ORCID Profile
0000-0001-8212-9884
Current Organisation
Florey Institute of Neuroscience and Mental Health
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Research Topics
In Research Link Australia (RLA), "Research Topics" refer to ANZSRC FOR and SEO codes. These topics are either sourced from ANZSRC FOR and SEO codes listed in researchers' related grants or generated by a large language model (LLM) based on their publications.
Top 5 Research Topics
ANZSRC Field of Research (FoR)
Central Nervous System | Biomaterials | Regenerative Medicine (incl. Stem Cells and Tissue Engineering) | Medical Biotechnology | Medical Biochemistry and Metabolomics | Neurosciences | Medical Biochemistry and Metabolomics not elsewhere classified | Nanobiotechnology | Cell Development, Proliferation and Death |
ANZSRC Socio-Economic Objective (SEO)
Expanding Knowledge in Engineering | Expanding Knowledge in the Biological Sciences | Cardiovascular System and Diseases | Nervous System and Disorders | Blood Disorders | Expanding Knowledge in the Medical and Health Sciences
Related Links
Publications
Creating a Ventral Midbrain Stem Cell Niche in an Animal Model for Parkinson's Disease
Publisher: Mary Ann Liebert Inc
Date: 12-2010
Abstract: Neural progenitor cells reside in many regions of the adult brain. However, their capacity to generate new neurons relies on stem cell niches, consisting of stem cells, niche support cells, and basal lamina, which maintain stem cells and direct their differentiation and migration into tissue structures. Neurospheres are thought to expose neural progenitor cells to an environment reminiscent of the stem cell niche. We show that embryonic day 14.5 ventral mesencephalon neurospheres grafted into the midbrain of 6-hydroxydopamine lesioned mice express markers of mesenchymal cells, such as CD29 and CD44, and enclose a core of host-derived proliferating cells that express nestin, polysialic acid-neural cell adhesion molecule, βIII-tubulin, and neuron-specific nuclear protein. Laminin was expressed between the grafted cells and the core of proliferating host-derived cells. Further, infusion of the anti-mitotic agent β-d-arabinofuroside into the midbrain resulted in the loss of host-derived core cells that gradually returned over many days following β-d-arabinofuroside withdrawal. Together, these data suggest that a stem cell niche had been formed. Tyrosine hydroxylase (TH+) cells, ectopic to the usual midbrain locations, were present 4 weeks after grafting and increased in numbers up to 12 weeks after grafting, resulting in significantly more TH cells than control animals. These data provide evidence that cells within the midbrain are capable of acquiring a TH phenotype when exposed to the appropriate environment. Whether these cells are a result of neurogenesis or phenotypic shift remains unanswered.
SFRP1 and SFRP2 dose-dependently regulate midbrain dopamine neuron development in vivo and in embryonic stem cells
Publisher: Oxford University Press (OUP)
Date: 09-04-2012
DOI: 10.1002/STEM.1049
Characterization of the stability and bio-functionality of tethered proteins on bioengineered scaffolds: Implications for stem cell biology and tissue repair
Publisher: Elsevier BV
Date: 05-2014
Biomimetic Materials and Their Utility in Modeling the 3-Dimensional Neural Environment.
Publisher: Elsevier BV
Date: 2019
Primary tissue for cellular brain repair in Parkinson's disease: Promise, problems and the potential of biomaterials
Publisher: Wiley
Date: 10-07-2019
DOI: 10.1111/EJN.14051
Abstract: The dopamine precursor, levodopa, remains the "gold standard" treatment for Parkinson's disease, and, although it provides superlative efficacy in the early stages of the disease, its long-term use is limited by the development of severe motor side effects and a significant abating of therapeutic efficacy. Therefore, there remains a major unmet clinical need for the development of effective neuroprotective, neurorestorative or neuroreparatory therapies for this condition. The relatively selective loss of dopaminergic neurons from the nigrostriatal pathway makes Parkinson's disease an ideal candidate for reparative cell therapies, wherein the dopaminergic neurons that are lost in the condition are replaced through direct cell transplantation into the brain. To date, this approach has been developed, validated and clinically assessed using dopamine neuron-rich foetal ventral mesencephalon grafts which have been shown to survive and reinnervate the denervated brain after transplantation, and to restore motor function. However, despite long-term symptomatic relief in some patients, significant limitations, including poor graft survival and the impact this has on the number of foetal donors required, have prevented this therapy being more widely adopted as a restorative approach for Parkinson's disease. Injectable biomaterial scaffolds have the potential to improve the delivery, engraftment and survival of these grafts in the brain through provision of a supportive microenvironment for cell adhesion, growth and immune shielding. This article will briefly review the development of primary cell therapies for brain repair in Parkinson's disease and will consider the emerging literature which highlights the potential of using injectable biomaterial hydrogels in this context.
Efficiently Specified Ventral Midbrain Dopamine Neurons from Human Pluripotent Stem Cells Under Xeno-Free Conditions Restore Motor Deficits in Parkinsonian Rodents
Publisher: Oxford University Press (OUP)
Date: 14-10-2016
Abstract: Recent studies have shown evidence for the functional integration of human pluripotent stem cell (hPSC)-derived ventral midbrain dopamine (vmDA) neurons in animal models of Parkinson's disease. Although these cells present a sustainable alternative to fetal mesencephalic grafts, a number of hurdles require attention prior to clinical translation. These include the persistent use of xenogeneic reagents and challenges associated with scalability and storage of differentiated cells. In this study, we describe the first fully defined feeder- and xenogeneic-free protocol for the generation of vmDA neurons from hPSCs and utilize two novel reporter knock-in lines (LMX1A-eGFP and PITX3-eGFP) for in-depth in vitro and in vivo tracking. Across multiple embryonic and induced hPSC lines, this “next generation” protocol consistently increases both the yield and proportion of vmDA neural progenitors (OTX2/FOXA2/LMX1A) and neurons (FOXA2/TH/PITX3) that display classical vmDA metabolic and electrophysiological properties. We identify the mechanism underlying these improvements and demonstrate clinical applicability with the first report of scalability and cryopreservation of bona fide vmDA progenitors at a time amenable to transplantation. Finally, transplantation of xeno-free vmDA progenitors from LMX1A- and PITX3-eGFP reporter lines into Parkinsonian rodents demonstrates improved engraftment outcomes and restoration of motor deficits. These findings provide important and necessary advancements for the translation of hPSC-derived neurons into the clinic.
Hydrogel oxygen reservoirs increase functional integration of neural stem cell grafts by meeting metabolic demands
Publisher: Springer Science and Business Media LLC
Date: 28-01-2023
DOI: 10.1038/S41467-023-36133-8
Abstract: Injectable biomimetic hydrogels have great potential for use in regenerative medicine as cellular delivery vectors. However, they can suffer from issues relating to hypoxia, including poor cell survival, differentiation, and functional integration owing to the lack of an established vascular network. Here we engineer a hybrid myoglobin:peptide hydrogel that can concomitantly deliver stem cells and oxygen to the brain to support engraftment until vascularisation can occur naturally. We show that this hybrid hydrogel can modulate cell fate specification within progenitor cell grafts, resulting in a significant increase in neuronal differentiation. We find that the addition of myoglobin to the hydrogel results in more extensive innervation within the host tissue from the grafted cells, which is essential for neuronal replacement strategies to ensure functional synaptic connectivity. This approach could result in greater functional integration of stem cell-derived grafts for the treatment of neural injuries and diseases affecting the central and peripheral nervous systems.
Temporally controlled growth factor delivery from a self-assembling peptide hydrogel and electrospun nanofibre composite scaffold
Publisher: Royal Society of Chemistry (RSC)
Date: 2017
DOI: 10.1039/C7NR05004F
Abstract: Tissue-specific self-assembling peptide (SAP) hydrogels designed based on biologically relevant peptide sequences have great potential in regenerative medicine.
GAPTrap: A Simple Expression System for Pluripotent Stem Cells and Their Derivatives
Publisher: Elsevier BV
Date: 09-2016
14-3-3ζ deficient mice in the BALB/c background display behavioural and anatomical defects associated with neurodevelopmental disorders
Publisher: Springer Science and Business Media LLC
Date: 24-07-2015
DOI: 10.1038/SREP12434
Abstract: Sequencing and expression analyses implicate 14-3-3ζ as a genetic risk factor for neurodevelopmental disorders such as schizophrenia and autism. In support of this notion, we recently found that 14-3-3ζ −/− mice in the Sv/129 background display schizophrenia-like defects. As epistatic interactions play a significant role in disease pathogenesis we generated a new congenic strain in the BALB/c background to determine the impact of genetic interactions on the 14-3-3ζ −/− phenotype. In addition to replicating defects such as aberrant mossy fibre connectivity and impaired spatial memory, our analysis of 14-3-3ζ −/− BALB/c mice identified enlarged lateral ventricles, reduced synaptic density and ectopically positioned pyramidal neurons in all subfields of the hippoc us. In contrast to our previous analyses, 14-3-3ζ −/− BALB/c mice lacked locomotor hyperactivity that was underscored by normal levels of the dopamine transporter (DAT) and dopamine signalling. Taken together, our results demonstrate that dysfunction of 14-3-3ζ gives rise to many of the pathological hallmarks associated with the human condition. 14-3-3ζ-deficient BALB/c mice therefore provide a novel model to address the underlying biology of structural defects affecting the hippoc us and ventricle and cognitive defects such as hippoc al-dependent learning and memory.
Transcriptional Profiling of Xenogeneic Transplants: Examining Human Pluripotent Stem Cell-Derived Grafts in the Rodent Brain
Publisher: Elsevier BV
Date: 11-2019
Extracellular Matrix Biomimetic Hydrogels, Encapsulated with Stromal Cell-Derived Factor 1, Improve the Composition of Foetal Tissue Grafts in a Rodent Model of Parkinson’s Disease
Publisher: MDPI AG
Date: 22-04-2022
DOI: 10.3390/IJMS23094646
Abstract: Clinical studies have provided evidence for dopamine (DA) cell replacement therapy in Parkinson’s Disease. However, grafts derived from foetal tissue or pluripotent stem cells (PSCs) remain heterogeneous, with a high proportion of non-dopaminergic cells, and display subthreshold reinnervation of target tissues, thereby highlighting the need to identify new strategies to improve graft outcomes. In recent work, Stromal Cell-Derived Factor-1 (SDF1), secreted from meninges, has been shown to exert many roles during ventral midbrain DA development and DA-directed differentiation of PSCs. Related, co-implantation of meningeal cells has been shown to improve neural graft outcomes, however, no direct evidence for the role of SDF1 in neural grafting has been shown. Due to the rapid degradation of SDF1 protein, here, we utilised a hydrogel to entrap the protein and sustain its delivery at the transplant site to assess the impact on DA progenitor differentiation, survival and plasticity. Hydrogels were fabricated from self-assembling peptides (SAP), presenting an epitope for laminin, the brain’s main extracellular matrix protein, thereby providing cell adhesive support for the grafts and additional laminin–integrin signalling to influence cell fate. We show that SDF1 functionalised SAP hydrogels resulted in larger grafts, containing more DA neurons, increased A9 DA specification (the subpopulation of DA neurons responsible for motor function) and enhanced innervation. These findings demonstrate the capacity for functionalised, tissue-specific hydrogels to improve the composition of grafts targeted for neural repair.
