Reversal of Cellular Phenotypes in Neural Cells Derived from Huntington’s Disease Monkey-Induced Pluripotent Stem Cells

Abstract

SummaryHuntington’s disease (HD) is a dominant neurodegenerative disorder caused by the expansion of glutamine residues in the N-terminal region of the huntingtin (HTT) protein. The disease results in progressive neuronal loss, leading to motor, cognitive, and psychiatric impairment. Here, we report the establishment of neural progenitor cell (NPC) lines derived from induced pluripotent stem cells (iPSCs) of transgenic HD monkeys. Upon differentiation to neurons, HD neural cells develop cellular features of HD, including the formation of nuclear inclusions and oligomeric mutant HTT (mHTT) aggregates, as well as increased apoptosis. These phenotypes are rescued by genetic suppression of HTT and pharmacological treatment, demonstrating the ability of our HD cell model to respond to therapeutic treatment. The development and reversal of HD-associated phenotypes in neural cells from HD monkeys provides a unique nonhuman primate (NHP) model for exploring HD pathogenesis and evaluating therapeutics that could be assessed further in HD monkeys.

Induced Pluripotent HD Monkey Stem Cells Derived Neural Cells for Drug Discovery

“…9 Similar observation was also described in NCs derived from HD-NPCs. 4 To examine if RI, ME and MB ameliorate apoptotic responses, they were administered into culture media of differentiated WT-NCs and HD-NCs for 24 hours followed by fixation and immunostaining using specific antibody that recognized cleaved caspase-3 and microtubule-associated protein-2 (Map2). Total cell count of Map-2 (green) and cleaved caspase-3 (red) positive cells was used to determine the ratio of apoptotic NC population (Fig.…”

“…We have shown that HD-NPCs and HD-NCs are highly susceptible to oxidative stress with HD cellular phenotypes including mHTT aggregates that can be reversed by genetic and biochemical approaches 4 . HD-NCs express glutamate receptors, which are the target of RI and ME, and a positive impact was observed similar to prior studies.…”

“…2, 3 Since the HD monkey model holds great promise as a preclinical animal model of HD, the current study has evolved based on the recent development of HD monkey NPCs (HD -NPCs) and their derivative neural cells (HD-NCs) that developed HD cellular features including increased susceptibility to oxidative stress, increased apoptosis, the formation of mHTT aggregate and intranuclear inclusions. 4 To further determine if HD-NPCs and HD-NCs derived from pluripotent stem cells (PSCs) of HD monkeys are a potential in vitro platform for HD drug discovery research, we examined the effect of three known drugs [Rilizole (2-amino-6-trifluoro-methoxy-benzothiazole; RI), Memantine (1-amino-3, 5-di-methyl-adamantane; ME), and Methylene blue (MB)] that had shown to have beneficial effects on HD. Cytotoxicity, apoptosis and mHTT aggregation were used as outcome measurements to determine if HD cellular pathologies were improved.…”

iPSC-based drug screening for Huntington׳s disease

“…These iPSCs were derived from skin cells of a transgenic monkey HD model with human HTT exon 1 with 84 CAG repeats (25). The iPSCs were able to differentiate into neurons and HD phenotypes were observed including aggregate formation of mutant HTT (mHTT) protein and increased cell death (38). These primate HD iPSC models provide another platform for drug screening with possibility of further exploring promising candidates in a non-human primate HD models.…”

“…As shown by immunocytochemical staining in Figure 2, phenotypes such as increase in cleaved HTT, activated caspases and decrease in BDNF and polymer F-actin as measured by phalloidin conjugated fluorophore are suitable endpoints for screening in HD-NSCs (Figure 2). ATP levels were lower in NSCs bearing a longer CAG expansion (38), which indicates impaired mitochondrial function, as mitochondria are the major power source inside cells that produce ATP. Furthermore, neurite outgrowth was impaired in neurons differentiated from HD patient derived iPSCs (26), a phenomenon resembling defects in neuronal maturation and axon outgrowth in HD.…”