|ognizant Communication Corporation|
The Regenerative Medicine Journal
VOLUME 15, NUMBER 6, 2006
Cell Transplantation, Vol. 15, pp. 455-462, 2006
0963-6897/06 $90.00 + 00
Copyright © 2006 Cognizant Comm. Corp.
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Myoblast Therapy: From Bench to Bedside
Zhongmin Liu,1 Yanling Wu,1 and Bing-guan Chen2
1Heart Center, Shanghai East Hospital, Tongji University,
Shanghai 200120, China
2Research Center, Shanghai East Hospital, Tongji University, Shanghai 200120, China
Myoblasts are defined as stem cells containing skeletal muscle cell precursors. A decade of experimental work has revealed many properties of myoblasts, including the stability of resulting hybrid myofibers without immune suppression, the persistence of transgene expression, and the lack of tumorigenicity. Early phase clinical trials also showed that myoblast-based therapy is a promising approach for many intractable clinical conditions, including both muscle-related and non-muscle-related diseases. The potential application of myoblast therapy may be in the treatment of genetic muscle diseases, cardiomyocyte damaged heart diseases, and urinary incontinence. This review will provide an overview of myoblast biology, along with discussion of the potential application in clinical medicine. In addition, problems in current myoblast therapy and possible future improvements will be addressed.
Key words: Myoblast therapy; Duchenne muscular dystrophy; Myocardiopathy; Incontinence; Gene therapy
Address correspondence to Bing-guan Chen, M.D., Ph.D., Medical Research Center, Shanghai East Hospital, Tongji University, 150 Jimo Rd, Shanghai 200120, China. Tel: +86-21-38804518; Fax: +86-21-58798999; E-mail: email@example.com
Cell Therapy for Parkinson's Disease: Only Young Onset Patients Allowed? Reflections About the Results of Recent Clinical Trials With Cell Therapy and the Progression of Parkinson's Disease
Centro de Investigación Parkinson (CIP), Policlínica Gipuzkoa, San Sebastián, Spain
The selection of the best candidates for surgery among Parkinson's disease (PD) patients is a debated topic. This could be particularly important for transplantation studies in which patients with advanced PD and motor complications refractory to conventional pharmacological treatments are usually included. The development of lesions in nondopaminergic structures, which apparently are unaffected by the intervention, could eventually lead to the appearance of disabling, treatment-resistant symptoms. This has been considered as the crucial factor responsible for the outcome of any therapeutic procedure. However, other factors might be involved. It is suggested in this article that the rate of progression of PD and the effects of ageing are more important than the extradopaminergic involvement in the final outcome. Rate of progression of PD is critically related to the power of compensatory mechanisms, which are age related and under the control of still unknown genes. Thus, patients with young onset parkinsonism (YOP), either caused by gene mutations or not, could be the best candidates for surgery because they have a slower disease progression and more competent compensatory mechanisms. On the other hand, this can also explain the appearance of unexpected side effects such as the "runaway" dyskinesias reported following transplantation.