Modulating Wnt signaling to improve cell replacement therapy for Parkinson's disease
Publisher: Oxford University Press (OUP)
Date: 13-12-2013
DOI: 10.1093/JMCB/MJT045
Abstract: Clinical trials have demonstrated the capacity for dopamine neurons, transplanted ectopically into the striatum, to structurally integrate, restore dopamine transmission, and induce long-term functional benefits for Parkinson's disease (PD) patients. Despite this proof of principle, a number of limitations have hindered the development of cell replacement therapy over the past 20 years, particularly tissue availability, graft survival, and adequate reinnervation of the host brain. With a greater understanding of failure in prior clinical trials, increased knowledge of midbrain dopamine development (now including Wnts), and the development of pluripotent stem cell technologies, we are better equipped than ever to re-address a number of these challenges. This review summarizes the trials, tribulations, and progress in cell replacement therapy for PD. We discuss the prospects of modulating canonical and non-canonical Wnt signaling to improve cell therapy based upon their roles in dopamine neural development and the adult brain. This will include the potential of Wnts to (i) expand fetally derived tissue in vitro and following transplantation, (ii) promote the differentiation of pluripotent stem cells, (iii) increase graft integration and restoration of neural circuitry, and finally (iv) enhance graft survival.
Functionalized composite scaffolds improve the engraftment of transplanted dopaminergic progenitors in a mouse model of Parkinson's disease
Publisher: Elsevier BV
Date: 2001
DOI: 10.1016/J.BIOMATERIALS.2015.09.039
Abstract: With the brain's limited capacity for repair there is a need for new and innovative therapies to promote regeneration. Stem rogenitor cell transplantation has received increasing attention, and whilst clinical trials demonstrating functional integration exist, inherent variability between patients has hindered development of this therapy. Variable outcomes have largely been attributed to poor survival and insufficient reinnervation of target tissues due in part to the suboptimal host environment. Here we examined whether improving the physical properties of the host milieu, by way of bioengineered scaffolds, may enhance engraftment. We developed a composite scaffold, incorporating electrospun poly(l-lactic acid) short nanofibers embedded within a thermo-responsive xyloglucan hydrogel, which could be easily injected into the injured brain. Furthermore, to improve the trophic properties of the host brain, glial derived neurotrophic factor (GDNF), a protein known to promote cell survival and axonal growth, was blended into and/or covalently attached onto the composite scaffolds to provide controlled delivery. In vitro we confirmed the ability of the scaffolds to support ventral midbrain (VM) dopamine progenitors, and provide sustained delivery of GDNF - capable of eliciting effects on cell survival and dopaminergic axon growth. In Parkinsonian mice, we show that these composite scaffolds, whilst having no deleterious impact on the host immune response, enhanced the survival of VM grafts and reinnervation of the striatum, an effect that was augmented through the scaffold delivery of GDNF. Taken together, these functionalized composite scaffolds provide a means to significantly improve the milieu of the injured brain, enabling enhanced survival and integration of grafted neurons.
Homophilic binding of the neural cell adhesion molecule CHL1 regulates development of ventral midbrain dopaminergic pathways
Publisher: Springer Science and Business Media LLC
Date: 24-08-2017
DOI: 10.1038/S41598-017-09599-Y
Abstract: Abnormal development of ventral midbrain (VM) dopaminergic (DA) pathways, essential for motor and cognitive function, may underpin a number of neurological disorders and thereby highlight the importance of understanding the birth and connectivity of the associated neurons. While a number of regulators of VM DA neurogenesis are known, processes involved in later developmental events, including terminal differentiation and axon morphogenesis, are less well understood. Recent transcriptional analysis studies of the developing VM identified genes expressed during these stages, including the cell adhesion molecule with homology to L1 ( Chl1 ). Here, we map the temporal and spatial expression of CHL1 and assess functional roles of substrate-bound and soluble-forms of the protein during VM DA development. Results showed early CHL1 in the VM, corresponding with roles in DA progenitor migration and differentiation. Subsequently, we demonstrated roles for CHL1 in both axonal extension and repulsion, selectively of DA neurons, suggestive of a role in guidance towards forebrain targets and away from hindbrain nuclei. In part, CHL1 mediates these roles through homophilic CHL1-CHL1 interactions. Collectively, these findings enhance our knowledge of VM DA pathways development, and may provide new insights into understanding DA developmental conditions such as autism spectrum disorders.
FGF-MAPK signaling regulates human deep-layer corticogenesis
Publisher: Elsevier BV
Date: 05-2021
Tailoring minimalist self-assembling peptides for localized viral vector gene delivery
Publisher: Springer Science and Business Media LLC
Date: 23-12-2016
Identification of cell barcodes from long-read single-cell RNA-seq with BLAZE
Publisher: Cold Spring Harbor Laboratory
Date: 16-08-2022
DOI: 10.1101/2022.08.16.504056
Abstract: Single-cell RNA sequencing (scRNA-seq) has revolutionised our ability to profile gene expression. However, short-read (SR) scRNAseq methodologies such as 10x are restricted to sequencing the 3’ or 5’ ends of transcripts, providing accurate gene expression but little information on the RNA isoforms expressed in each cell. Newly developed long-read (LR) scRNA-seq enables the quantification of RNA isoforms in in idual cells but LR scRNA-seq using the Oxford Nanopore platform has largely relied upon matched short-read data to identify cell barcodes and allow single cell analysis. Here we introduce BLAZE (Barcode identification from long-reads for AnalyZing single-cell gene Expression), which accurately and efficiently identifies 10x cell barcodes using only nanopore LR scRNA-seq data. We compared BLAZE to existing tools, including cell barcodes identified from matched SR scRNA-seq, on differentiating stem cells and 5 cancer cell lines. BLAZE outperforms existing tools and provides a more accurate representation of the cells present in LR scRNA-seq than using matched short-reads. BLAZE provides accurate cell barcodes over a wide range of experimental read depths and sequencing accuracies, while other methodologies commonly identify false-positive barcodes and cell clusters, disrupting biological interpretation of LR scRNA-seq results. In conclusion, BLAZE eliminates the requirement for matched SR scRNA-seq to interpret LR scRNA-seq, simplifying procedures and decreasing costs while also improving LR scRNA-seq results. BLAZE is compatible with downstream tools accepting a cell barcode whitelist file and is available at himlab/BLAZE .
Integrating Biomaterials and Stem Cells for Neural Regeneration
Publisher: Mary Ann Liebert Inc
Date: 02-2016
Abstract: The central nervous system has a limited capacity to regenerate, and thus, traumatic injuries or diseases often have devastating consequences. Therefore, there is a distinct need to develop alternative treatments that can achieve functional recovery without side effects currently observed with some pharmacological treatments. Combining biomaterials with pluripotent stem cells (PSCs), either embryonic or induced, has the potential to revolutionize the treatment of neurodegenerative diseases and traumatic injuries. Biomaterials can mimic the extracellular matrix and present a myriad of relevant biochemical cues through rational design or further functionalization. Biomaterials such as nanofibers and hydrogels, including self-assembling peptide (SAP) hydrogels can provide a superior cell culture environment. When these materials are then combined with PSCs, more accurate drug screening and disease modeling could be developed, and the generation of large number of cells with the appropriate phenotype can be achieved, for subsequent use in vitro. Biomaterials have also been shown to support endogenous cell growth after implantation, and, in particular, hydrogels and SAPs have effectively acted as cell delivery vehicles, increasing cell survival after transplantation. Few studies are yet to fully exploit the combination of PSCs and innovative biomaterials however, initial studies with neural stem cells, for ex le, are promising, and, hence, such a combination for use in vitro and in vivo is an exciting new direction for the field of neural regeneration.
Human stem cells harboring a suicide gene improve the safety and standardisation of neural transplants in Parkinsonian rats
Publisher: Springer Science and Business Media LLC
Date: 27-05-2021
DOI: 10.1038/S41467-021-23125-9
Abstract: Despite advancements in human pluripotent stem cells (hPSCs) differentiation protocols to generate appropriate neuronal progenitors suitable for transplantation in Parkinson’s disease, resultant grafts contain low proportions of dopamine neurons. Added to this is the tumorigenic risk associated with the potential presence of incompletely patterned, proliferative cells within grafts. Here, we utilised a hPSC line carrying a FailSafe TM suicide gene (thymidine kinase linked to cyclinD1) to selectively ablate proliferative cells in order to improve safety and purity of neural transplantation in a Parkinsonian model. The engineered FailSafe TM hPSCs demonstrated robust ventral midbrain specification in vitro, capable of forming neural grafts upon transplantation. Activation of the suicide gene within weeks after transplantation, by ganciclovir administration, resulted in significantly smaller grafts without affecting the total yield of dopamine neurons, their capacity to innervate the host brain or reverse motor deficits at six months in a rat Parkinsonian model. Within ganciclovir-treated grafts, other neuronal, glial and non-neural populations (including proliferative cells), were significantly reduced—cell types that may pose adverse or unknown influences on graft and host function. These findings demonstrate the capacity of a suicide gene-based system to improve both the standardisation and safety of hPSC-derived grafts in a rat model of Parkinsonism.
Biofunctionalisation of polymeric scaffolds for neural tissue engineering
Publisher: SAGE Publications
Date: 05-04-2012
Abstract: Patients who experience injury to the central or peripheral nervous systems invariably suffer from a range of dysfunctions due to the limited ability for repair and reconstruction of damaged neural tissue. Whilst some treatment strategies can provide symptomatic improvement of motor and cognitive function, they fail to repair the injured circuits and rarely offer long-term disease modification. To this end, the biological molecules, used in combination with neural tissue engineering scaffolds, may provide feasible means to repair damaged neural pathways. This review will focus on three promising classes of neural tissue engineering scaffolds, namely hydrogels, electrospun nanofibres and self-assembling peptides. Additionally, the importance and methods for presenting biologically relevant molecules such as, neurotrophins, extracellular matrix proteins and protein-derived sequences that promote neuronal survival, proliferation and neurite outgrowth into the lesion will be discussed.
