Biology subjects

Blackshaw, S.

Publications and source records attributed to Blackshaw, S..

At least 55 records · Page 3Linked to original sources

bioRxiv2021

Notch inhibition promotes regeneration and immunosuppression supports cone survival in a zebrafish model of inherited retinal dystrophy

Photoreceptor degeneration leads to irreversible vision loss in humans with retinal dystrophies such as Retinitis Pigmentosa. Whereas photoreceptor loss is permanent in mammals, zebrafish possesses the ability to regenerate retinal neurons and restore visual function. Following acute damage, Muller glia (MG) re-enter the cell cycle and produce multipotent progenitors whose progeny differentiate into mature neurons. Both MG reprogramming and proliferation of retinal progenitor cells require reactive microglia and associated inflammatory signaling. Paradoxically, MG in zebrafish models of photoreceptor degeneration fail to re-enter the cell cycle and regenerate lost cells. Here, we used the zebrafish cep290 mutant to demonstrate that progressive cone degeneration generates an immune response but does not stimulate MG proliferation. Acute light damage triggered photoreceptor regeneration in cep290 mutants but cones were only restored to pre-lesion densities. Using irf8 mutant zebrafish, we found that the chronic absence of microglia reduced inflammation and rescued cone degeneration in cep290 mutants. Finally, single-cell RNA-sequencing revealed sustained expression of notch3 in MG of cep290 mutants and inhibition of Notch signaling induced MG to re-enter the cell cycle. Our findings provide new insights on the requirements for MG to proliferate and the potential for immunosuppression to prolong photoreceptor survival.

neuroscience↗

bioRxiv2021

The neural cell adhesion molecule NrCAM regulates development of hypothalamic tanycytes

Hypothalamic tanycytes are neural stem and progenitor cells, but little is known of how they are regulated. Here we provide evidence that the cell adhesion molecule, NrCAM, regulates tanycytes in the adult niche. NrCAM is strongly expressed in adult mouse tanycytes. Immunohistochemical and in situ hybridization analysis revealed that NrCAM loss of function leads to both a reduced number of tanycytes and reduced expression of tanycyte-specific cell markers, along with a small reduction in tyrosine hydroxylase-positive arcuate neurons. Similar analyses of NrCAM mutants at E16 identify few changes in gene expression or cell composition, indicating that NrCAM regulates tanycytes, rather than early embryonic hypothalamic development. Neurosphere and organotypic assays support the idea that NrCAM governs cellular homeostasis. Single-cell RNA sequencing (scRNA-Seq) shows that tanycyte-specific genes, including a number that are implicated in thyroid hormone metabolism, show reduced expression in the mutant mouse. However, the mild tanycyte depletion and loss of markers observed in NrCAM-deficient mice were associated with only a subtle metabolic phenotype.

neuroscience↗

bioRxiv2021

IReNA: integrated regulatory network analysis of single-cell transcriptomes

Although single-cell RNA sequencing (scRNA-seq) and single-cell assay for transposase accessible chromatin using sequencing (scATAC-seq) have been widely used, few methods can reliably integrate these data to perform regulatory network analysis. Here, we developed IReNA (Integrated Regulatory Network Analysis) for network inference through integrated analysis of scRNA-seq and scATAC-seq data, network modularization, transcription factor enrichment, and construction of simplified intermodular regulatory networks. Using public datasets, we showed that integrated network analysis of scRNA-seq and scATAC-seq data using IReNA outperformed currently available methods in identifying known regulators. IReNA facilitates the systems-level understanding of biological regulatory mechanisms, and is available at https://github.com/jiang-junyao/IReNA.

bioinformatics↗

bioRxiv2021

Amyloid-beta and tau pathologies are both necessary to induce novel stage-specific microglia subtypes during Alzheimer's disease progression.

