further influenced by the fact that particulate material properties including chemistry, shape, and size are all known to influence APC behavior. In this study, peptide amphiphile micelles (PAMs) were employed as a modifiable platform to study the impact VPAC targeting and physical particle properties have on their association with macrophages. VIP amphiphile micelles (VIPAMs) and their scrambled peptide amphiphile micelle analogs (SVIPAMs) were fabricated from various chemistries yielding particle batches that were comprised of spheres (10 - 20 nm in diameter) and/or cylinders of varying lengths (i.e., 20 - 9000 nm). Micelle surface attachment to and internalization by macrophages were observed using confocal microscopy and their association was characterized by flow cytometry. The enclosed work provides strong evidence that macrophages rapidly bind VPAC specific micelles independent of physical properties though micelle shape and size as well as receptor-specificity all influence their long-term macrophage association. Specifically, a mixture of spherical and short cylindrical VIPAMs were able to achieve the greatest cell association which may correlate to their capacity to fully bind the VPAC receptors available on the surface of macrophages. These results provide the foundation of how nano- and microparticle physical properties and targeting capacity synergistically influence their capacity to associate with APCs.

bioengineering↗

bioRxiv2024

Myocardial infarction creates a critical time window for AAV gene therapy

Recombinant adeno-associated virus (AAV) is a major gene delivery vector for cardiac gene therapy. The factors that influence AAV-based cardiac gene transfer remain incompletely understood. This study showed that myocardial infarction (MI) enhanced cardiac AAV transduction and gene expression in mice after systemic administration, peaking at the third day post MI. These additional AAV vectors enriched at the infarcted region, correlated with the pathological permeabilization of the coronary vessels. The outcome of AAV-base gene therapy for MI, via Camk2d base editing, was significantly improved when AAV was injected at the third day post MI. Together, our findings uncovered a critical therapeutic time window after MI that facilitated AAV-based cardiac gene transfer, which could be harnessed to boost both basic and translational cardiology. MethodsAdult C57BL/6 mice were subjected to left anterior descending (LAD) coronary artery ligation. Permanent LAD ligation created the MI model while 30min ligation followed by reperfusion established the ischemia/reperfusion (I/R) model. 2x1011 AAV vectors were injected into the mice via tail vein. The AAV vectors carried transgenes that were activated by the cardiomyocyte-specific Tnnt2 promoter or the constitutively active CMV promoter. AAV expressed luciferase with the hemagglutinin (HA) tag. The amount of AAV vectors were quantified by real-time quantitative PCR (RT-qPCR) analysis of genomic DNA. Transgene expression was measured by RT-qPCR at the mRNA level or HA tag immunofluorescence imaging and western blot at the protein level. Luciferase activity was measured via bioluminescence imaging.

physiology↗

bioRxiv2023

In vivo proximity proteomics uncovers palmdelphin (PALMD) as a Z-line-associated mitigator of isoproterenol-induced cardiac injury

Z-lines are core ultrastructural organizers of cardiomyocytes that modulate many facets of cardiac pathogenesis. Yet a comprehensive proteomic atlas of Z-line-associated components remain incomplete. Here, we established an adeno-associated virus (AAV)-delivered, cardiomyocyte-specific, proximity-labeling approach to characterize the Z-line proteome in vivo. We found palmdelphin (PALMD) as a novel Z-line-associated protein in both adult murine cardiomyocytes and human pluripotent stem cell-derived cardiomyocytes. Germline and cardiomyocyte-specific palmd knockout mice were grossly normal at baseline but exhibited compromised cardiac hypertrophy and aggravated cardiac injury upon long-term isoproterenol treatment. By contrast, cardiomyocyte-specific PALMD overexpression was sufficient to mitigate isoproterenol-induced cardiac injury. PALMD ablation perturbed transverse tubules (T-tubules) and their association with sarcoplasmic reticulum, which formed the Z-line-associated junctional membrane complex (JMC) essential for calcium handling and cardiac function. These phenotypes were associated with disrupted localization of T-tubule markers caveolin-3 (CAV3) and junctophilin-2 (JPH2) and the reduction of nexilin (NEXN) protein, a crucial Z-line-associated protein that is essential for both Z-line and JMC structures and functions. PALMD was found to interact with NEXN and enhance its protein stability while the Nexn mRNA level was not affected. Together, this study discovered PALMD as a potential target for myocardial protection and highlighted in vivo proximity proteomics as a powerful approach to nominate novel players regulating cardiac pathogenesis. HighlightsO_LIIn vivo proximity proteomics uncover novel Z-line components that are undetected in in vitro proximity proteomics in cardiomyocytes. C_LIO_LIPALMD is a novel Z-line-associated protein that is dispensable for baseline cardiomyocyte function in vivo. C_LIO_LIPALMD mitigates cardiac dysfunction and myocardial injury after repeated isoproterenol insults. C_LIO_LIPALMD stabilizes NEXN, an essential Z-line-associated regulator of the junctional membrane complex and cardiac systolic function. C_LI