Shear Containment of BDNF within Molecular Hydrogels Promotes Human Stem Cell Engraftment and Postinfarction Remodeling in Stroke
Publisher: Wiley
Date: 18-07-2018
Long-Distance Axonal Growth and Protracted Functional Maturation of Neurons Derived from Human Induced Pluripotent Stem Cells After Intracerebral Transplantation
Publisher: Oxford University Press (OUP)
Date: 15-02-2017
DOI: 10.1002/SCTM.16-0198
Abstract: The capacity for induced pluripotent stem (iPS) cells to be differentiated into a wide range of neural cell types makes them an attractive donor source for autologous neural transplantation therapies aimed at brain repair. Translation to the in vivo setting has been difficult, however, with mixed results in a wide variety of preclinical models of brain injury and limited information on the basic in vivo properties of neural grafts generated from human iPS cells. Here we have generated a human iPS cell line constitutively expressing green fluorescent protein as a basis to identify and characterize grafts resulting from transplantation of neural progenitors into the adult rat brain. The results show that the grafts contain a mix of neural cell types, at various stages of differentiation, including neurons that establish extensive patterns of axonal growth and progressively develop functional properties over the course of 1 year after implantation. These findings form an important basis for the design and interpretation of preclinical studies using human stem cells for functional circuit re-construction in animal models of brain injury.
Neuronal activity regulates expression of tyrosine hydroxylase in adult mouse substantia nigra pars compacta neurons
Publisher: Wiley
Date: 19-01-2011
DOI: 10.1111/J.1471-4159.2010.07151.X
Abstract: Striatal delivery of dopamine (DA) by midbrain substantia nigra pars compacta (SNc) neurons is vital for motor control and its depletion causes the motor symptoms of Parkinson's disease. While membrane potential changes or neuronal activity regulates tyrosine hydroxylase (TH, the rate limiting enzyme in catecholamine synthesis) expression in other catecholaminergic cells, it is not known whether the same occurs in adult SNc neurons. We administered drugs known to alter neuronal activity to mouse SNc DAergic neurons in various experimental preparations and measured changes in their TH expression. In cultured midbrain neurons, blockade of action potentials with 1 μM tetrodotoxin decreased TH expression beginning around 20 h later (as measured in real time by green fluorescent protein (GFP) expression driven off TH promoter activity). By contrast, partial blockade of small-conductance, Ca(2+) -activated potassium channels with 300 nM apamin increased TH mRNA and protein between 12 and 24 h later in slices of adult midbrain. Two-week infusions of 300 nM apamin directly to the adult mouse midbrain in vivo also increased TH expression in SNc neurons, measured immunohistochemically. Paradoxically, the number of TH immunoreactive (TH+) SNc neurons decreased in these animals. Similar in vivo infusions of drugs affecting other ion-channels and receptors (L-type voltage-activated Ca(2+) channels, GABA(A) receptors, high K(+) , DA receptors) also increased or decreased cellular TH immunoreactivity but decreased or increased, respectively, the number of TH+ cells in SNc. We conclude that in adult SNc neurons: (i) TH expression is activity-dependent and begins to change ∼20 h following sustained changes in neuronal activity (ii) ion-channels and receptors mediating cell-autonomous activity or synaptic input are equally potent in altering TH expression and (iii) activity-dependent changes in TH expression are balanced by opposing changes in the number of TH+ SNc cells.
A Small Synthetic Cripto Blocking Peptide Improves Neural Induction, Dopaminergic Differentiation, and Functional Integration of Mouse Embryonic Stem Cells in a Rat Model of Parkinson's Disease
Publisher: Oxford University Press (OUP)
Date: 16-07-2010
DOI: 10.1002/STEM.458
Abstract: Cripto is a glycosylphosphatidylinositol-anchored coreceptor that binds Nodal and the activin type I (ALK)-4 receptor, and is involved in cardiac differentiation of mouse embryonic stem cells (mESCs). Interestingly, genetic ablation of cripto results in increased neuralization and midbrain dopaminergic (DA) differentiation of mESCs, as well as improved DA cell replacement therapy (CRT) in a model of Parkinson's disease (PD). In this study, we developed a Cripto specific blocking tool that would mimic the deletion of cripto, but could be easily applied to embryonic stem cell (ESC) lines without the need of genetic manipulation. We thus screened a combinatorial peptide library and identified a tetrameric tripeptide, Cripto blocking peptide (BP), which prevents Cripto/ALK-4 receptor interaction and interferes with Cripto signaling. Cripto BP treatment favored neuroectoderm formation and promoted midbrain DA neuron differentiation of mESCs in vitro and in vivo. Remarkably, Cripto BP-treated ESCs, when transplanted into the striatum of PD rats, enhanced functional recovery and reduced tumor formation, mimicking the effect of genetic ablation of cripto. We therefore suggest that specific blockers such as Cripto BP may be used to improve the differentiation of ESC-derived DA neurons in vitro and their engraftment in vivo, bringing us closer towards an application of ESCs in CRT.
Macrophages and Microglia Produce Local Trophic Gradients That Stimulate Axonal Sprouting Toward but Not beyond the Wound Edge
Publisher: Elsevier BV
Date: 11-2002
Abstract: Following injury to the mammalian CNS, axons sprout in the vicinity of the wound margin. Growth then ceases and axons fail to cross the lesion site. In this study, using dopaminergic sprouting in the injured striatum as a model system, we have examined the relationship of periwound sprouting fibers to reactive glia and macrophages. In the first week after injury we find that sprouting fibers form intimate relationships with activated microglia as they traverse toward the wound edge. Once at the wound edge, complicated plexuses of fibers form around in idual macrophages. Axons, however, fail to grow further into the interior of the wound despite the presence of many macrophages in this location. We find that the expression of BDNF by activated microglia progressively increases as the wound edge is approached, while GDNF expression by macrophages is highest at the immediate wound margin. In contrast, the expression of both factors is substantially reduced within the macrophage-filled interior of the wound. Our data suggest that periwound sprouting fibers grow toward the wound margin along an increasing trophic gradient generated by progressively microglial and macrophage activation. Once at the wound edge, sprouting ceases over macrophages at the point of maximal neurotrophic factor expression and further axonal growth into the relatively poor trophic environment of the wound core fails to occur.
Chronic corticotropin‐releasing factor type 1 receptor antagonism with antalarmin regulates the dopaminergic system of Fawn‐Hooded rats
Publisher: Wiley
Date: 08-08-2005
DOI: 10.1111/J.1471-4159.2005.03300.X
Abstract: Corticotropin-releasing factor is a neuropeptide associated with the integration of physiological and behavioural responses to stress and also in the modulation of affective state and drug reward. The selective, centrally acting corticotropin-releasing factor type 1 receptor antagonist, antalarmin, is a potent anxiolytic and reduces volitional ethanol consumption in Fawn-Hooded rats. The efficacy of antalarmin to reduce ethanol consumption increased with time, suggestive of adaptation to reinforcement processes and goal-directed behaviour. The aim of the present study was to examine the effects of chronic antalarmin treatment on reward-related regions of Fawn-Hooded rat brain. Bi-daily antalarmin treatment (20 mg/kg, i.p.) for 10 days increased tyrosine hydroxylase messenger RNA expression throughout the ventral mesencephalon. Following chronic antalarmin the density of dopaminergic terminals within the basal ganglia and amygdaloid complex were reduced, as was dopamine transporter binding within the striatum. Receptor autoradiography indicated an up-regulation of dopamine D2, but no change in D1, binding in striatum, and Golgi-Cox analysis of striatal medium spiny neurones indicated that chronic antalarmin treatment increased spine density. Thus, chronic antalarmin treatment modulates dopaminergic pathways and implies that chronic treatment with drugs of this class may ultimately alter postsynaptic signaling mechanisms within the basal ganglia.
Isolation of LMX1a Ventral Midbrain Progenitors Improves the Safety and Predictability of Human Pluripotent Stem Cell-Derived Neural Transplants in Parkinsonian Disease
Publisher: Society for Neuroscience
Date: 22-10-2019
DOI: 10.1523/JNEUROSCI.1160-19.2019
Abstract: Human pluripotent stem cells (hPSCs) are a promising resource for the replacement of degenerated ventral midbrain dopaminergic (vmDA) neurons in Parkinson's disease. Despite recent advances in protocols for the in vitro generation of vmDA neurons, the asynchronous and heterogeneous nature of the differentiations results in transplants of surprisingly low vmDA neuron purity. As the field advances toward the clinic, it will be optimal, if not essential, to remove poorly specified and potentially proliferative cells from donor preparations to ensure safety and predictable efficacy. Here, we use two novel hPSC knock-in reporter lines expressing GFP under the LMX1A and PITX3 promoters, to selectively isolate vm progenitors and DA precursors, respectively. For each cell line, unsorted, GFP + , and GFP − cells were transplanted into male or female Parkinsonian rodents. Only rats receiving unsorted cells, LMX1A-eGFP + , or PITX3-eGFP − cell grafts showed improved motor function over 6 months. Postmortem analysis revealed small grafts from PITX3-eGFP + cells, suggesting that these DA precursors were not compatible with cell survival and integration. In contrast, LMX1A-eGFP + grafts were highly enriched for vmDA neurons, and importantly excluded expansive proliferative populations and serotonergic neurons. These LMX1A-eGFP + progenitor grafts accelerated behavioral recovery and innervated developmentally appropriate forebrain targets, whereas LMX1A-eGFP − cell grafts failed to restore motor deficits, supported by increased fiber growth into nondopaminergic target nuclei. This is the first study to use an hPSC-derived reporter line to purify vm progenitors, resulting in improved safety, predictability of the graft composition, and enhanced motor function. SIGNIFICANCE STATEMENT Clinical trials have shown functional integration of transplanted fetal-derived dopamine progenitors in Parkinson's disease. Human pluripotent stem cell (hPSC)-derived midbrain progenitors are now being tested as an alternative cell source however, despite current differentiation protocols generating % correctly specified cells for implantation, resultant grafts contain a small fraction of dopamine neurons. Cell-sorting approaches, to select for correctly patterned cells before implantation, are being explored yet have been suboptimal to date. This study provides the first evidence of using 2 hPSC reporter lines (LMX1A-GFP and PITX3-GFP) to isolate correctly specified cells for transplantation. We show LMX1A-GFP + , but not PITX3-GFP + , cell grafts are more predictable, with smaller grafts, enriched in dopamine neurons, showing appropriate integration and accelerated functional recovery in Parkinsonian rats.
