IU Indianapolis ScholarWorks :: Browsing by Author "Meyer, Jason S."

Browsing by Author "Meyer, Jason S."

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Acquisition of neurodegenerative features in isogenic OPTN(E50K) human stem cell-derived retinal ganglion cells associated with autophagy disruption and mTORC1 signaling reduction
(Springer Nature, 2024-10-18) Huang, Kang‑Chieh; Gomes, Cátia; Shiga, Yukihiro; Belforte, Nicolas; VanderWall, Kirstin B.; Lavekar, Sailee S.; Fligor, Clarisse M.; Harkin, Jade; Hetzer, Shelby M.; Patil, Shruti V.; Di Polo, Adriana; Meyer, Jason S.; Biology, School of Science
The ability to derive retinal ganglion cells (RGCs) from human pluripotent stem cells (hPSCs) has led to numerous advances in the field of retinal research, with great potential for the use of hPSC-derived RGCs for studies of human retinal development, in vitro disease modeling, drug discovery, as well as their potential use for cell replacement therapeutics. Of all these possibilities, the use of hPSC-derived RGCs as a human-relevant platform for in vitro disease modeling has received the greatest attention, due to the translational relevance as well as the immediacy with which results may be obtained compared to more complex applications like cell replacement. While several studies to date have focused upon the use of hPSC-derived RGCs with genetic variants associated with glaucoma or other optic neuropathies, many of these have largely described cellular phenotypes with only limited advancement into exploring dysfunctional cellular pathways as a consequence of the disease-associated gene variants. Thus, to further advance this field of research, in the current study we leveraged an isogenic hPSC model with a glaucoma-associated mutation in the Optineurin (OPTN) protein, which plays a prominent role in autophagy. We identified an impairment of autophagic-lysosomal degradation and decreased mTORC1 signaling via activation of the stress sensor AMPK, along with subsequent neurodegeneration in OPTN(E50K) RGCs differentiated from hPSCs, and have further validated some of these findings in a mouse model of ocular hypertension. Pharmacological inhibition of mTORC1 in hPSC-derived RGCs recapitulated disease-related neurodegenerative phenotypes in otherwise healthy RGCs, while the mTOR-independent induction of autophagy reduced protein accumulation and restored neurite outgrowth in diseased OPTN(E50K) RGCs. Taken together, these results highlighted that autophagy disruption resulted in increased autophagic demand which was associated with downregulated signaling through mTORC1, contributing to the degeneration of RGCs.
Analysis of retinal ganglion cell development: from stem cells to synapses
(2018) Ohlemacher, Sarah K.; Meyer, Jason S.
Human pluripotent stem cells (hPSCs) have the ability to self renew indeﬁnitely while maintaining their pluripotency, allowing for the study of virtually any human cell type in a dish. The focus of the current study was the diﬀerentiation of hPSCs to retinal ganglion cells (RGCs), the primary cell type aﬀected in optic neuropathies. hPSCs were induced to become retinal cells using a stepwise diﬀerentiation protocol that allowed for formation of optic vesicle (OV)-like structures. Enrichment of OV like structures allowed for the deﬁnitive identiﬁcation of RGCs. RGCs displayed the proper temporal, spatial, and phenotypic characteristics of RGCs developing in vivo. To test the ability of hPSC-RGCs to serve as a disease model, lines were generated from a patient with an E50K mutation in the Optineurin gene, causative for normal tension primary open angle glaucoma. E50K RGCs displayed signiﬁcantly higher levels of apoptosis compared to a control lines. Apoptosis was reduced with exposure to neuroprotective factors. Lastly, hPSC-derived RGCs were studied for their ability to develop functional features possessed by mature in vivo RGCs. hPSC-derived RGCs displayed a few immature functional features and as such, strategies in which to expedite synaptogenesis using hPSC-derived astrocytes were explored. Astrocyte and RGG co-cultures displayed expedited synaptic and functional maturation, more closely resembling mature in vivo RGCs. Taken together, the results of this study have important implications for the study of RGC development and by extension, the advancement of translational therapies for optic neuropathies.