It is unknown whether specific microglia are selectively induced by amyloid-{beta} (A{beta}), tau pathologies, or both in combination. To address this, we use single-cell RNA-sequencing to profile mice bearing both A{beta} and tau pathologies during Alzheimers disease (AD) progression. We identify novel microglia subtypes induced in a disease stage-specific manner. We show that during early-stage disease, interferon signaling induces a subtype of microglia termed EADAM. During late-stage disease, a second microglia subtype termed LADAM is detected. While EADAM and LADAM-like microglia are observed in other neurodegenerative models, the magnitude and composition of subtype markers are distinct from microglia observed with AD-like pathology. The pattern of EADAM- and LADAM-associated gene expression is observed in microglia from human AD, during the early and late stages of disease, respectively. Furthermore, we observe that several siglec genes are selectively expressed in either EADAM or LADAM. Siglecg is expressed in white-matter-associated LADAM, and expression of the human orthologue of Siglecg is progressively elevated in AD-stage-dependent manner but not shown in non-AD tauopathy. Our findings imply that both A{beta} and tau pathologies are required for disease stage-specific induction of EADAM and LADAM.

neuroscience↗

bioRxiv2021

NFkB-signaling suppresses Müller glia-mediated neuron regeneration in the mammalian retina

Muller glia (MG) in mammalian retinas are incapable of regenerating neurons after damage, whereas the MG in lower vertebrates regenerate functional neurons. Identification networks that regulate MG-mediated regeneration is key to harnessing the regenerative potential of MG. Here we study how NFkB-signaling influences glial responses to damage and reprogramming of MG into neurons in the rodent retina. We find activation of NFkB and dynamic expression of NFkB-associated genes in MG after damage, however NFkB activity is inhibited by microglia ablation. Knockout of NFkB in MG suppressed the accumulation of immune cells after damage. Inhibition of NFkB following NMDA-damage significantly enhanced the reprogramming of Ascl1-overexpressing MG into neuron-like cells. scRNA-seq of retinal glia following inhibition of NFkB reveals coordination with signaling via TGF{beta}2 and suppression of NFI and Id transcription factors. Inhibition of Smad3 or Id transcription factors increased numbers of neuron-like cells produced by Ascl1-overexpressing MG. We conclude that NFkB is a key signaling hub that is activated in MG after damage, mediates the accumulation of immune cells, and suppresses the neurogenic potential of MG.

neuroscience↗

bioRxiv2021

Ptbp1 deletion does not induce glia-to-neuron conversion in adult mouse retina and brain.

Direct reprogramming of glia into neurons is a potentially promising approach for the replacement of neurons lost to injury or neurodegenerative disorders. Knockdown of the polypyrimidine tract-binding protein Ptbp1 has been recently reported to induce efficient conversion of retinal Muller glia and brain astrocytes into functional neurons. However, genetic analysis of Ptbp1 function in adult glia has not been conducted. Here, we use a combination of genetic lineage tracing, scRNA-Seq, and electrophysiological analysis to show that specific deletion of Ptbp1 in adult retinal Muller glia and brain astrocytes does not lead to any detectable level of glia-to-neuron conversion. Only a few changes in gene expression are observed in glia following Ptbp1 deletion, and glial identity is maintained. These findings highlight the importance of using genetic manipulation and lineage tracing methods in studying cell type conversion.

neuroscience↗

bioRxiv2021

Signaling and transcriptional networks governing late synovial joint development

BackgroundDuring synovial joint development, cavitation marks the end of the emergence of new cell types and the onset of the consolidation of cell type specific programs. However, the transcriptional programs that regulate this crucial stage prior to joint maturation are incompletely characterized. Gdf5-lineage cells give rise to the majority of joint constituents such as articular cartilage, meniscus, ligament, and tendon. Therefore, to explore pre-maturation of the synovial joint, we performed single cell RNA-Seq analysis of 1,306 Gdf5-lineage cells from the murine knee joint at E17.5. ResultsUsing computational analytics and in situ hybridization, we identified nine sub-states contributing to articular cartilage, meniscus, cruciate ligament, synovium, lining, and surrounding fibrous tissue. We identified a common progenitor population that is predicted to give rise to ligamentaocytes, articular chondrocytes, and lining cells. We further found that while a large number of signaling pathways orchestrate the differentiation of this progenitor to either ligamentocytes or to lining cells, only continued FGF signaling guides these cells to a default chondrocyte fate. ConclusionsOur single cell transcriptional atlas is a resource that can be used to better understand and further study synovial joint development and the reactivation of embryonic programs in diseases such as osteoarthritis.

developmental biology↗

bioRxiv2021

Fatty acid-binding proteins and fatty acid synthase influence glial reactivity and promote the formation of Müller glia-derived progenitor cells in the avian retina