Cripto as a Target for Improving Embryonic Stem Cell–Based Therapy in Parkinson's Disease
Publisher: Oxford University Press (OUP)
Date: 04-2005
DOI: 10.1634/STEMCELLS.2004-0294
Abstract: Embryonic stem (ES) cells have been suggested as candidate therapeutic tools for cell replacement therapy in neurodegenerative disorders. However, limitations for the use of these cells lie in our restricted knowledge of the molecular mechanisms involved in their specialized differentiation and in the risk of tumor formation. Recent findings suggest that the EGF-CFC protein Cripto is a key player in the signaling pathways controlling neural induction in ES cells. Here we show that in vitro differentiation of Cripto(-/-) ES cells results in increased dopaminergic differentiation and that, upon transplantation into Parkinsonian rats, they result in behavioral and anatomical recovery with no tumor formation. The use of knockout ES cells that can generate dopamine cells while eliminating tumor risk holds enormous potential for cell replacement therapy in Parkinson's disease.
Inhibition of JNK increases survival of transplanted dopamine neurons in Parkinsonian rats
Publisher: Springer Science and Business Media LLC
Date: 21-07-2006
Cell intrinsic and extrinsic factors contribute to enhance neural circuit reconstruction following transplantation in Parkinsonian mice
Publisher: Wiley
Date: 11-2013
Over-expression of meteorin drives gliogenesis following striatal injury
Publisher: Frontiers Media SA
Date: 05-07-2016
Wnt/β-Catenin Signaling Blockade Promotes Neuronal Induction and Dopaminergic Differentiation in Embryonic Stem Cells
Publisher: Oxford University Press (OUP)
Date: 09-2009
DOI: 10.1002/STEM.210
Abstract: Embryonic stem cells (ESCs) represent not only a promising source of cells for cell replacement therapy, but also a tool to study the molecular mechanisms underlying cellular signaling and dopaminergic (DA) neuron development. One of the main regulators of DA neuron development is Wnt signaling. Here we used mouse ESCs (mESCs) lacking Wnt1 or the low-density lipoprotein receptor-related protein 6 (LRP6) to decipher the action of Wnt/β-catenin signaling on DA neuron development in mESCs. We provide evidence that the absence of LRP6 abrogates responsiveness of mESCs to Wnt ligand stimulation. Using two differentiation protocols, we show that the loss of Wnt1 or LRP6 increases neuroectodermal differentiation and the number of mESC-derived DA neurons. These effects were similar to those observed following treatment of mESCs with the Wnt/β-catenin pathway inhibitor Dickkopf1 (Dkk1). Combined, our results show that decreases in Wnt/β-catenin signaling enhance neuronal and DA differentiation of mESCs. These findings suggest that: 1) Wnt1 or LRP6 are not strictly required for the DA differentiation of mESCs in vitro, 2) the levels of morphogens and their activity in ESC cultures need to be optimized to improve DA differentiation, and 3) by enhancing the differentiation and number of ESC-derived DA neurons with Dkk1, the application of ESCs for cell replacement therapy in Parkinson's disease may be improved. Disclosure of potential conflicts of interest is found at the end of this article.
Simple Complexity: Incorporating Bioinspired Delivery Machinery within Self-Assembled Peptide Biogels
Publisher: MDPI AG
Date: 06-03-2023
DOI: 10.3390/GELS9030199
Abstract: Bioinspired self-assembly is a bottom-up strategy enabling biologically sophisticated nanostructured biogels that can mimic natural tissue. Self-assembling peptides (SAPs), carefully designed, form signal-rich supramolecular nanostructures that intertwine to form a hydrogel material that can be used for a range of cell and tissue engineering scaffolds. Using the tools of nature, they are a versatile framework for the supply and presentation of important biological factors. Recent developments have shown promise for many applications such as therapeutic gene, drug and cell delivery and yet are stable enough for large-scale tissue engineering. This is due to their excellent programmability—features can be incorporated for innate biocompatibility, biodegradability, synthetic feasibility, biological functionality and responsiveness to external stimuli. SAPs can be used independently or combined with other (macro)molecules to recapitulate surprisingly complex biological functions in a simple framework. It is easy to accomplish localized delivery, since they can be injected and can deliver targeted and sustained effects. In this review, we discuss the categories of SAPs, applications for gene and drug delivery, and their inherent design challenges. We highlight selected applications from the literature and make suggestions to advance the field with SAPs as a simple, yet smart delivery platform for emerging BioMedTech applications.
Peptide-Based Scaffolds Support Human Cortical Progenitor Graft Integration to Reduce Atrophy and Promote Functional Repair in a Model of Stroke
Publisher: Elsevier BV
Date: 08-2017
DOI: 10.1016/J.CELREP.2017.07.069
Abstract: Stem cell transplants offer significant hope for brain repair following ischemic damage. Pre-clinical work suggests that therapeutic mechanisms may be multi-faceted, incorporating bone-fide circuit reconstruction by transplanted neurons, but also protection/regeneration of host circuitry. Here, we engineered hydrogel scaffolds to form "bio-bridges" within the necrotic lesion cavity, providing physical and trophic support to transplanted human embryonic stem cell-derived cortical progenitors, as well as residual host neurons. Scaffolds were fabricated by the self-assembly of peptides for a laminin-derived epitope (IKVAV), thereby mimicking the brain's major extracellular protein. Following focal ischemia in rats, scaffold-supported cell transplants induced progressive motor improvements over 9 months, compared to cell- or scaffold-only implants. These grafts were larger, exhibited greater neuronal differentiation, and showed enhanced electrophysiological properties reflective of mature, integrated neurons. Varying graft timing post-injury enabled us to attribute repair to both neuroprotection and circuit replacement. These findings highlight strategies to improve the efficiency of stem cell grafts for brain repair.
Local Injection of Endothelin-1 in the Early Neonatal Rat Brain Models Ischemic Damage Associated with Motor Impairment and Diffuse Loss in Brain Volume
Publisher: Elsevier BV
Date: 11-2018
DOI: 10.1016/J.NEUROSCIENCE.2018.09.043
Abstract: Cerebral palsy is an irreversible movement disorder resulting from cerebral damage sustained during prenatal or neonatal brain development. As survival outcomes for preterm injury improve, there is increasing need to model ischemic injury at earlier neonatal time-points to better understand the subsequent pathological consequences. Here we demonstrate a novel neonatal ischemic model using focal administration of the potent vasoconstrictor peptide, endothelin-1 (ET-1), in newborn rats. The functional and histopathological outcomes compare favourably to those reported following the widely used hypoxic ischemia (HI) model. These include a robust motor deficit sustained into adulthood and recapitulation of hallmark features of preterm human brain injury, including atrophy of subcortical white matter and periventricular fiber bundles. Compared to procedures involving carotid artery manipulation and periods of hypoxia, the ET-1 ischemia model represents a rapid and technically simplified model more amenable to larger cohorts and with the potential to direct the locus of ischemic damage to specific brain areas.
Neurons derived from human embryonic stem cells extend long-distance axonal projections through growth along host white matter tracts after intra-cerebral transplantation
Publisher: Frontiers Media SA
Date: 2012
The application of human pluripotent stem cells to model the neuronal and glial components of neurodevelopmental disorders
Publisher: Springer Science and Business Media LLC
Date: 27-08-2019
DOI: 10.1038/S41380-019-0495-0
Abstract: Cellular models of neurodevelopmental disorders provide a valuable experimental system to uncover disease mechanisms and novel therapeutic strategies. The ability of induced pluripotent stem cells (iPSCs) to generate erse brain cell types offers great potential to model several neurodevelopmental disorders. Further patient-derived iPSCs have the unique genetic and molecular signature of the affected in iduals, which allows researchers to address limitations of transgenic behavioural models, as well as generate hypothesis-driven models to study disorder-relevant phenotypes at a cellular level. In this article, we review the extant literature that has used iPSC-based modelling to understand the neuronal and glial contributions to neurodevelopmental disorders including autism spectrum disorder (ASD), Rett syndrome, bipolar disorder (BP), and schizophrenia. For instance, several molecular candidates have been shown to influence cellular phenotypes in three-dimensional iPSC-based models of ASD patients. Delays in differentiation of astrocytes and morphological changes of neurons are associated with Rett syndrome. In the case of bipolar disorders and schizophrenia, patient-derived models helped to identify cellular phenotypes associated with neuronal deficits (e.g., excitability) and mutation-specific abnormalities in oligodendrocytes (e.g., CSPG4). Further we provide a critical review of the current limitations of this field and provide methodological suggestions to enhance future modelling efforts of neurodevelopmental disorders. Future developments in experimental design and methodology of disease modelling represent an exciting new avenue relevant to neurodevelopmental disorders.
Specification of murine ground state pluripotent stem cells to regional neuronal populations
Publisher: Springer Science and Business Media LLC
Date: 22-11-2017
DOI: 10.1038/S41598-017-16248-X
Abstract: Pluripotent stem cells (PSCs) are a valuable tool for interrogating development, disease modelling, drug discovery and transplantation. Despite the burgeoned capability to fate restrict human PSCs to specific neural lineages, comparative protocols for mouse PSCs have not similarly advanced. Mouse protocols fail to recapitulate neural development, consequently yielding highly heterogeneous populations, yet mouse PSCs remain a valuable scientific tool as differentiation is rapid, cost effective and an extensive repertoire of transgenic lines provides an invaluable resource for understanding biology. Here we developed protocols for neural fate restriction of mouse PSCs, using knowledge of embryonic development and recent progress with human equivalents. These methodologies rely upon naïve ground-state PSCs temporarily transitioning through LIF-responsive stage prior to neural induction and rapid exposure to regional morphogens. Neural subtypes generated included those of the dorsal forebrain, ventral forebrain, ventral midbrain and hindbrain. This rapid specification, without feeder layers or embryoid-body formation, resulted in high proportions of correctly specified progenitors and neurons with robust reproducibility. These generated neural progenitors/neurons will provide a valuable resource to further understand development, as well disorders affecting specific neuronal subpopulations.