molecular biology↗

bioRxiv2023

Non-cell autonomous cardiomyocyte regulation complicates gene supplementation therapy for LMNA cardiomyopathy

AimsRecombinant adeno-associated viruses (rAAVs) are federally approved gene delivery vectors for in vivo gene supplementation therapy. Loss-of-function truncating variants of LMNA, the coding gene for Lamin-A/C, are one of the primary causes of inherited dilate cardiomyopathy (DCM). Here we aim to study whether AAV-based LMNA supplementation could treat LMNA deficiency-triggered cardiac defects. Methods and ResultsWe compared whole-body, cardiomyocyte-specific and genetic-mosaic mouse models that carry Lmna truncating variants at the same genetic loci and uncovered primarily a non-cell autonomous impact of Lmna on cardiomyocyte maturation. Whole-body lamin-A supplementation by rAAVs moderately rescued the cardiac defects in Lmna germline mutants. By contrast, cardiomyocyte-specific lamin-A addback failed to restore the cardiomyocyte growth defects. A Cre-loxP-based AAV vector that expresses lamin-A throughout the body but excluding the heart was able to restore cardiomyocyte growth in Lmna germline mutants. ConclusionsLmna regulates cardiomyocyte growth non-cell autonomously. Non-myocytes are the key cell targets for a successful gene therapy for LMNA-associated cardiac defects. Translational perspectiveLMNA truncating mutations are among the major causes of inherited DCM. AAV gene supplementation therapy is emerging as a promising strategy to treat genetic cardiomyopathy, but whether this strategy is suitable for LMNA cardiomyopathy remained unclear. Our study counterintuitively showed that the cardiomyocytes are not necessarily the correct therapeutic cell targets for AAV-based treatment of LMNA cardiomyopathy. By contrast, careful elucidation of cell-autonomous versus non-cell-autonomous gene functions is essential for the proper design of a gene supplementation therapy for cardiomyopathy. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=189 SRC="FIGDIR/small/549413v1_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@37cfdaorg.highwire.dtl.DTLVardef@2f2cd1org.highwire.dtl.DTLVardef@1278d08org.highwire.dtl.DTLVardef@19c4869_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2022

Efficient Human Germ Cell Specification from Stem Cells via Combinatorial Expression of Transcription Factors

