Biology subjects

Lippmann, E. S.

Publications and source records attributed to Lippmann, E. S..

11 recordsLinked to original sources

bioRxiv2026

A Gata4-Loxl2 Axis Controls ECM Remodeling to Enable Scar-Free Spinal Cord Regeneration in Zebrafish

Spinal cord injury in mammals causes permanent loss of sensory and motor function due to scar formation obstructing axon regrowth. In contrast, zebrafish fully regenerate their spinal cord without lasting fibrosis. Here, we identify the transcription factor Gata4 as a key positive regulator of regeneration. After injury, gata4 expression is predominantly upregulated in ependymo-radial glial cells and in glial cells extending processes toward sites of tissue bridging. Gata4 preserves glial identity and orchestrates regenerative extracellular matrix (ECM) remodeling. Gata4 loss biases glial cells toward a fibroblast-like state with upregulated Loxl2b, a collagen cross-linking enzyme, and can also increase Loxl2b in nearby non-glial cells. Excess Loxl2b activity promotes pathological crosslinking and formation of a rigid, fibrotic matrix impairing glial and axonal bridging across the lesion, greatly reducing motor recovery. Pharmacological inhibition of Loxl2b in gata4 mutants substantially rescues ECM organization, tissue bridging, and functional recovery. Injury-induced gata4 expression depends, at least in part, on Hif1 signaling and activation of a short injury-responsive enhancer. We identify the Gata4-Loxl2b axis as a central regulator of and potential therapeutic target for scar-free spinal cord repair.

neuroscience↗

bioRxiv2025

The role of MICOS in modulating mitochondrial dynamics and structural changes in vulnerable regions of Alzheimer's Disease

Mitochondrial contact site and cristae organizing system (MICOS) complexes are critical for maintaining the mitochondrial architecture, cristae integrity, and organelle communication in neurons. MICOS disruption has been implicated in neurodegenerative disorders, including Alzheimers disease (AD), yet the spatiotemporal dynamics of MICOS-associated neuronal alterations during aging remain unclear. Using three-dimensional reconstructions of hypothalamic and cortical neurons, we observed age-dependent fragmentation of mitochondrial cristae, reduced intermitochondrial connectivity, and compartment-specific changes in mitochondrial size and morphology. Notably, these structural deficits were most pronounced in neurons vulnerable to AD-related pathology, suggesting a mechanistic link between MICOS disruption and the early mitochondrial dysfunction observed in patients with AD. Our findings indicate that the loss of MICOS integrity is a progressive feature of neuronal aging, contributing to impaired bioenergetics and reduced resilience to metabolic stress and potentially facilitating neurodegenerative processes. MICOS disruption reduced neuronal firing and synaptic responsiveness, with miclxin treatment decreasing mitochondrial connectivity and inducing cristae disorganization. These changes link MICOS structural deficits directly to impaired neuronal excitability, highlighting vulnerability to AD-related neurodegeneration. These results underscore the importance of MICOS as a critical determinant of neuronal mitochondrial health and as a potential target for interventions aimed at mitigating AD-related mitochondrial dysfunction.

neuroscience↗

bioRxiv2025

Amyloid-β-regulated gene circuits for programmable Alzheimer's disease therapy

Alzheimer's disease (AD) is a neurodegenerative disease characterized in part by the accumulation of the protein amyloid-{beta} (A{beta}). Monoclonal antibodies (mAbs) that target A{beta} for clearance from the brain have received FDA approval; however, these therapies are accompanied by serious side effects, and their cognitive benefit for patients remains of tremendous debate. Here, we present a potential engineered cell therapy for AD in which we enlist cells of the central nervous system as programmable agents for sculpting the neurodegenerative niche toward one that mitigates glial reactivity and neuronal loss. We constructed a suite of A{beta}-sensitive synthetic Notch (synNotch) receptors from clinically tested anti-A{beta} mAbs and show that cells expressing these receptors can recognize synthetic A{beta}42 and A{beta}40 with differential sensitivity. We express these receptors in astrocytes, cells native to the brain that are known to become dysfunctional in AD. These synNotch astrocytes, which upregulate selected transgenes upon exposure to synthetic and human brain-derived amyloid, were engineered to express potential therapeutic transgenes in response to A{beta}, including brain-derived neurotrophic factor and antagonists of the cytokines tumor necrosis factor and interleukin-1. SynNotch astrocytes that express such antagonists in response to A{beta} partially attenuate a cytokine-induced reactive astrocyte phenotype and promote barrier properties in brain microvascular endothelial cells. Additionally, engineered A{beta}-synNotch cells potently upregulate transgene expression in response to A{beta} deposited in the 5xFAD mouse brain, demonstrating the capacity to recognize A{beta} in situ. Overall, our work supports A{beta}-synNotch receptors as promising tools to generate a cell-based therapy for AD with targeted functionalities to positively influence the AD niche.