The capacity for retinal regeneration varies greatly across vertebrates species. A recent comparative epigenetic and transcriptomic investigation of Muller glial (MG) in the retinas of fish, birds and mice revealed that Fatty Acid Binding Proteins (FABPs) are among the most highly up-regulated genes in activated chick MG (Hoang et al., 2020). Herein we provide an in-depth follow-up investigation to describe patterns of expression and how FABPs and fatty acid synthase (FASN) influence glial cells in the chick retina. During development, FABP7 is highly expressed by embryonic retinal progenitor cells (eRPCs) and maturing MG, whereas FABP5 is gradually up-regulated in maturing MG and remains elevated in mature glial cells. PMP2 (FABP8) is expressed by oligodendrocytes and FABP5 is expressed by non-astrocytic inner retinal glial cells, and both of these FABPs are significantly up-regulated in activated MG in damaged or growth factor-treated retinas. In addition to suppressing the formation of MGPCs, we find that FABP-inhibition suppressed the accumulation of proliferating microglia, although the microglia appeared highly reactive. scRNA-seq analyses of cells treated with FABP-inhibitor revealed distinct changes in patterns of expression suggesting that FABPs are involved in the transitions of MG from a resting state to a reactive state and conversion from reactive MG to MGPCs. Inhibition of FABPs in undamaged retinas had a significant impact upon the transcriptomic profiles of MG, with up-regulation of genes associated with gliogenesis, decreases in genes associated with neurogenesis, and suppression of the ability of MG to become MGPCs. scRNA-seq analyses of microglia indicated that FABP inhibition enhances gene modules related to reactivity, proliferation and cytokine signaling. We find that the proliferation of retinal progenitors in the circumferential marginal zone (CMZ) is unaffected by FABP-inhibitor. Upstream of FABP activity, we inhibited FASN in damaged retinas, which reduced numbers of dying cells, increased the proliferation of microglia, and potently suppressed the formation MGPCs in damaged retinas. We conclude that the activity of FASN and FABPs are required early during the formation of proliferating MGPCs. Fatty acid metabolism and cell signaling involving fatty acids are important in regulating glial homeostasis in the retina, and the dedifferentiation and proliferation of microglia and MGPCs.

developmental biology↗

bioRxiv2021

Gene regulatory networks controlling temporal patterning, neurogenesis and cell fate specification in the mammalian retina.

Gene regulatory networks (GRNs), consisting of transcription factors and their target cis- regulatory sequences, control neurogenesis and cell fate specification in the developing central nervous system, but their organization is poorly characterized. In this study, we performed integrated single-cell RNA- and scATAC-seq analysis in both mouse and human retina to profile dynamic changes in gene expression, chromatin accessibility and transcription factor footprinting during retinal neurogenesis. We identified multiple interconnected, evolutionarily-conserved GRNs consisting of cell type-specific transcription factors that both activate expression of genes within their own network and often inhibit expression of genes in other networks. These GRNs control state transitions within primary retinal progenitors that underlie temporal patterning, regulate the transition from primary to neurogenic progenitors, and drive specification of each major retinal cell type. We confirmed the prediction of this analysis that the NFI transcription factors Nfia, Nfib, and Nfix selectively activate expression of genes that promote late-stage temporal identity in primary retinal progenitors. We also used GRNs to identify additional transcription factors that promote (Insm1/2) and inhibit (Tbx3, Tcf7l1/2) rod photoreceptor specification in postnatal retina. This study provides an inventory of cis- and trans-acting factors that control retinal development, identifies transcription factors that control the temporal identity of retinal progenitors and cell fate specification, and will potentially guide cell-based therapies aimed at replacing retinal neurons lost due to disease.

neuroscience↗

bioRxiv2021

Multi-omic Analysis of Developing Human Retina and Organoids Reveals Cell-Specific Cis-Regulatory Elements and Mechanisms of Non-Coding Genetic Disease Risk.

Cis-regulatory elements (CREs) play a critical role in the development, maintenance, and disease-states of all human cell types. In the human retina, CREs have been implicated in a variety of inherited retinal disorders. To characterize cell-class-specific CREs in the human retina and elucidate their potential functions in development and disease, we performed single-nucleus (sn)ATAC-seq and snRNA-seq on the developing and adult human retina and on human retinal organoids. These analyses allowed us to identify cell-class-specific CREs, enriched transcription factor binding motifs, putative target genes, and to examine how these features change over development. By comparing DNA accessibility between the human retina and retinal organoids we found that CREs in organoids are highly correlated at the single-cell level, validating the use of organoids as a model for studying disease-associated CREs. As a proof of concept, we studied the function of a disease-associated CRE at 5q14.3 in organoids, identifying its principal target gene as the miR-9-2 primary transcript and demonstrating a dual role for this CRE in regulating neurogenesis and gene regulatory programs in mature glia. This study provides a rich resource for characterizing cell-class-specific CREs in the human retina and showcases retinal organoids as a model in which to study the function of retinal CREs that influence retinal development and disease. HIGHLIGHTSO_LISingle-cell map of cis-regulatory elements in developing and adult human retina. C_LIO_LICorrelation of single-cell DNA accessibility between human retina and retinal organoids. C_LIO_LIAssociation of disease risk loci with cell-class-specific accessibility. C_LIO_LIModeling of enhancer function at the 5q14.3 retinal disease-risk locus. C_LI

