Neural regeneration and cell replacement: a view from the eye

Neuronal degenerations in the retina are leading causes of blindness. Like most other areas of the CNS, the neurons of the mammalian retina are not replaced following degeneration. However, in nonmammalian vertebrates, endogenous repair processes restore neurons very efficiently, even after complete loss of the retina. We describe the phenomenon of retinal regeneration in nonmammalian vertebrates and attempts made in recent years to stimulate similar regenerative processes in the mammalian retina. In addition, we review the various strategies employed to replace lost neurons in the retina and the recent use of stem cell technologies to address problems of retinal repair.     

Stem cell-based approaches: Possible route to hearing restoration?

Disabling hearing loss is the most common sensorineural disability worldwide. It affects around 466 million people and its incidence is expected to rise to around900 million people by 2050, according to World Health Organization estimates.Most cases of hearing impairment are due to the degeneration of hair cells(HCs)in the cochlea, mechano-receptors that transduce incoming sound information into electrical signals that are sent to the brain. Damage to these cells is mainly caused by exposure to aminoglycoside antibiotics and to some anti-cancer drugs such as cisplatin, loud sounds, age, infections and genetic mutations. Hearing deficits may also result from damage to the spiral ganglion neurons that innervate cochlear HCs. Differently from what is observed in avian and nonmammalian species, there is no regeneration of missing sensory cell types in the adult mammalian cochlea, what makes hearing loss an irreversible process. This review summarizes the research that has been conducted with the aim of developing cell-based strategies that lead to sensory cell replacement in the adult cochlea and, ultimately, to hearing restoration. Two main lines of research are discussed, one directed toward the transplantation of exogenous replacement cells into the damaged tissue, and another that aims at reactivating the regenerative potential of putative progenitor cells in the adult inner ear. Results from some of the studies that have been conducted are presented and the advantages and drawbacks of the various approaches discussed.     

Identification of Tympanic Border Cells as Slow-Cycling Cells in the Cochlea

Mammalian cochlear sensory epithelial cells are believed to possess minimal regenerative potential because they halt proliferation during late stage of embryogenesis and never regenerate after birth. This means that sensorineural hearing loss caused by the death of cochlear sensory epithelial cells is a permanent condition. However, stem cells were recently identified in neonatal mice following dissociation of their inner ear organs. This suggests that regenerative therapy for sensorineural hearing loss may be possible. Unfortunately, dissociation distorts the microanatomy of the inner ear, making it difficult to determine the precise location of stem cells in unaltered specimens. To develop new therapeutic approaches based on sensory epithelial cell regeneration, the location of these stem cells must be elucidated. Stem cells normally proliferate at a slow rate in adult organs. In fact, so-called label-retaining cells, or slow-cycling cells, of the brain and skin are recognized as stem cells. In this study, using the exogenous proliferation marker, 5′-bromo-2′-deoxyuridine (BrdU) in combination with the endogenous proliferation marker Ki-67, we identified tympanic border cells. These cells, which are located beneath the basilar membrane in vivo, represent slow-cycling cells of the murine cochlea. Immunohistochemically, these cells stained positive for the immature cell marker Nestin. But it will be difficult to achieve regeneration of the cochlear function because these slow-cycling cells disappear in the mature murine cochlea.     

斑马鱼在再生医学研究中的应用及进展

组织器官的再生现象一直以来吸引着众多生物学家们的关注。再生能力在不同物种间差异很大, 与人及高等脊椎动物相比, 低等脊椎动物(如：斑马鱼)有着较高的再生能力。斑马鱼的鳍、心脏、视网膜、视神经、脊髓、肝脏及感觉毛细胞等都具有很强的再生能力。因此, 从斑马鱼再生过程的研究中将获得大量有用的信息, 促进对人类再生能力缺陷的认识, 进而推动再生医学的发展。文章就斑马鱼在心脏、神经系统、肝脏、鳍再生医学研究中的进展及应用做一综述。     

Otoacoustic emissions from insect ears: evidence of active hearing?