bioengineering↗

bioRxiv2023

Growth factor-free, peptide-functionalized gelatin hydrogel promotes arteriogenesis and attenuates tissue damage in a murine model of critical limb ischemia

Critical limb ischemia (CLI) occurs when blood flow is restricted through the arteries, resulting in ulcers, necrosis, and chronic wounds in the downstream extremities. The development of collateral arterioles (i.e. arteriogenesis), either by remodeling of pre-existing vascular networks or de novo growth of new vessels, can prevent or reverse ischemic damage, but it remains challenging to stimulate collateral arteriole development in a therapeutic context. Here, we show that a gelatin-based hydrogel, devoid of growth factors or encapsulated cells, promotes arteriogenesis and attenuates tissue damage in a murine CLI model. The gelatin hydrogel is functionalized with a peptide derived from the extracellular epitope of Type 1 cadherins. Mechanistically, these "GelCad" hydrogels promote arteriogenesis by recruiting smooth muscle cells to vessel structures in both ex vivo and in vivo assays. In a murine femoral artery ligation model of CLI, delivery of in situ crosslinking GelCad hydrogels was sufficient to restore limb perfusion and maintain tissue health for 14 days, whereas mice treated with gelatin hydrogels had extensive necrosis and autoamputated within 7 days. A small cohort of mice receiving the GelCad hydrogels were aged out to 5 months and exhibited no decline in tissue quality, indicating durability of the collateral arteriole networks. Overall, given the simplicity and off-the-shelf format of the GelCad hydrogel platform, we suggest it could have utility for CLI treatment and potentially other indications that would benefit from arteriole development.

bioengineering↗

bioRxiv2023

Amyloid-β exposed astrocytes induce iron transport from endothelial cells at the blood-brain barrier by altering the ratio of apo- and holo-transferrin

Excessive brain iron accumulation is observed in early in the onset of Alzheimers disease, notably prior to widespread proteinopathy. These findings suggest that increases in brain iron levels are due to a dysregulation of the iron transport mechanism at the blood-brain barrier. Astrocytes release signals (apo- and holo-transferrin) that communicate brain iron needs to endothelial cells in order to modulate iron transport. Here we use iPSC-derived astrocytes and endothelial cells to investigate how early-disease levels of amyloid-{beta} disrupt iron transport signals secreted by astrocytes to stimulate iron transport from endothelial cells. We demonstrate that conditioned media from astrocytes treated with amyloid-{beta} stimulates iron transport from endothelial cells and induces changes in iron transport pathway protein levels. The mechanism underlying this response begins with increased iron uptake and mitochondrial activity by the astrocytes which in turn increases levels of apo-transferrin in the amyloid-{beta} conditioned astrocyte media leading to increased iron transport from endothelial cells. These novel findings offer a potential explanation for the initiation of excessive iron accumulation in early stages of Alzheimers disease. Whats more, these data provide the first example of how the mechanism of iron transport regulation by apo- and holo-transferrin becomes misappropriated in disease to detrimental ends. The clinical benefit from understanding early dysregulation in brain iron transport in AD cannot be understated. If therapeutics can target this early process, they could possibly prevent the detrimental cascade that occurs with excessive iron accumulation. Significance StatementExcessive brain iron accumulation is hallmark pathology of Alzheimers disease that occurs early in the disease staging and before widespread proteinopathy deposition. This overabundance of brain iron has been implicated to contribute to disease progression, thus understandingthe mechanism of early iron accumulation has significant therapeutic potential to slow to halt disease progression. Here, we show that in response to low levels of amyloid-{beta} exposure, astrocytes increase their mitochondrial activity and iron uptake, resulting in iron deficient conditions. Elevated levels of apo (iron free)-transferrin stimulate iron release from endothelial cells. These data are the first to propose a mechanism for the initiation of iron accumulation and the misappropriation of iron transport signaling leading to dysfunctional brain iron homeostasis and resultant disease pathology.