Key words: Surgery; Transplantation; Parkinson's disease; Compensatory mechanisms; Neural plasticity
Address correspondence to Gurutz Linazasoro, Centro de Investigación Parkinson (CIP), Policlínica Gipuzkoa, Parque Tecnológico de Miramón, 20012 San Sebastián (Gipuzkoa), Spain. E-mail: firstname.lastname@example.org
Bridging Nigrostriatal Pathway With Fibroblast Growth Factor-Primed Peripheral Nerves and Fetal Ventral Mesencephalon Transplant Recuperates From Deficits in Parkinsonian Rats
Yung-Hsiao Chiang,1 Shinn-Zong Lin,2 and Feng C. Zhou3
1Department of Neurological Surgery, Tri-Service General
Hospital, National Defense Medical Center, Taipei, Taiwan
2Tzu-Chi General Hospital, Buddist Tzu-Chi University, Hua-Lien, Taiwan
3Department of Anatomy and Cell Biology, Paul Stark Neuroscience Institute, Indiana University School of Medicine, Indianapolis, IN, USA
Previous studies have indicated that the nigrostriatal dopaminergic (DA) pathway can be reconstructed in hemiparkinsonian rats with a bridge transplantation technique involving fetal ventral mesencephalic transplants and glial cell line-derived neurotrophic factor. In this study, we examined if the nigrostriatal pathway can be restored by combining peripheral nervous tissue with the fetal ventral mesencephalon transplants. Adult rats were injected with 6-hydroxydopamine into left median forebrain bundle. Those with marked rotational behavior, which has been previously shown to indicate complete DA dennervtion, were used for transplant treatments. One month after the lesion, fetal ventral mesencephalic cells were transplanted into the nigral region followed by nigral-striatal grafting of peripheral nerves as a bridge. The bridging nerves (sciatic or intercostals) were pretreated with basic fibrous growth factor (nerve+bFGF+) or Hank's saline (nerve+bFGF-). We found that (a) animals receiving transplants of VM and bFGF+ nerve had a reduction in rotational behavior; (b) animals receiving bFGF- nerve bridge only had a partial improvement in rotation. Reinnervation of tyrosine hydroxylase (TH)-immunoreactive (ir) fibers into the striatum was found in both of the above groups with more innervation in the former than in the latter. No TH-ir fibers in lesioned striatum or reduction in rotational behavior were found in animals receiving VM only, or VM plus bFGF. Taken together, our data indicate that peripheral nerve, with the aid of bFGF, greatly facilitates the reconstitution of the TH pathway from nigra to striatum and improves motor function in hemiparkinsonian rats.
Key words: Nigrostriatal pathway; Parkinsonian rats; Bridge transplantation; Peripheral nerves; Fetal ventral mesencephalon
Address correspondence to Yung Hsiao Chiang, M.D., Ph.D., Department of Neurological Surgery, Tri-Service General Hospital, National Defense Medical Center, 325 Cheng Kung Road, Sec. 2, Taipei, Taiwan. Tel: 886-2-87927177; E-mail: email@example.com
Inhibition of p38 MAP Kinase in the Early Posttransplantation Phase Redistributes Blood Vessels From the Surrounding Stroma Into the Transplanted Endocrine Tissue
Åsa Johansson,1 Daniel Sandvik,1 and Per-Ola Carlsson1,2
1Department of Medical Cell Biology, Uppsala University,
SE-751 23 Uppsala, Sweden
2Department of Medical Sciences, Uppsala University, SE-751 85 Uppsala, Sweden
Transplanted pancreatic islets attain a chronically decreased vascular density following transplantation, despite the increased concentrations of vascular endothelial growth factor (VEGF) secreted from beta-cells in response to hypoxia during culture and in the immediate posttransplantation phase. VEGF, however, exerts dual effects on endothelial cells, and in islet endothelial cells of the adult, the vascular permeability-inducing effects of VEGF seem normally more pronounced than those to induce angiogenesis. p38 MAP kinase activity has recently been shown to serve as a switch to separate these properties of VEGF; inhibition of p38 MAP kinase activity enhances VEGF-induced angiogenesis and, at the same time, abrogates VEGF-induced vascular permeability. We hypothesized that the revascularization of transplanted islets may be hampered by a predisposition of adult islet endothelial cells to react to VEGF by forming fenestrae rather than migrating and proliferating. We therefore administered the p38 MAP kinase inhibitor SB203580 by daily IP injections for the first 14 days following transplantation, and then studied the influence of this treatment on the oxygen tension, blood perfusion, and vascular density of the islet grafts 1 month posttransplantation. SB203580 treatment redistributed islet graft blood vessels from the stroma into the endocrine tissue, and this redistribution of blood vessels into the endocrine tissue was accompanied by an increased oxygenation of the islet cells. However, the total number of blood vessels in the tissue was not affected. The blood perfusion of the islet grafts was also similar in control and SB203580-treated animals. Our results suggest that effects of VEGF to preferentially induce vascular permeability may partially contribute to, but is not the main cause of, low revascularization of transplanted islets.
