Erin B. Lavik

Functional recovery following traumatic spinal cord injury mediated by a unique polymer scaffold seeded with neural stem cells

To better direct repair following spinal cord injury (SCI), we designed an implant modeled after the intact spinal cord consisting of a multicomponent polymer scaffold seeded with neural stem cells. Implantation of the scaffold–neural stem cells unit into an adult rat hemisection model of SCI promoted long-term improvement in function (persistent for 1 year in some animals) relative to a lesion-control group. At 70 days postinjury, animals implanted with scaffold-plus-cells exhibited coordinated, weight-bearing hindlimb stepping. Histology and immunocytochemical analysis suggested that this recovery might be attributable partly to a reduction in tissue loss from secondary injury processes as well as in diminished glial scarring. Tract tracing demonstrated corticospinal tract fibers passing through the injury epicenter to the caudal cord, a phenomenon not present in untreated groups. Together with evidence of enhanced local GAP-43 expression not seen in controls, these findings suggest a possible regeneration component. These results may suggest a new approach to SCI and, more broadly, may serve as a prototype for multidisciplinary strategies against complex neurological problems.

Differentiation of human embryonic stem cells on three-dimensional polymer scaffolds

Human embryonic stem (hES) cells hold promise as an unlimited source of cells for transplantation therapies. However, control of their proliferation and differentiation into complex, viable 3D tissues is challenging. Here we examine the use of biodegradable polymer scaffolds for promoting hES cell growth and differentiation and formation of 3D structures. We show that complex structures with features of various committed embryonic tissues can be generated, in vitro, by using early differentiating hES cells and further inducing their differentiation in a supportive 3D environment such as poly(lactic-co-glycolic acid)/poly(l-lactic acid) polymer scaffolds. We found that hES cell differentiation and organization can be influenced by the scaffold and directed by growth factors such as retinoic acid, transforming growth factor β, activin-A, or insulin-like growth factor. These growth factors induced differentiation into 3D structures with characteristics of developing neural tissues, cartilage, or liver, respectively. In addition, formation of a 3D vessel-like network was observed. When transplanted into severe combined immunodeficient mice, the constructs continue to express specific human proteins in defined differentiated structures and appear to recruit and anastamose with the host vasculature. This approach provides a unique culture system for addressing questions in cell and developmental biology, and provides a potential mechanism for creating viable human tissue structures for therapeutic applications.

Defect and transport properties of nanocrystalline CeO2−x

It is shown that unique defect thermodynamics and transport properties result for oxides of a few nanometers crystallite size. Fully‐dense CeO2−x polycrystals of ∼10 nm grain size were synthesized, and their electrical properties compared with those of samples coarsened from the same material. The nanocrystals showed reduced grain boundary resistance, 104 higher electronic conductivity, and less than one‐half the heat of reduction of its coarse‐grained counterpart. These properties are attributed to a dominant role of interfacial defect formation.

Tissue engineering: current state and perspectives

Abstract Tissue engineering is an interdisciplinary field that involves cell biology, materials science, reactor engineering, and clinical research with the goal of creating new tissues and organs. Significant advances in tissue engineering have been made through improving singular aspects within the overall approach, e.g., materials design, reactor design, or cell source. Increasingly, however, advances are being made by combining several areas to create environments which promote the development of new tissues whose properties more closely match their native counterparts. This approach does not seek to reproduce all the complexities involved in development, but rather seeks to promote an environment which permits the native capacity of cells to integrate, differentiate, and develop new tissues. Progenitors and stem cells will play a critical role in understanding and developing new engineered tissues as part of this approach.

Biodegradable polymer composite grafts promote the survival and differentiation of retinal progenitor cells.

Retinal progenitor cells (RPCs) are multipotent central nervous system precursors that give rise to all of the cell types of the retina during development. Several groups have reported that mammalian RPCs can be isolated and expanded in culture and can differentiate into retinal neurons upon grafting to the mature, diseased eye. However, cell delivery and survival remain formidable obstacles to application of RPCs in a clinical setting. Because biodegradable polymer/progenitor constructs have been shown to be capable of tissue generation in other compartments, we evaluated the survival, migration, and differentiation of RPCs delivered on PLLA/PLGA polymer substrates to the mouse subretinal space and compared these results to conventional injections of RPCs. Polymer composite grafts resulted in a near 10‐fold increase in the number of surviving cells after 4 weeks, with a 16‐fold increase in cell delivery. Grafted RPCs migrated into the host retina and expressed the mature markers neurofilament‐200, glial fibrillary acidic protein, protein kinase C‐α, recoverin, and rhodopsin. We conclude that biodegradable polymer/progenitor cell composite grafts provide an effective means of increasing progenitor cell survival and overall yield when transplanting to sites within the central nervous system such as the retina.

