Stem cells
Abstract
Dopamine (DA) cell replacement therapy in Parkinson disease (PD) can be achieved using human fetal mesencephalic tissue; however, limited tissue availability has hindered further developments. Embryonic stem cells provide a promising alternative, but poor survival and risk of teratoma formation have prevented their clinical application. We present here a method for generating large numbers of DA neurons based on expanding and differentiating ventral midbrain (VM) neural stem cells/progenitors in the presence of key signals necessary for VM DA neuron development. Mouse VM neurospheres (VMNs) expanded with FGF2, differentiated with sonic hedgehog and FGF8, and transfected with Wnt5a (VMN-Wnt5a) generated 10-fold more DA neurons than did conventional FGF2-treated VMNs. VMN-Wnt5a cells exhibited the transcriptional and biochemical profiles and intrinsic electrophysiological properties of midbrain DA cells. Transplantation of these cells into parkinsonian mice resulted in significant cellular and functional recovery. Importantly, no tumors were detected and only a few transplanted grafts contained sporadic nestin-expressing progenitors. Our findings show that Wnt5a improves the differentiation and functional integration of stem cell–derived DA neurons in vivo and define Wnt5a-treated neural stem cells as an efficient and safe source of DA neurons for cell replacement therapy in PD.
Authors
Clare L. Parish, Gonçalo Castelo-Branco, Nina Rawal, Jan Tonnesen, Andreas Toft Sorensen, Carmen Salto, Merab Kokaia, Olle Lindvall, Ernest Arenas
Abstract
Authors
Jeffrey J. Ross, Zhigang Hong, Ben Willenbring, Lepeng Zeng, Brett Isenberg, Eu Han Lee, Morayma Reyes, Susan A. Keirstead, E. Kenneth Weir, Robert T. Tranquillo, Catherine M. Verfaillie
Abstract
Smooth muscle formation and function are critical in development and postnatal life. Hence, studies aimed at better understanding SMC differentiation are of great importance. Here, we report that multipotent adult progenitor cells (MAPCs) isolated from rat, murine, porcine, and human bone marrow demonstrate the potential to differentiate into cells with an SMC-like phenotype and function. TGF-β1 alone or combined with PDGF-BB in serum-free medium induces a temporally correct expression of transcripts and proteins consistent with smooth muscle development. Furthermore, SMCs derived from MAPCs (MAPC-SMCs) demonstrated functional L-type calcium channels. MAPC-SMCs entrapped in fibrin vascular molds became circumferentially aligned and generated force in response to KCl, the L-type channel opener FPL64176, or the SMC agonists 5-HT and ET-1, and exhibited complete relaxation in response to the Rho-kinase inhibitor Y-27632. Cyclic distention (5% circumferential strain) for 3 weeks increased responses by 2- to 3-fold, consistent with what occurred in neonatal SMCs. These results provide evidence that MAPC-SMCs are phenotypically and functionally similar to neonatal SMCs and that the in vitro MAPC-SMC differentiation system may be an ideal model for the study of SMC development. Moreover, MAPC-SMCs may lend themselves to tissue engineering applications.
Authors
Jeffrey J. Ross, Zhigang Hong, Ben Willenbring, Lepeng Zeng, Brett Isenberg, Eu Han Lee, Morayma Reyes, Susan A. Keirstead, E. Kenneth Weir, Robert T. Tranquillo, Catherine M. Verfaillie
Abstract
Vision loss associated with ischemic diseases such as retinopathy of prematurity and diabetic retinopathy are often due to retinal neovascularization. While significant progress has been made in the development of compounds useful for the treatment of abnormal vascular permeability and proliferation, such therapies do not address the underlying hypoxia that stimulates the observed vascular growth. Using a model of oxygen-induced retinopathy, we demonstrate that a population of adult BM–derived myeloid progenitor cells migrated to avascular regions of the retina, differentiated into microglia, and facilitated normalization of the vasculature. Myeloid-specific hypoxia-inducible factor 1α (HIF-1α) expression was required for this function, and we also demonstrate that endogenous microglia participated in retinal vascularization. These findings suggest what we believe to be a novel therapeutic approach for the treatment of ischemic retinopathies that promotes vascular repair rather than destruction.
