SummaryThe utility of stem cells and their progeny in adult transplantation models has been limited by poor survival and integration. We designed an injectable and bioresorbable hydrogel blend of hyaluronan and methylcellulose (HAMC) and tested it with two cell types in two animal models, thereby gaining an understanding of its general applicability for enhanced cell distribution, survival, integration, and functional repair relative to conventional cell delivery in saline. HAMC improves cell survival and integration of retinal stem cell (RSC)-derived rods in the retina. The pro-survival mechanism of HAMC is ascribed to the interaction of the CD44 receptor with HA. Transient disruption of the retinal outer limiting membrane, combined with HAMC delivery, results in significantly improved rod survival and visual function. HAMC also improves the distribution, viability, and functional repair of neural stem and progenitor cells (NSCs). The HAMC delivery system improves cell transplantation efficacy in two CNS models, suggesting broad applicability.

Diseases and injuries of the central nervous system (CNS) including those in the brain, spinal cord and retina are devastating because the CNS has limited intrinsic regenerative capacity and currently available therapies are unable to provide significant functional recovery. Several promising therapies have been identified with the goal of restoring at least some of this lost function and include neuroprotective agents to stop or slow cellular degeneration, neurotrophic factors to stimulate cellular growth, neutralizing molecules to overcome the inhibitory environment at the site of injury, and stem cell transplant strategies to replace lost tissue. The delivery of these therapies to the CNS is a challenge because the blood-brain barrier limits the diffusion of molecules into the brain by traditional oral or intravenous routes. Injectable hydrogels have the capacity to overcome the challenges associated with drug delivery to the CNS, by providing a minimally invasive, localized, void-filling platform for therapeutic use. Small molecule or protein drugs can be distributed throughout the hydrogel which then acts as a depot for their sustained release at the injury site. For cell delivery, the hydrogel can reduce cell aggregation and provide an adhesive matrix for improved cell survival and integration. Additionally, by choosing a biodegradable or bioresorbable hydrogel material, the system will eventually be eliminated from the body. This review discusses both natural and synthetic injectable hydrogel materials that have been used for drug or cell delivery to the CNS including hyaluronan, methylcellulose, chitosan, poly(N-isopropylacrylamide) and Matrigel.

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SummarySelf-renewing, multipotential retinal stem cells (RSCs) reside in the pigmented ciliary epithelium of the peripheral retina in adult mammals. RSCs can give rise to rhodopsin positive-cells, which can integrate into early postnatal retina, and represent a potentially useful option for cellular therapy. The ability to purify a stem cell population and direct the differentiation toward a particular cell lineage is a challenge facing the application of stem cells in regenerative medicine. Here we use cell sorting to prospectively enrich mouse RSCs based on size, granularity and low expression of P-cadherin and demonstrate that only rare cells with defined properties proliferate to form colonies. We show that clonally-derived mouse and human RSC progeny are multipotent and can differentiate into mature rhodopsin-positive cells with high efficiency using combinations of exogenous culture additives known to influence neural retinal development, including taurine and retinoic acid. This directed RSC differentiation follows the temporal sequence of photoreceptor differentiation in vivo, and the cells exhibit morphology, protein and gene expression consistent with primary cultures of rods in vitro. These results demonstrate that the RSC, an adult stem cell, can be enriched and directed to produce photoreceptors as a first step toward a targeted cell replacement strategy to treat retinal degenerative disease.

No effective clinical treatment currently exists for traumatic spinal cord injury. Cell replacement therapy holds promise for attaining functional repair. Cells may be delivered directly or near the injury site; however, this strategy requires a delivery vehicle to maintain cell viability. We have identified an injectable, biocompatible, and biodegradable hydrogel scaffold composed of hyaluronan (HA) and methylcellulose (MC) that may be an effective scaffold for therapeutic cell delivery. The purpose of the present study was to determine the effects of polymer concentration on HAMC mechanical strength, gelation time, and cell viability. The yield stress of HAMC, a measure of mechanical stiffness, was tunable via manipulation of MC and HA content. Measurement of the elastic and storage moduli as functions of time revealed that HAMC gels in less than 5 min at physiological temperatures. Human umbilical tissue-derived cells encapsulated in HAMC were homogenously and stably distributed over 3 days in culture and extended processes into the scaffold. Cell viability was stable over this period in all but the most concentrated HAMC formulation. Because of its strength-tunability, rapid gelation, and ability to maintain cell viability, HAMC is a promising vehicle for cell delivery and is being tested in ongoing in vivo studies.

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Purpose: A number of indicators suggest that the physician scientist career track is threatened. As such, it is an opportune time to evaluate current training models. Perspectives on physician scientist education and career path were surveyed in trainees at the University of Toronto, home to Canada’s longest standing physician scientist training programs. Methods: Trainees from the Clinician Investigator Program (CIP) and MD/PhD Program at the University of Toronto were surveyed. Liekert-style closed-ended questions were used to assess future career goals, present and future perspectives and concerns about and beliefs on training. Demographic information was collected regarding year of study, graduate degree program and focus of clinical and health research. Statistical analysis included non-parametric tests for sub-group comparisons. Results: Both groups of trainees were motivated to pursue a career as a physician scientist. While confident in their decision to begin and complete physician scientist training, they expressed concerns about the level of integration between clinical and research training in the current programs. They also expressed concerns about career outlook, including the ability to find stable and sustainable careers in academic medicine. Trainees highlighted a number of factors, including career mentorship, as essential for career success. Conclusion: These findings indicate that while trainees at different stages consistently express career motivation, they identified concerns that are program- and training stage-specific. These concerns mirror those highlighted in the medical education literature regarding threats to the physician scientist career path. Understanding these different and changing perspectives and exploring those differences could form an important basis for trainee program improvements both nationally and internationally.

Retinal stem cells (RSCs) are present within the pigmented ciliary epithelium (CE) of the adult human eye and produce progeny that differentiate in vitro into all neural retinal subtypes and retinal pigmented epithelium (RPE). We hypothesized that a RSC population, similar to the adult CE-derived RSC, is contained within pigmented colonies that arise in long-term cultures of hESCs suggested to recapitulate retinal development in vitro. Single pigmented hESC-derived cells were isolated and plated in serum-free media containing growth factors and, after 2 weeks, clonal sphere colonies containing both pigmented and nonpigmented cells were observed. These colonies expressed the early retinal transcription factors Rx, Chx10 and Pax6, and could be dissociated and replated as single cells to form secondary clonal colonies. When allowed to differentiate, expression of markers for both RPE and neurons was observed. Rhodopsin expression was detected after explant coculture and transplantation into the developing mouse eye as well as following treatment with soluble factors in vitro. We show that RSCs emerge in an in vitro model of retinal development and are a potential source of human photoreceptors for use in transplantation.