Key words: Engraftment; Bandeiraea simplicifolia; Blood flow; Oxygen tension
Address correspondence to Per-Ola Carlsson, M.D., Ph.D., Department of Medical Cell Biology, Uppsala University, Husargatan 3, Box 571, SE-751 23 Uppsala, Sweden. Tel: +46 18 471 4425; Fax: +46 18?471 40 59; E-mail: Per-Ola.Carlsson@medcellbiol.uu.se
Isolated Pancreatic Islets in Three-Dimensional Matrices Are Responsive to Stimulators and Inhibitors of Angiogenesis
André Kidszun,1 Darius Schneider,1 Doris Erb,1 Gundula Hertl,1 Veronika Schmidt,2 Michael Eckhard,1 Klaus T. Preissner,2 Georg Breier,3 Reinhard G. Bretzel,1 and Thomas Linn1
1Medical Clinic and Policlinic 3, Justus Liebig University,
Rodthohl 6, 35392 Giessen, Germany
2Department of Biochemistry, Justus Liebig University, Friedrichstrasse 24, 35392 Giessen, Germany
3Department of Pathology, Medical Faculty Carl Gustav Carus, University of Technology Dresden, Fetscherstrasse 74, 01307 Dresden, Germany
The formation of a new microvasculature is essential for the long-term survival and function of the islet graft. In this study we examined endothelium of isolated pancreatic islets by stimulation with growth factors, different culture conditions, and genetic modification. We also inspected the effect of immunosuppressives used in human transplantation on angiogenesis. Isolated islets were embedded in a three-dimensional fibrin or Matrigel matrix. The effect of hyperglycemia, hypoxia, and the addition of VEGF and bFGF was investigated. We exposed islets from transgenic mice expressing the VEGF gene (RIP1VEGF-A) to high glucose (16.7 mmol/L) medium and tested the immunosuppressive agents rapamycin (100 ng/ml) and FK506 (100 ng/ml). To quantify angiogenesis the percentage of sprouting islets was determined. New endothelial capillary-like structures protruded from isolated pancreatic islets. Addition of VEGF to the islets and transgenic RIP-VEGF islets showed a two- to threefold increase of sprouting islets compared to control. Hypoxic culture conditions stimulated angiogenesis, resulting in a twofold increase of capillary sprouting. Rapamycin and FK506 proved to be potent inhibitors of angiogenesis in this system, because a decrease of sprouting islets of more than 20% by both agents was observed. Isolated pancreatic islets are capable of forming new capillary structures and are susceptible to pro- and antiangiogenic stimuli.