Chondroitinase improves midbrain pathway reconstruction by transplanted dopamine progenitors in Parkinsonian mice
Publisher: Elsevier BV
Date: 11-2015
DOI: 10.1016/J.MCN.2015.10.002
Abstract: Within the adult central nervous system the lack of guidance cues together with the presence of inhibitory molecules produces an environment that is restrictive to axonal growth following injury. Consequently, while clinical trials in Parkinson's disease (PD) patients have demonstrated the capacity of fetal-derived dopamine neurons to survive, integrate and alleviate symptoms, the non-permissive host environment has contributed to the incomplete re-innervation of the target tissue by ectopic grafts, and even more noticeable, the poor reconstruction of the midbrain dopamine pathways following homotopic midbrain grafting. One such inhibitory molecule is the chondroitin sulfate proteoglycan (CSPG), a protein that has been shown to impede axonal growth during development and after injury. Digestion of CSPGs, by delivery of the bacterial enzyme chondroitinase ABC (ChABC), can improve axonal regrowth following a number of neural injuries. Here we examined whether ChABC could similarly improve axonal growth of transplanted dopamine neurons in an animal model of PD. Acute delivery of ChABC, into the medial forebrain bundle, degraded CSPGs along the nigrostriatal pathway. Simultaneous homotopic transplantation of dopaminergic progenitors, into the ventral midbrain of ChABC treated PD mice, had no effect on graft survival but resulted in enhanced axonal growth along the nigrostriatal pathway and reinnervation of the striatum, compared to control grafted mice. This study demonstrates that removal of axonal growth inhibitory molecules could significantly enhance dopaminergic graft integration, thereby holding implications for future approaches in the development of cell replacement therapies for Parkinsonian patients.
Chronic cocaine administration reduces striatal dopamine terminal density and striatal dopamine release which leads to drug-seeking behaviour
Publisher: Elsevier BV
Date: 02-2011
DOI: 10.1016/J.NEUROSCIENCE.2010.11.055
Abstract: Drug addiction is associated with altered dopamine (DA) neurotransmission in the basal ganglia. We have previously shown that chronic stimulation of the dopamine D2 receptor (D(2)R) with cocaine results in reduced striatal DA terminal density. The aims of this study were to establish whether this reduction in DA terminal density results in reduced striatal DA release and increased cocaine-seeking behaviour and whether D(2)R antagonism can restore the cocaine-induced alterations in DA neurotransmission and drug-seeking behaviour. Rats were housed in idually and either control, cocaine, haloperidol (D(2)R antagonist), or cocaine and haloperidol was administered in the drinking water for 16 weeks. Chronic cocaine treatment, which reduced striatal DA terminal density by 20%, resulted in a reduction in basal (-34%) and cocaine-evoked (-33%) striatal DA release and increased cocaine-seeking behaviour. These cocaine-mediated effects on striatal DA terminal density, DA release and drug-seeking could be prevented by co-administration with haloperidol. Basal and cocaine-evoked DA release in the striatum directly correlated with DA terminal density and with preference for cocaine. We conclude that striatal DA terminal density and DA release is an important factor in maintaining drug preference and should be considered as a factor in drug-seeking behaviour and relapse.
Mice Lacking the α4 Nicotinic Receptor Subunit Fail to Modulate Dopaminergic Neuronal Arbors and Possess Impaired Dopamine Transporter Function
Publisher: American Society for Pharmacology & Experimental Therapeutics (ASPET)
Date: 02-08-2005
Abstract: Neuronal nicotinic acetylcholine receptors (nAChRs) at presynaptic sites can modulate dopaminergic synaptic transmission by regulating dopamine (DA) release and uptake. Dopaminergic transmission in nigrostriatal and mesolimbic pathways is vital for the coordination of movement and is associated with learning and behavioral reinforcement. We reported recently that the D2 DA receptor plays a central role in regulating the arbor size of substantia nigra dopaminergic neurons. Given the known effects of nAChRs on dopaminergic neurotransmission, we assessed the ability of the alpha4 nAChR subunit to regulate arbor size of dopaminergic neurons by comparing responses of wild-type and alpha4 nAChR subunit knockout [alpha4(-/-)] mice to long-term exposure to cocaine, hetamine, nicotine, and haloperidol, and after substantia nigra neurotoxic lesioning. We found that dopaminergic neurons in adult drug-naive alpha4(-/-) mice had significantly larger terminal arbors, and despite normal short-term behavioral responses to drugs acting on pre- and postsynaptic D2 DA receptors, they were unable to modulate their terminal arbor in response to pharmacological manipulation or after lesioning. In addition, although synaptosome DA uptake studies showed that the interaction of the D2 DA receptor and the dopamine transporter (DAT) was preserved in alpha4(-/-) mice, DAT function was found to be impaired. These findings suggest that the alpha4 subunit of the nAChR is an independent regulator of terminal arbor size of nigrostriatal dopaminergic neurons and that reduced functionality of presynaptic DAT may contribute to this effect by impairing DA uptake.
A Fully Human Inhibitory Monoclonal Antibody to the Wnt Receptor RYK
Publisher: Public Library of Science (PLoS)
Date: 18-09-2013
Cellular up-regulation of Nedd4 family interacting protein 1 (Ndfip1) using low levels of bioactive cobalt complexes
Publisher: Elsevier BV
Date: 03-2011
Dopamine D2 receptor knockout mice develop features of Parkinson disease
Publisher: Wiley
Date: 10-2009
DOI: 10.1002/ANA.21716
Abstract: This study questions whether increased dopamine (DA) turnover in nigral neurons leads to formation of Lewy bodies (LBs), the characteristic alpha-synuclein-containing cytoplasmic inclusion of Parkinson disease (PD). Mice with targeted deletion of the dopamine D(2) receptor gene (D(2)R[-/-]) have higher striatal and nigral dopamine turnover and elevated oxidative stress. These mice were examined for evidence of histological, biochemical, and gene expression changes consistent with a synucleinopathy. LB-like cytoplasmic inclusions containing alpha-synuclein and ubiquitin were present in substantia nigra pars compacta (SNpc) neurons of older D(2)R(-/-) mice, and were also occasionally seen in aged wild-type mice. These inclusions displaced the nucleus of affected cells and were eosinophilic. Diffuse cytosolic alpha-synuclein immunoreactivity in SNpc neurons increased with age in both wild-type and D(2)R(-/-) mice, most likely because of redistribution of alpha-synuclein from striatal terminals to SNpc cell bodies. Gene and protein expression studies indicated endoplasmic reticulum (ER) stress and changes in trafficking and autophagic pathways in D(2)R(-/-) SNpc. These changes were accompanied by a loss of DA terminals in the dorsal striatum, although there was no evidence of progressive cell death in the SNpc. Increased sprouting and DA turnover, as observed in PD and D(2)R(-/-) mice, augments LB-like inclusions and axonal degeneration of dopaminergic neurons. These changes are associated with ER stress and autophagy.
Functional Integration of Grafted Neural Stem Cell-Derived Dopaminergic Neurons Monitored by Optogenetics in an In Vitro Parkinson Model
Publisher: Public Library of Science (PLoS)
Date: 04-03-2011
Birth dating of midbrain dopamine neurons identifies A9 enriched tissue for transplantation into Parkinsonian mice
Publisher: Elsevier BV
Date: 07-2012
DOI: 10.1016/J.EXPNEUROL.2012.04.002
Abstract: Clinical trials have provided proof of principle that new dopamine neurons isolated from the developing ventral midbrain and transplanted into the denervated striatum can functionally integrate and alleviate symptoms in Parkinson's disease patients. However, extensive variability across patients has been observed, ranging from long-term motor improvement to the absence of symptomatic relief and development of dyskinesias. Heterogeneity of the donor tissue is likely to be a contributing factor in the variable outcomes. Dissections of ventral midbrain used for transplantation will variously contain progenitors for different dopamine neuron subtypes as well as different neurotransmitter phenotypes. The overall impact of the resulting graft will be determined by the functional contribution from these different cell types. The A9 substantia nigra pars compacta dopamine neurons, for ex le, are known to be particularly important for motor recovery in animal models. Serotonergic neurons, on the other hand, have been implicated in unwanted dyskinesias. Currently little knowledge exists on how variables such as donor age, which have not been controlled for in clinical trials, will impact on the final neuronal composition of fetal grafts. Here we performed a birth dating study to identify the time-course of neurogenesis within the various ventral midbrain dopamine subpopulations in an effort to identify A9-enriched donor tissue for transplantation. The results show that A9 neurons precede the birth of A10 ventral tegmental area dopamine neurons. Subsequent grafting of younger ventral midbrain donor tissue revealed significantly larger grafts containing more mitotic dopamine neuroblasts compared to older donor grafts. These grafts were enriched with A9 neurons and showed significantly greater innervation of the target dorso-lateral striatum and DA release. Younger donor grafts also contained significantly less serotonergic neurons. These findings demonstrate the importance of standardized methods to improve cell therapy for Parkinson's disease and have significant implications for the generation and selectivity of dopamine neurons from stem cell based sources.
Biomaterials for brain tissue engineering
Publisher: CSIRO Publishing
Date: 2010
DOI: 10.1071/CH10159
Abstract: Neurological disorders such as traumatic brain injuries or stroke result in neuronal loss and disruption of the brain parenchyma. Current treatment strategies are limited in that they can only mitigate the degeneration process or alleviate the symptoms but do not reverse the condition. In contrast, regenerative cell-based therapies offer long-term hope for many patients. Bioactive scaffolds are likely to reinforce the success of cell replacement therapies by providing a microenvironment that facilitates the survival, proliferation, differentiation, and connectivity of transplanted and/or endogenous cells. This Review outlines various biomaterials (including hydrogels, self-assembling peptides, and electrospun nanofibres) that have been investigated for the repair of brain tissue, and discusses strategies for the immobilization of biomolecules. An overview of the potential clinical applications of such scaffolds in neurodegenerative diseases is also provided.
Wnt5a regulates midbrain dopaminergic axon growth and guidance
Publisher: Public Library of Science (PLoS)
Date: 31-03-2011
VIRAL DELIVERY OF GDNF PROMOTES FUNCTIONAL INTEGRATION OF HUMAN STEM CELL GRAFTS IN PARKINSON’S DISEASE
Publisher: Cold Spring Harbor Laboratory
Date: 10-12-2019
DOI: 10.1101/870725
Abstract: The derivation of neurotransmitter and region-specific neuronal populations from human pluripotent stem cells (PSC) provides impetus for advancing cell therapies into the clinic. At the forefront is our ability to generate ventral midbrain (VM) dopaminergic (DA) progenitors, suitable for transplantation in Parkinson’s disease (PD). Pre-clinical studies, however, have highlighted the low proportion of DA neurons within these grafts and their inferior plasticity by comparison to human fetal donor transplants. Here we sought to examine whether modification of the host environment, through viral delivery of a developmentally critical molecule, glial cell line-derived neurotrophic factor (GDNF), could improve graft survival, integration and function in Parkinsonian rodents. Utilising LMX1A- and PITX3-GFP hPSC reporter lines, we tracked the response of DA progenitors implanted into either a GDNF-rich environment, or in a second group, after a 3-week delay in onset of exposure. We found that early exposure of the graft to GDNF promoted survival of DA and non-DA cells, leading to enhanced motor recovery in PD rats. Delayed overexpression of intrastriatal GDNF also promoted motor recovery in transplanted rats, through alternate selective mechanisms including enhanced A9/A10 specification, increased DA graft plasticity, greater activation of striatal neurons and elevated DA metabolism. Lastly, transcriptional profiling of the grafts highlighted novel genes underpinning these changes. Collectively these results demonstrate the potential of targeted neurotrophic gene therapy strategies to improve human PSC graft outcomes.