The generation of germline cell types from human induced pluripotent stem cells (hiPSCs) represents a key milestone toward in vitro gametogenesis, which has the potential to transform reproductive modeling and medicine. Methods to recapitulate advanced germline cell specification in vitro have relied on extensive, long term culture methods, the most notable of which is a four-month culture protocol employing xenogeneic reconstituted ovaries with mouse embryonic ovarian somatic cells. Recently, transcription factor (TF)-based methods have demonstrated the feasibility of exogenous factor expression to directly differentiate hiPSCs into cell types of interest, including various ovarian cell types. The protocols leveraged in these studies, however, utilize more local methods of factor selection, such as basic differential gene expression analysis, and lower-throughput screening strategies via iterative testing of a small set of TFs. In this work, we integrate our recently-described graph theory pipeline and highly-parallelized screening protocols to globally identify and screen 46 oogenesis-regulating TFs for their role in human germline formation. We identify ZNF281, LHX8, SOHLH1, ZGLP1, and ANHX whose combinatorial overexpression drives DDX4+ induced oogonia-like cell (iOLC) formation from hiPSCs. In contrast to previous methods, our protocol employs a simple four-day, feeder-free monolayer culture condition. We additionally demonstrate a method of post-isolation, feeder-free expansion of DDX4+ iOLCs that shows retained cell identity in vitro. We additionally identify DLX5, HHEX, and FIGLA whose individual overexpression enhances hPGCLC formation from hiPSCs. We characterize these TF-based iOLCs and hPGCLCs via gene and protein expression analyses and demonstrate their broad similarity to in vivo and in vitro-derived oogonia and primordial germ cells. Together, these results identify new regulatory factors that enhance in vitro human germ cell specification and further establish unique computational and experimental tools for human in vitro oogenesis research.

developmental biology↗

bioRxiv2022

Directed Differentiation of Human iPSCs to Functional Ovarian Granulosa-Like Cells via Transcription Factor Overexpression

An in vitro model of human ovarian follicles would greatly benefit the study of female reproduction. Ovarian development requires the combination of germ cells and their supporting somatic cells, known as granulosa cells. Whereas efficient protocols exist for generating human primordial germ cell-like cells (hPGCLCs) from human iPSCs, a method of generating granulosa cells has been elusive. Here we report that simultaneous overexpression of two transcription factors (TFs) can direct the differentiation of human iPSCs to granulosa-like cells. We elucidate the regulatory effects of several granulosa-related TFs, and establish that overexpression of NR5A1 and either RUNX1 or RUNX2 is necessary and sufficient to generate granulosa-like cells. Our granulosa-like cells form ovary-like organoids (ovaroids) when aggregated with hPGCLCs, and recapitulate key ovarian phenotypes including support of germ cell maturation, follicle formation, and steroidogenesis. This model system will provide unique opportunities for studying human ovarian biology, and may enable the development of therapies for female reproductive health.

developmental biology↗

bioRxiv2021

Deficits of Hippocampal RNA Editing and Social Interaction Resulting from Prenatal Stress are Mitigated by Clozapine.

BackgroundNeurodevelopmental deficits resulting from prenatal stress are associated with neurological disorders that include deficits of social behavior, such as schizophrenia1 and autism2-7. Studies of human brain and animal models indicate that an epitranscriptomic process known as RNA editing contributes to the pathophysiology of these disorders, which occur more frequently in males than in females8-20. RNA editing plays an important role in brain development through its modification of excitatory and inhibitory neurotransmission21. MethodsWe exposed pregnant mice to restraint stress three times daily during gestational weeks 2 and 3. We treated the adult male offspring with haloperidol (1mg/kg), clozapine (5mg/kg) or saline twice daily for 5 days. Subsequently we measured social interaction behavior (SI) and locomotor activity, followed by next-generation sequencing analyses of hippocampal RNA editing. ResultsMice exposed to PRS exhibited reduced SI, which correlated with hippocampal RNA editing of -amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor subunits GluA2, GluA3 and GluA4, the potassium channel Kv1.1, the calcium channel subunit Cav1.3, calcium-dependent secretion activator (CAPS-1) and the calcium-dependent cell adhesion protein, cadherin 22 (CDH22). Treatment with clozapine, but not haloperidol, normalized SI behavior, and selectively reduced the deficits in GluA2 RNA editing in PRS mice. ConclusionsRNA editing may contribute to impaired hippocampal function after exposure to PRS. The efficacy of clozapine in improving SI behavior may include indirect stimulation of GluA2 RNA editing in the hippocampus. Although these data are from male mice and not humans, the results suggest a new molecular pathway by which PRS leads to life-long impairments of hippocampal function.

neuroscience↗