Key words: Angiogenesis; Islet transplantation; VEGF; Rapamycin; FK506; Fibrin matrix; Rip1VEGF-A
Address correspondence to Thomas Linn, M.D., Ph.D, Medical Clinic and Policlinic 3, Justus Liebig University, Rodthohl 6, 35392 Giessen, Germany. Tel: 0049-641-99-42841; Fax: 0049-641-99-42489; E-mail: Thomas.Linn@innere.med.uni-giessen.de
Endothelial Cell Preservation at 10°C Minimizes Catalytic Iron, Oxidative Stress, and Cold-Induced Injury
Michael A. J. Zieger1,2 and Mahesh P. Gupta1
1Methodist Research Institute, Clarian Health Partners, Inc.,
Indianapolis, IN, USA
2Department of Medicine, Indiana University School of Medicine, Indianapolis, IN, USA
There is growing evidence that oxidative stress plays an important role in mediating the injury induced by hypothermia during the preservation of cells and tissues for clinical or research use. In cardiovascular allografts, endothelial cell loss or injury may lead to impaired control of vascular permeability and tone, thrombosis, and inflammation. We hypothesized that hypothermia-induced damage to the endothelium is linked to increases in intracellular catalytic iron pools and oxidative stress. In this study, bovine aortic endothelial cells and cell culture methods were used to model the response of the endothelium of cardiovascular tissues to hypothermia. Confluent cells were stored at 0°C to 25°C and cell damage was measured by lipid peroxidation (LPO) and lactate dehydrogenase release. Varying the bleomycin-detectible iron (BDI) in cells modulated cold-induced LPO and cell injury. In untreated cells, injury was highest at 0°C and a minimum at 10°C. A similar temperature-dependent trend was found in BDI levels and cell plating efficiencies. Arrhenius plots of cell killing and iron accumulation rates showed biphasic temperature dependence, with minima at 10°C and matching activation energies above and below 10°C. These findings imply that the mechanisms underlying the hypothermic increase in catalytic iron, oxidative stress, and cell killing are the same and that preservation of the endothelium may be optimized at temperatures above those routinely used.
Key words: Hypothermia; Oxidative stress; Iron; Endothelial cells
Address correspondence to Michael A. J. Zieger, Ph.D., Methodist Research Institute, Clarian Health Partners, Inc., 1701 North Senate Blvd., Indianapolis, IN 46202, USA. Tel: (317) 962-6940; Fax: (317) 962-9369; E-mail: firstname.lastname@example.org
Application of a New Subretinal Injection Device in the Dog
András M. Komáromy,1 Signe E. Varner,2 Eugene de Juan,3 Gregory M. Acland,4 and Gustavo D. Aguirre1
1School of Veterinary Medicine, University of Pennsylvania,
Philadelphia, PA 19104, USA
2SurModics, Inc., Irvine, CA 92606, USA
3Beckman Vision Center, University of California, San Francisco, CA 94143, USA
4Baker Institute, Cornell University, Ithaca, NY 14853, USA
The use of a new subretinal injection device (RetinaJectTM Subretinal Cannula, SurModics, Inc., Eden Prairie, MN) to access the subretinal space in the canine model was evaluated. Subretinal injections were performed in 33 mongrel dogs between 2 and 52 months of age (median = 9 months). In 5 normal dogs the injection of 150 μl saline or India ink occurred by using a conventional subretinal injection device (CSID) with a 30-gauge anterior chamber irrigating cannula. The sclera had to be surgically exposed and penetrated before the subretinal injection with the CSID could occur. After removing the CSID, the conjunctiva over the sclerotomy site had to be closed. In a second group of 28 dogs [16 normals, 10 RPE65 mutants, and 2 with progressive rod cone degeneration (prcd)], the 25-gauge needle of the RetinaJectTM was used to penetrate the conjunctiva and the sclera. Once the tip of the needle was close to the retinal surface, a 39-gauge polyimide cannula was extended and brought into apposition with the retina for the subsequent subretinal injection of 150 ml saline, India ink, or adeno-associated virus (AAV). No closure of the conjunctiva was required. The animals were clinically monitored between 1 and 59 weeks after surgery. From this second group 25 eyes were harvested for routine histological analysis either immediately after surgery or after a clinical observation time of between 1 and 40 weeks. Both devices provided equally successful access to the subretinal space. The main advantage of the RetinaJectTM was that no surgical dissection was required; this led to a shorter procedure time and milder postoperative conjunctival swelling. In summary, the use of the RetinaJectTM can be recommended as an alternative to the CSID for subretinal injections in dogs.