Tuning the amino acid sequence of minimalist peptides to present biological signals via charge neutralised self assembly
Publisher: Royal Society of Chemistry (RSC)
Date: 2013
DOI: 10.1039/C3SM27758E
Promoting engraftment of transplanted neural stem cells/progenitors using biofunctionalised electrospun scaffolds
Publisher: Elsevier BV
Date: 12-2012
DOI: 10.1016/J.BIOMATERIALS.2012.09.013
Abstract: With the brain's limited capacity for repair, new and innovative approaches are required to promote regeneration. While neural transplantation for a number of neural disease/injuries have been demonstrated, major limitations in the field include poor cell survival and integration. This, in part, is due to the non-conducive environment of the adult brain, failing to provide adequate chemical and physical support for new neurons. Here we examine the capacity of fibrous poly ε-caprolactone (PCL) scaffolds, biofunctionalised with immobilised glial cell-derived neurotrophic factor (GDNF), to influence primary cortical neural stem cells rogenitors in vitro and enhance integration of these cells following transplantation into the brain parenchyma. Immobilisation of GDNF was confirmed prior to in vitro culturing and at 28 days after implantation into the brain, demonstrating long-term delivery of the protein. In vitro, we demonstrate that PCL with immobilised GDNF (iGDNF) significantly enhances cell viability and neural stem cell rogenitor proliferation compared to conventional 2-dimensional cultureware. Upon implantation, PCL scaffolds including iGDNF enhanced the survival, proliferation, migration, and neurite growth of transplanted cortical cells, whilst suppressing inflammatory reactive astroglia.
Organized Development from Human Embryonic Stem Cells after Injection into Immunodeficient Mice
Publisher: Mary Ann Liebert Inc
Date: 08-2004
Abstract: Information concerning the development and differentiation of human embryonic stem (hES) cells in vivo is limited. The present study has focused on the in vivo outcome and differentiation of the hESC line HS181, after injection into SCID/beige mice. hES cell-derived teratomas were explored using histological evaluation and by the identification of markers for differentiated cells and tissues. The analyses identified predominant differentiation along a neuronal lineage, the formation of bone/cartilage and epithelia. Fluorescent in situ hybridization (FISH) analysis with a human-specific probe showed the teratomas to be mainly of human origin, with the most organized areas being exclusively human. Importantly, the study revealed interactions between mouse and human tissues, most notably in the formation of vessels. Both mouse and human cells contributed to specific microstructures in which mouse cells could be observed to take on the appropriate histiotypic appearance. Hence, HS181 cells were able to develop into defined mature tissues, supporting the relevant use of this hES cells model for studies of early human development, given the use of appropriate controls for host contribution. Although extensive mitotic activity implicated progenitor cell activity, no detectable multipotent or malignant areas were observed during the observation period. Persisting undifferentiated hESC were not detected.
In vivo assessment of grafted cortical neural progenitor cells and host response to functionalized self-assembling peptide hydrogels and the implications for tissue repair
Publisher: Royal Society of Chemistry (RSC)
Date: 2014
DOI: 10.1039/C4TB01391C
Abstract: Functionalized N -fluorenylmethyloxycarbonyl self-assembling peptides are biocompatible in vivo , demonstrating their utility as a cell delivery vehicle for tissue engineering.
Efficient expansion and dopaminergic differentiation of human fetal ventral midbrain neural stem cells by midbrain morphogens
Publisher: Elsevier BV
Date: 2013
DOI: 10.1016/J.NBD.2012.08.006
Abstract: Human fetal midbrain tissue grafting has provided proof-of-concept for dopamine cell replacement therapy (CRT) in Parkinson's disease (PD). However, limited tissue availability has hindered the development and widespread use of this experimental therapy. Here we present a method for generating large numbers of midbrain dopaminergic (DA) neurons based on expanding and differentiating neural stem rogenitor cells present in the human ventral midbrain (hVM) tissue. Our results show that hVM neurospheres (hVMN) with low cell numbers, unlike their rodent counterparts, expand the total number of cells 3-fold, whilst retaining their capacity to differentiate into midbrain DA neurons. Moreover, Wnt5a promoted DA differentiation of expanded cells resulting in improved morphological maturation, midbrain DA marker expression, DA release and electrophysiological properties. This method results in cell preparations that, after expansion and differentiation, can contain 6-fold more midbrain DA neurons than the starting VM preparation. Thus, our results provide evidence that by improving expansion and differentiation of progenitors present in the hVM it is possible to greatly enrich cell preparations for DA neurons. This method could substantially reduce the amount of human fetal midbrain tissue necessary for CRT in patients with PD, which could have major implications for the widespread adoption of this approach.
Transcriptome analysis reveals transmembrane targets on transplantable midbrain dopamine progenitors
Publisher: Proceedings of the National Academy of Sciences
Date: 09-03-2015
Abstract: An important challenge for improving cell-based approaches for Parkinson’s disease is the development of techniques that facilitate greater standardization of the donor material. This report describes the enrichment of transplantable progenitors for dopamine neurons from the ventral mesencephalon based on targeting of transmembrane proteins. It is an important step toward the development of clinically relevant techniques that allow for greater standardization of cell preparations used in transplantation and potentially, more predictable clinical outcomes. The findings are highly relevant for current efforts to develop stem cell-based therapies for Parkinson’s disease, where current techniques yield mixed cell populations that may contain unwanted cell types and thus, would benefit from a cell selection step prior to grafting.
Inhibition of amyloid beta toxicity in zebrafish with a chaperone-gold nanoparticle dual strategy
Publisher: Springer Science and Business Media LLC
Date: 22-08-2019
DOI: 10.1038/S41467-019-11762-0
Abstract: Alzheimer’s disease (AD) is the most prevalent form of neurodegenerative disorders, yet no major breakthroughs have been made in AD human trials and the disease remains a paramount challenge and a stigma in medicine. Here we eliminate the toxicity of amyloid beta (Aβ) in a facile, high-throughput zebrafish ( Danio rerio ) model using casein coated-gold nanoparticles (βCas AuNPs). βCas AuNPs in systemic circulation translocate across the blood brain barrier of zebrafish larvae and sequester intracerebral Aβ 42 and its elicited toxicity in a nonspecific, chaperone-like manner. This is evidenced by behavioral pathology, reactive oxygen species and neuronal dysfunction biomarkers assays, complemented by brain histology and inductively coupled plasma-mass spectroscopy. We further demonstrate the capacity of βCas AuNPs in recovering the mobility and cognitive function of adult zebrafish exposed to Aβ. This potent, safe-to-use, and easy-to-apply nanomedicine may find broad use for eradicating toxic amyloid proteins implicated in a range of human diseases.
An Efficient Method for the Derivation of Mouse Embryonic Stem Cells
Publisher: Oxford University Press (OUP)
Date: 04-2006
DOI: 10.1634/STEMCELLS.2005-0444
Abstract: Mouse embryonic stem cells (mESCs) represent a unique tool for many researchers however, the process of ESC derivation is often very inefficient and requires high specialization, training, and expertise. To circumvent these limitations, we aimed to develop a simple and efficient protocol based on the use of commercially available products. Here, we present an optimized protocol that we successfully applied to derive ESCs from several knockout mouse strains (Wnt-1, Wnt-5a, Lrp6, and parkin) with 50%–75% efficiency. The methodology is based on the use of mouse embryonic fibroblast feeders, knockout serum replacement (SR), and minimal handling of the blastocyst. In this protocol, all centrifugation steps (as well as the use of trypsin inhibitor) were avoided and replaced by an ESC medium containing fetal calf serum (FCS) after the trypsinizations. We define the potential advantages and disadvantages of using SR and FCS in in idual steps of the protocol. We also characterize the ESCs for the expression of ESC markers by immunohistochemistry, Western blot, and a stem cell focused microarray. In summary, we provide a simplified and improved protocol to derive mESCs that can be useful for laboratories aiming to isolate transgenic mESCs for the first time.
Genome-wide binding and mechanistic analyses of Smchd1-mediated epigenetic regulation
Publisher: Proceedings of the National Academy of Sciences
Date: 19-06-2015
Abstract: Structural maintenance of chromosomes flexible hinge domain containing 1 (Smchd1) is a protein that plays an important role in maintaining gene silencing in many biological circumstances, including facioscapulohumeral muscular dystrophy however, how it brings about gene silencing is unknown. Understanding the molecular mechanism by which Smchd1 contributes to stable transcriptional silencing is critical to appreciate how it functions in normal biology and when it is mutated in facioscapulohumeral muscular dystrophy. This study reveals, for the first time to our knowledge, where Smchd1 binds genome-wide, its hitherto unappreciated functional interaction with chromatin organizer CCCTC-binding factor in gene regulation, and which part of the protein is required for chromatin binding. These data lead to a new model of Smchd1 function, where it directly binds DNA to mediate 3D chromatin architecture.
Development of an In Vitro Model to Evaluate the Regenerative Capacity of Adult Brain-Derived Tyrosine Hydroxylase-Expressing Dopaminergic Neurons
Publisher: Springer Science and Business Media LLC
Date: 06-03-2011
DOI: 10.1007/S11064-011-0435-0
Abstract: The loss of nigral dopaminergic (DA) neurons is the disease-defining pathological change responsible for progressive motor dysfunction in Parkinson's disease. In this study, we sought to establish a culture method for adult rat tyrosine hydroxylase (TH)-immunoreactive DA neurons. In this context, we investigated the role of fibroblast growth factor 2 (FGF2), brain-derived neurotrophic factor (BDNF), transforming growth factor-β3 (TGF-β3), glial-derived neurotrophic factor (GDNF) and dibutyryl-cyclic AMP (dbcAMP) in these cultures. Culturing in the presence of FGF2, BDNF and GDNF enhanced the survival of DA neurons by 15-fold and promoted neurite growth. In contrast, dbcAMP promoted neurite growth in all neurons but did not enhance DA cell survival. This study demonstrates that long-term cultures of DA neurons can be established from the mature rat brain and that survival and regeneration of DA neurons can be manipulated by epigenetic factors such as growth factors and intracellular cAMP pathways.