neuroscience↗

bioRxiv2022

Nuclear receptor ligand screening in an iPSC-derived in vitro blood-brain barrier model identifies new contributors to leptin transport

BackgroundThe peptide hormone leptin exerts its function in the brain to reduce food intake and increase energy expenditure to prevent obesity. However, most obese subjects reflect the resistance to leptin even with elevated serum leptin. Considering that leptin must cross the blood-brain barrier (BBB) in several regions to enter the brain parenchyma, altered leptin transport through the BBB might play an important role in leptin resistance and other biological conditions. Here, we report the use of a human induced pluripotent stem cell (iPSC)-derived BBB model to explore mechanisms that influence leptin transport. MethodsiPSCs were differentiated into brain microvascular endothelial cell (BMEC)-like cells using standard methods. BMEC-like cells were cultured in Transwell filters, treated with ligands from a nuclear receptor agonist library, and assayed for leptin transport using an enzyme-linked immune sorbent assay. RNA sequencing was further used to identify differentially regulated genes and pathways. The role of a select hit in leptin transport was tested with the competitive substrate assay and after gene knockdown using CRISPR techniques. ResultsFollowing a screen of 73 compounds, 17{beta}-estradiol was identified as a compound that could significantly increase leptin transport. RNA sequencing revealed many differentially expressed transmembrane transporters after 17{beta}-estradiol treatment. Of these, cationic amino acid transporter-1 (CAT-1, encoded by SLC7A1) was selected for follow-up analyses due to its high and selective expression in BMECs in vivo. Treatment of BMEC-like cells with CAT-1 substrates, as well as knockdown of CAT-1 expression via CRISPR-mediated epigenome editing, yielded significant increases in leptin transport. ConclusionsA major female sex hormone, as well as an amino acid transporter, were revealed as regulators of leptin BBB transport in the iPSC-derived BBB model. Outcomes from this work provide insights into regulation of peptide hormone transport across the BBB.

neuroscience↗

bioRxiv2021

CRISPRi screens in human astrocytes elucidate regulators of distinct inflammatory reactive states

In response to central nervous system injury or disease, astrocytes become reactive, adopting context-dependent states and functional outputs. Certain inflammatory insults induce reactive astrocytes that lose homeostatic functions and gain harmful outputs through cellular pathways that are not fully understood. Here, we combined single-cell transcriptomics with CRISPRi screening in human iPSC-derived astrocytes to systematically interrogate inflammatory astrocyte reactivity. We found that autocrine-paracrine IL-6 and interferon signaling downstream of canonical NF-{kappa}B activation drove two distinct inflammatory reactive signatures - one promoted by and the other inhibited by STAT3. These signatures overlapped with those observed in other experimental contexts, including mouse models, and their markers were upregulated in the human brain in Alzheimers disease and hypoxic ischemic encephalopathy. Furthermore, we validated that these signatures were regulated by Stat3 in vivo. These results and the platform we established have the potential to guide the development of therapeutics to selectively modulate different aspects of inflammatory astrocyte reactivity.

neuroscience↗

bioRxiv2019

Machine learning on drug-specific data to predict small molecule teratogenicity

Pregnant women are an especially vulnerable population, given the sensitivity of a developing fetus to chemical exposures. However, prescribing behavior for the gravid patient is guided on limited human data and conflicting cases of adverse outcomes due to the exclusion of pregnant populations from randomized, controlled trials. These factors increase risk for adverse drug outcomes and reduce quality of care for pregnant populations. Herein, we propose the application of artificial intelligence to systematically predict the teratogenicity of a prescriptible small molecule from information inherent to the drug. Using unsupervised and supervised machine learning, our model probes all small molecules with known structure and teratogenicity data published in research-amenable formats to identify patterns among structural, meta-structural, and in vitro bioactivity data for each drug and its teratogenicity score. With this workflow, we discovered three chemical functionalities that predispose a drug towards increased teratogenicity and two moieties with potentially protective effects. Our models predict three clinically-relevant classes of teratogenicity with AUC = 0.8 and nearly double the predictive accuracy of a blind control for the same task, suggesting successful modeling. We also present extensive barriers to translational research that restrict data-driven studies in pregnancy and therapeutically "orphan" pregnant populations. Collectively, this work represents a first-in-kind platform for the application of computing to study and predict teratogenicity.