Modeling the neurovascular niche: VEGF- and BDNF-mediated cross-talk between neural stem cells and endothelial cells: An in vitro study

Neural stem cells (NSCs) exist in vascularized niches. Although there has been ample evidence supporting a role for endothelial cell‐derived soluble factors as modulators of neural stem cell self‐renewal and neuronal differentiation there is a paucity of data reported on neural stem cell modulation of endothelial cell behavior. We show that co‐culture of NSCs with brain‐derived endothelial cells (BECs) either in direct contact or separated by a porous membrane elicited robust vascular tube formation and maintenance, mediated by induction of vascular vascular endothelial growth factor (VEGF) and brain‐derived neurotrophic factor (BDNF) and activation of vascular VEGFR2 and TrkB by NSC NO. Nitric oxide (NO) scavengers and sequestration of VEGF and BDNF blunted this induction of tube formation, whereas addition of exogenous NO donor, rBDNF and rVEGF rescued the induction of tube formation. Further, rBDNF enhanced NSC eNOS activation and NO generation, suggesting an inducible positive feed‐back signaling loop between NSCs and BECs, providing for homeostasis and responsiveness of the resident NSCs and BECs comprising the neurovascular niche. These findings show the importance of reciprocal modulation of NSCs and BECs in induction and maintenance of the neurovascular niche and underscores their dynamic interactions.

A tissue-engineered approach towards retinal repair: Scaffolds for cell transplantation to the subretinal space

BackgroundSeveral mechanisms of retina degeneration result in the deterioration of the outer retina and can lead to blindness. Currently, with the exception of anti-angiogenic treatments for wet age-related macular degeneration, there are no treatments that can restore lost vision. There is evidence that photoreceptors and embryonic retinal tissue, transplanted to the subretinal space, can form new synapses with surviving host neurons. However, these transplants have yet to result in a clinical treatment for retinal degeneration.MethodsThis article reviews the current literature on the transplantation of scaffolds with retinal and retinal pigmented epithelial (RPE) cells to the subretinal space. We discuss the types of cells and materials that have been investigated for transplantation to the subretinal space, summarize the current findings, and present opportunities for future research and the next generation of scaffolds for retinal repair.ResultsChallenges to cell transplantation include limited survival upon implantation and the formation of abnormal cell architectures in vivo. Scaffolds have been shown to enhance cell survival and direct cell differentiation and organization in a number of models of retinal degeneration.ConclusionsThe transplantation of cells within a scaffold represents a possible treatment to repair retinal degeneration and restore vision in effected patients. Materials have been developed for the delivery of retinal and RPE cells separately however, the development of a combined tissue-engineered scaffold targeting both cell populations represents a promising direction for retinal repair.