Generation of four iPSC lines from peripheral blood mononuclear cells (PBMCs) of an attention deficit hyperactivity disorder (ADHD) individual and a healthy sibling in an Australia-Caucasian family
Publisher: Elsevier BV
Date: 2019
DOI: 10.1016/J.SCR.2018.11.014
Abstract: Peripheral blood mononuclear cells were donated by a male teenager with clinically diagnosed attention deficit hyperactivity disorder (ADHD) under the Diagnostic and Statistical Manual of Mental Disorders IV criteria and his unaffected male sibling. Induced pluripotent stem cells were developed using integration-free Sendai Reprogramming factors containing OCT4, SOX2, KLF4, and c-MYC. All four iPSC lines displayed pluripotent cell morphology, pluripotency-associated factors at the DNA and protein level, alkaline phosphatase enzymatic activity and a male karyotype of 46, XY. All lines had capacity for in vitro differentiation into all the three germ layers. All were negative for Mycoplasma.
Diverse roles for Wnt7a in ventral midbrain neurogenesis and dopaminergic axon morphogenesis
Publisher: Mary Ann Liebert Inc
Date: 09-2014
Abstract: During development of the central nervous system, trophic, together with genetic, cues dictate the balance between cellular proliferation and differentiation. Subsequent to the birth of new neurons, additional intrinsic and extrinsic signals regulate the connectivity of these cells. While a number of regulators of ventral midbrain (VM) neurogenesis and dopaminergic (DA) axon guidance are known, we identify a number of novel roles for the secreted glycoprotein, Wnt7a, in this context. We demonstrate a temporal and spatial expression of Wnt7a in the VM, indicative of roles in neurogenesis, differentiation, and axonal growth and guidance. In primary VM cultures, and validated in Wnt7a-deficient mice, we show that the early expression within the VM is important for regulating VM progenitor proliferation, cell cycle progression, and cell survival, thereby dictating the number of midbrain Nurr1 precursors and DA neurons. During early development of the midbrain DA pathways, Wnt7a promotes axonal elongation and repels DA neurites out of the midbrain. Later, Wnt7a expression in the VM midline suggests a role in preventing axonal crossing while expression in regions flanking the medial forebrain bundle (thalamus and hypothalamus) ensured appropriate trajectory of DA axons en route to their forebrain targets. We show that the effects of Wnt7a in VM development are mediated, at least in part, by the β-catenin/canonical pathways. Together, these findings identify Wnt7a as a new regulator of VM neurogenesis and DA axon growth and guidance.
Stem-Cell-Based Strategies for the Treatment of Parkinson’s Disease
Publisher: S. Karger AG
Date: 2007
DOI: 10.1159/000101892
Abstract: i Background: /i Cell transplantation to replace lost neurons in neurodegenerative diseases such as Parkinson’s disease (PD) offers a hopeful prospect for many patients. Previously, fetal grafts have been shown to survive, integrate and induce functional recovery in PD patients. However, limited tissue availability has haltered the widespread use of this therapy and begs the demand for alternative tissue sources. In this regard, stem cells may constitute one such source. i Objective/Methods: /i In this review we outline various types of stem cells currently available and provide an overview of their possible application for PD. We address not only the obvious possibility of using stem cells in cell replacement therapy but also the benefits of stem cell lines in drug discovery. i Results/Conclusion: /i Stem cells carrying reporters or mutations in genes linked to familial PD are likely to contribute to the identification of new drug targets and subsequent development of new drugs for PD. Thus, stem cells are, and will be more so in the future, invaluable tools in the quest for new therapies against neurodegenerative diseases such as PD.
Ryk, a receptor regulating wnt5a-mediated neurogenesis and axon morphogenesis of ventral midbrain dopaminergic neurons
Publisher: Mary Ann Liebert Inc
Date: 08-2013
Abstract: Ryk is an atypical transmembrane receptor tyrosine kinase that has been shown to play multiple roles in development through the modulation of Wnt signaling. Within the developing ventral midbrain (VM), Wnts have been shown to contribute to the proliferation, differentiation, and connectivity of dopamine (DA) neurons however, the Wnt-related receptors regulating these events remain less well described. In light of the established roles of Wnt5a in dopaminergic development (regulating DA differentiation as well as axonal growth and repulsion), and its interaction with Ryk elsewhere within the central nervous system, we investigated the potential role of Ryk in VM development. Here we show temporal and spatial expression of Ryk within the VM, suggestive of a role in DA neurogenesis and axonal plasticity. In VM primary cultures, we show that the effects of Wnt5a on VM progenitor proliferation, DA differentiation, and DA axonal connectivity can be inhibited using an Ryk-blocking antibody. In support, Ryk knockout mice showed reduced VM progenitors and DA precursor populations, resulting in a significant decrease in DA cells. However, Ryk(-/-) mice displayed no defects in DA axonal growth, guidance, or fasciculation of the MFB, suggesting other receptors may be involved and/or compensate for the loss of this receptor. These findings identify for the first time Ryk as an important receptor for midbrain DA development.
Transplantation of fetal midbrain dopamine progenitors into a rodent model of Parkinson's disease
Publisher: Humana Press
Date: 2013
DOI: 10.1007/978-1-62703-574-3_15
Abstract: Cell therapy is a promising experimental treatment for Parkinson's disease (PD). It is based on the idea that new dopamine neurons transplanted directly into the forebrain of the patient can structurally and functionally compensate for those lost to the disease in order to restore motor function. While there is a highly active field of research focused on the development of stem cell-based procedures, fetal tissue remains the "gold standard" as a safe and reliable source of dopamine neuron progenitors capable of structural and functional integration with existing motor circuitry following transplantation. This chapter describes the basic procedures for preparation of dopamine progenitor rich cell suspensions of ventral mesencephalon as well as implantation into the unilateral 6-hydroxydopamine model of PD and assessment of functional impact according to drug-induced rotational behavior. The description assumes a basic knowledge of animal handling and stereotaxic surgical procedures in rodents.
The human testis-determining factor SRY localizes in midbrain dopamine neurons and regulates multiple components of catecholamine synthesis and metabolism
Publisher: Wiley
Date: 08-06-2012
Combined immunohistochemical and retrograde tracing reveals little evidence of innervation of the rat dentate gyrus by midbrain dopamine neurons
Publisher: Springer Science and Business Media LLC
Date: 06-2016
Huntingtin Inclusions Trigger Cellular Quiescence, Deactivate Apoptosis, and Lead to Delayed Necrosis
Publisher: Elsevier BV
Date: 05-2017
DOI: 10.1016/J.CELREP.2017.04.029
Abstract: Competing models exist in the literature for the relationship between mutant Huntingtin exon 1 (Httex1) inclusion formation and toxicity. In one, inclusions are adaptive by sequestering the proteotoxicity of soluble Httex1. In the other, inclusions compromise cellular activity as a result of proteome co-aggregation. Using a biosensor of Httex1 conformation in mammalian cell models, we discovered a mechanism that reconciles these competing models. Newly formed inclusions were composed of disordered Httex1 and ribonucleoproteins. As inclusions matured, Httex1 reconfigured into amyloid, and other glutamine-rich and prion domain-containing proteins were recruited. Soluble Httex1 caused a hyperpolarized mitochondrial membrane potential, increased reactive oxygen species, and promoted apoptosis. Inclusion formation triggered a collapsed mitochondrial potential, cellular quiescence, and deactivated apoptosis. We propose a revised model where sequestration of soluble Httex1 inclusions can remove the trigger for apoptosis but also co-aggregate other proteins, which curtails cellular metabolism and leads to a slow death by necrosis.
The potential of stem cells and tissue engineered scaffolds for repair of the central nervous system
Publisher: Springer Netherlands
Date: 24-12-2012
Using minimalist self-assembling peptides as hierarchical scaffolds to stabilise growth factors and promote stem cell integration in the injured brain
Publisher: Hindawi Limited
Date: 27-11-2018
DOI: 10.1002/TERM.2582
Abstract: Neurotrophic growth factors are effective in slowing progressive degeneration and/or promoting neural repair through the support of residual host and/or transplanted neurons. However, limitations including short half-life and enzyme susceptibility of growth factors highlight the need for alternative strategies to prolong localised delivery at a site of injury. Here, we establish the utility of minimalist N-fluorenylmethyloxycarbonyl (Fmoc) self-assembling peptides (SAPs) as growth factor delivery vehicle, targeted at supporting neural transplants in an animal model of Parkinson's disease. The neural tissue-specific SAP, Fmoc-DIKVAV, demonstrated sustained release of glial cell line derived neurotrophic factor, up to 172 hr after gel loading. This represents a significant advance in drug delivery, because its lifetime in phosphate buffered saline was less than 1 hr. In vivo transplantation of neural progenitor cells, together with our growth factor-loaded material, into the injured brain improved graft survival compared with cell transplants alone. We show for the first time the use of minimalist Fmoc-SAP in an in vivo disease model for sustaining the delivery of neurotrophic growth factors, facilitating their spatial and temporal delivery in vivo, whilst also providing an enhanced niche environment for transplanted cells.
Viral Delivery of GDNF Promotes Functional Integration of Human Stem Cell Grafts in Parkinson's Disease.
Publisher: Elsevier BV
Date: 04-2020
Axonal Growth of Midbrain Dopamine Neurons is Modulated by the Cell Adhesion Molecule ALCAM Through Trans-Heterophilic Interactions with L1cam, Chl1, and Semaphorins
Publisher: Society for Neuroscience
Date: 12-07-2019
Meningeal cells influence midbrain development and the engraftment of dopamine progenitors in Parkinsonian mice
Publisher: Elsevier BV
Date: 05-2015
DOI: 10.1016/J.EXPNEUROL.2015.02.017
Abstract: Dopaminergic neuroblasts, isolated from ventral midbrain fetal tissue, have been shown to structurally and functionally integrate, and alleviate Parkinsonian symptoms following transplantation. The use of donor tissue isolated at an age younger than conventionally employed can result in larger grafts - a consequence of improved cell survival and neuroblast proliferation at the time of implantation. However studies have paid little attention to removal of the meninges from younger tissue, due to its age-dependent tight attachment to the underlying brain. Beyond the protection of the central nervous system, the meninges act as a signaling center, secreting a variety of trophins to influence neural development and additionally impact on neural repair. However it remains to be elucidated what influence these cells have on ventral midbrain development and grafted dopaminergic neuroblasts. Here we examined the temporal role of meningeal cells in graft integration in Parkinsonian mice and, using in vitro approaches, identified the mechanisms underlying the roles of meningeal cells in midbrain development. We demonstrate that young (embryonic day 10), but not older (E12), meningeal cells promote dopaminergic differentiation as well as neurite growth and guidance within grafts and during development. Furthermore we identify stromal derived factor 1 (SDF1), secreted by the meninges and acting on the CXCR4 receptor present on dopaminergic progenitors, as a contributory mediator in these effects. These findings identify new and important roles for the meningeal cells, and SDF1/CXCR4 signaling, in ventral midbrain development as well as neural repair following cell transplantation into the Parkinsonian brain.