Sensitive hearing organs often employ nonlinear mechanical sound processing which generates distortion-product otoacoustic emissions (DPOAE). Such emissions are also recordable from tympanal organs of insects. In vertebrates (including humans), otoacoustic emissions are considered by-products of active sound amplification through specialized sensory receptor cells in the inner ear. Force generated by these cells primarily augments the displacement amplitude of the basilar membrane and thus increases auditory sensitivity. As in vertebrates, the emissions from insect ears are based on nonlinear mechanical properties of the sense organ. Apparently, to achieve maximum sensitivity, convergent evolutionary principles have been realized in the micromechanics of these hearing organs-although vertebrates and insects possess quite different types of receptor cells in their ears. Just as in vertebrates, otoacoustic emissions from insects ears are vulnerable and depend on an intact metabolism, but so far in tympanal organs, it is not clear if auditory nonlinearity is achieved by active motility of the sensory neurons or if passive cellular characteristics cause the nonlinear behavior. In the antennal ears of flies and mosquitoes, however, active vibrations of the flagellum have been demonstrated. Our review concentrates on experiments studying the tympanal organs of grasshoppers and moths; we show that their otoacoustic emissions are produced in a frequency-specific way and can be modified by electrical stimulation of the sensory cells. Even the simple ears of notodontid moths produce distinct emissions, although they have just one auditory neuron. At present it is still uncertain, both in vertebrates and in insects, if the nonlinear amplification so essential for sensitive sound processing is primarily due to motility of the somata of specialized sensory cells or to active movement of their (stereo-)cilia. We anticipate that further experiments with the relatively simple ears of insects will help answer these questions.     

Cell- and gene-therapy approaches to inner ear repair

Sensorineural hearing loss is the most common sensory disorder in humans. It is primarily due to the degeneration of highly specialised mechanosensory cells in the cochlea, the so-called hair cells. Hearing problems can also be caused or further aggravated by the death of auditory sensory neurons that convey the information from the hair cells to the brain stem. Despite the discovery of stem/progenitor cells in the mammalian cochlea, no regeneration of either damaged hair cells or auditory neurons has been observed in mammals, in contrast to what is seen in avians and non-mammalian vertebrates. The reasons for this divergence have not yet been elucidated, although loss of stem cells and/or loss of their phenotypic plasticity in adult mammals have been put forward as possible explanations. Given the high incidence of this disorder and its economic and social implications, a considerable number of research lines have been set up aimed towards the regeneration of cochlear sensory cell types. This review summarizes the various routes that have been explored, ranging from the genetic modification of endogenous cells remaining in the inner ear in order to promote their transdifferentiation, to the implantation of exogenous stem or progenitor cells and their subsequent differentiation within the host tissue. Prophylactic treatments to fight against progressive sensory cell degeneration in the inner ear are also discussed.     

The potential of stem cell research for the treatment of neuronal damage in glaucoma

Stem cell research offers a wide variety of approaches for the advancement of our understanding of basic mechanisms of neurodegeneration and tissue regeneration and for the discovery and development of new therapeutic strategies to prevent and restore neuronal cell loss. Similar to most other regions of our central nervous system, degenerative diseases of the retina lead to the loss of neurons, which are not replaced. Recent work in animals has provided proof-of-concept evidence for the restoration of photoreceptor cells by cell transplantation and neuronal cell replacement by regeneration from endogenous cell sources. However, efficient therapeutic prevention of neuronal cell loss has not been achieved. Moreover, successful cell replacement of retinal neurons in humans, including that of ganglion cells, remains a major challenge. Future successes in the discovery and translation of neuroprotective drug and gene therapies and of cell-based regenerative therapies will depend on a better understanding of the underlying disease pathomechanisms. Existing stem cell and cell-reprogramming technologies offer the potential to generate human retina cells, to develop specific human-cell-based retina disease models, and to open up novel therapeutic strategies. Further, we might glean substantial knowledge from species that can or cannot regenerate their neuronal retina, in the search for new therapeutic approaches. Thus, stem cell research will pave the way toward clinical translation. In this review, I address some of the major possibilities presently on offer and speculate about the power of stem cell research to gain further insights into the pathomechanisms of retinal neurodegeneration (with special emphasis on glaucoma) and to advance our therapeutic options.     

Müller Glia maintain their regenerative potential despite degeneration in the aged zebrafish retina

Ageing is a significant risk factor for degeneration of the retina. Müller glia cells (MG) are key for neuronal regeneration, so harnessing the regenerative capacity of MG in the retina offers great promise for the treatment of age‐associated blinding conditions. Yet, the impact of ageing on MG regenerative capacity is unclear. Here, we show that the zebrafish retina undergoes telomerase‐independent, age‐related neurodegeneration but that this is insufficient to stimulate MG proliferation and regeneration. Instead, age‐related neurodegeneration is accompanied by MG morphological aberrations and loss of vision. Mechanistically, yes‐associated protein (Yap), part of the Hippo signalling, has been shown to be critical for the regenerative response in the damaged retina, and we show that Yap expression levels decline with ageing. Despite this, morphologically and molecularly altered aged MG retain the capacity to regenerate neurons after acute light damage, therefore, highlighting key differences in the MG response to high‐intensity acute damage versus chronic neuronal loss in the zebrafish retina.     