developmental biology↗

bioRxiv2021

Transcriptomic profiling of human orbital fat and differentiating orbital fibroblasts

Structured AbstractO_ST_ABSPurposeC_ST_ABSOrbital fat hyperplasia has a central role in the manifestations of thyroid-associated orbitopathy (TAO). To better understand the pathways involved in adipogenesis in TAO, we have used transcriptomic methods to analyze gene expression in control and TAO patients, as well as in differentiating orbital fibroblasts (OFs). MethodsWe performed bulk RNA sequencing (RNA-Seq) on intraconal orbital fat to compare gene expression in control and TAO patients. We treated cultured OFs derived from TAO patients with media containing dexamethasone, insulin, rosiglitazone, and isobutylmethylxanthine (IBMX) to induce adipogenesis. We used single nuclear RNA-Seq (snRNA-Seq) profiling of treated OFs to compare gene expression over time in order to identify pathways that are involved in orbital adipogenesis in vitro and compared the dynamic patterns of gene expression identify differences in gene expression in control and TAO orbital fat. ResultsOrbital fat from TAO and control patients segregate with principal component analysis (PCA). Numerous signaling pathways are enriched in orbital fat isolated from TAO patients. SnRNA-Seq of orbital fibroblasts undergoing adipogenesis reveals differential expression of adipocyte-specific genes over the developmental time course. Furthermore, genes that are enriched in TAO orbital fat are also upregulated in orbital adipocytes that differentiate in vitro, while genes that are enriched in control orbital fat are enriched in orbital fibroblasts prior to differentiation. ConclusionsDifferentiating orbital fibroblasts serve as a model to study orbital fat hyperplasia seen in TAO. We demonstrate that the insulin-like growth factor-1 receptor (IGF-1R) and Wnt signaling pathways are differentially expressed early in orbital adipogenesis. PrecisTo understand the pathways involved in adipogenesis in TAO, we used transcriptomic methods to analyze gene expression in control and TAO patients, as well as in differentiating OFs. We demonstrate that the IGF-1R and Wnt signaling pathways are differentially expressed during orbital adipogenesis.

molecular biology↗

bioRxiv2021

Lhx2 is a progenitor-intrinsic modulator of Sonic Hedgehog signaling during early retinal neurogenesis

An important question in organogenesis is how tissue-specific transcription factors interact with signaling pathways. In some cases, transcription factors define the context for how signaling pathways elicit tissue- or cell-specific responses, and in others, they influence signaling through transcriptional regulation of signaling components or accessory factors. We previously showed that during optic vesicle patterning, the Lim-homeodomain transcription factor Lhx2 has a contextual role by linking the Sonic Hedgehog (Shh) pathway to downstream targets without regulating the pathway itself. Here, we show that during early retinal neurogenesis, Lhx2 is a multilevel regulator of Shh signaling. Specifically, Lhx2 acts cell autonomously to control the expression of pathway genes required for efficient activation and maintenance of signaling in retinal progenitor cells. The Shh co-receptors Cdon and Gas1 are candidate direct targets of Lhx2 that mediate pathway activation, whereas Lhx2 directly or indirectly promotes the expression of other pathway components important for activation and sustained signaling. We also provide genetic evidence suggesting that Lhx2 has a contextual role by linking the Shh pathway to downstream targets. Through these interactions, Lhx2 establishes the competence for Shh signaling in retinal progenitors and the context for the pathway to promote early retinal neurogenesis. The temporally distinct interactions between Lhx2 and the Shh pathway in retinal development illustrate how transcription factors and signaling pathways adapt to meet stage-dependent requirements of tissue formation.

developmental biology↗

bioRxiv2021

Single-cell analysis of early chick hypothalamic development reveals that hypothalamic cells are induced from prethalamic-like progenitors.