Astrocytes modulate neurodegenerative phenotypes associated with glaucoma in OPTN(E50K) human stem cell-derived retinal ganglion cells
(Elsevier, 2022) Gomes, Cátia; VanderWall, Kirstin B.; Pan, Yanling; Lu, Xiaoyu; Lavekar, Sailee S.; Huang, Kang-Chieh; Fligor, Clarisse M.; Harkin, Jade; Zhang, Chi; Cummins, Theodore R.; Meyer, Jason S.; Medical and Molecular Genetics, School of Medicine
Although the degeneration of retinal ganglion cells (RGCs) is a primary characteristic of glaucoma, astrocytes also contribute to their neurodegeneration in disease states. Although studies often explore cell-autonomous aspects of RGC neurodegeneration, a more comprehensive model of glaucoma should take into consideration interactions between astrocytes and RGCs. To explore this concept, RGCs and astrocytes were differentiated from human pluripotent stem cells (hPSCs) with a glaucoma-associated OPTN(E50K) mutation along with corresponding isogenic controls. Initial results indicated significant changes in OPTN(E50K) astrocytes, including evidence of autophagy dysfunction. Subsequently, co-culture experiments demonstrated that OPTN(E50K) astrocytes led to neurodegenerative properties in otherwise healthy RGCs, while healthy astrocytes rescued some neurodegenerative features in OPTN(E50K) RGCs. These results are the first to identify disease phenotypes in OPTN(E50K) astrocytes, including how their modulation of RGCs is affected. Moreover, these results support the concept that astrocytes could offer a promising target for therapeutic intervention in glaucoma.
Astrocytes Regulate the Development and Maturation of Retinal Ganglion Cells Derived from Human Pluripotent Stem Cells
(Elsevier, 2019-02-12) VanderWall, Kirstin B.; Vij, Ridhima; Ohlemacher, Sarah K.; Sridhar, Akshayalakshmi; Fligor, Clarisse M.; Feder, Elyse M.; Edler, Michael C.; Baucum, Anthony J.; Cummins, Theodore R.; Meyer, Jason S.; Biology, School of Science
Retinal ganglion cells (RGCs) form the connection between the eye and the brain, with this connectivity disrupted in numerous blinding disorders. Previous studies have demonstrated the ability to derive RGCs from human pluripotent stem cells (hPSCs); however, these cells exhibited some characteristics that indicated a limited state of maturation. Among the many factors known to influence RGC development in the retina, astrocytes are known to play a significant role in their functional maturation. Thus, efforts of the current study examined the functional maturation of hPSC-derived RGCs, including the ability of astrocytes to modulate this developmental timeline. Morphological and functional properties of RGCs were found to increase over time, with astrocytes significantly accelerating the functional maturation of hPSC-derived RGCs. The results of this study clearly demonstrate the functional and morphological maturation of RGCs in vitro, including the effects of astrocytes on the maturation of hPSC-derived RGCs.
Autophagy disruption reduces mTORC1 activation leading to retinal ganglion cell neurodegeneration associated with glaucoma
(Cold Spring Harbor Laboratory, 2023-01-04) Huang, Kang-Chieh; Gomes, Cátia; Shiga, Yukihiro; Belforte, Nicolas; VanderWall, Kirstin B.; Lavekar, Sailee S.; Fligor, Clarisse M.; Harkin, Jade; Di Polo, Adriana; Meyer, Jason S.; Biology, School of Science
Autophagy dysfunction has been associated with several neurodegenerative diseases including glaucoma, characterized by the degeneration of retinal ganglion cells (RGCs). However, the mechanisms by which autophagy dysfunction promotes RGC damage remain unclear. Here, we hypothesized that perturbation of the autophagy pathway results in increased autophagic demand, thereby downregulating signaling through mammalian target of rapamycin complex 1 (mTORC1), a negative regulator of autophagy, contributing to the degeneration of RGCs. We identified an impairment of autophagic-lysosomal degradation and decreased mTORC1 signaling via activation of the stress sensor adenosine monophosphate-activated protein kinase (AMPK), along with subsequent neurodegeneration in RGCs differentiated from human pluripotent stem cells (hPSCs) with a glaucoma-associated variant of Optineurin (OPTN-E50K). Similarly, the microbead occlusion model of glaucoma resulting in ocular hypertension also exhibited autophagy disruption and mTORC1 downregulation. Pharmacological inhibition of mTORC1 in hPSC-derived RGCs recapitulated disease-related neurodegenerative phenotypes in otherwise healthy RGCs, while the mTOR-independent induction of autophagy reduced protein accumulation and restored neurite outgrowth in diseased OPTN-E50K RGCs. Taken together, these results highlight an important balance between autophagy and mTORC1 signaling essential for RGC homeostasis, while disruption to these pathways contributes to neurodegenerative features in glaucoma, providing a potential therapeutic target to prevent neurodegeneration.
Defining Inner Ear Cell Type Specification at Single-Cell Resolution in a Model of Human Cranial Development
(2022-07) Steinhart, Matthew Reed; Meyer, Jason S.; Koehler, Karl R.; Herbert, Brittney-Shea; Landreth, Gary E.; Shearer, A. Eliot; Yates, Charles W.