The Link between Autotomy and CNS Regeneration: Echinoderms as Non-Model Species for Regenerative Biology

Achieving regeneration of the central nervous system (CNS) is a major challenge for regenerative medicine. The inability of mammals to regrow a severed CNS contrasts with the amazing regenerative powers of their deuterostome kin, the echinoderms. Rapid CNS regeneration from a specialized autotomy plane in echinoderms presents a highly tractable and suitable non-model system for regenerative biology and evolution. Starfish arm autotomy triggers mass cell migration and local proliferation, facilitating rapid CNS regeneration. Many regeneration events in nature are preceded by autotomy and there are striking parallels between autotomy and regeneration in starfish and lizards. Comparison of these systems holds promise to provide insight into regeneration deficiency in higher vertebrates and to uncover evolutionarily conserved deuterostome-chordate regenerative processes. This will help identify mechanisms that may be present but inactive in higher vertebrates to address the problem of their poor regenerative capacities and the challenge to achieve CNS repair and regrowth.     

Expression and function of pannexins in the inner ear and hearing

Pannexin (Panx) is a gene family encoding gap junction proteins in vertebrates. So far, three isoforms (Panx1, 2 and 3) have been identified. All of three Panx isoforms express in the cochlea with distinct expression patterns. Panx1 expresses in the cochlea extensively, including the spiral limbus, the organ of Corti, and the cochlear lateral wall, whereas Panx2 and Panx3 restrict to the basal cells of the stria vascularis in the lateral wall and the cochlear bony structure, respectively. However, there is no pannexin expression in auditory sensory hair cells. Recent studies demonstrated that like connexin gap junction gene, Panx1 deficiency causes hearing loss. Panx1 channels dominate ATP release in the cochlea. Deletion of Panx1 abolishes ATP release in the cochlea and reduces endocochlear potential (EP), auditory receptor current/potential, and active cochlear amplification. Panx1 deficiency in the cochlea also activates caspase-3 cell apoptotic pathway leading to cell degeneration. These new findings suggest that pannexins have a critical role in the cochlea in regard to hearing. However, detailed information about pannexin function in the cochlea and Panx mutation induced hearing loss still remain largely undetermined. Further studies are required.     

Stem‐cell‐based therapies for retinal degenerative diseases: Current challenges in the establishment of new treatment strategies

Various advances have been made in the treatment of retinal diseases, including new treatment strategies and innovations in surgical devices. However, the treatment of degenerative retinal diseases, such as retinitis pigmentosa (RP) and age‐related macular degeneration (AMD), continues to pose a significant challenge. In this review, we focus on the use of embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) to treat retinal diseases by harnessing the ability of stem cells to differentiate into different body tissues. The retina is a tissue specialized for light sensing, and its degradation leads to vision loss. As part of the central nervous system, the retina has very low regenerative capability, and therefore, treatment options are limited once it degenerates. Nevertheless, innovations in methods to induce the generation of retinal cells and tissues from ESCs/iPSCs enable the development of novel approaches for these irreversible diseases. Here we review some historical background and current clinical trials involving the use of stem‐cell‐derived retinal pigment epithelial cells for AMD treatment and stem cell‐derived retinal cells/tissues for RP therapy. Finally, we discuss our future vision of regenerative treatment for retinal diseases with a partial focus on our studies and introduce other interesting approaches for restoring vision.     

Limb regeneration in axolotl: is it superhealing?

The ability of axolotls to regenerate their limbs is almost legendary. In fact, urodeles such as the axolotl are the only vertebrates that can regenerate multiple structures like their limbs, jaws, tail, spinal cord, and skin (the list goes on) throughout their lives. It is therefore surprising to realize, although we have known of their regenerative potential for over 200 years, how little we understand the mechanisms behind this achievement of adult tissue morphogenesis. Many observations can be drawn between regeneration and other disciplines such as development and wound healing. In this review, we present new developments in functional analysis that will help to address the role of specific genes during the process of regeneration. We also present an analysis of the resemblance between wound healing and regeneration, and discuss whether axolotls are superhealers. A better understanding of these animals' regenerative capacity could lead to major benefits by providing regenerative medicine with directions on how to develop therapeutic approaches leading to regeneration in humans.     

