OBrien Institute

Fitzroy, Australia

OBrien Institute

Fitzroy, Australia
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Tomaskovic-Crook E.,Agency for Science, Technology and Research Singapore | Crook J.M.,OBrien Institute | Crook J.M.,University of Melbourne
CNS and Neurological Disorders - Drug Targets | Year: 2011

There is a renewed enthusiasm for the clinical translation of human embryonic stem (hES) cells. This is abetted by putative clinically-compliant strategies for hES cell maintenance and directed differentiation, greater understanding of and accessibility to cells through formal cell registries and centralized cell banking for distribution, the revised US government policy on funding hES cell research, and paradoxically the discovery of induced pluripotent stem (iPS) cells. Additionally, as we consider the constraints (practical and fiscal) of delivering cell therapies for global healthcare, the more efficient and economical application of allogeneic vs autologous treatments will bolster the clinical entry of hES cell derivatives. Neurodegenerative disorders such as Parkinson's disease are primary candidates for hES cell therapy, although there are significant hurdles to be overcome. The present review considers key advances and challenges to translating hES cells into novel therapies for neurodegenerative diseases, with special consideration given to Parkinson's disease and Alzheimer's disease. Importantly, despite the focus on degenerative brain disorders and hES cells, many of the issues canvassed by this review are relevant to systemic application of hES cells and other pluripotent stem cells such as iPS cells. © 2011 Bentham Science Publishers.

Wu Y.,University of Western Ontario | Li J.,University of Western Ontario | Saleem S.,University of Western Ontario | Yee S.-P.,University of Western Ontario | And 3 more authors.
Laboratory Investigation | Year: 2010

Recent evidence has shown that stem cell factor (SCF) and its receptor, c-Kit, have an important role in pancreatic islet development by promoting islet cell differentiation and proliferation. In this study, we examined the role of c-Kit and SCF in the differentiation and proliferation of insulin- and glucagon-producing cells using a human pancreatic duct cell line (PANC-1). Our study showed that increased expression of endocrine cell markers (such as insulin and glucagon) and transcription factors (such as PDX-1 and PAX-6) coincided with a decrease in CK19 and c-Kit cells (P0.001) during PANC-1 cell differentiation, determined by immunofluorescence and qRT-PCR. Cells cultured with exogenous SCF showed an increase in insulin (26%) and glucagon (35%) cell differentiation (P0.01), an increase in cell proliferation (P0.05) and a decrease in cell apoptosis (P0.01). siRNA knockdown of c-Kit resulted in a decrease in endocrine cell differentiation with a reduction in PDX-1 and insulin mRNA, as well as the number of cells immunostaining for PDX-1 and insulin. Taken together, these results show that c-Kit/SCF interactions are involved in mediating islet-like cluster formation and islet-like cell differentiation in a human pancreatic duct cell line. © 2010 USCAP, Inc All rights reserved.

Joglekar M.V.,National Center for Cell Science | Joglekar M.V.,St. Vincent's Institute | Hardikar A.A.,National Center for Cell Science | Hardikar A.A.,OBrien Institute | Hardikar A.A.,University of Melbourne
Cell Cycle | Year: 2010

In vitro generation of insulin-producing cells from stem/progenitor cells presents a promising approach to overcome the scarcity of donor pancreas for cell replacement therapy in diabetes. In this regard, pancreatic islet-derived progenitors are proposed to be a better alternative as they are obtained from cells that can efficiently produce insulin under physiological conditions and are supposed to retain the epigenetic memory for producing 'insulin' even after transition to a mesenchymal-like cell type. However, in last few years there has been significant debate in understanding the origin of such islet-derived mesenchymal-like progenitor cells in vitro. The initial idea proposed that human insulin-producing β-cells contribute to generation of a population of islet-derived endocrine progenitor cells by a process of epithelial-to- mesenchymal transition (EMT) in vitro. This idea was challenged by a series of lineage-tracing studies in mice demonstrating the non-beta origin of mesenchymal cells in culture. However, recent observations made by two independent groups confirm that human islet insulin-producing cells can proliferate and contribute to mesenchymal-like cell populations in vitro. Here, we provide a fact sheet about the observations that are till now reported by several groups regarding origin of mesenchymal-like cells in the cultures of pancreatic islets. © 2010 Landes Bioscience.