The hypothalamus is an evolutionarily ancient brain region that regulates many innate behaviors, but its development is still poorly understood. To identify molecular mechanisms controlling hypothalamic specification and patterning, we used single-cell RNA-Seq to profile multiple stages of early hypothalamic development in the chick. We observe that hypothalamic neuroepithelial cells are initially induced from prethalamic-like cells. Two distinct hypothalamic progenitor populations emerge later, which give rise to paraventricular/mammillary and tuberal hypothalamus. At later developmental stages, the regional organization of the chick and mouse hypothalamus closely resembles one another. This study identifies selective markers for major subdivisions of the developing chick hypothalamus and many uncharacterized candidate regulators of hypothalamic patterning and neurogenesis. As proof of concept for the power of the dataset, we demonstrate that follistatin, a novel prethalamic progenitor-like marker, inhibits hypothalamic induction. This study both clarifies the organization of the early developing hypothalamus and identifies novel molecular mechanisms controlling hypothalamic induction, regionalization, and neurogenesis. HighlightsO_LIEarly hypothalamic development was profiled in chick using scRNA-Seq and multiplexed HCR. C_LIO_LIHypothalamic cells are induced from prethalamic-like neuroepithelial cells. C_LIO_LIDistinct paraventricular/mammillary and tuberal progenitor populations emerge later, and hypothalamic organization is evolutionarily conserved. C_LIO_LIPrethalamic progenitor-derived follistatin inhibits hypothalamic specification. C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/438683v1_ufig1.gif" ALT="Figure 1"> View larger version (73K): org.highwire.dtl.DTLVardef@18ac4c2org.highwire.dtl.DTLVardef@17bb90aorg.highwire.dtl.DTLVardef@1b17cceorg.highwire.dtl.DTLVardef@178e769_HPS_FORMAT_FIGEXP M_FIG C_FIG

developmental biology↗

bioRxiv2021

Cannabinoid signaling promotes the reprogramming of Muller glia into proliferating progenitor cells.

Endocannabinoids (eCB) are lipid-based neurotransmitters that are known to influence synaptic function in the visual system. eCBs are also known to suppress neuroinflammation in different pathological states. However, nothing is known about the roles of the eCB system during reprogramming of Muller glia (MG) into proliferating progenitor-like cells in the retina. Accordingly, we used the chick and mouse model to characterize expression patterns of eCB-related genes and applied pharmacological agents to examine how the eCB system impacts glial reactivity and the capacity of MG to become Muller glia-derived progenitor cells (MGPCs). We probed single cell RNA-seq libraries to identify eCB-related genes and identify cells with dynamic patterns of expression in damaged retinas. MG and inner retinal neurons expressed the eCB receptor CNR1, as well as enzymes involved in eCB metabolism. In the chick, intraocular injections of 2-Arachidonoylglycerol (2-AG) and Anandamide (AEA) potentiated the formation of MGPCs. Consistent with these findings, CNR1-agonists and MGLL-inhibitor promoted reprogramming, whereas CNR1-antagonist and inhibitors of eCB synthesis suppressed reprogramming. Surprisingly, retinal microglia were largely unaffected by increases or decreases in eCB signaling in both chick and mouse models. However, eCB-signaling suppressed the activation of NFkB-reporter in MG in damaged mouse retinas. We conclude that the eCB system in the retina influences the reactivity of MG and is important for regulating glial reactivity and the reprogramming of MG into proliferating MGPCs, but not for regulating the reactivity of immune cells in the retina. Main PointsMuller glia express CNR1 receptor and endocannabinoid synthesis genes. Endocannabinoids after retinal damage promote the formation of Muller glia derived progenitor cells in chick. Endocannabinoids reduce NFkB activity in mouse Muller glia.

molecular biology↗

bioRxiv2020

Control of neurogenic competence in mammalian hypothalamic tanycytes.

Hypothalamic tanycytes, radial glial cells that share many features with neuronal progenitors, generate small numbers of neurons in the postnatal hypothalamus, but the identity of these neurons and the molecular mechanisms that control tanycyte-derived neurogenesis are unknown. In this study, we demonstrate that tanycyte-specific disruption of the NFI family of transcription factors (Nfia/b/x) robustly stimulates tanycyte proliferation and tanycyte-derived neurogenesis. Single-cell RNA- and ATAC-Seq analysis reveals that NFI factors repress Shh and Wnt signaling in tanycytes, and small molecule modulation of these pathways blocks proliferation and tanycyte-derived neurogenesis in Nfia/b/x-deficient mice. We show that Nfia/b/x-deficient tanycytes give rise to multiple mediobasal hypothalamic neuronal subtypes that can mature, integrate into hypothalamic synaptic circuitry, and selectively respond to changes in internal states. These findings identify molecular mechanisms that control tanycyte-derived neurogenesis, suggesting a new therapeutic approach to selectively remodel the hypothalamic neural circuitry that controls homeostatic physiological processes.

neuroscience↗

bioRxiv2020

Temperature and species-dependent regulation of browning in retrobulbar fat.