Modelling the dopamine and noradrenergic cell loss that occurs in Parkinson's disease and the impact on hippocampal neurogenesis
Publisher: Wiley
Date: 24-02-2018
DOI: 10.1002/HIPO.22835
Functional Characterization of Friedreich Ataxia iPS-Derived Neuronal Progenitors and Their Integration in the Adult Brain
Publisher: Public Library of Science (PLoS)
Date: 07-07-2014
Midbrain dopaminergic neurogenesis and behavioural recovery in a salamander lesion-induced regeneration model
Publisher: The Company of Biologists
Date: 08-2007
DOI: 10.1242/DEV.002329
Abstract: Death and lack of functional regeneration of midbrain dopaminergic (DA)neurons, decreased DA input in the target striatum and movement anomalies characterise Parkinson's disease (PD). There is currently no cure for PD. One way to promote recovery would be to induce or enhance DA neurogenesis. Whether DA neurogenesis occurs in the adult midbrain is a matter of debate. Here, we describe the creation of a salamander 6-hydroxydopamine model of PD to examine midbrain DA regeneration. We demonstrate a robust and complete regeneration of the mesencephalic and diencephalic DA system after elimination of DA neurons. Regeneration is contributed by DA neurogenesis, leads to histological restoration, and to full recovery of motor behaviour. Molecular analyses of the temporal expression pattern of DA determinants indicate that the regenerating DA neurons mature along a similar developmental program as their mammalian counterparts during embryogenesis. We also find that the adult salamander midbrain can reactivate radial glia-like ependymoglia cells that proliferate. The salamander model provides insights into the mechanisms of DA regeneration/neurogenesis and may contribute to the development of novel regenerative strategies for the mammalian brain.
Locomotor hyperactivity in 14-3-3ζ KO mice is associated with dopamine transporter dysfunction
Publisher: Springer Science and Business Media LLC
Date: 03-12-2013
DOI: 10.1038/TP.2013.99
Developing stem cell-based therapies for neural repair
Publisher: Frontiers Media SA
Date: 2013
Motor and behavioral phenotype in conditional mutants with targeted ablation of cortical D1 dopamine receptor-expressing cells
Publisher: Elsevier BV
Date: 04-2015
DOI: 10.1016/J.NBD.2015.02.006
Abstract: D1-dopamine receptors (Drd1a) are highly expressed in the deep layers of the cerebral cortex and the striatum. A number of human diseases such as Huntington disease and schizophrenia are known to have cortical pathology involving dopamine receptor expressing neurons. To illuminate their functional role, we exploited a Cre/Lox molecular paradigm to generate Emx-1(tox) MUT mice, a transgenic line in which cortical Drd1a-expressing pyramidal neurons were selectively ablated. Emx-1(tox) MUT mice displayed prominent forelimb dystonia, hyperkinesia, ataxia on rotarod testing, heightened anxiety-like behavior, and age-dependent abnormalities in a test of social interaction. The latter occurred in the context of normal working memory on testing in the Y-maze and for novel object recognition. Some motor and behavioral abnormalities in Emx-1(tox) MUT mice overlapped with those in CamKIIα(tox) MUT transgenic mice, a line in which both striatal and cortical Drd1a-expressing cells were ablated. Although Emx-1(tox) MUT mice had normal striatal anatomy, both Emx-1(tox) MUT and CamKIIα(tox) MUT mice displayed selective neuronal loss in cortical layers V and VI. This study shows that loss of cortical Drd1a-expressing cells is sufficient to produce deficits in multiple motor and behavioral domains, independent of striatal mechanisms. Primary cortical changes in the D1 dopamine receptor compartment are therefore likely to model a number of core clinical features in disorders such as Huntington disease and schizophrenia.
Resolving pathobiological mechanisms relating to Huntington disease: Gait, balance, and involuntary movements in mice with targeted ablation of striatal D1 dopamine receptor cells
Publisher: Elsevier BV
Date: 02-2014
DOI: 10.1016/J.NBD.2013.09.015
Abstract: Progressive cell loss is observed in the striatum, cerebral cortex, thalamus, hypothalamus, subthalamic nucleus and hippoc us in Huntington disease. In the striatum, dopamine-responsive medium spiny neurons are preferentially lost. Clinical features include involuntary movements, gait and orofacial impairments in addition to cognitive deficits and psychosis, anxiety and mood disorders. We utilized the Cre-LoxP system to generate mutant mice with selective postnatal ablation of D1 dopamine receptor-expressing striatal neurons to determine which elements of the complex Huntington disease phenotype relate to loss of this neuronal subpopulation. Mutant mice had reduced body weight, locomotor slowing, reduced rearing, ataxia, a short stride length wide-based erratic gait, impairment in orofacial movements and displayed haloperidol-suppressible tic-like movements. The mutation was associated with an anxiolytic profile. Mutant mice had significant striatal-specific atrophy and astrogliosis. D1-expressing cell number was reduced throughout the rostrocaudal extent of the dorsal striatum consistent with partial destruction of the striatonigral pathway. Additional striatal changes included up-regulated D2 and enkephalin mRNA, and an increased density of D2 and preproenkephalin-expressing projection neurons, and striatal neuropeptide Y and cholinergic interneurons. These data suggest that striatal D1-cell-ablation alone may account for the involuntary movements and locomotor, balance and orofacial deficits seen not only in HD but also in HD phenocopy syndromes with striatal atrophy. Therapeutic strategies would therefore need to target striatal D1 cells to ameliorate deficits especially when the clinical presentation is dominated by a bradykinetic/ataxic phenotype with involuntary movements.
Three-dimensional nanofibrous scaffolds incorporating immobilized bdnf promote proliferation and differentiation of cortical neural stem cells
Publisher: Mary Ann Liebert Inc
Date: 06-2010
Abstract: Attempts to repair the central nervous system damaged as a result of trauma or disease will depend on the ability to restore the appropriate neuronal connectivity. This will rely on establishing appropriate chemical and physical environments for supporting neural cells and their processes and in this regard, engineering of biomaterials is of increasing interest. It will be important to understand how cells behave on these biomaterials in vitro, prior to future in vivo application. We reveal that modification of 3-dimensional (3D) electrospun poly-epsilon-caprolactone (PCL) nanofiber scaffolds by fiber alignment and aminolysation is superior to classical 2-dimensional (2D) culture-ware in promoting in vitro proliferation and differentiation of cortical cells. Many studies have examined the importance of exogenous soluble factors to promote cell fate specification. Here, we demonstrate that tethering the neurotrophin, brain-derived neurotrophic factor (BDNF), onto modified nanofibers is superior to culturing in the presence of soluble BDNF. Functional immobilization of BDNF to polymer nanofibers enhances neural stem cell (NSC) proliferation and directs cell fate toward neuronal and oligodendrocyte specification, essential for neural tissue repair. These findings indicate that modified PCL nanofibrous 3D scaffolds are capable of supporting NSCs and their derivatives and may present a new avenue for encouraging neural repair in the future.
Wnt5a-treated midbrain neural stem cells improve dopamine cell replacement therapy in parkinsonian mice
Publisher: American Society for Clinical Investigation
Date: 02-01-2008
DOI: 10.1172/JCI32273
Temporally controlled release of multiple growth factors from a self-assembling peptide hydrogel
Publisher: IOP Publishing
Date: 12-08-2016
DOI: 10.1088/0957-4484/27/38/385102
Abstract: Protein growth factors have demonstrated great potential for tissue repair, but their inherent instability and large size prevents meaningful presentation to biologically protected nervous tissue. Here, we create a nanofibrous network from a self-assembling peptide (SAP) hydrogel to carry and stabilize the growth factors. We significantly reduced growth factor degradation to increase their lifespan by over 40 times. To control the temporal release profile we covalently attached polysaccharide chitosan molecules to the growth factor to increase its interactions with the hydrogel nanofibers and achieved a 4 h delay, demonstrating the potential of this method to provide temporally controlled growth factor delivery. We also describe release rate based analysis to examine the growth factor delivery in more detail than standard cumulative release profiles allow and show that the chitosan attachment method provided a more consistent release profile with a 60% reduction in fluctuations. To prove the potential of this system as a complex growth factor delivery platform we demonstrate for the first time temporally distinct release of multiple growth factors from a single tissue specific SAP hydrogel: a significant goal in regenerative medicine.
A PITX3-EGFP Reporter Line Reveals Connectivity of Dopamine and Non-dopamine Neuronal Subtypes in Grafts Generated from Human Embryonic Stem Cells
Publisher: Elsevier BV
Date: 09-2017
Harnessing stem cells and biomaterials to promote neural repair
Publisher: Wiley
Date: 21-12-2018
DOI: 10.1111/BPH.14545
Dopamine Receptor Antagonists Enhance Proliferation and Neurogenesis of Midbrain Lmx1a-expressing Progenitors
Publisher: Springer Science and Business Media LLC
Date: 06-2016
DOI: 10.1038/SREP26448
Abstract: Degeneration of dopamine neurons in the midbrain causes symptoms of the movement disorder, Parkinson disease. Dopamine neurons are generated from proliferating progenitor cells localized in the embryonic ventral midbrain. However, it remains unclear for how long cells with dopamine progenitor character are retained and if there is any potential for reactivation of such cells after cessation of normal dopamine neurogenesis. We show here that cells expressing Lmx1a and other progenitor markers remain in the midbrain aqueductal zone beyond the major dopamine neurogenic period. These cells express dopamine receptors, are located in regions heavily innervated by midbrain dopamine fibres and their proliferation can be stimulated by antagonizing dopamine receptors, ultimately leading to increased neurogenesis in vivo . Furthermore, treatment with dopamine receptor antagonists enhances neurogenesis in vitro , both from embryonic midbrain progenitors as well as from embryonic stem cells. Altogether our results indicate a potential for reactivation of resident midbrain cells with dopamine progenitor potential beyond the normal period of dopamine neurogenesis.
Related Organisations
Florey Institute Of Neuroscience And Mental Health
Location: Australia
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View Funded ActivityKnowledge, Identification And Exploitation Of Dopaminergic Axon Guidance Cues Will Improve Cell Replacement Therapy For ParkinsonÍs Disease.
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