Key words: Animal model; Dog; Retinitis pigmentosa; Subretinal injection
Address correspondence to András M. Komáromy, D.M.V., Ph.D., Department of Clinical Studies, School of Veterinary Medicine, University of Pennsylvania, 3900 Delancey Street, Philadelphia, PA 19104, USA. Tel: 215-573-2695; Fax: 215-573-2162; E-mail: email@example.com
Periosteal Cell Pellet Culture System: A New Technique for Bone Engineering
Maril Akiyama,1,2 Hidehiko Nonomura,1 Syed H. Kamil,1 and Ronald A. Ignotz1
1Center for Tissue Engineering, University of Massachusetts
Medical School, Worcester, MA 01655, USA
2Department of Biomaterials, Osaka Dental University, Hirakata-shi, Osaka 573-1121, Japan
To treat bone loss that is induced by disease or wounds, bone grafts are commonly used. In dentistry, guided tissue regeneration is effective in the treatment of periodontal diseases. However, bone resorption after implantation is a major problem with the bone graft and guided tissue regeneration technique. This study examines a cell pellet culture system without exogenous scaffolds for bone regeneration. First, we examined the effect of ascorbic acid on cells. Transmission electron microscopic observation revealed that cells formed a three-dimensional structure of multiple cell layers after 5 weeks of culturing in medium containing 50 mg/ml ascorbic acid with the medium changed every 7 days. A single cell pellet was produced by centrifuging cells that were gathered from 10 tissue culture dishes. Van Gieson staining and collagen type I immunostaining showed that the pellet contained collagen fibers and cells that adhered to the collagen fibers. Several of these cell pellets were implanted subcutaneously on the backs of nude mice for 6 weeks. Histology and immunohistochemistry results indicated new bone formation, vascular invasion, and insular areas of calcification. Bone tissue was surrounded by osteoblasts. The appearance of new bone formation is similar to that seen in intramembranous ossification. The present pellet system is reliable and might solve problems of bone resorption after implantation.
Key words: Pellet culture system; Bovine periosteal cells; Bone regeneration; Three-dimensional structure
Address correspondence to Mari Akiyama, Department of Biomaterials, Osaka Dental University, Hirakata-shi, Osaka 573-1121, Japan Tel: +81-72-864-3056; Fax: +81-72-864-3156; E-mail: firstname.lastname@example.org
Directed Three-Dimensional Growth of Microvascular Cells and Isolated Microvessel Fragments
Carlos C. Chang1 and James B. Hoying1,2,3
1Biomedical Engineering Program, University of Arizona, Tucson,
AZ 85724, USA
2Arizona Research Laboratories, University of Arizona, Tucson, AZ 85724, USA
3The BIO5 Institute, University of Arizona, Tucson, AZ 85724, USA
Tissue engineering has promise as a means for repairing diseased and damaged tissues. A significant challenge in tissue construction relates to the constraints placed on tissue geometries resulting from diffusion limitations. An ability to incorporate a premade vasculature would overcome these difficulties and promote construct viability once implanted. Most in vitro microvascular fabrication strategies rely on surface-associated cell growth, manipulated cell monolayers, or random arrangement of cells within matrix materials. In contrast, we successfully suspended microvascular cells and isolated microvessel fragments within collagen and then microfluidically drove the mixtures into microfabricated network topologies. Developing within the 3D collagen matrix, patterned cells progressed into cord-like morphologies. These geometries were directed by the surrounding elastomer mold. With similar techniques, suspended fragments formed endothelial sprouts. By avoiding the addition of exogenous growth factors, we allowed constituent cells and fragments to autonomously develop within the constructs, providing a more physiologically relevant system for in vitro microvascular development. In addition, we present the first examples of directed endothelial cell sprouting from parent microvessel fragments. We believe this system may serve as a foundation for future in vivo fabrication of microvascular networks for tissue engineering applications.
Key words: Microvascular; Tissue engineering; Microcirculation; Cell patterning
Address correspondence to James B. Hoying, University of Arizona, UA PO Box 245084, Tucson, AZ 85724-0001, USA. Tel: 520-626-5273; Fax: 520-626-2890; E-mail: email@example.com