Diseases that have been modeled for reprogramming can be divided into rare, monocausal genetic diseases and the large group of sporadic and multifactorial diseases. No large-scale disease modeling is currently available for the latter group. It has been more difficult and challenging to obtain conclusive results from this group due to the complexity of the different genetic backgrounds and environmental clues involved in these diseases. However, even patients with monogenetic diseases within families display large genotype–phenotype variability (2
), likely due to environmental influence. It will be interesting to determine whether the same variability can be reproduced in iPSC-derived neural cells or if reprogramming in culture eliminates environmental ‘noise’.
Reprogramming of fibroblasts for several neurologic diseases has been reported (Table ), but few studies have actually recapitulated the phenotype of diseases in the iPSC-derived neuronal population. Successful generation of iPSC-derived neurons has been reported for sporadic middle- or late-age onset neurodegenerative diseases like amyotrophic lateral sclerosis (ALS) and Parkinson's disease (PD) (3
). From the reprogramming point of view, it is remarkable that aged fibroblasts (up to 85 years old) from ALS and PD patients could still be reset with similar efficiency as fibroblasts from younger patients.
iPSC for modeling human neurological diseases
Nonetheless, demonstration of disease-related phenotypes in the relevant cell type (i.e. the specific cells that are affected in the disease) has been a major challenge during the past 2 years. It may well be that iPSC-derived neural cells from age-dependent neurodegenerative diseases will not show a phenotypic difference compared with normal control cells with regard to morphology, differentiation and survival. Test assays for challenging these cells for characterization of disease phenotypes could be required. For example, culturing cells under increased oxidative stress may reveal and/or accelerate aberrant neuronal phenotypes in late-onset diseases.
Partial disease modeling with spontaneous induction of a disease phenotype was reported in two young childhood-onset monogenetic diseases: spinal muscular atrophy (SMA) and familial dysautonomia (FD) (6
). Both diseases are autosomal recessive and share the common trait of rapid disease progression within the first years of life. In addition, both diseases are associated with loss of function of the gene as well as a role in RNA processing.
SMA is a group of autosomal recessive diseases caused by large deletions or point mutations in the survival motor neuron (SMN
) genes, leading to loss of function of the survival motor neuron protein. SMN1
gene encodes a 20 kb protein, spans 9 exons and has a role in RNA processing (8
). SMA type 1 is characterized by mutations in the SMN1
gene and promotes fast progressing degeneration of motor neurons, inducing muscular atrophy and symmetric proximal lower limb weakness. SMA type 1 onset occurs around the sixth month of life, and death often occurs due to respiratory failure before the age of 2. Ebert et al
) were able to derive iPSC from fibroblasts from a single SMA patient and showed a decrease in iPSC-derived motor neuron survival after 6 weeks of differentiation, compared with iPSC-derived neurons from the patient's unaffected mother . It remains to be shown whether the neuronal cells derived were indeed functional (i.e. able to fire action potentials or make neuromuscular junctions). Moreover, authors detected an increase in the nuclear number of SMA ‘gems’—protein aggregates that correlate with disease intensity—in fibroblasts and iPSC derived from the SMA patient. Interestingly, this deficiency could be reversed in fibroblasts or SMA-iPSC by increasing wild-type SMA protein levels using non-specific SMN-inducing compounds (6
). This work showed for the first time a proof-of-principle for a potential future drug-screening platform using the iPSC technology. However, initial enthusiasm was decreased by the absence of other SMA patients and controls. Incorporation of additional control and patient cells would have reduced the concern that the observed phenotype is a consequence of the intrinsic iPSC variability system (discussed in what follows).