Hair cell regeneration

The mammalian inner ear largely lacks the capacity to regenerate hair cells, the sensory cells required for hearing and balance. Recent studies in both lower vertebrates and mammals have uncovered genes and pathways important in hair cell development and have suggested ways that the sensory epithelia could be manipulated to achieve hair cell regeneration. These approaches include the use of inner ear stem cells, transdifferentiation of nonsensory cells, and induction of a proliferative response in the cells that can become hair cells.     

Intraflagellar transport genes are essential for differentiation and survival of vertebrate sensory neurons

Cilia play diverse roles in vertebrate and invertebrate sensory neurons. We show that a mutation of the zebrafish oval (ovl) locus affects a component of the ciliary transport (IFT) mechanism, the IFT88 polypeptide. In mutant retina, cilia are generated but not maintained, producing the absence of photoreceptor outer segments. A loss of cilia also occurs in auditory hair cells and olfactory sensory neurons. In all three sense organs, cilia defects are followed by degeneration of sensory cells. Similar phenotypes are induced by the absence of the IFT complex B polypeptides, ift52 and ift57, but not by the loss of complex A protein, ift140. The degeneration of mutant photoreceptor cells is caused, at least partially, by the ectopic accumulation of opsins. These studies reveal an essential role for IFT genes in vertebrate sensory neurons and implicate the molecular components of intraflagellar transport in degenerative disorders of these cells.     

Cytokine interplay among the diseased retina,inflammatory cells and mesenchymal stem cells - a clue to stem cell-based therapy

Retinal degenerative disorders, such as diabetic retinopathy, retinitis pigmentosa, age-related macular degeneration or glaucoma, represent the most common causes of loss of vision and blindness. In spite of intensive research, treatment options to prevent, stop or cure these diseases are limited. Newer therapeutic approaches are offered by stem cell-based therapy. To date, various types of stem cells have been evaluated in a range of models. Among them, mesenchymal stem/stromal cells (MSCs) derived from bone marrow or adipose tissue and used as autologous cells have been proposed to have the potential to attenuate the negative manifestations of retinal diseases. MSCs delivered to the vicinity of the diseased retina can exert local anti-inflammatory and repair-promoting/regenerative effects on retinal cells. However, MSCs also produce numerous factors that could have negative impacts on retinal regeneration. The secretory activity of MSCs is strongly influenced by the cytokine environment. Therefore, the interactions among the molecules produced by the diseased retina, cytokines secreted by inflammatory cells and factors produced by MSCs will decide the development and propagation of retinal diseases. Here we discuss the interactions among cytokines and other factors in the environment of the diseased retina treated by MSCs, and we present results supporting immunoregulatory and trophic roles of molecules secreted in the vicinity of the retina during MSC-based therapy.     

Towards gene therapy for deafness

Many hearing disorders are associated with the damage or loss of sensory hair cells (HC) which can produce a profound and irreversible deafness. Apoptosis pathway is reported to play an important role leading to rapid expansion of the HC lesion after exposure to intense noise. Furthermore, progress made over the last year in understanding molecular mechanisms involved in the proliferative and regenerative capacity of sensory cells in the mammalian inner ear has raised the possibility that targeted therapies might prevent the loss of these cells and preserve the patient's hearing. A first step towards the successful therapeutic exploitation is a better understanding of the different pathways that control survival and proliferation of sensory cells. In this review, we provide an overview of recent findings concerning the possibility to prevent apoptosis in auditory cells. We also show the current knowledge on the molecular mechanisms involved in the potential regenerative behavior of these cells and the progress of gene therapy to prevent deafness noise-induced.     

Lgr5+ amacrine cells possess regenerative potential in the retina of adult mice

Current knowledge indicates that the adult mammalian retina lacks regenerative capacity. Here, we show that the adult stem cell marker, leucine‐rich repeat‐containing G‐protein‐coupled receptor 5 (Lgr5), is expressed in the retina of adult mice. Lgr5⁺ cells are generated at late stages of retinal development and exhibit properties of differentiated amacrine interneurons (amacrine cells). Nevertheless, Lgr5⁺ amacrine cells contribute to regeneration of new retinal cells in the adult stage. The generation of new retinal cells, including retinal neurons and Müller glia from Lgr5⁺ amacrine cells, begins in early adulthood and continues as the animal ages. Together, these findings suggest that the mammalian retina is not devoid of regeneration as previously thought. It is rather dynamic, and Lgr5⁺ amacrine cells function as an endogenous regenerative source. The identification of such cells in the mammalian retina may provide new insights into neuronal regeneration and point to therapeutic opportunities for age‐related retinal degenerative diseases.