Authors
Matthew R. Ritter, Eyal Banin, Stacey K. Moreno, Edith Aguilar, Michael I. Dorrell, Martin Friedlander
Abstract
Tissue regeneration requires the recruitment of adult stem cells and their differentiation into mature committed cells. In this study we describe what we believe to be a novel approach for tendon regeneration based on a specific signalling molecule, Smad8, which mediates the differentiation of mesenchymal stem cells (MSCs) into tendon-like cells. A biologically active Smad8 variant was transfected into an MSC line that coexpressed the osteogenic gene bone morphogenetic protein 2 (BMP2). The engineered cells demonstrated the morphological characteristics and gene expression profile of tendon cells both in vitro and in vivo. In addition, following implantation in an Achilles tendon partial defect, the engineered cells were capable of inducing tendon regeneration demonstrated by double quantum filtered MRI. The results indicate what we believe to be a novel mechanism in which Smad8 inhibits the osteogenic pathway in MSCs known to be induced by BMP2 while promoting tendon differentiation. These findings may have considerable importance for the therapeutic replacement of tendons or ligaments and for engineering other tissues in which BMP plays a pivotal developmental role.
Authors
Andrea Hoffmann, Gadi Pelled, Gadi Turgeman, Peter Eberle, Yoram Zilberman, Hadassah Shinar, Keren Keinan-Adamsky, Andreas Winkel, Sandra Shahab, Gil Navon, Gerhard Gross, Dan Gazit
Abstract
CD34+ bone marrow–derived progenitor cells contribute to tissue repair by differentiating into endothelial cells, vascular smooth muscle cells, hematopoietic cells, and possibly other cell types. However, the mechanisms by which circulating progenitor cells home to remodeling tissues remain unclear. Here we show that integrin α4β1 (VLA-4) promotes the homing of circulating progenitor cells to the α4β1 ligands VCAM and cellular fibronectin, which are expressed on actively remodeling neovasculature. Progenitor cells, which express integrin α4β1, homed to sites of active tumor neovascularization but not to normal nonimmune tissues. Antagonists of integrin α4β1, but not other integrins, blocked the adhesion of these cells to endothelia in vitro and in vivo as well as their homing to neovasculature and outgrowth into differentiated cell types. These studies describe an adhesion event that facilitates the homing of progenitor cells to the neovasculature.
Authors
Hui Jin, Aparna Aiyer, Jingmei Su, Per Borgstrom, Dwayne Stupack, Martin Friedlander, Judy Varner
Abstract
Ischemia causes kidney tubular cell damage and abnormal renal function. The kidney is capable of morphological restoration of tubules and recovery of function. Recently, it has been suggested that cells repopulating the ischemically injured tubule derive from bone marrow stem cells. We studied kidney repair in chimeric mice expressing GFP or bacterial β-gal or harboring the male Y chromosome exclusively in bone marrow-derived cells. In GFP chimeras, some interstitial cells but not tubular cells expressed GFP after ischemic injury. More than 99% of those GFP interstitial cells were leukocytes. In female mice with male bone marrow, occasional tubular cells (0.06%) appeared to be positive for the Y chromosome, but deconvolution microscopy revealed these to be artifactual. In β-gal chimeras, some tubular cells also appeared to express β-gal as assessed by X-gal staining, but following suppression of endogenous (mammalian) β-gal, no tubular cells could be found that stained with X-gal after ischemic injury. Whereas there was an absence of bone marrow–derived tubular cells, many tubular cells expressed proliferating cell nuclear antigen, which is reflective of a high proliferative rate of endogenous surviving tubular cells. Upon i.v. injection of bone marrow mesenchymal stromal cells, postischemic functional renal impairment was reduced, but there was no evidence of differentiation of these cells into tubular cells of the kidney. Thus, our data indicate that bone marrow–derived cells do not make a significant contribution to the restoration of epithelial integrity after an ischemic insult. It is likely that intrinsic tubular cell proliferation accounts for functionally significant replenishment of the tubular epithelium after ischemia.