Denham M.,University of Melbourne | Denham M.,Lund University | Thompson L.H.,Lund University | Thompson L.H.,Florey Neuroscience Institute | And 5 more authors.
Stem Cells | Year: 2010

Generation of mesencephalic dopamine (mesDA) neurons from human embryonic stem cells (hESCs) requires several stages of signaling from various extrinsic and intrinsic factors. To date, most methods incorporate exogenous treatment of Sonic hedgehog (SHH) to derive mesDA neurons. However, we and others have shown that this approach is inefficient for generating FOXA2+ cells, the precursors of mesDA neurons. As mesDA neurons are derived from the ventral floor plate (FP) regions of the embryonic neural tube, we sought to develop a system to derive FP cells from hESC. We show that forced expression of the transcription factor GLI1 in hESC at the earliest stage of neural induction, resulted in their commitment to FP lineage. The GLI1+ cells coexpressed FP markers, FOXA2 and Corin, and displayed exocrine SHH activity by ventrally patterning the surrounding neural progenitors. This system results in 63% FOXA2+ cells at the neural progenitor stage of hESC differentiation. The GLI1-transduced cells were also able to differentiate to neurons expressing tyrosine hydroxylase. This study demonstrates that GLI1 is a determinant of FP specification in hESC and describes a highly robust and efficient in vitro model system that mimics the ventral neural tube organizer. © AlphaMed Press.

Chan E.C.,University of Melbourne | Peshavariya H.M.,University of Melbourne | Liu G.-S.,University of Melbourne | Jiang F.,Shandong University | And 2 more authors.
Biochemical and Biophysical Research Communications | Year: 2013

The synthesis of extracellular matrix including collagen during wound healing responses involves signaling via reactive oxygen species (ROS). We hypothesized that NADPH oxidase isoform Nox4 facilitates the stimulatory effects of the profibrotic cytokine transforming growth factor (TGF) β1 on collagen production in vitro and in vivo. TGFβ1 stimulated collagen synthesis and hydrogen peroxide generation in mouse cardiac fibroblasts, and both responses were attenuated by a scavenger of superoxide and hydrogen peroxide (EUK-134). Furthermore, by expressing a dominant negative form of Nox4 (Adv-Nox4ΔNADPH) in fibroblasts, TGFβ1-induced hydrogen peroxide production and collagen production were abrogated, suggesting that Nox4-dependent ROS are important for TGFβ1 signaling in collagen production. This was confirmed by the inhibitory effect of an adenovirus carrying siRNA targeting Nox4 (Adv-Nox4i) on TGFβ1-induced collagen synthesis and expression of activated myofibroblasts marker smooth muscle alpha actin. Finally we used a mouse model of subcutaneous sponge implant to examine the role of Nox4 in the local stimulatory effects of TGFβ1 on collagen accumulation in vivo. TGFβ1-induced collagen accumulation was significantly reduced when the sponges were instilled with Adv-Nox4ΔNADPH. In conclusion, Nox4 acts as an intermediary in the signaling of TGFβ1 to facilitate collagen synthesis. © 2012 Elsevier Inc.

Williams M.D.,OBrien Institute | Williams M.D.,University of Melbourne | Mitchell G.M.,OBrien Institute | Mitchell G.M.,University of Melbourne | Mitchell G.M.,Australian Catholic University
Experimental Diabetes Research | Year: 2012

MicroRNAs (miRNAs) are a class of short, single-stranded non-protein coding gene products which can regulate the gene expression through post-transcriptional inhibition of messenger RNA (mRNA) translation. They are known to be involved in many essential biological processes including development, insulin secretion, and adipocyte differentiation. miRNAs are involved in complex metabolic processes, such as energy and lipid metabolism, which have been studied in the context of diabetes and obesity. Obesity, hyperlipidemia (elevated levels of blood lipids), and insulin resistance are strongly associated with the onset of type 2 diabetes. These conditions are also associated with aberrant expression of multiple essential miRNAs in pancreatic islets of Langerhans and peripheral tissues, including adipose tissue. A thorough understanding of the physiological role these miRNAs play in these tissues, and changes to their expression under pathological conditions, will allow researchers to develop new therapeutics with the potential to correct the aberrant expression of miRNAs in type 2 diabetes and obesity. Copyright © 2012 Michael D. Williams and Geraldine M. Mitchell.

Phillips B.W.,Singapore Institute of Medical Biology | Crook J.M.,OBrien Institute
BioDrugs | Year: 2010

The need for new and improved pharmacotherapies in medicine, high late-stage compound attrition in drug discovery, and upcoming patent expirations is driving interest by the pharmaceutical industry in pluripotent stem cells for in vitro modeling and early-stage testing of toxicity and target engagement. In particular, human embryonic and induced pluripotent stem cells represent potentially cost-effective and accessible sources of organ-specific cells that foretell in vivo human tissue response to new chemical entities. Here we consider the potential of these cells as novel tools for drug development, including toxicity screening and metabolic profiling. We hold that despite various challenges to translating proof-of-concept screening platforms to industrial use, the promise of research is considerable, and close to being realized. © 2010 Adis Data Information BV.