Intravenous Hemostat: Nanotechnology to Halt Bleeding

Synthetic platelets composed of functionalized nanoparticles halve bleeding time in a rat injury model and may prove useful in treating human trauma victims. Basic first aid teaches us to put immediate pressure on a bleeding wound to stop the blood flow and allow natural clotting to occur. But what about when the wound is inside the body or difficult to compress? By coating polymer particles with peptides that promote platelet aggregation, Bertram et al. have made a synthetic “platelet” that accelerates natural platelet clotting and can be administered directly into the blood system to get access to internal organs. These tiny spheres can markedly decrease bleeding time in a rodent model with a serious injury to the femoral artery. Because blood clotting is well understood, the authors knew to choose the peptide arginine-glycine-aspartic acid (RGD) to attach to ~600 ends of the polyethylene glycol arms that extended from their 170-nm polylysine spheres. RGD binds to receptors on the surface of activated platelets, so the particles with multiple RGDs specifically adhered to multiple platelets, facilitating their aggregation. The authors optimized other features of the nanoparticles to guarantee that they would be useful in the emergency room or on the battlefield. The materials used to make the particles have all been used in devices previously approved by the U.S. Food and Drug Administration. The small RGD peptide can be inexpensively synthesized, and its size makes it unlikely to cause immunological problems. When stored dry, the platelet-like nanoparticles are stable and remain effective for at least 2 weeks, far surpassing the 5- to 7-day shelf life of donated platelets. They are cleared from the system (in rats) within 24 hours. To test how well these particles augmented blood clotting, the authors injected them into rats with a wound in the femoral artery. Whether the particles were injected before or, more realistically, after the wound was created, they reduced the bleeding time by 25% to 50%. Even the current standard of care for traumatic uncontrolled bleeding, a recombinant version of the natural clotting molecular factor VIIa, was less effective than the nanoparticles. Scanning electron micrographs of the blood clots from these treated rats confirmed that they contained numerous RGD-coated nanoparticles, nestled among blood cells, and a fibrin network. These nanoparticles augment only one of the many functions of real platelets—injury-induced aggregation—but, in a traumatic situation, that could be the critical function that is needed. Blood loss is the major cause of death in both civilian and battlefield traumas. Methods to staunch bleeding include pressure dressings and absorbent materials. For example, QuikClot effectively halts bleeding by absorbing large quantities of fluid and concentrating platelets to augment clotting, but these treatments are limited to compressible and exposed wounds. An ideal treatment would halt bleeding only at the injury site, be stable at room temperature, be administered easily, and work effectively for internal injuries. We have developed synthetic platelets based on Arg-Gly-Asp functionalized nanoparticles, which halve bleeding time after intravenous administration in a rat model of major trauma. The effects of these synthetic platelets surpass other treatments, including recombinant factor VIIa, which is used clinically for uncontrolled bleeding. Synthetic platelets were cleared within 24 hours at a dose of 20 mg/ml, and no complications were seen out to 7 days after infusion, the longest time point studied. These synthetic platelets may be useful for early intervention in trauma and demonstrate the role that nanotechnology can have in addressing unmet medical needs.

Novel drug delivery systems for glaucoma

Reduction of intraocular pressure (IOP) by pharmaceutical or surgical means has long been the standard treatment for glaucoma. A number of excellent drugs are available that are effective in reducing IOP. These drugs are typically applied as eye drops. However, patient adherence can be poor, thus reducing the clinical efficacy of the drugs. Several novel delivery systems designed to address the issue of adherence and to ensure consistent reduction of IOP are currently under development. These delivery systems include contact lenses-releasing glaucoma medications, injectables such as biodegradable micro- and nanoparticles, and surgically implanted systems. These new technologies are aimed at increasing clinical efficacy by offering multiple delivery options and are capable of managing IOP for several months. There is also a desire to have complementary neuroprotective approaches for those who continue to show progression, despite IOP reduction. Many potential neuroprotective agents are not suitable for traditional oral or drop formulations. Their potential is dependent on developing suitable delivery systems that can provide the drugs in a sustained, local manner to the retina and optic nerve. Drug delivery systems have the potential to improve patient adherence, reduce side effects, increase efficacy, and ultimately, preserve sight for glaucoma patients. In this review, we discuss benefits and limitations of the current systems of delivery and application, as well as those on the horizon.

Photopolymerized poly(ethylene glycol)/poly(L-lysine) hydrogels for the delivery of neural progenitor cells

Neural progenitor cells (NPCs) have shown promise in a number of models of disease and injury, but for these cells to be safe and effective, they must be directed to differentiate appropriately following transplantation. We have developed a photopolymerized hydrogel composed of macromers of poly(ethylene glycol) (PEG) bound to poly(L-lysine) (PLL) that supports NPC survival and directs differentiation. Green fluorescent protein (GFP) positive NPCs were encapsulated in these gels and demonstrated survival up to 17 days. When encapsulated in the gels at a photoinitiator concentration of 5.0 mg/ml, few NPCs (0.5 ± 0.25%) demonstrated apoptosis. Furthermore, 55 ± 6% of the NPCs cultured within the gels in epidermal growth factor (EGF) containing media differentiated into a mature neuronal cell type (neurofilament 200 positive) while the remainder 44 ± 8% were undifferentiated (nestin positive). A small percentage, 1 ± 0.4%, expressed the astrocytic marker glial acidic fibrilary protein. Photopolymerized PEG/PLL gels promote the survival and direct the differentiation of NPCs, making this system a promising delivery vehicle for NPCs in the treatment of injuries and diseases of the central nervous system.