Authors
Jeremy S. Duffield, Kwon Moo Park, Li-Li Hsiao, Vicki R. Kelley, David T. Scadden, Takaharu Ichimura, Joseph V. Bonventre
Abstract
Ischemic injury to the kidney produces acute tubular necrosis and apoptosis followed by tubular regeneration and recovery of renal function. Although mitotic cells are present in the tubules of postischemic kidneys, the origins of the proliferating cells are not known. Bone marrow cells (BMCs) can differentiate across lineages to repair injured organs, including the kidney. However, the relative contribution of intrarenal cells and extrarenal cells to kidney regeneration is not clear. We produced transgenic mice that expressed enhanced GFP (EGFP) specifically and permanently in mature renal tubular epithelial cells. Following ischemia/reperfusion injury (IRI), EGFP-positive cells showed incorporation of BrdU and expression of vimentin, which provides direct evidence that the cells composing regenerating tubules are derived from renal tubular epithelial cells. In BMC-transplanted mice, 89% of proliferating epithelial cells originated from host cells, and 11% originated from donor BMCs. Twenty-eight days after IRI, the kidneys contained 8% donor-derived cells, of which 8.4% were epithelial cells, 10.6% were glomerular cells, and 81% were interstitial cells. No renal functional improvement was observed in mice that were transplanted with exogenous BMCs. These results show that intrarenal cells are the main source of renal repair, and a single injection of BMCs does not make a significant contribution to renal functional or structural recovery.
Authors
Fangming Lin, Ashley Moran, Peter Igarashi
Abstract
Parkinson disease (PD) is a neurodegenerative disorder characterized by loss of midbrain dopaminergic (DA) neurons. ES cells are currently the most promising donor cell source for cell-replacement therapy in PD. We previously described a strong neuralizing activity present on the surface of stromal cells, named stromal cell–derived inducing activity (SDIA). In this study, we generated neurospheres composed of neural progenitors from monkey ES cells, which are capable of producing large numbers of DA neurons. We demonstrated that FGF20, preferentially expressed in the substantia nigra, acts synergistically with FGF2 to increase the number of DA neurons in ES cell–derived neurospheres. We also analyzed the effect of transplantation of DA neurons generated from monkey ES cells into 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine–treated (MPTP-treated) monkeys, a primate model for PD. Behavioral studies and functional imaging revealed that the transplanted cells functioned as DA neurons and attenuated MPTP-induced neurological symptoms.
Authors
Yasushi Takagi, Jun Takahashi, Hidemoto Saiki, Asuka Morizane, Takuya Hayashi, Yo Kishi, Hitoshi Fukuda, Yo Okamoto, Masaomi Koyanagi, Makoto Ideguchi, Hideki Hayashi, Takayuki Imazato, Hiroshi Kawasaki, Hirofumi Suemori, Shigeki Omachi, Hidehiko Iida, Nobuyuki Itoh, Norio Nakatsuji, Yoshiki Sasai, Nobuo Hashimoto
Abstract
The chemokine stromal cell–derived factor–1 (SDF-1) and its receptor, CXCR4, play a major role in migration, retention, and development of hematopoietic progenitors in the bone marrow. We report the direct involvement of atypical PKC-ζ in SDF-1 signaling in immature human CD34+-enriched cells and in leukemic pre-B acute lymphocytic leukemia (ALL) G2 cells. Chemotaxis, cell polarization, and adhesion of CD34+ cells to bone marrow stromal cells were found to be PKC-ζ dependent. Overexpression of PKC-ζ in G2 and U937 cells led to increased directional motility to SDF-1. Interestingly, impaired SDF-1–induced migration of the pre-B ALL cell line B1 correlated with reduced PKC-ζ expression. SDF-1 triggered PKC-ζ phosphorylation, translocation to the plasma membrane, and kinase activity. Furthermore we identified PI3K as an activator of PKC-ζ, and Pyk-2 and ERK1/2 as downstream targets of PKC-ζ. SDF-1–induced proliferation and MMP-9 secretion also required PKC-ζ activation. Finally, we showed that in vivo engraftment, but not homing, of human CD34+-enriched cells to the bone marrow of NOD/SCID mice was PKC-ζ dependent and that injection of mice with inhibitory PKC-ζ pseudosubstrate peptides resulted in mobilization of murine progenitors. Our results demonstrate a central role for PKC-ζ in SDF-1–dependent regulation of hematopoietic stem and progenitor cell motility and development.
Authors
Isabelle Petit, Polina Goichberg, Asaf Spiegel, Amnon Peled, Chaya Brodie, Rony Seger, Arnon Nagler, Ronen Alon, Tsvee Lapidot
Copyright © 2019 American Society for Clinical Investigation
ISSN: 0021-9738 (print), 1558-8238 (online)