Choi Y.S.,OBrien Institute | Choi Y.S.,University of Melbourne | Matsuda K.,OBrien Institute | Dusting G.J.,OBrien Institute | And 6 more authors.
Biomaterials | Year: 2010

Cardiac tissue engineering offers promise as a surgical approach to cardiac repair, but requires an adequate source of cardiomyocytes. Here we evaluate the potential for generating human cardiac muscle cells in vivo from adipose-derived stem cells (ASC) by co-implanting in a vascularised tissue engineering chamber with inducing rat cardiomyocytes (rCM). Co-implantation (ASC-rCM) was compared with rCM or ASC controls alone after 6 weeks. Immunostaining using human nucleus specific antibody and cardiac markers revealed several fates for ASC in the chamber; (1) differentiation into cardiomyocytes and integration with co-implanted rCM; (2) differentiation into smooth muscle cells and recruitment into vascular structures; (3) adipogenic differentiation. ASC-rCM and ASC groups grew larger tissue constructs than rCM alone (212 ± 25 μl, 171 ± 16 μl vs. 137 ± 15 μl). ASC-rCM and rCM groups contracted spontaneously at up to 140 bpm and generated a 10-15-fold larger volume of cardiac muscle (14.5 ± 4.8 μl and 18.5 ± 2.6 μl) than ASC alone group (1.3 ± 0.5 μl). Vascular volume in ASC-rCM group was twice that of the rCM group (28.7 ± 5.0 μl vs. 14.8 ± 1.8 μl). The cardiac tissue engineered by co-implanting human ASC with neonatal rCM showed in vivo plasticity of ASC and their cardiomyogenic potential in tissue engineering. ASC contribution to vascularisation also promoted the growth of engineered tissue, confirming their utility in this setting. © 2009 Elsevier Ltd. All rights reserved.

Dilley R.J.,University of Western Australia | Dilley R.J.,OBrien Institute | Dilley R.J.,University of Melbourne | Morrison W.A.,OBrien Institute | And 2 more authors.
International Journal of Biochemistry and Cell Biology | Year: 2014

Cardiac tissue engineering is developing as an alternative approach to heart transplantation for treating heart failure. Shortage of organ donors and complications arising after orthotopic transplant remain major challenges to the modern field of heart transplantation. Engineering functional myocardium de novo requires an abundant source of cardiomyocytes, a biocompatible scaffold material and a functional vasculature to sustain the high metabolism of the construct. Progress has been made on several fronts, with cardiac cell biology, stem cells and biomaterials research particularly promising for cardiac tissue engineering, however currently employed strategies for vascularisation have lagged behind and limit the volume of tissue formed. Over ten years we have developed an in vivo tissue engineering model to construct vascularised tissue from various cell and tissue sources, including cardiac tissue. In this article we review the progress made with this approach and others, together with their potential to support a volume of engineered tissue for cardiac tissue engineering where contractile mass impacts directly on functional outcomes in translation to the clinic. It is clear that a scaled-up cardiac tissue engineering solution required for clinical treatment of heart failure will include a robust vascular supply for successful translation. This article is part of a directed issue entitled: Regenerative Medicine: the challenge of translation. © 2014 Elsevier Ltd. All rights reserved.

Chan E.C.,University of Melbourne | Chan E.C.,Obrien Institute | van Wijngaarden P.,University of Melbourne | Liu G.-S.,University of Melbourne | And 6 more authors.
Investigative Ophthalmology and Visual Science | Year: 2013

Purpose: The proliferation of new blood vessels in the retina is a leading cause of vision impairment. Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (Nox) is involved in cell signaling for ischemia-induced angiogenesis, but its role in retinal neovascularization is unclear. We have analyzed the dependence of retinal neovascularization on the Nox2 isoform in oxygen-induced retinopathy (OIR) in mice.Methods: Neonatal C57BL/6 mice aged 7 days (P7) were placed in a hyperoxic chamber (75% O2) for 5 days, followed by 5 days of exposure to room air. Eyes were harvested on P8 and P17 for the quantification of retinal vaso-obliteration and neovascularization, respectively. The retinal expression of Nox2 and VEGF-A were measured by RT-PCR, while superoxide generation was detected by in situ dihydroethidium (DHE) staining of fresh frozen sections.Results: In wild type (WT) mice, OIR was characterized by central retinal vaso-obliteration at P8 and neovascularization at P17, which was associated with increases in Nox2 and VEGF-A gene expression, superoxide generation, and accumulation of Iba-1 positive cells in the inner retina. In contrast, Nox2 knockout mice exhibited markedly less retinal neovascularization and VEGF-A mRNA expression at P17, despite showing comparable vaso-obliteration at P8. These changes were accompanied by reductions in DHE fluorescence and Iba-1–positive cell accumulation in the hypoxic retina.Conclusions: The Nox2-generated reactive oxygen species (ROS) facilitate the retinal expression of VEGF-A and neovascularization in this mouse model of OIR. Therapies targeting Nox2 could be of value to reduce aberrant retinal neovascularization in retinopathy of prematurity, diabetes, and other disease processes driven by VEGF. © 2013 The Association for Research in Vision and Ophthalmology, Inc.

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