bioinformatics↗

bioRxiv2019

Development of an N-Cadherin Biofunctionalized Hydrogel to Support the Formation of Synaptically Connected Neural Networks

In vitro models of the human central nervous system (CNS), particularly those derived from induced pluripotent stem cells (iPSCs), are becoming increasingly recognized as useful complements to animal models for studying neurological diseases and developing therapeutic strategies. However, current 3D CNS models suffer from deficits that limit their research utility. Notably, it remains difficult to drive iPSC-derived neurons to a mature and synaptically connected state. Moreover, the most common extracellular matrices (ECMs) used to fabricate 3D CNS models are either difficult to pattern into complex structures due to their mechanical properties or lack appropriate bioinstructive cues. Here, we describe the functionalization of gelatin methacrylate (GelMA) with an N-cadherin extracellular peptide epitope to create a biomaterial termed GelMA-Cad. After photopolymerization, GelMA-Cad forms soft hydrogels that can maintain patterned architectures. The N-cadherin functionality promotes survival and maturation of iPSC-derived glutamatergic neurons into synaptically connected networks as determined by viral tracing and electrophysiology. Immunostaining reveals a pronounced increase in presynaptic and postsynaptic marker expression in GelMA-Cad relative to Matrigel, as well as extensive co-localization of these markers, thus highlighting the biological activity of the N-cadherin peptide. Overall, given its ability to enhance iPSC-derived neuron maturity and connectivity, GelMA-Cad should be broadly useful for in vitro studies of neural circuitry in health and disease.

bioengineering↗

bioRxiv2019

Spin{infty} an improved miniaturized spinning bioreactor for the generation of human cerebral organoids from pluripotent stem cells

Three-dimensional (3D) brain organoids derived from human pluripotent stem cells (hPSCs), including human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs), have become a powerful system to study early development events and to model human disease. Cerebral organoids are generally produced in static culture or in a culture vessel with active mixing, and the two most widely used systems for mixing are a large spinning flask and a miniaturized multi-well spinning bioreactor (also known as Spin Omega (Spin{Omega})). The Spin{Omega} provides a system that is amenable to drug testing, has increased throughput and reproducibility, and utilizes less culture media. However, technical limitations of this system include poor stability of select components and an elevated risk of contamination due to the inability to sterilize the device preassembled. Here, we report a new design of the miniaturized bioreactor system, which we term Spin{infty} that overcomes these concerns to permit long-term experiments.

bioengineering↗

bioRxiv2018

A 3D model of human skeletal muscle innervated with stem cell-derived motor neurons enables epsilon-subunit targeted myasthenic syndrome studies

Two-dimensional (2D) human skeletal muscle fiber cultures are ill equipped to support the contractile properties of maturing muscle fibers. This limits their application to the study of adult human neuromuscular junction (NMJ) development, a process requiring maturation of muscle fibers in the presence of motor neuron endplates. Here we describe a three-dimensional (3D) co-culture method whereby human muscle progenitors mixed with human pluripotent stem cell-derived motor neurons self-organize to form functional NMJ connections within two weeks. Functional connectivity between motor neuron endplates and muscle fibers is confirmed with calcium transient imaging and electrophysiological recordings. Notably, we only observed epsilon acetylcholine receptor subunit protein upregulation and activity in 3D co-culture. This demonstrates that the 3D co-culture system supports a developmental shift from the embryonic to adult form of the receptor that does not occur in 2D co-culture. Further, 3D co-culture treatments with myasthenia gravis patient sera shows the ease of studying human disease with the system. This work delivers a simple, reproducible, and adaptable method to model and evaluate adult human NMJ de novo development and disease in culture.

bioengineering↗