Human skeletal muscle-derived CD133(+) cells form functional satellite cells after intramuscular transplantation in immunodeficient host mice.

Stem cell therapy is a promising strategy for treatment of muscular dystrophies. In addition to muscle fiber formation, reconstitution of functional stem cell pool by donor cells is vital for long-term treatment. We show here that some CD133(+) cells within human muscle are located underneath the basal lamina of muscle fibers, in the position of the muscle satellite cell. Cultured hCD133(+) cells are heterogeneous and multipotent, capable of forming myotubes and reserve satellite cells in vitro. They contribute to extensive muscle regeneration and satellite cell formation following intramuscular transplantation into irradiated and cryodamaged tibialis anterior muscles of immunodeficient Rag2-/γ chain-/C5-mice. Some donor-derived satellite cells expressed the myogenic regulatory factor MyoD, indicating that they were activated. In addition, when transplanted host muscles were reinjured, there was significantly more newly-regenerated muscle fibers of donor origin in treated than in control, nonreinjured muscles, indicating that hCD133(+) cells had given rise to functional muscle stem cells, which were able to activate in response to injury and contribute to a further round of muscle regeneration. Our findings provide new evidence for the location and characterization of hCD133(+) cells, and highlight that these cells are highly suitable for future clinical application.

Neurotoxic and neurotrophic roles of proNGF and the receptor sortilin in the adult and ageing nervous system.

The precursor form of the nerve growth factor (proNGF), forms a heterotrimeric complex with the receptors p75 and sortilin; this complex has been implicated in neuron cell death. However, it is not known whether proNGF and the receptors p75 and sortilin contribute to age- and disease-related neurodegeneration. Here we show that proNGF induces cell death in subpopulations of basal forebrain and peripheral sympathetic neurons of old, but not of young, adult rodents. In contrast, proNGF appears to induce neurite outgrowth rather than cell death of young adult sympathetic neurons. We have examined the neurotoxic role of proNGF in old age, and find that proNGF protein is elevated during ageing in the projection areas of some populations of vulnerable central and peripheral neurons; caloric restriction, which has known neuroprotective effects, partially prevents these increases. Sortilin was found to play a significant part in the observed patterns of age-related proNGF-mediated neurotoxicity. In particular, survival of aged neurons was rescued by neurotensin, an alternative sortilin ligand that blocks the sortilin-mediated effects of proNGF. Furthermore, sortilin immunoreactivity increases markedly in ageing rodent basal forebrain and sympathetic neurons; in contrast, p75 levels are either unchanged or reduced. From these data we propose that selective age-related neuronal atrophy and neurodegeneration may be mediated by increased sortilin expression in neurons, together with elevated levels of proNGF expression in some targets.

ProNGF, sortilin, and age-related neurodegeneration.

Several studies have sought to demonstrate that neurodegeneration during disease and in old age is associated with reduced neurotrophic support. Little positive evidence has been forthcoming, either in relation to the availability of neurotrophins or to expression and function of the relevant receptors. Recently, a novel way in which neurotrophins could contribute to neurodegeneration has been suggested. In contrast to the well-known neurotrophic functions of the mature beta-form of nerve growth factor (mNGF), its precursor proNGF has recently been shown to be abundant in the adult brain and in the brains of patients with Alzheimer's disease. proNGF is synthesized as 25 and 32 kDa isoforms, which are glycosylated to form a principal 40 kDa species. Studies of the cortical targets of NGF-responsive basal forebrain neurons show that the 40 kDa form of proNGF is secreted in response to nerve stimulation, along with the proteases needed to generate the 13 kDa mNGF, or to degrade it. We have recently found that levels of 40 kDa proNGF are elevated in the aging brain and also in targets of peripheral NGF-responsive neurons. proNGF has been shown to be neurotoxic when bound in a heterotrimer with the p75 receptor and the receptor sortilin (identical to the neurotensin receptor NTS3). Interestingly, we find that sortilin levels increase in aged central and peripheral neurons, perhaps making these neurons more vulnerable to age-related increases in proNGF. Whether elevated levels of proNGF in targets or of sortilin in neurons contribute to known patterns of age- and disease-related neurodegeneration has not been previously investigated. Using in vitro models, our preliminary data now indicate that proNGF is indeed neurotoxic for aged, but not young, NGF-responsive basal forebrain and sympathetic neurons and that blockade of sortilin rescues proNGF-induced cell death. We therefore propose that increased proNGF in targets combined with increased sortilin expression in projecting neurons contributes to age-related neuronal atrophy and degeneration.

The effects of short-term JNK inhibition on the survival and growth of aged sympathetic neurons.

During the course of normal aging, certain populations of nerve growth factor (NGF)-responsive neurons become selectively vulnerable to cell death. Studies using dissociated neurons isolated from neonates have shown that c-Jun N-terminal kinases (JNKs) are important in regulating the survival and neurite outgrowth of NGF-responsive sympathetic neurons. Unlike neonatal neurons, adult sympathetic neurons are not dependent on NGF for their survival. Moreover, the NGF precursor, proNGF, is neurotoxic for aging but not young adult NGF-responsive neurons. Because of these age-related differences, the effects of JNK inhibition on the survival and growth of sympathetic neurons isolated from aged mice were studied. Aged neurons, as well as glia, were found to be dependent on JNK for their growth but not their survival. Conversely, proNGF neurotoxicity was JNK-dependent and mediated by the p75-interacting protein NRAGE, whereas neurite outgrowth was independent of NRAGE. These results have implications for the potential use of JNK inhibitors as therapies for ameliorating age-related neurodegenerative disease.