Retrobulbar fat deposits surround the posterior retina and optic nerve head, but their function and origin are obscure. We report that mouse retrobulbar fat is a neural crest-derived tissue histologically and transcriptionally resembles interscapular brown fat. In contrast, human retrobulbar fat closely resembles white adipose tissue. Retrobulbar fat is also brown in other rodents, which are typically housed at temperatures below thermoneutrality, but is white in larger animals. We show that retrobulbar fat in mice housed at thermoneutral temperature show reduced expression of the brown fat marker Ucp1, and histological properties intermediate between white and brown fat. We conclude that retrobulbar fat can potentially serve as a site of active thermogenesis, that this capability is both temperature and species-dependent, and that this may facilitate regulation of intraocular temperature.

physiology↗

bioRxiv2020

Regulation of retinal neurogenesis by somatostatin signaling.

Neuropeptides have been reported to regulate progenitor proliferation and neurogenesis in the central nervous system. However, these studies have typically been conducted using pharmacological agents in ex vivo preparations, and in vivo evidence for their developmental function is generally lacking. Recent scRNA-Seq studies have identified multiple neuropeptides and their receptors as being selectively expressed in neurogenic progenitors of the embryonic mouse and human retina. This includes Sstr2, whose ligand somatostatin is transiently expressed by immature retinal ganglion cells. By analyzing retinal explants treated with selective ligands that target these receptors, we found that Sstr2-dependent somatostatin signaling induces a dose-dependent inhibition of photoreceptor generation while increasing the relative fraction of primary progenitor cells. These effects were confirmed by scRNA-Seq analysis of retinal explants and abolished in Sstr2-deficient retinas. Although no changes in the relative fraction of primary progenitors or photoreceptor precursors were observed in Sstr2-deficient retinas in vivo, scRNA-Seq analysis demonstrated accelerated differentiation of neurogenic progenitors. We conclude that Sstr2 signaling may act to negatively regulate retinal neurogenesis in combination with other retinal ganglion cell-derived secreted factors such as Shh, although in vivo Sstr2 is dispensable for normal retinal development.

neuroscience↗

bioRxiv2020

Midkine in chick and mouse retinas: neuroprotection, glial reactivity and the formation of Müller glia-derived progenitor cells

Recent studies have shown that midkine (MDK), a basic heparin-binding growth factor, is involved in the development and regeneration of the zebrafish retina. However, very little is known about MDK in the retinas of warm-blooded vertebrates. We investigate the expression patterns of MDK and related factors, roles in neuronal survival, and influence upon the formation of Muller glia-derived progenitor cells (MGPCs) in chick and mouse model systems. By using single-cell RNA-sequencing, we find that MDK is upregulated during Muller glia (MG) maturation in chick development and when stimulated to reprogram into MGPCs after NMDA damage or FGF2/Insulin treatment. Interestingly, MDK is significantly up-regulated by MG in damaged chick retinas, but down-regulated by MG in damaged mouse retinas. In both chick and mouse retinas, exogenous MDK selectively up-regulates cFOS and pS6 (a readout of mTOR-signaling) in MG. In the chick, intraocular injections of MDK before injury is neuroprotective with an observed decrease in dying neurons and microglial reactivity, inducing fewer proliferating MGPCs. Blocking MDK signaling with Na3VO4 following blocks neuroprotective effects with an increase the number of dying cells and negates the pro-proliferative effects on MGPCs. Inhibitors of PP2A and Pak1 associated with MDK integrin {beta}1 signaling had MG specific inhibitory effects on MGPC formation. In mice, MDK administration with NMDA damage drives a small but significant increase in MGPCs. We conclude that MDK expression is dynamically regulated in reactive Muller glia and during reprogramming into MGPCs. MDK acts to coordinate glial activity, neuronal survival, and may act in an autocrine manner to influence the re-programming of Muller glia into proliferating MGPCs.

neuroscience↗