Inner ear development requires the complex interaction of numerous cell types arising from multiple embryologic origins. Current knowledge of inner ear organogenesis is limited primarily to animal models. Although most mechanisms of cellular development show conservation between vertebrate species, there are uniquely human aspects of inner ear development which remain unknown. Our group recently described a model of in vitro human inner ear organogenesis using pluripotent stem cells in a 3D organoid culture system. This method promotes the formation of an entire sensorineural circuit, including hair cells, inner ear neurons, and Schwann cells. Our past work has characterized certain aspects of this culture system, however we have yet to fully define all the cell types which contribute to inner ear organoid assembly. Here, our goal was to reconstruct a time-based map of in vitro development during inner ear organoid induction to understand the developmental elements captured in this system. We analyzed inner ear organoid development using single-cell RNA sequencing at ten time points during the first 36 days of induction. We reconstructed the on-target progression of undifferentiated pluripotent stem cells to surface ectoderm, pre-placodal, and otic epithelial cells, including supporting cells, hair cells, and neurons, following treatment with FGF, BMP, and WNT signaling modulators. Our data revealed endogenous signaling pathwayrelated gene expression that may influence the course of on-target differentiation. In addition, we classified a diverse array of off-target ectodermal cell types encompassing the neuroectoderm, neural crest, and mesenchymal lineages. Our work establishes the Inner ear Organoid Developmental Atlas (IODA), which can provide insights needed for understanding human biology and refining the guided differentiation of in vitro inner ear tissue.
Derivation of endothelial colony forming cells from human cord blood and embryonic stem cells
(2013) Meador, J. Luke; Kapur, Ruben; Yoder, Mervin C.; Corson, Timothy W.; Meyer, Jason S.
Endothelial Colony Forming Cells (ECFCs) are highly proliferative endothelial progenitor cells with clonal proliferative potential and in vivo vessel forming ability. While endothelial cells have been derived from human induced pluripotent stem cells (hiPS) or human embryonic stem cells (hES), they are not highly proliferative and require ectopic expression of a TGFβ inhibitor to restrict plasticity. Neuropilin-1 (NRP-1) has been reported to identify the emergence of endothelial precursor cells from human and mouse ES cells undergoing endothelial differentiation. However, the protocol used in that study was not well defined, used uncharacterized neuronal induction reagents in the culture medium, and failed to fully characterize the endothelial cells derived. We hypothesize that NRP-1 expression is critical for the emergence of stable endothelial cells with ECFC properties from hES cells. We developed a novel serum and feeder free defined endothelial differentiation protocol to induce stable endothelial cells possessing cells with cord blood ECFC-like properties from hES cells. We have shown that Day 12 hES cell-derived endothelial cells express the endothelial markers CD31+ NRP-1+, exhibit high proliferative potential at a single cell level, and display robust in vivo vessel forming ability similar to that of cord blood-derived ECFCs. The efficient production of the ECFCs from hES cells is 6 logs higher with this protocol than any previously published method. These results demonstrate progress towards differentiating ECFC from hES and may provide patients with stable autologous cells capable of repairing injured, dysfunctional, or senescent vasculature if these findings can be repeated with hiPS.
Development of a three-dimensional organoid model to explore early retinal phenotypes associated with Alzheimer’s disease
(Springer Nature, 2023-08-24) Lavekar, Sailee S.; Harkin, Jade; Hernandez, Melody; Gomes, Cátia; Patil, Shruti; Huang, Kang‑Chieh; Puntambekar, Shweta S.; Lamb, Bruce T.; Meyer, Jason S.; Biology, School of Science
Alzheimer's disease (AD) is a progressive neurodegenerative disorder characterized by the accumulation of Aβ plaques and neurofibrillary tangles, resulting in synaptic loss and neurodegeneration. The retina is an extension of the central nervous system within the eye, sharing many structural similarities with the brain, and previous studies have observed AD-related phenotypes within the retina. Three-dimensional retinal organoids differentiated from human pluripotent stem cells (hPSCs) can effectively model some of the earliest manifestations of disease states, yet early AD-associated phenotypes have not yet been examined. Thus, the current study focused upon the differentiation of hPSCs into retinal organoids for the analysis of early AD-associated alterations. Results demonstrated the robust differentiation of retinal organoids from both familial AD and unaffected control cell lines, with familial AD retinal organoids exhibiting a significant increase in the Aβ42:Aβ40 ratio as well as phosphorylated Tau protein, characteristic of AD pathology. Further, transcriptional analyses demonstrated the differential expression of many genes and cellular pathways, including those associated with synaptic dysfunction. Taken together, the current study demonstrates the ability of retinal organoids to serve as a powerful model for the identification of some of the earliest retinal alterations associated with AD.