FD is an autosomal recessive disease mostly occurring in persons of Ashkenazi Jewish descent (12
). The disease is characterized by degeneration of sensory and autonomic neurons, leading to severe and often lethal autonomic dysfunction. Common clinical features include alacrima, hypoactivity and relative indifference to pain and temperature. A splicing defect in the IkB kinase complex-associated protein (IKBKAP
) gene results in a tissue-specific splicing defect, inducing a loss of function or reduced levels of the IKAP protein (13
). IPSC derived from three patients with FD revealed that neural crest precursors, specifically, had low levels of IKBKAP
expression. In addition, a defect in neuronal differentiation and migration was reported. A drug candidate, kinetin, was able to reduce the levels of mutant IKBKAP
splice forms and improved neuronal differentiation, but not cell migration, in iPSC-derived neural crest precursors, suggesting incomplete phenotype complementation (7
). Drug screening using kinetin-like variations could be performed using recovery of both of neuronal differentiation and cell migration phenotypes as readout in future studies.
PD is the second most common neurodegenerative disease. Prominent clinical features are motor symptoms (bradykinesia, tremor, rigidity and postural instability) and non-motor-related PD symptoms (olfactory deficits, autonomic dysfunction, depression and sleep disorders). PD is a synucleinopathy, with accumulation of misfolded alpha-synuclein, that forms intracellular inclusions: Lewy bodies and Lewy neurites. Loss of dopaminergic (DA) neurons in the substantia nigra of the midbrain and in other brain regions is a characteristic neuropathological hallmark (14
). Several different techniques to produce DA neurons in culture from human embryonic stem cells (HESC) are currently available. They include co-culture systems [such as mesencephalic astrocytes (16
) and stromal cell-derived inducing activity (17
)] and direct differentiation protocols (18
). When these cells were transplanted into animal models of PD, functional integration was observed, although technical issues were reported (19
). Disease modeling in the dish using embryonic stem cells (ESC) is still limited. Even though specific toxicity and cell death of α-synuclein overexpression were shown in mouse ESC-derived DA neurons (20
), α-synuclein overexpressed in human neural embryonic cells resulted in patterns of degeneration that recapitulate PD features only in some cases (22
). Mouse iPSC-derived precursors were differentiated into DA neurons and transplanted into 6-OHDA-lesioned rats, a rat model of DA depletion. The authors showed that a striatal graft of 1−3 × 105
iPSC-derived neurons expressed midbrain DA markers and functionally integrated after transplantation (23
). Primary fibroblasts from sporadic PD patients were successfully reprogrammed and differentiated into DA neurons as efficiently as those from healthy individuals (3
). These results are promising, as age does not seem to interfere with reprogramming. Phenotypic differences were not reported, indicating that more subtle analysis or even strong stressors or toxins will be necessary to reveal phenotypes of diseases with late onset.
ALS or Lou Gehrig's disease is a progressive fatal neurodegenerative disease affecting mainly motor neurons. The most common clinical features of ALS are degeneration of motor neurons producing fasciculation, muscle wasting and hyper-reflexia. Respiratory complications usually develop in patients with advanced disease and the cause of death is generally paralysis of the respiratory muscles and diaphragm. With a projected lifetime risk of 1/2000, ALS is considered one of the most common motor neuron diseases (24
). ALS is universally fatal, with a median age of onset of 55 years and survival of 2–5 years from symptoms onset. Although the exact pathophysiological mechanisms underlying neurodegeneration in ALS remain uncertain, the presence of a persistent inflammatory reaction prompted researchers to study the involvement of a non-cell-autonomous component in motor neuron death. HESC have been used for modeling both the autonomous and the non-cell-autonomous effects of ALS in vitro
, using a gene that is mutated in 20% of the familial cases, superoxide dismutase 1 (SOD1
). IPSC technology allows for the unprecedented opportunity to also include patient genetic background in the cell-modeling system. Dimos et al
) successfully reprogrammed cells from two familial ALS patients. The iPSC generated were able to differentiate in motor neurons (the affected neuronal subtype in ALS pathology), but no phenotype has yet been observed or reported. It remains to be determined whether iPSC-derived neurons have the potential to recapitulate ALS late-onset pathology in vitro
and if both familial and sporadic (vast majority of ALS) cases share common phenotypic traits in culture.