Differential susceptibility of retinal ganglion cell subtypes in acute and chronic models of injury and disease
(Nature Publishing Group, 2020-10-15) VanderWall, Kirstin B.; Lu, Bin; Alfaro, Jorge S.; Allsop, Anna R.; Carr, Alexa S.; Wang, Shaomei; Meyer, Jason S.; Medical and Molecular Genetics, School of Medicine
Retinal ganglion cells (RGCs) are a heterogeneous population of neurons, comprised of numerous subtypes that work synchronously to transmit visual information to the brain. In blinding disorders such as glaucoma, RGCs are the main cell type to degenerate and lead to loss of vision. Previous studies have identified and characterized a variety of RGC subtypes in animal models, although only a handful of studies demonstrate the differential loss of these RGC subtypes in response to disease or injury. Thus, efforts of the current study utilized both chronic (bead occlusion) and acute (optic nerve crush, ONC) rat models to characterize disease response and differential loss of RGC subtypes. Bead occlusion and ONC retinas demonstrated significant RGC loss, glial reactivity and apoptosis compared to control retinas. Importantly, bead occlusion and ONC retinas resulted in differential subtype-specific loss of RGCs, with a high susceptibility for alpha- and direction selective-RGCs and preferential survival of ipRGCs. Results of this study serve as an important foundation for future experiments focused on the mechanisms resulting in the loss of RGCs in optic neuropathies, as well as the development of targeted therapeutics for RGC subtype-specific neuroprotection.
Differentiation and characterization of cell types associated with retinal degenerative diseases using human induced pluripotent stem cells
(2014-07-31) Gupta, Manav; Meyer, Jason S.; Belecky-Adams, Teri; Randall, Stephen Karl, 1953-
Human induced pluripotent stem (iPS) cells have the unique ability to differentiate into 200 or so somatic cell types that make up the adult human being. The use of human iPS cells to study development and disease is a highly exciting and interdependent field that holds great promise in understanding and elucidating mechanisms behind cellular differentiation with future applications in drug screening and cell replacement studies for complex and currently incurable cellular degenerative disorders. The recent advent of iPS cell technology allows for the generation of patient-specific cell lines that enable us to model the progression of a disease phenotype in a human in vitro model. Differentiation of iPS cells toward the affected cell type provides an unlimited source of diseased cells for examination, and to further study the developmental progression of the disease in vitro, also called the “disease-in-a-dish” model. In this study, efforts were undertaken to recapitulate the differentiation of distinct retinal cell affected in two highly prevalent retinal diseases, Usher syndrome and glaucoma. Using a line of Type III Usher Syndrome patient derived iPS cells efforts were undertaken to develop such an approach as an effective in vitro model for studies of Usher Syndrome, the most commonly inherited disorder affecting both vision and hearing. Using existing lines of iPS cells, studies were also aimed at differentiation and characterization of the more complex retinal cell types, retinal ganglion cells (RGCs) and astrocytes, the cell types affected in glaucoma, a severe neurodegenerative disease of the retina leading to eventual irreversible blindness. Using a previously described protocol, the iPS cells were directed to differentiate toward a retinal fate through a step-wise process that proceeds through all of the major stages of neuroretinal development. The differentiation process was monitored for a period of 70 days for the differentiation of retinal cell types and 150 days for astrocyte development. The different stages of differentiation and the individually derived somatic cell types were characterized by the expression of developmentally associated transcription factors specific to each cell type. Further approaches were undertaken to characterize the morphological differences between RGCs and other neuroretinal cell types derived in the process. The results of this study successfully demonstrated that Usher syndrome patient derived iPS cells differentiated to the affected photoreceptors of Usher syndrome along with other mature retinal cell types, chronologically analogous to the development of the cell types in a mature human retina. This study also established a robust method for the in vitro derivation of RGCs and astrocytes from human iPS cells and provided novel methodologies and evidence to characterize these individual somatic cell types. Overall, this study provides a unique insight into the application of human pluripotent stem cell biology by establishing a novel platform for future studies of in vitro disease modeling of the retinal degenerative diseases: Usher syndrome and glaucoma. In downstream applications of this study, the disease relevant cell types derived from human iPS cells can be used as tools to further study disease progression, drug screening and cell replacement strategies.

Browsing by Author "Meyer, Jason S."

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